Brenner and Rector's The Kidney, 8th ed.

CHAPTER 49. Hematologic Aspects of Kidney Disease

Steven Fishbane



Anemia of Kidney Failure, 1728



Definition and Prevalence of Anemia of Chronic Kidney Disease, 1728



Normal Erythropoiesis, 1729



Pathobiology of the Anemia of Kidney Disease, 1732



Consequences of Anemia in Chronic Kidney Disease, 1733



Disorders of Hemostasis in Chronic Kidney Disease, 1737


The disturbed metabolism caused by chronic kidney failure adversely affects many of the body's organs and systems. The erythropoietic system is affected in a number of ways, with the primary clinical manifestation being reduced erythrocyte mass, or anemia. This complication of renal failure was first observed by Richard Bright in 1836,[1] when he observed, in regard to nephritis, “after a time the healthy colour of the countenance fades.” A more complete description of renal anemia was provided by Sir Robert Christison (1797–1882). In patients with advanced nephritis, he observed the blood to be progressively diminished in color, “in the progress of time its proportion sinks; and at length it is reduced so much as to form less than a third of average.” Christison considered and then dismissed the possible etiologic role of blood letting. He concluded that no other natural disease came as close to hemorrhage for “impoverishing the red particles of the blood.” [2] [3] Anemia may be a relatively early complication in chronic kidney disease (CKD) and one that becomes more prevalent as kidney disease progresses. The detrimental impact of anemia on patients' quality of life makes it one of the most important complications of renal failure.

Definition and Prevalence of Anemia in Chronic Kidney Disease

Anemia is a state of deficient mass of red blood cells and hemoglobin (Hgb), resulting in insufficient oxygen delivery to the body's tissues and organs. [4] [5] [6] The normal values for Hgb and hematocrit (Hct) depend on gender, race, and other factors. In addition, the reference range changes in older individuals, with an unexpected increase in the prevalence of anemia. [7] [8] [9] [10] The National Kidney Foundation's clinical practice guidelines define anemia as a Hgb level less than 13.5 g/dL for adult men and less than 12.0 g/dL for adult women.[11] For patients living at altitude, a greater red cell mass is required to maintain tissue oxygen delivery given the reduced ambient oxygen tension.[12] [13] [14] [15] [16] In ad-dition, permanent residents at high altitude develop reduced Hgb oxygen affinity. [17] [18]

The prevalence of anemia in patients with CKD has been widely studied. In general, anemia becomes more frequent as renal function declines, becoming almost universal in end-stage renal disease (ESRD). [19] [20] [21] The prevalence reported in different studies depends on how anemia was defined and on the characteristics of the population studied. Most recent studies report on observations obtained from large databases of patients with varying levels of kidney function. The most useful analyses are those that were community based, avoiding biases inherent in studies of clinic-based populations. Hsu and co-workers[22] studied 12,055 adult ambulatory subjects from health clinics in Boston, using the Cockcroft-Gault equation to estimate creatinine clearance and the Modification of Diet in Renal Disease (MDRD) formula to estimate the glomerular filtration rate (GFR) indexed to body surface area. The authors found that mean Hct values decreased progressively when creatinine clearance was below 60 mL/min in men and below 40 mL/min in women. Moderately severe anemia, Hct less than 33%, was common (present in >20% of patients) only when GFR was severely depressed, less than 30 mL/min in women and 20 mL/min in men.[22] Hsu and associates[23] conducted a second study, using the third National Health and Nutrition Examination Survey (NHANES III) (1988–1994) of 15,971 adults aged over 18 years, with measurements of serum creatinine, Hgb, and iron indices. Creatinine clearance was estimated using the Cockcroft-Gault formula. A statistically significant decrease in mean Hgb was found in men and women with creatinine clearance less than 70 mL/min and 50 mL/min, respectively, compared with those with creatinine clearance greater than 80 mL/min. However, a mean decrease of 1.0 g/dL was found only for those with creatinine clearance less than 30 ml/min. Anemia was more common among women and non-Hispanic blacks. Among non-Hispanic blacks, the risk for anemia was generally more than twice that in non-Hispanic whites[23] ( Fig. 49-1 ). The authors extrapolated the data to the general U.S. population and estimated that approximately 1,590,000 Americans with creatinine clearance less than 50 mL/min are probably anemic with Hgb less than 12 g/dL.[23] Astor and colleagues[24] studied the same NHANES III data as Hsu and co-workers but restricted analysis to a different age range, 15,419 participants 20 years and older. Anemia (World Health Organization definition, Hgb < 13 g/dL in men, <12 g/dL in women) was present in 7.3% of all subjects ( Fig. 49-2 ). Both functional and absolute iron deficiency were found to be important predictors of anemia.[24]Interestingly, the prevalence of anemia decreased somewhat from NHANES III to NHANES IV, suggesting greater use of recombinant human erythropoietin treatment (rHuEPO) in CKD in recent years ( Fig. 49-3 ).[25]

FIGURE 49-1  A, The percentage of non-Hispanic white and black women, age 61 to 70, with hemoglobin less than 11 g/dL. B, The percentage of non-Hispanic white and black men, age 61 to 70, with hemoglobin less than 11 g/dL.  (Adapted from Hsu C, McCulloch C, Curhan G: Epidemiology of anemia associated with chronic renal insufficiency among adults in the United States: Results from the Third National Health and Nutrition Examination Survey. J Am Soc Nephrol 13:504–510, 2002.)



FIGURE 49-2  Prevalence of hemoglobin level less than 11 g/dL, 12 g/dL, and 13 g/dL among men (A) and women (B) 20 years and older from the Third National Health and Nutrition Examination Survey (NHANES III) (1988–1994). All values are adjusted to the age of 60 years.  (Adapted from Astor B, Muntner P, Levin A, et al: Association of kidney function with anemia. Arch Intern Med 162:1401–1408, 2002.)



FIGURE 49-3  Prevalence of hemoglobin less than 13.0 g/dL in males and less than 12.0 g/dL in females by chronic kidney disease stage in NHANES IV compared with NHANES III results.  (From U.S. Renal Data System: USRDS 2004 Annual Data Report. Bethesda, MD, National Institutes of Health, National Institute of Diabetes and Digestive and Kidney Diseases, 2004.)



Anemia develops earlier in CKD among patients with diabetes mellitus, and the magnitude of anemia tends to be more severe than in nondiabetic subjects. [26] [27] [28] [29] [30] [31] [32] [33] El-Achkar and colleagues[26] studied 5380 individuals in the community, surveyed as part of the Kidney Early Evaluation Program (KEEP), a community-based screening initiative for patients at high risk for kidney disease. Anemia was more prevalent among patients with diabetes and developed earlier than in nondiabetic subjects ( Fig. 49-4 ). In stage 3 CKD (GFR 30-59 mL/min), 22.2% of diabetics were anemic; in stage 4 disease (GFR 15-29 mL/min), the prevalence increased to 52.4%. The difference between diabetics and nondiabetics was greatest in stage 3 kidney disease, in which the prevalence of anemia was nearly threefold greater among diabetic subjects. The increased risk for anemia in diabetics was more pronounced among male compared with female subjects. The mechanism for the greater prevalence of anemia in diabetes was explored by Symeonidis and co-workers.[33] These investigators studied 694 anemic individuals, of whom 237 were diabetic. Serum erythropoietin levels were found to be lower in diabetic subjects, particularly in relation to the degree of anemia present. There was a negative correlation (r = -0.0446) between serum erythropoietin and the concentration of glycosylated Hgb. Thomas and associates[27] studied the contribution of proteinuria to anemia in diabetes among 315 Australian patients with type 1 diabetes. The prevalence of anemia was found to increase greatly in patients with macroalbuminuria compared with those with microalbuminuria or no albuminuria (52% vs. 24% vs. 8%).

FIGURE 49-4  Anemia prevalence (anemia defined as hemoglobin values < 12.0 g/dL for men and for postmenopausal women [>50 yr old], and <11.0 g/dL for premenopausal women [≤50 yr old] at different levels of glomerular filtration rate (GFR). Anemia develops earlier in patients with diabetes, and at any given level of GFR, the prevalence of anemia is greater in patients with diabetes.  (From El-Achkar TM, Ohmit SE, McCullough PA, et al: Higher prevalence of anemia with diabetes mellitus in moderate kidney insufficiency: The Kidney Early Evaluation Program. Kidney Int 67:1483–1488, 2005.)



With aging, renal function tends to progressively decline. The interaction of aging and loss of renal function might be expected to increase the prevalence of anemia. In actuality, the relationship is more complex. Whereas men with CKD tend to have greater prevalence of anemia with increasing age, among women, anemia is more frequent in younger than in older individuals.[23] It is likely that the high prevalence of iron deficiency in menstruating women accounts for the gender age discrepancy. If analysis is limited to older men and women, the effect of aging on risk for anemia becomes clearer. Ble and colleagues[34] studied 1005 community living individuals over 65 years old in Italy. The prevalence of anemia was found to increase with age in both sexes. By multivariate analysis, much of the risk for anemia segregated to individuals with creatinine clearance less than 30 mL/min, who also had lower mean serum erythropoietin levels.

Taken together, the studies discussed previously lead to several conclusions regarding the prevalence of anemia in CKD. First, anemia is relatively uncommon in earlier stages of CKD. Second, the prevalence of anemia begins to increase significantly with GFR less than 60 mL/minute, but anemia is generally not a frequent or severe complication of CKD until GFR is less than 30 mL/min. Third, anemia is a more significant problem for younger women, older men, and non-Hispanic blacks. Fourth, anemia occurs earlier and is more severe among patients whose kidney disease is due to diabetes mellitus. Screening for anemia (measurement of Hgb levels) should generally begin at stage 3 of CKD and should become more frequent with progressive disease.

Normal Erythropoiesis

The erythropoietic system functions to maintain a relatively stable level of serum Hgb and, thereby, blood oxygen transport and delivery. In the steady state, this requires production of a sufficient number of erythrocytes to replace the number lost each day owing to senescence. In addition, if anemia or hypoxia develops, then erythrocyte production must increase significantly. The system's afferent limb senses tissue oxygen delivery, conducting continuing surveillance for tissue hypoxia. The efferent limb is mediated via the glycoprotein hormone erythropoietin, a 30.4-kDa glycoprotein that is the major hormone regulator of erythrocyte production. It is an obligatory growth factor for erythroid development; erythroid progenitors do not survive without it. Erythropoietin functions as a true hormone in that it is produced in the kidneys, circulates in the bloodstream, and acts at receptors in bone marrow. The interaction of erythropoietin with its receptor on the surface of certain erythrocyte precursor cells in the bone marrow leads to enhanced cellular survival and completion of the erythropoietic process.

The first recognition of the linkage between hypoxia and erythrocyte quantity arose from astute 19th century observations on the effects of living at altitude. [35] [36] Carnot and Deflandre[37] first postulated that a humoral factor (a “hemopoietin”) might regulate erythropoiesis. Serum from anemic rabbits was injected into normal animals, resulting in increased reticulocyte counts.[37] These authors termed the circulating factor hematopoietine, but interest in the area waned as attempts to confirm their results were often unsuccessful.[3] Forty-four years later, Reissmann[38] rekindled interest in the field with ingenious experiments in parabiotic rats (i.e., artificial conjoined twins). In this model, rats were joined by skin and muscle, ear to tail, living for 3 months in parabiosis. When one animal breathed air with low oxygen tension and the other breathed normal air, both animals developed increased reticulocyte number and increased bone marrow erythropoiesis. This provided strong evidence that a humoral factor was the stimulus for erythropoiesis. In 1953, Erslev[39] definitively demonstrated the erythropoietic role of the serum factor, now termed erythropoietin. He infused 100 to 200 mL of plasma from bled anemic rabbits into normal rabbit recipients. Reticulocyte count increased rapidly and dramatically, with a fourfold increase in cell count within 4 days of infusion. [39] [40] In 1957, Jacobson and co-workers[41] provided indirect evidence to suggest that the kidneys were the primary source of erythropoietin. After demonstrating that removal of a variety of different organs did not affect erythropoietin production after phlebotomy, the investigators turned their attention to the kidneys. They found that nephrectomized rats and rabbits failed to increase erythropoietin production (incorporation of iron-59 into erythrocytes) after blood loss.[41] Further studies by Koury and associates[42] and Fisher and colleagues[43] demonstrated that the site of erythropoietin production within the kidneys was likely peritubular interstitial cells. Purification and identification of erythropoietin is credited to Miyake and co-workers.[44] These investigators spent years collecting urine of patients with aplastic anemia. From 2550 L of urine, using multiple isolation steps, a small quantity of pure glycoprotein was obtained.[44] Purification of human erythropoietin led to successful cloning of the gene, reported in 1985. Lin and associates[45] used erythropoietin supplied by Goldwasser's laboratory, derived from the urine of patients with aplastic anemia. After tryptase digestion, two fragments were sequenced, six and seven amino acids in length. These were used to create gene probes that were employed to screen a library of phage clones, resulting in three samples that contained portions of the erythropoietin gene; one contained the entire erythropoietin gene sequence. The gene was found to encode a protein of 193 amino acids, of which the last 166 residues corresponded to mature erythropoietin. When the gene was introduced into Chinese hamster ovary cells, erythropoietin with full biologic activity was produced.[45] Almost simultaneous to Lin's publication, Jacobs and colleagues[46] also reported isolation of the erythropoietin gene using similar techniques. These developments led in short order to the development of techniques to produce recombinant erythropoietin. On December 2, 1985, Eschbach and colleagues first administered rHuEPO at a low dose of 2.5 U/kg to an 18-year-old hemodialysis patient in Seattle, Washington. The patient's Hct did not increase. Soon thereafter, in another hemodialysis patient, at a higher dose of 15 U/kg thrice weekly, the first positive response was noted, an increase in Hct from 15% to 25% (personal communication, J. Eschbach). By 1989, clinical trials of rHuEPO demonstrated its remarkable efficacy, leading to regulatory approval and routine clinical erythropoietin replacement treatment.

The afferent limb of the erythropoietin system, centered on detection of tissue hypoxia, has been rapidly elucidated over the past decade.[47] A critical protein has been identified and extensively studied, hypoxia inducible-factor 1 (HIF-1). [4] [46] [47] [48] This complex is a heterodimer composed of two helix-loop-helix PAS proteins, HIF-1α and HIF-1β.[49] The b subunit is constitutively produced, with levels that do not respond to hypoxia. [50] [51] The α subunit (actually three members, 1α, 2α, and 3α) is produced constitutively but is rapidly degraded in the presence of oxygen by the ubiquitin-proteosome system. [52] [53] [54] [55] [56] In contrast, when hypoxia is present, degradation is inhibited, leading to rapid increases in HIF-1α levels. [47] [57] [58] [59] In animal knockout models, where no HIF-1α is produced, death occurs in midgestation.[59] In mice containing only one HIF-1α allele, the animals survive, but the erythropoietic response to hypoxia is blunted.[60] The rapid degradation of HIF-1α in the presence of oxygen depends on binding of the tumor suppressor protein von Hippel-Lindau (VHL) to two oxygen-dependent degradation domains (ODD) of HIF-1α.[61] This process results in the molecule being tagged for proteasomal degradation.[62] Recognition of HIF-1α by VHL requires two proline residues in the ODD to be hydroxylated. [64] [65] [66] Recently, three enzymes that drive this process have been identified, the HIF-prolyl hydroxylases (PHDs). [67] [68] The activity of the PHDs depends on the presence of oxygen, placing these enzymes in a central role for sensing oxygen and detecting hypoxia.[57]

When activated by tissue hypoxia, HIF-1 binds to the hypoxia response elements (HRE) of oxygen-regulated genes, including the erythropoietin gene, stimulating increased erythropoietin production. [69] [70] [71] [72] Rosenberger and associates [73] [74] studied HIF-1 in rats to assess the renal response to ischemia. HIF-1α was induced by ischemia in tubular cells, with different tubular locations depending on the cause of ischemia. HIF-2α was induced in some glomerular endothelial cells and in peritubular fibroblasts and endothelial cells. The location of HIF-2α induction, in cells known to produce erythropoietin, suggests a role in homeostatic erythropoietin regulation.

