SURFACE STAINING AND CYTOTOXIC ACTIVITY OF HEAT-SHOCK PROTEIN-60 ANTIBODY IN STRESSED AORTIC ENDOTHELIAL-CELLS

Heat-shock protein (hsp) expression can be induced by high temperature , exposure to cytokines or oxygen radicals, ischemia, hemodynamic over load, or viral infections. To determine whether surface expression of hsp60 occurs in aortic endothelial cells stressed by high temperature or cytokines, cells from rat aortas were cultivated and stained with s everal types of monoclonal antibodies against hsp60. Other antibodies, eg, those against intercellular adhesion molecule-1 (ICAM-1), or immu ne response-associated antigens were also used as controls. Positive s taining of endothelial cells on the surface and in the cytoplasm was o bserved after pretreatment of the cells with cytokine-containing mediu m, tumor necrosis factor-alpha (TNF-alpha), or interleukin-1 alpha and labeling with a specific monoclonal antibody against hsp60 (II-13). F luorescence-activated cell sorter analyses showed that over 80% of liv ing endothelial cells stressed by cytokine-containing medium, by TNF-a lpha, or at 42 degrees C, but not by interleukin-1 alpha, were positiv ely surface stained with this antibody. Increased intensity of immunos taining with antibodies to ICAM-1 and immune response-associated antig en was also seen on the cytokine-stressed endothelial cells. Furthermo re, when TNF-alpha stimulated endothelial cells labeled with Cr-51 wer e incubated with antibody II-13 in the presence of complement, signifi cant lysis occurred. In summary, endothelial cells stressed by high te mperature or certain cytokines, eg, TNF-alpha, express hsp60 in the cy toplasm and on their surfaces, and these cells were susceptible to com plement-dependent lysis by hsp60-specific antibody. These observations may be significant for elucidating the mechanisms of the involvement of immune reactions to hsp65/60 in initiating atherosclerosis.