HIF-1介导的线粒体能量代谢参与异丙酚调控肾透明细胞癌细胞的增殖和凋亡

Zhen-guo Li, Yu-ming Zhang, Z. Zhen, Ru Zhang, Jing Zhu, Jun Wang
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HIF-1 protein expression was detected by Western blotting in the two kinds of cells, cell proliferation activity and apoptosis rate were detected by flow cytometry, and mitochondrial energy metabolism was detected by energy metabolism analyzer. \n \n \nResults \nCompared with RCC4-VHL(-) cells, the relative expression of HIF-1α protein in RCC4-VHL(+ ) cells was significantly decreased (0.05±0.02 vs. 1.23±0.10, t=16.016, P<0.001). When propofol concentrations were 50 μmol/L and 100 μmol/L, the proliferation activity of RCC4-VHL(+ ) cells was significantly lower than that of RCC4-VHL(-) cells (50 μmol/L: 0.10±0.02 vs. 0.13±0.04, t=3.502, P=0.032; 100 μmol/L: 0.05±0.02 vs. 0.10±0.01, t=6.771, P=0.017), and the apoptotic rate was significantly higher than that of RCC4-VHL (-) cells [50 μmol/L: (35.50±1.84)% vs. (22.15±1.06)%, t=7.082, P=0.004; 100 μmol/L: (54.35±2.97)% vs. (35.10±3.25)%, t=10.241, P<0.001). Compared with 0 μmol/L propofol, 100 μmol/L propofol increased HIF-1α protein expression in RCC4-VHL (+ ) cells (0.93±0.05 vs. 0.04±0.02, t=18.500, P<0.001). Compared with RCC4-VHL(-) cells, the oxygen consumption rate (OCR) [(130.42±11.81) pmol/min vs. (48.27±7.66) pmol/min, t=11.672, P<0.001], basal aerobic respiration [(98.55±8.09) pmol/min vs. (41.63±6.21) pmol/min, t=11.162, P<0.001], aerobic maximum [(226.79±13.51) pmol/min vs. (70.18±6.82) pmol/min, t=20.697, P<0.001], non-mitochondrial respiration [(28.36±4.29) pmol/min vs. (8.92±1.70) pmol/min, t=8.426, P=0.001] and oxygen consumption rate of proton leak [(23.85±5.08) pmol/min vs. (7.80±1.24) pmol/min, t=6.139, P=0.006] were significantly increased in RCC4-VHL(+ ) cells, while the extracellular acidification rate (ECAR) was significantly decreased [(26.76±4.35) mpH/min vs. (39.48±5.17) mpH/min, t=3.765, P=0.010]. Compared with 0 μmol/L propofol added in RCC4-VHL(+ ) cells, 100 μmol/L propofol decreased OCR [(72.44±8.15) pmol/min vs. (131.56±9.04) pmol/min, t=9.751, P<0.001], basal aerobic respiration [(54.31±5.35) pmol/min vs. (96.49±6.86) pmol/min, t=9.697, P<0.001], aerobic maximum [(116.71±12.39) pmol/min vs. (219.53±11.80) pmol/min, t=12.019, P<0.001], non-mitochondrial respiration [(13.25±4.01) pmol/min vs. (29.04±5.11) pmol/min, t=4.862, P=0.002] and oxygen consumption rate of proton leak [(10.24±3.79) pmol/min vs. (22.92±4.12) pmol/min, t=4.530, P=0.003], and increased ECAR significantly [(37.69±3.75) mpH/min vs. (25.87±4.03) mpH/min, t=4.294, P=0.004]. \n \n \nConclusion \nLoss of VHL up-regulates expression of HIF-1 protein, and mitochondrial energy metabolism mediated via HIF-1 involves the proliferation and apoptosis of RCC4 cells regulated by propofol. \n \n \nKey words: \nHypoxia-inducible factor 1; Energy metabolism; Propofol; Cell proliferation; Renal clear cell carcinoma","PeriodicalId":16120,"journal":{"name":"国际肿瘤学杂志","volume":"6 1","pages":"711-717"},"PeriodicalIF":0.0000,"publicationDate":"2019-12-08","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":"0","resultStr":"{\"title\":\"Mitochondrial energy metabolism mediated via HIF-1 involves the proliferation and apoptosis of renal clear cell carcinoma cells regulated by propofol\",\"authors\":\"Zhen-guo Li, Yu-ming Zhang, Z. 