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Fetal microcirculation of abnormal human placenta I. Scanning electron microscopy of placental vascular casts from small for gestational age fetus
BASIC SCIENCE SECTION
Fetal microcirculation of abnormal human placenta I. Scanning electron microscopy of placental vascular casts from small for gestational age fetus Mary M. L. Lee, Ph.D., and Ming-neng Yeh, M.D. New York, New York The latex injection-eorrosion cast technique coupled with scanning electron microscopy was applied to the study of small for gestational age placentas. The main ultrastructural changes observed in the fetal vasculature of small for gestational age placentas were less branching of arteries and veins, capillaries with variable diameters, and many capillary bud projections; also numerous "H"-shaped anastomoses were present in the capillary network. The corrosion/cast technique approach enabled us to visualize the three-dimensional distribution of arteries, veins, and capillary network as well as the ultrastructural alterations among the various-sized vessels in the abnormal placenta. Light microscopic examination of these placentas revealed numerous syncytial knots and fibrinoid deposits pressing on arteriolar and capillary vessels at different locations. We suggest that the numerous capillary buds and anastomoses are characteristics of neovascularization and may be a compensatory phenomenon of the capillary network. This information could shed new light on alterations of physiologic functions of the pathologic placenta. (AM J OSSTET GVNECOL 1986;154:1133-9.)
Key words: Fetal microcirculation, placenta, scanning electron microscopy, intrauterine growth retardation, neovascularization
Since the placenta is involved in the basic functions of metabolism, transfer of nutrients, exchange of gases, and endocrine secretion, it is essential for maintaining pregnancy and promoting normal embryonic-to-fetal development. An examination of the three-dimensional distribution and ultrastructural appearance of normal fetal placental vasculature helps in understanding these physiologic functions. The morphologic characteristics of fetal blood vessels in the human placenta have recently been examined by the latex injectioncorrosion cast technique coupled with the scanning electron microscope.': The latex injection-corrosion cast technique, when applied to the study of the pathologic small for gestational age (SGA) placenta, revealed new data on the ultrastructural appearance of abnormal fetal-placental
From the Department of Physiology and the Department of Obstetrics and Gynecology, Columbia University College of Physicians and Surgeons. Receivedfor publication September 27, 1985; revised December 17, 1985; acceptedJanuary 28, 1986. Reprint requests: Dr. Mary M. L. Lee, Department of Physiology, Columbia University College ofPhysicians and Surgeons, 630 West 168th St., New York, NY 10032.
vasculature. The purpose of the present study was to elucidate the ultrastructural changes in the fetal-placental vasculature that are present in the pathologic SGA placenta by scanning electron microscopy.
Material and methods Seven human placentas from mothers who were delivered of SGA babies (less than tenth percentile in weight at birth) were used for the study (Table I). The same latex injection-corrosion cast technique used in the study of normal human placentas was applied to this investigation.' The umbilical cord was threaded by a small catheter in the umbilical artery and vein and provided a route of entrance for the injection medium and a route of egress. After catheterization, the placenta was suspended in water and then flushed with warm saline at 50 mm Hg injection pressure. This was followed by an injection of 70% Cementex (Cementex Corp., New York, NY 10013). After polymerization by air drying, the latex inside the blood vessel hardened into a vascular cast. For further hardening, tissue was left in 10% formalin for 2 weeks. Then the placental tissue was macerated by treatment with sodium hypochlorite. The resulting vascular cast was air dried and 1133
1134 Lee and Yeh
May, 1986 Am J Obstet Gynecol
Fig. 1. a, Low-magnification scanning electron micrograph of cast from fetus with intrauterine growth retardation. An arteriole is shown on the right (A) leading into a fairly dense capillary network (C) located on the left. Bar = 100 urn. b, Scanning electron micrograph taken at high magnification of the capillary vessels in the placenta of an SGA fetus. Several H-shaped anastomoses are shown. Bar = 10 urn.
Table I. Clinical information Gravidity and parity
Patient
Mode of delivery
c.L.
24 27 27 27 37 20
G2PO GIPO G5P2 G6P4 G2P2 GIPO
40 37'/2 38V2 38 37 31
Female Female Female Female Female Male
2500 1500 2040 2250 2400 1000
Cesearan Cesearan Vaginal Vaginal Cesarean Cesarean
section section
6-9 9-9-9 8-9 8-9-9 8-9 7-8-8
J. H.
37
G3PO
39V2
Female
2210
Cesarean section
8-8-8
B. M. M.R. L. F. M.K. S. F.
CST = Contraction stress test.
