Ezio Fulcheri1, 2 , Maria Pia Brisigotti3, 4 and Leonardo Resta5
Division of Anatomic Pathology, Departement of Surgical and Diagnostic Science, University of Genoa, Genoa, Italy
UOSD Feto-perinatal Pathology, IRCCS Istituto Giannina Gaslini, University of Genoa, Largo G.Gaslini 5, Genoa, 16147, Italy
Ph.D course in Paediatric Science, Fetal-Perinatal and Paediatric Pathology, University of Genoa, Genoa, Italy
UOSD Feto-perinatal Pathology, IRCCS Istituto Giannina Gaslini, Genova, Largo G.Gaslini 5, Genoa, 16147, Italy
Section of Pathological Anatomy, Department of Emergency and Organ Transplantation (DETO), University of Bari, Bari, Italy
Ezio Fulcheri (Corresponding author)
A miscarriage can be defined as a pregnancy that ends spontaneously before the foetus has reached a viable gestational age . The term miscarriage is often utilised for both a clinical and a biochemical pregnancy loss. Clinical pregnancies are the ones that can be identified by ultrasound or histological evidence, while biochemical pregnancies occur earlier and can only be identified by a raised bHCG. In practice, the majority of biochemical pregnancy losses may go unnoticed. In fact, evidence suggests that the actual biochemical loss rate in the general population may even reach 60 % .
A miscarriage can be sporadic if there is only a single occurrence within the reproductive life of the woman, but if there are two miscarriages, it is repeated; however, if there are three or more, it is defined as recurrent miscarriage (or habitual) by most clinicians .
Though miscarriage is a rather common event, it is not easy to determine the true incidence due to impossibility of correctly evaluating signs and symptoms. Miscarriages, according to the American College of Obstetricians and Gynecologists, are the most frequent type of interruption of the pregnancy, reporting that miscarriages after artificial insemination are about 50 % of the total. The overall risk of miscarriage is estimated to be from 10 to 25 % and it increases with age. For women over age 35, the probability is 15 %, while from age 35–45 the risk is much higher being from 20 to 35 %. However, for women over age 45, the probability of a miscarriage reaches about 50 %. In the case of women who have already experienced a sporadic miscarriage, the probability of a second event is around 20 %, which in fact is not that much different from women who have not miscarried [20, 28]. If biochemical pregnancy loss were fully taken into consideration, the rate of spontaneous recurrent miscarriage would reach 20 %.
Miscarriage can be separated into early pregnancy loss (EPL) and late pregnancy loss (LPL) but not only because of when the miscarriage occurs but also because of its cause. EPL is defined as in the period from conception to the tenth conception week (12th gestation week) so including the blastogenesis and organogenesis phases and also the initial period of xenogenesis. From the 12th gestational week to the 22nd week, a miscarriage is classed as a LPL. In a late pregnancy loss, the cause is almost always to be found in some type of infection, while in early loss, for the majority of events, there is some structural alteration of the villi due to karyotype anomalies.
3.2 Histopathological Diagnostics of Early Pregnancy Loss
The histopathological diagnostics of EPL as part of the pathology of human reproduction has today become even more important than before because of the radical changes which have taken place in the last 30 years concerning the conditions leading to EPL. These can be defined as intrinsic (those pathological conditions of the mother and of the embryo/foetus and its placenta) and extrinsic (those environmental conditions from the artificial environment created by humans). Within intrinsic causes we find the mother’s age, dyslipidaemias and autoimmunity while within extrinsic causes we can list increases in radiation, pollution, chemicals found in soils or in industrial processes and of course medicinal drugs whether administered voluntarily or involuntarily as in adulterated or contaminated foodstuffs.
Today we must be even more aware of the anthropological and psychological aspects of miscarriage as a pregnancy is sought for and programmed and nonsuccess is not part of the programme, especially if this lack of success is repeated. The emotional impact can be damaging to the extent in which a person believes that disease and death can be beaten back and no longer accepts defeat. Every miscarriage must be seen as an event for grieving, an event which can undermine not only the psychological and behavioural equilibrium of the mother but also the equilibrium and future of the parents’ relationship.
People look more and more to histopathological diagnostics to find more precise answers to the question of how much risk there is of a recurrence of the event in future pregnancies or if the mother has a latent pathology.
3.3 The Diagnostic Approach
The diagnostics of EPL cannot have the same criteria as those of “surgical pathology”. The approach must be broad and open-minded with all the characteristics of a postmortem examination of an adult. Today the autopsy of the embryo requires all the aspects of a complex and multi-specialist diagnostic approach. Only such an approach allows a global vision of the pathology of the pregnancy which includes the conditions of the mother and of the foetus, the normal or pathological conditions of the foetal adnexa, including the gestational sac and the placenta and the even more delicate relationships in the interactions with the decidua implantation site .
This all-round approach permits the making of diagnoses not only speculatively useful in the determination of the cause of the miscarriage (post hoc) but above all useful for the discovery of any possible latent maternal pathologies, parental genetic anomalies or even extrinsic natural/unnatural environmental conditions (human–nature relationship).
3.4 The Thanatology of the Embryo and Its Adnexa
Determining the time of arrest in the development of the embryo is extremely important not only for diagnosis but also for the clinical anamnestic aspects of the case management .
In the large majority of cases, the moment of arrest of development (the death of the embryo) is different to the moment of expulsion from the gestational sac or what remains of it.