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HIF-1 protein expression was detected by Western blotting in the two kinds of cells, cell proliferation activity and apoptosis rate were detected by flow cytometry, and mitochondrial energy metabolism was detected by energy metabolism analyzer. \\n \\n \\nResults \\nCompared with RCC4-VHL(-) cells, the relative expression of HIF-1α protein in RCC4-VHL(+ ) cells was significantly decreased (0.05±0.02 vs. 1.23±0.10, t=16.016, P<0.001). When propofol concentrations were 50 μmol/L and 100 μmol/L, the proliferation activity of RCC4-VHL(+ ) cells was significantly lower than that of RCC4-VHL(-) cells (50 μmol/L: 0.10±0.02 vs. 0.13±0.04, t=3.502, P=0.032; 100 μmol/L: 0.05±0.02 vs. 0.10±0.01, t=6.771, P=0.017), and the apoptotic rate was significantly higher than that of RCC4-VHL (-) cells [50 μmol/L: (35.50±1.84)% vs. (22.15±1.06)%, t=7.082, P=0.004; 100 μmol/L: (54.35±2.97)% vs. (35.10±3.25)%, t=10.241, P<0.001). Compared with 0 μmol/L propofol, 100 μmol/L propofol increased HIF-1α protein expression in RCC4-VHL (+ ) cells (0.93±0.05 vs. 0.04±0.02, t=18.500, P<0.001). Compared with RCC4-VHL(-) cells, the oxygen consumption rate (OCR) [(130.42±11.81) pmol/min vs. (48.27±7.66) pmol/min, t=11.672, P<0.001], basal aerobic respiration [(98.55±8.09) pmol/min vs. (41.63±6.21) pmol/min, t=11.162, P<0.001], aerobic maximum [(226.79±13.51) pmol/min vs. (70.18±6.82) pmol/min, t=20.697, P<0.001], non-mitochondrial respiration [(28.36±4.29) pmol/min vs. (8.92±1.70) pmol/min, t=8.426, P=0.001] and oxygen consumption rate of proton leak [(23.85±5.08) pmol/min vs. (7.80±1.24) pmol/min, t=6.139, P=0.006] were significantly increased in RCC4-VHL(+ ) cells, while the extracellular acidification rate (ECAR) was significantly decreased [(26.76±4.35) mpH/min vs. (39.48±5.17) mpH/min, t=3.765, P=0.010]. Compared with 0 μmol/L propofol added in RCC4-VHL(+ ) cells, 100 μmol/L propofol decreased OCR [(72.44±8.15) pmol/min vs. (131.56±9.04) pmol/min, t=9.751, P<0.001], basal aerobic respiration [(54.31±5.35) pmol/min vs. (96.49±6.86) pmol/min, t=9.697, P<0.001], aerobic maximum [(116.71±12.39) pmol/min vs. (219.53±11.80) pmol/min, t=12.019, P<0.001], non-mitochondrial respiration [(13.25±4.01) pmol/min vs. (29.04±5.11) pmol/min, t=4.862, P=0.002] and oxygen consumption rate of proton leak [(10.24±3.79) pmol/min vs. (22.92±4.12) pmol/min, t=4.530, P=0.003], and increased ECAR significantly [(37.69±3.75) mpH/min vs. (25.87±4.03) mpH/min, t=4.294, P=0.004]. \\n \\n \\nConclusion \\nLoss of VHL up-regulates expression of HIF-1 protein, and mitochondrial energy metabolism mediated via HIF-1 involves the proliferation and apoptosis of RCC4 cells regulated by propofol. \\n \\n \\nKey words: \\nHypoxia-inducible factor 1; Energy metabolism; Propofol; Cell proliferation; Renal clear cell carcinoma\",\"PeriodicalId\":16120,\"journal\":{\"name\":\"国际肿瘤学杂志\",\"volume\":\"6 1\",\"pages\":\"711-717\"},\"PeriodicalIF\":0.0000,\"publicationDate\":\"2019-12-08\",\"publicationTypes\":\"Journal