section section
Complications Oligohydramnios Hypertension
Severe preeclampsia Meconium, positive CST
Scanning electron microscopy of SGA placental casts
Volu me 154 Nu mber 5
1135
Fig. 2. Latex cast of the capillary network in the placenta of an SGA fetus. Note "the numerous anastomoses and the presence of frequent vascular bud projections (arrows). Bar = 10 urn,
Fig. 3. Scanning electron micrograph taken at low magnification of latex cast from fetal-placental vasculature in a fetus with intrauterine growth retardation. The central vessel is a vein with collecting venules, The vein has very few branches. Bar = 100 urn. tea sed into sma ll pie ces for examination by scanning electron microscop y. The details of th e injection -corrosion cas t tech nique have been d escribed in previou s articles."! For exam ination by scanning elect ron microscopy, the casts were attac hed to aluminum stubs by silver conductive paint and the n coated with go ld- pallad iu m in a Hummer I Sputterer (Anatech , Ltd. , Alexandria,
VA 2231 0). On com pletio n of the preparat ions , th e re sulting vascular casts were viewed and ph otographed on a JEOL- 25 scan ning elect ron micro scope at 15 kV and 0- to 45-d egr ee tilt. A sma ll porti on of placenta was removed for regular light microscop ic exam inat ion . T he placental tissue was put int o for ma lin and rem ained th ere for 2 weeks before processing for light microscopy. The sections for
1136
Lee and Yeh
May, 1986 Am
J Obstet Gynecol
Fig. 4. An area of chorionic villi from an SGA placenta shows a number of capillaries that have compressed or occluded lumina. Bar = 10 11m.
light microscopy were cut at 8 urn and stained with hematoxylin and eosin for observation and photography.
Observations Scanning electron microscopic observations of vascular casts from SGA placentas revealed the following alterations from normal morphologic features: the capillary network was more dense and contained many "H"-shaped anastomoses (Fig. 1 a and b); the capillaries were of variable diameters and contained numerous budding projections (Fig. 2); also less branching occurred among the arteries and veins than is found in normal human fetal-placental vessels (Fig. 3). Regarding the H-shaped capillary anastomoses, we found that they were numerous in the SGA placental microcirculation. The capillary network appeared more dense and more tortuous than that of the normal human placenta (Fig. I, a). Furthermore, the number offused loops was definitely increased over norma\. While the number of capillary anastomoses increased, the amount of arterial and venous branching was decreased as compared with that of the normal placenta. Scanning electron microscopy of whole-surface SGA placental tissue showed that large portions of the chorionic villi had occluded capillaries (Fig. 4) while a more normal-appearing villous capillary network area was seen elsewhere (Fig. 5). Also seen in the pathologic specimen were areas of fibrinoid-like material compressing portions of capillaries; many of the lumina of capillaries appeared collapsed or clogged. The umbilical cord from our SGA placenta was cut and processed for scanning electron microscopy. Fig. 6 shows a cross section of the cord with one patent umbilical vein and
two umbilical arteries. One of the arteries is patent while the other has an occluded lumen; this was observed in two of the three umbilical cords. Light microscopic observations of the pathologic placental tissue revealed that the chorionic villi had numerous syncytial knots (Fig. 7) on the villous surface. The cytotrophoblast was distinct while the syncytiotrophoblast was so thinly attenuated that it was not clearly seen. Fibrinoid deposits of variable size were observed around some villous capillaries. The villi themselves had a nonedematous appearance by light and scanning electron microscopy.
Comment Vascular anatomy of the abnormal placenta with fetal growth retardation has been investigated by many researchers.v" However, there have been no previous reports on vascular cast studies from abnormal placentas by scanning electron microscopy. This approach enabled us to visualize the three-dimensional distribution of arteries, veins, and capillary network in the pathologic placenta as well as the ultrastructural alterations among the various-sized vessels in the abnormal placenta. In addition, this approach could elucidate the alterations of the physiologic functions of the pathologic placenta. Scanning electron microscopic analysis of fetal placental vascular casts revealed the following new findings: The capillaries have frequent H-shaped anastomoses and numerous forming vascular buds. Both phenomena are involved in neovascularization and may represent efforts by the abnormal placental capillary network to increase its density so that the fetus will have an increased capacity for exchange of nutrients and
Scanning electron microscopy of SGA placental casts
Volume 154
1137
Number 5
Fig. 5. A sectio n of the cho rio nic villi fro m ano the r portion of the SGA placen ta showi ng a mo re norma l-ap pea ring cluster of ca pillaries with pat ent lumina, man y of them filled with red blood cells. B ar = 10 11m.