This discrepancy is important to be able to correctly interpret the autolytic phenomena which have taken place in the embryo but more crucially in the amniochorial structures necessary for a correct differential diagnosis with morphological or structural anomalies as the cause of the miscarriage.
With this operational method, it is in fact possible to identify four major conditions:
3.5 The Morphological Definition of the Gestational Sac
The second most important diagnostic aspect is the morphological definition of the gestational sac. In the literature we find different classifications [22, 36], some of which are now historic, but it seems there is neither a universally accepted valid classification system nor one which reflects actual clinical needs.
It needs to be said that all the diagnostics of EPL are not an exercise in virtuosity for the pathologist but in fact an important tool in the management of the infertile parents and especially of the woman.
Many classification systems are based almost exclusively on the macroscopic aspects of the expelled material, but without knowledge of the diagnostic procedure, it is difficult for them to be validated. Before taking up this theme, a few words are needed on how the material is collected.
3.6 Procedure for the Collection of the Material
The aspects of the material that reaches the pathologist have been influenced by the situations outlined above and especially by the method of expulsion (spontaneous or pharmacologically induced or via suction or scraping).
In a miscarriage with spontaneous expulsion, a large quantity of blood clots is usually present and not infrequently all of the uterine mucosa (decidua) detached in one piece. Such an event allows a complete examination of the gestational sac to be carried out together with a precise analysis of the implantation site. It must be remembered that the material could be from a second expulsion of the decidua and remaining parts of the chorial structure after a spontaneous miscarriage which is often unrecognised by the patient.
In a pharmacologically induced expulsion of the foetus through the use of prostaglandin, there will be much material complete of all structures. However, the alterations such as the oedema of the mucosa and the congestion of the blood vessels of the decidua are directly correlated with the effect of the drugs and they must not be considered in the report.
In a surgically induced evacuation of the products of conception, we must distinguish between the material obtained by suction and that obtained by scraping. In the first case, all the villous structures will be distorted and twisted, and the embryo, if developed beyond the 18th week, will necessarily be fragmented and lacerated. In the second case, the material will be characterised by a large quantity of blood clots and often the edges of the decidua will be lacerated and delaminated in a mix with the chorial tissue.
It is evident that any macroscopic classification of the gestational sac will initially be affected by the method of collection and by the thanatologic chronology.
The listed above procedures were stated by .
3.7 The Main Classifications
The main classifications for early pregnancy loss had the first of them originally defined in 1966 followed by others but without any of them ever having a significant impact on the diagnostics. A brief analysis of these will be useful before we propose a new classification system.
The Fujikura classification  is based on the integrity of the gestational sac, on the presence or absence of the embryo and on the characteristics of this, if it is amorphous, cylindrical or stunted; the completeness or incompleteness of the material which reaches the pathologist is emphasised together with the integrity or not of the gestational sac.
The Poland classification  concentrates on the examination of the embryo, completely ignoring the characteristics of the gestational sac; embryos are divided into three categories, stunted (dimensions between 1 and 4 mm), cylindrical (dimensions less than 10 mm) and if the embryo has a perfect correlation between the cranium-caudal length and the stage and if there are abnormal pathologies present in the embryo. Differently to Fujikura’s scheme, the three types of embryo are defined based on both the macroscopic and microscopic aspects, including some parts of the details of Carnegie’s classification  and part of the characteristics of the adnexa (yolk sac and amniotic sac).
Great weight is given to the gestational sac if it is found to be intact and empty.
Later, a classification by Mall condensed and simplified various features of Poland’s classification. Rushton proposed a fourth classification  based principally on the morphological and histological aspects of the placenta. Three categories were identified, the first relative to the early phases of conception, the second to the presence of a macerated foetus and the third with a non-macerated foetus. The third category is primarily interested in late pregnancy loss, while the second focuses on the diagnostics of early pregnancy loss at the end of the phase of embryonal development.
3.8 Normal Histology of the Gestational Sac and the Embryonic Adnexa
Before talking about the histopathological picture of EPLs, it is necessary to define the important aspects of a normal histological picture.
The decidua is composed of the decidua basalis, the implantation site, the parietalis and the capsularis.
The decidua basalis is characterised (and recognisable) for the thick fibrinoid streak above it. It is composed of markedly modified stromal cells thanks to the effects of progesterone (physiological hyperprogesteronism). The cells have a pale cytoplasm slightly distant between themselves in a loose interstitial matrix. We find modified endometrial glands lined by a simple cuboidal epithelium, while the vessels have walls of irregular thickness, maintained by infrequent smooth muscle cells (IICH positive for smooth muscle actin) . The wall, like the endometrial modified stroma (decidua cells), is infiltrated by a proliferation of extravillous intermediate trophoblasts (EVIT) which have a discretely enlarged roundish nucleus with an abundant eosinophilic cytoplasm. These elements (IICH – positive for CD 146 and CK 19) infiltrate the vessel walls substituting the normal constituents and not infrequently they are found in clumps inside the vessels [6, 17, 23].
Other EVIT colonise the decidua stroma arranging themselves in band-shaped clumps. In this situation it is not possible to find a tissue-type structure as the cells remain isolated and disaggregated like “flocks of birds”.
