Continuing growth and development of the third-trimester human placenta

Two hundred and nineteen human placentae of well ascertained gestational age were measured for weight and nuclear number. Contrary to previous reports, analysis of the results showed no faltering in either parameter, however expressed. The placentae from babies exhibiting intrauterine growth retardation were appropriate to the size of the babies. However else the placenta ages it does not do so in respect of the rate of increase in the number of its nuclei.     

Angiotensin I-converting enzyme in human placenta

Angiotensin I-converting enzyme (ACE, peptidyldipeptide hydrolase, kininase II, EC 3.4.15.I) from human placenta was purified 6297-fold and characterized. ACE could be extensively purified by affinity chromatography with Captopril (D-3-mercapto-2-methylpropanoyl-L-proline), an orally active antihypertensive agent and a potent inhibitor of this enzyme. Its molecular weight and subunit size were estimated to be 300 000 by high-performance gel permeation chromatography and 85 000 by sodium dodecyl sulphate gel electrophoresis, respectively, indicating its polymeric structure.     

Receptors for oestradiol in human placenta

The binding of oestradiol in human placental tissue obtained from spontaneous and caesarean deliveries was investigated. Villous tissue was homogenized and the 800 g 'cytosol' supernatant treated with ammonium sulphate. The precipitate was redissolved and the preparation then equilibrated with [3H]oestradiol. After separation of bound and free steroid with dextran coated charcoal, receptor binding site concentrations and dissociation constants were determined. The presence of high-affinity, saturable binding was clearly demonstrated. Mean values (+/- s.d.) for receptor binding site concentrations were 64.6 +/- 22.9 (n = 6) and 33.8 +/- 32.9 (n = 8) fmol/mg protein for spontaneous and caesarean deliveries, respectively; respective dissociation constants were 16.0 +/- 7.6 and 7.0 +/- 6.0 pM. Heat denaturation studies, equilibration time studies and certain of the Scatchard plots suggested that forms of the oestradiol receptor of differing stability may be present in human placental tissue.     

Localization and distribution of unconjugated steroid hormones in normal placenta at term

The localization and distribution of unconjugated oestrone, oestradiol-17 beta, oestriol and progesterone were studied in normal term placentae, using subcellular fractionation and indirect immunofluorescence techniques. Radioimmunoassay of these steroids in each subcellular fraction showed that they were detectable mainly in the cytosol fraction. The efficiency of fixatives for retaining steroids in placental tissue during immunohistochemical procedures was studied in order to validate the present experimental techniques. As compared with other fixatives, glutaraldehyde solution produced minimal leakage of steroid hormones from placental tissue. Using an indirect immunofluorescence technique oestrone, oestriol and progesterone were detectable in the cytoplasm of villous syncytiotrophoblast of fixed term placentae.     

Transferrin receptor affinity and iron transport in the human placenta

Transferrin receptor activity has been investigated on isolated human placental syncytiotrophoblast microvillous plasma membrane preparations following 3 M KCl washing of membranes to remove endogenous receptor-bound maternal transferrin. The calculated binding parameters for essentially apo- and diferric human transferrin, and also for rabbit transferrin, were closely comparable (ranges: Ka, 3.5 to 5.0 X 10(7) M-1; n, 2.6 to 4.0 X 10(14)/mg membrane protein). Iron transport in pregnancy is thus unlikely to involve a simple process of transferrin displacement following iron release at the placental trophoblast plasma membrane receptor site.     

Myofibroblast as a major cellular constituent of villous stroma in human placenta

The human placenta requires contractile structures to generate energy for blood propulsion. Smooth muscle cells are not present in significant numbers in the human placenta while fibroblasts lack effective contractile properties. This study provides the following evidence that the stromal cells of the placental villi, cotyledonary septa and perivascular connective tissue are myofibroblasts. (I) Stromal cells are strongly positive for dipeptidylpeptidase IV (EC 3.4.14.5) which occurs exclusively in myofibroblasts as far as connective tissue cells are concerned. (2) The isoenzyme pattern of the placental dipeptidylpeptidase IV is identical to that of myofibroblasts in palmar fibromatosis on isoelectric focusing. (3) Antibodies raised against isolated placental dipeptidylpeptidase IV cross-react with dipeptidylpeptidase IV from myofibroblasts of palmar fibromatosis as shown by immunohistochemistry. (4) On electron microscopic examination, stromal cells present all the ultrastructural features of myofibroblasts. It is concluded that, except for the vascular component and a negligible number of Hofbauer cells, myofibroblasts make up nearly all the cellular constituents of human placental villous stroma.     