Erythropoietin itself is a 30.4-kDa glycoprotein, a member of the family of class 1 cytokines.[74] The carbohydrate moiety is probably important for molecular stability, whereas the 165 amino acid protein component is critical for receptor binding. [76] [77] [78] There are four discrete carbohydrate chains, three are N-linked oligosaccharides important for circulatory stability, and a small O-linked chain of unclear function. [78] [79] [80] [81] [82] [83] In the fetal period, erythropoietin is primarily produced by the liver; after birth, the kidneys are the major source of production. [75] [84] [85] [86] Clearance of circulating erythropoietin occurs by mechanisms that have not yet been full elucidated. The liver, kidneys, and bone marrow have all been studied as possible sources of erythropoietin elimination. None, however, appears to play an important role. Instead, it has been suggested that a more likely route of clearance is receptor-mediated uptake of erythropoietin, mainly in bone marrow.[86]

The quantity of erythropoietin is traditionally expressed in units, with 1 unit representing the same erythropoietic effect in animals as occurs after stimulation with 5 mmol cobalt chloride.[74] Steady-state production of small amounts of erythropoietin maintains the serum erythropoietin concentration at approximately 10 to 30 U/L, enough to stimulate sufficient production of erythrocytes to replace those lost to senescence. [57] [88] When anemia or hypoxia is present, serum erythropoietin levels increase rapidly to as much as 10,000 U/L. [69] [70] [89] A clear demonstration of the relationship between acute anemia, serum erythropoietin levels, and Hgb response was provided by the work of Al-Huniti and co-workers.[89] Sheep underwent acute phlebotomy to lower Hgb from 10 g/dL to less than 4 g/dL. Serum erythropoietin levels rapidly increased, from 15 U/L at baseline to a peak of 836 U/L by 1.5 to 3.7 days after the phlebotomy. Surprisingly, the response was temporary; serum levels began to decrease after approximately 3 to 4 days, declining to near baseline levels as the Hgb level began to increase.[89] In contrast, human studies indicate a more sustained increase in serum erythropoietin levels after phlebotomy, with levels remaining elevated for several weeks. [91] [92] With chronic anemia, as occurs with pure red cell aplasia and aplastic anemia, serum erythropoietin remains chronically elevated, with levels as great as 1000-fold higher compared with normal. [93] [94] [95] [96] [97]

In bone marrow, erythropoietin works to enhance survival of certain erythrocyte progenitor cells. [75] [98] [99] [100] Hematopoietic cells of the bone marrow develop along lines that lead to progressive and irreversible commitment to certain blood cell lines ( Fig. 49-5 ). Erythropoietic maturation begins with the pluripotent stem cell, which is influenced by nonspecific cytokines such as insulin-like growth factor-1 and interleukin-3. [101] [102] Stimulated stem cells proliferate and develop into early erythroid progenitor cells. Maturation continues through cells increasingly committed to expressing erythrocytic characteristics. Although schematics often portray erythrocyte development as a series of discrete steps, in actuality, the process is more of a continuous gradient of progressive phenotypic maturation.

FIGURE 49-5  Erythrocyte progenitors and precursors. The stages at which different cytokines and growth factors act to influence development. BFU-E, burst forming unit erythroid; CFU-E, colony forming unit; EPO, erythropoietin; IGF-1, insulin-like growth factor-1; IL-3, interleukin-3. (From Fisher JW: Erythopoietin: Physiology and pharmacology update. Exp Biol Med (Maywood) 288(1):1–14, 2003.)



Burst-forming units-erythroid (BFU-E), named for their capacity to generate multiclustered colonies of cells, are the earliest cell type exclusively committed to the erythrocyte line.[102] It is believed that these cells are produced stochastically from pluripotent stem cells. Only a minority of BFU-E, 10% to 20%, are in cell cycle at any given time; the rest remain an inert reserve of progenitor cells. With continued maturation of BFU-E, receptors for cytokines are lost, and there is a dramatic increase in the number of cell surface receptors for erythropoietin. [103] [104] Later in this stage, cells begin to take on the characteristics of colony-forming units-erythroid (CFU-E).[104] BFU-E contain only small quantities of GATA-1, an key transcription factor for erythroid development, whereas CFU-E have much higher concentrations. [106] [107] CFU-E begin to express some attributes of mature erythrocytes, including blood group and Rh antigens. [108] [109] It is at the CFU-E stage that erythropoietin exerts its greatest influence; CFU-E cells express the highest surface concentration of erythropoietin receptors of any erythrocyte precursor.[109]Without erythropoietin present, these cells are rapidly lost to programmed cell death. [111] [112] [113] [114] [115] Under the influence of erythropoietin, these cells survive and mature into Hgb-producing erythrocyte precursors and, finally, into the first circulating erythrocyte form, the reticulocyte. The interaction of erythropoietin with CFU-E is the key process determining net erythrocyte production. After this stage, the number of erythropoietin receptors per cell declines progressively, with none found on the surface of reticulocytes or mature erythrocytes. [116] [117]

The erythropoietin receptor is a 55-D transmembrane protein that belongs to the cytokine receptor super-family. [118] [119] [120] [121] [122] [123] [124] [125] The receptor undergoes homodimerization after binding to erythropoietin, with a change in conformation that leads to activation of JAK2 (janus kinase 2), a tyrosine kinase that is preassociated with the receptor. [126] [127] [128] [129] [130] [131] [132] [133] [134] This is a key activation step; animals lacking the gene for JAK2 die as embryos.[134] Activated JAK2 phosphorylates several tyrosine molecules of the erythropoietin receptor, exposing binding sites for key signaling proteins. [136] [137] The result is a cascade of signal transduction, with activation of multiple pathways including Ras/MAP kinase, JNK/p38 MAP kinase, JAK/STAT, and PI-3 kinase. [138] [139] [140] [141] [142] [143] The interaction of JAK2 with an important intermediary in signal transduction, STAT, has been extensively studied. After phosphorylation, STAT5 becomes activated and undergoes homodimerization and may translocate to the nucleus where it activates erythropoietin-inducible genes.[143] The net effect of the signaling cascade is promotion of mitogenesis and differentiation, with enhanced survival of committed erythrocyte progenitors and precursors, in particular the CFU-E. [145] [146] When stimulation by erythropoietin is terminated, it appears to take 30 to 60 minutes for the signaling cascade to return to basal levels.[146]

Pathobiology of the Anemia of Kidney Disease

Anemia in CKD can develop because of any of the hematologic diseases that may afflict individuals without kidney disease such as iron deficiency, vitamin B12 [148] [149] or folic acid deficiency,[148] and chronic blood loss.[149] But the form of anemia most common in CKD is a normocytic, normochromic anemia with insufficient production of erythrocytes. [151] [152] [153] The etiology is multifactorial, with contributors including relative erythropoietin deficiency, iron deficiency, blood loss, hemolysis, and other factors. In addition, it has long been noted that kidney disease is associated with circulating inhibitors of erythropoiesis. [154] [155] [156] But the preponderance of evidence demonstrates the primary role of erythropoietin deficiency as the major cause of anemia in CKD. [157] [158] [159] Eschbach and colleagues[159] elegantly demonstrated the importance of erythropoietin deficiency in experiments with sheep made uremic by subtotal nephrectomy. Plasma rich in erythropoietin was obtained from anemic sheep and infused into the uremic animals. The response was a brisk reticulocytosis and subsequent correction of anemia. Furthermore, the erythropoietic response in uremic sheep was not significantly different from the response in anemic sheep that were not uremic. This demonstrated that circulating inhibitors of erythropoiesis in uremia are not of great pathogenic importance.[159] Ultimately, the greatest proof of the primacy of erythropoietin deficiency in the pathogenesis of renal anemia has been the consistent success of treatment with rHuEPO.

Erythropoietin deficiency in CKD cannot be reliably demonstrated by measurement of serum erythropoietin levels. Serum levels may be the same or higher than levels of normal, nonanemic individuals. The deficiency is relative, not absolute; levels are insufficient for the degree of anemia present. [161] [162] [163] [164] [165] The adequacy of erythropoietin production in response to anemia appears to decline in rough proportion to the degree of reduced nephron mass. [166] [167] [168] Radtke and co-workers[168] measured serum erythropoietin levels in 135 patients with kidney disease and 59 normal subjects. At all levels of renal insufficiency, serum erythropoietin levels were found to be elevated compared with those in patients with normal renal function ( Fig. 49-6 ). However, the relationship between Hgb and serum erythropoietin level depended on the severity of renal failure. Among subjects with mild to moderate renal insufficiency, the investigators found a normal negative correlation; lower Hgb concentrations were associated with higher levels of serum erythropoietin. However, among patients with creatinine clearance less than 40 mL/min, mean serum erythropoietin levels were severely depressed and not correlated with the degree of anemia present. In these patients, there was a positive correlation between creatinine clearance and serum erythropoietin concentration, indicating a parallel loss of renal excretory and endocrine function.[168] Fehr and associates[169] recently studied 395 patients undergoing coronary angiography, 84% of whom had reduced levels of creatinine clearance. Similar to the findings of Ratke and colleagues,[168] serum erythropoietin levels were higher in patients with lower levels of Hgb, except when creatinine clearance was less than 40 mL/min. In this group with severe renal insufficiency, the concentration of erythropoietin was dissociated from Hgb.[169] Interestingly, despite the severely diminished erythropoietin response with advanced renal insufficiency, some small degree of sustained feedback remains. In the 6 months prior to starting dialysis, Radtke and colleagues[168] found that as anemia worsened, serum erythropoietin levels increased, and in the 6 months after starting dialysis, the opposite occurred. This continued response to anemia in advanced kidney disease was also demonstrated by Walle and co-workers,[170] who found that serum erythropoietin levels increased after hemorrhage and declined after blood transfusion in dialysis patients. Taken together, the literature indicates that patients with CKD (1) generally have higher levels of serum erythropoietin than patients without kidney disease, (2) mean serum erythropoietin levels increase with worsening anemia in mild to moderate renal insufficiency (although to an insufficient degree), (3) mean serum erythropoietin levels become more a function of creatinine clearance than Hgb concentration when creatinine clearance is less than 40 mL/min, and (4) even with severe renal insufficiency, some degree of responsiveness to lower Hgb is retained, although blunted in magnitude. The pronounced breakdown of erythropoietin production in response to anemia when creatinine clearance is less than 40 mL/min fits well with the observation (discussed previously) that clinically relevant anemia becomes common only with severe renal insufficiency. [23] [24]

FIGURE 49-6  135 patients with kidney disease and 59 healthy subjects were studied. Serum erythropoietin (EPO) levels in patients with kidney disease were generally higher than in normal subjects. With decreasing levels of creatinine clearance in the range of 20-90 ml/min, mean serum erythropoietin concentration increased as mean hematocrit (Hct) decreased. With severe renal insufficiency (creatinine clearance < 20 ml/min), serum erythropoietin levels were markedly decreased for the degree of anemia present.  (Adapted from Radtke HW, Claussner A, Erbes PM, et al: Serum erythropoietin concentration in chronic renal failure: Relationship to degree of anemia and excretory renal function. Blood 54(4):877–884, 1979.)



In addition to the hypoproliferative anemia caused by relative erythropoietin deficiency, patients with CKD may also have shortened erythrocyte life owing to low-grade hemolysis[171] and other factors. Red cell half-life in CKD, as measured by 51chromium labeling, has varied widely between studies. Erslev and Besarab[156] found that many patients had significantly reduced erythrocyte circulating half-life, but more than half of patients actually had normal red cell survival (106–122 days). Others have found significantly reduced survival, with half-life as low as 22 days.[172] The diminution of erythrocyte survival in CKD may be partially reversible by treatment with rHuEPO. [173] [174] Polenakovich and Sikole[173] studied 40 chronic hemodialysis patients. Prior to initiation of rHuEPO treatment, the mean erythrocyte half-life was 23.3 ± 2.6 days. After 12 months of treatment with rHuEPO, the mean erythrocyte half-life increased slightly to 27.2 ± 4.1 days, and after discontinuing rHuEPO for 12 months, the erythrocyte half-life decreased again to 22.1 ± 3.6 days.[173] Increased erythrocyte antioxidant content after rHuEPO treatment may contribute to improved cellular survival. [175] [176]

The etiology of impaired red cell survival in CKD has not been fully elucidated. Erythrocytes from renal failure patients have nearly normal survival after being transfused into nonuremic recipients.[176] Furthermore, red cell survival significantly improves with intensive dialysis treatment.[156] These observations suggest the possibility of a circulating uremic toxin. There is evidence that lipid peroxidation of the red cell membrane with premature clearance of cells by the reticuloendothelial system may play a role. [178] [179] [180] Because reduced erythrocyte survival has been postulated to result from circulating uremic toxins, Ly and colleagues[179a] recently analyzed the impact of more effective modern dialysis modalities. Twenty-two patients with ESRD treated with three forms of dialysis—high-efficiency thrice-weekly hemodialysis, nocturnal, and short daily hemodialysis—were studied. The primary finding was that erythrocyte survival was markedly decreased to 14.5 to 17.1 days and was not significantly different between the three dialysis modalities. An additional contributor during rHuEPO therapy may be a novel process termed neocytolysis, which is selective hemolysis of young circulating erythrocytes during periods of erythropoietin withdrawal.[180] Rice and co-workers[181] observed this phenomenon in four of five dialysis patients studied. When rHuEPO treatment was withheld, survival of red cells during their first 9 days in circulation was significantly reduced. These investigators subsequently demonstrated that splenic endothelial cells may be the site of detecting young erythrocytes and of initiating neocytolysis during erythropoietin withdrawal.[182]

Consequences of Anemia in Chronic Kidney Disease

Impact of Anemia on Quality of Life

Because anemia results in reduced delivery of oxygen to the body's tissues, it results in symptoms such as fatigue, dyspnea, and reduced mental acuity that degrade the individual's overall experience and quality of life. In fact, this may be the greatest single adverse effect of anemia. The importance of quality of life as a dimension of health is declared in the constitution of the World Health Organization, “health is a state of complete physical, mental and social well-being and not merely the absence of disease or infirmity.”[183]

Whereas quality of life is a vitally important aspect of health, it is one that is less objective and difficult to quantify than many other health outcomes. Therefore, a number of survey tools have been developed to permit more rigorous investigation of the various components that make up quality of life. These instruments may be broadly divided into nonspecific tools that may be useful in a variety of disease states as well as in health and tools that were designed for specific diseases such as kidney disease.[184] Examples of the nonspecific tools include the Karnofsky Scale,[185] the Sickness Impact Profile,[186] and the Medical Outcomes Study Short-Form Health Survey (SF-36).[187] The most widely used instrument designed specifically for kidney disease is the KDQOL (Kidney Disease Quality of Life survey).[188] This questionnaire includes scales examining the effect of kidney disease on daily life, the burden of kidney disease, symptoms/problems, cognitive function, sexual function, work status, sleep, and social interaction. [190] [191] Other kidney disease instruments include the Kidney Disease Questionnaire[190] and the EuroQol-5D.[191]

Quality of life in CKD has been most extensively studied among hemodialysis patients. Moreno and associates[192] administered the Karnofsky Scale and Sickness Impact Profile to 1013 hemodialysis patients and found 26% to 31% had severely diminished results.[192] Important aspects of function were impaired, including work, recreation, home management, sleep, and rest. In this study, a number of clinical characteristics were related to quality of life including Hgb; diminished quality of life was significantly associated with lower Hgb levels.

