高糖及氟伐他汀对HK-2细胞增殖、细胞凋亡及klotho蛋白的影响
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摘要
随着社会经济发展和人们生活方式改变,人类疾病谱正在发生变化。2007年全球糖尿病患者已达246,000,000,到2025年WHO预计全球糖尿病患者将达380,000,000。在美国、欧洲等主要的发达国家,糖尿病尤其是2型糖尿病已成为终末期肾脏病(ESRD)最常见的病因;在中国、东南亚等发展中国家,糖尿病导致ESRD的比率也逐渐上升,给家庭和社会带来巨大的经济负担和医疗压力。
据全美健康与营养调查显示近10%成人服用他汀类药物,超过1/4老人正服用他汀类药物。他汀类药物即羟甲基戊二酰辅酶A(hydroxy -methyl-glutaryl coenzyme A, HMG-CoA)还原酶抑制剂,它通过竞争性抑制HMG-CoA转为甲羟戊酸(mevalonate,MVA),抑制胆固醇生物合成,发挥降低低密度脂蛋白胆固醇(low -density lipoprotein cholesterol, LDL-C)、降低甘油三酯、升高高密度脂蛋白胆固醇(high-density lipoprotein cholesterol, HDL-C)的调脂作用。既往认为他汀类药物的益处来源于其降低血清总胆固醇及LDL-C,近来研究认为他汀类药物具有多样性作用,此作用独立于其降脂作用而存在。其原因目前认为与MVA通路的类戊二烯化合物等有关。他汀抑制MVA合成的同时,也抑制MVA通路中辅酶Q、鲨烯、焦磷酸法尼酯(farnesyl pyrophosphate, FPP)、焦磷酸牛龙牛儿基牛龙牛儿酯(granylgeranyl pyrophosphate, GGPP)等合成;其中FPP、GGPP可经过翻译后修饰参与细胞分化、增殖,调节多种信号转导过程,从而发挥他汀的多样性作用。
目的:本研究主要通过体外实验探讨氟伐他汀(Flv)及高糖(HG)对人肾小管上皮细胞(HK-2)细胞增殖、细胞凋亡及klotho蛋白的影响,并探讨其作用机制。
方法:正常糖(5.5mmol/L, NG)、甘露醇、HG(25.0mmol/L)伴/不伴不同浓度的Flv(0.01μmol/L~1mmol/L)培养HK-2,伴/不伴MVA、GGPP、FPP,干预不同时间(0h、12h、24h、48h、72h),采用苯唑氮蓝法(MTT)检测HG及Flv对细胞增殖的影响,Hoechst33258试剂盒检测细胞凋亡形态学变化,western blot检测半胱氨酸天冬氨酸特异性蛋白酶-3(caspase-3)、半胱氨酸天冬氨酸特异性蛋白酶-8(caspase-8)、半胱氨酸天冬氨酸特异性蛋白酶-9(caspase-9)、klotho、RhoA蛋白表达。
结果:HG培养HK-2 24~48h促进细胞增殖P<0.05,HG培养72h细胞增殖下降P<0.05。NG环境下(0.01μmol/L~1mmol/L)Flv抑制HK-2细胞增殖。HG环境下(0.1μmol/L ~10μmol/L)Flv抑制HK-2细胞增殖,细胞增殖抑制率具有时间-浓度依赖性。10μmol/L Flv干预72h,细胞增殖抑制率>50%。
HG诱导HK-2细胞凋亡小体增加,与0h相比P<0.05;HG环境下10μmol/L Flv干预后,HK-2细胞凋亡率也增加,两者均具有时间依赖性。HG环境下,不同浓度Flv干预72h,荧光显微镜下见0.1、1μmol/L Flv细胞凋亡减少P<0.05; 10μmol/L Flv诱导HK-2细胞凋亡变化不明显,与HG组相比P>0.05。western blot结果显示HG、Flv分别培养HK-2,caspase-3、caspase-8、caspase-9均活化增加,HG环境下10μmol/L Flv活化caspase-3、caspase-8、caspase-9表达。HG环境下不同浓度Flv比较,0.1、1μmol/L Flv干预后caspase-3、caspase-9、caspase-8均表达减少;10μmol/L Flv干预后,于HG相比caspase-3、caspase-9表达减少,caspase-8无显著差别。
NG培养下Flv上调klotho蛋白。HG下调HK-2 klotho蛋白表达,HG环境Flv干预后,HK-2 klotho蛋白表达部分恢复,与HG比较P<0.05;此作用可被MVA、GGPP逆转,FPP不能逆转。同时HK-2 RhoA蛋白表达在高糖环境下增加P<0.05,Flv干预后RhoA蛋白表达下降,与未干预组比较P<0.05。
结论:HG对HK-2细胞增殖出现双重效应,HG干预24~48h促进细胞增殖;HG干预72h细胞增殖下降。NG培养下,0.01μmol/L~1mmol/L Flv抑制HK-2细胞增殖。HG环境下0.1~10μmol/L Flv抑制HK-2细胞增殖,且具有时间、浓度依赖性。10μmol/L Flv干预HK-2 72h,细胞增殖抑制率>50%。HG诱导HK-2细胞凋亡;Flv诱导NG培养HK-2细胞凋亡,两者均具有时间依赖性。10μmol/L Flv与HG对HK-2细胞凋亡无明显协同作用;HG环境下低浓度Flv(0.1、1μmol/L)可改善HG诱导的HK-2细胞凋亡。HG抑制HK-2 klotho蛋白表达,NG、HG环境下,Flv均上调klotho表达。Flv通过MVA通路、影响GGPP异戊烯化,抑制RhoA/Rho激酶信号,上调klotho蛋白表达。低浓度Flv通过上调klotho蛋白表达,下调caspase-9、caspase-8,下调caspase-3蛋白表达,改善HG诱导的HK-2细胞凋亡。
The 21st century has the most diabetogenic environment in human history. In 2007, there were 246 million people with diabetes in the world, but by 2025, that number is estimated to reach 380 million. The increase in prevalence of diabetes will be greater in the developing countries, and the problems of diabetes are now seen as a major global public health concern. Therefore,diabetes is now the major cause of end-stage kidney failure throughout the world in both developed and emerging nations,and it is the primary diagnosis causing kidney disease in 20~40% of people starting treatment for end stage renal disease worldwide.