Insulin-induced receptor regulation in early gestation and term human placental cell cultures

Human placentae of different gestational ages have been used to investigate the binding and degradation of insulin as well as the regulation of insulin membrane receptors. Bacitracin was found to be an effective inhibitor of insulin degradation in human early gestation and term placental cell cultures. In the presence of bacitracin, [125I]-insulin bound rapidly and reversibly: maximal binding occurred at 4 degrees C, with a sharp pH optimum at 7.5, and exhibited a high degree of specificity. The extent of binding was proportional to cell protein and [125I]-insulin concentrations. Term (38 to 41 weeks; n = 25) placental cell cultures possessed receptors for insulin that were increased three-fold compared to early gestation (8 to 18 weeks; n = 17). This was due to an increase in receptor number with no significant alteration in affinity. A decrease in insulin binding in both early gestation and term placental cells was related to both the insulin and bacitracin concentrations present during 12 to 20 h of preincubation at 37 degrees C. The receptor loss was due to a decrease in the number of receptors per mg cell protein with no apparent change in their affinity. We conclude that our in vitro system, which utilizes human placental cells in monolayer culture, will permit a more direct study of the metabolic effects of insulin in both early gestation and term placentae.     

Placental hormones: I. Immunofluorescence studies of the localization of chorionic gonadotrophin, placental lactogen and prolactin in human and rat placenta and in the endometrium of pregnant rats

Chorionic gonadotrophin (CG), placental lactogen (PL) and prolactin (PRL) were localized in sections of human and rat placenta using the indirect immunofluorescence technique. Antisera were absorbed with homologous and unrelated antigens by affinity chromatography or complexed with their respective antigens. To exclude binding of IgG and immune complexes to Fc-receptor in chorionic tissue, thus leading to erroneous results, sections were preincubated with an excess of rat IgG. The results indicate that, in the human placenta, human CG (hCG) and human PL (hPL) can be identified in the syncytiotrophoblast. Using antibodies to hPRL fluorescence was found in giant cells within the cytotrophoblastic islands in the intervillous space and in cytotrophoblastic cells within placental septa. In the rat placenta rat PL (rPL) and rat PRL (rPRL) could be identified in the giant cells of the basal zone. Furthermore rPRL/rPL was localized in the endometrium and adjacent decidial cells of rat pregnancy. No staining of chorionic cells was detected using antibodies against hCG and bLH, which cross-reacted with rLH. These experiments provide further evidence that the rat placenta is unlikely to produce a CG/LH-like hormone.     

Quantification of aryl hydrocarbon hydroxylase and 7-ethoxycoumarin O-deethylase activity in human placentae: development of a protocol suitable for studying effects of environmental exposures on human metabolism

Several issues regarding measurement of placental AHH and 7ECD activity were studied, and standardized procedures that appeared more suitable than previous assay procedures for measurement of MFO induction in epidemiological studies were adopted. In the AHH assay, deletion of the rat-liver supernatant eliminated a possible extraneous contribution to measurement of low levels of AHH activity and did not substantially affect measurement of higher levels of activity. Increasing the protein concentration of placentae homogenate from 2 to 6 mg, and the length of the incubation time from 20 to 60 min, allowed for accumulation of more BaP products, potentially maximizing the detection of low levels of AHH activity. Use of tissue homogenates made the procedure more convenient and did not appear to interfere with interindividual comparisons of activity. Assay of homogenates of fresh and frozen tissue from the same placenta gave similar results, so that frozen tissue was adopted for convenience and replicability. Although a potential problem for specimens with high AHH activity, degradation of product(s) was modest in AHH assays of human placenta. The efficiency of extraction of fluorescent products declined with increasing protein concentrations in the reaction mixture of AHH assays, but it was stable for a range of product concentrations, and could be controlled by the use of a constant amount of protein per assay. Recovery of product for the 7ECD assay was more complete and was not affected by protein concentration. Additionally, the 7ECD activity was easily detected in every placenta, regardless of smoking exposure. However, in this study the AHH assay appeared to be better at discriminating between non-smokers and smokers. These observations, and the potential differences in the spectrum of agents causing induction of mixed-function oxidases, suggest that both assays are potentially useful measures of human MFO induction in clinical or epidemiological studies.     