There are also loci relatively limited by fibrinoid necroses of the endometrial stroma with an inflammatory infiltrate principally made up of lymphocytes. Such loci can be compared to the normal mechanism of expansion of the placenta bed and not infrequently they are characterised peripherally by a valley of ischaemic necrosis which involves both the stroma cells and the EVIT that colonised the sector.
The decidua capsularis lacks the fibrinoid streak while there is the presence of scattered villi, without a trophoblast layer and with unrecognisable vessels. The villous stroma which makes up the majority of the structure is compact and slightly basophilic. The endometrial stroma is also compact with rare small vessels and sparse EVIT cells.
The decidua parietalis is sometimes the larger part of the abortive material. It is a thick endometrium, completely decidualised with dilated glands, lined with simple cuboidal epithelium, often separated. The endometrial vessels are modified, with normal or apparently thickened walls by a contraction of the smooth muscle cells. The endothelium is always integral and well preserved.
EVIT cells are few and sparse. Some elements have a hyperchromatic nucleus which is slightly irregular or pyknotic, a sign of a cell in functional exhaustion.
During the phase of what is known as the Arias Stella reaction, the endometrium is characterised by lushness and crowded secretory glands with a sinuous lumen which has a ragged profile, all due to the hyperprogesteronism, causing hyperplastic and secretory modifications of the glands further from the implantation site. The cells are enlarged with an abundant clear cytoplasm of the secretory type, and they jut out into the lumen in tufts. Atypical cytology is common enough to have a situation known as an atypical Arias Stella phenomenon.
The gestational sac gives us the chorionic plate with branching villi, the amniochorial membranes and the funiculus and very rarely the yolk sac.
The chorion has two distinct parts:
· The chorion laeve is made up of a layer of fibrinoid and a chorionic plate from which short irregular shaped villi branch off, principally attributable to intermediate or pre-stem elements. The stroma of the villi is characterised by a considerable level of degeneration. The trophoblast lining is irregular if not completely absent. Commonly, especially in the weeks after the fifth week, the villi are walled inside the aforesaid fibrinoid layer. The decidua capsularis which lines the chorion laeve is thin, a modified stroma for the most part without glands but with a complete blood vessel network.
· The chorion frondosum is made up for the most part of villi that will go on to create the chorionic plate; only from the 12° gestational week can we talk about a chorionic plate and therefore a placenta.
The villi are crowded but branch regularly and will eventually form the stem villi and the intermediate type. They exhibit a bilayer epithelium of cytotrophoblasts and syncytiotrophoblasts, the former being positive for the stain IIC with p63 while the latter being positive for HCG and PLAP. The villous stroma is a reticular fibrous network of cells with a small nucleus with interspaces in which Hofbauer cells are suspended, them being typical. Blood vessels are generally without walls, with a regular and slightly prominent endothelium (positive for IIC per CD 31).
At the superior part of the immature intermediate villi, we find buds of intermediate trophoblasts which are strongly positive for Ki67 e per CD 146 .
The amniochorial membranes are difficult to identify as the free membranes are strongly influenced by the presence of chorion laeve in which numerous chorial villi still remain, while the membranes which line the chorion frondosum are not yet well anchored to the villous part which will later go on to form the chorionic plate as an integral part of the completed placenta structure. Therefore, in the expelled material, we can find ribbons of amniotic epithelia sustained by homogeneous sub-amniotic connective tissue which is lightly coloured. The amniotic epithelium is simple, cylindrical and cuboidal.
The funiculus is distinguishable only from Carnegie stage ten. Prior to this we find the body stalk in which we can find two arteries and a vein. Occasionally there are reports of a second vein in involution or parts of a vessel in the Wharton jelly.
Remains of the allantois and the ductus omphaloentericus are very rarely found because they are not still functionally active.
The yolk sac is the part of the embryo adnexa which is the most important in the diagnostics of early pregnancy loss. Responsible for the production of red blood cells, it has a honeycomb structure that is an anastomosed network with small sacs and lacunae wherein we find the immature elements of erythropoietic production (foetal erythroblasts). The foetal erythroblasts are released together with the mature erythrocytes in a percentage which varies from week to week. The major peak is at the seventh gestational week (fifth conception week, Carnegie stage 13) which sees the progressive expansion of the amniotic sac inside the chorionic sac with the initial fusion of the two elements. At the eighth gestational week, the nucleated erythrocytes are 75 % of the total erythrocytes in circulation; this percentage goes down to 50 % at the ninth week, 20 % at the tenth week and 10 % at the 11th week. In the placenta, in its definite form at the 12th week, nucleated erythrocytes are only 1 % of the total population.
3.9 Interpretative Errors: Main Pitfalls to Avoid on Normal Examples
For the decidua:
For the chorial structures:
3.9.1 The Embryo
For a morphological description of the embryo, see the classification of the Carnegie Institute of Embryology .
3.10 The Type of Material
Before taking up the argument of the diagnostics of EPL, it is necessary to draw a conclusion on what has been said so far.
3.11 Criteria and Diagnostic Elements
The famous classifications utilised for the definition of miscarriage essentially refer to a macroscopic evaluation of the expelled tissues.
In the case of massive expulsion of the integral gestational sac complete with the endometrial layer, we are in the ideal conditions to carry out a detailed assessment of all the components of the mother–embryo complex. Only in this case can we diagnose a “blighted ovum”, an early arrest of the development of the embryo, anomalies of development and formation of the embryo and more generally all the conditions of the large and complex chapter of embryo pathology.