A number of studies have evaluated the effect of rHuEPO treatment on quality of life. These studies are discussed in greater detail in the chapter on erythropoietin treatment. Interventional studies have demonstrated a relatively linear improvement in quality of life results with higher attained Hgb levels.[11] Delano[193] studied 37 hemodialysis patients after treatment with rHuEPO. The mean Hct increased from 19.8% before therapy to 31.5% after therapy. Sense of well-being improved in 84% of patients. Other improvements included appetite (81%), sexual function (62%), socializing (70%), and sleep (68%). In one of the larger studies, Moreno and colleagues[194] treated 115 anemic hemodialysis patients with rHuEPO for 6 months, resulting in an increase in mean Hct from 30.9% to 38.4%. Significant improvements were found in mean physical dimension (sickness impact profile [SIP]), psychosocial dimension, global SIP, and mean Karnofsky scale score.[194] Among randomized controlled trials, several have been published in which the sample size was greater than 100 subjects. The Canadian Erythropoietin Study Group randomized 118 anemic hemodialysis patients to treatment with placebo or low-dose or high-dose rHuEPO. After 6 months of study, patients in the placebo group had a mean Hgb of 7.4 g/dL, whereas patients treated with rHuEPO had an increase in Hgb of approximately 3 to 4 g/dL. Treated patients had significant reduction in fatigue and improvement in several other facets of quality of life, although other quality of life measures such as the 6-minute walk test failed to improve. [197] [198] Besarab and co-workers[197] randomized 1233 hemodialysis patients with cardiac disease to treatment with rHuEPO to a target Hct of 30% or 42%. Patient's SF-36 physical function improved as a function of higher Hct, an increase of 0.6 for every 1.0% increase in Hct.[197] Furuland and associates[198] studied 416 anemic kidney disease patients (predialysis, hemodialysis, and peritoneal dialysis), randomized to rHuEPO treatment to two Hgb levels, 9 to 12 g/dL or 13.5 to 16 g/dL. A subset of 253 patients had a kidney disease questionnaire administered. At study week 48, there were significant improvements in some but not all quality of life measures including fatigue, depression, and physical symptoms.[198] Parfrey and colleagues[199] studied 324 anemic, incident hemodialysis patients randomized to rHuEPO treatment to achieve a Hgb level of 9.5 to 11 g/dL or 13.5 to 14.5 g/dL. The mean SF-36 vitality score was found to significantly improve in the higher Hgb group in most study weeks. However, there were no significant differences between the groups in the KDQOL quality of social interaction score or the Functional Assessment of Chronic Illness Therapy (FACIT)-fatigue score.[199] Among CKD stage 3-4 patients, Roger and colleagues[200] studied 155 subjects with initial Hgb less than 13 g/dL. Patients were randomized to rHuEPO treatment as necessary to achieve a target Hgb level of 9 to 10 g/dL or 12 to 13 g/dL. Quality of life was studied by SF-36 Health Survey and the Renal Quality of Life Profile. There were no significant differences at study conclusion in physical health score, mental health score, or the Renal Quality of Life Profile total score.[200]

One difficulty in analyzing the effect of anemia on quality of life is the problem of translating findings of quality of life surveys into some quantitative measure of the true impact on patients' experience. For example, in the Furuland and co-workers study,[198] the baseline mean kidney disease questionnaire score for fatigue was 4.67. After 48 weeks of rHuEPO treatment, with partial anemia correction, the mean score increased by 0.16 to 4.83. Whereas any reduction in fatigue is desirable, it is unclear what a 3% to 4% increase in a survey score means to a patient in terms of functional abilities. In addition, reporting on quality of life in rHuEPO treatment studies has been inconsistent and suboptimal. Some studies have used unvalidated scales, quality of life measures were often not prespecified outcomes, and reporting has tended to focus on only positive outcomes.[201]

Impact of Anemia on Mortality Risk

It is certainly plausible that anemia in CKD could increase the risk of death. Anemia causes reduced delivery of oxygen to the body's tissues and organs. [204] [205] [206] The resulting tissue hypoxia could adversely affect the structure and function of vital organs. This could be particularly true among patients with preexisting cardiac or vascular disease, in which tissue hypoxia might be poorly tolerated. In addition, anemia results in a variety of compensatory mechanisms including increased heart rate and cardiac stroke volume[205] and reduced systemic vascular resistance. The immediate effect is to maintain tissue oxygen delivery, but chronic activation could be maladap-tive, leading to development of left ventricular hypertro-phy (LVH), [111] [208] which could secondarily increase the risk for death. [209] [210]

The relationship of anemia to mortality risk has been evaluated through both observational and interventional studies. Neither approach is entirely satisfying in its ability to clearly define the relationship. Observational studies lead to inconclusive results because of the indirect and incomplete nature of the databases studied and the important muddling effect of the multiple confounders. The latter point is particularly difficult because patients who are sicker, and therefore at greater risk for poor outcomes, tend to be anemic and have lower Hgb levels. Therefore, it would be expected that lower levels of Hgb would be associated with increased mortality risk. However, to establish a causal role for anemia requires the ability to completely adjust for the confounding effects of health status, comorbidity, inflammation, and nutrition. Even the most sophisticated analyses struggle to overcome the intrinsic intertwining of anemia with other processes linked to outcomes. Furthermore, it is likely that there are a number of unknown confounders. As a result, the published observational studies are useful for describing the relationship between anemia and mortality risk, but in no way do they establish causality. Several of the key studies are reviewed later. In the chapter on treatment with rHuEPO, interventional studies, including those targeting full normalization of Hgb, are reviewed.

Ma and colleagues[209] studied 96,369 prevalent hemodialysis patients, with data obtained from Medicare claims. Baseline Hct was found to be significantly associated with subsequent mortality risk. Compared with a reference Hct range of 30% to 33%, patients with Hct 27% to 30% had a relative risk of 1.12; with Hct less than 27%, the relative risk was 1.33. Among patients with Hct 33% to 36%, there was relative risk of 0.96 compared with the reference range.[209] Locatelli and associates[210] reported on results from the Dialysis Outcomes and Practice Patterns Study (DOPPS), involving 4951 hemodialysis patients. The data source was clinical records, and data elements were predefined to ensure capture of many relevant covariates. The primary finding was that for every 1 g/dL increase in Hgb, there was a 4% reduction in relative risk for mortality. Levin and co-workers[211] studied the relationship between Hgb and mortality among patients with earlier stages of CKD. For each 1 g/dL increase in Hgb, a relative risk for mortality of 0.754 was noted. Furthermore, patients who received consistent rHuEPO replacement treat ment in the 2 years before initiating dialysis had reduced cardiac and overall mortality risk. Sandgren and colleagues[212] studied more than 1 million unselected Medicare beneficiaries and evaluated mortality risk over a 2-year period. Compared with individuals without comorbid conditions, the presence of CKD led to a 100% increased risk of death. The addition of anemia to CKD led to a further 270% increase in 2-year mortality risk.

Impact of Anemia on Cardiac Health

Cardiac disease has a grave impact on patients with kidney disease, reducing quality of life and increasing risk for hospitalizations and death. Among hemodialysis patients, mortality risk due to cardiovascular disease is more than 15 times greater than in the normal population.[213] Approximately 50% of deaths in CKD are related to cardiovascular disease, owing to congestive heart failure (CHF), acute myocardial infarction, and sudden cardiac death.[214]Indeed, patients with CKD are far more likely to die of cardiac events than to progress to ESRD.[215] This is true even in moderate to severe CKD, National Kidney Foundation stages 3 to 4.[215] The outcome of cardiac events is particularly devastating in CKD patients, after myocardial infarction the 1- and 5-year mortality are 59% and 90%, respectively.[216] The great burden of cardiac disease in CKD can be partially explained by the fact that traditional risk factors for atherosclerosis and cardiovascular disease tend to overlap with risk factors for kidney disease.[213] But, in addition, it is plausible that anemia, a common complication in CKD, may play a key role in incrementing risk.

Anemia in CKD results in chronic changes in the cardiovascular system. Part of the body's compensation for anemia is a high cardiac output and vasodilated state, which partially mitigates the effect of reduced oxygen carriage by the bloodstream. Related neurohormonal changes include increased plasma norepinephrine, renin activity, aldosterone, growth hormone, and atrial natriuretic peptide, all of which may be stimulated by anemia-induced vasodilatation and reduction in blood pressure.[217] Reduced systemic vascular resistance, owing to diminished arteriolar tone and blood viscosity, improves tissue perfusion. Increased endothelial production of nitric oxide appears to contribute to the arteriolar dilatation. [220] [221] Augmented venous return to the heart, increased sympathetic tone, and other factors contribute to increased cardiac stroke volume and heart rate, which combine to chronically increase cardiac output (Fig. 49-7 ). [222] [223] In the short term, these compensatory processes diminish the effect of reduced oxygen carriage by the circulation, but in the long term, maladaptive cardiac changes may occur.

FIGURE 49-7  Anemia triggers a multitude of processes designed to compensate for reduced oxygen carriage and tissue hypoxia. Chronic elevation of cardiac output may be maladaptive, increasing cardiac work and resulting in left ventricular hypertrophy and increased risk for cardiovascular events.


LVH is the cardiac abnormality most often found in association with chronic anemia. It is readily diagnosed by characteristic echocardiographic findings,[222] with left ventricular mass index greater than 134 and 110 g/m2 in men and women, respectively.[223] It is a particularly important finding in that it is a strong independent predictor of mortality risk. [226] [227] [228] Silberberg and co-workers[227] studied the relationship of anemia to LVH in a group of 78 hemodialysis patients, prior to the era of routine rHuEPO treatment. There was a strong association between worsening anemia and LVH risk, with a correlation coefficient of -0.81. In the lowest quartile of Hgb, the mean left ventricular mass index was 158 g/m2 compared with 120 g/m2 among patients in the highest Hgb quartile.[227] The relationship between anemia and cardiac disease was studied in earlier stages of CKD by Levin and co-workers.[228] Echocardiograms were performed in 175 patients attending a renal insufficiency clinic. LVH was found to be present in 38.9% of patients. The prevalence of LVH progressively increased with declining levels of renal function; 26.7% of patients with creatinine clearance (CrCl) greater than 50 mL/min, 30.8% with CrCl 25 to 49 mL/min, and 45.2% with CrCl less than 25 mL/min. Each 1 g/dL decrease in Hgb was associated with a 6% increase in risk for LVH. Furthermore, these investigators performed two echocardiograms 1 year apart on 246 patients with early stages of CKD to determine factors responsible for subsequent worsening of LVH. Worsening anemia proved to be an important predictor, with Hgb decreasing 0.85 g/dL in patients with ventricular growth compared with a decrease of 0.11 g/dL among patients with stable LVH.[229] Rigatto and colleagues[230] studied LVH in 416 renal transplant patients. Electrocardiographically demonstrated LVH was found to be present in 14% of patients (note the lower sensitivity for the diagnosis compared with that of echocardiography), and lower levels of Hgb were found to be an independent predictor of risk.[230] A substudy of the Atherosclerosis Risk in Communities (ARIC) Study examined left ventricular morphology in African Americans. Echocardiograms were performed in 1968 subjects 6 to 9 years after initial Hgb and creatinine measurements. Lower Hgb levels were found to be strongly associated with left ventricular end-diastolic diameter and weakly associated with the presence of LVH.[231] Ayus and associates[232]studied Spanish patients with severe renal insufficiency and found 68.3% had LVH. The prevalence of LVH was 87.5% among anemic patients, and 55.7% among nonanemic subjects. The adjusted odds ratio for LVH was 0.69 g/dL of Hgb.[232] Foley and co-workers[224] studied cardiac structure and function in 433 new hemodialysis patients during the rHuEPO treatment era. Echocardiography was performed a mean of 3.3 months after the initiation of hemodialysis. Most patients had abnormal hearts, 73.9% had LVH. In 44.3%, the hypertrophy was concentric; another 29.6% had eccentric hypertrophy. Survival of patients with LVH was significantly reduced compared with those with normal left ventricular mass. In contrast to other studies, this study did not find that lower Hgb was associated with LVH.

Taken together, this literature indicates a fairly consistent association between anemia and LVH. The association spans the spectrum of CKD from relatively early stages through ESRD and includes patients with renal transplants. The association seems to hold for patients of different ethnic and racial groups. Because the literature discussed is entirely composed of observational studies, it is not possible to de-termine whether the relationship is causal. Interventional studies aimed at treatment of anemia with assessment of the impact on left ventricular mass are discussed in the chapter on rHuEPO treatment. However, it should be noted here that these studies have had mixed results. Smaller studies with correction of severe anemia have demonstrated at least partial regression of LVH. [235] [236] [237] [238] [239] In contrast, a series of larger randomized trials of rHuEPO treatment for milder degrees of anemia have not found improvement in LVH. [240] [241] [242] [243]

The discrepancy in findings between observational and interventional studies raises fundamental questions regarding the relationship between anemia and LVH. A possible explanation is that the relationship may not be causal; anemia could be a marker for LVH without contributing to its development. This would seem unlikely because so many of the adaptive responses to anemia contribute to chronically increased cardiac workload, the typical root cause of LVH. An alternative explanation would be that the boundaries of therapeutic benefit could be limited. If the greatest impact of anemia on left ventricular mass were at very low levels of Hgb, then treatment studies targeting more severe degrees of anemia would be more likely to demonstrate benefit. In fact, as discussed previously, this has generally been the case. Finally, other effects of rHuEPO treatment, independent of raising Hgb level, could interfere with a potential beneficial effect on left ventricular mass. For example, if blood pressure was increased by treatment, then the benefit of raising Hgb might be obscured.

There have been several limited investigations into the effect of anemia on ischemic cardiac disease. For example, Wizemann and colleagues[242] studied eight hemodialysis patients with coronary artery disease and Hct less than 27%. Exercise stress testing was performed at baseline and then again after treatment with rHuEPO to a target Hct of 34%. During maximal exercise, treatment resulted in a reduction in ST segment depression from 2.1 mm to 0.4 mm and improved exercise duration. Similarly, Macdougall and co-workers[243] found reduced exercise-induced electrocardiographic changes of ischemia in seven of eight patients after treatment with rHuEPO.[243] Hase and associates[244] studied nine hemodialysis patients with coronary artery disease treated with rHuEPO to increase the mean Hgb from 7.9 g/dL to 10.4 g/dL. Maximal exercise-induced ST segment depression decreased after treatment from 1.3 to 0.5 mm.[244]

Anemia appears to play an important role in CHF. This is particularly relevant to nephrologists because of the large number of patients who have coexisting CHF and CKD. McClellan and co-workers[245] found a mean GFR of 55.7 mL/min among 627 patients admitted to hospitals with decompensated CHF. The proportion of patients with CKD was 60.4% ( Fig. 49-8 ). Anemia is a common finding in patients with CHF and CKD, with prevalence, defined as Hgb less than 12 g/dL, of approximately 50%.[246] To some extent, the anemia may represent dilution on the basis of volume expansion. However, Mancini and co-workers[247] found 60% of anemic CHF patients to have truly reduced packed red cell mass.[247] Anemia is closely associated with prognosis in CHF. Al-Ahmad and colleagues[248] performed a post hoc analysis of the Studies of Left Ventricular Dysfunction (SOLVD) database and found that each decrease of 1% in Hct was associated with a 2.7% increase in risk for mortality.[248] Furthermore, the presence of anemia in CHF is associated with risk for rehospitalization and more rapid progression of associated CKD. Finally, studies of treatment with rHuEPO in patients with coexistent CHF and CKD demonstrate improved quality of life and a reduced number of hospitalizations. [152] [251] In the study noted previously, Mancini and co-workers[247] randomized 26 patients with CHF to treatment with epoetin alfa or placebo for 3 months. Treatment resulted in significant improvement in several measures of physical function including exercise parameters and physiologic markers.