Data from the National Health and Nutrition Examination Surveys showed that approximately 10% of all adults and nearly one-quarter of adults over 60 reporting statin use. Statins modify circulating lipid levels by inhibiting the conversion of HMG-CoA to mevalonate. Thus,statins have a well-established role in reducing levels of total cholesterol,low-density lipoprotein cholesterol,triglycerides and increasing high-density lipoprotein cholesterol. The clinical benefits of statins were generally assumed to result from their ability to lower serum total- and LDL -cholesterol levels; however, recent observations suggest that some of the clinical benefits associated with statin therapy may be independent of their cholesterol -inhibiting action. Statins not only inhibit cholesterol synthesis, but also affect production of many other compounds with mevalonic acid pathway, such as ubiquinone, dolochol, farnesyl pyrophosphate (FPP) and geranylgeranyl pyropho -sphate (GGPP). The last two compounds are involved in a number of cellular processes including cell signalling, differentiation and proliferation, and that might be the reason of protective effects against kidney disease.
Objective: The purpose of the study was to investigate the effects of high glucose and fluvastatin on cell proliferation, apoptosis and klotho protein of human renal tubular epithelial cell(HK-2), and explored the mechanism.
Methods: HK-2 cells were incubated with normal glucose (5.5mmol/L), mannitol, high glucose (25.0mmol/L), fluvastatin in different concentrations(0.01μmol/L~1mmol/L)for different time(0h、12h、24h、48h、72h)with or without MVA, GGPP or FPP. The proliferation of HK-2 was determined by MTT colorimetry. The apoptotic morphology of HK-2 was detected by Hoechst 33258 staining. The protein expression of caspase-3, caspase-8, caspase-9, klotho and RhoA were detected by western blot.
Results: High glucose induced HK-2 cell proliferation at an early phase of (24~48h), decreased HK-2 cell proliferation later at 72h. Incubation with normal glucose, fluvastatin in different concentrations(0.01μmol/L~1mmol/L)decreased HK-2 cell proliferation. The inhibitory of(0.1μmol/L~10μmol/L)fluvastatin on HK-2 was prominent in high glucose status. The percentage of antiproliferation of fluvastatin in the high glucose status was in a time-dose dependent effect, especially fluvastatin at the concentration of 10μmol/L for 72h.
In the normal glucose status, 10μmol/L fluvastatin induced HK-2 apoptosis in a time-dependent manner. High glucose induced apoptosis; fluvastatin in low concentrations (0.1, 1μmol/L) decreased apoptosis in the high glucose status, while apoptotic ratio of 10μmol/L fluvastatin was the similar to HG. HG stimulated the activities of caspase-3, caspase-8 and caspase-9 in a time-dependent manner. In the high and normal glucose status, fluvastatin induced the activities of caspase-3, caspase-8 and caspase-9 in a time-dependent manner. (0.1, 1μmol/L) fluvastatin inhibited the activities of caspase-3, caspase-8 and caspase-9 in the high glucose status, while 10μmol/L fluvastatin reduced caspase-3, caspase-9 protein at 48h in the high glucose status, caspase-8 protein expression was the similar to high glucose without fluvastain.
High glucose decreased klotho protein expression P<0.05, fluvastatin induced significantly over-expression of klotho protein P<0.05. The effect of fluvastatin on klotho protein was abrogated by MVA and GGPP, but not by FPP, meanwhile fluvastatin inhibited high glucose-induced RhoA expression P<0.05.
