Heat shock protein 27 (Hsp27), one of the small heat shock proteins, is an intracellular stress protein that confers cytoprotection to a variety of cellular stressors. Recent reports have established that the overexpression of Hsp27 mammalian cell lines confers protection against several noxious stresses like ischemia/reperfusion, hyperthermia, hypertonic stress, and various cytotoxic agents, including cytoskeletal disruptors. Several lines of evidence have suggested that augmentation of the heat shock response by increasing the expression of Hsp27 represents a potential therapeutic strategy for the treatment of critically ill patients. In this study, we used a novel approach to study the potential cytoprotective effect of Hsp27 as a therapeutic agent in rat myogenic cell line H9c2 and rat vascular smooth muscle cell (rVSMC). In the current study, we cloned a human Hsp27 cDNA gene and fused it with a gene fragment encoding the basic domain from 49-57 of HIV-1 TAT protein called the protein transduction domain (PTD). We then analyzed the efficiency of transduction of the resulting TAT-Hsp27 fusion protein into rat myogenic cell line H9c2. Finally we determined the ability of TAT-mediated transduction of the Hsp27 protein to confer cytoprotection against a serum starvation and hypoxic condition. The transduction of TAT-Hsp27 to cardiac cells protects against hypoxia-mediated cell death. The experimental data we reported here suggest that the PTD-mediated delivery of therapeutic proteins may represent a novel strategy of protein therapy.