Ultrastructural development of the human decidua

Ultrastructural development of the human decidua

Ultrastructural development of the human decidua RALPH M. WYNN, M.D. Chicago, Illinois The ultrastructure of the human decidua in all three trimesters...

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Ultrastructural development of the human decidua RALPH M. WYNN, M.D. Chicago, Illinois The ultrastructure of the human decidua in all three trimesters of normal pregnancy, in the tube in a case of ectopic pregnancy, and in hydatidiform mole is described. By Day 25 of the menstrual cycle, most of the ultrastructural characteristics associated with pregnancy have begun to appear: The morphologic manifestations of cellular activity in the true decidua are maximal by Day 70 of pregnancy. By Day 100, ultrastructural evidence of metabolic activity begins to decline. During the first 100 days of pregnancy, the ultrastructure of the stroma in the decidua capsularis is virtually identical with that in the decidua parietalis. Later in pregnancy, a material very similar to both basal lamina (basement membrane) and fibrinoid appears to be elaborated by the stromal cells. The decidual reaction in early tubal pregnancy is ultrastructurally indistinguishable from that in uterine pregnancy. The endometrial epithelial responses in normal uterine and tubal pregnancies also are similar. The reaction (Arias-Stella) appears to reflect hyperstimulation rather than degeneration and is further exaggerated in cases of hydatidiform mole. Previously undescribed ultrastructural features of the endometrial epithelium in the presence of a mole include intramitochondrial crystals and oval electron-rare intranuclear bodies with a fine fibrillar structure. Similar juxtanucleolar bodies are found in association with certain viral diseases.

the decidua has dealt almost exclusively with experimental animals, mainly rodents, 11 and has provided only isolated fine-structure details of the human decidua, 3 • 5 · 13 often incidental to a discussion of the trophoblast. This electron microscopic study of the human decidua was undertaken to bridge the gap between conventional histologic and histochemical studies and recent biochemical investigations. Although comparative studies may suggest biological principles, the great diversity in structure and function of the decidua among even closely related species requires knowledge of the human decidua to draw valid conclusions about such phenomena as implantation and its inhibition in man. In this discussion "decidua" refers to the pregnant endometrium (epithelium and stroma), some of which is shed at parturition, and a "decidual cell" refers specifically to the transformed polygonal glycogen-containing stromal cell. According to these definitions, even the closely related macaque,

F 0 R M A T I 0 N 0 F decidua IS a unique example of rapid histogenesis m a normal adult mammal. Therefore, it is surpnsmg that a search of the world's literature failed to uncover a previous systematic ultrastructural study of this tissue in the human being. The absence of such studies is even more startling in vie\v of the numerous published papers dealing with ultrastructure of the human placenta and cycling nonpregnant endometrium. Electron microscopy of From the Department of Obstetrics and Gynecology of The Abraham Lincoln School of Medicine, University of Illinois at the Medical Center. Supported in part by Grant HD04152 from the National Institutes of Health. Presented by invitation at the Eighty-fourth Annual Meeting of the American Association of Obstetricians and Gynecologists, Hot Springs, Virginia, September 6-8, 1973. Reprint requests: Dr. Ralph Wynn, Department of Obstetrics and Gynecology, University of Illinois Medical Center, 840 S. Wood St., Chicago, Illinois 60612.

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Fig. l. Endometrial stromal (predecidual) cells from Day 25 of the menstrual cycle. A homogeneous cytoplasmic process ( P) of one cell indents the cytoplasm of a typical fibrocytic element) (F). Abundant collagen (arrow) surrounds the stromal cells. (Negative 14,331. Original magnification x 16,500.)

which forms an epithelial plaque rather than an extensive stromal reaction, is a poor model for the study of human decidual development. Other animals such as carnivores and ungulates have no true decidua, in the sense of maternal tissue that is shed with the placenta. To eliminate problems of histogenetic identification, this study was confined to the parietal and capsular deciduas. The basal decidua and the complex elements that are often confused with trophoblast have been considered elsewhere. 12 ' 14 Analysis of the development of the decidua from the premenstrual endometrium throughout pregnancy was attempted to reveal the ultrastructural adaptations that permit the decidua to effect placental separation, restrict

trophoblastic invasion, and nourish the conceptus. Finally, comparison of the ultrastructural changes in normal pregnancy with those in hydatidiform mole was designed to provide information about the genesis of trophoblastic growths and the significance of their effects on the human endometrium.