In the case of expulsion with lacerations of the gestational sac (as in the case of haemorrhage of a prior pregnancy loss or in the case of a cleaning out of the cavity), the material is not really representative of all the constituents and even less representative of the embryo or what remains of it due to autolytic activity. The absence of the embryo does not permit a complete diagnosis to be carried out.
The histopathological diagnostics is therefore based on expelled material or that collected from scraping in a very altered and fragmented condition from which it is often difficult to identify the topographical anatomical relationships of individual components, and so the reconstruction of the histo-architecture of the various tissues becomes difficult.
All the anatomic parts making up the mother–embryo complex, even if fragmented, incomplete and mixed together, are essential to define the adequacy or not of the material; in the absence of one or more parts, the diagnosis will necessarily be only partial and incomplete, less comprehensive or even only approximate .
This distinction must be kept to the forefront whenever one speaks of diagnostic criteria and must of course be mentioned in the diagnostic report.
3.12 The Classification of the Principal Histopathological Features
A basic classification of the principal histopathologic categories identifies eight principal groups of lesions.
3.12.1 Histo-architectural Alterations Indicative of Karyotype Anomalies
This category is the most important in miscarriages in the first trimester of gestation [16, 29, 34].
22.214.171.124 Clinical Findings
The incidence of major chromosomopathy in miscarriage varies between 40 and 60 %. The distribution of said anomalies sees in the first place autosomal trisomies (52 % approx.) followed by 19 % of a missing X chromosome (45,X0), 22 % of polyploidy (of which 16 % is triploidy), 7 % of structural aberrations and 8 % of mosaicism or autosomal monosomy. The clinical meaning of the chromosomic aberrations of number is only relative in that there is a low recurrence in successive pregnancies. There are however families with a significant recurrence of aneuploidy: in these cases the genetic mechanisms are suspect, but they are difficult to evaluate clinically as they are such as disjunction defects and germinal mosaicism. Fifty percent of the structural aberrations are inherited from a parent carrier of balanced structural rearrangement. In couples with recurrent miscarriage, the prevalence of a parent carrier of balanced structural rearrangement (reciprocal and robertsonian translocations, inversion) is estimated to be at about 5 % with a direct and apparently significant proportionality between the number of miscarriages and the frequency of parental chromosome rearrangements. The constant progress in molecular genetics can open a new chapter in aetiopathogeneses of miscarriage, particularly in recurrent miscarriage. We still do not have definite data, but mechanisms such as uniparental disomy, genomic imprinting, monogenic anomalies and the alterations of random X chromosome inactivation will probably assume an important role in the aetiopathogenetic definition of a still unexplained quota of recurrent unsuccessful pregnancies.
126.96.36.199 Principal Macroscopic Morphological Characteristics
Usually there is abundant material, with the chorionic component being predominant and formed of oedematous villous structures which give a swollen aspect to the tissue. Readily identifiable pale and translucent villous groups can be seen; however, this cannot be used diagnostically as it is well known that in some karyotype anomalies, villi are fibrotic, thin and compact.
Consideration must be given to the extraction method because in the case of suction, the macroscopic chorionic component is severely altered, but with little change to the decidua.
188.8.131.52 Principal Microscopic Morphological Characteristics
Chorionic villi have an irregular pattern of growth and branching, characteristically having large and swollen alternating with smaller villi (Figs. 3.1 and 3.2). There is no unique pattern, there being an extreme variety of shapes including some that seem bizarre and others that seem more shapeless but with deep indentations. The trophoblastic layer loses its regular linearity of the two types of trophoblasts though not in a circumferential way but from a focal point and so presents a hyperplasia of the trophoblast even with macro-nucleated forms or with vacuoles in the cytoplasm. Some hyperplastic aspects, with high-eosinophil cytoplasm cells, are ascribable to the intermediate villous trophoblast. The stroma is loose, with reduced cellularity, or frankly oedematous with small aggregates of cells of a histiocytic macrophagic nature. Degenerative phenomena consist mainly of basophil degeneration and of the mineralisation of the basal membrane of the trophoblast covering.
Picture of a group of crowded chorionic villi with evident histo-architectural alterations. The villar profile is irregular. Bizarre forms and different size of villi are present. A small amount of fibrinoid may be observed in the intervillar space. These characteristics are referred to a genetic disease of the embryo
Chorionic villi in an abortive pregnancy with anomalous karyotype of the embryo. The stroma is oedematous and shows a basophilic degeneration. Small collapsed vessels are evident in the sub-trophoblastic area. The trophoblast shows a minimal and focal hyperplasia: this aspect is different from the circumferential proliferation present in the hydatidiform mole
Characteristic and pathognomonic are the inclusion of trophoblasts (determined by the intersection of a cut line with a deep invagination) (Fig. 3.3). The vessels are irregularly distributed and it is difficult to recognise the affluent vascular structure within the main branches of the villi, precursors of the stem villi. Some of the villi are hypo-vascularised or with an extremely irregular or reduced vascularisation (differential diagnosis in a complete mole). If present, the erythrocytes in circulation do not correspond to the gestational period (ratio between erythroblasts and erythrocytes).
