Induction of Anti-Apoptotic Genes in Cultured Small, Large, and Mixed Ovine Luteal Cells in Response to Interferon-Tau.

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Ovine interferon-t (oIFNT) is released from the conceptus into the uterine vein by Day 15 of pregnancy and has endocrine action through inducing IFN-stimulated genes (ISGs) in the corpus luteum (CL). ISG15 is localized to large and small luteal cells based on immunohistochemical staining. Large luteal cells contain prostaglandin F2a (PGF) receptors that stimulate apoptotic signalling and release of oxytocin (OXT) when bound to ligand. Small luteal cells contain OXT receptors, and when activated also induce apoptotic signalling. Infusion of 200 μg of recombinant (r) oIFNT/day into the uterine vein of ewes induces cell survival genes in the CL such as B cell lymphoma extra large (Bcl-XL), protein kinase B (Akt), and X-linked inhibitor of apoptosis (XIAP). Endocrine delivery of 20 μg roIFNT/day into the uterine vein or subcutaneously to the ewes on Day 10 of the estrous cycle also protects the CL against the decline in serum progesterone concentrations caused by injection with PGF on Day 11 of the estrous cycle. Others have described the presence of PGE2 in uterine vein blood and its luteotrophic action on the CL. The present in vitro studies tested the hypothesis that IFNT, rather than PGE2, induces ISG15 and cell survival genes in cultured luteal cells. CLs were collected from adult ewes on Day 10 of the estrous cycle and prepared as mixed (1 x 106/ml) or separated by elutriation into small (2 x 106/ml ) or large (5 x 105/ml) luteal cells. Luteal cells were cultured for 24 h in M199 media supplemented with 10% FBS. The media was replaced with serum-free media and treatments were added to cells. Differences are described as P < 0.05. Culture with 0.1, 1, 10 or 100 ng/ml of roIFNT (108 U/mg) revealed that 1 ng/ml induced ISG15 mRNA concentrations (determined by semi-quantitative RTPCR) in small, large and mixed cells that were in the upper (~80-90% of maximum) linear response to these doses of roIFNT. Cells also were cultured with 10 μM OXT, 3.5 ng/ml PGF and 3.5 ng/ml PGE2, based on effective doses published by others. The roIFNT induced greater BCL-XL, Akt and XIAP mRNA concentrations in small, mixed and large cell cultures when compared to controls. PGE2, PGF and OXT had no effect on ISG15 mRNA concentrations. PGF caused a slight induction of Bcl-XL in large luteal cells and a slight reduction of XIAP mRNA concentrations in mixed luteal cells, but otherwise had no effect. OXT had no effect on Bcl-XL or Akt, but slightly upregulated XIAP mRNA concentrations in small luteal cells. PGE2 had no effect on XIAP, but slightly decreased Bcl-XL and Akt mRNA concentrations in small and large cells. It is concluded that ISG15, Bcl-XL, Akt, and XIAP mRNAs are up regulated within 24h in response to roIFNT treatment of isolated small, large and mixed luteal cells, and that this induction is independent of the actions of PGE2, OXT and PGF. These results are similar to the results obtained in previous in vivo studies using osmotic infusion of roIFNT into the uterine vein of ewes. Endocrine action of IFNT might entail up regulation of ISGs as well as cell survival genes to assist with providing resistance of the CL to PGF, but also in longer-term resilience in context of continued production of progesterone. USDA-NIFA-AFRI 2011–67015–20067. Dr. Sereno is on sabbatical leave from Embrapa Cerrados, Planaltina, DF, Brazil.