FIGURE 49-8  Among 627 patients admitted to a hospital with decompensated congestive heart failure, chronic kidney disease was found to coexist in 60.4%. The estimated GFR at hospital admission was abnormal in most patients, with more than 80% having eGFR less than 90%.  (Adapted from McClellan WM, Langston RD, Presley R: Medicare patients with cardiovascular disease have a high prevalence of chronic kidney disease and a high rate of progression to end-stage renal disease. J Am Soc Nephrol 15:1912–1919, 2004.)



Other Effects of Anemia in Chronic Kidney Disease

Anemia and its direct consequence, reduced oxygen carriage and delivery, may have other detrimental effects in patients with CKD. For example, worsening anemia could potentially accelerate the progression of kidney disease by depriving diseased kidneys of oxygen. It has been difficult to directly study the relationship without excessive confounding because of the fundamental intertwining of diminished renal function with worsening anemia. A post hoc analysis of the RENAAL (Reduction of Endpoints in NIDDM with the Angiotensin II Antagonist Losartan) trial was reported recently by Mohanram and associates.[250] Among 1513 subjects with type 2 diabetes mellitus, initial Hgb was an important predictor of renal outcome, including time to ESRD or doubling of serum creatinine. The risk was increased by 11% for every 1 g/dL decrease in Hgb concentration.[250] Occasional rHuEPO treatment studies have been adequately powered to allow exploratory analyses of the impact of treatment on renal disease progression. In the study of Roger and colleagues[200] described previously, there was no difference in the rate of progression between the high and the low Hgb groups.[200] In contrast, Kuriyama and co-workers[251] randomized 73 patients with serum creatinine 2 to 4 mg/dL and Hct less than 30% to treatment with rHuEPO or to no specific anemia treatment. Patients in the treatment group had increase in Hct from 27% to 32.1%. The primary end point, doubling in serum creatinine, occurred in 84% of untreated subjects compared with 52% of those treated with rHuEPO.

Anemia in CKD may have other adverse effects as well. Hoen and colleagues[252] prospectively studied 988 hemodialysis patients and found anemia to be associated with an increased risk for bacteremia. For every 1 g/dL decrease in Hgb, there was a 30% increase in infection risk. In addition, anemia in advanced kidney disease has repeatedly been demonstrated to be associated with impaired hemostasis (see later). A number of studies have assessed the effects of anemia on brain and cognitive function. The results have consistently linked anemia to impaired function and rHuEPO treatment to measurable improvements. [255] [256] [257] [258] [259] [260] Pickett and associates[255] treated 20 hemodialysis patients with increased doses of rHuEPO to raise mean Hct from 31.6% to 42.8%. Electroencephalogram results were found to improve, as did other measures of neurocognitive function.


Excessive bleeding has long been recognized as an important complication of the uremic state. [261] [262] This was particularly true prior to the advent of dialysis and the availability of rHuEPO and, to a lesser degree, today. Events may be as minor as epistaxis, excessive bleeding with tooth brushing, or easy bruisability. More severe, clinically relevant bleeding episodes tend to occur with trauma or after invasive procedures, such as renal biopsy, rather than spontaneously.[261] Prior to the availability of routine dialysis, catastrophic gastrointestinal hemorrhage was the major cause of death with uremia.[260]

The circulatory system is a closed, high-pressure circuit, which when disrupted, could result in massive hemorrhage and death. The coagulation process is the host defense that leads to “sealing” of the injured area. This requires the coordinated activation of vascular endothelial cells, platelets, and circulating coagulation proteins. Normal function requires clotting to be rapid, controlled, and localized, resulting in effective hemostasis without inducing excessive thrombotic activity.

Coagulation begins with formation of the platelet plug. In an intact blood vessel, the endothelium resists binding and adherence by platelets. With vascular injury and disruption, the subendothelial vessel wall is exposed, providing a receptive surface for platelet attachment.[262] Platelet adhesion is mediated by the interaction of receptor glycoprotein 1β with vessel wall collagen. Other vessel wall proteins such as von Willebrand factor (vWF), fibronectin, thrombospondin-1, and laminins play important roles. Of these, vWF is particularly important for adhesion in the presence of increased sheer force.[263] Upon adherence, platelets secrete thromboxane A2 and adenosine diphosphate, initiating and amplifying a cascade of activation and aggregation of large numbers of platelets. Individual platelets undergo a dramatic change in shape as their cytoskeleton rearranges to yield a discoid form with multiple filopodial projections.[264] In the process, platelet surface glycoprotein IIb/IIIa transforms and binds to circulating fibrinogen.[265] The second key phase of blood coagulation is the formation of a fibrin clot that enmeshes, stabilizes, and strengthens the platelet plug. The clotting cascade is a series of sequentially amplifying enzymatic reactions involving circulating clotting factors. A key step is the activation of thrombin, which catalyzes the conversion of soluble fibrinogen into the cross-linked fibrin clot. The plasma clotting factors are produced in the liver, and factors II, VII, IX, and X are dependent on the availability of sufficient vitamin K for carboxylation of glutamic acid residues.[265a]Deficiencies in individual clotting factors or of Vitamin K result in a variety of perturbations of blood coagulation.

It has long been noted that bleeding in uremic patients occurs despite normal or elevated circulating levels of coagulation factors.[260] This observation suggested that platelet abnormalities were the primary cause of the bleeding diathesis. Whereas the number of circulating platelets is generally normal, the function of platelets is often impaired (thrombasthenia). Evidence for platelet dysfunction includes elevated bleeding time,[260] diminished in vitro response to adenosine diphosphate and epinephrine,[266] and reduced ristocetin-induced platelet agglutination.[267]

The most consistent abnormality in platelet function in uremia is impaired interaction of platelets with the vascular subendothelium.[260] As a result, platelet adhesion and aggregation are hindered. The cause of this dysfunction is incompletely understood and could be related to abnormalities of the vessel wall, platelets, or plasma constituents. As for the vessel wall, it appears that its function may be altered in uremia. In particular, it has been noted that endothelial production of nitric oxide, a powerful platelet inhibitor, is increased. [271] [272] [273] In uremic rats, treatment with a nitric oxide inhibitor partially restores platelet function.[271] Interestingly, guanidinosuccinic acid, long postulated to play a role in uremic platelet dysfunction, has recently been found to up-regulate nitric oxide production by the vascular endothelium.[272]

The platelet itself is intrinsically altered in uremia. For example, the content of serotonin and adenosine diphosphate is reduced in uremic platelet granules.[266] Secretion of mediators may also be impaired, although this may be a function of repeated activation during hemodialysis.[273] Platelet receptors that play a critical role in adhesion to the vessel wall and aggregation, GP1b and GPIIb-IIIa, are probably not significantly reduced in quantity in uremia.[274] However, interaction of the receptors with vessel wall proteins may be abnormal.[275] In particular, activation of GPIIb-IIIa to facilitate its adhesion to vWF may be impaired.[276] Finally, the platelet cytoskeleton may be altered, with diminished actin incorporation and suboptimal intracellular trafficking of molecules. [280] [281] [282]

Whereas the platelet itself is not entirely normal in uremia, it appears that a more important pathogenic factor in platelet dysfunction may be the effect that uremic plasma has on platelet responsiveness. Platelets from normal individuals develop impaired adhesive function on exposure to uremic plasma.[280] In contrast, platelets from uremic subjects regain some function on exposure to normal plasma.[280] Certain molecules with molecular weights that preclude adequate clearance with hemodialysis accumulate in uremia and may contribute to platelet dysfunction.[281] A variety of toxins including quinolinic acids and guanidine substances have been implicated.[260] In addition, a role for hyperparathy-roidism has been suggested. Benigni and colleagues[282] found parathyroid hormone to impair platelet aggregation induced by a variety of substances. The plasma content of the major adhesive proteins, vWF and fibrinogen, are normal in uremia. However, the multimeric structure of vWF may be altered, with a sporadic decrease in high-molecular-weight multimers.[283]

It is generally accepted that dialysis reduces uremic platelet dysfunction and the risk for bleeding. But dialysis does not completely eliminate the problem. Moreover, hemodialysis may induce a transient worsening in platelet function. Sloand and Sloand[284] measured a variety of indicators of platelet function immediately before and after treatments. Bleeding time was found to increase and ristocetin response was impaired after hemodialysis. By the day after treatment, these abnormalities were reversed. The investigators noted a transient decrease of platelet membrane expression of glycoprotein 1β after hemodialysis. Other potential detrimental consequences of hemodialysis might include the enervating effect of repeated platelet activation, [288] [289] removal of younger platelets with greater function, [290] [291] and impaired platelet function due to a secondary effect of activated leukocytes.[289]

Anemia is an important contributor to uremic platelet dysfunction.[290] During normal circulation, erythrocytes tend to force the flow of platelets radially, away from the center of flow and toward the endothelial surfaces. When vascular injury occurs, platelets are in closer apposition to the vessel wall, facilitating platelet adherence and activation by vessel wall constituents such as collagen. With anemia, more platelets circulate in the center of the vessel, further from endothelial surfaces, hindering efficient platelet activation.[290] In addition, anemia may contribute to platelet dysfunction because adenosine diphosphate release by erythrocytes normally stimulates platelet interaction with collagen. [294] [295] Treatment of anemia may help reverse platelet dysfunction, as both transfusion of blood [293] [296] and rHuEPO therapy[294] have been found to be beneficial.

The treatment of uremic patients experiencing bleeding episodes requires (1) an assessment of the severity of blood loss, (2) hemodynamic stabilization, (3) replacement of blood products as needed, (4) identification of the bleeding source and etiology, and (5) correction of platelet dysfunction and other factors contributing to the bleeding diathesis. The first four aspects are routine components of clinical care and are not discussed further here; the fifth extends from the previous discussion on the pathobiology of uremic bleeding. It should be clear, however, that the intensity of interventions to correct uremic platelet dysfunction hinges on the degree of bleeding severity.

The first aspect of treatment to correct uremic platelet dysfunction is provision of adequate dialysis. Initiation of dialysis will lead to some improvement in thrombasthenia and bleeding risk. [269] [298] Bleeding time is improved in approximately 67% of dialyzed patients.[266] Bleeding episodes appear to be less common and of reduced severity. No studies have fully elucidated the relative effectiveness of hemodialysis versus other dialytic modalities. If hemodialysis or hemofiltration is to be used, then anticoagulation must be minimized. The relationship of dose of dialysis to improvement of platelet function has not been well studied. It would be reasonable clinical practice to provide a dose of dialysis consistent with current guidelines.

Treatment of anemia with rHuEPO may be the most effective treatment of uremic platelet dysfunction (see earlier). Cases and associates[296] found that treatment with epoetin alfa, 40 U/kg intravenously resulted in improvement in several parameters of platelet function as the Hgb level rose.[296] Others have found the same salutary effect of rHuEPO treatment. [297] [300] The mechanism of improved platelet function related to erythropoietin treatment is probably most directly related to vessel flow phenomena (as discussed earlier). However, it is also possible that erythropoietin treatment itself may directly affect platelet function. Tassies and colleagues[298] found that platelet function improved in some patients very soon after erythropoietin treatment was initiated, before Hgb levels increased. The authors attributed this effect to an increase in young circulating forms of platelets, with improved functional characteristics. Other potential direct beneficial effects of rHuEPO include improved platelet intracellular calcium mobilization,[299] increased expression of GP1b, [262] [303] and repaired platelet signal transduction.[301]

Desmopressin (DDAVP) is a synthetic form of antidiuretic hormone that is often used to treat uremic bleeding. The drug has little vasopressor activity and is only rarely associated with induced hyponatremia. The mechanism of improved platelet function is not completely known, but enhanced release of larger vWF multimers by endothelial cells probably plays an important role. [305] [306] Other factors may include improved platelet aggregation on contact with collagen[302] and increased levels of platelet glycoprotein Ib/IX.[304] Intravenous infusion of 0.3 mcg/kg desmopressin (or 3.0 mcg/kg subcutaneously) results in improved bleeding time in 50% to 100% of treated patients, although with little response in others. [305] [308] The reason for the heterogeneity of response is unknown. When bleeding time improves, the response is brief, lasting only 4 to 8 hours. [309] [310] DDAVP may also be administered by the intranasal route, at a dose approximately tenfold greater than that given intravenously. [311] [312] Rydzewski and co-workers[309] administered the drug twice weekly intranasally at a dose of 2 mcg/kg to 17 uremic patients, finding an approximate 30% improvement in bleeding time.[309] Repeated administrations of DDAVP may result in a diminished response with development of tachyphylaxis. [313] [314] Other treatments for uremic bleeding include infusion of cryoprecipitate, a plasma product rich in vWF and fibrinogen. [315] [316] There is very little published evidence to support the use of cryoprecipitate, and response appears to be highly variable. In one study of five patients with active bleeding, only two had normalization of bleeding time and a favorable clinical outcome after treatment.[314] Cryoprecipitate use should be reserved for life-threatening bleeding due to risk for infectious complications. Estrogens improve platelet function in both men and women. [318] [319] Liu and colleagues[317] treated six uremic patients with 25 mg of oral conjugated estrogen (Premarin). In four of six subjects, bleeding time normalized after 5 days of treatment. After discontinuation of drug, bleeding time remained normal for 3 to 10 days. In the remaining two subjects, bleeding time was significantly shortened after a week of treatment.[317] After intravenous infusion, Livio and associates[318] found the beneficial effect of conjugated estrogens to begin early and last for up to 2 weeks.[318] The mechanism of action of estrogen treatment is not fully known, but it may be related to inhibition of vascular nitric oxide production.[319]


1. Bright R: Cases and observations, illustrative of renal disease accompanied with the secretion of albuminous urine.  Guy's Hosp Rep  1836; 1:338-379.

2. Christison R: On granular degeneration of the kidnies and its connexions with dropsy inflammations and other diseases,  Edinburgh: Black; 1839:63-74.

3. Cameron JS: Towards the millennium: A history of renal anaemia and the optimal use of epoetin.  Nephrol Dial Transplant  1999; 14(suppl 2):10-21.

4. Brazier J, Cooper N, Maloney Jr JV, Buckberg G: The adequacy of myocardial oxygen delivery in acute normovolemic anemia.  Surgery  1974; 75(4):508-516.

5. Cain SM: Oxygen delivery and uptake in dogs during anemic and hypoxic hypoxia.  J Appl Physiol  1977; 42(2):228-234.

6. Malmberg PO, Woodson RD: Effect of anemia on oxygen transport in hemorrhagic shock.  J Appl Physiol  1979; 47(4):882-888.

7. Tsang CW, Lazarus R, Smith W, et al: Hematological indices in an older population sample: Derivation of healthy reference values.  Clin Chem  1998; 44:96-101.

