Biomedical subjects

T L Atkins

Publications and source records attributed to T L Atkins.

2 recordsLinked to original sources

PubMed1995

Activated alpha 2-macroglobulin promotes mitogenesis in rat vascular smooth muscle cells by a mechanism that is independent of growth-factor-carrier activity.

Vascular smooth muscle cell (vSMC) proliferation is important in atherosclerosis. We previously demonstrated that methylamine-activated alpha 2-macroglobulin (alpha 2M) and transforming growth factor beta 1 (TGF-beta 1) cause a synergistic proliferative response in quiescent rat aortic vSMCs [Stouffer, G. A., La-Marre, J., Gonias, S. L. & Owens, G. K. (1993) J. Biol. Chem. 268, 18,340-18,344]. The first goal of this study was to determine whether the synergy is due to the ability of alpha 2M-methylamine (alpha 2M-MeNH2) to bind TGF-beta 1 and target the growth factor to vSMCs that express the alpha 2M receptor. Receptor-recognized alpha 2M derivatives without TGF-beta 1-binding activity, including ternary alpha 2M-trypsin, an 18-kDa proteolytic fragment of the alpha 2M subunit, and the corresponding recombinant receptor-binding fragment (rRBF) increased vSMC [3H]thymidine incorporation and cell number in a manner similar to alpha 2M-MeNH2. In combination with TGF-beta 1, each alpha 2M derivative caused a synergistic vSMC proliferative response. vSMCs responded comparably when treated with alpha 2M-MeNH2 and TGF-beta 1 simultaneously or in sequence. Furthermore, alpha 2M-MeNH2-TGF-beta 1 complexes increased [3H]thymidine incorporation no more than alpha 2M-MeNH2 alone. These results indicate that TGF-beta 1 binding to alpha 2M is not responsible for the synergistic mitogenic activity. Additional studies were undertaken to determine whether activated alpha 2M independently induces a signal-transduction response in vSMCs. alpha 2M-MeNH2 and rRBF caused a rapid, transient increase in vSMC inositol 1,4,5-trisphosphate. This response was pertussis-toxin insensitive. Receptor-associated protein (RAP; 170 nmol/L) inhibited 91-95% of the specific binding of 125I-alpha 2M-MeNH2 and 125I-rRBF to vSMC; however, RAP did not affect the inositol 1,4,5-trisphosphate response or the mitogenic response. These studies suggest that vSMCs express a receptor, other than low-density-lipoprotein-receptor-related protein, that transduces a signal in response to activated alpha 2M. This receptor may mediate the mitogenic activity of alpha 2M in vSMC culture.

Animals↗

PubMed1994

Transforming growth factor beta isoform 2-specific high affinity binding to native alpha 2-macroglobulin. Chimeras identify a sequence that determines affinity for native but not activated alpha 2-macroglobulin.

Transforming growth factor beta 2 (TGF-beta 2) is less potent than TGF-beta 1 in some endothelial cell proliferation assays due to the greater tendency of TGF-beta 2 to bind alpha 2-macroglobulin (alpha 2M). Substitution of TGF-beta 1 residues 40-47 into the TGF-beta 2 sequence yields a chimeric molecule that, like TGF-beta 1, expresses activity that is not substantially affected by serum alpha 2M (Burmester, J. K., Qian, S. W., Roberts, A. B., Huang, A., Amatayakul-Chantler, S., Suardet, L., Odartchenko, N., Madri, J. A., and Sporn, M. B. (1993) Proc. Natl. Acad. Sci. U.S.A. 90, 8628-8632). In this investigation, we studied the binding of TGF-beta chimeras, which contain TGF-beta 1 residues 40-47, to both major conformations of human alpha 2M under apparent equilibrium conditions. Native alpha 2M, the primary form of this protein in serum, bound TGF-beta 2/beta 1 (40-82) and TGF-beta 2/beta 1 (40-47) with low affinity. The apparent KD values for the two chimeras and native alpha 2M were 310 and 330 nM, respectively. These values were much higher than the KD determined for TGF-beta 2 and native alpha 2M (11 nM) and equivalent to the KD determined for TGF-beta 1 and native alpha 2M. By contrast, both TGF-beta chimeras bound alpha 2M-methylamine, an altered conformation of alpha 2M, with high affinity (16 and 19 nM), which is characteristic of TGF-beta 2 and not TGF-beta 1. Fetal bovine heart endothelial cell DNA synthesis was inhibited to a similar degree by TGF-beta 1, TGF-beta 2, TGF-beta 2/beta 1 (40-82), and TGF-beta 2/beta 1 (40-47) in the presence of dilute (0.2%) fetal bovine serum. When 0.07 microM alpha 2M-methylamine was added, the activities of TGF-beta 2, TGF-beta 2/beta 1 (40-82), and TGF-beta 2/beta 1 (40-47) were significantly counteracted while the activity of TGF-beta 1 was unchanged, as would be predicted by the equilibrium binding analyses. These studies indicate that the TGF-beta structural elements, which mediate binding to native alpha 2M and conformationally transformed alpha 2M, are not equivalent. Residues 40-47 are critical in determining affinity for native alpha 2M but are less important in determining affinity for alpha 2M-methylamine.

Animals↗