Rodak's Hematology: Clinical Principles and Applications, 5th Ed.

CHAPTER 6. Cellular structure and function

Elaine M. Keohane*

OUTLINE

Cell Organization

Plasma Membrane

Membrane Proteins

Membrane Carbohydrates

Nucleus

Chromatin

Nuclear Envelope

Nucleoli

Cytoplasm

Ribosomes

Endoplasmic Reticulum

Golgi Apparatus

Mitochondria

Lysosomes

Microfilaments and Intermediate Filaments

Microtubules

Centrosomes

Hematopoietic Microenvironment

Cell Cycle

Regulation of the Cell Cycle

Cell Death by Necrosis and Apoptosis

Objectives

After completion of this chapter, the reader will be able to:

1. Describe the structure, composition, and general function of cellular membranes.

2. Describe the structure, composition, and function of components of the nucleus, including staining qualities visible by light microscopy.

3. Describe the structure, composition, and general function of the cytoplasmic organelles in the cell, including staining qualities visible by light microscopy, if applicable.

4. Describe the general structure and function of the hematopoietic microenvironment.

5. Associate the stages of the cell cycle with activities of the cell.

6. Describe the role of cyclins and cyclin-dependent kinases in cell cycle regulation.

7. Discuss the function of checkpoints in the cell cycle and where in the cycle they occur.

8. Differentiate between apoptosis and necrosis.

Knowledge of the normal structure, composition, and function of cells is fundamental to the understanding of blood cell pathophysiology covered in later chapters. From the invention of the microscope and the discovery of cells in the 1600s to the present-day highly sophisticated analysis of cell ultrastructure with electron microscopy and other technologies, a remarkable body of knowledge is available about the structure of cells and their varied organelles. Complementing these discoveries were other advances in technology that enabled detailed understanding of the biochemistry, metabolism, and genetics of cells at the molecular level. Today, highly sophisticated analysis of cells using flow cytometry, cytogenetics, and molecular genetic testing (Chapters 30, 31, and 32) has become the standard of care in diagnosis and management of many malignant and non-malignant blood cell diseases. This new and ever-expanding knowledge has revolutionized the diagnosis and treatment of hematologic diseases resulting in a dramatic improvement in patient survival for many conditions that previously had a dismal prognosis. With all these advances, however, the visual examination of blood cells on a peripheral blood film by light microscopy still remains the hallmark for the initial evaluation of hematologic abnormalities.

This chapter will provide an overview of the structure, composition, and function of the components of the cell, the hematopoietic microenvironment, the cell cycle and its regulation, and the process of cell death by apoptosis and necrosis.

Cell organization

Cells are the structural units that constitute living organisms ( and Figures 6-16-2). Cells have specialized functions and contain the components necessary to perform and perpetuate these functions. Regardless of shape, size, or function, human cells contain:

• A plasma membrane that separates the cytoplasm and cellular components from the extracellular environment;

• A membrane-bound nucleus (with the exception of mature red blood cells and platelets); and

• Other unique subcellular structures and organelles that support the various cellular functions.1

Image 

FIGURE 6-1 Cell organization and components.

Image 

FIGURE 6-2 Electron micrograph of a cell. Source: (From Carr JH, Rodak BF: Clinical hematology atlas, ed 4, St. Louis, 2013, Saunders.)

Table 6-1 summarizes the cellular components and their functions, which are explained in more detail later.

TABLE 6-1

Summary of Cellular Components and Functions

Organelle

Location

Appearance and Size

Function

Plasma membrane

Outer boundary of cell

Lipid bilayer consisting of phospholipids, cholesterol, proteins; glycolipids and glycoproteins form a glycocalyx

Provides physical barrier for cell; facilitates and restricts cellular exchange of substances; maintains electrochemical gradient and receptors for signal transduction

Nucleus

Within cell

Round or oval; varies in diameter; composed of DNA and proteins

Controls cell division and functions; and contains genetic code

Nucleolus

Within nucleus

Usually round or irregular in shape; 2-4 μm in diameter; composed of ribosomal RNA and the genes coding it, and accessory proteins; there may be one to several within the nucleus

Synthesizes ribosomal RNA and assembles ribosome subunits

Ribosomes

Free in cytoplasm; also on outer surface of rough endoplasmic reticulum

Macromolecular complex composed of protein and ribosomal RNA; composed of large and small subunits

Synthesizes proteins

Rough endoplasmic reticulum

Membranous network throughout cytoplasm

Membrane-lined tubules that branch and connect to nuclear membrane; studded with ribosomes

Synthesizes most membrane-bound proteins

Smooth endoplasmic reticulum

Membranous network throughout cytoplasm

Membrane-lined tubules contiguous with rough endoplasmic reticulum; does not have ribosomes

Synthesizes phospholipids and steroids; detoxifies drugs; stores calcium

Golgi apparatus

Next to nucleus and rough endoplasmic reticulum

System of stacked, membrane-bound, flattened sacs

Modifies and packages macromolecules for other organelles and for secretion

Mitochondria

Randomly distributed in cytoplasm

Round or oval structures; 3-14 nm in length, 2-10 nm in width; membrane has two layers; inner layer has folds called cristae