Material and methods Samples of premenstrual (Day 25) endometrium were obtained by biopsy. Endo· metrial tissues were removed during interruption of pregnancy by curettage or hysterotomy at the following stages of gestation: 6, 8, 10, and 12 weeks. Decidual tissues of patients prematurely delivered at 22 and 28 weeks were compared with those delivered at term. The decidual reaction was

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Fig. 2. Parietal decidua from the sixth week of gestation. Golgi complexes (G), endoplasmic reticulum ( R), and glycogen (thin arrow) are increasing in prominence. A cilium (thick arrow ) is seen in a stromal cell. (Negative 14,631. x6,000. )

studied in a Fallopian tube removed for ectopic pregnancy. Endometrial tissues from normal third-trimester and molar pregnancies were obtained during cesarean section and hysterotomy, respectively. Fragments of tissues were fixed immediately in 3.4 to 5 per cent glutaraldehyde in phosphate buffer at 700 to 850 mOsm., pH 7.2 to 7.5, and 0° to 5° C. After rinsing in phosphate buffer raised to the same os-

molality with sucrose, the specimens were postfixed in buffered 1 per cent osmium tetroxide. Tissues were dehydrated in an ascending ethanolic series, transferred to propylene oxide, and embedded in Araldite 502. Thick (2 p. ) sections were examined with phase-contrast microscopy. Representative areas of stroma and epithelium were stained with toluidine blue for morphologic orientation . Thin sections were prepared

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Fig. 3. Decidua parietalis at 6 weeks, showing parallel channels of rough-surfaced endoplasmic reticulum (R) and smooth-contoured nucleus (N) . Lysosome-like bodies (thin arrow) are found in projections from the surface of the decidual cell into the intercellular space. Fibrillar material (thick arrow) is condensed around the surface of the stromal cell. (negative 4,085. x15,000.)

with an ultramicrotome* and stained with uranyl acetate and lead citrate. Sections were examined with an electron microscopet at initial magnifications of 900 to 20,000.

Observations Predecidual changes. The ultrastructural features of the human decidua are foreshadowed in the stromal cells of the endometrium during the second postovulatory *Porter-Blum. tEM-9 or -9A, Carl Zeiss, Inc. , #4 Fifth Ave ., New York, New York 10018,

week. The epithelial cells have convoluted plasma membranes, small slender mitochondria, large Golgi complexes, and relatively sparse microvilli. The major changes in endometrial development affect the stroma. Many of the cells are typical fusiform fibrocytic elements with moderately well-differentiated cytoplasm. The laminae externae of these stromal elements are well developed. Between stromal cells, tight junctions (maculae, zonae, and fasciae occludentes ) but not true desmosomes are found. In addition, gap junctions may form between processes of

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Fig. 4. UeCJdua parietaiis at 8 weeks, showing epithelium (B) and stroma ( ;';). The complex epithelium is packed with endoplasmic reticulum ( R) and lipid droplets ( L). The stromal cells have a fibrocytic appearance with numerous slender mitochondria. (Negative 9,466. x4,500.)

the same cell. Unusual features of predecidual cells that are shared by the early true decidua are deep projections of one cell into the cytoplasm of another (Fig. 1 ) . These invaginations of homogeneous cytoplasm resemble inclusions at low magnification, but intercellular membranes are seen Yvith high resolution. "'\bundant collagen and fibrinoid, some of which appears to be intracellular, surround the stromal cells. Ciliated stromal elements are found in the secretory phase of the menstrual cycle as well as in true decidua (Fig. 2). Decidua) development. During the first few months of gestation, the decidua parietalis and the decidua capsularis resemble each other in detail. Apparent secretory activity is maximal during the first I 0 weeks. A transitional phase occurs during the tenth to twelfth weeks, and ultrastructural evi-

dence of diminution in actiVIty follows during the twelfth to fourteenth weeks. The capsularis usually disappears by the fourteenth week. The decidual cell of early pregnancy is surrounded by fine reticular fibers and occasionally by fibrinoid. Its cytoplasm is generally homogeneous and moderately \vel! developed. Some of the smaller cells appear better differentiated. The typical stromal cell has a large pale nucleus. The epithelium contains sparsely granular endoplasmic reticulum, most of it in vesicular form. Fine filaments course throughout the epithelium, occasionally forming a terminal web beneath the microvillous border. However, these fine filaments do not form bundles, as they do in the connective tissue. The epithelial Golgi complexes are moderately well developed, and the mitochondria are