Conclusion: High glucose induced a dual effect involving an early phase of increasing cell proliferation (24~48h), and a late phase of decreasing cell prolifera -tion (72h). Incubation with normal and high glucose, fluvastatin inhibited HK-2 cell proliferation. High glucose induced HK-2 apoptosis, and 10μmol/L fluvastatin induced HK-2 apoptosis in a time-dependant manner both in the normal and high glucose status. (0.1, 1μmol/L) fluvastatin decreased high glucose-induced apoptosis, while HG induced proapoptotic protein (caspase-3, caspase-8 and caspase-9) above and beyond 10μmol/L fluvastatin. HG reduced anti-apoptotic klotho protein expression, Fluvastatin inhibited RhoA/Rho kinase pathway activated by high glucose, resulting in up-regulation of klotho expression in HK-2. It was concluded that low-level fluvastatin could inhibit HK-2 apoptosis induced by high glucose.
据全美健康与营养调查显示近10%成人服用他汀类药物,超过1/4老人正服用他汀类药物。他汀类药物即羟甲基戊二酰辅酶A(hydroxy -methyl-glutaryl coenzyme A, HMG-CoA)还原酶抑制剂,它通过竞争性抑制HMG-CoA转为甲羟戊酸(mevalonate,MVA),抑制胆固醇生物合成,发挥降低低密度脂蛋白胆固醇(low -density lipoprotein cholesterol, LDL-C)、降低甘油三酯、升高高密度脂蛋白胆固醇(high-density lipoprotein cholesterol, HDL-C)的调脂作用。既往认为他汀类药物的益处来源于其降低血清总胆固醇及LDL-C,近来研究认为他汀类药物具有多样性作用,此作用独立于其降脂作用而存在。其原因目前认为与MVA通路的类戊二烯化合物等有关。他汀抑制MVA合成的同时,也抑制MVA通路中辅酶Q、鲨烯、焦磷酸法尼酯(farnesyl pyrophosphate, FPP)、焦磷酸牛龙牛儿基牛龙牛儿酯(granylgeranyl pyrophosphate, GGPP)等合成;其中FPP、GGPP可经过翻译后修饰参与细胞分化、增殖,调节多种信号转导过程,从而发挥他汀的多样性作用。
目的:本研究主要通过体外实验探讨氟伐他汀(Flv)及高糖(HG)对人肾小管上皮细胞(HK-2)细胞增殖、细胞凋亡及klotho蛋白的影响,并探讨其作用机制。
方法:正常糖(5.5mmol/L, NG)、甘露醇、HG(25.0mmol/L)伴/不伴不同浓度的Flv(0.01μmol/L~1mmol/L)培养HK-2,伴/不伴MVA、GGPP、FPP,干预不同时间(0h、12h、24h、48h、72h),采用苯唑氮蓝法(MTT)检测HG及Flv对细胞增殖的影响,Hoechst33258试剂盒检测细胞凋亡形态学变化,western blot检测半胱氨酸天冬氨酸特异性蛋白酶-3(caspase-3)、半胱氨酸天冬氨酸特异性蛋白酶-8(caspase-8)、半胱氨酸天冬氨酸特异性蛋白酶-9(caspase-9)、klotho、RhoA蛋白表达。
结果:HG培养HK-2 24~48h促进细胞增殖P<0.05,HG培养72h细胞增殖下降P<0.05。NG环境下(0.01μmol/L~1mmol/L)Flv抑制HK-2细胞增殖。HG环境下(0.1μmol/L ~10μmol/L)Flv抑制HK-2细胞增殖,细胞增殖抑制率具有时间-浓度依赖性。10μmol/L Flv干预72h,细胞增殖抑制率>50%。
HG诱导HK-2细胞凋亡小体增加,与0h相比P<0.05;HG环境下10μmol/L Flv干预后,HK-2细胞凋亡率也增加,两者均具有时间依赖性。HG环境下,不同浓度Flv干预72h,荧光显微镜下见0.1、1μmol/L Flv细胞凋亡减少P<0.05; 10μmol/L Flv诱导HK-2细胞凋亡变化不明显,与HG组相比P>0.05。western blot结果显示HG、Flv分别培养HK-2,caspase-3、caspase-8、caspase-9均活化增加,HG环境下10μmol/L Flv活化caspase-3、caspase-8、caspase-9表达。HG环境下不同浓度Flv比较,0.1、1μmol/L Flv干预后caspase-3、caspase-9、caspase-8均表达减少;10μmol/L Flv干预后,于HG相比caspase-3、caspase-9表达减少,caspase-8无显著差别。
NG培养下Flv上调klotho蛋白。HG下调HK-2 klotho蛋白表达,HG环境Flv干预后,HK-2 klotho蛋白表达部分恢复,与HG比较P<0.05;此作用可被MVA、GGPP逆转,FPP不能逆转。同时HK-2 RhoA蛋白表达在高糖环境下增加P<0.05,Flv干预后RhoA蛋白表达下降,与未干预组比较P<0.05。
结论:HG对HK-2细胞增殖出现双重效应,HG干预24~48h促进细胞增殖;HG干预72h细胞增殖下降。NG培养下,0.01μmol/L~1mmol/L Flv抑制HK-2细胞增殖。HG环境下0.1~10μmol/L Flv抑制HK-2细胞增殖,且具有时间、浓度依赖性。10μmol/L Flv干预HK-2 72h,细胞增殖抑制率>50%。HG诱导HK-2细胞凋亡;Flv诱导NG培养HK-2细胞凋亡,两者均具有时间依赖性。10μmol/L Flv与HG对HK-2细胞凋亡无明显协同作用;HG环境下低浓度Flv(0.1、1μmol/L)可改善HG诱导的HK-2细胞凋亡。HG抑制HK-2 klotho蛋白表达,NG、HG环境下,Flv均上调klotho表达。Flv通过MVA通路、影响GGPP异戊烯化,抑制RhoA/Rho激酶信号,上调klotho蛋白表达。低浓度Flv通过上调klotho蛋白表达,下调caspase-9、caspase-8,下调caspase-3蛋白表达,改善HG诱导的HK-2细胞凋亡。
The 21st century has the most diabetogenic environment in human history. In 2007, there were 246 million people with diabetes in the world, but by 2025, that number is estimated to reach 380 million. The increase in prevalence of diabetes will be greater in the developing countries, and the problems of diabetes are now seen as a major global public health concern. Therefore,diabetes is now the major cause of end-stage kidney failure throughout the world in both developed and emerging nations,and it is the primary diagnosis causing kidney disease in 20~40% of people starting treatment for end stage renal disease worldwide.
