The present study evaluated the secretions of interleukin (IL)-1beta and tumor necrosis factor (TNF) alpha by fetal membranes stimulated with group B streptococci (GBS) and lipopolysaccharide (LPS). The aim was to evaluate the initial response of full-thickness membranes to the microbial insult using an in vitro experimental model that allowed testing of the individual contributions of amnion and choriodecidua to stimulation. Full-thickness membranes were obtained after delivery by elective cesarean section from women at 37-40 wk of gestation without evidence of active labor. The membranes were mounted in Transwell devices, physically separating the upper and lower chambers. The LPS (500 ng/ml) or GBS (1 x 10(6) colony-forming units/ml) was added to either the amniotic or choriodecidual surface, and accumulation of IL-1beta and TNFalpha were measured in both compartments using a specific ELISA. Fetal membranes followed different patterns of secretion of proinflammatory cytokines that depended on the side to which the stimulus was added or the nature of the stimulus itself. The TNFalpha was secreted by amnion and choriodecidua in the presence of LPS or GBS, and stimulation with GBS induced a greater synthesis of IL-1beta than did stimulation with LPS. Choriodecidual tissue was more responsive than amniotic tissue, and this response tended to be higher even when the stimulation was only on the amniotic side. However, the amnion plays an active role in recognizing LPS or GBS, contributing a significant amount of TNFalpha. Thus, cooperative and bidirectional communications occur between amnion and choriodecidua in response to bacterial products, which include intermembranous cytokine traffic and signaling between tissues.

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MDCK cells (epithelioid line derived from the kidney of a normal dog) form monolayers which retain the properties of transporting epithelia. In these cells viruses bud asymmetrically: influenza from the apical, and vesicular stomatitis (VSV from the basolateral membrane (E. Rodríguez-Boulán and D.D. Sabatini, Proc. Natl. Acad. Sci. USA 75: 5071-5075, 1978; E. Rodríguez-Boulán and M. Pendergast, Cell 20: 45-54, 1980). In the present study, we analyzed whether these viruses affect specific ion-translocating mechanisms located in the plasma membrane. We studied the effect of infection on membrane and transepithelial conductance, passive and active unidirectional fluxes of Na+ and K+, intracellular potentials, cellular content of Na+ and K+, and formation of blisters which, in these preparations, are due to the vectorial transport of fluid. Two main observations are derived from these studies. First, infection with VSV caused an increase in transepithelial electrical conductance, due to the opening of tight junctions, 5 to 6 hr after the start of infection, coincident with the accumulation of envelope protein in the cell surface and with the rise in the curve of virus budding. Infection with influenza, on the other hand, increased the transepithelial conductance only late in the infection (12 to 14 hr) when virus production has already stopped. Second, viruses did affect membrane permeability. Yet, the changes observed may not be ascribed to a perturbation of the specific translocating mechanisms for Na+ and K+ which operate in the same region of the plasma membrane that the viruses use to penetrate and leave MDCK cells. The methods used in the present study are not suitable to decide whether the nonspecific changes in permeability elicited by the viruses occur over the whole cell membrane or are restricted to a given region.

We examined the molecular mechanisms of the cytotoxicity of Entamoeba histolytica, using the loss of transepithelial electrical resistance (TER) of monolayers of Madin-Darby canine-kidney (MDCK) cells on their incubation with axenic trophozoites of the HM1-IMSS strain. Such loss of TER occurs very early (in 2-5 min) and is caused by the opening of tight junctions and the detachment of cells. We used specific inhibitors for three of the four molecules currently accepted as being responsible for cytotoxicity: galactose-specific adhesin(s), phospholipase A, and cysteine proteinases. We also used inhibitors of calcium channels. Axenic trophozoites of E. histolytica strain HM1-IMSS were preincubated with the different inhibitors for 1 h prior to their coincubation with MDCK-cell monolayers. The only inhibitor that effectively blocked the loss of TER caused by the parasite was galactose. We suggest that in this experimental model, galactose-specific adhesin(s) are essential for amebic cytotoxicity.