gases at the capillary level. Neovascu lar izatio n was similarly observed in vascular casts by scanning electron microscop y by Burger et al. 12 in the rat corneal capillary network after injury to the cornea by che mical cauterization . Afte r I day, the injec ted vascular cast showed formatio n of new vascular buds from cap illaries an d venules. After a lon g time, microsco py of the casts revealed that a rich an asto mosing capi llary network had formed from the earlier vascular buds. T he findings of Burger et al. are in conformity with our findings from placental capillary casts with regard to the frequent H-shap ed anasto moses and fre quent vascular bud formation . Although the placental type of neovascularization was not the result of chemical injury, it had a very similar ultrastr uctural appearance. The stimulus for the cap illary ne ovascularization in a SGA placent a may be the initially reduced number of blood vessels in the placenta. This would impair nutrient and gaseo us excha nge for the develop ing fet us. Un der this condition , an increase in th e den sity of the capillary net wor k would result in improving placenta l-feta l excha nge. Presently we are wor king on an SGA an ima l model to ex plore th is hypot hesis. Fro m light micro scopic exa minations of th ese placentas, numerous syncytial knots and fibr inoid deposits (Kirby et a1. 2 ) were observed pressi ng on the arteriolar and capi llary vessels in the chorionic villi. These findings may accoun t for the var iability of capillary diameters an d some arteriolar const rictions as seen in th e lat ex casts by scanning electro n micro scop y. The cause of the fibr inoid deposits is not yet fully understood, but McCormick et al." and Fox J4 have sugg ested an im-
Fig. 6. Scanning electron micro gra ph of the umbilical cord from the placenta of an SGA fetus . The u mbilical vein (V) is dilated. while one of the umbilical arteries (A) is patent and the other artery (a) is occlude d . Bar = 1 mm . .
munologic source. Addi tiona l stu dies for furt he r exploration of this possible mechan ism may be warra nte d. Also seen by light micro scopy were hyperplasia of the cytotrophoblastic cells and a th inly attenuat ed syncytiotrophob lastic layer. This finding confirms the light microscopi c observatio ns of Van Der Veen and Fox. II Normally at term the cytotrop hob last can no t be so clea rly seen by ligh t microscopy; such large, distinct cells probably represent highl y active cells. Perhaps a genetic signal to these cells to red uce their functioning activity as term approached was lost or a time sequence of deve lopment was delayed. Ano ther possibility is that
1138 Lee and Yeh
May, 1986 Am J ObstetGynecol
Fig. 7. Light microgr aph of a section of the cho rionic villi from an SGA placent a sho wing numerou s syncytial knots (5) on the villi. The cytotro phoblast is d istinct. while the syncytiotrophoblast layer is not clearl y seen . (He matoxylin and eosin. x 225.)
the hyperplasia of the cytotrophoblastic cells could be a compensatory repair mechanism. Reduced branching of the large vessels (arteries and veins) was also observed, which may indicate that the re was an arrest of development during an early stage of placental blood vessel formation. With fewer large vessels, the volume of blood exchange to the gr owing fetus is decreased . The capillary network would the reb y tr y to compensate by forming more capillaries to help increase the amount of surface area for nutrient and gaseous exchange. Our vascular casts of the term placenta of course cannot reveal the reason for a possible interruption of early development. We favor the concept th at some event occurring earl y in blood vessel development caused alterations in the macrocirculation and microcirculation. Cross section of the umbilical cord from an SGA placenta showed that while the umbili cal vein appeared normal, the umbilical arteries had one patent and on e occluded lumen. A possible expl an ation of this peculiar ar rangement of one patent and one occluded umbilical artery is that there was a reduction of the amount of blood returning to the fetu s, and the fetu s became hypo volemic. As a consequence of thi s condition, both umbilical arteries cannot be ma intained in the patent condition. O~r present scanning electron micro scopic observation s oflatex casts provide an ultrastructural de scription of the alterations seen in the pathologic placentas, and light microscopy done in pa rallel supports the scanning electron microscope observations. In the future, transmission electron microsc op y of specific vascular areas of the pathologic placentas will provide further
ultrastructural data on the changes already observed by the other microscopic techniques. We wish to thank Dr. Shu Chien, Department of Physiology, for the kind use of his laboratory facilities for this study. We also thank Honor O'Sullivan and Gerard Norw ich for their fine technical assistance. REFERENCES I. I-Iabashi S, Burton GJ, Steven DH. Morphologic study of the fetal vasculature of the human term placenta: scanning electron microscopy of corrosion casts. Placenta 1983;4:41. 2. Kirby DRS, Billington WD, Bradbury S, Goldstein DJ. Anti gen barrier of the mouse placenta. Nature (London) 1964 ;204:548. 3. Lee ML, Yeh MN. Fetal circulation of the placenta. A compara tive study on human and baboon placenta by scanning electron microscop y of vascular casts. Placenta 1983;4:515. 4. Lee ML, Purkerson ML, Agate FJ, Dempsey EW. Ultrastructural changes in renal glomeru li of rats during experimentally induced hypertension and uremia. Am J Anat 1972; 135 :191. 5. Lee ML. Morphological effects of procaine amide on mou se kidne y as observed by scanning and transmission electron microscopy. Lab Inve st 1974;31:321. 6. Lee ML, Dempsey EW. Microcirculation of the rat placenta. AM J OBSTET GYNECOL 1976;126:495. 7. WigglesworthJS. The Langhans layer in late pregnancy: a histological study of normal and abnormal cases. J Obstet Gynaecol Br Commonw 1962 ;69:355. 8. Wigglesworth JS. Vascular anatomy of the human placenta and its significance for placent al pathology.J Obstet Gynaecol Br Commonw 1969;76:979. 9. Fox H. Fibrinoid necrosis of placental villi. J Obstet Gynaecol Br Commonw 1968;75:448. 10. Altshuler G, Russell P, Ermscilla R. The placental pathology of SGA infants. AM J OBSTET GYNECOL 1975; 121:35 1.