In this particularly we observe a scalloped villus with marked irregularity of the profile and pseudo-inclusion of the syncytio-trophoblast (arrows)
184.108.40.206 Differential Diagnosis
The most basic differential diagnosis is the hydropic abortus in which the proliferative aspects of the trophoblast are missing and above all there is the irregular profile of the villi with fiord-like indentations. However, the main differential diagnosis is that of molar degeneration, both total (hydatidiform) and partial (triploidy) [4, 5].
3.12.2 Infarcted Haemorrhagic Lesions Responsible for the Separation of the Gestational Sac
220.127.116.11 Clinical Findings
This category of miscarriage includes numerous pathologies such as diabetes, gestational and essential hypertension and even genetic factors which seem to play an important role (genes Stox1, COMT and CORIN). These pathologies can have diverse symptoms and have a varying impact on the pregnancy. From recent studies, smoking also has an important role in this problem.
18.104.22.168 Principal Macroscopic Morphological Characteristics
The material contains a large quantity of blood and blood clots either recent or stratified with fibrin; often the chorionic villi are bathed in clotted blood and the decidua material, which has a wine-red colour and is more abundant than the chorionic material. The quantity and the characteristics of the blood clots vary depending on whether the material comes from a complete and sudden separation of the gestational sac or from a separation in steps over time.
22.214.171.124 Principal Microscopic Morphological Characteristics
The main histological alterations affect the decidua. In the decidua basalis, we find haemorrhages (Fig. 3.4) which are recent or mixed over time accompanied by thrombi of the decidua lacunae or of the modified spiral arteries. In some cases, it is possible to see the breakage of the vessel walls with haemorrhagic apoplectic type expansion which disassociates the cells from the cytogen stroma.
Decidua with recent haemorrhages
A second characteristic element is the haemorrhagic flooding of the inert glandular structure. There is also an accompanying inflammatory infiltrate which is more marked when there have been a series of haemorrhagic events over time before the final event of the complete separation. In this case we observe (a) blood clotting or fresh thrombi alternating with reabsorbed clotting which leave traces in the histiocytic haemosiderophagic elements (Fig. 3.5) and (b) recent thrombi alternating with completely organised thrombi (Fig. 3.6).
Case of miscarriage due to abruption of the ovular sac. The picture shows in the decidua a large haemorrhagic focus, partially organised, for several repetitive and more serious events
Decidual haemorrhage with a partially organised thrombus and reactive inflammatory infiltrate. The pattern is characteristics of the subacute and repetitive haemorrhagic events. A differential diagnosis with several types of maternal thrombophilia or coagulative disorders is necessary
In the decidua parietalis, we find interstitial oedema and haemorrhagic vessel outflows. The decidua vessels far from the implantation site can be normal or have thickened walls because of hyperplasia of the smooth muscle cells or for hyalinosis of the media. The complete separation of the gestational sac can be due to varying causes including maternal circulatory pathologies such as thrombophilia and all of the maternal hypertensive states. This can determine both a haemorrhage behind the plate with successive miscarriage and a state of hypoxic hypo-perfusive suffering which provokes villous lesions due to the hypoxia or even ischaemic infarction.
The chorial components include more or less normal villi in the case of massive recent separation or, depending on the type and on the chronology of the haemorrhagic events, three main alterations. In case of a recent massive membrane separation, the villi are normal. A recent but not massive separation shows swollen oedematous villi, while in cases with a series of partial separations which have been long time, we find villi with fibrotic stroma with a slight hyperplasia of the syncytiotrophoblast even in syncytial knot shapes. These findings are consequent to the hypo-perfusive state determined in the chorionic plate by the disconnection between the chorionic structure and the maternal vascular bed caused by the interposition of the clots. In these conditions there is an increase in the intervillous fibrin with moderate inflammatory lymphoplasmacellular infiltrate.
126.96.36.199 Differential Diagnosis
The differential diagnosis comes from the haemorrhage consequent to the endometritis or to the vasculitis of whatever origin or aetiology, considering that the relationship between haemorrhage and inflammation is always a consequence and rarely they do manifest themselves in isolation or in a pure form.
3.12.3 Decidua Vascular Lesions or Primitive Maternal Systemic Lesions Responsible for Hypo-perfusion and Hypo-oxygenation of the Chorionic Plate
Such conditions form in a maternal pre-existing hypertensive state, of maternal degenerative vasculopathies, especially atheromatous, or from smoking. Particular attention should be given to smoking as a cause of increased risk to vascular pathologies especially in young women and those with a genetic predisposition to heart problems [15, 27]. The damage to placental vessels is antecedent to the pregnancy and to the fertilisation.
188.8.131.52 Principal Macroscopic Morphological Characteristics
As the concluding event of the miscarriage is usually a haemorrhage with separation of the gestational sac, the macroscopic view is not much different from that of decidua haemorrhage. In this case the villous component is normally present and the villi appear thin and often compacted in clumps of fibrin.
184.108.40.206 Principal Microscopic Morphological Characteristics
In the decidua basalis we find recent haemorrhages cause a terminal separation of the gestational sac (Fig. 3.7); the decidua arteries appear normally modified by the proliferation of extravillous intermediate trophoblasts with a loss of the smooth muscle component of the walls. In the most striking cases, we can find a vascular dilation with fibrinoid deposits in the intima. There is also a minimum infiltration of foamy histiocytes  accompanied by rare lymphocytes which dissect the sub-intimal portion of the vessels among the remains of the rare smooth muscle cells. The decidualisation of the endometrial stroma appears normal as is the proliferation of the extravillous intermediate trophoblasts.
