ABSTRACT
During luteolysis, luteal cell apoptosis is induced by the Fas ligand (FasL)/Fas system. In murine luteal bodies, we demonstrated the expression of mRNA of soluble form of Fas (FasB), which binds to FasL and prevents apoptosis induction. By in situ hybridization, strong expression of FasB mRNA was observed in normal luteal bodies, in which no apoptotic cells were detected, but negative/trace expression in regressing luteal bodies, in which many apoptotic cells were observed. Immunohistochemical staining revealed that Fas and TNF-alpha were localized in both normal and regressing luteal bodies, but IFN-gamma was localized only in regressing luteal bodies. Apoptosis was induced in primary cultured luteal cells, when they were pretreated with TNF-alpha and IFN-gamma and then incubated with TNF-alpha, IFN-gamma, and mouse recombinant FasL (rFasL). However, no apoptosis was detected in the cells, when they were treated with rFasL alone, TNF-alpha alone, IFN-gamma alone, TNF-alpha and rFasL, IFN-gamma and rFasL, or TNF-alpha and IFN-gamma. Fas mRNA expression in cultured luteal cells was up-regulated by the treatment of TNF-alpha, IFN-gamma, or TNF-alpha and IFN-gamma. The expression of FasB mRNA was down-regulated, when the cells were treated with TNF-alpha and IFN-gamma, but its expression was not changed by the treatment of TNF-alpha alone or IFN-gamma alone. We conclude that FasB inhibits the apoptosis induction in luteal cells of normal luteal bodies, and that decreased FasB production induced by TNF-alpha and IFN-gamma made possible the apoptosis induction in the luteal cells of regressing luteal bodies.
Subject(s)
Apoptosis , Luteal Cells/physiology , Luteolysis , Tumor Necrosis Factor-alpha/pharmacology , fas Receptor/physiology , Animals , Apoptosis/drug effects , Cell Survival , Cells, Cultured , Corpus Luteum/chemistry , Corpus Luteum/cytology , Fas Ligand Protein , Female , Gene Expression/drug effects , Interferon-gamma/analysis , Interferon-gamma/pharmacology , Luteal Cells/drug effects , Luteal Cells/ultrastructure , Membrane Glycoproteins/genetics , Membrane Glycoproteins/metabolism , Membrane Glycoproteins/pharmacology , Mice , Mice, Inbred ICR , Ovary/chemistry , RNA, Messenger/analysis , Recombinant Proteins/pharmacology , Reverse Transcriptase Polymerase Chain Reaction , Solubility , Tumor Necrosis Factor-alpha/analysis , fas Receptor/genetics , fas Receptor/metabolismABSTRACT
Immune cells, which constitute a significant cell mass in the corpora lutea (CLs), are considered to play critical roles in luteolysis, but the details are not fully understood. We histochemically investigated the changes in distribution and cell density of macrophages and T lymphocytes and in tumor necrosis factor (TNF)-alpha and interferon (IFN)-gamma, which can induce apoptosis in the luteal cells in murine CLs during luteal regression. No macrophages or T lymphocytes were observed in functionally healthy CLs. Abundant macrophages and increasing T lymphocytes were demonstrated in CLs at the functional regression stage (early stage of regression). At the structural regression stage (late stage of regression), abundant T lymphocytes but no macrophages were demonstrated in the CLs. A moderate amount of TNF-alpha was detected in all CLs at all stages. No IFN-gamma was detected in either healthy or early regressing CLs, but a large amount of IFN-gamma was detected at the late regression stage. Moreover, in cultured luteal cells, reactivity against Fas-ligand (FasL) was caused by pretreatment with TNF-alpha and IFN-gamma and apoptosis was induced by FasL treatment. These findings support the hypothesis that macrophages initiate T lymphocyte aggregation at the early stage of luteal regression, and then T lymphocytes induce apoptosis on luteal cells, which in turn develop sensitivity against FasL by TNF-alpha and IFN-gamma.
Subject(s)
Corpus Luteum/cytology , Corpus Luteum/immunology , Interferon-gamma/metabolism , Luteolysis/physiology , Macrophages/metabolism , T-Lymphocytes/metabolism , Tumor Necrosis Factor-alpha/metabolism , Animals , Apoptosis/drug effects , Cell Count , Cell Survival/drug effects , Cells, Cultured , Corpus Luteum/drug effects , Fas Ligand Protein , Female , Immunohistochemistry , Interferon-gamma/pharmacology , Luteolysis/immunology , Membrane Glycoproteins/metabolism , Membrane Glycoproteins/pharmacology , Mice , Tumor Necrosis Factor-alpha/pharmacologyABSTRACT
Senescence accelerated mouse-prone (SAMP) mice with a shortened life span show accelerated changes in many of the signs of aging and a shorter reproductive life span than SAM-resistant (SAMR) controls. We previously showed that functional regression (progesterone dissimilation) occurs in abnormally accumulated luteal bodies (aaLBs) of SAMP mice, but structural regression of luteal cells in aaLB is inhibited. A deficiency of luteal cell apoptosis causes the abnormal accumulation of LBs in SAMP ovaries. In the present study, to show the abnormality of Fas ligand (FasL)/Fas-mediated apoptosis signal transducing factors in the aaLBs of the SAMP ovaries, we assessed the changes in the expression of FasL, Fas, caspase-8 and caspase-3 mRNAs by reverse transcription-polymerase chain reaction, and in the expression and localization of FasL, Fas and activated caspase-3 proteins by Western blotting and immunohistochemistry, respectively, during the estrus cycle/luteolysis. These mRNAs and proteins were expressed in normal LBs of both SAMP and SAMR ovaries, but not at all or only in trace amounts in aaLBs of SAMP, indicating that structural regression is inhibited by blockage of the expression of these transducing factors in luteal cells of aaLBs in SAMP mice.