Volume 154 Number 5
11. Van Der Veen F, Fox H. The human placenta in idiopathic intrauterine growth retardation: a light and electron microscopic study. Placenta 1983;4:65. 12. Burger P, Chandler D, Klintworth GK. Corneal neovascularization as studied by scanning electron microscopy of vascular casts. Lab Invest 1983;48: 160.
Scanning electron microscopy of SGA placental casts
13. McCormick]N, Faulk WP, Fox H, Fundenberg HH. Immunohistological and elution studies of the human placenta.} Exp Med 1971;91:1. 14. Fox H. Histological abnormalities of the placenta. In: Fox H, ed. Pathology of the placenta. London: WB Saunders, 1978: 149.
Fetal microcirculation of abnormal human placenta II. Scanning electron microscopy of placental vascular casts from fetus with severe erythroblastosis fetalis Mary M. L. Lee, Ph.D., and Ming-neng Yeh, M.D. New York, New York The latex injection-corrosion cast technique coupled with scanning electron microscopy was applied to the study of severe Rh sensitization of the placenta. The main observed ultrastructural changes in fetal vasculature in this type of placenta were very similar to those of the small for gestational age placenta; large-sized vessels exhibited less branching than those of the normal placenta, and the capillary network showed numerous vascular buds and "H"-shaped anastomoses. In addition, in Rh-sensitized placentas, the capillary network was remarkably sparse, and the cast also revealed smooth-surfaced veins with distinct U shapes undulating through the placental stroma. These U-shaped veins may be the result of umbilical flow impedance with subsequent umbilical venous hypertension, which stretches the veins into undulating form. (AM J OSSTET GYNECOL 1986;154:1139-46.)
Key words: Fetal microcirculation, placenta, scanning electron microscopy, Rh sensitization, neovascularization The latex injection-corrosion cast technique coupled with scanning electron microscopy enables investigators to examine vascular casts from normal" 2 and abnormal human placentas in a three-dimensional form and to observe the ultrastructural changes in the fetal-placental vasculature. Several investigators':' have described, using light and transmission electron microscopy, the appearance of Rh-incompatible placentas. In part one data were presented on ultrastructural changes observed in vascular casts from the placentas of small for gestational age fetuses." This article will describe the vascular alterations that occurred in the placentas of fetuses with severe erythroblastosis fetalis and will discuss possible effects on the fetal-placental microcirculation resulting from such changes. To date, there has not been a scanning electron microscopic analysis of corrosion casts from Rh-incompatible placentas. From the Department of Physiology and the Department of Obstetrics and Gynecology, Columbia University College of Physicians and Surgeons. Received for publication September 27, 1985; revised December 17, 1985; acceptedJanuary 28, 1986. Reprint requests: Dr. Mary M. L. Lee, Department of Physiology, Columbia University College ofPhysicians and Surgeons, 630 West 168th St., New York, NY 10032.
