Proliferation assays Tissue culture plates were coated with purified fibronecti

Nevertheless, as stated in the Background section, analy sis of clinical melanomas suggests large INNO-406 溶解度 variations in expression of Bcl 2 and related proteins, which possibly depend on the growth rate, the TNM staging, andor the type of therapies applied before clinical samples were obtained. Obviously, potential adaptive changes of pro tein expression, under in vivo conditions, could be reflected by changes in the anti death resistance potential of melanoma cells. Studies by Stein and coworkers showed that the Bcl 2 protein profoundly affects the ability of human 518A2 melanoma cells to grow as human tumor xenografts. However, in vivo expression or silencing of other death related proteins was not evaluated in these studies.<br><br> In fact it appears reasonable to expect that, in addition to Bcl 2, other Bcl 2 related proteins may also play growth regulating effects in human melanoma. We have addressed these questions in the A375 mela noma model showing that A375 cells, in vivo, down reg ulate pro death bax expression, and up regulate anti death bcl 2, bcl xl, and mcl 1 relative to cells cultured Lapatinib 分子量 in vitro. The impact of these changes were inves tigated by reversing them in vivo using modified A375 cells grown as xenografts. Forced bax overex pression or mcl 1 silencing did not affect A375 growth as compared to controls. A375 melanoma cells with reduced bcl xl grew in vivo, but more slowly than A375 controls. In parallel, we found that A375 with reduced bcl 2 expression failed to grow after implantation and tended to regress.<br><br> Thus suggesting that Bcl xl and Bcl 2 are not functionally interchangeable in vivo, and that Bcl 2 alone could be a relevant target in melanoma therapy. Why would Bcl 2, which is only one of the many regu lators of apoptosis, be essential for melanoma progres sion in vivo The more aggressive behaviour of different Bcl 2 overexpressing LY2109761 700874-71-1 melanomas was asso ciated to an increase in several metalloproteases expression, and to an elevated microvessel density as compared to parental cells. In agreement with those findings it was shown, e. g. that a Bcl 2 promotes invasion and lung metastasis of Bcl 2 overexpressing non small cell lung cancer cells by inducing matrix metalloproteinase 2. b mem brane type 1 matrix metalloproteinase promotes human melanoma invasion and growth.<br><br> and c Bcl 2 overex pression in human melanoma cells increases angiogenesis through VEGF mRNA stabilization and HIF 1 mediated transcriptional activity, and regulates HIF 1alpha protein stabilization in hypoxic melanoma cells via HSP90. Nevertheless these possibilities have been questioned sincea Bcl 2 down regulation in 518A2 melanoma cells did not associate with down regulation of MMP 2 or MMP 9. and b also in 518A2 tumor cells, transfected with the Bcl 2 plasmid and growing as xeno grafts, extensive necrosis in the setting of very poor vas cularity was observed. Therefore, correlations between Bcl 2 overexpression and MMPs or angiogenesis lack experimental evidence. Besides it is known that Bcl 2 and Bcl xl are antiproliferative by facilitating arrest at G0 G1. The dual functions in apoptosis and cell cycle are coordinately regulated by the multi domain Bcl 2 family members and suggest that survival is maintained at the expense of proliferation.