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Fig. 5. Detail of a parietal decidua cell at 8 weeks. Stroma (S) and epithelium (E) are intimately apposed. Numerous granules (arrow) are found in the extracellular space between the stroma and epithelium. Prominent basal lamina (B) is shown. (Negative 5, 176. x50,000.)

slender and dense. True desmosomes are found between adjacent epithelial cells. By 6 weeks, the endoplasmic reticulum of the stromal cell has become maximally differentiated. The nucleus remains smooth, and the collagenous extracellular material is compact. Unusual cytoplasmic features at this stage are projections of the plasma membrane containing extremely osmiophilic bodies (Fig. 3) . Many of these bodies look like lysosomes and appear to be associated with the formation of extracellular coats. The delicate mitochondria remain small and numerous. A few of the better differentiated elements resemble fibroblasts . The low ratio of nuclear to cytoplasmic size explains the apparent absence of nuclei from many sections of decidual cells. By 8 weeks, the granular endoplasmic

reticulum of the epithelial cell is maximally developed (Fig. 4). Mitochondria are small and most abundant above the nucleus, which appears to be invaginated by numerous lipoprotein granules. The microvilli are low and scanty. The Golgi complexes are moderately well developed and distended. The basal plasma membrane is highly convoluted. True desmosomes and tight junctions are prominent. At 8 weeks, many of the stromal cells lie very close to the basal lamina, and some seem to share certain ultrastructural features with the epithelium. The stromal cells near the capillaries are best developed. A few of the cells form extracellular spaces between the decidua and basal lamina that contain granules, some of which are consistent morphologically with procollagen (Fig.5).

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Fig. 6. Decidua capsularis at 10 weeks. The stromal cells, identical with those of the parietal decidua at this stage, contain slender mitochondria and numerous small vesicles of endoplasmic reti culum . (Negative 9, 17'! . x4500.)

At 10 weeks, the capsular decidua, which still resembles the parietal decidua, begins to regress (Fig. 6) . The epithelium is laden with lipid and glycogen but contains relatively few organelles. Microvilli are found in reduced numbers. The stroma is still rich in granular endoplasmic reticulum, much of it in vesicular form, but more free ribosomes are noted at this stage. A few stromal cells are ultrastructurally complex, resembling basal decidual cells, but larger numbers a re degenerate. Some of the fusiform decidual cells have cytoplasmic streamers. By 12 weeks, even in the parietal decidua,

degeneration of the epithelium is extensive (Fig. 7) . There is no further ultrastructural differentiation of the stroma beyond this stage. Processes of decidual cytoplasm make very close contact with capillary endothelium . By 22 weeks, degeneration of the fusiform parietal stromal cells as well is noted, along with deposition of considerable basement membrane-like material (Fig. 8) . Some of this deposit appears to be intracellular but more likely represents indentation of the cytoplasm from without. At 28 weeks, the decidual cell contains

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Fig. 7. Decidua parietalis at 12 weeks. The stromal cell has already reached maximal ultrastructural development. The cytoplasm contains numerous mitochondria (M) , Golgi complexes (G), and vesicles of endoplasmic reticulum (R) . Processes of decidual cytoplasm make intimate contact with capillary endothelium (C) . (Negative 5,116. x l5,000 .)

short fragments of endoplasmic reticulum and Golgi complexes with prominent condensed tubules (Fig. 9). The mitochondria remain small and delicate. Lamina externa, collagen, and fibrinoid are prominent. Fine fibrillar material appears to be discharged into the extracellular space from the stromal cytoplasm. A terminal web forms near the surface, as in epithelium. At term, a thin layer of epithelium may

be distinguished in certain portions of the decidua parietalis (Fig. 10). The epithelial cells have low microvilli and prominent deposits of glycogen. Many of the stromal cells that lie close to the epithelium are surrounded by a lamina externa that closely resembles the basal lamina of the epithelium. The cytoplasm of the parietal decidual cell is homogeneous and not highly differentiated. Such an element cannot easily be con-