Material and methods Four placentas from Rh-sensitized mothers who had given birth to babies with severe anemia were used for the study (Table I). The same latex injection-corrosion cast technique used in the normal human placenta was applied to this investigation." The umbilical cord was threaded by a small catheter into the umbilical artery and vein, providing a route of entrance of injection medium and a route of egress. After catheterization, the placenta was suspended in water, flushed with warm saline solution at an injection pressure of 50 mm Hg, and followed by an injection of 70% Cementex (Cementex Corp., New York, NY 10013). After polymerization by air drying, the latex inside the blood vessel hardened into a vascular cast. For further hardening the tissue was left in 10% formalin for 2 weeks. Then the placenta was macerated by treatment with sodium hypochlorite. The resulting vascular cast was air dried and teased into small pieces for examination in the scanning electron microscope as previously described.v" For scanning electron microscopic examination, the casts were attached onto aluminum stubs by silver conductive paint and then coated with gold-palladium in 1139
Fetal microcirculation of abnormal human placenta I. Scanning electron microscopy of placental vascular casts from small for gestational age fetus Mary M. L. Lee, Ph.D., and Ming-neng Yeh, M.D. New York, New York The latex injection-eorrosion cast technique coupled with scanning electron microscopy was applied to the study of small for gestational age placentas. The main ultrastructural changes observed in the fetal vasculature of small for gestational age placentas were less branching of arteries and veins, capillaries with variable diameters, and many capillary bud projections; also numerous "H"-shaped anastomoses were present in the capillary network. The corrosion/cast technique approach enabled us to visualize the three-dimensional distribution of arteries, veins, and capillary network as well as the ultrastructural alterations among the various-sized vessels in the abnormal placenta. Light microscopic examination of these placentas revealed numerous syncytial knots and fibrinoid deposits pressing on arteriolar and capillary vessels at different locations. We suggest that the numerous capillary buds and anastomoses are characteristics of neovascularization and may be a compensatory phenomenon of the capillary network. This information could shed new light on alterations of physiologic functions of the pathologic placenta. (AM J OSSTET GVNECOL 1986;154:1133-9.)
Key words: Fetal microcirculation, placenta, scanning electron microscopy, intrauterine growth retardation, neovascularization
Since the placenta is involved in the basic functions of metabolism, transfer of nutrients, exchange of gases, and endocrine secretion, it is essential for maintaining pregnancy and promoting normal embryonic-to-fetal development. An examination of the three-dimensional distribution and ultrastructural appearance of normal fetal placental vasculature helps in understanding these physiologic functions. The morphologic characteristics of fetal blood vessels in the human placenta have recently been examined by the latex injectioncorrosion cast technique coupled with the scanning electron microscope.': The latex injection-corrosion cast technique, when applied to the study of the pathologic small for gestational age (SGA) placenta, revealed new data on the ultrastructural appearance of abnormal fetal-placental
From the Department of Physiology and the Department of Obstetrics and Gynecology, Columbia University College of Physicians and Surgeons. Receivedfor publication September 27, 1985; revised December 17, 1985; acceptedJanuary 28, 1986. Reprint requests: Dr. Mary M. L. Lee, Department of Physiology, Columbia University College ofPhysicians and Surgeons, 630 West 168th St., New York, NY 10032.
vasculature. The purpose of the present study was to elucidate the ultrastructural changes in the fetal-placental vasculature that are present in the pathologic SGA placenta by scanning electron microscopy.
Material and methods Seven human placentas from mothers who were delivered of SGA babies (less than tenth percentile in weight at birth) were used for the study (Table I). The same latex injection-corrosion cast technique used in the study of normal human placentas was applied to this investigation.' The umbilical cord was threaded by a small catheter in the umbilical artery and vein and provided a route of entrance for the injection medium and a route of egress. After catheterization, the placenta was suspended in water and then flushed with warm saline at 50 mm Hg injection pressure. This was followed by an injection of 70% Cementex (Cementex Corp., New York, NY 10013). After polymerization by air drying, the latex inside the blood vessel hardened into a vascular cast. For further hardening, tissue was left in 10% formalin for 2 weeks. Then the placental tissue was macerated by treatment with sodium hypochlorite. The resulting vascular cast was air dried and 1133
1134 Lee and Yeh
May, 1986 Am J Obstet Gynecol
Fig. 1. a, Low-magnification scanning electron micrograph of cast from fetus with intrauterine growth retardation. An arteriole is shown on the right (A) leading into a fairly dense capillary network (C) located on the left. Bar = 100 urn. b, Scanning electron micrograph taken at high magnification of the capillary vessels in the placenta of an SGA fetus. Several H-shaped anastomoses are shown. Bar = 10 urn.
Table I. Clinical information Gravidity and parity
Patient
Mode of delivery
c.L.
24 27 27 27 37 20
G2PO GIPO G5P2 G6P4 G2P2 GIPO
40 37'/2 38V2 38 37 31
Female Female Female Female Female Male
2500 1500 2040 2250 2400 1000
Cesearan Cesearan Vaginal Vaginal Cesarean Cesarean
section section
6-9 9-9-9 8-9 8-9-9 8-9 7-8-8
J. H.
37
G3PO
39V2
Female
2210
Cesarean section
8-8-8
B. M. M.R. L. F. M.K. S. F.
CST = Contraction stress test.
