In the present study, we examined the activity of p53 protein in Jurkat cells treated with high mobility group box-1 protein (HMGB1), thereafter we investigated the mechanism of extracellular HMGB1 mediated p53 expression in immune dysfunction of T lymphocytes. mRNA expression of p53, mdm2, and p21 was determined by Real-time reverse transcription-polymerase chain reaction(RT-PCR). The apoptotic rate of Jurkat cells was analyzed by flow cytometry. Expressions of bcl-2, bax, caspase-3, phosphorylated (p) extracellular signal-regulated kinase (ERK)1/2, ERK1/2, p-p38 mitogen-activated protein kinase (MAPK), p38 MAPK, and p-c-jun amino-terminal kinase (JNK)1/2 and JNK1/2 were simultaneously determined by Western blotting. After treatment with HMGB1 (100 ng/ml or 1000 ng/ml), the proliferative activity of Jurkat cells was significantly decreased, and a low and medium concentration of HMGB1 induced an up-regulation of p53 mRNA, p-p53 and p53 protein expression. Meanwhile, levels of mdm2 and p21 were elevated by incubated with HMGB1 (100 ng/ml) for 24 or 48 hours. Moreover, the proliferation of Jurkat cells in response to HMGB1 (100 ng/ml) in the vector group was significantly depressed. The bax and caspase-3 levels in p53 shRNA-expressed cells treated with HMGB1 (100 ng/ml) was markedly decreased, whereas expression of bcl-2 was obviously enhanced. Among ERK1/2, p38 MAPK and JNK1/2 signaling, only p38 MAPK pathway could be significantly activated by treatment with HMGB1, and the specific inhibitor of p38 MAPK was used, p53 and p-p53 expression induced by HMGB1 were significantly down-regulated. Taken together, our data strongly indicated that HMGB1 might enhance p53 expression, which was associated with both the proliferative activity as well as apoptosis of T cells.