Produces most of the cell’s ATP by oxidative phosphorylation

Lysosomes

Randomly distributed in cytoplasm

Membrane-bound sacs; diameter varies

Contains hydrolytic enzymes that degrade unwanted material in the cell

Microfilaments

Near nuclear envelope, plasma membrane, and mitotic processes

Double-stranded, intertwined solid structures of actin; 5-7 nm in diameter

Supports cytoskeleton and motility

Intermediate filaments

Cytoskeleton

Solid structures 8-10 nm in diameter; self-assemble into larger bundles

Provides strong structural support

Microtubules

Cytoskeleton and centrioles, near nuclear envelope and Golgi apparatus

Hollow cylinder of α- and β-tubulin forming 13 protofilaments; 20-25 nm in diameter

Maintains cell shape, motility, and mitotic process

Centrosome

Near nucleus

Composed of two centrioles, each having nine sets of triplet microtubules; 150 nm in diameter, 300-500 nm in length

Contains centrioles that serve as insertion points for mitotic spindle fibers

Plasma membrane

The plasma membrane serves as a semipermeable outer boundary separating the cellular components from their surrounding environment. The cell membrane serves four basic functions: (1) it provides a physical but flexible barrier to contain and protect cell components from the extracellular environment; (2) it regulates and facilitates the interchange of substances with the environment by endocytosis, exocytosis, and selective permeability (using various membrane channels and transporters); (3) it establishes electrochemical gradients between the interior and exterior of the cell; and (4) it has receptors that allow the cell to respond to a multitude of signaling molecules through signal transduction pathways.2

Relevant to hematology, the membrane is also the location of cell surface glycoprotein and glycolipid molecules (surface markers or antigens) used for blood cell identity. Each type of blood cell expresses a unique repertoire of surface markers at different stages of differentiation.3 Monoclonal antibodies are used to identify a blood cell’s surface antigens using flow cytometry (Chapter 32). An international nomenclature was developed, called the cluster of differentiation, or CD, system, in which a CD number was assigned to each identified blood cell surface antigen.4 Over 350 CD antigens have been identified on blood cells.3 The CD nomenclature allows scientists, clinicians, and laboratory practitioners to communicate in a universal language for hematology research and diagnostic and therapeutic practice.

In addition to the plasma membrane, many components found within the cell (e.g., the mitochondria, Golgi apparatus, nucleus, and endoplasmic reticulum) have similarly constructed membrane systems. The red blood cell membrane has been the most widely studied and serves as an example of a cell membrane (Figure 9-2).

To accomplish its many requirements, the cell membrane must be resilient and elastic. It achieves these qualities by being a fluid structure of proteins floating in lipids. The lipids are phospholipids and cholesterol arranged in two layers. The phosphate end of the phospholipid and the hydroxyl radical of cholesterol are polar-charged hydrophilic (water-soluble) structures that orient toward the extracellular and cytoplasmic surfaces of the cell membrane. The fatty acid chains of the phospholipids and the steroid nucleus of cholesterol are non-polar-charged hydrophobic (water-insoluble) structures and are directed toward each other in the center of the bilayer (Figure 13-10).2 The phospholipids are distributed asymmetrically in the membrane with mostly phosphatidylserine and phosphatidylethanolamine in the inner layer and sphingomyelin and phosphatidylcholine in the outer layer (Chapters 9 and 13). In the outer layer, carbohydrates (oligosaccharides) are covalently linked to some membrane proteins and phospholipids (forming glycoproteins and glycolipids, respectively).2 These also contribute to the membrane structure and function.

Membrane proteins

Cell membranes contain two types of proteins: transmembrane and cytoskeletal. Transmembrane proteins may traverse the entirety of the lipid bilayers in one or more passes and penetrate the plasma and cytoplasmic layers of the membrane. The transmembrane proteins serve as channels and transporters of water, ions, and other molecules between the cytoplasm and the external environment. They also function as receptors and adhesion molecules. Cytoskeletal proteins are found only on the cytoplasmic side of the membrane and form the lattice of the cytoskeleton. The cytoplasmic ends of transmembrane proteins attach to the cytoskeletal proteins at junctional complexes to provide structural integrity to the cell and vertical support in linking the membrane to the cytoskeleton (Figure 9-4).5 Inherited mutations in genes coding for transmembrane or cytoskeletal proteins can disrupt membrane integrity, decrease the life span of red blood cells, and lead to a hemolytic anemia. An example is hereditary spherocytosis (Chapter 24).

Membrane carbohydrates

The carbohydrate chains of the glycoproteins and glycolipids extend beyond the outer cell surface, giving the cell a carbohydrate coat often called the glycocalyx. These carbohydrate moieties function in cell-to-cell recognition and provide a negative surface charge, surface receptor sites, and cell adhesion capabilities. The function of the red blood cell membrane is discussed in detail in Chapter 9.