section section
Complications Oligohydramnios Hypertension
Severe preeclampsia Meconium, positive CST
Scanning electron microscopy of SGA placental casts
Volu me 154 Nu mber 5
1135
Fig. 2. Latex cast of the capillary network in the placenta of an SGA fetus. Note "the numerous anastomoses and the presence of frequent vascular bud projections (arrows). Bar = 10 urn,
Fig. 3. Scanning electron micrograph taken at low magnification of latex cast from fetal-placental vasculature in a fetus with intrauterine growth retardation. The central vessel is a vein with collecting venules, The vein has very few branches. Bar = 100 urn. tea sed into sma ll pie ces for examination by scanning electron microscop y. The details of th e injection -corrosion cas t tech nique have been d escribed in previou s articles."! For exam ination by scanning elect ron microscopy, the casts were attac hed to aluminum stubs by silver conductive paint and the n coated with go ld- pallad iu m in a Hummer I Sputterer (Anatech , Ltd. , Alexandria,
VA 2231 0). On com pletio n of the preparat ions , th e re sulting vascular casts were viewed and ph otographed on a JEOL- 25 scan ning elect ron micro scope at 15 kV and 0- to 45-d egr ee tilt. A sma ll porti on of placenta was removed for regular light microscop ic exam inat ion . T he placental tissue was put int o for ma lin and rem ained th ere for 2 weeks before processing for light microscopy. The sections for
1136
Lee and Yeh
May, 1986 Am
J Obstet Gynecol
Fig. 4. An area of chorionic villi from an SGA placenta shows a number of capillaries that have compressed or occluded lumina. Bar = 10 11m.
light microscopy were cut at 8 urn and stained with hematoxylin and eosin for observation and photography.
Observations Scanning electron microscopic observations of vascular casts from SGA placentas revealed the following alterations from normal morphologic features: the capillary network was more dense and contained many "H"-shaped anastomoses (Fig. 1 a and b); the capillaries were of variable diameters and contained numerous budding projections (Fig. 2); also less branching occurred among the arteries and veins than is found in normal human fetal-placental vessels (Fig. 3). Regarding the H-shaped capillary anastomoses, we found that they were numerous in the SGA placental microcirculation. The capillary network appeared more dense and more tortuous than that of the normal human placenta (Fig. I, a). Furthermore, the number offused loops was definitely increased over norma\. While the number of capillary anastomoses increased, the amount of arterial and venous branching was decreased as compared with that of the normal placenta. Scanning electron microscopy of whole-surface SGA placental tissue showed that large portions of the chorionic villi had occluded capillaries (Fig. 4) while a more normal-appearing villous capillary network area was seen elsewhere (Fig. 5). Also seen in the pathologic specimen were areas of fibrinoid-like material compressing portions of capillaries; many of the lumina of capillaries appeared collapsed or clogged. The umbilical cord from our SGA placenta was cut and processed for scanning electron microscopy. Fig. 6 shows a cross section of the cord with one patent umbilical vein and
two umbilical arteries. One of the arteries is patent while the other has an occluded lumen; this was observed in two of the three umbilical cords. Light microscopic observations of the pathologic placental tissue revealed that the chorionic villi had numerous syncytial knots (Fig. 7) on the villous surface. The cytotrophoblast was distinct while the syncytiotrophoblast was so thinly attenuated that it was not clearly seen. Fibrinoid deposits of variable size were observed around some villous capillaries. The villi themselves had a nonedematous appearance by light and scanning electron microscopy.
Comment Vascular anatomy of the abnormal placenta with fetal growth retardation has been investigated by many researchers.v" However, there have been no previous reports on vascular cast studies from abnormal placentas by scanning electron microscopy. This approach enabled us to visualize the three-dimensional distribution of arteries, veins, and capillary network in the pathologic placenta as well as the ultrastructural alterations among the various-sized vessels in the abnormal placenta. In addition, this approach could elucidate the alterations of the physiologic functions of the pathologic placenta. Scanning electron microscopic analysis of fetal placental vascular casts revealed the following new findings: The capillaries have frequent H-shaped anastomoses and numerous forming vascular buds. Both phenomena are involved in neovascularization and may represent efforts by the abnormal placental capillary network to increase its density so that the fetus will have an increased capacity for exchange of nutrients and
Scanning electron microscopy of SGA placental casts
Volume 154
1137
Number 5
Fig. 5. A sectio n of the cho rio nic villi fro m ano the r portion of the SGA placen ta showi ng a mo re norma l-ap pea ring cluster of ca pillaries with pat ent lumina, man y of them filled with red blood cells. B ar = 10 11m.