Old ischaemic infarctual lesion characterised by collapsed villi, with interposition of a minimal amount of fibrinoid material, is present in a decidual sheet. A necrotic and haemorrhagic band with a reactive inflammatory infiltrate is interposed between the lesion and the endometrium
For the decidua parietalis, the most relevant finding is the already reported hyperplasia and hypertrophy of the tunica media accompanied by a reactive thickening of the intima itself (Fig. 3.8). The lumen is restricted if not virtual while the thickening by both a hyperplasia of the smooth muscle cells and a fibro-sclerosis of the wall itself.
Histological picture of a fragment of decidua parietalis in which the spiral arteries are not modified by the trophoblast, as in the normal pregnancy. The arterial lumen is reduced and/or obliterated by a cell proliferation in the intima and the sclerosis of the media layer. This aspect is typical of the endometrial arteriopathies correlated with the maternal older age, hypertension or smoking abuse
The most characteristic lesions are from the chorionic component. The villi appear hyper-mature and hyper-branched, having stem villi (normally present in the third trimester) that are completely out of sync even if the miscarriage takes place slightly later (9–10 weeks). The villi have buds and syncytial knots while the stromal support has a lightly diffused fibrosis. The excessive branching is supported by a proliferation of capillaries and pre-capillaries which are dilated especially in the peripheral parts.
In the case of a hypoxic ischaemic suffering which persists in the embryo or in the case of arrested development or a protracted retention, the vessels appear in a collapsed state with a virtual lumen. Usually the circulating foetal erythrocytes in the ratio of erythroblasts/erythrocytes are not congruent with the gestational time.
220.127.116.11 Differential Diagnosis
The differential diagnosis is found in all the defects of superficial implantation either due to an inadequate proliferation of extravillous intermediate trophoblasts or due to inadequate and insufficient implantation because of an incomplete decidualisation of the endometrium mainly caused by luteal deficits. The histology is complicated and complex and involves the decidua basalis which presents arteries at the implantation site which are unmodified or only a little modified, or there is a scarce decidualisation of the stromal cells in which elements of the endometrial stroma can still be seen.
At the chorionic level, the villi are thin, little branched, far apart and with a fibrous stroma. The only element common to the damage by hypo-perfusion is found in the presence of the syncytial knots, in the terminal parts of the villi and along the length of the stem villi.
3.12.4 Histo-architectural Alterations Indicative of Inadequate and Insufficient Implantation
These conditions include:
18.104.22.168 Clinical Findings
A lutein insufficiency, otherwise known as corpus luteum deficiency, is a defect found in 4–5 % of women with unexplainable infertility or recurrent miscarriage. They often suffer from problems with the rhythm of the menstrual cycle with a shorter progestin phase.
22.214.171.124 Principal Macroscopic Morphological Characteristics
There are no particular macroscopic aspects. There is an average quantity of material, haemorrhagic in part due to the separation or how the material was obtained.
126.96.36.199 Principal Microscopic Morphological Characteristics
As was said in the introduction, there are two distinct situations: the first mainly due to a maternal alteration (luteal phase defect) and the second mainly due to an alteration in the gestational sac.
188.8.131.52 Differential Diagnosis
The differential diagnosis does not treat the different situations which cause the miscarriage but treat the two different categories described in this paragraph or the hypo-perfusion states described previously (Figs. 3.12 and 3.13). What is essential is the evaluation of the vascular wall by immunohistochemical staining with antibody anti-smooth muscle actin and desmin and by typing the extravillous intermediate trophoblast with antibody anti-CD 146.
Miscarriage for insufficient and inadequate implantation. The villi are hypo-branching with syncytial knots
Miscarriage for insufficient and inadequate implantation. The villi are hypo- branching with syncytial knots. The figure shows a thin and elongated villus in which the immunohistochemical reaction for p63 demonstrates the regular presence of cytotrophoblast cells. The regularity of the cytotrophoblast layer excludes an anomalous karyotype and witnesses a regular even if hypo-perfusion inhibited development of the villi
The defect in trophoblast proliferation can also be correlated with a misreactive state or autoimmunity of the mother that in fact reduces the growth and organisation of the trophoblasts especially that part destined to invade and modify the maternal vessel network. Each form of misreactivity is able to interfere in the relationship between maternal immunology and foetal immunology, modifying their interactions and the subsequent morphological expression of their interface.
We must remember that a raw defect in the proliferation of the extravillous trophoblast causes damage exclusively to the gestational sac. This situation opens an important chapter linked to the intrinsic characteristics of the embryo and of its own pathologies. The examination of the embryo will not be dealt with here as embryo pathology deserves a chapter to itself.
3.12.5 Infective States
184.108.40.206 Clinical Findings
These situations are extremely serious and devastating and so it is often very difficult to identify the pathogen. Microbiology and virology can lead to a sure aetiologic diagnosis. However, a precise diagnosis of the pathogen involved is not very useful in a context of the pathology of reproduction and rarely is it identified.
The signs and symptoms of miscarriages caused by infections are properly those of the infection (shivering, fever, septicaemia, peritonitis) and of the threat of miscarriage or incomplete miscarriage. There is a leucocytosis (WBC 16,000–22,000/ml). Those severely ill patients can show signs of septic or endotoxin shock with vasomotor collapse, hypothermia, oliguria or anuria and respiratory failure. The causes include Escherichia coli, Enterobacter aerogenes, Streptococcus haemolyticus, staphylococci and some anaerobic microorganisms like Clostridium perfringens. If the sepsis is due to C. perfringens, an intravascular haemolysis is possible.
