Differential inhibition of recombinant bovine GH (rbGH) activity in vitro by in vivo enhancing monoclonal antibodies.
We have previously described the effects of complexing recombinant bovine growth hormone (rbGH) with the in vivo enhancing monoclonal antibodies (Mabs) OA11 and OA15 and the non-enhancing Mab OA14 on the subsequent activity of GH in different tissue culture models. We reported that all of these Mabs caused the inhibition of GH-stimulated Jak-2 tyrosine kinase phosphorylation in the GH responsive pre-adipocyte cell line 3T3-F442A. However, using the mouse myeloid cell line FDC-P1 transfected with the full length ovine GH receptor (GHR), we subsequently found that OA11 and OA14 remained inhibitory with respect to the end point measurement of GH stimulated mitogenesis but that OA15 had no inhibitory effect on GH stimulated mitogenesis in this cell line. In order to correlate longer term mitogenic effects of Mab-GH complexes with signalling events in this transfected cell line model, we now report on the effects of complexing with Mab on the subsequent GH stimulated phosphorylation of Stat5b (signal transducer and activator of transcription). In agreement with our data for the mitogenic activity of GH-Mab complexes, we found that OA11 and OA14 inhibit GH activation of Stat5b but that OA15 is not inhibitory. Further to this, the dose-response effect of both OA11 and OA14 on the GH stimulation of Stat5b in the FDC-P1-oGHR transfected cells correlates with the previously described dose-response effects for both Mabs in the context of GH stimulation of mitogenic effects. We conclude that in this oGHR transfected cell line model, Mab effects on short and long term GH signalling events are tightly correlated. The observation that neither of the in vivo enhancing Mabs--OA11 or OA15--amplifies the response to GH in our transfected cell line model, coupled with the differential nature of Mab effects on GH activity (OA11--inhibition; OA15--no effect) may argue for an in vivo mechanism for enhancement of GH activity.
Beattie
J
J
Molecular Recognition Department, Hannah Research Institute, Ayr, Scotland, UK.
Phillips
K
K
Borromeo
V
V
D000818:Animals; D000911:Antibodies, Monoclonal / Q000378:metabolism*; D015153:Blotting, Western; D002417:Cattle; D002478:Cells, Cultured; D004268:DNA-Binding Proteins / Q000378:metabolism; D004305:Dose-Response Relationship, Drug; D013006:Growth Hormone / Q000037:antagonists & inhibitors* / Q000737:chemistry*; D008894:Milk Proteins; D011485:Protein Binding; D011994:Recombinant Proteins / Q000037:antagonists & inhibitors*; D050799:STAT5 Transcription Factor; D012756:Sheep; D015398:Signal Transduction; D013997:Time Factors; D015534:Trans-Activators / Q000378:metabolism; D014162:Transfection
D000911:Antibodies, Monoclonal; D004268:DNA-Binding Proteins; D008894:Milk Proteins; D011994:Recombinant Proteins; D050799:STAT5 Transcription Factor; D015534:Trans-Activators; D013006:Growth Hormone