8. Nilsson-Ehle H, Jagenburg R, Landahl S, Svanborg AL: Blood haemoglobin values in the elderly: Implications for reverence intervals from age 70 to 88.  Eur J Haematol  2000; 65:297-305.

9. Beutler E, West C: Hematologic differences between African-Americans and whites: The roles of iron deficiency and alpha-thalassemia on hemoglobin levels and mean corpuscular volume.  Blood  2005; 106:740-745.

10. Guralnik JM, Eisenstardt RS, Ferrucci L, et al: Prevalence of anemia in persons 65 years and older in the United States: Evidence for a high rate of unexplained anemia.  Blood  2004; 104:2263-2268.

11. National Kidney Foundation: Kidney disease outcome quality initiative anemia guidelines.  Am J Kidney Dis  2006; 47:516-585.

12. Pugh LG: Haemoglobin levels on the British Himalayan expeditions to Cho Oyu in 1952 and Everest in 1953.  J Physiol  1954; 126(2):38P-39P.

13. Pugh LG: Blood volume and haemoglobin concentration at altitudes above 18,000 ft. (5500 m).  J Physiol  1964; 170:344-354.

14. Dill DB, Terman JW, Hall FG: Hemoglobin at high altitude as related to age.  Clin Chem  1963; 12:710-716.

15. Okin JT, Treger A, Overy HR, et al: Hematologic response to medium altitude.  Rocky Mt Med J  1966; 63(1):44-47.

16. Miao G, Xinping L, Haiyan F, et al: Normal reference value of hemoglobin of middleaged women and altitude.  Yale J Biol Med  2004; 77(5-6):117-123.

17. Aste-salazar H, Hurtado A: The affinity of hemoglobin for oxygen at sea level and high altitudes.  Am J Physiol  1944; 142:733-743.

18. Hebbel RP, Eaton JW, Kronenberg RS, et al: Human llamas: Adaptation to altitude in subjects with high hemoglobin oxygen affinity.  J Clin Invest  1978; 62(3):593-600.

19. Eschbach JJ, Funk D, Adamson J, et al: Erythropoiesis in patients with renal failure undergoing chronic dialysis.  N Engl J Med  1967; 276:653-688.

20. Rambach WA, Kurtides E, Alt HL, Del Greco F: Azotemic anemia and the effect of hemodialysis.  Trans Am Soc Artif Intern Organs  1963; 9:57-61.

21. Korbet SM: Comparison of hemodialysis and peritoneal dialysis in the management of anemia related to chronic renal disease.  Semin Nephrol  1989; 9(1 suppl 1):9-15.

22. Hsu C, Bates D, Kuperman G, Curhan G: Relationship between hematocrit and renal function in men and women.  Kidney Int  2001; 59:725-731.

23. Hsu C, McCulloch C, Curhan G: Epidemiology of anemia associated with chronic renal insufficiency among adults in the United States: Results from the Third National Health and Nutrition Examination Survey.  J Am Soc Nephrol  2002; 13:504-510.

24. Astor B, Muntner P, Levin A, et al: Association of kidney function with anemia.  Arch Intern Med  2002; 162:1401-1408.

25. U.S. Renal Data System: USRDS 2004 Annual Data Report,  Bethesda, MD, National Institutes of Health, National Institute of Diabetes and Digestive and Kidney Diseases, 2004.

26. El-Achkar TM, Ohmit SE, McCullough PA, et al: Higher prevalence of anemia with diabetes mellitus in moderate kidney insufficiency: The Kidney Early Evaluation Program.  Kidney Int  2005; 67:1483-1488.

27. Thomas MC, MacIsaac RJ, Tsalamandris C, et al: Anemia in patients with type 1 diabetes.  J Clin Endocrinol Metab  2004; 89(9):4359-4363.

28. Guralnik JM, Eisenstaedt RS, Ferrucci L, et al: Prevalence of anemia in persons 65 years and older in the United States: Evidence for a high rate of unexplained anemia.  Blood  2004; 104:2263-2268.

29. Ishimura E, Nishizawa Y, Okuno S, et al: Diabetes mellitus increases the severity of anemia in non-dialyzed patients with renal failure.  J Nephrol  1998; 11:83-86.

30. Bosman DR, Winkler AS, Marsden JT, et al: Anemia with erythropoietin deficiency occurs early in diabetic nephropathy.  Diabetes Care  2001; 24:495-499.

31. Thomas MC, MacIsaac RJ, Tsalamandris C, et al: Unrecognized anemia in patients with diabetes: A cross-sectional survey.  Diabetes Care  2003; 26:1164-1169.

32. Thomas MC, MacIsaac RJ, Tsalamandris C, et al: The burden of anaemia in type 2 diabetes and the role of nephropathy: A cross-sectional audit.  Nephrol Dial Transplant  2004; 19:1792-1797.

33. Symeonidis A, Kouraklis-Symeonidis A, Psiroyiannis A, et al: Inappropriately low erythropoietin response for the degree of anemia in patients with noninsulin-dependent diabetes mellitus.  Ann Hematol  2005; 85:79-85.

34. Ble A, Fink JC, Woodman RC, et al: Renal function, erythropoietin, and anemia of older persons: the InCHIANTI study.  Arch Intern Med  2005; 165(19):2222-2227.

35. Bert P: Sur la richesse en hemoglobine du sang des animaux vivant sur les hauts lieux.  C R Acad Sci Paris  1882; 94:805-807.

36. Viault F: Sur l'augmentation considerable du nombre des globules rouge dans le sang chez les inhabitants des hauts plateux l'Amerique du Sud.  C R Acad Sci Paris  1890; 111:917-918.

37. Carnot P, Deflandre C: Sur l'activité hémopoiétique du sérum au cours de la régénération du sang.  C R Acad Sci Paris  1906; 143:384-386.

38. Reissmann KR: Studies on the mechanism of erythropoietin stimulation in parabiotic rats during hypoxia.  Blood  1950; 5:347-380.

39. Erslev AJ: Humoral regulation of red cell production.  Blood  1953; 8:349-357.

40. Bonsdorff E, Jalavisto E: A humoral mechanism in anoxic erythrocytosis.  Acta Physiol Scand  1948; 16:150-170.

41. Jacobson LO, Goldwasser E, Fried W, Plzak L: Role of the kidney in erythropoiesis.  Nature  1957; 179:633-634.

42. Koury ST, Bondurant MS, Koury MJ: Localization of erythropoietin synthesizing cells in murine kidneys by in situ hybridization.  Blood  1988; 71:524-537.

43. Fisher JW, Koury S, Ducey T, Mendel S: Erythropoietin (Epo) production by interstitial cells of hypoxic monkey kidneys.  Br J Haematol  1996; 95:27-32.

44. Miyake T, Kung CK, Goldwasser E: Purification of human erythropoietin.  J Biol Chem  1977; 252:5558-5564.

45. Lin FK, Suggs S, Lin CH, et al: Cloning and expression of the human erythropoietin gene.  Proc Natl Acad Sci U S A  1985; 82:7580-7584.

46. Jacobs K, Shoemaker C, Rudersdorf R, et al: Isolation and characterization of genomic and cDNA clones of human erythropoietin.  Nature  1985; 313:806-810.

47. Wenger RH: Cellular adaptation to hypoxia: O2-sensing protein hydroxylases, hypoxia-inducible transcription factors, and O2-regulated gene expression.  FASEB J  2002; 16(10):1151-1162.

48. Wenger RH: Mammalian oxygen sensing, signalling and gene regulation.  J Exp Biol  2000; 203(pt 8):1253-1263.

49. Wang GL, Jiang BH, Rue EA, Semenza GL: Hypoxia-inducible factor 1 is a basic-helix-loop-helix-PAS heterodimer regulated by cellular O2 tension.  Proc Natl Acad Sci U S A  1995; 92(12):5510-5514.

50. Gradin K, McGuire J, Wenger RH, et al: Functional interference between hypoxia and dioxin signal transduction pathways: Competition for recruitment of the Arnt transcription factor.  Mol Cell Biol  1996; 16(10):5221-5231.

51. Wood SM, Gleadle JM, Pugh CW, et al: The role of the aryl hydrocarbon receptor nuclear translocator (ARNT) in hypoxic induction of gene expression. Studies in ARNT-deficient cells.  J Biol Chem  1996; 271(25):15117-15123.

52. Huang LE, Gu J, Schau M, Bunn HF: Regulation of hypoxia-inducible factor 1alpha is mediated by an O2-dependent degradation domain via the ubiquitin-proteasome pathway.  Proc Natl Acad Sci U S A  1998; 95(14):7987-7992.

53. Salceda S, Caro J: Hypoxia-inducible factor 1alpha (HIF-1alpha) protein is rapidly degraded by the ubiquitin-proteasome system under normoxic conditions. Its stabilization by hypoxia depends on redox-induced changes.  J Biol Chem  1997; 272(36):22642-22647.

54. Huang LE, Gu J, Schau M, Bunn HF: Regulation of hypoxia-inducible factor 1alpha is mediated by an O2-dependent degradation domain via the ubiquitin-proteasome pathway.  Proc Natl Acad Sci U S A  1998; 95(14):7987-7992.

55. Ohh M, Park CW, Ivan M, et al: Ubiquitination of hypoxia-inducible factor requires direct binding to the beta-domain of the von Hippel-Lindau protein.  Nat Cell Biol  2000; 2(7):423-427.

56. Maxwell PH, Wiesener MS, Chang GW, et al: The tumour suppressor protein VHL targets hypoxia-inducible factors for oxygen-dependent proteolysis.  Nature  1999; 399(6733):271-275.

57. Fandrey J: Oxygen-dependent and tissue-specific regulation of erythropoietin gene expression.  Am J Physiol Regul Integr Comp Physiol  2004; 286(6):R977-R988.

58. Jewell UR, Kvietikova I, Scheid A, et al: Induction of HIF-1 in response to hypoxia is instantaneous.  FASEB J  2001; 15:1312-1314.

58a. Sowter HM, Raval RR, Moore JW, et al: Predominant role of hypoxia-inducible transcription factor (Hif)-1alpha versus Hif-2alpha in regulation of the transcriptional response to hypoxia.  Cancer Res  2003; 63(19):6130-6134.

59. Ryan HE, Lo J, Johnson RS: HIF-1 alpha is required for solid tumor formation and embryonic vascularization.  EMBO J  1998; 17(11):3005-3015.

60. Yu AY, Shimoda LA, Iyer NV, et al: Impaired physiological responses to chronic hypoxia in mice partially deficient for hypoxia-inducible factor 1alpha.  J Clin Invest  1999; 103(5):691-696.

61. Maxwell PH, Wiesener MS, Chang GW, et al: The tumour suppressor protein VHL targets hypoxia-inducible factors for oxygen-dependent proteolysis.  Nature  1999; 399:271-275.

62. Stebbins CE, Kaelin Jr WG, Pavletich NP: Structure of the VHL-ElonginC-ElonginB complex: implications for VHL tumor suppressor function.  Science  1999; 284:455-461.

63. Masson N, Willam C, Maxwell PH, et al: Independent function of two destruction domains in hypoxia-inducible factor-chains activated by prolyl hydroxylation.  EMBO J  2001; 20:5197-5206.

64. Ivan M, Kondo K, Yang H, et al: HIF targeted for VHL-mediated destruction by proline hydroxylation: implications for O2 sensing.  Science  2001; 292:464-468.

65. Jaakkola P, Mole DR, Tian YM, et al: Targeting of HIF- to the von Hippel-Lindau ubiquitylation complex by O2-regulated prolyl hydroxylation.  Science  2001; 292:468-472.

66. Bruick RK, McKnight SL: A conserved family of prolyl-4-hydroxylases that modify HIF.  Science  2001; 294:1337-1340.

67. Epstein AC, Gleadle JM, McNeill LA, et al: C. elegans EGL-9 and mammalian homologs define a family of dioxygenases that regulate HIF by prolyl hydroxylation.  Cell  2001; 107:43-54.

68. Fandrey J, Bunn HF: In vivo and in vitro regulation of erythropoietin mRNA: Measurement by competitive polymerase chain reaction.  Blood  1993; 81(3):617-623.

69. Schuster SJ, Badiavas EV, Costa-Giomi P, et al: Stimulation of erythropoietin gene transcription during hypoxia and cobalt exposure.  Blood  1989; 73(1):13-16.

70. Zhu H, Bunn HF: Oxygen sensing and signaling: Impact on the regulation of physiologically important genes.  Respir Physiol  1999; 115(2):239-247.

71. Semenza GL, Agani F, Booth G, et al: Structural and functional analysis of hypoxia-inducible factor 1.  Kidney Int  1997; 51(2):553-555.

72. Rosenberger C, Mandriota S, Jurgensen JS, et al: Expression of hypoxia-inducible factor-1alpha and -2alpha in hypoxic and ischemic rat kidneys.  J Am Soc Nephrol  2002; 13(7):1721-1732.

73. Rosenberger C, Heyman SN, Rosen S, et al: Up-regulation of HIF in experimental acute renal failure: Evidence for a protective transcriptional response to hypoxia.  Kidney Int  2005; 67(2):531-542.

74. Jelkmann W: Molecular biology of erythropoietin.  Intern Med  2004; 43(8):649-659.

75. Dordal MS, Wang FF, Goldwasser E: The role of carbohydrate in erythropoietin action.  Endocrinology  1985; 116(6):2293-2299.

76. Imai N, Kawamura A, Higuchi M, et al: Physicochemical and biological comparison of recombinant human erythropoietin with human urinary erythropoietin.  J Biochem (Tokyo)  1990; 107(3):352-359.

77. Sasaki H, Ochi N, Dell A, Fukuda M: Site-specific glycosylation of human recombinant erythropoietin: Analysis of glycopeptides or peptides at each glycosylation site by fast atom bombardment mass spectrometry.  Biochemistry  1988; 27(23):8618-8626.

78. Sasaki H, Bothner B, Dell A, Fukuda M: Carbohydrate structure of erythropoietin expressed in Chinese hamster ovary cells by a human erythropoietin cDNA.  J Biol Chem  1987; 262(25):12059-12076.

79. Rush RS, Derby PL, Smith DM, Merry C, et al: Microheterogeneity of erythropoietin carbohydrate structure.  Anal Chem  1995; 67(8):1442-1452.

80. Takeuchi M, Inoue N, Strickland TW, et al: Relationship between sugar chain structure and biological activity of recombinant human erythropoietin produced in Chinese hamster ovary cells.  Proc Natl Acad Sci U S A  1989; 86(20):7819-7822.

81. Misaizu T, Matsuki S, Strickland TW, et al: Role of antennary structure of N-linked sugar chains in renal handling of recombinant human erythropoietin.  Blood  1995; 86(11):4097-4104.

82. Takeuchi M, Takasaki S, Shimada M, Kobata A: Role of sugar chains in the in vitro biological activity of human erythropoietin produced in recombinant Chinese hamster ovary cells.  J Biol Chem  1990; 265(21):12127-12130.

83. Maxwell PH, Osmond MK, Pugh CW, et al: Identification of the renal erythropoietin-producing cells using transgenic mice.  Kidney Int  1993; 44(5):1149-1162.

84. Bachmann S, Le Hir M, Eckardt KU: Co-localization of erythropoietin mRNA and ecto-5′-nucleotidase immunoreactivity in peritubular cells of rat renal cortex indicates that fibroblasts produce erythropoietin.  J Histochem Cytochem  1993; 41(3):335-341.

85. Zucali JR, Stevens V, Mirand EA: In vitro production of erythropoietin by mouse fetal liver.  Blood  1975; 46(1):85-90.