220.127.116.11 Principal Macroscopic Morphological Characteristics
Material is abundant and the decidua material is pale and spongy, while the chorionic component is normal or swollen.
18.104.22.168 Principal Microscopic Morphological Characteristics
These can be divided into two principal categories:
22.214.171.124 Differential Diagnosis
The differential diagnosis is the exaggerated inflammatory and necrobiotic reaction at the implantation site that is at times particularly marked and characterised by scarce necrotic tissue. In this case the material from the decidua parietalis is spared by the inflammatory infiltrate.
Particular attention must be paid to those low-level nonspecific inflammatory states that can easily be present in miscarriages long believed to be either reactive to structural alterations or dystrophic in the chorionic components.
3.12.6 Autoimmune Disorders
Pregnancy in itself is a pro-inflammatory state and produces a protracted hypercoagulatory state [13, 30] and so can activate a latent immunitary defect or modulate an already known autoimmune disease. The most common acquired thrombophilia  in cases of recurrent miscarriage that needs to be considered is antiphospholipid syndrome (APS). The diagnosis of APS requires at least one of the clinical criteria and one of the laboratory criteria. Laboratory criteria must be positive on two or more occasions, at least 12 weeks apart. In a large meta-study on couples with RM, the incidence of APS was 15–20 % compared to 5 % in nonpregnant women without a history of obstetric complications. In particular, the antiphospholipids (aPL) are being intensively researched as the mechanisms leading to miscarriage are not well known. Though aPLs are associated with thrombosis, these events cannot explain all the aPL-related miscarriages. There is in vitro evidence of a direct link between aPLs on the trophoblast which provokes a reduction in cell proliferation, in the release of human chorionic gonadotropin, in the invasiveness of the trophoblasts and in the expression of adhesion molecules. It also induces an increase in apoptosis. The damage caused by the aPL seems to be mediated by the beta2 glycoprotein I (ß2GPI) which acts as a cofactor in the bonding of circulating antibodies. An ulterior effect of the bonding of the anti-ß2GPI antibodies is given by the induction of a pro-inflammatory phenotype at the level of decidua, so fostering the activation of the complement and the local secretion of pro-inflammatory cytokines/chemokines .
The autoimmune state of the mother can play an important role in miscarriages of the first trimester. Even if we still cannot define a correlation with the individual autoimmune diseases, their effects are sufficiently clear to permit their inclusion in a nosographic work. In this way the doctor can examine such problems and at the same time avoid wasting time and energy in exploring all the possible causes of the miscarriage. For this reason, each and every diagnostic effort must be made with the aim of characterising the condition. It is worth repeating that the expulsion event is often only the result of a pre-existing disease condition that had determined the failure of the pregnancy and more that sudden dramatic events like endometritis or acute villitis can establish themselves more easily in a compromised autoimmune mother. Finally, even in the case of the identification of the cause of the miscarriage which seems unrelated to the autoimmune field, for example, a structural anomaly in the villi or an alteration in karyotype, it is still necessary to be able to exclude a coexisting autoimmune disorder which could again provoke other future miscarriages.
126.96.36.199 Principal Macroscopic Morphological Characteristics
There are no particular macroscopic aspects. There is an average quantity of material, haemorrhagic in part due to the separation or how the material was obtained.
188.8.131.52 Principal Microscopic Morphological Characteristics [18, 32, 33]
The basal layer of the endometrium appears normally decidualised and the spiral arteries are also normally modified while the proliferation of the extravillous trophoblast is only moderate. We find inflammatory infiltrates characteristically in loci and even more so in the decidua parietalis where small aggregates of lymphocytes crowd the non-modified vessel walls (Figs. 3.17 and 3.18). Some arterioles present thickened intima with hyperplasia of the endothelium and this finding is relatively more evident in the states of transition from decidua basalis to decidua parietalis where you can find larger calibre vessels not yet modified by trophoblastic proliferation. In the chorionic plate, abundant fibrinoid deposits englobe small groups of villi or individual villi (Fig. 3.19). There is also a slight inflammatory infiltrate in the stroma of the villi, while the characteristic histo-architecture of all the chorionic villi is particularly normal components. Only the major vascular branches of the amnio-chorionic network can show alterations in the intima with hyperplasia and endothelium proliferation determining a sub-stenosis of the lumen or a complete vascular occlusion actioned by recent or organised parietal microthrombi.
Miscarriage due to a maternal autoimmune pathology. The decidua shows partially modified spiral arteries and focal inflammatory infiltration. In the borderline between the decidua basalis and parietalis, some more large vessels are not still modified by the trophoblast proliferation. The pattern is very complex with ambiguous findings: the lesions of an inadequate implantation (low or inadequate modification of the arterial wall) are prevalent. On the other hand, the inflammatory reaction and the lesions of the vessel intima suggest a disreactive maternal disease which could be the reason of all the described modifications
Miscarriage due to an autoimmune maternal disease. The small arteries of the decidua parietalis are characterised by a thick intimal layer and some perivascular aggregation of inflammatory infiltrate. The endometrial stroma is very oedematous
(a) Miscarriage due to an autoimmune maternal disease. In the chorionic disc, a large amount of fibrinoid circumscribes the villi (the s. c. fibrinoid pseudo-infarcts). The trophoblast aggregates and small calcifications are present. (b) Another field of the same case: the picture shows the villi embedded in the fibrinoid material. The immunohistochemical reaction for ck 18 (expressed by the trophoblast) designs the villar contour
184.108.40.206 Differential Diagnosis
The differential diagnosis is the inadequate and insufficient implantations that coexist and are mixed with the misreactive and autoimmune states of the mother and which not infrequently are the effect of the latter. We should always remember that the extravillous trophoblast proliferation is orthologic and efficient only when guaranteed by a normal reactive maternal immune system.
