Experimental study of compound BAM15 alleviating cryopreservation-induced primary hepatocyte injury in rats / 器官移植

Objective To investigate the effect of compound BAM15 on the primary hepatocyte injury induced by cold storage in rats. Methods The primary rat hepatocytes were extracted by collagenase perfusion method. According to different cell culture conditions, the cells were divided into 4 groups: group A (Hibernate cell culture solution containing 250 nmol/L BAM15), group B (Hibernate cell culture solution containing 500 nmol/L BAM15), group C (Hibernate cell culture solution containing 1 000 nmol/ L BAM15), control group (Hibernate cell culture solution). The cells of each group were cryopreserved for 12 h. The purity of primary hepatocytes was observed under fluorescence microscope. The changes in the cell proliferation ability, cell apoptosis rate and mitochondrial reactive oxygen species (ROS) were measured in each group. Results The cell proliferation ability in groups B and C was significantly higher than that in the control group (both P < 0.05). The apoptosis rates in groups A, B and C were (33.7±2.2)%, (19.7±1.1)% and (28.7±1.2)%, which were significantly lower than (82.7±4.2)% in the control group (all P < 0.05). The positive rates of intracellular ROS in groups A, B and C were (11.8±4.0)%, (7.6±1.3)% and (8.9±1.6)%, remarkably lower than (27.4±4.5)% in the control group (all P < 0.05). Conclusions Compound BAM15 can effectively mitigate the primary hepatocyte injury in rats induced by cryopreservation. The underlying mechanism is probably associated with the role of BAM15 in reducing ROS generation during cold ischemia.

Mitochondrial uncoupler BAM15 inhibits artery constriction and potently activates AMPK in vascular smooth muscle cells

Our previous studies found that mitochondrial uncouplers CCCP and niclosamide inhibited artery constriction and the mechanism involved AMPK activation in vascular smooth muscle cells. BAM15 is a novel type of mitochondrial uncoupler. The aim of the present study is to identify the vasoactivity of BAM15 and characterize the BAM15-induced AMPK activation in vascular smooth muscle cells (A10 cells). BAM15 relaxed phenylephrine (PE)-induced constricted rat mesenteric arteries with intact and denuded endothelium. Pretreatment with BAM15 inhibited PE-induced constriction of rat mesenteric arteries with intact and denuded endothelium. BAM15, CCCP, and niclosamide had the comparable IC value of vasorelaxation in PE-induced constriction of rat mesenteric arteries. BAM15 was less cytotoxic in A10 cells compared with CCCP and niclosamide. BAM15 depolarized mitochondrial membrane potential, induced mitochondrial fission, increased mitochondrial ROS production, and increased mitochondrial oxygen consumption rate in A10 cells. BAM15 potently activated AMPK in A10 cells and the efficacy of BAM15 was stronger than that of CCCP, niclosamide, and AMPK positive activators metformin and AICAR. In conclusion, BAM15 activates AMPK in vascular smooth muscle cells with higher potency than that of CCCP, niclosamide and the known AMPK activators metformin and AICAR. The present work indicates that BAM15 is a potent AMPK activator.