OBJECTIVE:
    LPS (endotoxin) and tumor necrosis factor - a (TNF- a) are both elevated in the amniotic fluid of women during infection associated preterm labor and premature rupture of the membranes (PROM). Our laboratory has shown that apoptosis is associated with PROM but not PTL. The exact pathway that leads to apoptosis mediated PROM is still unclear. Since infection and increased inflammatory cytokine response is associated with the majority of cases of PROM, we examined the role of bacterial LPS and TNF- a in inducing the pro-apoptotic caspase pathway in fetal membranes.

METHODS:
    Amniochorionic membrane collected from women undergoing elective repeat C-section at term were placed in an organ explant system. At the end of 48 hours incubation membranes were stimulated with LPS (50 ng/ml) and recombinant TNF (50 ng/ml). Total RNA extracted from these samples was subjected to reverse transcription and two separate sets of multiple PCR (MPCR). One set studied the expression of Fas, Fas ligand (FasL), caspase 8, Fas associated death domain (FADD) and TNF receptor associated death domain (TRADD) genes and the second set studied the expression of caspase 2,4, 6, 7 and 10. Caspase 2, 3 and 9 expression was also studied by RT-PCR.

RESULTS:
    Multiple PCR and RT-PCR documented the induction of Fas, caspase 2, 7, 8 and 9 genes in amniochorion after LPS and TNF stimulation compared to the non stimulated controls. Neither LPS nor TNF induced Fas L expression in human fetal membranes. Caspase 3, 4, 6, FADD and TRADD expressions were constitutive in all the tissues tested, however, TRADD expression appeared stronger in TNF stimulated tissues.

CONCLUSION:
    The presence of the signal docking proteins TRADD, FADD and induction of caspase cascade initiators (caspase 2, 8 and 10), effector caspases (3, 6, 7 and 9) by LPS and TNF suggests that TNF-TNF receptor mediated apoptosis may occur in the human fetal membranes