Nucleus

The nucleus is composed of three components: the chromatin, the nuclear envelope, and the nucleoli. It is the control center of the cell and the largest organelle within the cell. The nucleus is composed largely of deoxyribonucleic acid (DNA) and is the site of DNA replication and transcription (Chapter 31). It is responsible for the chemical reactions within the cell and the cell’s reproductive process. The nucleus has an affinity for basic dyes because of the nucleic acids contained within it; it stains deep purple with Wright stain (Chapters 8 and 12).

Chromatin

The chromatin consists of one long molecule of double-stranded DNA in each chromosome that is tightly folded with histone and nonhistone proteins. The first level of folding is the formation of nucleosomesalong the length of the DNA molecule (Figure 30-3). Each nucleosome is 11 nm in length and consists of approximately 150 base pairs of DNA wrapped around a histone protein core.6 The positive charge of the histones facilitates binding with the negatively charged phosphate groups of DNA. The nucleosomes are folded into 30 nm chromatin fibers, and these fibers are further folded into loops, then supercoiled chromatin fibers that greatly condense the DNA (Figure 30-3). This highly structured folding allows the long strands of DNA to be tightly condensed in the nucleus when inactive and enables segments of the DNA to be rapidly unfolded for active transcription when needed. This complex process of gene expression is controlled by transcription factors and other regulatory proteins and processes. Inappropriate silencing of genes needed for blood cell maturation contributes to the molecular pathophysiology of myelodysplastic syndromes and acute leukemias (Chapters 34 and 35).

Morphologically, chromatin is divided into two types: (1) the heterochromatin, which is represented by the more darkly stained, condensed clumping pattern and is the transcriptionally inactive area of the nucleus, and (2) the euchromatin, which has diffuse, uncondensed, open chromatin and is the genetically active portion of the nucleus where DNA transcription into mRNA occurs. The euchromatin is loosely coiled and turns a pale blue when stained with Wright stain. More mature cells have more heterochromatin because they are less transcriptionally active.

Nuclear envelope

Surrounding the nucleus is a nuclear envelope consisting of two phospholipid bilayer membranes. The inner membrane surrounds the nucleus, and the outer membrane is continuous with an extension of the endoplasmic reticulum.1 Between the two membranes is a 30- to 50-nm perinuclear space that is continuous with the lumen of the endoplasmic reticulum.6 Nuclear pore complexes penetrate the nuclear envelope, which allows passage of molecules between the nucleus and the cytoplasm.

Nucleoli

The nucleus contains one to several nucleoli. The nucleolus is the site of ribosomal RNA (rRNA) production and assembly into ribosome subunits. Because the ribosomes synthesize proteins, the number of nucleoli in the nucleus is proportional to the amount of protein synthesis that occurs in the cell. As blood cells mature, protein synthesis decreases, and the nucleoli eventually disassemble.

Nucleoli contain a large amount of rRNA, the genes that code for rRNA (or rDNA), and ribosomal proteins. In ribosome biogenesis, rDNA is first transcribed to rRNA precursors. The rRNA precursors are processed into smaller RNA molecules and subsequently complexed with proteins forming the small and large ribosome subunits.6 The ribosomal proteins enter the nucleus through the nuclear pores after being synthesized in the cytoplasm. After the ribosome subunits are synthesized and assembled, they are transported out of the nucleus through the nuclear pores. Once in the cytoplasm, the large and small ribosome subunits self-assemble into a functional ribosome during protein synthesis (Chapter 31).6

Cytoplasm

The cytoplasmic matrix is a homogeneous, continuous, aqueous solution called the cytosol. It is the environment in which the organelles exist and function. These organelles are discussed individually.

Ribosomes

Ribosomes are macromolecular complexes composed of a small and large subunit of rRNA and many accessory ribosomal proteins. Ribosomes are found free in the cytoplasm or on the surface of rough endoplasmic reticulum. They may exist singly or form chains (polyribosomes). Ribosomes serve as the site of protein synthesis. This is accomplished with transfer RNA (tRNA) for amino acid transport to the ribosome, and specific messenger RNA (mRNA) molecules. The mRNA provides the genetic code for the sequence of amino acids for the protein being synthesized (Chapter 31). Cells that actively produce proteins have many ribosomes in the cytoplasm which give it a dark blue color (basophilia) when stained with Wright stain. Cytoplasmic basophilia is particularly prominent in RBC precursor cells when hemoglobin and other cell components are actively synthesized (Chapter 8).

Endoplasmic reticulum

The endoplasmic reticulum () is a membranous network found throughout the cytoplasm and appears as flattened sheets, sacs, and tubes of membrane. The outer membrane of the nuclear envelope is continuous with the endoplasmic reticulum membrane and it specializes in making and transporting lipids and membrane proteins. Figure 6-3

Image 

FIGURE 6-3 Endoplasmic reticulum. Note rough endoplasmic reticulum with attached ribosomes.