gases at the capillary level. Neovascu lar izatio n was similarly observed in vascular casts by scanning electron microscop y by Burger et al. 12 in the rat corneal capillary network after injury to the cornea by che mical cauterization . Afte r I day, the injec ted vascular cast showed formatio n of new vascular buds from cap illaries an d venules. After a lon g time, microsco py of the casts revealed that a rich an asto mosing capi llary network had formed from the earlier vascular buds. T he findings of Burger et al. are in conformity with our findings from placental capillary casts with regard to the frequent H-shap ed anasto moses and fre quent vascular bud formation . Although the placental type of neovascularization was not the result of chemical injury, it had a very similar ultrastr uctural appearance. The stimulus for the cap illary ne ovascularization in a SGA placent a may be the initially reduced number of blood vessels in the placenta. This would impair nutrient and gaseo us excha nge for the develop ing fet us. Un der this condition , an increase in th e den sity of the capillary net wor k would result in improving placenta l-feta l excha nge. Presently we are wor king on an SGA an ima l model to ex plore th is hypot hesis. Fro m light micro scopic exa minations of th ese placentas, numerous syncytial knots and fibr inoid deposits (Kirby et a1. 2 ) were observed pressi ng on the arteriolar and capi llary vessels in the chorionic villi. These findings may accoun t for the var iability of capillary diameters an d some arteriolar const rictions as seen in th e lat ex casts by scanning electro n micro scop y. The cause of the fibr inoid deposits is not yet fully understood, but McCormick et al." and Fox J4 have sugg ested an im-
Fig. 6. Scanning electron micro gra ph of the umbilical cord from the placenta of an SGA fetus . The u mbilical vein (V) is dilated. while one of the umbilical arteries (A) is patent and the other artery (a) is occlude d . Bar = 1 mm . .
munologic source. Addi tiona l stu dies for furt he r exploration of this possible mechan ism may be warra nte d. Also seen by light micro scopy were hyperplasia of the cytotrophoblastic cells and a th inly attenuat ed syncytiotrophob lastic layer. This finding confirms the light microscopi c observatio ns of Van Der Veen and Fox. II Normally at term the cytotrop hob last can no t be so clea rly seen by ligh t microscopy; such large, distinct cells probably represent highl y active cells. Perhaps a genetic signal to these cells to red uce their functioning activity as term approached was lost or a time sequence of deve lopment was delayed. Ano ther possibility is that
1138 Lee and Yeh
May, 1986 Am J ObstetGynecol
Fig. 7. Light microgr aph of a section of the cho rionic villi from an SGA placent a sho wing numerou s syncytial knots (5) on the villi. The cytotro phoblast is d istinct. while the syncytiotrophoblast layer is not clearl y seen . (He matoxylin and eosin. x 225.)
the hyperplasia of the cytotrophoblastic cells could be a compensatory repair mechanism. Reduced branching of the large vessels (arteries and veins) was also observed, which may indicate that the re was an arrest of development during an early stage of placental blood vessel formation. With fewer large vessels, the volume of blood exchange to the gr owing fetus is decreased . The capillary network would the reb y tr y to compensate by forming more capillaries to help increase the amount of surface area for nutrient and gaseous exchange. Our vascular casts of the term placenta of course cannot reveal the reason for a possible interruption of early development. We favor the concept th at some event occurring earl y in blood vessel development caused alterations in the macrocirculation and microcirculation. Cross section of the umbilical cord from an SGA placenta showed that while the umbili cal vein appeared normal, the umbilical arteries had one patent and on e occluded lumen. A possible expl an ation of this peculiar ar rangement of one patent and one occluded umbilical artery is that there was a reduction of the amount of blood returning to the fetu s, and the fetu s became hypo volemic. As a consequence of thi s condition, both umbilical arteries cannot be ma intained in the patent condition. O~r present scanning electron micro scopic observation s oflatex casts provide an ultrastructural de scription of the alterations seen in the pathologic placentas, and light microscopy done in pa rallel supports the scanning electron microscope observations. In the future, transmission electron microsc op y of specific vascular areas of the pathologic placentas will provide further
ultrastructural data on the changes already observed by the other microscopic techniques. We wish to thank Dr. Shu Chien, Department of Physiology, for the kind use of his laboratory facilities for this study. We also thank Honor O'Sullivan and Gerard Norw ich for their fine technical assistance. REFERENCES I. I-Iabashi S, Burton GJ, Steven DH. Morphologic study of the fetal vasculature of the human term placenta: scanning electron microscopy of corrosion casts. Placenta 1983;4:41. 2. Kirby DRS, Billington WD, Bradbury S, Goldstein DJ. Anti gen barrier of the mouse placenta. Nature (London) 1964 ;204:548. 3. Lee ML, Yeh MN. Fetal circulation of the placenta. A compara tive study on human and baboon placenta by scanning electron microscop y of vascular casts. Placenta 1983;4:515. 4. Lee ML, Purkerson ML, Agate FJ, Dempsey EW. Ultrastructural changes in renal glomeru li of rats during experimentally induced hypertension and uremia. Am J Anat 1972; 135 :191. 5. Lee ML. Morphological effects of procaine amide on mou se kidne y as observed by scanning and transmission electron microscopy. Lab Inve st 1974;31:321. 6. Lee ML, Dempsey EW. Microcirculation of the rat placenta. AM J OBSTET GYNECOL 1976;126:495. 7. WigglesworthJS. The Langhans layer in late pregnancy: a histological study of normal and abnormal cases. J Obstet Gynaecol Br Commonw 1962 ;69:355. 8. Wigglesworth JS. Vascular anatomy of the human placenta and its significance for placent al pathology.J Obstet Gynaecol Br Commonw 1969;76:979. 9. Fox H. Fibrinoid necrosis of placental villi. J Obstet Gynaecol Br Commonw 1968;75:448. 10. Altshuler G, Russell P, Ermscilla R. The placental pathology of SGA infants. AM J OBSTET GYNECOL 1975; 121:35 1.
