Support for an infection-induced apoptotic pathway in human fetal membranes.
OBJECTIVE
Lipopolysaccharide and tumor necrosis factor alpha levels are both elevated in the amniotic fluid of women during infection-associated preterm labor and premature rupture of fetal membranes. Our laboratory has shown that apoptosis is associated with premature rupture of fetal membranes but is not associated with preterm labor. The exact pathway that leads to apoptosis-mediated premature rupture of fetal membranes is still unclear. Because infection and increased inflammatory cytokine response are associated with the majority of cases of premature rupture of fetal membranes, we examined the roles of bacterial lipopolysaccharide and tumor necrosis factor alpha in inducing the proapoptotic caspase pathway in fetal membranes.


METHODS
Amniochorionic membranes collected from women undergoing elective repeat cesarean delivery at term were placed in an organ explant system. At the end of a 48-hour incubation period, membranes were stimulated with lipopolysaccharide (50 ng/mL) and recombinant tumor necrosis factor (50 ng/mL). Total ribonucleic acid extracted from these samples was subjected to reverse transcription and two separate sets of multiple polymerase chain reaction. One set studied the expression of Fas, Fas ligand, caspase 8, Fas-associated death domain, and tumor necrosis factor receptor-associated death domain 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 reverse transcriptase-polymerase chain reaction.


RESULTS
Multiple polymerase chain reactions and reverse transcriptase-polymerase chain reactions documented the induction of Fas and caspase 2, 3, 7, 8, and 9 genes in amniochorion after lipopolysaccharide and tumor necrosis factor stimulation compared with the nonstimulated controls. Neither lipopolysaccharide nor tumor necrosis factor induced Fas ligand expression in human fetal membranes. Caspase 3, 4, and 6, Fas-associated death domain, and tumor necrosis factor receptor-associated death domain expressions were constitutive in all the tissues tested; however, tumor necrosis factor receptor-associated death domain expression appeared stronger in tumor necrosis factor-stimulated tissues.


CONCLUSIONS
The presence of the signal docking proteins tumor necrosis factor receptor-associated death domain and Fas-associated death domain and the induction of caspase cascade initiators (caspase 2, 8, and 10) and effector caspases (caspase 3, 6, 7, and 9) by lipopolysaccharide and tumor necrosis factor suggest that tumor necrosis factor-tumor necrosis factor receptor-mediated apoptosis may occur in the human fetal membrane.
Fortunato
S J
SJ
Perinatal Research Center of The Women's Health Research and Education Foundation, the Maternal-Fetal Group, and Aquinas College, Nashville, Tennessee 37203, USA.
Menon
R
R
Lombardi
S J
SJ
D048868:Adaptor Proteins, Signal Transducing; D000650:Amnion / Q000187:drug effects / Q000382:microbiology* / Q000503:physiopathology*; D015703:Antigens, CD / Q000378:metabolism; D017209:Apoptosis; D002352:Carrier Proteins / Q000378:metabolism; D020169:Caspases / Q000235:genetics; D002823:Chorion / Q000187:drug effects / Q000382:microbiology* / Q000503:physiopathology*; D053222:Fas Ligand Protein; D053200:Fas-Associated Death Domain Protein; D005786:Gene Expression Regulation; D006801:Humans; D007239:Infections / Q000503:physiopathology*; D008070:Lipopolysaccharides / Q000494:pharmacology; D008562:Membrane Glycoproteins / Q000378:metabolism; D016133:Polymerase Chain Reaction; D018124:Receptors, Tumor Necrosis Factor / Q000378:metabolism; D047888:Receptors, Tumor Necrosis Factor, Type I; D020133:Reverse Transcriptase Polymerase Chain Reaction; D014409:Tumor Necrosis Factor-alpha / Q000494:pharmacology; D019014:fas Receptor / Q000235:genetics
D048868:Adaptor Proteins, Signal Transducing; D015703:Antigens, CD; D002352:Carrier Proteins; C505533:FADD protein, human; C505661:FASLG protein, human; D053222:Fas Ligand Protein; D053200:Fas-Associated Death Domain Protein; D008070:Lipopolysaccharides; D008562:Membrane Glycoproteins; D018124:Receptors, Tumor Necrosis Factor; D047888:Receptors, Tumor Necrosis Factor, Type I; D014409:Tumor Necrosis Factor-alpha; D019014:fas Receptor; D020169:Caspases