Rough endoplasmic reticulum (RER) has a studded look on its outer surface caused by the presence of ribosomes engaged in the synthesis of mainly membrane-bound proteins.2 Smooth endoplasmic reticulum (SER) is contiguous with the RER, but it does not contain ribosomes. It is involved in synthesis of phospholipids and steroids, detoxification or inactivation of harmful compounds or drugs, and calcium storage and release.2

Golgi apparatus

The Golgi apparatus is a system of stacked, membrane-bound, flattened sacs called cisternae that are involved in modifying, sorting, and packaging macromolecules for secretion or delivery to other organelles. It contains numerous enzymes for these activities. The Golgi apparatus is normally located in close proximity to the rough endoplasmic reticulum (RER) and the nucleus. In stained bone marrow smears of developing white blood cell precursors, the Golgi area may be observed as an unstained region next to the nucleus.

Vesicles containing membrane-bound and soluble proteins from the RER enter the Golgi network on the “cis face” and are directed through the stacks where the proteins are modified, as needed, by enzymes for glycosylation, sulfation, or phosphorylation.12 Vesicles with processed proteins exit the Golgi on the “trans face” to form lysosomes or secretory vesicles bound for the plasma membrane.12

Mitochondria

The mitochondrion () has a continuous outer membrane. Running parallel to the outer membrane is an inner membrane that invaginates at various intervals, giving the interior a shelflike or ridgelike appearance. These internal ridges, termed Figure 6-4cristae, are where oxidative enzymes are attached. The convolution of the inner membrane increases the surface area to enhance the respiratory capability of the cell. The interior of the mitochondrion consists of a homogeneous material known as the mitochondrial matrix, which contains many enzymes for the extraction of energy from nutrients.

Image 

FIGURE 6-4 Mitochondrion.

The mitochondria generate most of the adenosine triphosphate (ATP) for the cell. Mitochondrial enzymes oxidize pyruvate and fatty acids to acetyl CoA, and the citric acid cycle oxidizes the acetyl CoA producing electrons for the electron-transport pathway. This pathway generates ATP through oxidative phosphorylation.2

The mitochondria are capable of self-replication. This organelle has its own DNA and RNA for the mitochondrial division cycle. There may be fewer than 100 or up to several thousand mitochondria per cell. The number is directly related to the amount of energy required by the cell.

Lysosomes

Lysosomes contain hydrolytic enzymes bound within a membrane and are involved in the cell’s intracellular digestive process. The membrane prevents the enzymes from digesting cellular components and macromolecules. Lysosomal enzymes are active at the acidic pH of the lysosome and are inactivated at the higher pH of the cytosol.2 This also protects the cell in case lysosomal enzymes are released into the cytoplasm. Lysosomes fuse with endosomes and phagosomes (Chapter 12); this allows the lysosome hydrolytic enzymes to safely digest their contents.1 With Wright stain, lysosomes are visualized as granules in white blood cells and platelets (Chapters 12 and 13). Lysosomal lipid storage diseases result from inherited mutations in genes for enzymes that catabolize lipids. Gaucher disease and Tay-Sachs disease are examples of these disorders (Chapter 29).

Microfilaments and intermediate filaments

Actin microfilaments are double-stranded, intertwined solid structures approximately 5 to 7 nm in diameter. They associate with myosin to enable cell motility, contraction, and intracellular transport. They locate near the nuclear envelope or in the proximity of the nucleus and assist in cell division. They also are present near the plasma membrane and provide cytoskeletal support.

Intermediate filaments, with a diameter of approximately 8 to 10 nm, self-assemble into larger bundles.2 They are the most durable element of the cytoskeleton and provide structural stability for the cells, especially those subjected to more physical stress, such as the epidermal layer of skin.1 Examples include the keratins and lamins.

Microtubules

Microtubules are hollow cylindrical structures that are approximately 25 nm in diameter and vary in length. These organelles are organized from α- and β-tubulin through self-assembly.2 The tubulin polypeptides form protofilaments, and the microtubule usually consists of 13 protofilaments.1 This arrangement gives the microtubules structural strength. Tubulins can rapidly polymerize and form microtubules and then rapidly depolymerize them when no longer needed by the cell.

Microtubules have several functions. They help support the cytoskeleton to maintain the cell’s shape and are involved in the movement of some intracellular organelles. Microtubules also form the mitotic spindle fibers during mitosis and are the major components of centrioles.

Centrosomes

The centrosome consists of two cylinder-shaped centrioles that are typically oriented at right angles to each other. A centriole consists of nine bundles of three microtubules each. They serve as insertion points for the mitotic spindle fibers during mitosis.