Article\",\"fieldsOfStudy\":null,\"isOpenAccess\":false,\"openAccessPdf\":\"\",\"citationCount\":\"0\",\"resultStr\":null,\"platform\":\"Semanticscholar\",\"paperid\":null,\"PeriodicalName\":\"国际肿瘤学杂志\",\"FirstCategoryId\":\"3\",\"ListUrlMain\":\"https://doi.org/10.3760/CMA.J.ISSN.1673-422X.2019.12.002\",\"RegionNum\":0,\"RegionCategory\":null,\"ArticlePicture\":[],\"TitleCN\":null,\"AbstractTextCN\":null,\"PMCID\":null,\"EPubDate\":\"\",\"PubModel\":\"\",\"JCR\":\"\",\"JCRName\":\"\",\"Score\":null,\"Total\":0}","platform":"Semanticscholar","paperid":null,"PeriodicalName":"国际肿瘤学杂志","FirstCategoryId":"3","ListUrlMain":"https://doi.org/10.3760/CMA.J.ISSN.1673-422X.2019.12.002","RegionNum":0,"RegionCategory":null,"ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":null,"EPubDate":"","PubModel":"","JCR":"","JCRName":"","Score":null,"Total":0}
引用次数: 0

摘要

目的探讨缺氧诱导因子-1 (HIF-1)介导的线粒体能量代谢在异丙酚调控肾透明细胞癌细胞增殖和凋亡中的作用。方法选择不表达VHL基因的人肾透明细胞癌细胞株RCC4作为研究对象。将pcDNA3-VHL质粒和pcDNA3空质粒分别转染到RCC4细胞中,得到稳定表达外源VHL蛋白的RCC4-VHL(+)细胞和不表达VHL蛋白的RCC4-VHL(-)细胞。然后分别以0、25、50和100 μmol/L剂量的异丙酚处理这两种细胞。Western blotting检测两种细胞中HIF-1蛋白的表达,流式细胞术检测细胞增殖活性和凋亡率,能量代谢分析仪检测线粒体能量代谢。结果与RCC4-VHL(-)细胞相比,RCC4-VHL(+)细胞中HIF-1α蛋白的相对表达量显著降低(0.05±0.02∶1.23±0.10,t=16.016, P<0.001)。异丙酚浓度分别为50 μmol/L和100 μmol/L时,RCC4-VHL(+)细胞的增殖活性显著低于RCC4-VHL(-)细胞(50 μmol/L: 0.10±0.02 vs. 0.13±0.04,t=3.502, P=0.032;100 μmol/L: 0.05±0.02比0.10±0.01,t=6.771, P=0.017),且凋亡率显著高于RCC4-VHL(-)细胞[50 μmol/L:(35.50±1.84)%比(22.15±1.06)%,t=7.082, P=0.004;100μmol / L:(54.35±2.97)%和(35.10±3.25)%,t = 10.241, P < 0.001)。与0 μmol/L异丙酚相比,100 μmol/L异丙酚可提高RCC4-VHL(+)细胞中HIF-1α蛋白的表达(0.93±0.05 vs. 0.04±0.02,t=18.500, P<0.001)。与RCC4-VHL(-)细胞相比,耗氧量(OCR)[(130.42±11.81)pmol/min vs(48.27±7.66)pmol/min, t=11.672, P<0.001],基础有氧呼吸[(98.55±8.09)pmol/min vs(41.63±6.21)pmol/min, t=11.162, P<0.001],有氧最大值[(226.79±13.51)pmol/min vs(70.18±6.82)pmol/min, t=20.697, P<0.001],非线粒体呼吸[(28.36±4.29)pmol/min vs(8.92±1.70)pmol/min, t=8.426,RCC4-VHL(+)细胞质子泄漏耗氧量[(23.85±5.08)pmol/min vs(7.80±1.24)pmol/min, t=6.139, P=0.006]显著升高,细胞外酸化率(ECAR)显著降低[(26.76±4.35)mpH/min vs(39.48±5.17)mpH/min, t=3.765, P=0.010]。与RCC4-VHL(+)细胞中添加0 μmol/L异丙酚相比,100 μmol/L异丙酚降低了OCR[(72.44±8.15)pmol/min vs(131.56±9.04)pmol/min, t=9.751, P<0.001]、基础有氧呼吸[(54.31±5.35)pmol/min vs(96.49±6.86)pmol/min, t=9.697, P<0.001]、有氧最大值[(116.71±12.39)pmol/min vs(219.53±11.80)pmol/min, t=12.019, P<0.001]、非线粒体呼吸[(13.25±4.01)pmol/min vs(29.04±5.11)pmol/min, t=4.862,质子泄漏耗氧量[(10.24±3.79)pmol/min vs(22.92±4.12)pmol/min, t=4.530, P=0.003], ECAR显著升高[(37.69±3.75)mpH/min vs(25.87±4.03)mpH/min, t=4.294, P=0.004]。结论VHL缺失可上调HIF-1蛋白的表达,HIF-1介导的线粒体能量代谢参与异丙酚调控RCC4细胞的增殖和凋亡。关键词:缺氧诱导因子1;能量代谢;异丙酚;细胞增殖;肾透明细胞癌
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Mitochondrial energy metabolism mediated via HIF-1 involves the proliferation and apoptosis of renal clear cell carcinoma cells regulated by propofol
