Immunochemical characteristics of a novel cell death receptor and a decoy receptor on granulosa cells of porcine ovarian follicles

Previously, we prepared an IgM monoclonal antibody (PFG-1) which specifical ly recognized a cell-membrane glycoprotein (PFG-1 antigen; 55 kD, pI 5.9), immunohistochemically reacted with granulosa cells of healthy follicles but not of atretic follicles, and induced granulosa cell apoptosis. In the pre sent study, an IgM monoclonal antibody (PFG-3) capable of inducing granulos a cell apoptosis and an IgG monoclonal antibody (PFG-4) not capable of indu cing apoptosis were produced against granulosa cells prepared from healthy antral follicles of porcine ovaries. Two-dimensional Western blotting analy sis revealed that PFG-3 specifically recognized two cell-membrane proteins (named PFG-3-1 and PFG-3-2/PFG-1 antigens; 42 kD, pI 5.2 and 55 kD, pI 5.9, respectively) of healthy granulosa cells, and that PFG-4 recognized the sa me two cell-membrane proteins. In atretic granulosa cells, PFG-3-2/PFG-1 an tigen disappeared. Immunochemical reactions of these antibodies were only d etected in follicular granulosa cells but not any other ovarian tissues or organs. PFG-3 and PFG-4 immunohistochemically reacted with granulosa cells of healthy and atretic follicles. When the isolated granulosa cells prepare d from healthy follicles were cultured in medium containing PFG-3, the cell s underwent apoptosis, and co-incubation with PFG-4 inhibited PFG-3-inducib le apoptosis. These observations suggested that PFG-3-2/PFG-1 antigen is a novel cell death receptor which is different from the apoptosis-mediating r eceptors (Fas/Apo-1/CD95 or TNF receptor), and that PFG-3-1 antigen may act as a decoy receptor and inhibit apoptotic signal transmission.