Hematopoietic microenvironment

Hematopoiesis occurs predominantly in the bone marrow from the third trimester of fetal life through adulthood (Chapter 7). The bone marrow microenvironment must provide for hematopoietic stem cell self-renewal, proliferation, differentiation, and apoptosis and support the developing progenitor cells. This protective environment is provided by stromal cells, which is a broad term for specialized endothelial cells; reticular adventitial cells (fibroblasts); adipocytes (fat cells); lymphocytes and macrophages; osteoblasts; and osteoclasts.7 The stromal cells secrete substances that form an extracellular matrix, including collagen, fibronectin, thrombospondin, laminin, and proteoglycans (such as hyaluronate, chondroitin sulfate, and heparan sulfate).78 The extracellular matrix is critical for cell growth and for anchoring developing blood cell progenitors in the bone marrow. Hematopoietic progenitor cells have many receptors for cytokines and adhesion molecules. One purpose of these receptors is to provide a mechanism for attachment to extracellular matrix. This provides an avenue for cell-cell interaction, which is essential for regulated hematopoiesis.

Stromal cells also secrete many different growth factors required for stem, progenitor, and precursor cell survival (Chapter 7). Growth factors participate in complex processes to regulate the proliferation and differentiation of progenitor and precursor cells. Growth factors must bind to specific receptors on their target cells to exert their effect. Most growth factors are produced by cells in the hematopoietic microenvironment and exert their effects in local cell-cell interactions. One growth factor, erythropoietin, has a hormone-type stimulation in that it is produced in another location (kidney) and exerts its effect on erythroid progenitors in the bone marrow (Chapter 8). An important feature of growth factors is their use of synergism to stimulate a cell to proliferate or differentiate. In other words, several different growth factors work together to generate a more effective response.9 Growth factors are specific for their corresponding receptors on target cells.

Growth factor receptors are transmembrane proteins.9 When the growth factor (or ligand) binds the extracellular domain of the receptor, a signal is transmitted to the nucleus in the cell through the cytoplasmic domain. For example, when erythropoietin binds with its receptor, it causes a conformational change in the receptor which activates a kinase (Janus kinase 2 or JAK2) associated with its cytoplasmic domain.9 The activated kinase in turn activates other intracellular signal transduction molecules that ultimately interact with the DNA in the nucleus to promote expression of genes required for cell growth and proliferation (Figure 33-9).

Cell cycle

The purpose of the cell cycle is to replicate DNA once and distribute identical chromosome copies equally to two daughter cells during mitosis.10 The cell cycle is a biochemical and morphologic four-stage process through which a cell passes when it is stimulated to divide (Figure 6-5). These stages are G1 (gap 1), S (DNA synthesis), G2 (gap 2), and M (mitosis). G1 is a period of cell growth and synthesis of components necessary for replication. G1 lasts about 10 hours.10 In the S stage, DNA replication takes place, a process requiring about 8 hours (Chapter 31). An exact copy of each chromosome is produced and they pair together as sister chromatids. The centrosome is also duplicated during the S stage.10 In G2, the tetraploid DNA is checked for proper replication and damage (discussed later). G2 takes approximately 4 hours. The time spent in each stage can be variable, but mitosis takes approximately 1 hour.10 During G0 (quiescence) the cell is not actively in the cell cycle.

Image 

FIGURE 6-5 Stages of the cell cycle. A, Diagrams of cellular morphology and chromosome structure across the cell cycle; B, Time scale of cell cycle stages; C, Length of cell cycle stages in cultured cells. Source: (From Pollard TD, Earnshaw WC. Chapter 40 Introduction to the cell cycle. In: Cell Biology, e2. Philadelphia, 2008, Saunders, An imprint of Elsevier.)

The mitosis or M stage involves the division of chromosomes and cytoplasm into two daughter cells. It is divided into six phases (Figure-6-5):10

1. Prophase: the chromosomes condense, the duplicated centrosomes begin to separate, and mitotic spindle fibers appear.

2. Prometaphase: the nuclear envelope disassembles, the centrosomes move to opposite poles of the cell and serve as a point of origin of the mitotic spindle fibers; the sister chromatids (chromosome pairs) attach to the mitotic spindle fibers.

3. Metaphase: the sister chromatids align on the mitotic spindle fibers at a location equidistant from the centrosome poles.

4. Anaphase: the sister chromatids separate and move on the mitotic spindles toward the centrosomes on opposite poles.

5. Telophase: the nuclear membrane reassembles around each set of chromosomes and the mitotic spindle fibers disappear.

6. Cytokinesis: the cell divides into two identical daughter cells

Interphase is a term used for the non-mitosis stages of the cell cycle, that is, G1, S, and G2.

Regulation of the cell cycle

A regulatory mechanism is needed to prevent abnormal or mutated cells from going through the cell cycle and producing an abnormal clone. The cell cycle is a highly complicated process that can malfunction. There are four major checkpoints in the cell cycle (Figure 6-5).110 The first is a restriction point late in G1 that checks for the appropriate amount of nutrients and appropriate cell volume. The second checkpoint at the end of G1 (called the G1 DNA damage checkpoint) checks the DNA for damage and makes the cell wait for DNA repair or initiates apoptosis. The third checkpoint, G2 DNA damage checkpoint, takes place after DNA synthesis at the end of G2, and its purpose is to verify that replication took place without error or damage. If abnormal or malformed replication occurred, then mitosis is blocked. The last checkpoint is during mitosis at the time of metaphase (metaphase checkpoint). Here the attachment and alignment of chromosomes on the mitotic spindle and the integrity of the spindle apparatus are checked.110 Anaphase will be blocked if any defects are detected.

