Estudo das células endoteliais humanas frente a estímulos inflamatórios: urease de Helicobacter pylori e o tecido adiposo na obesidade

Abstract

Alterations in endothelial cells (ECs) play a central role in the pathogenesis of a broad human pathologies, such as obesity and infections. During the onset of many diseases, the continuous activation of endothelium by pro-inflammatory stimuli favors the persistence of inflammation, becoming a chronic process. Our group has been studying the mechanisms that lead to changes in ECs functions induced by pathogenic factors produced or released during chronic inflammation, caused by bacterial infections or chronic degenerative diseases. In this study, human endothelial cell cultures (HMEC1) were used in an in vitro model to evaluate: (1) the effect of Helicobacter pylori urease (HPU), a key virulence factor with proinflammatory properties, essential for colonization gastric mucosa, and (2) the effect of the conditioned media and microparticles isolated from cultures explants adipose tissue (AT) obtained from obese and non-obese subjects. In the first part of this study, we have shown that HPU, independenly of its ureolytic activity, enhanced permeability in HMEC1 and reduced trasendothelial resistance, causing disruption of cell–cell contact, with dissociation of cadherin from adherens junctions and VE-cadherin phosphorylation, accompanied by alterations in actin cytoskeleton dynamics and increased FAK phosphorylation. HPU also stimulated NO and ROS production by ECs and induced NF-kB activation, enhancing the expression of COX-2, HO-1, pro-IL-1β and adhesion molecules. Higher expression of ICAM-1 and E-selectin allowed an increased adhesion of human neutrophils to HMEC1 monolayer, confirming the pro-inflammatory properties of HPU. Cells treated with HPU also presented increasing expression of VEGFR-2, and underwent differentiation and tubulogenesis. The effects induced by HPU in HMECs are, at least in part, modulated by integrin signaling activation and dependent on redox metabolism and lipoxygenase pathway activation, being attenuated by RGD peptides and esculetin, respectively. The data support the role of HPU as a pro-inflammatory virulence factor that contributes to chronic inflammation induced by the bacteria. In the second part, we have shown that the conditioned media and microparticles, obtained from obese AT explants cultures, increased endothelial permeability, which was associated to VE-cadherin phosphorylation and actin cytoskeleton rearrangement in CEs. Additionally, these stimuli also enhanced invasion and the proliferative capacity of these cells, allowing the increase of tubulogenesis. ECs stimulated by obese adipose microenvironment also showed an increase in NO and ROS production, which resulted in an up-regulated expression of HO-1. The data indicate that obese AT, contrasting with the AT from lean subjects, release soluble pro-inflammatory and pro-angiogenic mediators that may modulate endothelial dysfunction and differentiation. Together, our data contribute to a better understanding of the role of different factors, either derived from microorganisms (HPU) or produced by AT during obesity, able to induce ECs modifications and contribute to the onset inflammatory process and also to the persistence and progression of diseases.