Análise fenotípica e funcional das células T que expressam receptores do tipo toll (TLR)-2 e TLR-4 e células B no sangue periférico de pacientes com cavernomas cerebrais e sua relação com a gravidade da doença

Abstract

Cerebral cavernous malformations (CCM) are vascular lesions containing grossly dilated capillaries and altered permeability, predisposing patients to a risk of hemorrhagic stroke, epilepsy and various neurological deficiencies. The outcome of the disease appears to be influenced by elements of the immune response. This hypothesis is reinforced by the finding of intense T and B cell infiltration in the cavernoma regions and the involvement of the toll-like receptor (TLR) 4 with the development of CCM. Therefore, the aim of the present study was to evaluate the cytokine profile of T cells expressing TLR2 and TLR4, as well as subsets of B cells, in asymptomatic (CCMAssint, n=14) and symptomatic (CCMSint, n=23) patients. For that, blood samples were collected and the mononuclear cells, or T lymphocytes, were stimulated with polyclonal activators of T cells (anti-CD3/anti-CD28 antibodies) or with TLR2 (Pam3C) and TLR4 (lipopolysaccharide, LPS) ligands. Phenotypic characterization of T cells and B cells was conducted by flow cytometry. The release of cytokines in the culture supernatants was quantified using the ELISA technique. In the present study, it was observed that, when compared to CCMAssint patients, CCMSint patients had a higher frequency and density of TLR4 expressed in CD4+ and CD8+ T cells. The percentage of Th17 cells expressing TLR2 and TLR4 was also higher in CCMSint patients. Furthermore, the occurrence of symptoms in patients with CCM was associated with greater expansion of Tc-1TLR4+ cells, as well as Tc-17 and Th17.1 cells expressing TLR2 and TLR4. On the other hand, the percentage of CD4+TLR4+IL-10+ T cells was higher in the CCMAssint group. In vitro, Pam3C and LPS were more efficient in increasing the frequency of CD4+IL-6+ T cells and Th17.1 cells in the CCMSint group. In addition, compared to CCMAssint, purified T cells from the CCMSint group released higher levels of Th17-related cytokines in response to Pam3C and, especially, in response to LPS. The same behavior was observed in the T cell cultures of these patients after stimulation via TCR/CD28. Regarding B cell subtypes, a higher frequency of activated memory B cells was observed in the CCMSint group. Although preliminary, our findings reveal an increase in circulating Th17/Tc-17 cell subtypes expressing functional TLR2 molecules, and especially TLR4, associated with an increase in memory B cell subsets in CCM patients with clinical disease activity