Cell cycle control is under the direction of cyclin and cyclin-dependent kinases (CDKs). The cyclin/CDK complexes phosphorylate key substrates that assist the cell through the cell cycle. Cyclin is named appropriately because the concentration of the cyclin/CDK complex moves the cell through the different stages of the cell cycle. G1 begins with a combination of the cyclin D family (D1, D2, D3) with CDK4 and CDK6.10 To transition the cell from G1 to S, cyclin E increases and binds to CDK2, producing the cyclin E/CDK2 complex. In the S stage cyclin E decreases and cyclin A increases and complexes with CDK2, forming cyclin A/CDK2. This complex takes the cell through the S and G2 stage. Cyclin A also partners with CDK1 (cyclin A/CDK1). For mitosis to occur, cyclin B must replace cyclin A and bind to CDK1, forming the cyclin B/CDK1 complex. This complex takes the cell through the intricate process of mitosis.1011 Inhibitors of the cyclin/CDK complexes also play a primary role in cell cycle regulation.11

Tumor suppressor proteins are needed for the proper function of the checkpoints. One of the first tumor suppressor genes recognized was TP53. It codes for the TP53 protein that detects DNA damage during G1. It can also assist in triggering apoptosis. Many tumor suppressor genes have been described.11 When these genes are mutated or deleted, abnormal cells are allowed to go through the cell cycle and replicate. Some of these cells simply malfunction, but others form neoplasms, often with aggressive characteristics. For example, patients with chronic lymphocytic leukemia have a more aggressive disease with a shorter survival time when their leukemic cells lose TP53 activity either through gene mutation or deletion (Chapter 36). Patients whose leukemia cells have normal TP53 function have a better prognosis.

Cell death by necrosis and apoptosis

Cell death occurs as a normal physiologic process in the body or as a response to injury. Events that injure cells include ischemia (oxygen deprivation), mechanical trauma, toxins, drugs, infectious agents, autoimmune reactions, genetic defects including acquired and inherited mutations, and improper nutrition.12 There are two major mechanisms for cell death: necrosis and apoptosis. Necrosis is a pathologic process caused by direct external injury to cells—for example, from burns, radiation, or toxins.12 Apoptosis is a self-inflicted cell death originating from the activation signals within the cell itself.13 Most apoptosis occurs as a normal physiologic process to eliminate potentially harmful cells (e.g., self-reacting lymphocytes [Chapter 7]), cells that are no longer needed (e.g., excess erythroid progenitors in oxygen-replete states (Chapter 8) or neutrophils after phagocytosis), and aging cells.12 Apoptosis of older terminally differentiated cells balances with new cell growth to maintain needed numbers of functional cells in organs, hematopoietic tissue, and epithelial cell barriers, particularly in skin and the intestines. On the other hand, apoptosis also initiates in response to internal or external pathologic injury to a cell. For example, if DNA damage occurred during the replication phase of the cell cycle and the damage is beyond the capability of the DNA repair mechanisms, the cell will activate apoptosis to prevent its further progression through the cell cycle. Apoptosis can also be triggered in virally infected cells by the virus itself or by the body’s immune response.12 This is one of the mechanisms to remove virally infected cells from the body.

The first morphologic manifestation of necrosis is a swelling of the cell. The cell may be able to recover from minor injury at that point. More severe damage, however, disrupts organelles and membranes; enzymes leak out of lysosomes that denature and digest DNA, RNA, and intracellular proteins; and ultimately the cell lyses.12 An inflammatory response usually accompanies necrosis due to the release of cell contents into the extracellular space.

The morphologic manifestation of apoptosis is shrinkage of the cell. The nucleus condenses and undergoes systematic fragmentation due to cleavage of the DNA between nucleosome subunits (multiples of 180 to 200 base pairs). The plasma membrane remains intact, but the phospholipids lose their asymmetric distribution and “flip” phosphatidylserine (PS) from the inner to the outer leaflet.14 The cytoplasm and nuclear fragments bud off in membrane-bound vesicles. Macrophages, recognizing the PS and other signals on the membranes, rapidly phagocytize the vesicles. Thus, cellular products are not released into the extracellular space and an inflammatory response is not elicited.12 Figure 6-6 and Table 6-2 summarize the differences between necrosis and apoptosis.

Image 

FIGURE 6-6 Schematic illustration of the morphologic changes in cell injury culminating in necrosis or apoptosis. Source: (From: Kumar V, Abbas AK, Fausto, N, et al. Chapter 1 Cellular Responses to Stress and Toxic Insults: Adaptation, Injury, and Death. In: Robbins and Cotran Pathologic Basis of Disease, e8. Philadelphia, 2009, Saunders, an Imprint of Elsevier.)