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Fig. 8. Decidua parietalis at 22 weeks. Degeneration of the fusiform elements (F) around the vessles (V) is already in evidence. Fibrinoid basement membrane-like material (arrowi surrounds decidual cells. (Negative 3,649. x4,500.)

fused with trophoblast, as can a complex basal decidual cell. Ectopic decidua. In the Fallopian tube at 6 weeks' gestation, the stromal reaction is ultrastructurally indistinguishable from that of the endometrium at an equivalent stage of intrauterine gestation (Fig. 11). Golgi complexes, granular endoplasmic reticulum, and mitochondria, many \Vithout obvious crests, are seen. Collagen is abundant, and cytoplasmic lipoprotein granules are prominent.

Hydatidiform mole. The endometrial epithelium associated with hydatidiform mole is highly complex ultrastructurally, although the stroma resembles rather closely that of the endometrium associated with normal pregnancy (Fig. ] 2) . In certain stromal cells, whorls of endoplasmic reticulum, perhaps precursors of myelin figures, are prominent (Fig. 13). The ultrastructural features

of the epithelium are consistent with a hypersecretory interpretation of the AriasStella reaction (Fig. 14) . The nuclei are

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Fig. 9. Decidua parietalis at 28 weeks. The prominent features of the cytoplasm are short fragments of endoplasmic reticulum ( R), numerous delicate mitochondria ( M) , and Golgi complexes (G) with abundant tubules. (Negative 3,521. xl5,000.)

large and irregular, but the chromatin is rather evenly distributed. Microvilli are low, and zonae adherentes are prominent. Parallel channels of granular endoplasmic reticulum, large mitochondria, and microtubules are abundant, particularly in supranuclear regions. The cytoplasm is crowded with many small oval mitochondria and lipoprotein granules. Some of the mitochondria lack crests, and others have prominent transverse crests. The Golgi complexes have numerous

parallel tubules, and the plasma membranes are complexly interdigitated. Previously undescribed features of the epithelium include electron-rare intranuclear bodies with a fine filamentous structure and intramitochondrial crystalloids. The nuclear membrane is tortuous, and the prominent nucleolus is skeinlike. Near the nucleolus are electron-rare fields about 2 Jl. in diameter. Within these fields are round or oval structures containing numerous fine filaments

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Fig. 10. Decidua parietalis at term. Epithelial cells (E) of varying density contain patches of glycogen (arrow). Typical stromal cells (S) lie extremely close to the epithelium. (Negative 758. x4,500.)

(Fig. 15 ). Within the mitochondria, crystalloids about 0.5 1-'- in largest diameter appear to lie between the crests (Fig. 16). Comment

The premenstrual endometrium. In the normal menstrual cycle, cellular growth in response to estrogen ceases at or shortly after ovulation. 10 • 16 • 1 7 Thus, progesterone appears to brake the estrogen-induced endometrial proliferation. 1 3 With few exceptions, 10 • 11 ultrastructural studies of the hu-

man endometrium have focused on the epithelium. As a result, the changes leading to development of true decidua have not been carefully documented. Because of the wide differences in patterns of endometrial development among even closely related mammalian species, comparative studies required confirmation by detailed observations of human decidual formation . Even the baboon6 fails to form the nucleolar channel systems and giant mitochondria that are characteristic of the postovulatory human

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Fig. 11. Decidual reaction in a Fallopian tube removed for ectopic pregnancy at 6 weeks' gestation. The stromal cells are indistinguishable from those of intrauterine pregnancy of the same gestational age. Lipid granules (L) and lysosomes (arrow ) are seen. (Negative 12,05 7. x !S,OOO.)

endometrium. Wynn and Woolley 17 found that after the first few postovulatory days there was no further increase in Golgi complexes or granular endoplasmic reticulum in the human endometrial epithelium. Only in the third week of the menstrual cycle do the stromal changes that culminate in the decidual reaction appear. Specifically, on Day 22, broader contacts betwen the decidual cells are found. On Day 23, the individual

stromal cells enlarge and the cytoplasm is packed with glycoprotein. On Day 25, the ground substance is more compact, and the first signs of regression occur if the patient is not pregnant. If pregnancy occurs, endometrial development progresses to form true decidua. In their study of the stromal cells during the second half of the cycle, Wienke and co-workers 10 noted a "margin of condensation," which we interpret to be similar

