NONGENOMIC EFFECTS OF AN ANTIIDIOTYPIC ANTIBODY AS AN ESTROGEN MIMETIC IN FEMALE HUMAN AND RAT OSTEOBLASTS

We investigated the early effects of the anti-idiotypic antibody (clon e 1D(5)), which recognized the estrogen receptor (ER), on cytosolic fr ee calcium concentration ([Ca2+]i) and its long term effects on creati ne kinase (CK) specific activity in female human and rat osteoblasts. These actions were compared to the known membrane and genomic effects of 17 beta estradiol (E-2). Like E-2, clone 1D(5) increased within 5 s [Ca2+]i in both cell types by two mechanisms: i) Ca2+ influx through voltage-gated Ca2+ channels as shown by using EGTA, a chelator of extr acellular Ca2+, and nifedipine, a Ca2+ channel blocker; 2) Ca2+ mobili zation from the endoplasmic reticulum as shown by using phospholipase C inhibitors, such as neomycin and U-73122, which involved a Pertussis toxin-sensitive G-protein. Clone 1D(5) and E-2 stimulated CK specific activity in human and rat osteoblasts with ten fold higher concentrat ions than those needed for the membrane effects (0.1 mu g/ml and 10 pM , respectively). Both effects were gender-specific since testosterone and 5 alpha-dihydotesterone were uneffective. Tamoxifen and Raloxifene , two estrogen nuclear antagonists, inhibited CK response to 1D(5) and E-2 and Ca2+ response to 1D(5), but not Ca2+ response to E-2. By cont rast, (Fab')(2) dimer, a proteolytic fragment of 1D(5) with antagonist properties, inhibited both membrane and genomic effects of 1D(5) and E-2. In conclusion, these results imply that clone 1D(5) has an estrog en like activity both at the membrane and nuclear levels in female hum an and rat osteoblasts. 1D(5) must therefore interact with membrane bi nding sites, penetrate the cells, and reach the nuclear receptors by a n as yet uncharacterized mechanism. (C) 1997 Wiley-Liss, Inc.