Data from the National Health and Nutrition Examination Surveys showed that approximately 10% of all adults and nearly one-quarter of adults over 60 reporting statin use. Statins modify circulating lipid levels by inhibiting the conversion of HMG-CoA to mevalonate. Thus,statins have a well-established role in reducing levels of total cholesterol,low-density lipoprotein cholesterol,triglycerides and increasing high-density lipoprotein cholesterol. The clinical benefits of statins were generally assumed to result from their ability to lower serum total- and LDL -cholesterol levels; however, recent observations suggest that some of the clinical benefits associated with statin therapy may be independent of their cholesterol -inhibiting action. Statins not only inhibit cholesterol synthesis, but also affect production of many other compounds with mevalonic acid pathway, such as ubiquinone, dolochol, farnesyl pyrophosphate (FPP) and geranylgeranyl pyropho -sphate (GGPP). The last two compounds are involved in a number of cellular processes including cell signalling, differentiation and proliferation, and that might be the reason of protective effects against kidney disease.
Objective: The purpose of the study was to investigate the effects of high glucose and fluvastatin on cell proliferation, apoptosis and klotho protein of human renal tubular epithelial cell(HK-2), and explored the mechanism.
Methods: HK-2 cells were incubated with normal glucose (5.5mmol/L), mannitol, high glucose (25.0mmol/L), fluvastatin in different concentrations(0.01μmol/L~1mmol/L)for different time(0h、12h、24h、48h、72h)with or without MVA, GGPP or FPP. The proliferation of HK-2 was determined by MTT colorimetry. The apoptotic morphology of HK-2 was detected by Hoechst 33258 staining. The protein expression of caspase-3, caspase-8, caspase-9, klotho and RhoA were detected by western blot.
Results: High glucose induced HK-2 cell proliferation at an early phase of (24~48h), decreased HK-2 cell proliferation later at 72h. Incubation with normal glucose, fluvastatin in different concentrations(0.01μmol/L~1mmol/L)decreased HK-2 cell proliferation. The inhibitory of(0.1μmol/L~10μmol/L)fluvastatin on HK-2 was prominent in high glucose status. The percentage of antiproliferation of fluvastatin in the high glucose status was in a time-dose dependent effect, especially fluvastatin at the concentration of 10μmol/L for 72h.
In the normal glucose status, 10μmol/L fluvastatin induced HK-2 apoptosis in a time-dependent manner. High glucose induced apoptosis; fluvastatin in low concentrations (0.1, 1μmol/L) decreased apoptosis in the high glucose status, while apoptotic ratio of 10μmol/L fluvastatin was the similar to HG. HG stimulated the activities of caspase-3, caspase-8 and caspase-9 in a time-dependent manner. In the high and normal glucose status, fluvastatin induced the activities of caspase-3, caspase-8 and caspase-9 in a time-dependent manner. (0.1, 1μmol/L) fluvastatin inhibited the activities of caspase-3, caspase-8 and caspase-9 in the high glucose status, while 10μmol/L fluvastatin reduced caspase-3, caspase-9 protein at 48h in the high glucose status, caspase-8 protein expression was the similar to high glucose without fluvastain.
High glucose decreased klotho protein expression P<0.05, fluvastatin induced significantly over-expression of klotho protein P<0.05. The effect of fluvastatin on klotho protein was abrogated by MVA and GGPP, but not by FPP, meanwhile fluvastatin inhibited high glucose-induced RhoA expression P<0.05.
