Type VII collagen associated with the basement membrane of amniotic epithelium forms giant anchoring rivets which penetrate a massive lamina reticularis.

In human amnion a simple cuboidal epithelium and underlying fibroblast layer are separated by an almost acellular compact layer rich in collagen types I and III. This (>10 µm) layer, which may be a thick lamina reticularis, apparently presents an unusual set of conditions. Integration of the multilaminous tissue across it is apparently achieved by waisted structures which we have observed with the light microscope in frozen, paraffin-wax and semi-thin resin sections. We have also captured transmission and scanning electron micrographs of the structures. These structures which cross the compact layer we call "rivets". The composition of these "rivets" has been examined immunocytochemically and in three dimensions using the confocal laser scanning epi-fluorescence microscope. The rivets contain type VII collagen and an α6 integrin. They associate with type IV collagen containing structures (basement membrane lamina densa and spongy coils) and a special population of fibroblasts which may generate, maintain or anchor rivets to the underlying mesenchymal layer. Although type VII collagen is well known to anchor basal lamina to underlying mesodermal collagen fibres these "rivets" are an order of magnitude larger than any previously described type VII collagen containing anchoring structures. Intriguing possible functions of these features include nodes for growth of fibrous collagen sheets and sites of possible enzymatic degradation during regulated amnion weakening approaching term. If these sites are confirmed to be involved in amnion degradation and growth they may represent important targets for therapeutic agents that are designed to delay preterm premature rupture of the membranes a major cause of fetal morbidity and mortality.

Selective estrogenic effects of a novel triphenylethylene compound, FC1271a, on bone, cholesterol level, and reproductive tissues in intact and ovariectomized rats.

FC1271a is a novel triphenylethylene compound with a tissue-selective profile of estrogen agonistic and weak antagonistic effects. It specifically binds to the estrogen receptor alpha and beta with affinity closely similar to that of toremifene and tamoxifen. To study the in vivo effects of the compound, 4-month-old rats were sham operated (sham) or ovariectomized (OVX) and treated daily for 4 weeks with various doses of FC1271a or vehicle (orally). FC1271a was able to oppose OVX-induced bone loss by maintaining the trabecular bone volume of the distal femur. Accordingly, the OVX-induced loss of bone strength was prevented at doses of 1 and 10 mg/kg. FC1271a also prevented the OVX-induced increase in serum cholesterol in a dose-dependent manner. No significant changes in uterine wet weight or morphology were observed in the OVX-rats treated with 0.1 or 1 mg/kg FC1271a, but at a dose of 10 mg/kg it had a slightly estrogenic effect. In immature rats the effect of FC1271a on uterine wet weight was less stimulatory than that of toremifene or tamoxifen, but more stimulatory than that of raloxifene or droloxifene. The appearance of the dimethylbenzanthracene (DMBA)-induced mammary tumors was inhibited by treatment of DMBA-treated rats with FC1271a in a dose-dependent manner. In human MCF-7 breast cancer cell tumors raised in nude mice in the presence of estrogen, the growth and expression of pS2 marker gene could not be maintained after estrogen withdrawal by treatment with FC1271a. No formation of DNA adducts was observed in the liver of the FC1271a-treated rats. In conclusion, the bone-sparing, antitumor, and cholesterol-lowering effects of FC1271a combined with a low uterotropic activity and lack of liver toxicity indicate that FC1271a could be an important alternative in planning antiosteoporosis therapy for estrogen deficiency.

Albumin endocytosis is regulated by heterotrimeric GTP-binding protein G alpha i-3 in opossum kidney cells.

Proteinuria is an adverse feature in patients with renal disease, possibly due to toxicity of albumin to proximal tubular cells. Albumin is reabsorbed from tubular fluid by receptor-mediated endocytosis. The mechanism of regulation of the endocytosis is unknown. The large quantities of G proteins in proximal tubular cell apical membranes suggests that they may have a regulatory role in endocytosis. 125I-labeled albumin uptake was measured in opossum kidney (OK) cells. This is a saturable process with high-affinity [apparent dissociation constant (Kd) = 24.3 mg/l] and low-affinity (Kd = 15.9 g/l) components. The endocytic uptake of gold-albumin into OK cells was confirmed by electron microscopy. 125I-albumin endocytosis in OK cells was inhibited by pertussis toxin, but cholera toxin had no effect. Pertussis toxin also inhibited uptake of [3H]inulin. OK cells were stably transfected with a cDNA for the G protein subunit G alpha i-3 and transfectants were screened by immunoblotting. Several G alpha i-3-overexpressing clones were detected. OK cells overexpressing G alpha i-3 demonstrate increased 125I-albumin uptake, which is abolished by pertussis toxin, in both a concentration- and time-dependent manner. These results suggest that albumin endocytosis in OK cells is regulated by the G protein G alpha i-3.

Placental transfer of the hypolipidemic drug, clofibrate, induces CYP4A expression in 18.5-day fetal rats.

Rats at day 15.5 of gestation were dosed intraperitoneally with 300 mg.kg-1 of clofibrate for three consecutive days at 24-hr intervals and were culled 24 hr after the final injection. This regime produced maximal induction of the cytochrome P4504A (CYP4A) mRNAs in the maternal liver and kidney and in 18.5-day fetal tissues. The maternal hepatic and renal CYP4A mRNA levels had risen 12- and 2-fold, respectively, above the constitutive levels seen in untreated pregnant rats at an equivalent stage of gestation. Clofibrate was capable of traversing the placenta and modulating the fetal CYP4A mRNA expression as demonstrated by a 3-fold elevation in the mRNA levels in those fetuses explanted from drug-induced mothers, compared with those fetuses removed from untreated mothers. The CYP4A mRNAs were demonstrated in the fetal liver via dot-blot and Northern blot analyses. In addition, low levels of CYP4A mRNA expression were detected in the induced placenta via Northern blot analysis. Western blot analysis revealed that the CYP4A protein levels increased in the maternal liver and in the kidney and fetal livers after exposure to clofibrate. Peroxisome proliferation, a phenomenon associated with induction of CYP4A1 expression in rodents, was demonstrated in both maternal and fetal livers, with the use of light and electron microscopy.