In this report, we have developed a panel of monoclonal anti-idiotypic antibodies to pGH by immunising BALB/c mice with a purified monoclonal anti-pGH antibody (1A3), among which one mAb, termed CG-8F, was selected for further characterisation. We found that CG-8F behaved as a typical Ab2β, not only conformationally competing with pGH for 1A3 but also exhibiting recognition for GHR in a rat hepatocyte model. We next examined the resulting signal transduction pathways triggered by this antibody in rat hepatocytes and found that both pGH and CG-8F could trigger the JAK2-STAT1/3/5-mediated signal transduction pathway. Furthermore, the phosphorylation kinetics of pSTAT1/3/5 induced by either pGH or CG-8F were remarkably similar in the dose-response and time course rat hepatocyte experiments. In contrast, only pGH, but not CG-8F, was capable of inducing ERK phosphorylation. Further experimental studies indicated that the two functional binding sites on CG-8F are required for GHR activation. This study partially reveals the mechanism of action of GH anti-idiotypic antibodies and also indicates that monoclonal anti-idiotypic antibodies represent an effective way to produce GH mimics, suggesting that it is possible to produce signal-specific cytokine agonists using an anti-idiotypic antibody approach.
Keywords: Ab2; Anti-idiotypic monoclonal antibody; CLSM; FITC; GHR; Growth hormone; JAK; Janus kinase; MAPK; PBS; PRL; STAT; Signal transduction; TMB; TPO; anti-idiotypic antibody; confocal laser scanning microscope; fluorescein isothiocyanate; growth hormone receptor; mAb; mitogen-activated protein kinase; monoclonal antibody; pGH; phosphate-buffered saline; porcine growth hormone; prolactin; signal transducers and activators of transcription; tetramethylbenzidine; thrombopoietin.
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