Conclusion: High glucose induced a dual effect involving an early phase of increasing cell proliferation (24~48h), and a late phase of decreasing cell prolifera -tion (72h). Incubation with normal and high glucose, fluvastatin inhibited HK-2 cell proliferation. High glucose induced HK-2 apoptosis, and 10μmol/L fluvastatin induced HK-2 apoptosis in a time-dependant manner both in the normal and high glucose status. (0.1, 1μmol/L) fluvastatin decreased high glucose-induced apoptosis, while HG induced proapoptotic protein (caspase-3, caspase-8 and caspase-9) above and beyond 10μmol/L fluvastatin. HG reduced anti-apoptotic klotho protein expression, Fluvastatin inhibited RhoA/Rho kinase pathway activated by high glucose, resulting in up-regulation of klotho expression in HK-2. It was concluded that low-level fluvastatin could inhibit HK-2 apoptosis induced by high glucose.
引文
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[20] Konturek PC, Burnat G, Hahn EG. Inhibition of Barret's adenocarcinoma cell growth by simvastatin: involvement of COX-2 and apoptosis-related proteins. J Physiol Pharmacol,2007, 58 Suppl 3:141-148.
[21] Lee SK, Kim YC, Song SB, et al. Stabilization and translocation of p53 to mitochondria is linked to Bax translocation to mitochondria in simvastatin -induced apoptosis. Biochem Biophys Res Commun,2010 , 391(4):1592-1597.
[22] Gauthaman K, Fong CY, Bongso A. Statins, stem cells, and cancer. J Cell Biochem,2009, 106(6):975-983.
[23] Uruno A, Sugawara A, Kudo M, et al. Stimulatory effects of low-dose 3-hydroxy-3-methylglutaryl coenzyme a reductase inhibitor fluvastatin on hepatocyte growth factor-induced angiogenesis: involvement of p38 mitogen -activated protein kinase. Hypertens Res,2008, 31(11):2085-2096.
[24] Berube C, Boucher LM, Ma W, et al. Apoptosis caused by p53-induced protein with death domain (PIDD) depends on the death adapter protein RAIDD. Proc Natl Acad Sci U S A,2005, 102(40):14314-14320.
[25] Hoffman B, Liebermann DA. Apoptotic signaling by c-MYC. Oncogene, 2008, 27(50):6462-6472.
[26] Shankar B, Krishnan S, Malladi V, et al. Outer membrane proteins of wild-type and intimin-deficient enteropathogenic Escherichia coli induce Hep-2 cell death through intrinsic and extrinsic pathways of apoptosis. Int J Med Microbiol, 2009, 299(2):121-132.
[27] Meguid El Nahas A, Bello AK. Chronic kidney disease: the global challenge. Lancet,2005, 365(9456):331-340.
[28] Kurosu H, Yamamoto M, Clark JD, et al. Suppression of aging in mice by the hormone Klotho. Science,2005, 309(5742):1829-1833.
[29] Li SA, Watanabe M, Yamada H, et al. Immunohistochemical localization of Klotho protein in brain, kidney, and reproductive organs of mice. Cell Struct Funct,2004, 29(4):91-99.
[30] Mitani H, Ishizaka N, Aizawa T, et al. In vivo klotho gene transfer ameliorates angiotensin II-induced renal damage. Hypertension,2002, 39(4):838-843.
[31] Wang Y, Sun Z. Current understanding of klotho. Ageing Res Rev,2009, 8 (1):43-51.
[32] Hanayama R, Shimizu H, Nakagami H, et al. Fluvastatin improves osteoporosis in fructose-fed insulin resistant model rats through blockade of the classical mevalonate pathway and antioxidant action. Int J Mol Med , 2009, 23 (5):581-588.
[33] Xu H, Zeng L, Peng H, et al. HMG-CoA reductase inhibitor simvastatin miti -gates VEGF-induced "inside-out" signaling to extracellular matrix by preventing RhoA activation. Am J Physiol Renal Physiol,2006, 291(5):F995-1004.
[34] Narumiya H, Sasaki S, Kuwahara N, et al. HMG-CoA reductase inhibitors up -regulate anti-aging klotho mRNA via RhoA inactivation in IMCD3 cells. Cardiovasc Res,2004, 64(2):331-336.
[35] Nagai T, Yamada K, Kim HC, et al. Cognition impairment in the klotho gene mutant mice and oxidative stress. Nihon Shinkei Seishin Yakurigaku Zasshi, 2003, 23(5):211-217.
[36] Ikushima M, Rakugi H, Ishikawa K, et al. Anti-apoptotic and anti-senescence effects of Klotho on vascular endothelial cells. Biochem Biophys Res Commun, 2006, 339(3):827-832.
[37] Yamamoto M, Clark JD, Pastor JV, et al. Regulation of oxidative stress by theanti-aging hormone klotho. J Biol Chem,2005, 280(45):38029-38034.
[38] Mitobe M, Yoshida T, Sugiura H, et al. Oxidative stress decreases klotho expression in a mouse kidney cell line. Nephron Exp Nephrol,2005, 101 (2):e67-74.
[39] Sugiura H, Yoshida T, Tsuchiya K, et al. Klotho reduces apoptosis in experimental ischaemic acute renal failure. Nephrol Dial Transplant,2005, 20(12):2636-2645.