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Fig. 12. Low-power electron micrograph of endometrial epithelium associated with hydatidiform mole. The epithelium (E) contains large irregular nuclei (N) with prominent electron-rare patches. The cytoplasm is filled with lipid droplets and small mitochondria. (Negative 11,248. x4,500.)

to a lamina externa. The cytoplasmic membranes may disappear focally to form margins of condensation of tropocollagen, or the precursor of collagen may be actively secreted into the extracellular space, where polymerization occurs. The mechanism of formation of collagen remains to be clarified, but the association with increased numbers of lysosomes suggests autolysis as one factor. Whatever its origin, the collagen very likely functions to support the decidua, and the accumulated glycogen and glycoprotein nourish the early conceptus. Decidualization. The human blastocyst attaches to the endometrium between the fifth and sixth days of age. The stromal cells show some enlargement by the seventh

day, increasing in size and gradually differentiating into large epithelioid polyhedral elements rich in glycogen and lipid. The ovum is completely embedded by the eleventh day of age. Regardless of whether implantation occurs, stromal edema is followed within 3 to 4 days by a decidual reaction, beginning around the spiral arterioles and spreading to involve the upper two thirds of the endometrium. If implantation occurs, the reaction persists. If it does not, the endometrium sloughs during menses. All Mullerian tissues respond similarly to the presence of trophoblast. On the one hand, in ectopic pregnancy the reaction in the subepithelial connective tissue of the tube is similar to that in the endometrium. On the

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Fig. 13. Endometrial stromal cell associated with hydatidiform mole. Whorls of endoplasmic reticulum, perhaps indicating early degeneration, are seen in an otherwise unremarkable cytoplasm. Proximity of the capillary (C) is shown. (Negative 11,256. xlS,OOO.)

other hand, there is a decidual reaction in the connective tissue of the oviduct in cases of intrauterine pregnancy. Decidualization in the human being is a spontaneous phenomenon that occurs in the presence of a functioning corpus luteum.

The predecidual, or pseudodecidual, reaction in the second half of the normal menstrual cycle persists as long as the trophoblast maintains luteal secretion, through its early and prolonged production of chorionic gonadotropin. In brief, the decidual cells

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Hg. 14. Detail of endometrial epithelium associated with hydatidiform mole. The cytoplasm is packed with mitochondria (arrow), many without crests. The ultrastructural features of the Arias-Stella reaction are shown. (Negative 11,240. x12,500.)

display ultrastructural features foreshadowed on Day 24 of the menstrual cycle: increase in size, decrease in the ratio of nucleus to cytoplasm, a nd further development of typical slender mitochondria and elaborate Golgi complexes. Lawn and co-workers" studied the junctions between decidual cells and concluded that they connected processes of the same cell rather than those between adjacent cells.

Since these gap junctions in the human decidua could not form a significant barrier, they reasoned that the extracellular matrix was a more important mecha nical barri er in human placentation. The dense extracellular material surrounding the mature human decidual cell is ultrastructurally similar to the basal lamina of the epithelium. The osmiophilic membrane-bound inclusions associated with well-developed endoplasmic reticu-

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Fig. 15. Detail of endometrial epithelial nucleus in molar pregnancy. Skeinlike nucleolus(NO) and electron-rare field containing an oval area with fine filaments (arrow) are shown. (Negative I 1,243. x15,000.)

lum and intracellular fibrils suggest intracellular secretion of this material or intracellular digestion, or a combination of these two processes. In the basal decidua, Enders 3 and Wynn 12 described, in addition, occasional tight junctions and desmosome-like attachments. The Arias-Stella reaction. In 1954, Arias-Stella 1 described atypical endometrial changes associated with the presence of

chorionic tissue. These changes have been found in ectopic pregnancy but have also been identified in normal intrauterine pregnancy, abortion, 2 and even more prominently in the invasive mole. 15 Arias Stella1 described " . . . irregular nuclear hypertrophy of isolated glandular cells usually accompanied by marked proliferative activity, together with simultaneous secretion changes in variable degree."

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Fig. 16. Detail of cytoplasm of endometrial epithelial cell associated with hydatidiform mole. Numerous mitochondria (M) and well-developed Golgi complexes (G) are shown. Some mitochondria contain crystalloid structures (arrow). (Negative 11,244. Original magnification x50,000.)

