TNF-α promotes caspase activation and apoptosis in human fetal membranes

Ramkumar Menon, Salvatore J. Lombardi, Stephen J. Fortunato

Research output: Contribution to journalArticle

53 Citations (Scopus)

Abstract

Purpose: Increased amniotic fluid tumor necrosis factor (TNF) is a marker of infection when associated with preterm labor and preterm premature rupture of the amniochorionic membranes (PROM). We have noted increased apoptosis in membranes derived from women with PROM. This study examines the role of TNF in promoting fetal membrane apoptosis. Methods: Amniochorion (n = 8), collected at the time of elective repeat cesarean section prior to labor from normal term gestation, were placed in an organ explant system. After 48 h in culture, the membranes were stimulated with recombinant TNF-α (20 ng/mL) for 24 h. Tissue frozen after stimulation was subjected to RT-PCR to study the expression of TNF-induced caspase genes. ELISA assayed the levels of proapoptotic p53 in tissues and cell death related nuclear matrix protein (NMP) in tissue culture supernatants. The activity of caspases in tissue homogenates was measured using substrates specific for caspase 2, 3, 6, 8, and 9. Results were analyzed by using the Wilcoxon nonparametric test for paired samples. A p < 0.05 was considered significant. Results: RT-PCR showed induction of caspases 2, 8, and 9 (caspase cascade initiators) in human fetal membranes after TNF stimulation. Caspases 3 and 6 (effector caspases) expression was constitutive in both TNF stimulated- and control membranes. Caspases, 2, 3, 8, and 9 activity was significantly higher in TNF-stimulated tissues compared with control, whereas, no significant change in caspase 6 activity was noticed. TNF-stimulated tissues released increased levels of NMP (24.03 U/mL) compared with control (13.5U/mL) (p = 0.03). TNF also increased p53 levels in the tissues (0.05 ng/mL) compared with control cultures (0.03 ng/mL; p = 0.02). Conclusions: TNF increases proapoptotic p53 levels and caspase activities in fetal membranes. Increased NMP reflects cell death.

Original languageEnglish (US)
Pages (from-to)201-204
Number of pages4
JournalJournal of Assisted Reproduction and Genetics
Volume19
Issue number4
DOIs
StatePublished - 2002
Externally publishedYes

Fingerprint

Extraembryonic Membranes
Caspases
Tumor Necrosis Factor-alpha
Apoptosis
Nuclear Matrix-Associated Proteins
Caspase 2
Caspase 6
Caspase 3
Membranes
Cell Death
Repeat Cesarean Section
Initiator Caspases
Effector Caspases
Polymerase Chain Reaction
Caspase 9
Caspase 8
Premature Obstetric Labor
Amniotic Fluid
Rupture
Enzyme-Linked Immunosorbent Assay

Keywords

  • Amniochorion
  • Caspases
  • p53
  • Preterm labor
  • PROM
  • TNF

ASJC Scopus subject areas

  • Obstetrics and Gynecology
  • Developmental Biology
  • Genetics
  • Reproductive Medicine

Cite this

TNF-α promotes caspase activation and apoptosis in human fetal membranes. / Menon, Ramkumar; Lombardi, Salvatore J.; Fortunato, Stephen J.

In: Journal of Assisted Reproduction and Genetics, Vol. 19, No. 4, 2002, p. 201-204.

Research output: Contribution to journalArticle

Menon, Ramkumar ; Lombardi, Salvatore J. ; Fortunato, Stephen J. / TNF-α promotes caspase activation and apoptosis in human fetal membranes. In: Journal of Assisted Reproduction and Genetics. 2002 ; Vol. 19, No. 4. pp. 201-204.
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AB - Purpose: Increased amniotic fluid tumor necrosis factor (TNF) is a marker of infection when associated with preterm labor and preterm premature rupture of the amniochorionic membranes (PROM). We have noted increased apoptosis in membranes derived from women with PROM. This study examines the role of TNF in promoting fetal membrane apoptosis. Methods: Amniochorion (n = 8), collected at the time of elective repeat cesarean section prior to labor from normal term gestation, were placed in an organ explant system. After 48 h in culture, the membranes were stimulated with recombinant TNF-α (20 ng/mL) for 24 h. Tissue frozen after stimulation was subjected to RT-PCR to study the expression of TNF-induced caspase genes. ELISA assayed the levels of proapoptotic p53 in tissues and cell death related nuclear matrix protein (NMP) in tissue culture supernatants. The activity of caspases in tissue homogenates was measured using substrates specific for caspase 2, 3, 6, 8, and 9. Results were analyzed by using the Wilcoxon nonparametric test for paired samples. A p < 0.05 was considered significant. Results: RT-PCR showed induction of caspases 2, 8, and 9 (caspase cascade initiators) in human fetal membranes after TNF stimulation. Caspases 3 and 6 (effector caspases) expression was constitutive in both TNF stimulated- and control membranes. Caspases, 2, 3, 8, and 9 activity was significantly higher in TNF-stimulated tissues compared with control, whereas, no significant change in caspase 6 activity was noticed. TNF-stimulated tissues released increased levels of NMP (24.03 U/mL) compared with control (13.5U/mL) (p = 0.03). TNF also increased p53 levels in the tissues (0.05 ng/mL) compared with control cultures (0.03 ng/mL; p = 0.02). Conclusions: TNF increases proapoptotic p53 levels and caspase activities in fetal membranes. Increased NMP reflects cell death.

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