[1] Koh N, Fujimori T, Nishiguchi S , et al . Severely reduced production of klotho in human chronic renal failure kidney. Biochem Biophys Res Commun, 2001, 280 (4): 1015 - 1020.
[2] Yamada Y, Ando F, Niino N, et al. Association of polymorphisms of the andr -ogen receptor and klotho genes with bone mineral density in Japanese women. J. Mol. Med, 2005, 83(1), 50–57.
[3] Kurosu H, Yamamoto M, Clark JD, et al. Suppression of aging in mice by the hormone Klotho. Science, 2005, 309(5742), 1829–1833.
[4] Duce JA, Podvin S, Hollander W, et al. Gene profile analysis implicates Klotho as an important contributor to aging changes in brain white matter of the rhesus monkey. Glia, 2008, 56(1), 106–117.
[5] Vonend O, Apel T, Amann K, Sellin L, et al. Modulation of gene expression by moxonidine in rats with chronic renal failure. Nephrol Dial Transplant, 2004, 19(9), 2217–2222.
[6] Kuwahara N, Sasaki S, Kobara M, et al. HMG-CoA reductase inhibition impr -oves anti-aging klotho protein expression and arteriosclerosis in ratswith chronicinhibition of nitric oxide synthesis. Int J Cardiol, 2008, 123(2), 84–90.
[7] Haruna Y, Kashihara N, Satoh M, et al. Amelioration of progressive renal injury by genetic manipulation of Klotho gene. Proc. Natl. Acad. Sci. USA, 2007, 104 (7), 2331–2336.
[8] Urakawa I, Yamazaki Y, Shimada T, et al. Klotho converts canonical FGF recap -tor into a specific receptor for FGF23. Nature, 2006, 444(7120), 770–774.
[9] Liu H, Fergusson MM, Castilho RM, et al. Augmented Wnt signaling in a mam -malian model of accelerated aging. Science, 2007, 317(5839), 803–806.
[10] Medici D, Razzaque MS, Deluca S, et al. FGF-23-Klotho signaling stimulates proliferation and prevents vitamin D-induced apoptosis. J Cell Biol, 2008, 182(3), 459–465.
[11] Hesse M, Frohlich LF, Zeitz U, et al. Ablation of vitamin D signaling rescues bone, mineral, and glucose homeostasis in Fgf-23 deficient mice. Matrix Biol, 2007, 26(2), 75–84.
[12] Kirstetter P, Anderson K, Porse BT, et al. Activation of the canonical Wnt path -way leads to loss of hematopoietic stem cell repopulation and multilineage differ -entiation block. Nat. Immunol.2006, 7(10), 1048–1056.
[13] Kuro-o, M. Klotho as a regulator of oxidative stress and senescence. Biol Chem, 2008, 389(3), 233–241.
[14] De Oliveira RM. Klotho RNAi induces premature senescence of human cells via a p53/p21 dependent pathway. FEBS Lett, 2006, 580(24), 5753–5758.
[15] Rakugi H, Matsukawa N, Ishikawa K, et al. Anti-oxidative effect of Klotho on endothelial cells through cAMP activation. Endocrine, 2007, 31(1), 82–87.
[16] Imai M, Ishikawa K, Matsukawa N, et al. Klotho protein activates the PKC pathway in the kidney and testis and suppresses 25-hydroxyvitamin D3 1alpha -hydroxylase gene expression. Endocrine, 2004, 25(3), 229–234.
[2] Haruna Y, Kashihara N, Satoh M, et al. Amelioration of progressive renal injury by genetic manipulation of Klotho gene. Proc Natl Acad Sci U S A ,2007,104 (7):2331-2336.
[3] Sugiura H, Yoshida T, Mitobe M, et al. Klotho reduces apoptosis in experimental ischaemic acute kidney injury via HSP-70. Nephrol Dial Transplant,2010,25 (1):60-68.
[4] Xing L, Muxun Z. Expression of c-met stimulated by high glucose in human renal tubular epithelial cells and its implication. J Huazhong Univ Sci Technolog Med Sci,2007,27(2):161-163.
[5] Musunuru K, Nasir K, Pandey S, et al. A synergistic relationship of elevated low-density lipoprotein cholesterol levels and systolic blood pressure with coronary artery calcification. Atherosclerosis,2008,200(2):368-373.
[6] Samikkannu T, Thomas JJ, Bhat GJ, et al. Acute effect of high glucose on long -term cell growth: a role for transient glucose increase in proximal tubule cell injury. Am J Physiol Renal Physiol,2006,291(1):F162-175.
[7] Lorz C, Benito-Martin A, Justo P, et al. Modulation of renal tubular cell survival: where is the evidence? Curr Med Chem,2006,13(4):449-454.
[8] Mou S, Wang Q, Shi B, et al. Hepatocyte growth factor ameliorates progression of interstitial injuries in tubular epithelial cells. Scand J Urol Nephrol,2010,44 (2):121-128.
[9] Kim YJ, Kim YA, Yokozawa T. Protection against oxidative stress, inflammation, and apoptosis of high-glucose-exposed proximal tubular epithelial cells byastaxanthin. J Agric Food Chem,2009,57(19):8793-8797.
