Efeito do ATL-1, um análogo sintético de 15-epi-Lipoxina A4, em células endoteliais moduladas pela interação com células de melanoma humano

Abstract

Tumors arise from a pathological disorder characterized by excessive cell proliferation and are supported by the establishment of a favorable microenvironment. Melanoma has a metastatic profile and originates from melanocytes. Angiogenesis is necessary for tumor growth and metastasis. The vascular endothelial growth factor (VEGF) is the main pro-angiogenic and permeable factor. Tumor microenvironment presents a chronic inflammation profile and an activated extracellular matrix (ECM). Lipoxins (LX) and 15-epi-LX are lipids with potent anti-inflammatory and antiangiogenic effects in different models in vivo and in vitro. However, their effects on endothelial cell (EC) in the tumor microenvironment are unknown. In the present study we investigated the effect of ATL-1, a synthetic analog of 15-epi-LX, on EC, in a model that mimics the tumor microenvironment. Initially, the inhibitory effect of ATL-1 on VEGF-induced endothelial permeability was characterized. This action was related to the stabilization of VE-cadherin and reduction of β-catenin nuclear translocation. Additionally, in co-culture assays, the pretreatment of EC (HUVEC) with ATL-1 reduced the transendothelial migration of highly metastatic melanoma cells (MV3), but not normal melanocytes (NGM). The secretion of VEGF by HUVEC, MV3 and NGM was determined by ELISA. The MV3 cells secreted large amounts of VEGF in a time-dependent manner and ATL-1 did not alter this action. The correlation between the efficiency of MV3 cells to migrate through an endothelial monolayer and the accumulation of VEGF was observed in an assay where the endothelial monolayer was stimulated with exogenous VEGF, in a concentration comparable to levels secreted by MV3 cells. In this test, we observed an increase in cell migration of both cells which was inhibited by ATL-1. The influence of the reactive stroma of the endothelium was also evaluated using the ECM extracted from melanoma. Tumor matrix supports endothelial growth inducing HUVEC monolayer permeability, increasing VEGFR-2, the pro-angiogenic cytokine IL-8 and its receptor, CXCR-2, gene expression. Also, tumor-derived ECM induced a pro-inflammatory profile in HUVEC characterized by the production of nitric oxide and the adhesion molecule VCAM-1 gene and protein expression, which was followed by an increased adhesion of activated mononuclear cells. These actions were inhibited by the treatment of endothelial cells with ATL-1. The increase in the tubulogenic capacity presented by endothelium in contact with the matrix was also inhibited by ATL-1. Taken together, these data suggest an intrinsic relation between endothelium and melanoma-derived ECM and a role in melanoma progression and metastasis. Moreover, in this scenario, ATL-1 appears as a powerful anti-inflammatory and anti-metastatic tool.