86. Jelkmann W: The enigma of the metabolic fate of circulating erythropoietin (Epo) in view of the pharmacokinetics of the recombinant drugs rhEpo and NESP.  Eur J Haematol  2002; 69(5-6):265-274.

87. Jelkmann W: Molecular biology of erythropoietin.  Intern Med  2004; 43(8):649-659.

88. Cotes PM: Immunoreactive erythropoietin in serum. I. Evidence for the validity of the assay method and the physiological relevance of estimates.  Br J Haematol  1982; 50(3):427-438.

89. Al-Huniti NH, Widness JA, Schmidt RL, Veng-Pedersen P: Pharmacokinetic/pharmacodynamic analysis of paradoxal regulation of erythropoietin production in acute anemia.  J Pharmacol Exp Ther  2004; 310(1):202-208.

90. Maeda H, Hitomi Y, Hirata R, et al: The effect of phlebotomy on serum erythropoietin levels in normal healthy subjects.  Int J Hematol  1992; 55(2):111-115.

91. Goodnough LT, Price TH, Parvin CA: The endogenous erythropoietin response and the erythropoietic response to blood loss anemia: The effects of age and gender.  J Lab Clin Med  1995; 126(1):57-64.

92. Rege AB, Brookins J, Fisher JW: A radioimmunoassay for erythropoietin: serum levels in normal human subjects and patients with hemopoietic disorders.  J Lab Clin Med  1982; 100(6):829-843.

93. de Klerk G, Rosengarten PC, Vet RJ, Goudsmit R: Serum erythropoietin (EST) titers in anemia.  Blood  1981; 58(6):1164-1170.

94. Pavlovic-Kentera V, Milenkovic P, Ruvidic R, et al: Erythropoietin in aplastic anemia.  Blut  1979; 39(5):345-350.

95. Takeichi N, Umemura T, Nishimura J, et al: Regulation of erythropoietin and burst-promoting activity production in patients with aplastic anemia and iron deficiency anemia.  Acta Haematol  1988; 80(3):145-152.

96. Zaroulis CG, Hoffman BJ, Kourides IA: Serum concentrations of erythropoietin measured by radioimmunoassay in hematologic disorders and chronic renal failure.  Am J Hematol  1981; 11(1):85-92.

97. Kelley LL, Green WF, Hicks GG, et al: Apoptosis in erythroid progenitors deprived of erythropoietin occurs during the G1 and S phases of the cell cycle without growth arrest or stabilization of wild-type p53.  Mol Cell Biol  1994; 14(6):4183-4192.

98. Silva M, Grillot D, Benito A, et al: Erythropoietin can promote erythroid progenitor survival by repressing apoptosis through Bcl-XL and Bcl-2.  Blood  1996; 88(5):1576-1582.

99. Jelkmann W, Metzen E: Erythropoietin in the control of red cell production.  Ann Anat  1996; 178(5):391-403.

100. Smith BR: Regulation of hematopoiesis.  Yale J Biol Med  1990; 63(5):371-380.

101. Ratajczak J, Zhang Q, Pertusini E, et al: The role of insulin (INS) and insulin-like growth factor-I (IGF-I) in regulating human erythropoiesis. Studies in vitro under serum-free conditions—comparison to other cytokines and growth factors.  Leukemia  1998; 12:371-381.

102. Sawada K, Krantz SB, Dai CH, et al: Purification of human blood burst-forming units-erythroid and demonstration of the evolution of erythropoietin receptors.  J Cell Physiol  1990; 142(2):219-230.

103. Broudy VC, Lin N, Brice M, et al: Erythropoietin receptor characteristics on primary human erythroid cells.  Blood  1991; 77(12):2583-2590.

104. Testa NG: Structure and regulation of the erythroid system at the level of progenitor cells.  CRC Crit Rev Oncol Hematol  1989; 9:17.

105. Suzuki N, Suwabe N, Ohneda O, et al: Identification and characterization of 2 types of erythroid progenitors that express GATA-1 at distinct levels.  Blood  2003; 102(10):3575-3583.

106. Chiba T, Ikawa Y, Todokoro K: GATA-1 transactivates erythropoietin receptor gene, and erythropoietin receptor-mediated signals enhance GATA-1 gene expression.  Nucleic Acids Res  1991; 19(14):3843-3848.

107. Blacklock HA, Katz F, Michalevicz R, et al: A and B blood group antigen expression on mixed colony cells and erythroid precursors: relevance for human allogeneic bone marrow transplantation.  Br J Haematol  1984; 58(2):267-276.

108. Nakahata T, Okumura N: Cell surface antigen expression in human erythroid progenitors: Erythroid and megakaryocytic markers.  Leuk Lymphoma  1994; 13(5-6):401-409.

109. Fisher JW: Erythropoietin: Physiology and pharmacology update.  Exp Biol Med (Maywood)  2003; 228(1):1-14.

110. Sawyer ST, Penta K: Erythropoietin cell biology.  Hematol Oncol Clin North Am  1994; 8:895-911.

111. Muta K, Krantz SB: Apoptosis of human erythroid colony-forming cells is decreased by stem cell factor and insulin-like growth factor I as well as erythropoietin.  J Cell Physiol  1993; 156(2):264-271.

112. Nijhof W, de Haan G, Pietens J, Dontje B: Mechanistic options of erythropoietin-stimulated erythropoiesis.  Exp Hematol  1995; 23(4):369-375.

113. Koury MJ, Bondurant MC: The molecular mechanism of erythropoietin action.  Eur J Biochem  1992; 210(3):649-663.

114. De Maria R, Zeuner A, Eramo A, et al: Negative regulation of erythropoiesis by caspase-mediated cleavage of GATA-1.  Nature  1999; 401(6752):489-493.

115. Wickrema A, Krantz SB, Winkelmann JC, Bondurant MC: Differentiation and erythropoietin receptor gene expression in human erythroid progenitor cells.  Blood  1992; 80(8):1940-1949.

116. Adamson JW: Regulation of red blood cell production.  Am J Med  1996; 101(2A):4S-6S.

117. Jones SS, D'Andrea AD, Haines LL, Wong GG: Human erythropoietin receptor: Cloning, expression, and biologic characterization.  Blood  1990; 76(1):31-35.

118. Atkins HL, Broudy VC, Papayannopoulou T: Characterization of the structure of the erythropoietin receptor by ligand blotting.  Blood  1991; 77(12):2577-2582.

119. Kaczmarski RS, Mufti GJ: The cytokine receptor superfamily.  Blood Rev  1991; 5(3):193-203.

120. Watowich SS, Yoshimura A, Longmore GD, et al: Homodimerization and constitutive activation of the erythropoietin receptor.  Proc Natl Acad Sci U S A  1992; 89(6):2140-2144.

121. Jelkmann W, Metzen E: Erythropoietin in the control of red cell production.  Ann Anat  1996; 178(5):391-403.

122. Farrell F, Lee A: The erythropoietin receptor and its expression in tumor cells and other tissues.  Oncologist  2004; 9(suppl 5):18-30.

123. D'Andrea AD, Zon LI: Erythropoietin receptor. Subunit structure and activation.  J Clin Invest  1990; 86(3):681-687.

124. Youssoufian H, Longmore G, Neumann D, et al: Structure, function, and activation of the erythropoietin receptor.  Blood  1993; 81(9):2223-2236.

125. Kubatzky KK, Ruan W, Gurezka R, et al: Self assembly of the transmembrane domain promotes signal transduction through the erythropoietin receptor.  Curr Biol  2001; 11:110-115.

126. Constantinescu JN, Keren T, Socolovsky M, et al: Ligand-independent oligomerizatin of cell-surface erythropoietin receptor is mediated by the transmembrane domain.  Proc Natl Acad Sci U S A  2001; 98:4379-4384.

127. Watowich SS, Hilton DJ, Lodish HF: Activation and inhibition of erythropoietin receptor function: Role of receptor dimerization.  Mol Cell Biol  1994; 14(6):3535-3549.

128. Schneider H, Chaovapong W, Matthews DJ, et al: Homodimerization of erythropoietin receptor by a bivalent monoclonal antibody triggers cell proliferation and differentiation of erythroid precursors.  Blood  1997; 89(2):473-482.

129. Mulcahy L: The erythropoietin receptor.  Semin Oncol  2001; 28(2 suppl 8):19-23.

130. Remy I, Wilson IA, Michnick SW: Erythropoietin receptor activation by a ligand-induced conformation change.  Science  1999; 283(5404):990-993.

131. Livnah O, Stura EA, Middleton SA, et al: Crystallographic evidence for preformed dimers of erythropoietin receptor before ligand activation.  Science  1999; 283(5404):987-990.

132. Cheetham JC, Smith DM, Aoki KH, et al: NMR structure of human erythropoietin and a comparison with its receptor bound conformation.  Nat Struct Biol  1998; 5(10):861-866.

133. Witthuhn BA, Quelle FW, Silvennoinen O, et al: JAK2 associates with the erythropoietin receptor and is tyrosine phosphorylated and activated following stimulation with erythropoietin.  Cell  1993; 74(2):227-236.

134. Krempler A, Qi Y, Triplett AA, et al: Generation of a conditional knockout allele for the Janus kinase 2 (Jak2) gene in mice.  Genesis  2004; 40(1):52-57.

135. Tauchi T, Feng GS, Shen R, et al: Involvement of SH2-containing phosphotyrosine phosphatase Syp in erythropoietin receptor signal transduction pathways.  J Biol Chem  1995; 270(10):5631-5635.

136. Barber DL, Beattie BK, Mason JM, et al: A common epitope is shared by activated signal transducer and activator of transcription-5 (STAT5) and the phosphorylated erythropoietin receptor: Implications for the docking model of STAT activation.  Blood  2001; 97(8):2230-2237.

137. Pallard C, Fabrice G, Martine C, et al: Interleukin-3, erythropoietin, and prolactin activate a STAT5 line factor in lymphoid cells.  J Biol Chem  1995; 270:15942-15945.

138. Penta K, Sawyer ST: Erythropoietin induces the tyrosine phosphorylation nuclear translocation and DNA binding of STAT1 and STAT5 in erythroid cells.  J Biol Chem  1995; 270:31282-31287.

139. Damen JE, Wakao H, Miyajima A, et al: Tyrosine 343 in the erythropoietin-receptor positively regulates erythropoietin-induced cell proliferation and STAT5 activation.  EMBO J  1995; 14:5557-5568.

140. Klingmuller U, Bergelson S, Hsiao JG, Lodish HC: Multiple tyrosine residues in the cytosolic domain of the erythropoietin receptor promote activation of STAT5.  Proc Natl Acad Sci U S A  1996; 93:8324-8328.

141. Klingmuller U, Lorenz U, Cantley LC, et al: Specific recruitment of SH-PTP1 to the erythropoietin receptor causes activation of JAK2 and termination of proliferative signals.  Cell  1995; 80:729-738.

142. Shan R, Price JO, Guarde WA, et al: Distinct roles of JNKs/p38 MAP kinase and ERKs in apoptosis and survival of HCP-57 cells induced by withdrawal or addition of erythropoietin.  Blood  1999; 94:4067-4076.

143. Oda A, Sawada K, Druker BJ, et al: Erythropoietin induces tyrosine phosphorylation of Jak2, STAT5A, and STAT5B in primary cultured human erythroid precursors.  Blood  1998; 92(2):443-451.

144. Spivak JL, Pham T, Isaacs M, Hankins WD: Erythropoietin is both a mitogen and survival factor.  Blood  1991; 77:1228.

145. Koury MJ, Bondurant MC: Erythropoietin retards DNA breakdown and prevents programmed death in erythroid progenitor cells.  Science  1990; 248:378.

146. Verdier F, Walrafen P, Hubert N, et al: Proteasomes regulate the duration of erythropoietin receptor activation by controlling down-regulation of cell surface receptors.  J Biol Chem  2000; 275(24):18375-18381.

147. Obeid R, Kuhlmann M, Kirsch CM, Herrmann W: Cellular uptake of vitamin B12 in patients with chronic renal failure.  Nephron Clin Pract  2005; 99(2):c42-c48.

148. Tabernero Romo JM, Bondia Roman A, Corbacho Becerra L, et al: Plasma B12 vitamin and folic acid during chronic renal failure and hemodialysis.  Med Clin (Barc)  1979; 73(4):129-132.

149. Posner GL, Fink SM, Huded FV, et al: Endoscopic findings in chronic hemodialysis patients with upper gastrointestinal bleeding.  Am J Gastroenterol  1983; 78(11):720-721.

150. Obrador GT, Ruthazer R, Arora P, et al: Prevalence of and factors associated with suboptimal care before initiation of dialysis in the United States.  J Am Soc Nephrol  1999; 10(8):1793-1800.

151. Loge JP, Lange RD, Moore CV: Characterization of the anemia associated with chronic renal insufficiency.  Am J Med  1958; 24:4-18.

152. Paganini EP: Overview of anemia associated with chronic renal disease: Primary and secondary mechanisms.  Semin Nephrol  1989; 9(1 suppl 1):3-8.

153. Radtke HW, Rege AB, Lamarche B, et al: Identification of spermine as an inhibitor of erythropoiesis in patients with chronic renal failure.  J Clin Invest  1981; 67:1623-1629.

154. Massry SG: Is parathyroid hormone an uremic toxin?.  Nephron  1977; 19:125-130.

155. Fisher JW, Hatch FE, Roh BL, et al: Erythropoietin inhibitor in kidney extracts and plasma from anemic uremic human subjects.  Blood  1968; 31(4):440-452.

156. Erslev AJ, Besarab A: The rate and control of baseline red cell production in hematologically stable patients with uremia.  J Lab Clin Med  1995; 126:283-286.

157. Eschbach JW: The anemia of chronic renal failure: Pathophysiology and the effects of recombinant erythropoietin.  Kidney Int  1989; 35:134-148.

158. Eschbach JW, Detter JC, Adamson JW: Physiologic studies in normal and uremic sheep. II. Changes in erythropoiesis and oxygen transport.  Kidney Int  1980; 18(6):732-745.

159. Eschbach JW, Mladenovic J, Garcia JF, et al: The anemia of chronic renal failure in sheep. Response to erythropoietin-rich plasma in vivo.  J Clin Invest  1984; 74(2):434-441.

160. Erslev AJ, Wilson J, Caro J: Erythropoietin titers in anemic, nonuremic patients.  J Lab Clin Med  1987; 109(4):429-433.

161. Besarab A, Caro J, Jarrell BE, et al: Dynamics of erythropoiesis following renal transplantation.  Kidney Int  1987; 32(4):526-536.

162. Radtke HW, Frei U, Erbes PM, et al: Improving anemia by hemodialysis: Effect of serum erythropoietin.  Kidney Int  1980; 17(3):382-387.

163. Seguchi C, Shima T, Misaki M, et al: Serum erythropoietin concentrations and iron status in patients on chronic hemodialysis.  Clin Chem  1992; 38(2):199-203.

164. Teruel JL, Marcen R, Navarro JF, et al: Evolution of serum erythropoietin after androgen administration to hemodialysis patients: A prospective study.  Nephron  1995; 70(3):282-286.

165. Eschbach JW: Erythropoietin 1991—An overview.  Am J Kidney Dis  1991; 18(4 suppl 1):3-9.

166. Erslev AJ: Erythropoietin.  N Engl J Med  1991; 324(19):1339-1344.

167. McGonigle RJ, Wallin JD, Shadduck RK, Fisher JW: Erythropoietin deficiency and inhibition of erythropoiesis in renal insufficiency.  Kidney Int  1984; 25(2):437-444.