[10] Rane MJ, Song Y, Jin S, et al. Interplay between Akt and p38 MAPK pathways in the regulation of renal tubular cell apoptosis associated with diabetic nephropathy. Am J Physiol Renal Physiol,2010,298(1):F49-61.
[11] Liu F, Brezniceanu ML, Wei CC, et al. Overexpression of angiotensinogen increases tubular apoptosis in diabetes. J Am Soc Nephrol,2008,19 (2):269-280.
[12] Verzola D, Bertolotto MB, Villaggio B, et al. Oxidative stress mediates apopt -otic changes induced by hyperglycemia in human tubular kidney cells. J Am Soc Nephrol,2004,15 Suppl 1:S85-87.
[13] Brezniceanu ML, Liu F, Wei CC, et al. Attenuation of interstitial fibrosis and tubular apoptosis in db/db transgenic mice overexpressing catalase in renal proximal tubular cells. Diabetes,2008,57(2):451-459.
[14] Allen DA, Harwood S, Varagunam M, et al. High glucose-induced oxidative stress causes apoptosis in proximal tubular epithelial cells and is mediated by multiple caspases. Faseb J,2003,17(8):908-910.
[15] Li YM, Schilling T, Benisch P, et al. Effects of high glucose on mesenchymal stem cell proliferation and differentiation. Biochem Biophys Res Commun, 2007,363(1):209-215.
[16] Minn AH, Hafele C, Shalev A. Thioredoxin-interacting protein is stimulated by glucose through a carbohydrate response element and induces beta-cell apoptosis. Endocrinology,2005,146(5):2397-2405.
[17]Hwang KE, Na KS, Park DS, et al. Apoptotic induction by simvastatin in human lung cancer A549 cells via Akt signaling dependent down-regulation of survivin. Invest New Drugs,2010,Epub ahead of print.
[18] Acquavella N, Quiroga MF, Wittig O, et al. Effect of simvastatin on endothelial cell apoptosis mediated by Fas and TNF-alpha. Cytokine,2010, 49(1):45-50.
[19] Kato S, Smalley S, Sadarangani A, et al. Lipophilic but not hydrophilic statins selectively induce cell death in gynecological cancers expressing high levels of HMGCoA reductase. J Cell Mol Med,2009, Epub ahead of print.
[20] Konturek PC, Burnat G, Hahn EG. Inhibition of Barret's adenocarcinoma cell growth by simvastatin: involvement of COX-2 and apoptosis-related proteins. J Physiol Pharmacol,2007, 58 Suppl 3:141-148.
[21] Lee SK, Kim YC, Song SB, et al. Stabilization and translocation of p53 to mitochondria is linked to Bax translocation to mitochondria in simvastatin -induced apoptosis. Biochem Biophys Res Commun,2010 , 391(4):1592-1597.
[22] Gauthaman K, Fong CY, Bongso A. Statins, stem cells, and cancer. J Cell Biochem,2009, 106(6):975-983.
[23] Uruno A, Sugawara A, Kudo M, et al. Stimulatory effects of low-dose 3-hydroxy-3-methylglutaryl coenzyme a reductase inhibitor fluvastatin on hepatocyte growth factor-induced angiogenesis: involvement of p38 mitogen -activated protein kinase. Hypertens Res,2008, 31(11):2085-2096.
[24] Berube C, Boucher LM, Ma W, et al. Apoptosis caused by p53-induced protein with death domain (PIDD) depends on the death adapter protein RAIDD. Proc Natl Acad Sci U S A,2005, 102(40):14314-14320.
[25] Hoffman B, Liebermann DA. Apoptotic signaling by c-MYC. Oncogene, 2008, 27(50):6462-6472.
[26] Shankar B, Krishnan S, Malladi V, et al. Outer membrane proteins of wild-type and intimin-deficient enteropathogenic Escherichia coli induce Hep-2 cell death through intrinsic and extrinsic pathways of apoptosis. Int J Med Microbiol, 2009, 299(2):121-132.
[27] Meguid El Nahas A, Bello AK. Chronic kidney disease: the global challenge. Lancet,2005, 365(9456):331-340.
[28] Kurosu H, Yamamoto M, Clark JD, et al. Suppression of aging in mice by the hormone Klotho. Science,2005, 309(5742):1829-1833.
[29] Li SA, Watanabe M, Yamada H, et al. Immunohistochemical localization of Klotho protein in brain, kidney, and reproductive organs of mice. Cell Struct Funct,2004, 29(4):91-99.
[30] Mitani H, Ishizaka N, Aizawa T, et al. In vivo klotho gene transfer ameliorates angiotensin II-induced renal damage. Hypertension,2002, 39(4):838-843.
[31] Wang Y, Sun Z. Current understanding of klotho. Ageing Res Rev,2009, 8 (1):43-51.
[32] Hanayama R, Shimizu H, Nakagami H, et al. Fluvastatin improves osteoporosis in fructose-fed insulin resistant model rats through blockade of the classical mevalonate pathway and antioxidant action. Int J Mol Med , 2009, 23 (5):581-588.
