The Effect Of Tunicamycin Pretreatment On Septic Neonatal Rat Cardiomyocytes Activity And Mitochondrial Function

Objective:To investigate the effect of tunicamycin pretreatment on septic neonatal rat cardiomyocytes activity and mitochondrial function.Methods: Neonatal rat cardiomyocytes were isolated and cultured in vitro. Section 1 - Cardiomyocytes were exposed to different concentrations of tunicamycin (control, 0.05μg/ml, 0.1μg/ml, 0.5μg/ml). GRP78 expressions were determined by Western Blot. Chose an appropriate concentration which can induce effective endoplasmic reticulum stress. Section 2 - Cardiomyocytes were divided into four groups and cultured by mediums with different drugs : control(normal DMEM), T(0.5μg/ml tunicamycin), L(1μg/ml LPS), T+L(0.5μg/ml tunicamycin followed by 1μg/ml LPS). Variability of myocytes were measured by MTT assay, potential changes of mitochondrial were observed by fluorescence microscope, intracellular ROS levels were monitored by flow cytometry.Results:Section 1– Only 0.5μg/ml tunicamycin can induce effective ER Stress with dramatically up-regulated production of GRP78 ( P<0.01 ). Section 2– 0.5μg/ml tunicamycin alone did not affect myocytes activity and mitochondrial function. 1μg/ml LPS did not affect the variability of myocytes either, but it could cause depolarization of mitochondrial, and dramatically increased production of ROS (P<0.05). Variability of myocytes preincubated by tunicamycin (T+L) was dramatically increased compared with LPS group (P<0.05), depolarization of mitochondrial were moderated, and production of ROS was also dramatically decreased (P<0.01).Conclusion:0.5μg/ml tunicamycin induced effective ER Stress with dramatically up-regulated production of GRP78, at the same time, did not affect myocytes activity and mitochondrial function. 1μg/ml LPS did not affect the variability of myocytes, but it caused mitochondrial impairment and oxidative stress. Appropriate ER Stress induced by sub-lethal tunicamycin pretreatment protects neonatal rat cardiomyocytes from consequential LPS injury.