168. Radtke HW, Claussner A, Erbes PM, et al: Serum erythropoietin concentration in chronic renal failure: Relationship to degree of anemia and excretory renal function.  Blood  1979; 54(4):877-884.

169. Fehr T, Ammann P, Garzoni D, et al: Interpretation of erythropoietin levels in patients with various degrees of renal insufficiency and anemia.  Kidney Int  2004; 66(3):1206-1211.

170. Walle AJ, Wong GY, Clemons GK, et al: Erythropoietin-hematocrit feedback circuit in the anemia of end-stage renal disease.  Kidney Int  1987; 31(5):1205-1209.

171. Lerner R, Werner B, Asaba H, et al: Assessment of hemolysis in regular hemodialysis patients by measuring carbon monoxide production rate.  Clin Nephrol  1983; 20(5):239-243.

172. Schwartz AB, Kelch B, Terzian L, et al: One year of rHuEPO therapy prolongs RBC survival and may stabilize RBC membranes despite natural progression of chronic renal failure to uremia and need for dialysis.  ASAIO Trans  1990; 36(3):M691-M696.

173. Polenakovic M, Sikole A: Is erythropoietin a survival factor for red blood cells?.  J Am Soc Nephrol  1996; 7(8):1178-1182.

174. Cavdar C, Camsari T, Semin I, et al: Lipid peroxidation and antioxidant activity in chronic haemodialysis patients treated with recombinant human erythropoietin.  Scand J Urol Nephrol  1997; 31(4):371-375.

175. Boran M, Kucukaksu C, Balk M, Cetin S: Red cell lipid peroxidation and antioxidant system in haemodialysed patients: Influence of recombinant human erythropoietin (r-HuEPO) treatment.  Int Urol Nephrol  1998; 30(4):507-512.

176. Eschbach JW, Haley NR, Adamson JW: The anemia of chronic renal failure: Pathophysiology and effects of recombinant erythropoietin.  Contrib Nephrol  1990; 78:24-36.

177. Martino F, Lubrano R, Ruberto U, et al: Peroxidation of erythrocyte lipids and levels of vitamin E in red blood cells in children with chronic renal insufficiency.  Minerva Pediatr  1989; 41(6):315-317.

178. Usberti M, Gerardi G, Bufano G, et al: Effects of erythropoietin and vitamin E-modified membrane on plasma oxidative stress markers and anemia of hemodialyzed patients.  Am J Kidney Dis  2002; 40(3):590-599.

179. Miguel A, Miguel A, Linares M, et al: Evidence of an increased susceptibility to lipid peroxidation in red blood cells of chronic renal failure patients.  Nephron  1988; 50(1):64-65.

179a. Ly J, Marticorena R, Donnelly S: Red blood cell survival in chronic renal failure.  Am J Kidney Dis  2004; 44(4):715-719.

180. Alfrey CP, Rice L, Udden MM, Driscoll TB: Neocytolysis: Physiological down-regulator of red-cell mass.  Lancet  1997; 349(9062):1389-1390.

181. Rice L, Alfrey CP, Driscoll T, et al: Neocytolysis contributes to the anemia of renal disease.  Am J Kidney Dis  1999; 33(1):59-62.

182. Trial J, Rice L, Alfrey CP: Erythropoietin withdrawal alters interactions between young red blood cells, splenic endothelial cells, and macrophages: An in vitro model of neocytolysis.  J Investig Med  2001; 49(4):335-345.

183.   World Health Organization Constitution. URL—

184. Rettig RA, Sadler JH, Meyer KB, et al: Assessing health and quality of life outcomes in dialysis: A report on an Institute of Medicine workshop.  Am J Kidney Dis  1997; 30:140-155.

185. Karnofsky DA: The clinical evaluation of chemotherapeutic agents in cancer.   In: Macleod CM, ed. Evaluation of Chemotherapeutic Agents,  New York: Columbia University Press; 1949:191-205.

186. Bergner M, Bobbitt RA, Carter WB, Gilson BS: The sickness impact profile: Development and final revision of a health status measure.  Med Care  1981; 19(8):787-805.

187. Ware Jr JE, Sherbourne CD: The MOS 36-item short-form health survey (SF-36). I. Conceptual framework and item selection.  Med Care  1992; 30(6):473-483.

188. Hays RD, Kallich JD, Mapes DL, et al: Development of the kidney disease quality of life (KDQOL) instrument.  Qual Life Res  1994; 3(5):329-338.

189. Valderrabano F: Quality of life benefits of early anaemia treatment.  Nephrol Dial Transplant  2000; 15(suppl 3):23-28.

190. Laupacis A, Muirhead N, Keown P, Wong C: A disease-specific questionnaire for assessing quality of life in patients on hemodialysis.  Nephron  1992; 60(3):302-306.

191. Korevaar JC, Jansen MA, Merkus MP, et al: Quality of life in predialysis end-stage renal disease patients at the initiation of dialysis therapy. The NECOSAD Study Group.  Perit Dial Int  2000; 20:69-75.

192. Moreno F, Lopez Gomez JM, Sanz-Guajardo D, et al: Quality of life in dialysis patients. A Spanish multicentre study. Spanish Cooperative Renal Patients Quality of Life Study Group.  Nephrol Dial Transplant  1996; 11(suppl 2):125-129.

193. Delano BG: Improvements in quality of life following treatment with r-HuEPO in anemic hemodialysis patients.  Am J Kidney Dis  1989; 14(2 suppl 1):14-18.

194. Moreno F, Sanz-Guajardo D, Lopez-Gomez JM, et al: Increasing the hematocrit has a beneficial effect on quality of life and is safe in selected hemodialysis patients. Spanish Cooperative Renal Patients Quality of Life Study Group of the Spanish Society of Nephrology.  J Am Soc Nephrol  2000; 11(2):335-342.

195. Canadian Erythropoietin Study Group : Association between recombinant human erythropoietin and quality of life and exercise capacity of patients receiving haemodialysis.  BMJ  1990; 300(6724):573-578.

196. Laupacis A, Wong C, Churchill D: The use of generic and specific quality-of-life measures in hemodialysis patients treated with erythropoietin. The Canadian Erythropoietin Study Group.  Control Clin Trials  1991; 12(4 suppl):168S-179S.

197. Besarab A, Bolton WK, Browne JK, et al: The effects of normal as compared with low hematocrit values in patients with cardiac disease who are receiving hemodialysis and epoetin.  N Engl J Med  1998; 339(9):584-590.

198. Furuland H, Linde T, Ahlmen J, et al: A randomized controlled trial of haemoglobin normalization with epoetin alfa in pre-dialysis and dialysis patients.  Nephrol Dial Transplant  2003; 18(2):353-361.

199. Parfrey PS, Foley RN, Wittreich BH, et al: Double-blind comparison of full and partial anemia correction in incident hemodialysis patients without symptomatic heart disease.  J Am Soc Nephrol  2005; 16(7):2180-2189.

200. Roger SD, McMahon LP, Clarkson A, et al: Effects of early and late intervention with epoetin alpha on left ventricular mass among patients with chronic kidney disease (stage 3 or 4): Results of a randomized clinical trial.  J Am Soc Nephrol  2004; 15(1):148-156.

201. Strippoli GF, Craig JC, Manno C, Schena FP: Hemoglobin targets for the anemia of chronic kidney disease: A meta-analysis of randomized, controlled trials.  J Am Soc Nephrol  2004; 15(12):3154-3165.

202. Brazier J, Cooper N, Maloney Jr JV, Buckberg G: The adequacy of myocardial oxygen delivery in acute normovolemic anemia.  Surgery  1974; 75(4):508-516.

203. Pianosi P, D'Souza SJ, Charge TD, et al: Cardiac output and oxygen delivery during exercise in sickle cell anemia.  Am Rev Respir Dis  1991; 143(2):231-235.

204. Cilley RE, Scharenberg AM, Bongiorno PF, et al: Low oxygen delivery produced by anemia, hypoxia, and low cardiac output.  J Surg Res  1991; 51(5):425-433.

205. Fowler NO, Holmes JC: Blood viscosity and cardiac output in acute experimental anemia.  J Appl Physiol  1975; 39(3):453-456.

206. Levin A, Thompson CR, Ethier J, et al: Left ventricular mass index increase in early renal disease: Impact of decline in hemoglobin.  Am J Kidney Dis  1999; 34(1):125-134.

207. Brown DW, Giles WH, Croft JB: Left ventricular hypertrophy as a predictor of coronary heart disease mortality and the effect of hypertension.  Am Heart J  2000; 140(6):848-856.

208. Benjamin EJ, Levy D: Why is left ventricular hypertrophy so predictive of morbidity and mortality?.  Am J Med Sci  1999; 317(3):168-175.

209. Ma JZ, Ebben J, Xia H, Collins AJ: Hematocrit level and associated mortality in hemodialysis patients.  J Am Soc Nephrol  1999; 10(3):610-619.

210. Locatelli F, Pisoni RL, Combe C, et al: Anaemia in haemodialysis patients of five European countries: Association with morbidity and mortality in the Dialysis Outcomes and Practice Patterns Study (DOPPS).  Nephrol Dial Transplant  2004; 19(1):121-132.

211. Levin A, Djurdev O, Duncan J, et al: Hemoglobin levels prior to therapy predict survival in chronic kidney disease patients.  J Am Soc Nephrol  2002; 13:461a.

212. Sandgren PE, Murray AM, Herzog CA, et al: Anemia and new-onset congestive heart failure in the general Medicare population.  J Card Fail  2005; 11(2):99-105.

213. Sarnak MJ, Levey AS: Cardiovascular disease and chronic renal disease: A new paradigm.  Am J Kidney Dis  2000; 35(4 suppl 1):S117-S131.

214. Wali RK, Henrich WL: Chronic kidney disease: A risk factor for cardiovascular disease.  Cardiol Clin  2005; 23(3):343-362.

215. Keith DS, Nichols GA, Gullion CM: Longitudinal follow-up and outcomes among a population with chronic kidney disease in a large managed care organization.  Arch Intern Med  2004; 164:659-663.

216. Herzog CA, Ma JZ, Collins AJ: Poor long-term survival after acute myocardial infarction among patients on long-term dialysis.  N Engl J Med  1998; 339(12):799-805.

217. Anand IS, Chandrashekhar Y, Ferrari R, et al: Pathogenesis of oedema in chronic severe anaemia: Studies of body water and sodium, renal function, haemodynamic variables, and plasma hormones.  Br Heart J  1993; 70(4):357-362.

218. Anand IS, Chandrashekhar Y, Wander GS, Chawla LS: Endothelium-derived relaxing factor is important in mediating the high output state in chronic severe anemia.  J Am Coll Cardiol  1995; 25:1402.

219. Chung IJ, Lee JJ, Nam CE, et al: Increased inducible nitric oxide synthase expression and nitric oxide concentration in patients with aplastic anemia.  Ann Hematol  2003; 82(2):104-108.

220. Weiskopf RB, Feiner J, Hopf H, et al: Heart rate increases linearly in response to acute isovolemic anemia.  Transfusion  2003; 43(2):235-240.

221. Woodson RD, Auerbach S: Effect of increased oxygen affinity and anemia on cardiac output and its distribution.  J Appl Physiol  1982; 53(5):1299-1306.

222. Lorell BH, Carabello BA: Left ventricular hypertrophy: Pathogenesis, detection, and prognosis.  Circulation  2000; 102:470.

223. Abergel E, Tase M, Bohlender J, et al: Which definition for echocardiographic left ventricular hypertrophy?.  Am J Cardiol  1995; 75(7):498-502.

224. Foley RN, Parfrey PS, Harnett JD, et al: Clinical and echocardiographic disease in patients starting end-stage renal disease therapy.  Kidney Int  1995; 47(1):186-192.

225. Frohlich E: Left ventricular hypertrophy as a risk factor.  Cardiol Clin  1986; 4:137-145.

226. Kannel WB, Abbott R: A prognostic comparison of asymptomatic left ventricular hypertrophy and unrecognised myocardial infarction: The Framingham Study.  Am Heart J  1986; 111:391-397.

227. Silberberg JS, Rahal DP, Patton DR, Sniderman AD: Role of anemia in the pathogenesis of left ventricular hypertrophy in end-stage renal disease.  Am J Cardiol  1989; 64(3):222-224.

228. Levin A, Singer J, Thompson CR, et al: Prevalent left ventricular hypertrophy in the predialysis population: Identifying opportunities for intervention.  Am J Kidney Dis  1996; 27(3):347-354.

229. Levin A, Thompson CR, Ethier J, et al: Left ventricular mass index increase in early renal disease: Impact of decline in hemoglobin.  Am J Kidney Dis  1999; 34(1):125-134.

230. Rigatto C, Foley R, Jeffery J, et al: Electrocardiographic left ventricular hypertrophy in renal transplant recipients: Prognostic value and impact of blood pressure and anemia.  J Am Soc Nephrol  2003; 14(2):462-468.

231. Astor BC, Arnett DK, Brown A, Coresh J: Association of kidney function and hemoglobin with left ventricular morphology among African Americans: The Atherosclerosis Risk in Communities (ARIC) study.  Am J Kidney Dis  2004; 43(5):836-845.

232. Ayus JC, Go AS, Valderrabano F, et al: Spanish Group for the Study of the Anemia and Left Ventricular Hypertrophy in Pre-dialysis Patients: Effects of erythropoietin on left ventricular hypertrophy in adults with severe chronic renal failure and hemoglobin <10 g/dL.  Kidney Int  2005; 68(2):788-795.

233. Hayashi T, Suzuki A, Shoji T, et al: Cardiovascular effect of normalizing the hematocrit level during erythropoietin therapy in predialysis patients with chronic renal failure.  Am J Kidney Dis  2000; 35(2):250-256.

234. Zehnder C, Zuber M, Sulzer M, et al: Influence of long-term amelioration of anemia and blood pressure control on left ventricular hypertrophy in hemodialyzed patients.  Nephron  1992; 61:21-25.

235. Portoles J, Torralbo A, Martin P, et al: Cardiovascular effects of recombinant human erythropoietin in predialysis patients.  Am J Kidney Dis  1997; 29:541-548.

236. London GM, Pannier B, Guerin AP, et al: Alterations of left ventricular hypertrophy in and survival of patients receiving hemodialysis: Follow-up of an interventional study.  J Am Soc Nephrol  2001; 12:2759-2767.

237. Hayashi T, Suzuki A, Shoji T, et al: Cardiovascular effect of normalizing the hematocrit level during erythropoietin therapy in predialysis patients with chronic renal failure.  Am J Kidney Dis  2000; 35:250-256.

238. Roger SD, McMahon LP, Clarkson A, et al: Effects of early and late intervention with epoetin alpha on left ventricular mass among patients with chronic kidney disease (stage 3 or 4): Results of a randomized clinical trial.  J Am Soc Nephrol  2004; 15(1):148-156.

239. Parfrey PS, Foley RN, Wittreich BH, et al: Double-blind comparison of full and partial anemia correction in incident hemodialysis patients without symptomatic heart disease.  J Am Soc Nephrol  2005; 16(7):2180-2189.

240. Foley RN, Parfrey PS, Morgan J, et al: Effect of hemoglobin levels in hemodialysis patients with asymptomatic cardiomyopathy.  Kidney Int  2000; 58(3):1325-1335.

241. Levin A, Djurdjev O, Thompson C, et al: Canadian randomized trial of hemoglobin maintenance to prevent or delay left ventricular mass growth in patients with CKD.  Am J Kidney Dis  2005; 46(5):799-811.

