Department of Anesthesiology, Chi Mei Medical Center, Tainan, Taiwan.
Anesthesiology. 2012 Apr;116(4):868-81. doi: 10.1097/ALN.0b013e31824af68a.
Overdose propofol treatment with a prolong time causes injury to multiple cell types; however, its molecular mechanisms remain unclear. Activation of glycogen synthase kinase (GSK)-3β is proapoptotic under death stimuli. The authors therefore hypothesize that propofol overdose induces macrophage apoptosis through GSK-3β.
Phagocytic analysis by uptake of Staphylococcus aureus showed the effects of propofol overdose on murine macrophages RAW264.7 and BV2 and primary human neutrophils in vitro. The authors further investigated cell apoptosis in vitro and in vivo, lysosomal membrane permeabilization, and the loss of mitochondrial transmembrane potential (MTP) by propidium iodide, annexin V, acridine orange, and rhodamine 123 staining, respectively. Protein analysis identified activation of apoptotic signals, and pharmacologic inhibition and genetic knockdown using lentiviral-based short hairpin RNA were further used to clarify their roles.
A high dose of propofol caused phagocytic inhibition and apoptosis in vitro for 24 h (25 μg/ml, in triplicate) and in vivo for 6 h (10 mg/kg/h, n = 5 for each group). Propofol induced lysosomal membrane permeabilization and MTP loss while stabilizing MTP and inhibiting caspase protected cells from mitochondrial apoptosis. Lysosomal cathepsin B was required for propofol-induced lysosomal membrane permeabilization, MTP loss, and apoptosis. Propofol decreased antiapoptotic Bcl-2 family proteins and then caused proapoptotic Bcl-2-associated X protein (Bax) activation. Propofol-activated GSK-3β and inhibiting GSK-3β prevented Mcl-1 destabilization, MTP loss, and lysosomal/mitochondrial apoptosis. Forced expression of Mcl-1 prevented the apoptotic effects of propofol. Decreased Akt was important for GSK-3β activation caused by propofol.
These results suggest an essential role of GSK-3β in propofol-induced lysosomal/mitochondrial apoptosis.
长时间大剂量的丙泊酚处理会导致多种细胞类型受损；然而，其分子机制尚不清楚。在死亡刺激下，糖原合酶激酶（GSK）-3β的激活具有促凋亡作用。因此，作者假设丙泊酚过量会通过 GSK-3β诱导巨噬细胞凋亡。
通过金黄色葡萄球菌摄取来进行吞噬分析，研究丙泊酚过量对体外鼠源巨噬细胞 RAW264.7 和 BV2 以及原代人中性粒细胞的影响。作者进一步研究了细胞凋亡，包括体外和体内，溶酶体膜通透性，以及用碘化丙啶、膜联蛋白 V、吖啶橙和罗丹明 123 染色分别检测线粒体跨膜电位（MTP）的丧失。蛋白质分析确定了凋亡信号的激活，并用基于慢病毒的短发夹 RNA 进行药理学抑制和基因敲低，以进一步阐明其作用。
高剂量丙泊酚在体外 24 小时（25μg/ml，重复 3 次）和体内 6 小时（10mg/kg/h，每组 5 只）引起吞噬抑制和细胞凋亡。丙泊酚诱导溶酶体膜通透性和 MTP 丧失，而稳定 MTP 和抑制半胱天冬酶可保护细胞免受线粒体凋亡。溶酶体组织蛋白酶 B 是丙泊酚诱导的溶酶体膜通透性、MTP 丧失和凋亡所必需的。丙泊酚降低了抗凋亡 Bcl-2 家族蛋白，然后导致促凋亡 Bcl-2 相关 X 蛋白（Bax）的激活。丙泊酚激活 GSK-3β，抑制 GSK-3β，可防止 Mcl-1 不稳定、MTP 丧失和溶酶体/线粒体凋亡。强制表达 Mcl-1 可防止丙泊酚的凋亡作用。丙泊酚引起的 GSK-3β 激活需要 Akt 的减少。
这些结果表明 GSK-3β 在丙泊酚诱导的溶酶体/线粒体凋亡中起重要作用。