The present study of the endometrium associated with benign hydatidiform mole revealed several heretofore undescribed nuclear changes, the significance of whi<:h remains to be ascertained. The hypersecretory reaction of the endometrial epithelium to the presence of neoplastic trophoblast is either a direct response to chorionic gonadotropin or an indirect effect mediated through secretion of estrogen and progesterone. Thrasher and Richart 8 point out that the Arias-Stella reaction differs ultrastructurally from carcinoma of the endometrium in that the distribution of chromatin is more homogeneous in the Arias-Stella endometrium. Wagner and Richart~ conclude that polyploidy explains the abnormal appearance of

the nuclei. The excess secretion of hormones beyond that of the normal menstrual cycle was offered as an explanation of the increased size of the nucleus, the hyperchromatism, the polymorphism, and the loss of polarity. The unique features of the endometrium associated with the mole in this study are the intramitochondrial crystalloids and the fibrillar intranuclear electron-rare bodies. Nakao and co-workers 7 described cytoplasmic endometrial crystalloids that they considered to be progesterone-specific protein crystals. These structures measured 0.2 to 0.6 by 4 to 7 J-L in the rabbit and 0.2 to 0.5 by 1 /! in the human being. These crystalloids differed from the structures described here

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in their larger size and their free location in the cystoplasm. Krishnan and co-workers 4 described eceltron-rare intranuclear bodies 1 to 2.5 p. in diameter in a variety of animal tissues in cases of viral infections, Shope papillomas, leukemia, and Hodgkin's disease. The structures were thought to be proteins or histones rather than nucleoproteins and were found in certain normal tissues as

REFERENCES

1. Arias-Stella, J.: Arch. Pathol. 58: 112, 1954. 2. De Brux, J., and Ancla, M.: AM. J. 0BSTET. GYNECOL. 89: 661, 1964. 3. Enders, A. C.: Am. J. Anat. 122:419, 1968. 4. Krishnan, A., Uzman, B. G., and HedleyWhyte, E. T.: J. Ultrastruct. Res. 19: 563, 1967. 5. Lawn, A. M., Wilson, E. W., and Finn, C. A.: J. Reprod. Fertil. 26: 85, 1971. 6. MacLennan, A. H., Harris, J. A., and Wynn, R. M.: Obstet. Gynecol. 38: 359, 1971. 7. Nakao, K., Meyer, C. J., and Noda, Y.: AM. J. 0BSTET. GYNECOL. 11: 1034, 1971. 8. Thrasher, T. V., and Richart, R. M.: AM. J. 0BSTET. GYNECOL. 112: 113, 1972. 9. Wagner, D., and Richart, R. M.: Arch. Pathol. 85: 475, 1968.

Discussion DR. RlCHARD F. MATTINGLY, Milwaukee, Wisconsin. This ultrastructural study of the pregnant endometrium, including both epithelium and stroma, was carried out in order to document those adaptation characteristics which permit decidual tissue to: ( 1 ) effect placental separation, (2) restrict trophoblast invasion, and ( 3 ) nourish the conceptus. Unique findings in the decidua associated with molar trophoblastic disease also have been described in this presentation. The author has documented skillfully the ultrastructural changes of the stromal cell at the sixth, eighth, tenth, twelfth, twenty-second, and twenty-eighth weeks of gestation and compared these changes with the term placenta. This is one of the first studies to define systematically the intracellular changes of the decidual cell that transpire throughout the three trimesters of pregnancy. The study clearly illustrates the intrastructural development of the decidua and demonstrates the hypersecretory reaction of the endometrial epithelium, typical of the AriasStella reaction. This latter observation is an im-

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well. The size and number of these bodies, which were considered to be possibly of nucleolar origin, were related to altered metabolism of the nucleus. These ultrastructural findings by no means prove or even strongly suggest a viral origin of hydatidiform mole. However, they are consistent with such a hypothesis, which must, of course, be tested by more direct techniques.

