Huang Bi-jun, Cheng Xiao-shu
Department of Cardiovasology, Second Affiliated Hospital of Nanchang University, Jiangxi Provincial Key Laboratory of Molecular Medicine, Nanchang 330006, China.
Department of Cardiovasology, Second Affiliated Hospital of Nanchang University, Jiangxi Provincial Key Laboratory of Molecular Medicine, Nanchang 330006, China. Email:
Zhonghua Xin Xue Guan Bing Za Zhi. 2013 Sep;41(9):785-9.
OBJECTIVE: To investigate the expression changes and effects of hypoxia inducible factor-1α (HIF-1α) on non-lethal high temperature induced thermotolerance and its role on thermotolerance protection. METHODS: H9c2 cardiomyocytes were cultured and pretreated with the HIF-1α inhibitor YC-1, the cells were then subjected to normal temperature (37 °C), thermotolerance induction (40 °C, 3 h), or hyperthermia (43 °C, 2 h). The cells were divided into 8 groups (n = 3 each): normal temperature control group; thermotolerance group; thermotolerance/hyperthermia group; hyperthermia group; DMSO+normal temperature group; YC-1+thermotolerance group; YC-1+thermotolerance/hyperthermia group; YC-1+hyperthermia group. Cell apoptotic rate was assessed by flow cytometry. Western blot was used to detect the expression of HIF-1α and caspase-3. RESULTS: Flow cytometry results showed that apoptosis rate was similar between control group and thermotolerance group, between DMSO+normal temperature group and YC-1+thermotolerance group, between YC-1+thermotolerance/hyperthermia group and YC-1+hyperthermia group, but was significantly higher in hyperthermia group [(17.35 ± 1.07)%] than in control group [(7.52 ± 1.55)%, P < 0.01] which was partly reduced in thermotolerance/hyperthermia group [(12.58 ± 1.97)%, P < 0.01 vs. thermotolerance group]. Cell apoptosis rate of YC-1+thermotolerance/hyperthermia group (23.75 ± 1.92)% was significantly higher than that of thermotolerance/hyperthermia group [(12.58 ± 1.97)%, P < 0.01], and in YC-1+hyperthermia group [(24.89 ± 1.83)%] than in hyperthermia group [(17.35 ± 1.07)%, P < 0.01]. HIF-1α expression was obviously upregulated in thermotolerance cells compared with control cells, in thermotolerance/hyperthermia cells than in hyperthermia cells, in YC-1+thermotolerance group, YC-1+thermotolerance/hyperthermia group and YC-1+hyperthermia group than in DMSO group (all P < 0.05). Caspase-3 expression was similar between control group and thermotolerance group, but was significantly lower in thermotolerance/hyperthermia group than in hyperthermia group (P < 0.05), significantly higher in YC-1+thermotolerance group, YC-1+thermotolerance/hyperthermia group and YC-1+hyperthermia group than in DMSO group (all P < 0.05) and significantly higher in YC-1+thermotolerance/hyperthermia group than in thermotolerance/hyperthermia group (P < 0.01) and in YC-1+hyperthermia group than in hyperthermia group (P < 0.01). CONCLUSION: Non-lethal high temperature induced thermotolerance can reduce heat stress-induced cardiomyocytes apoptosis rate via upregulating the expression of HIF-1α and inhibiting caspase-3 signalling pathways.
目的：探讨缺氧诱导因子-1α（HIF-1α）的表达变化及其对非致死性高温诱导的热耐受性的影响及其在热耐受性保护中的作用。 方法：培养H9c2心肌细胞，并用HIF-1α抑制剂YC-1预处理，然后将细胞置于正常温度（37℃）、热耐受性诱导（40℃，3小时）或热疗（43℃，2小时）条件下。细胞分为8组（每组n = 3）：正常温度对照组；热耐受性组；热耐受性/热疗组；热疗组；二甲基亚砜+正常温度组；YC-1+热耐受性组；YC-1+热耐受性/热疗组；YC-1+热疗组。采用流式细胞术评估细胞凋亡率。采用蛋白质免疫印迹法检测HIF-1α和半胱天冬酶-3的表达。 结果：流式细胞术结果显示，对照组与热耐受性组、二甲基亚砜+正常温度组与YC-1+热耐受性组、YC-1+热耐受性/热疗组与YC-1+热疗组之间的凋亡率相似，但热疗组[（17.35±1.07）%]的凋亡率明显高于对照组[（7.52±1.55）%，P<0.01]，热耐受性/热疗组[（12.58±1.97）%，与热耐受性组相比P<0.01]的凋亡率有所降低。YC-1+热耐受性/热疗组的细胞凋亡率（23.75±1.92）%明显高于热耐受性/热疗组[（12.58±1.97）%，P<0.01]，YC-1+热疗组[（24.89±1.83）%]的凋亡率高于热疗组[（17.35±1.07）%，P<0.01]。与对照细胞相比，热耐受性细胞中HIF-1α表达明显上调，热耐受性/热疗细胞中HIF-1α表达高于热疗细胞，YC-1+热耐受性组、YC-1+热耐受性/热疗组和YC-1+热疗组中的HIF-1α表达高于二甲基亚砜组（均P<0.05）。对照组与热耐受性组之间半胱天冬酶-3表达相似，但热耐受性/热疗组中的半胱天冬酶-3表达明显低于热疗组（P<0.05），YC-1+热耐受性组、YC-1+热耐受性/热疗组和YC-1+热疗组中的半胱天冬酶-3表达明显高于二甲基亚砜组（均P<0.05），YC-1+热耐受性/热疗组中的半胱天冬酶-3表达明显高于热耐受性/热疗组（P<0.01），YC-/+热疗组中的半胱天冬酶-3表达高于热疗组（P<0.01）。 结论：非致死性高温诱导的热耐受性可通过上调HIF-1α表达并抑制半胱天冬酶-3信号通路来降低热应激诱导的心肌细胞凋亡率。
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