TABLE 6-2

Comparison of Necrosis and Apoptosis1213

 

Necrosis

Apoptosis

Cell size

Enlarged due to swelling

Reduced due to shrinkage

Nucleus

Random breaks and lysis (karyolysis)

Condensation and fragmentation between nucleosomes

Plasma membrane

Disrupted with loss of integrity

Intact with loss phospholipid asymmetry

Inflammation

Enzyme digestion and leakage of cell contents; inflammatory response occurs

Release of cell contents in membrane-bound apoptotic bodies which are phagocytized by macrophages; no inflammation occurs

Physiologic or pathologic function

Pathologic; results from cell injury

Mostly physiologic to remove unwanted cells; pathologic in response to cell injury

Activation of apoptosis occurs through extrinsic and intrinsic pathways. Both pathways involve the activation of proteins called caspases. The extrinsic pathway, also called the death receptor pathway, initiates with the binding of ligand to a death receptor on the cell membrane. Examples of death receptors and their ligands include Fas and Fas ligand, and tumor necrosis factor receptor 1 (TNFR1) and tumor necrosis factor.15The binding activates caspase-8. The intrinsic pathway is initiated by intracellular stressors (such as hypoxia, DNA damage, or membrane disruption) that stimulate the release of cytochrome c from mitochondria.15 Cytochrome c binds to apoptotic protease-activating factor-1 (APAF-1) and caspase-9, forming an apoptosome, which activates caspase-9. Both pathways converge when the “initiator” caspases (8 or 9) activate “executioner” caspases 3, 6, and 7, which leads to apoptosis.1315

Various cellular proapoptotic and antiapoptotic proteins tightly regulate apoptosis. Examples of antiapoptotic proteins include some members of the BCL-2 family of proteins (such as Bcl-2, Bcl-XL) as well as various growth factors (such as erythropoietin, granulocyte-colony stimulating factor, granulocyte-macrophage-colony stimulating factor, interleukin-3, and FLT3 ligand).14 BAX, BAK, and BID are examples of proapoptotic proteins.14 The ratio of these intracellular proteins plays a primary role in regulating apoptosis. Any dysregulation, mutation, or translocation can cause inhibition or overexpression of apoptotic proteins, which can lead to hematopoietic malignancies or malfunctions.1214

Summary

• The cell contains cytoplasm that is separated from the extracellular environment by a plasma membrane; a membrane-bound nucleus (with the exception of mature red blood cells and platelets); and other unique subcellular structures and organelles.

• The plasma membrane is a bilayer of phospholipids, cholesterol, and transmembrane proteins. Glycolipids and glycoproteins on the outer surface form the glycocalyx.

• The cytoplasm contains ribosomes for protein synthesis, which can be free in the cytoplasm or located on rough endoplasmic reticulum (RER). The RER makes most of the membrane proteins. Smooth endoplasmic reticulum (SER) lacks ribosomes; the SER is involved in synthesis of phospholipids and steroids, detoxification or inactivation of harmful compounds or drugs, and calcium storage and release.

• The Golgi apparatus modifies and packages macromolecules for secretion and for other cell organelles. Mitochondria make ATP to supply energy for the cell. Lysosomes contain hydrolytic enzymes involved in the cell’s intracellular digestive process.

• The bone marrow provides a suitable microenvironment for hematopoietic stem cell self-renewal, proliferation, differentiation, and apoptosis. Stromal cells secrete substances that form an extracellular matrix to support cell growth and function and help to anchor developing cells in the bone marrow. Growth factors participate in complex processes to regulate the proliferation and differentiation of hematopoietic stem and progenitor cells.

• The cell cycle involves four active stages: G1 (gap 1), S (DNA synthesis), G2 (gap 2), and M (mitosis). The cell cycle is under the direction of cyclins and CDKs. Checkpoints in the cell cycle recognize abnormalities and initiate apoptosis.

• Two major mechanisms for cell death are necrosis and apoptosis. Necrosis is a pathologic process caused by direct external injury to cells, while apoptosis is a self-inflicted cell death originating from the activation signals within the cell itself. Most apoptosis occurs as a normal physiologic process to eliminate unwanted cells, but it can also be initiated in response to internal or external pathologic injury to a cell.

Review questions

Answers can be found in the Appendix.