242. Wizemann V, Kaufmann J, Kramer W: Effect of erythropoietin on ischemia tolerance in anemic hemodialysis patients with confirmed coronary artery disease.  Nephron  1992; 62(2):161-165.

243. Macdougall IC, Lewis NP, Saunders MJ, et al: Long-term cardiorespiratory effects of amelioration of renal anaemia by erythropoietin.  Lancet  1990; 335(8688):489-493.

244. Hase H, Imamura Y, Nakamura R, et al: Effects of rHuEPO therapy on exercise capacity in hemodialysis patients with coronary artery disease.  Jpn Circ J  1993; 57(2):131-137.

245. McClellan WM, Langston RD, Presley R: Medicare patients with cardiovascular disease have a high prevalence of chronic kidney disease and a high rate of progression to end-stage renal disease.  J Am Soc Nephrol  2004; 15:1912-1919.

246. Silverberg DS, Wexler D, Blum M, et al: The use of subcutaneous erythropoietin and intravenous iron for the treatment of the anemia of severe, resistant congestive heart failure improves cardiac and renal function, functional cardiac class, and markedly reduces hospitalizations.  J Am Coll Cardiol  2000; 35:1737-1744.

247. Mancini DM, Katz SD, Lang C: Effect of erythropoietin on exercise capacity in patients with moderate to severe chronic heart failure.  Circulation  2003; 107:294-299.

248. Al-Ahmad A, Levey A, Rand W, et al: Reduced kidney function and anemia as risk factors for mortality in patients with left ventricular dysfunction.  J Am Coll Cardiol  2001; 38:955-962.

249. Silverberg DS, Wexler D, Blum M, et al: The effect of correction of anaemia in diabetics and non-diabetics with severe resistant congestive heart failure and chronic renal failure by subcutaneous erythropoietin and intravenous iron.  Nephrol Dial Transplant  2003; 18(1):141-146.

250. Mohanram A, Zhang Z, Shahinfar S, et al: Anemia and end-stage renal disease in patients with type 2 diabetes and nephropathy.  Kidney Int  2004; 66(3):1131-1138.

251. Kuriyama S, Tomonari H, Yoshida H, et al: Reversal of anemia by erythropoietin therapy retards the progression of chronic renal failure, especially in nondiabetic patients.  Nephron  1997; 77(2):176-185.

252. Hoen B, Paul-Dauphin A, Hestin D, Kessler M: EPIBACDIAL: A multicenter prospective study of risk factors for bacteremia in chronic hemodialysis patients.  J Am Soc Nephrol  1998; 9(5):869-876.

253. Marsh JT, Brown WS, Wolcott D, et al: rHuEPO treatment improves brain and cognitive function of anemic dialysis patients.  Kidney Int  1991; 39(1):155-163.

254. Temple RM, Deary IJ, Winney RJ: Recombinant erythropoietin improves cognitive function in patients maintained on chronic ambulatory peritoneal dialysis.  Nephrol Dial Transplant  1995; 10(9):1733-1738.

255. Pickett JL, Theberge DC, Brown WS, et al: Normalizing hematocrit in dialysis patients improves brain function.  Am J Kidney Dis  1999; 33(6):1122-1130.

256. Crowell JW, Frod RG, Lews VM: Oxygen transport in hemorrhagic shock as a function of the hematocrit ratio.  Am J Physiol  1959; 196(5):1033-1038.

257. Jan KM, Chien S: Effect of hematocrit variations on coronary hemodynamics and oxygen utilization.  Am J Physiol  1977; 233(1):106-113.

258. Benz RL, Pressman MR, Hovick ET, Peterson DD: A preliminary study of the effects of correction of anemia with recombinant human erythropoietin therapy on sleep, sleep disorders, and daytime sleepiness in hemodialysis patients (The SLEEPO study).  Am J Kidney Dis  1999; 34(6):1089-1095.

259. Ferguson JH, Lewis JH, Zucker MB: Bleeding tendency in uremia.  Blood  1956; 11(12):1073-1076.

260. Escolar G, Diaz-Ricart M, Cases A: Uremic platelet dysfunction: past and present.  Curr Hematol Rep  2005; 4(5):359-367.

261. Diaz-Buxo JA, Donadio Jr JV: Complications of percutaneous renal biopsy: An analysis of 1,000 consecutive biopsies.  Clin Nephrol  1975; 4(6):223-227.

262. Brass LF, Bensusan HB: The role of collagen quaternary structure in the platelet: Collagen interaction.  J Clin Invest  1974; 54:1480-1487.

263. Clemetson KJ: Platelet GPIb-V-IX complex.  Thromb Haemost  1997; 78:266.

264. Ma AD, Abrams CS: Pleckstrin homology domains and phospholipid-induced cytoskeletal reorganization.  Thromb Haemost  1999; 82:399.

265. Li R, Mitra N, Gratkowski H, et al: Activation of integrin alphaIIbbeta3 by modulation of transmembrane helix associations.  Science  2003; 300:795.

265a. Furie B, Bouchard BA, Furie BC: Vitamin K-dependent biosynthesis of gamma-carboxyglutamic acid.  Blood  1999; 93(6):1798-1808.

266. Di Minno G, Martinez J, McKean ML, et al: Platelet dysfunction in uremia. Multifaceted defect partially corrected by dialysis.  Am J Med  1985; 79(5):552-559.

267. Castaman G, Rodeghiero F, Lattuada A, et al: Multimeric pattern of plasma and platelet von Willebrand factor is normal in uremic patients.  Am J Hematol  1993; 44(4):266-269.

268. Remuzzi G, Perico N, Zoja C, et al: Role of endothelium-derived nitric oxide in the bleeding tendency of uremia.  J Clin Invest  1990; 86:1768-1771.

269. Noris M, Benigni A, Boccardo P, et al: Enhanced nitric oxide synthesis in uremia: Implications for platelet dysfunction and dialysis hypotension.  Kidney Int  1993; 44:445-450.

270. Gordge MP, Neild GH: Platelets from patients on haemodialysis show impaired responses to nitric oxide.  Clin Sci  1992; 83:313-318.

271. Aiello S, Noris M, Remuzzi G: Nitric oxide synthesis and L-arginine in uremia.  Miner Electrolyte Metab  1997; 23:151-156.

272. Noris M, Remuzzi G: Uremic bleeding: Closing the circle after 30 years of controversies?.  Blood  1999; 94:2569-2574.

273. Gordge MP, Neild GH: Platelet function in uraemia.  Platelets  1991; 2:115-123.

274. Escolar G, Monteagudo J, Castillo R, et al: Ultrastructural immunolocalization and morphometric quantitation of platelet membrane GP1b and GPIIb-IIIa in uremic patients.  Prog Clin Biol Res  1988; 283:197-201.

275. Benigni A, Boccardo P, Galbusera M, et al: Reversible activation defect of the platelet glycoprotein IIb-IIIa complex in patients with uremia.  Am J Kidney Dis  1993; 22:668-676.

276. Gawaz MP, Dobos G, Spath M, et al: Impaired function of platelet membrane glycoprotein IIb-IIIa in end-stage renal disease.  J Am Soc Nephrol  1994; 5:36-46.

277. Escolar G, Diaz-Ricart M, Cases A, et al: Abnormal cytoskeletal assembly in platelets from uremic patients.  Am J Pathol  1993; 143:823-831.

278. Ordinas A, Escolar G, White JG: Ultrastructure of platelets and platelet-surface interactions.   In: Missirlis YF, Wautier JL, ed. The Role of Platelets in Blood-Biomaterial Interactions,  Amsterdam: Kluwer Academic Publishers; 1993:3-13.

279. Diaz-Ricart M, Estebanell E, Cases A, et al: Abnormal platelet cytoskeletal assembly in hemodialyzed patients results in deficient tyrosine phosphorylation signaling.  Kidney Int  2000; 57:1905-1914.

280. Castillo R, Lozano T, Escolar G, et al: Defective platelet adhesion on vessel subendothelium in uremic patients.  Blood  1986; 68(2):337-342.

281. Horowitz HI: Uremic toxins and platelet function.  Arch Intern Med  1970; 126(5):823-826.

282. Benigni A, Livio M, Dodesini P, et al: Inhibition of human platelet aggregation by parathyroid hormone. Is cyclic AMP implicated?.  Am J Nephrol  1985; 5(4):243-247.

283. Gralnick HR, Mckeown LP, Williams SB, Shafer B: Plasma and platelet von Willenbrand factor defects in uremia.  Am J Med  1989; 85:806-810.

284. Sloand JA, Sloand EM: Studies on platelet membrane glycoproteins and platelet function during hemodialysis.  J Am Soc Nephrol  1997; 8(5):799-803.

285. Cases A, Reverter JC, Escolar G, et al: In vivo evaluation of platelet activation by different cellulosic membranes.  Artif Organs  1997; 21:330-334.

286. Hernandez MR, Galan AM, Cases A, et al: Biocompatibility of cellulosic and synthetic membranes assessed by leukocyte activation.  Am J Nephrol  2004; 24:235-241.

287. Cases A, Reverter JC, Escolar G, et al: Platelet activation on hemodialysis: Influence of dialysis membranes.  Kidney Int Suppl  1993; 41:S217-S220.

288. Mason RG, Zucker WH, Bilinsky RT, et al: Blood components deposited on used and reused dialysis membranes.  Biomater Med Devices Artif Organs  1976; 4:333-358.

289. Hernandez MR, Galan AM, Lozano M, et al: Platelet-leukocyte activation during hemodialysis detected with a monoclonal antibody to leukocyte integrin CD11b.  Nephron  1998; 80:197-203.

290. Livio M, Gotti E, Marchesi D, et al: Uraemic bleeding: Role of anaemia and beneficial effect of red cell transfusions.  Lancet  1982; 2(8306):1013-1015.

291. Valles J, Santos MT, Aznar J, et al: Erythrocytes metabolically enhance collagen-induced platelet responsiveness via increased thromboxane production, adenosine diphosphate release, and recruitment.  Blood  1991; 78:154-162.

292. Weiss HJ, Lages B, Hoffmann T, Turitto VT: Correction of the platelet adhesion defect in delta-storage pool deficiency at elevated hematocrit: possible role of adenosine diphosphate.  Blood  1996; 87:4214-4222.

293. Fernandez F, Goudable C, Sie P, et al: Low hematocrit and prolonged bleeding time in uraemic patients: Effect of red cell transfusions.  Br J Haematol  1985; 59:139-148.

294. Moia M, Mannucci PM, Vizzotto L, et al: Improvement in the haemostatic defect of uraemia after treatment with recombinant human erythropoietin.  Lancet  1987; 2(8570):1227-1229.

295. Lindsay RM, Friesen M, Aronstam A, et al: Improvement of platelet function by increased frequency of hemodialysis.  Clin Nephrol  1978; 10(2):67-70.

296. Cases A, Escolar G, Reverter JC, et al: Recombinant human erythropoietin treat-ment improves platelet function in uremic patients.  Kidney Int  1992; 42(3):668-672.

297. Eschbach JW, Egrie JC, Downing MR, et al: Correction of the anemia of end-stage renal disease with recombinant human erythropoietin. Results of a combined phase I and II clinical trial.  N Engl J Med  1987; 316:73-78.

298. Tassies D, Reverter JC, Cases A, et al: Effect of recombinant human erythropoietin treatment on circulating reticulated platelets in uremic patients: Association with early improvement in platelet function.  Am J Hematol  1998; 59(2):105-109.

299. Van Geet C, Van Damme-Lombaerts R, Vanrusselt M, et al: Recombinant human erythropoietin increases blood pressure, platelet aggregability and platelet free calcium mobilization in uremic children: A possible link?.  Thromb Haemost  1990; 64:7-10.

300. Malyszko J, Malyszko JS, Borawski J, et al: A study of platelet functions, some hemostatic and fibrinolytic parameters in relation to serotonin in hemodialyzed patients under erythropoietin therapy.  Thromb Res  1995; 77:133-143.

301. Diaz-Ricart M, Estebanell E, Cases A, et al: Erythropoietin improves signaling through tyrosine phosphorylation in platelets from uremic patients.  Thromb Haemost  1999; 82:1312-1317.

302. Zeigler ZR, Megaludis A, Fraley DS: Desmopressin (d-DAVP) effects on platelet rheology and von Willebrand factor activities in uremia.  Am J Hematol  1992; 39:90.

303. Ruggeri ZM, Mannucci PM, Lombardi R, et al: Multimeric composition of factor VIII/von Willebrand factor following administration of DDAVP: implications for pathophysiology and therapy of von Willebrand's disease subtypes.  Blood  1982; 59(6):1272-1278.

304. Gordz S, Mrowietz C, Pindur G, et al: Effect of desmopressin (DDAVP) on platelet membrane glycoprotein expression in patients with von Willebrand's disease.  Clin Hemorheol Microcirc  2005; 32(2):83-87.

305. Soslau G, Schwartz AB, Putatunda B, et al: Desmopressin-induced improvement in bleeding times in chronic renal failure patients correlates with platelet serotonin uptake and ATP release.  Am J Med Sci  1990; 300(6):372-379.

306. Mannucci PM, Remuzzi G, Pusineri F, et al: Deamino-8-D-arginine vasopressin shortens the bleeding time in uremia.  N Engl J Med  1983; 308(1):8-12.

307. Vigano GL, Mannucci PM, Lattuada A, et al: Subcutaneous desmopressin (DDAVP) shortens the bleeding time in uremia.  Am J Hematol  1989; 31(1):32-35.

308. Shapiro MD, Kelleher SP: Intranasal deamino-8-D-arginine vasopressin shortens the bleeding time in uremia.  Am J Nephrol  1984; 4(4):260-261.

309. Rydzewski A, Rowinski M, Mysliwiec M: Shortening of bleeding time after intranasal administration of 1-deamino-8-D-arginine vasopressin to patients with chronic uremia.  Folia Haematol Int Mag Klin Morphol Blutforsch  1986; 113(6):823-830.

310. Mannucci PM, Bettega D, Cattaneo M: Patterns of development of tachyphylaxis in patients with haemophilia and von Willebrand disease after repeated doses of desmopressin (DDAVP).  Br J Haematol  1992; 82(1):87-93.

311. Lusher JM: Response to 1-deamino-8-D-arginine vasopressin in von Willebrand disease.  Haemostasis  1994; 24(5):276-284.

312. Couch P, Stumpf JL: Management of uremic bleeding.  Clin Pharm  1990; 9(9):673-681.

313. Davenport R: Cryoprecipitate for uremic bleeding.  Clin Pharm  1991; 10(6):429.

314. Triulzi DJ, Blumberg N: Variability in response to cryoprecipitate treatment for hemostatic defects in uremia.  Yale J Biol Med  1990; 63(1):1-7.

315. Vigano G, Gaspari F, Locatelli M, et al: Dose-effect and pharmacokinetics of estrogens given to correct bleeding time in uremia.  Kidney Int  1988; 34(6):853-858.

316. McCarthy ML, Stoukides CA: Estrogen therapy of uremic bleeding.  Ann Pharmacother  1994; 28(1):60-62.

317. Liu YK, Kosfeld RE, Marcum SG: Treatment of uraemic bleeding with conjugated oestrogen.  Lancet  1984; 2(8408):887-890.

318. Livio M, Mannucci PM, Vigano G, et al: Conjugated estrogens for the management of bleeding associated with renal failure.  N Engl J Med  1986; 315:731-735.

319. Noris M, Todeschini M, Zappella S, et al: 17beta-estradiol corrects hemostasis in uremic rats by limiting vascular expression of nitric oxide synthases.  Am J Physiol Renal Physiol  2000; 279(4):F626-F635.