Reactivity of γ/δ T cells to human 60-kd heat-shock protein and their cytotoxicity to aortic endothelial cells in Takayasu arteritis

Abstract

Objective: Increased numbers of circulating γ/δ T cells with a restricted T cell receptor repertoire, as well as colocalization of the expression of heat-shock protein Hsp60/65 and γ/δ T cells in the arterial lesions of patients with Takayasu arteritis (TA), indicate that γ/δ T cells may react to Hsp60 and cause damage to the arterial endothelium. In this study we investigated the proliferative responses of γ/δ T cells to human Hsp60 and their cytotoxicity to human aortic endothelial cells (ECs) in patients with TA.Methods: Blood samples were obtained from 12 patients with TA, 8 patients with systemic lupus erythematosus (SLE) (as disease controls), and 10 healthy control subjects. Proliferative responses of circulating γ/δ T cells to human Hsp60 were detected by flow cytometry-based bromodeoxyuridine incorporation assay. Cytotoxicity of the γ/δ T cells to human aortic ECs was analyzed by colorimetric lactate dehydrogenase release assay.Results: The γ/δ T cells of 11 of 12 patients with TA exhibited reactivity to Hsp60, whereas none of the γ/δ T cells from patients with SLE or healthy controls showed reactivity (both P < 0.001). The mean ± SD proliferative response of γ/δ T cells in patients with TA was 21.4 ± 11.3%, compared with 4.2 ± 1.2% in patients with SLE and 4.01 ±1.82% in healthy controls (both P < 0.001). In addition, compared with the control groups, the γ/δ T cells of patients with TA had increased spontaneous cytotoxicity to aortic ECs (22.1 ± 15.0% versus 9.6 ± 2.13% in SLE patients and 8.1 ± 4.7% in healthy controls; both P < 0.005), which was further enhanced following stimulation of γ/δ T cells with Hsp60. The cytotoxicity of the γ/δ T cells was significantly inhibited by treatment of these cells with concanamycin A and anti-Fas ligand-blocking antibodies.Conclusion: The results show that γ/δ T cells in patients with TA are reactive to Hsp60 and exhibit cytotoxicity to aortic ECs, suggesting a key role of Hsp60 and γ/δ T cells in the pathogenesis of TA. Takayasu arteritis (TA) is a chronic granulomatous arteritis characterized by intimal thickening, fibrosis, and stenosis as well as aneurysm of the large elastic arteries, predominantly the aorta and its major branches. Its etiology is largely unknown, but most of the available data suggest that immune-mediated dysfunction of the arterial endothelium is a primary event in the pathogenesis of the disease (1-3). We previously have demonstrated an increased number of circulating activated γ/δ T cells with a restricted T cell receptor (TCR) repertoire in patients with TA (4). Similarly, Seko et al observed the predominance of TCR-restricted γ/δ T cells in the arterial lesions of patients with TA (5). These findings indicate an antigen-driven activation and involvement of these cells in the pathogenesis of the disease. Since a major human heat-shock protein (HSP) antigenic target of γ/δ T cells is Hsp60 (6), and Hsp60/65 is highly expressed in the arterial lesions of patients with TA (7), it appears that γ/δ T cells recognize HSPs or some homologous arterial antigens and transmigrate from the circulation to the vascular wall, thus causing arterial damage that culminates in different clinical manifestations of the disease. In a recent study (8) we detected the presence of anti-endothelial cell (anti-EC) antibodies that were predominantly directed against the 60-65-kd aortic EC antigen, which was suggestive of endothelial Hsp60. These findings indicate that γ/δ T cells may react to this 60-65-kd EC antigen and lead to damage of the endothelium in TA. However, there are no data available on the reactivity of γ/δ T cells to Hsp60 and their cytotoxicity to arterial endothelium in TA. We therefore undertook this study to investigate the proliferative responses of γ/δ T cells to human Hsp60 and their cytotoxicity to human aortic ECs, as well as the mechanism of their cytotoxicity, in patients with TA