Tumor necrosis factor-alpha induces clustering in ovarian theca-interstitial cells in vitro

Tumor necrosis factor-alpha (TNF) has been implicated in the regulation of steroidogenesis in theca-interstitial cells (TIC). The purpose of this study was to evaluate any change in TIC morphology during the time course of TNF-induced inhibition of LH-stimulated androstenedione production. Ovaries from immature hypophysectomized rats were enzymatically digested and highly purified TIC were obtained by density gradient centrifugation. TIC treated with TNF (0.1-10 ng/ml) demonstrated distinct clustering in the presence and absence of LH (50 ng/ml). The number of clusters and the mean area per cluster were greatest after 4 days as a result of treatment with 1 or 10 ng TNF/ml. In addition, a dose-dependent inhibition of LH-supported androstenedione production was induced by TNF. TNF also inhibited LH-induced androstenedione in TIC after 2, 4, or 6 days of continuous LH treatment, and TIC clustering still occurred. TIC clustering was impeded by the protein kinase inhibitor H7 at 10 microM; however, the protein kinase inhibitor, HA 1004 (5 microM), did not inhibit TNF-induced clustering in TIC. Since H7 blocked TNF induced clustering, but did not block TNF inhibition of LH stimulated androstenedione synthesis, it is suggested that alternate signal transduction pathways for TNF induced inhibition of LH-stimulated androstenedione and stimulation of clustering of TIC may exist. The results also indicate that the TNF-induced TIC clustering may be independent of the TNF-induced inhibition of LH-stimulated androstenedione production and states of LH-induced differentiation of TIC.