Avaliação de mecanismos celulares e moleculares envolvidos na ação cicatrizante gástrica de frações do látex de Vasconcellea cundinamarcensis

Abstract

P1G10 is a cysteine protease rich fraction from Vasconcellea cundinamarcensis latex obtained through chromatographic separation in Sephadex G10 column. Previous studies demonstrated the gastric healing activity of P1G10 in lesions induced by acetic acid in rats. The effects were dependent on its proteolytic property and related to an increase of cell proliferation and angiogenesis. Continuing the pharmacological studies of V. cundinamarcensis latex, here, we evaluated the gastric healing properties of P1G10 subfractions, CMS1 e CMS2, as well as some pathways involved. Both fractions – CMS1 at 1mg/kg and CMS2 at 30 mg/kg – display healing activities in lesions caused by acetic acid, with similar intensity to P1G10 10 mg/kg and Ranitidine 100 mg/kg, and were independent on their proteolytic action. Histological and immunohistochemical analysis showed higher proliferation, angiogenesis and tissue remodeling in gastric lesions treated with P1G10, CMS2 and, in special, CMS1. These effects were related to a modulation on growth factors and mediators gene expression by the fractions; CMS1 increased mRNA copies for bFGF, VEGF and COX-1, and reduced COX-2 and TIMP-2 ones. Both MMP-2 gene expression and activity were increased by CMS1 treatment. P1G10 and CMS2 increased EGF and VEGF transcripts, reduced COX-2 and TIMP-2, but did not alter expression/activity of MMP-2. In vitro investigation confirmed the proliferative action of CMS1 and CMS2 on epithelial (AGS) and endothelial (HUVEC-CS) cells, beyond it demonstrated a cell migratory stimulus by the fractions. The proliferative and migratory effects by CMS1 and CMS2 were dependent on the activation of Ras/Raf/MEK/ERK pathway, and seem to involve the participation of EGFR. The proteolytic activity is important for proliferative actions by the fractions, however it just influence the migration activity of CMS2. Both fractions promote cytoskeleton modifications in AGS cells through actin polymerization and stress fiber formation. As completion, CMS1 and CMS2 display interesting gastric healing activities, however CMS1 showed a greater potency comparing to CMS2 or P1G10 and more intensive pro-healing effects.