Melanoma represents the deadliest of all skin cancers. The comparison of melanoma incidence and mortality rates indicated a worse prognosis with increasing age. Patients with melanoma demonstrated an inverse correlation of melanoma mortality and incidence of sentinel node metastasis in young and old patients. The Wnt5A and HAPLN1 proteins are altered in the aged microenvironment, they seem to be contributors to alterations in extracellular matrix (ECM) integrity, and can influence broadly to the metastatic progression, resistance therapy, and poor prognosis. This project aims to investigate the age-related ECM alterations. Specifically, in vitro studies will be focused on the role of Wnt5A and HAPLN1 proteins in young and aged fibroblasts evaluated by production of cross-linking alterations of collagen protein and ECM integrity. Whether the activation of angiogenesis process compromises the lymphatics vessels and if it results in a hematogeneous route of dissemination will be also a main question of this work. For this, we will build 3D reconstructed skin models using dermal endothelial cells, young and aged fibroblasts with manipulated genes and melanoma cells and we will determine the ECM integrity by specific markers (CK10, CK14, and DDR1) and by microscopy images (Second Harmonic Generation Microscopy). Quantification of invasion of melanoma cells will be performed using ImageJ software. Furthermore, we will also assess the angiogenic (VEGF, bFGF) and lymphangiogenic markers (VEGFC, VEGFD and EGF) by ELISA and by IHC (CD31 and thrombospondin-1, respectively). Using this aged capillarized reconstructed skin, we expect to demonstrate the role of aged microenvironment modulating/impairing the formation of capillary-like tubes, thus modifying the angiogenesis and lymphangiogenesis process.
News published in Agência FAPESP Newsletter about the scholarship: