Isolation and characterization of Hofbauer cells from human placental villi

Summary Hofbauer cells are a major cell type of the human placental villous core and they are particularly numerous at the beginning of pregnancy. In the present study we describe a method suitable to obtain HC suspensions in a highly purified form. These suspensions have been analyzed for surface markers using a battery of monoclonal antibodies. Of all the surface markers used, Hofbauer cells were only positive for 4F2, LeuM2 and LeuM3 monoclonals which mainly detect cells of the monocyte-macrophage lineage. Hofbauer cells were consistently negative for HLA-DR antigens, C3bR and T- or B-cell markers. Hofbauer cells appeared capable of phagocytosing latex beads, adhering to and spreading over plastic surface and secreting lysozyme. In contrast, they failed to originate an efficient respiratory burst in response to appropriate stimulation. Hofbauer cells were positive for ANAE with a perinuclear localization of the enzyme activity, but consistently negative for peroxidase. These observations suggest that they share a number of features with cells of the monocyte-macrophage lineage and yet have some distinctive properties.     

Identification of arginase in human placental villi

l-Arginine is the common substrate for arginase and nitric oxide synthase (NOS). Arginase converts l-arginine to urea and ornithine, which is the principal precursor for production of polyamines required for cell proliferation. Human placenta expresses endothelial NOS (eNOS) in syncytiotrophoblasts, but the expression of arginase has not been fully elucidated. Our aim was to investigate the expression and distribution patterns of arginase-I (A-I) and arginase-II (A-II) in human placental villi in the first trimester and at term using immunohistochemistry, RT-PCR and Western blot analysis. The arginase enzyme activity in placental villi was also measured. Immunohistochemistry showed different distribution patterns of the arginase isoforms during gestation: A-I was observed only in cytotrophoblasts, while A-II was observed in both cytotrophoblasts and syncytiotrophoblasts. RT-PCR and Western blot analysis showed expression of A-I and A-II in the first trimester and at term in human placental villi. Expression of A-II and arginase activity was greater in the first trimester than at term. Differentiation of cytotrophoblasts into syncytiotrophoblasts may be associated with l-arginine metabolism through modulation of l-arginine availability for eNOS and A-I. And elevated arginase activity in the early gestational period may be responsible for proliferation of trophoblasts by increasing polyamines production. These results suggest that the l-arginine-ornithine-polyamine and l-arginine-nitric oxide pathways play a role in placental growth and development.     

Immunocytochemical identification of mitotic Hofbauer cells in cultures of first trimester human placental villi

Summary In cultures of first trimester human placental villi, mitotic Hofbauer cells have been identified using a combined autoradiographic and immunostaining technique for the demonstration of HCG, a marker for Hofbauer cells.     

Steroid production by early pregnancy human placental villi in culture

Organ cultures prepared from human placentae obtained at 7-12 weeks of gestation were maintained for 3-13 days in Dulbecco's modified Eagle medium (DMEM). The addition of pregnenolone to the medium resulted in a dose-related increase in progesterone production and the addition of androstenedione resulted in a dose related increase in oestrogen production. More oestrone than oestradiol was measured in medium collected at the end of the first day of culture, but, on all subsequent days, oestradiol was the predominant oestrogen produced when androstenedione was added to the culture medium. When villi were incubated with [3H]androstenedione immediately after dissection most of the radiolabelled oestrogen recovered from the tissue and medium was oestrone; however, more [3H]oestradiol was recovered when villi were tested after 3 days of culture in DMEM. The addition of oestrone to the culture medium resulted in a dose related increase in oestradiol production with oestradiol accounting for a larger proportion of the total oestrogen in the day 2 and 3 medium samples than in the day 1 samples. These data demonstrate that the enzymes required for biosynthesis of progesterone and oestrogen from exogenous substrate are maintained for at least 13 days when early pregnancy placental villi are cultured in serum-free DMEM. However, a temporal change in the pattern of oestrogen synthesis does occur in culture, such that oestradiol rather than oestrone becomes the major product of androstenedione metabolism.     

Cross-sectional features and three-dimensional structure of human placental villi

Human placental villi from both normal and complicated pregnancies were examined by both light and scanning electron microscopy. The findings provide evidence that histological features such as syncytial sprouts, bridges, and a net-like arrangement of villi represent tangential sections of irregularly shaped villi rather than proliferative activity of the villous surface. Hence the two-dimensional appearance of paraffin and semithin sections has to be interpreted three-dimensionally in comparison with the respective scanning electron micrographs. In the light of these findings the various types of villous maldevelopment are summarized in a diagram which may be used as an aid for pathological diagnosis.     

Lectin binding of membrane proteins from human placental villi

The binding of membrane proteins from human placental villi to lectins such as concanavalin A (Con A), wheat germ agglutinin (WGA) and Riccinus communis agglutinin (RCA) was examined. Five major components of 66K, 69K, 100K, 130K and 170K dalton showed specific binding to WGA, whereas two other major components of 43K and 40K dalton had only weak binding. There were no membrane proteins which bound specifically to either Con A or RCA. The WGA-Sepharose column is considered useful for the purification of human placental membrane proteins. Although the function of these proteins is still unknown, a diagnostic method not only for determining the pathological state of the placenta, but also for cancer will possibly be developed by taking advantage of them.     

Expression of Fas-ligand in first trimester and term human placental villi

The expression of Fas-ligand (FasL) on trophoblast cells is thought to play a role in immune regulation during human pregnancy. However, there are some discrepancies in the published data concerning the cell types expressing FasL in the placental villi. Therefore, we examined the expression of FasL on cryosections of first trimester and term placental tissue with three different anti-sera against FasL, which are in common use. By immunohistochemistry, all three anti-sera principally gave the same staining result. In the first trimester of pregnancy, villous cytotrophoblast cells underlying the syncytium, as well as all extravillous trophoblast cells of cell columns and cell islands, gave a clear, mainly membrane-located staining, whereas the syncytiotrophoblast, which forms the borderline to the maternal blood flow, only gave a spot-like reaction in distinct areas. The same result was obtained with term placental villi; however, in this tissue, the staining of the villous cytotrophoblast cells was less pronounced. From our results, we suggest that in placental villi, an important role of FasL in immune regulation is not very conclusive because this molecule is mainly expressed on trophoblast with no access to maternal blood or tissue. This is in contrast to the uterine part of the placenta, where FasL expressing trophoblast cells are in close contact with apoptotic maternal leukocytes.