Department of Cardiology, Tangdu Hospital, Fourth Military Medical University, Xi'an 710038, China.
Life Sci. 2013 Jul 30;93(2-3):116-24. doi: 10.1016/j.lfs.2013.06.002. Epub 2013 Jun 14.
Hyperglycemia leads to cytotoxicity in the heart. Although theories were postulated for glucose toxicity-induced cardiomyocyte dysfunction including oxidative stress, the mechanism involved still remains unclear. Recent evidence has depicted a role of protein kinase C (PKC) in diabetic complications while high concentrations of glucose stimulate PKC. This study examined the role of PKCβII in glucose toxicity-induced cardiomyocyte contractile and intracellular Ca(2+) aberrations.
Adult rat cardiomyocytes were maintained in normal (NG, 5.5 mM) or high glucose (HG, 25.5 mM) medium for 12 h. Contractile and intracellular Ca(2+) properties were measured using a video edge-detection system including peak shortening (PS), maximal velocity of shortening/relengthening (±dL/dt), time-to-PS (TPS), time-to-90% relengthening (TR90), rise in intracellular Ca(2+) Fura-2 fluorescence intensity and intracellular Ca(2+) decay. Production of ROS/O2(-) and mitochondrial integrity were examined using fluorescence imaging, aconitase activity and Western blotting.
High glucose triggered abnormal contractile and intracellular Ca(2+) properties including reduced PS, ±dL/dt, prolonged TR90, decreased electrically-stimulated rise in intracellular Ca(2+) and delayed intracellular Ca(2+) clearance, the effects of which were ablated by the PKCβII inhibitor LY333531. Inhibition of PKCβII rescued glucose toxicity-induced generation of ROS and O2(-), apoptosis, cell death and mitochondrial injury (reduced aconitase activity, UCP-2 and PGC-1α). In vitro studies revealed that PKCβII inhibition-induced beneficial effects were mimicked by the NADPH oxidase inhibitor apocynin and were canceled off by mitochondrial uncoupling using FCCP.
These findings suggest the therapeutic potential of specific inhibition of PKCβII isoform in the management of hyperglycemia-induced cardiac complications.
高血糖可导致心脏细胞毒性。虽然已有理论提出葡萄糖毒性诱导的心肌细胞功能障碍的机制包括氧化应激，但具体机制仍不清楚。最近的证据表明蛋白激酶 C（PKC）在糖尿病并发症中发挥作用，而高浓度葡萄糖可刺激 PKC。本研究旨在探讨 PKCβII 在葡萄糖毒性诱导的心肌细胞收缩和细胞内 Ca(2+)异常中的作用。
成年大鼠心肌细胞在正常葡萄糖（NG，5.5 mM）或高葡萄糖（HG，25.5 mM）培养基中培养 12 小时。使用视频边缘检测系统测量心肌细胞的收缩和细胞内 Ca(2+)特性，包括峰缩短（PS）、最大缩短/复长速度（±dL/dt）、PS 时间（TPS）、90%复长时间（TR90）、细胞内 Ca(2+)荧光强度上升和细胞内 Ca(2+)衰减。通过荧光成像、乌头酸酶活性和 Western blot 检测 ROS/O2(-)的产生和线粒体完整性。
高葡萄糖可触发异常的收缩和细胞内 Ca(2+)特性，包括 PS、±dL/dt 降低，TR90 延长，电刺激引起的细胞内 Ca(2+)上升减少，细胞内 Ca(2+)清除延迟，这些作用可被 PKCβII 抑制剂 LY333531 消除。PKCβII 抑制可挽救葡萄糖毒性引起的 ROS 和 O2(-)产生、细胞凋亡、细胞死亡和线粒体损伤（乌头酸酶活性、UCP-2 和 PGC-1α 降低）。体外研究表明，PKCβII 抑制诱导的有益作用可被 NADPH 氧化酶抑制剂 apocynin 模拟，并用 FCCP 阻断线粒体解偶联而消除。
这些发现提示，特异性抑制 PKCβII 同工型可能具有治疗高血糖诱导的心脏并发症的潜力。