Objective To investigate the role of mitochondrial energy metabolism mediated via hypoxia-inducible factor-1 (HIF-1) in the proliferation and apoptosis of renal clear cell carcinoma cells regulated by propofol. Methods We chose human renal clear cell carcinoma cell line RCC4 as the research object, which did not express VHL gene. The pcDNA3-VHL plasmid and the pcDNA3 empty plasmid were respectively transfected into RCC4 cells to obtain RCC4-VHL(+ ) cells stably expressing the exogenous VHL protein and RCC4-VHL(-) cells without expressing the VHL protein. These two kinds of cells were then exposed to propofol at dosage of 0, 25, 50 and 100 μmol/L. HIF-1 protein expression was detected by Western blotting in the two kinds of cells, cell proliferation activity and apoptosis rate were detected by flow cytometry, and mitochondrial energy metabolism was detected by energy metabolism analyzer. Results Compared with RCC4-VHL(-) cells, the relative expression of HIF-1α protein in RCC4-VHL(+ ) cells was significantly decreased (0.05±0.02 vs. 1.23±0.10, t=16.016, P<0.001). When propofol concentrations were 50 μmol/L and 100 μmol/L, the proliferation activity of RCC4-VHL(+ ) cells was significantly lower than that of RCC4-VHL(-) cells (50 μmol/L: 0.10±0.02 vs. 0.13±0.04, t=3.502, P=0.032; 100 μmol/L: 0.05±0.02 vs. 0.10±0.01, t=6.771, P=0.017), and the apoptotic rate was significantly higher than that of RCC4-VHL (-) cells [50 μmol/L: (35.50±1.84)% vs. (22.15±1.06)%, t=7.082, P=0.004; 100 μmol/L: (54.35±2.97)% vs. (35.10±3.25)%, t=10.241, P<0.001). Compared with 0 μmol/L propofol, 100 μmol/L propofol increased HIF-1α protein expression in RCC4-VHL (+ ) cells (0.93±0.05 vs. 0.04±0.02, t=18.500, P<0.001). Compared with RCC4-VHL(-) cells, the oxygen consumption rate (OCR) [(130.42±11.81) pmol/min vs. (48.27±7.66) pmol/min, t=11.672, P<0.001], basal aerobic respiration [(98.55±8.09) pmol/min vs. (41.63±6.21) pmol/min, t=11.162, P<0.001], aerobic maximum [(226.79±13.51) pmol/min vs. (70.18±6.82) pmol/min, t=20.697, P<0.001], non-mitochondrial respiration [(28.36±4.29) pmol/min vs. (8.92±1.70) pmol/min, t=8.426, P=0.001] and oxygen consumption rate of proton leak [(23.85±5.08) pmol/min vs. (7.80±1.24) pmol/min, t=6.139, P=0.006] were significantly increased in RCC4-VHL(+ ) cells, while the extracellular acidification rate (ECAR) was significantly decreased [(26.76±4.35) mpH/min vs. (39.48±5.17) mpH/min, t=3.765, P=0.010]. Compared with 0 μmol/L propofol added in RCC4-VHL(+ ) cells, 100 μmol/L propofol decreased OCR [(72.44±8.15) pmol/min vs. (131.56±9.04) pmol/min, t=9.751, P<0.001], basal aerobic respiration [(54.31±5.35) pmol/min vs. (96.49±6.86) pmol/min, t=9.697, P<0.001], aerobic maximum [(116.71±12.39) pmol/min vs. (219.53±11.80) pmol/min, t=12.019, P<0.001], non-mitochondrial respiration [(13.25±4.01) pmol/min vs. (29.04±5.11) pmol/min, t=4.862, P=0.002] and oxygen consumption rate of proton leak [(10.24±3.79) pmol/min vs. (22.92±4.12) pmol/min, t=4.530, P=0.003], and increased ECAR significantly [(37.69±3.75) mpH/min vs. (25.87±4.03) mpH/min, t=4.294, P=0.004]. Conclusion Loss of VHL up-regulates expression of HIF-1 protein, and mitochondrial energy metabolism mediated via HIF-1 involves the proliferation and apoptosis of RCC4 cells regulated by propofol. Key words: Hypoxia-inducible factor 1; Energy metabolism; Propofol; Cell proliferation; Renal clear cell carcinoma
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