PARENT SESSION
Apoptosis

FACTORS INVOLVED IN TUMOR NECROSIS FACTOR--INDUCED RAT CORPUS LUTEUM APOPTOSIS.

Abdo, Michael¹, Dharmarajan, Arun¹, ¹

ABSTRACT- Tumor necrosis factor-alpha (TNF) is a cytokine with diverse biological outcomes that are governed by the presence of its receptors (p55 and p75) and second messengers. To date a number of factors have been identified that contribute to TNF-induced responses and in particular apoptosis. Apoptosis, a discrete form of cell death is associated with regular tissue turnover, development and disease. The focus of our research is the role of signaling molecules in corpus luteum (CL) apoptosis and ovarian function. The aim of this study was to identify several specific factors associated with TNF-induced CL apoptosis in the rat. Here we report the selective involvement of the caspases in TNF-induced apoptosis, differential expression of the TNF receptors in the rat CL, and the role of both nitric oxide (NO) and nuclear factor-kappaB (NF-B) in CL apoptosis. The role of the caspases (1 to 10) was assessed through incubation of luteal cells with specific caspase inhibitors (50M) following TNF treatment (75ng/mL). TNF receptor and NF-B expression was assessed through immunocytochemistry and Western blot analysis. NO was investigated through incubation of whole CL with L-arginine (NO substrate; 1-10mg/mL) and L-NAME (inhibitor of NO synthase; 1-10mg/mL) in the presence and absence of TNF (75ng/mL). Inhibition of caspases 3, 6, and 8 resulted in a significant reduction (p<0.05) of TNF-induced apoptosis, assessed through DNA fragmentation analysis. TNF receptors (p55 and p75) were localized within the rat CL compartment and their expression correlated with a decline in CL function and increased apoptosis. NF-B expression was inversely correlated with CL apoptosis. Treatment of CL with L-NAME had no effect on TNF-induced apoptosis, which was taken to suggest that the NO pathway is independent of TNF in effecting CL apoptosis. Together these results further clarify the involvement of TNF and signaling molecules in rat CL apoptosis.

KEY WORDS: Apoptosis, Tumor Necrosis Factor - alpha, Corpus Luteum, Signal Transduction