1. The organelle involved in packaging and trafficking of cellular products is the:

a. Nucleus

b. Golgi apparatus

c. Mitochondria

d. Rough endoplasmic reticulum

2. The glycocalyx is composed of membrane:

a. Phospholipids and cholesterol

b. Glycoproteins and glycolipids

c. Transmembrane and cytoskeletal proteins

d. Rough and smooth endoplasmic reticulum

3. The “control center” of the cell is the:

a. Nucleus

b. Cytoplasm

c. Membrane

d. Microtubular system

4. The nucleus is composed largely of:

a. RNA

b. DNA

c. Ribosomes

d. Glycoproteins

5. Protein synthesis occurs in the:

a. Nucleus

b. Mitochondria

c. Ribosomes

d. Golgi apparatus

6. The shape of a cell is maintained by which of the following?

a. Microtubules

b. Spindle fibers

c. Ribosomes

d. Centrioles

7. Functions of the cell membrane include all of the following except:

a. Regulation of molecules entering or leaving the cell

b. Receptor recognition of extracellular signals

c. Maintenance of electrochemical gradients

d. Lipid production and oxidation

8. The energy source for cells is the:

a. Golgi apparatus

b. Endoplasmic reticulum

c. Nucleolus

d. Mitochondrion

9. Ribosomes are synthesized by the:

a. Endoplasmic reticulum

b. Mitochondrion

c. Nucleolus

d. Golgi apparatus

10. Euchromatin functions as the:

a. Site of microtubule production

b. Transcriptionally active DNA

c. Support structure for nucleoli

d. Attachment site for centrioles

11. The cell cycle is regulated by:

a. Cyclins and CDKs

b. Protooncogenes

c. Apoptosis

d. Growth factors

12. The transition from the G1 to S stage of the cell cycle is regulated by:

a. Cyclin B/CDK1 complex

b. Cyclin A/CDK2 complex

c. Cyclin D1

d. Cyclin E/CDK2 complex

13. Apoptosis is morphologically identified by:

a. Cellular swelling

b. Nuclear condensation

c. Rupture of the cytoplasm

d. Rupture of the nucleus

14. Regulation of the hematopoietic microenvironment is provided by the:

a. Stromal cells and growth factors

b. Hematopoietic stem cells

c. Liver and spleen

d. Cyclins and caspases

References

1.  Pollard T.D, Earnshaw W.C. Introduction to cells. In Cell Biology. 2nd ed. Philadelphia : Saunders, An imprint of Elsevier 2008.

2.  Mescher A.L. The cytoplasm. In: Mescher A.L. Junqueira’s Basic Histology. 13th ed. New York : McGraw-Hill 2013 Available at: http://www.accessmedicine.com.libproxy2.umdnj.edu/content.aspx?aID=57330175 Accessed January 5.01.14.

3.  Kipps T.J. The cluster of differentiation antigens. In: Prchal J.T, Kaushansky K, Lichtman M.A, Kipps T.J, Seligsohn U. Williams Hematology. 8th ed. New York : McGraw-Hill 2010.

4.  Zola H, Swart B, Nicholson I, et al. CD molecules 2005 human cell differentiation molecules ; 2005; 106:3123-3126.

5.  Mohandas N, Gallagher P.G. Red cell membrane past, present, and future. Blood; 2008; 112:3939-3948.

6.  Mescher A.L. The nucleus. In: Mescher A.L. Junqueira’s Basic Histology. 13th ed. New York : McGraw-Hill 2013.

7.  Koury M.J, Lichtman M.A. Structure of the marrow and the hematopoietic microenvironment. In: Prchal J.T, Kaushansky K, Lichtman M.A, Kipps T.J, Seligsohn U. Williams Hematology. 8th ed. New York : McGraw-Hill 2010.

8.  Kaushansky K. Hematopoietic stem cells, progenitors, and cytokines. In: Prchal J.T, Kaushansky K, Lichtman M.A, Kipps T.J, Seligsohn U. Williams Hematology. 8th ed. New York : McGraw-Hill 2010.

9.  Kaushansky K. Signal transduction pathways. In: Prchal J.T, Kaushansky K, Lichtman M.A, Kipps T.J, Seligsohn U. Williams Hematology. 8th ed. New York : McGraw-Hill 2010.

10.  Pollard T.D, Earnshaw W.C. Introduction to the cell cycle. In Cell Biology. 2nd ed. Philadelphia : Saunders, An imprint of Elsevier 2008 Chapter 40.

11.  Schmid M, Carson D.A. Cell-cycle regulation and hematologic disorders. In: Prchal J.T, Kaushansky K, Lichtman M.A, Kipps T.J, Seligsohn U. Williams Hematology. 8th ed. New York : McGraw-Hill 2010.

12.  Kumar V, Abbas A.K, Fausto N, et al. Cellular responses to stress and toxic insults adaptation, injury, and death. In Robbins and Cotran Pathologic Basis of Disease. 8th ed. Philadelphia : Saunders, an Imprint of Elsevier 2009 Chapter 1.

13.  Danial N.K, Hockenbery D.M. Cell death. In: Hoffman R, Benz E.J, Jr. Silberstein L.E, et al. Hematology Basic Principles and Practice 6th ed. Philadelphia : Saunders, an imprint of Elsevier 2013.

14.  Gottlieb R.A. Apoptosis. In: Prchal J.T, Kaushansky K, Lichtman M.A, Kipps T.J, Seligsohn U. Williams Hematology. 8th ed. New York : McGraw-Hill 2010 Chapter 12.

15.  McIlwain D.R, Berger T, Mak T.W. Caspase functions in cell death and diseaseCold Spring Harb Perspect Biol; 2013; 5:1-28.


*The author extends appreciation to Keila B. Poulsen, whose work in prior editions provided the foundation for this chapter.