Tetramethylpyrazine confers cardioprotection against I/R injury by the Hsp70/NF-κB dependent pathway

Background and Objective: Tetramethylpyrazine (TMP), one of the active ingredients of the Chinese herb Lingusticum Wallichii Frantchat (Chuan Xiong), plays an important role in cardioprotection against I/R injury. However, the underling mechanisms remain unknown. This study investigated the effects of TMP on myocardial ischemia-reperfusion (I/R) injury in rats and explored its possible related mechanism. Methods: The mice were anesthetized with pentobarbital intraperitoneally and mice were subjected to myocardial ischemia (6 h) and reperfusion (24 h). Animals were randomly classified into 3 groups (n = 6 each). (1) sham-operated mice (2) ischemia (6 h) and reperfusion (24 h) (3) 10 mg/kg TMP iv into the right jugular vein at the time of reperfusion. A ischemia reperfusion model was constructed in vitro using H9c2 cells. The hearts were taken for histopathological examination and infarct size was detected with 2, 3, 5-triphenyltetrazolium chloride staining. Expression of Hsp70, NF-kappa B p65, PUMA, pro-caspase-3, cleaved caspase-3, IL-6, IL-1 beta, and TNF-alpha protein and mRNA were detected by Western blot and RT-PCR assay. Caspase-3 activity was detected along with IL-6, IL-1 beta, and TNF-alpha protein determined by ELISA. NF-kappa B activity was detected by EMSA. Cell apoptosis was detected using terminal deoxynucleotidyl nick-end labeling (TUNEL). Results: The mice displayed larger infarct sizes and enhanced cardiac myocyte apoptosis following I/R. The effects were associated with a marked activation of NF-kappa B and increased IL-6, IL-1 beta. and TNF-alpha protein as well as pro-apoptotic PUMA protein. TMP significantly reduced infarct size compared to the sham-treated group (14.6% +/- 2.3% vs. 41.3% +/- 7.4%, P < 0.01). These beneficial effects were associated with a marked increase of Hsp70 in the cardiac myocyte in vivo and in vitro, followed by inactivation of myocardial induced I/R NF-kappa B and PUMA as well as reduction of IL-6, IL-1 beta, and TNF-alpha level. TMP increased Hsp70 expression and reduced expression of NF-kappa B and PUMA genes. Conclusions: TMP protects myocardial I/R injury by inhibiting cell apoptosis and reducing inflammatory infiltration. The protective effects of TMP are associated with activation of Hsp70, resulting in the inactivation of NF-kappa B and PUMA, as well as reduction of inflammatory factors. Drug development strategies specifically targeting Hsp70 may be beneficial in treating ischemic heart disease.