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11. Van Der Veen F, Fox H. The human placenta in idiopathic intrauterine growth retardation: a light and electron microscopic study. Placenta 1983;4:65. 12. Burger P, Chandler D, Klintworth GK. Corneal neovascularization as studied by scanning electron microscopy of vascular casts. Lab Invest 1983;48: 160.
Scanning electron microscopy of SGA placental casts
13. McCormick]N, Faulk WP, Fox H, Fundenberg HH. Immunohistological and elution studies of the human placenta.} Exp Med 1971;91:1. 14. Fox H. Histological abnormalities of the placenta. In: Fox H, ed. Pathology of the placenta. London: WB Saunders, 1978: 149.
Fetal microcirculation of abnormal human placenta II. Scanning electron microscopy of placental vascular casts from fetus with severe erythroblastosis fetalis Mary M. L. Lee, Ph.D., and Ming-neng Yeh, M.D. New York, New York The latex injection-corrosion cast technique coupled with scanning electron microscopy was applied to the study of severe Rh sensitization of the placenta. The main observed ultrastructural changes in fetal vasculature in this type of placenta were very similar to those of the small for gestational age placenta; large-sized vessels exhibited less branching than those of the normal placenta, and the capillary network showed numerous vascular buds and "H"-shaped anastomoses. In addition, in Rh-sensitized placentas, the capillary network was remarkably sparse, and the cast also revealed smooth-surfaced veins with distinct U shapes undulating through the placental stroma. These U-shaped veins may be the result of umbilical flow impedance with subsequent umbilical venous hypertension, which stretches the veins into undulating form. (AM J OSSTET GYNECOL 1986;154:1139-46.)
Key words: Fetal microcirculation, placenta, scanning electron microscopy, Rh sensitization, neovascularization The latex injection-corrosion cast technique coupled with scanning electron microscopy enables investigators to examine vascular casts from normal" 2 and abnormal human placentas in a three-dimensional form and to observe the ultrastructural changes in the fetal-placental vasculature. Several investigators':' have described, using light and transmission electron microscopy, the appearance of Rh-incompatible placentas. In part one data were presented on ultrastructural changes observed in vascular casts from the placentas of small for gestational age fetuses." This article will describe the vascular alterations that occurred in the placentas of fetuses with severe erythroblastosis fetalis and will discuss possible effects on the fetal-placental microcirculation resulting from such changes. To date, there has not been a scanning electron microscopic analysis of corrosion casts from Rh-incompatible placentas. From the Department of Physiology and the Department of Obstetrics and Gynecology, Columbia University College of Physicians and Surgeons. Received for publication September 27, 1985; revised December 17, 1985; acceptedJanuary 28, 1986. Reprint requests: Dr. Mary M. L. Lee, Department of Physiology, Columbia University College ofPhysicians and Surgeons, 630 West 168th St., New York, NY 10032.
Material and methods Four placentas from Rh-sensitized mothers who had given birth to babies with severe anemia were used for the study (Table I). The same latex injection-corrosion cast technique used in the normal human placenta was applied to this investigation." The umbilical cord was threaded by a small catheter into the umbilical artery and vein, providing a route of entrance of injection medium and a route of egress. After catheterization, the placenta was suspended in water, flushed with warm saline solution at an injection pressure of 50 mm Hg, and followed by an injection of 70% Cementex (Cementex Corp., New York, NY 10013). After polymerization by air drying, the latex inside the blood vessel hardened into a vascular cast. For further hardening the tissue was left in 10% formalin for 2 weeks. Then the placenta was macerated by treatment with sodium hypochlorite. The resulting vascular cast was air dried and teased into small pieces for examination in the scanning electron microscope as previously described.v" For scanning electron microscopic examination, the casts were attached onto aluminum stubs by silver conductive paint and then coated with gold-palladium in 1139