Caracterização morfológica e molecular de células epiteliais tímicas humanas após estímulo com TGF-β1 in vitro

Abstract

The thymus is a lymphoid organ responsible for fundamental events for the development of the organism’s cellular immunity, such as maturation, differentiation and selection of T lymphocytes. In this context, thymic epithelial cells (TECs), which make up the majority of the thymic microenvironment, they actively participate in the differentiation stages of functionally competent T lymphocytes. TGF-β1 is a protein related to epithelial-mesenchymal transistion (EMT) and oxidative stress, important events for the process of thymus involution with age. Therefore, the objective of this work was to evaluate the morphological and molecular changes induced by TGF-β1 in thymic epithelial cells in vitro. For this, TECs from the human THPN lineage were used and the parameters of morphology, laminin production, activation of the canonical TGF-β pathway and expression of genes related to TEM and oxidative stress were evaluated. Furthermore, expression of the leucemia inhibitory factor (LIF) and its receptor, LIF-R, genes was seen. TGF-β1 was able to induce changes in the morphology of TECs, which presented a larger and more elongated morphology compared to cells without stimulation. Furthermore, an increase in fluorescence intesity for phosphorylated SMAD3 was seen when cells were stimulated with TGF-β1 and an increase in laminin production. Through qPCR, it was possible to observe that TGF-β1 increased the relative expression of the gene for N-cadherin, a marker of EMT, and decreased the expression of the LIFR gene. Finally, a reduction in the relative expression of the gene for the Catalase enzyme (CAT) was seen when TECs were stimulated with TGF-β1. Thus, it can be concluded that TECs, when stimulated with TGF-β1, present cellular and molecular modifications similar to those found in TECs during thymic involution, such as an increase in extracellular matrix, modification to a mesenchymal profile and loss of antioxidant homeostasis, making it a possible instrument for studying changes in TECs during aging.