Molecular response to ischemia-reperfusion of rat skin: Study of expression of p53, p21(WAF-1), and Bax proteins, and apoptosis

The authors investigated the expression of p53, p21(WAF-1), and Bax protein s, and apoptosis to elucidate the cellular response to ischemia-reperfusion of the skin. The rat left lower limb was dissected at the inguinal region retaining the bone and femoral vessels, and the vessels were clamped to pro duce an ischemic condition. After 6 hours the clamps were removed, and the plantar skin was resected at various times up to 72 hours after reperfusion . Five skin specimens were obtained at each time point from 5 rats. When a rat died during the study, additional rats were used until five specimens c ould be obtained from 5 rats at each time point. The expression of the thre e proteins was detected by Western blot analysis. The apoptotic cells were detected using the terminal deoxytransferase-mediated dUDP nick-end labelin g assay. After reperfusion, the levels of p53 and p21(WAF-1) were significa ntly higher in the ischemia-reperfusion rats compared with the sham-operate d rats. However, the levels of Bax protein did not show a noticeable increa se at any period. The apoptotic cells in both the epidermis and dermis were not evident compared with the sham skin, which were similar to those in th e nontreated, normal skin. These results demonstrate that p53 and p21(WAF-1 ) proteins accumulate after 6 hours of ischemia of the skin during reperfus ion. Moreover, it is speculated that accumulation of these proteins plays a n important role in the survival of the skin by inducing growth arrest of t he cells, but not apoptosis.