Placental transport of hexoses: a comparative study with antipyrine and amino acids

A comparative study of the transplacental passage of some labelled hexoses, amino acids and antipyrine leads to the following conclusions: The maternal-fetal transfer of D-glucose, which amounted to 90 per cent of antipyrine, is very efficient. D-glucose, 3-O-methylglucose and 2-deoxyglucose, for which equivalent transfers were obtained, appear to share the same carrier system. L-glucose transport occurs by simple diffusion. 3-O-methylglucose did not accumulate in the placental tissue. Similarly, maternal and fetal concentrations of 2-deoxyglucose were nearly at equilibrium. It is concluded that the transport of D-glucose is not concentrative. These observations provide further evidence of a facilitated diffusion process for the transport of D-glucose across the human placental membrane.     

Hydatidiform mole in Nottingham: a 12-year retrospective epidemiological and morphological study

The results of a 12-year retrospective study on the epidemiology of hydatidiform mole (HM) in Nottingham are presented. We have reviewed the histology of our cases of HM, tried to minimize selection bias, and have used the most reliable data available. Using data from two different sources we have calculated the frequency of HM as 1 in 1400 deliveries. The frequency of HM in this study is one quarter of that reported in an excellent study from Japan. We suggest that, with accurate epidemiological studies, the difference in frequency of HM between 'high risk' and 'low risk' areas is less than previously accepted. The present study also shows a lower incidence of persistent trophoblastic disease than previously generally accepted. We confirm that partial HM is a distinct clinicopathological entity and that two forms are distinguishable histologically. The malignant potential of partial HM is uncertain, and we suggest that the clinical management of partial HM should be no different from that of complete HM until further studies dictate otherwise.     

Amyloid P component in normal human placentae

The presence of tissue amyloid P component was determined by using a direct immunofluorescence technique on frozen sections of normal human placentae and umbilical cords from gestations of various durations. Amyloid P component was first detected at the 16th week of gestation and appeared to increase progressively in amount so as to be present in abundance in the term placenta. Placental amyloid P component was present in the perifetal capillary zone where basement membrane-like material and reticulin fibres are also found. Amyloid P component may be related to the maturation of the placenta.     

An immunofluorescence study of extracellular matrix associated with cytotrophoblast of the chorion laeve

Using immunofluorescence we have studied the distribution within and beneath the cytotrophoblast of chorion laeve of five extracellular matrix components: types I, III and IV collagen, fibronectin and laminin. Fibronectin, laminin and types I and IV collagen are located in a network of extracellular matrix which encapsulates cells of the cytotrophoblast multilayer. The results suggest that cytotrophoblast in all layers is active in matrix biosynthesis. The pseudo-basement membrane, which links this intercellular matrix to the underlying reticular layer, contains rich deposits of fibronectin and type III collagen along with more restricted foci of laminin and type IV collagen. The reticular layer contains fibronectin, type III collagen and small amounts of type I collagen, but no laminin or type IV collagen. Type III collagen is also found in the maternal decidua. Extracellular matrix markers may be useful in defining trophoblast differentiation pathways. Possible reasons are discussed for the unusual architecture of extracellular matrix of cytotrophoblast of the chorion laeve. Cytoplasmic actin occurs in cells throughout the chorion laeve, while cytokeratin appears only in cytotrophoblast. Plasminogen also occurs throughout cytotrophoblast in pericellular locations.     

Modification of protein kinase pattern in human placentae during gestation

The cAMP-dependent and cAMP-independent histone kinases have been studied in the two subcellular compartments (cytosol and particulate fraction) from placentae of different gestational age. The total protein kinase activity, as well as its distribution between the two compartments, changes during the period of gestation. The total activity is significantly increased in full-term placentae. The increase is much greater for the cAMP-dependent (400 per cent), than for the cAMP-independent (270 per cent) protein kinases. It is much higher (400 per cent) in the cytosol than in the particulate fraction (170 per cent); consequently, the particulate fraction of term placentae shows a relatively lower proportion of protein kinase activity (26 per cent of the total activity) than the corresponding fraction of young placentae (37 per cent). DEAE-cellulose chromatography revealed the presence of two cAMP-dependent protein kinase peaks which correspond to Type I and Type II isoenzymes described in many mammalian tissues (Corbin, Keely and Park, 1975). The Type II isoenzyme is predominant in both first- and third-trimester placentae. The increase in protein kinase activity in term placentae is due to the selective activation of the Type II kinase only. The activity of the Type I isoenzyme remained unchanged throughout the period of gestation. The third peak eluted from the DEAE-cellulose column corresponds to a cAMP-independent sucrose-gradient ultracentrifugation into two distinct peaks similar to those already observed in several rat tissues (Toru-Delbauffe, Ohayon and Pavlovic-Hournac, 1983). The protein kinase patterns of both young and term placentae remain stable during the incubation of the tissues 'in vitro' for three hours.