Ação da disintegrina insularina (GST-INS) na expressão de genes pró-metastáticos de células de melanoma (SK-MEL-28) estimuladas por plaquetas

Abstract

The αIIbβ3 integrin expressed on the surface of platelets and the αvβ3 integrin, expressed in tumor cells, mainly melanomas, modulate cell adhesion, migration, cell growth and survival. In addition, it generates activation of tumorigenic factors, that is, genes expressed by the tumor machinery capable of generating aggressive phenotypes favoring adhesion, angiogenesis and tumor proliferation. Therefore, this study aimed to evaluate the ability of activated platelet membranes to stimulate SK-MEL-28 melanoma cells to express pro-metastatic genes such as αV, MCAM, β3, MMP9, MMP2, NFκb and PIK3 and to identify the ability of the recombinant disintegrin, insularin GST-INS to negatively modulate these mechanisms. For this, real-time PCR standardization assays were performed with cDNA extracted from the incubation of platelet membranes obtained from whole blood of healthy donors with SK-MeL-28 cells treated or not with Insularin (GST-INS) and GST (control protein).; the activated membrane was collected by centrifugation of PRP (platelet-rich plasma) followed by heat shock (37oC and PBS at 4oC). The results of real time PCR assays showed that platelet membranes stimulate the expression of αlphaV integrin, NFκb and PIK3 genes in melanoma cells, but do not stimulate MMP9, MMP2, MCAM and β3 genes, and the GST-INS does not appear to have a modulatory action in these mechanisms. We conclude that platelets are a major pro-metastatic modulating factor and further assays with consolidated standardization and evaluation of other genes expressed by the tumor that generate metastatic potentials are still needed.