10. Wienke, E. C., Jr., Cavazos, F., Hall, D. G., and Lucas, F. V.: AM. J. 0BSTET. GYNECOL. 102: 65, 1968. 11. Wynn, R. M.: Fertil. Steril. 16: 16, 1965. 12. Wynn, R. M.: AM. J. 0BSTET. GYNECOL. 97: 832, 1967. 13. Wynn, R. M.: Cellular Biology of the Uterus, New York, 1967, Appleton-Century-Crofts, Inc., p. 475. 14. Wynn, R. M.: AM. J. 0BSTET. GvNECOL. 114: 339, 1972. 15. Wynn, R. M., and Harris, J. A.: AM. J. 0BSTET. GYNECOL. 99: 1125, 1967. 16. Wynn, R. M., and Harris, J. A.: Fertil. Steril. 18: 632, 1967. 17. Wynn, R. M., and Woolley, R. S.: Fertil. Steril. 18: 721, 1967.

portant one to note as a normal component of human pregnancy and should lay to rest the prolonged debate regarding the nature of these histologic changes concerning secretion or degeneration. Dynamic biological studies are still needed in order to document clearly the contributions of the decidua and trophoblast to the phenomena of placental separation and restriction of trophoblastic invasion. The present paper emphasizes the parallel changes occurring in both endometrial and Fallopian tube epithelium at comparable periods of gestation. The phenomenon of trophoblast separation from endometrial cells was shown in a tissue culture environment from our laboratory in which invading trophoblastic tumor cells were explanted with the stroma of endometrium during pregnancy. Under these circumstances, a markedly exaggerated interaction results in the development of an "acellular zone" reminiscent of the fibrinoid layer of the placenta. Electron microscopic examination of single trophoblastic cells stained with colloidal iron reveals a distinct mucopolysaccharide "coating"

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of the microvilli and cell surfaces of these cells. In light microscopy, periodic acid-Schiff stains and diastase digests confirm the ''cell surface coatings" of individual trophoblastic tumor cells. A similar study conducted during the midtrimester of pregnancy by Azab and coauthors 1 demonstrated both histochemical and electron microscopic changes in the human placenta with an inseparable relationship of collagen and fibrinoid to decidual cells. Evidence from this study sug.gests that there is active secretion of specific mucoprotein-mucopolysaccharide complexes by the decidual cell which may providr the prerequisite immunologic tolerance between mother and fetus which is necessary for maintenance of this unique homograft. To date, electron microscopic studies are absent in cases of placenta accreta to determine the exact role of this mucopolysaccharide material, termed ''fibrinoid,'' in controlling trophoblastic invasion of the uterus. The findings of intranuclear fibrils in endometrium associated with benign trophoblastic tumors is of great interest. Present studies in our laboratory and studies by Kalter and co-workers 1 in the Virus Cancer Program of the National Cancer Institute have identified "C"-type viral particles in primate and human placentas. However, any relationship of these particles to trophoblastic tumors remains to be identified. This study is of unquestionable value in the field nf human reproduction, and the author deserws great credit for his diligent approach to thr function of the decidual cell. However. two questions remain unanswered from the original plan of study. What changes have been oh-

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~erved in this investigation concerning the role (lf the decidual cell with placental separation, and how does this cell restrict trophoblastic invasion:' Although the author has addressed his research to these questions in previous studies, is there new information from the present investigation that assists in the resolution of these basic biological enigmas!

REFERENCES

I. Azab, N., Okamura, M. D., and Beer, A.: Obstet. Gynecol. 40: 186, 1972. ~. Kalter, S. S., Helmke, R. ]., Panigel, M., Heberling, R. L., Felsburg, P. J., and Axelrod, L. R.: Science 179: 1132, 1973.

DR. WYNN (Closing). The mechanisms by which the decidua appears to restrict trophoblastic invasion vary widely among species. In rodents, tight junctions between stromal cells an~ probably of mechanical importancf', whereas in the human decidua the stromal cells form gap junctions, which could not be expected to resi~t trophoblastic invasion. A more important anatomic adaptation of the human decidua appears more likely to be the deposition of dense fibrinoid-like extracellular mucopolysaccharides. At one point, we suggested that the increased penetration of trophoblast in cases of invasive hydatidiform mol<' might reflect a less extensive deposition of extracellular mucopolysaccharide. I am no longt>r convinced of the validity of that hypothesis, hut recPnt <·vidence suggest' that glycoprotein deposits on the trophoblast may play a crucial role in forestalling immunologic rejection of the trophoblast by maternal tissues.