Expression analysis of Notch-related molecules in peripheral blood T helper cells of patients with rheumatoid arthritis

Abstract

Objective: Expression of Notch homologues in local tissue inflammation in rheumatoid arthritis (RA) and cultured synoviocytes has been reported, but the expression profile of Notch-related molecules in peripheral lymphocytes in RA remains unclear. In this study, we measured the expression of Notch receptors and downstream molecules in peripheral lymphocytes from RA patients.Methods: Expression of Notch receptors in peripheral lymphocytes of RA patients was assessed by both flow cytometry and real-time polymerase chain reaction (PCR). Expression of the representative Notch target gene HES-1 and the regulatory gene NUMB in purified T helper cells from RA patients was determined by real-time PCR, and expression of Notch intracellular domain (ICD) was determined by immunoblot analysis.Results: There was an increased expression of Notch 2, Notch 3, and Notch 4 in T helper cells from active RA patients, among which increased expression of Notch 3 was mainly by activated T cells. Notably, expression of Notch 3 in T cells decreased in inactive RA patients and the level was similar to that of healthy controls (HC). Notch receptors were rarely observed on B cells and no difference in expression was found between RA patients and HC. T helper cells from RA patients exhibited increased expression of the target gene HES-1 but decreased expression of the negative modulation gene NUMB of Notch signalling. There was also an increased nuclear translocation of Notch-ICD in T helper cells from active RA disease.Conclusion: The present study demonstrated that T helper cells from RA patients display a significantly altered expression profile of Notch receptors and enhanced activation of Notch signalling compared with HC.