[33] Xu H, Zeng L, Peng H, et al. HMG-CoA reductase inhibitor simvastatin miti -gates VEGF-induced "inside-out" signaling to extracellular matrix by preventing RhoA activation. Am J Physiol Renal Physiol,2006, 291(5):F995-1004.
[34] Narumiya H, Sasaki S, Kuwahara N, et al. HMG-CoA reductase inhibitors up -regulate anti-aging klotho mRNA via RhoA inactivation in IMCD3 cells. Cardiovasc Res,2004, 64(2):331-336.
[35] Nagai T, Yamada K, Kim HC, et al. Cognition impairment in the klotho gene mutant mice and oxidative stress. Nihon Shinkei Seishin Yakurigaku Zasshi, 2003, 23(5):211-217.
[36] Ikushima M, Rakugi H, Ishikawa K, et al. Anti-apoptotic and anti-senescence effects of Klotho on vascular endothelial cells. Biochem Biophys Res Commun, 2006, 339(3):827-832.
[37] Yamamoto M, Clark JD, Pastor JV, et al. Regulation of oxidative stress by theanti-aging hormone klotho. J Biol Chem,2005, 280(45):38029-38034.
[38] Mitobe M, Yoshida T, Sugiura H, et al. Oxidative stress decreases klotho expression in a mouse kidney cell line. Nephron Exp Nephrol,2005, 101 (2):e67-74.
[39] Sugiura H, Yoshida T, Tsuchiya K, et al. Klotho reduces apoptosis in experimental ischaemic acute renal failure. Nephrol Dial Transplant,2005, 20(12):2636-2645.
[1] Koh N, Fujimori T, Nishiguchi S , et al . Severely reduced production of klotho in human chronic renal failure kidney. Biochem Biophys Res Commun, 2001, 280 (4): 1015 - 1020.
[2] Yamada Y, Ando F, Niino N, et al. Association of polymorphisms of the andr -ogen receptor and klotho genes with bone mineral density in Japanese women. J. Mol. Med, 2005, 83(1), 50–57.
[3] Kurosu H, Yamamoto M, Clark JD, et al. Suppression of aging in mice by the hormone Klotho. Science, 2005, 309(5742), 1829–1833.
[4] Duce JA, Podvin S, Hollander W, et al. Gene profile analysis implicates Klotho as an important contributor to aging changes in brain white matter of the rhesus monkey. Glia, 2008, 56(1), 106–117.
[5] Vonend O, Apel T, Amann K, Sellin L, et al. Modulation of gene expression by moxonidine in rats with chronic renal failure. Nephrol Dial Transplant, 2004, 19(9), 2217–2222.
[6] Kuwahara N, Sasaki S, Kobara M, et al. HMG-CoA reductase inhibition impr -oves anti-aging klotho protein expression and arteriosclerosis in ratswith chronicinhibition of nitric oxide synthesis. Int J Cardiol, 2008, 123(2), 84–90.
[7] Haruna Y, Kashihara N, Satoh M, et al. Amelioration of progressive renal injury by genetic manipulation of Klotho gene. Proc. Natl. Acad. Sci. USA, 2007, 104 (7), 2331–2336.
[8] Urakawa I, Yamazaki Y, Shimada T, et al. Klotho converts canonical FGF recap -tor into a specific receptor for FGF23. Nature, 2006, 444(7120), 770–774.
[9] Liu H, Fergusson MM, Castilho RM, et al. Augmented Wnt signaling in a mam -malian model of accelerated aging. Science, 2007, 317(5839), 803–806.
[10] Medici D, Razzaque MS, Deluca S, et al. FGF-23-Klotho signaling stimulates proliferation and prevents vitamin D-induced apoptosis. J Cell Biol, 2008, 182(3), 459–465.
[11] Hesse M, Frohlich LF, Zeitz U, et al. Ablation of vitamin D signaling rescues bone, mineral, and glucose homeostasis in Fgf-23 deficient mice. Matrix Biol, 2007, 26(2), 75–84.
[12] Kirstetter P, Anderson K, Porse BT, et al. Activation of the canonical Wnt path -way leads to loss of hematopoietic stem cell repopulation and multilineage differ -entiation block. Nat. Immunol.2006, 7(10), 1048–1056.
[13] Kuro-o, M. Klotho as a regulator of oxidative stress and senescence. Biol Chem, 2008, 389(3), 233–241.
[14] De Oliveira RM. Klotho RNAi induces premature senescence of human cells via a p53/p21 dependent pathway. FEBS Lett, 2006, 580(24), 5753–5758.
[15] Rakugi H, Matsukawa N, Ishikawa K, et al. Anti-oxidative effect of Klotho on endothelial cells through cAMP activation. Endocrine, 2007, 31(1), 82–87.
[16] Imai M, Ishikawa K, Matsukawa N, et al. Klotho protein activates the PKC pathway in the kidney and testis and suppresses 25-hydroxyvitamin D3 1alpha -hydroxylase gene expression. Endocrine, 2004, 25(3), 229–234.