Alpha 2-macroglobulin and serum preferentially counteract the mitoinhibitory effect of transforming growth...

Transforming growth factors-beta 1 and beta 2 (TGF-beta 1 and TGF-beta 2) are potent, multifunctional modifiers of cellular proliferation and differentiation in many cell types. To evaluate factors which may alter the activity of the TGF-beta s during hepatocyte proliferation, we examined the influences of bovine serum and purified bovine or human alpha 2-macroglobulin (alpha 2M) on the mitoinhibitory effects of the TGF-beta in primary cultures of rat hepatocytes. The inhibitory activity of TGF-beta was evaluated by autoradiographic labeling index at 48 hours in hepatocyte cultures exposed to [3H]thymidine between hours 24 and 48 in culture. In the absence of serum or alpha 2M, TGF-beta 1 and TGF-beta 2 were equivalently potent in inhibiting S-phase DNA synthesis in hepatocytes cultured with or without epidermal growth factor. However, bovine serum and purified bovine or human alpha 2M consistently counteracted the mitoinhibitory effects of TGF-beta 2. S-phase DNA synthesis increased five- to six-fold when bovine serum (15%) or alpha 2M (200 micrograms/ml) were included with TGF-beta 2 (0.1 ng/ml) and epidermal growth factor (20 ng/ml). The mitoinhibitory effect of TGF-beta 1 was not influenced by the addition of purified bovine alpha 2M. Human alpha 2M or bovine serum counteracted inhibition by TGF-beta 1 to a lesser extent. [125I]TGF-beta 1 and 125I]TGF-beta 2 formed complexes with purified bovine alpha 2M and serum proteins migrating at identical positions to purified alpha 2M during nondenaturing polyacrylamide gel electrophoresis. However, [125I]TGF-beta 1 associated preferentially with the "fast" migrating form of alpha 2M, whereas [125I]TGF-beta 2 associated with both the "slow" and "fast" migrating forms. Inhibitory activity of TGF-beta 1 and TGF-beta 2 coeluted with alpha 2M in the high molecular weight fractions from a Sephacryl S-200 column. These results support the hypothesis that purified alpha 2M and alpha 2M in bovine serum binds both TGF-betas but preferentially counteracts the mitoinhibitory effect of TGF-beta 2 on rat hepatocytes.