3.12.7 Pathology of the Embryonic Adnexa
The pathology of the adnexa is principally that of the yolk sac (Fig. 3.20) .
(a) Yolk sac. In this figure the characteristic structure of the yolk sac (organ of the primary haemopoiesis) is evident. Its wall is thick for the presence of a rich network of sacs and lacunae, often lacking of red blood cells. Note in the top left some normal villi and in the bottom right a sheet of amniotic membrane. (b) Yolk sac. A particular area of the structure of the yolk sac with an evidence of the network of lacunae and sacs in which the immature haemopoietic cells are evident. A minimal inflammatory infiltrate and signs of fibrosis evoke an initial involution of this foetal organ
In the embryonic period between the Carnegie stages 13–22, early involutions of the umbilical vesicle can occur so causing a reduction in erythropoiesis that is critical for the embryo. The involutions are normally determined by moderate and subclinical inflammatory states which are insufficient to provoke an immediate miscarriage.
The opposite situation is created when for pathological reasons of the embryo or mostly for genetic anomalies, there is an early arrest in development of the embryo and only the umbilical vesicle remains. This can occur between the Carnegie stages four to ten (period of blastogenesis, second/third conception week). For a long time, this was confused with a blighted ovum and only a meticulous examination technique of an integral and closed gestational sac permits an incisive differential diagnosis. With the yolk sac remaining, the presence of the embryo is indirectly documented even if there is an early arrest in development.
3.12.8 Pathology of the Embryo
Leaving aside the classifications of Poland, Mall and Rushton that refer only to the macroscopic aspects of the embryo after being deformed by the autolytic processes with its retention in the uterus, this is not the place to embark on an analysis of the pathologies of the embryo which should be treated exhaustively and comprehensively in another place. Embryo and foetal pathology is a study which requires high levels of competence and above all good technique and diagnostic methodologies. Therefore, to fulfil the obligations of the present work, we define the following diagnostic categories as basic and important:
3.13 Closing Considerations
The pathologist’s activities are more and more important in the diagnosis of miscarriage and abortion and also in the field of infertility to which it is often linked. Socio-anthropological change in recent decades has changed the characteristics of the female population, and we are also seeing significant ethnic changes in both urban and rural areas. The most obvious changes in procreation are the increasing age of women when deciding to become a mother and the decision to have only one child at a determined time, while the new immigrants have desires and needs both in maternity and fertility which are different to the local population of today and of yesterday.
Evidently, a general diagnostic formulation carried out on abortive material is no longer sufficient, but it is also evident that a timely and correct diagnosis can be made only in optimal conditions and with the support of cytogenetic examinations and clinic-anamnestic information.
Therefore, taking into account the abovementioned conditions and the fact that in the case of an occasional miscarriage there is no possibility of gathering all the information necessary, we are led to consider the problem from another viewpoint.
There are four different situations:
It remains to ask, with the whys and wherefores of these situations, what can be the minimum or maximum diagnostic level and the most correct type of finding. A diagnosis which is certain, exhaustive and complete is rare as is almost as rare a diagnosis with a high probability of being correct as can be seen from the reviews available. Then there are the diagnoses which are indicative and which have a high value in the delicate fields of infertility and recurrent miscarriages. In no field except this one does the indicative diagnosis become so important in the anatomo-clinical dialogue and in consultations on the pathologies of human reproduction. There remain the diagnoses of exclusion which are still very useful in evaluating the problems of recurrent miscarriage.
In following this scale of diagnostic complexity, the ability to interpret the signs of the gestational sac and of the membranes is paramount, but first it is necessary to exactly establish the adequateness of the material to study in the histological examination:
The basic difference between cause and effect must be clear in the generic diagnostic definition of separation of the gestational sac. In fact, the separation that always follows any type of abortion can in some cases be its own cause, as abruptio placentae, while in other cases it is the normal consequence of a defunct gestational sac.
In conclusion it is worthwhile to summarise and underline the most important aspects.
· Anatomo-clinical integrated diagnostics with the histological examination of the abortive product is a determining moment in the management of spontaneous miscarriage in the first trimester whether it is sporadic or recurrent.
· Missing the diagnostic moment makes it impossible to treat not only the woman but also, in the widest sense, the couple.
· In a spontaneous miscarriage, as in all diagnostic activities in infertility of the couple, current or retrospective investigation on recovered biological material is significant for the health of the mother, becoming a real tool in preventive medicine for those diseases present at the start of the pregnancy (diabetes and hypertension) though latent or not recognised by the woman.
· The reduction to the minimum or the abolition of anatomo-pathologic diagnostic activity has been transformed into an exorbitant increase in costs for nonspecific examinations that could be avoided instead of specific focused investigations.
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