Beta-hydroxy-beta-methylbutyrate (HMB), a leucine catabolite, has been shown to prevent exercise-induced protein degradation and muscle damage. We hypothesized that HMB would directly regulate muscle-cell proliferation and differentiation and would attenuate apoptosis, the latter presumably underlying satellite-cell depletion during muscle degradation or atrophy. Adding various concentrations of HMB to serum-starved myoblasts induced cell proliferation and MyoD expression as well as the phosphorylation of MAPK/ERK. HMB induced differentiation-specific markers, increased IGF-I mRNA levels and accelerated cell fusion. Its inhibition of serum-starvation- or staurosporine-induced apoptosis was reflected by less apoptotic cells, reduced BAX expression and increased levels of Bcl-2 and Bcl-X. Annexin V staining and flow cytometry analysis showed reduced staurosporine-induced apoptosis in human myoblasts in response to HMB. HMB enhanced the association of the p85 subunit of PI3K with tyrosine-phosphorylated proteins. HMB elevated Akt phosphorylation on Thr308 and Ser473 and this was inhibited by Wortmannin, suggesting that HMB acts via Class I PI3K. Blocking of the PI3K/Akt pathway with specific inhibitors revealed its requirement in mediating the promotive effects of HMB on muscle cell differentiation and fusion. These direct effects of HMB on myoblast differentiation and survival resembling those of IGF-I, at least in culture, suggest its positive influence in preventing muscle wasting.
Several T cell abnormalities have been described in the course of acute Trypanosoma cruzi infection in mice, including severe effects on the thymus. In the present study, looking at the expression of extracellular matrix ligands in the thymus, we observed that deposits of fibronectin and laminin increased progressively during the course of infection, reaching a maximum at the peak of parasitemia and thymic atrophy. Concomitantly, membrane expression of fibronectin and laminin receptors (VLA-4, VLA-5 and VLA-6) was also enhanced on thymocyte subsets of infected mice. These results correlated with changes in intrathymic thymocyte migration ability during the acute phase of infection, when a higher fibronectindependent transmigratory activity of CD4 + CD8 + thymocytes was observed. Strikingly, we detected higher frequency of immature and high VLA-expressing CD4 + CD8 + T cells in the peripheral lymphoid organs of infected mice at the peak of parasitemia. These cells seemed to be thymus dependent, since significantly lower amounts of them were found in thymectomized mice, and some of them carry "prohibited" V g segments of the TCR. Our data suggest an imbalance in the intrathymic cell trafficking following acute T. cruzi infection, likely due to dysregulated extracellular matrix-dependent interactions.
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