AIM:To investigate the effect of MEK/ERK1/2 pathway on peroxisome proliferator-activated receptors(PPARγ)agonist-induced alterations in Δ6-desaturase(Δ6D)and stearoyl-CoA desaturase 1(SCD1)in hepatocellular carcino...AIM:To investigate the effect of MEK/ERK1/2 pathway on peroxisome proliferator-activated receptors(PPARγ)agonist-induced alterations in Δ6-desaturase(Δ6D)and stearoyl-CoA desaturase 1(SCD1)in hepatocellular carcinoma cell line HepG2.METHODS:HepG2 cells cultured in RPMI-1640 were exposed to the commonly used ERK1/2 pathway inhibitor PD98059 and PPARγ agonist,pioglitazone.Total RNA was isolated and reverse transcribed from treated cells.Changes in gene expression and metabolites ratio,as activity index for Δ6D and SCD1,were then determined using reverse transcriptionpolymerase chain reaction and gas liquid chromatography,respectively.RESULTS:The expression of both Δ6D(P = 0.03)and SCD1(P = 0.01)increased following PD98059 treatment,with a higher impact on SCD1(24.5%vs 62.5%).Although pioglitazone increased the mRNA level(1.47 ± 0.10 vs 0.88 ± 0.02,P = 0.006)and activity index(1.40 ± 0.07 vs 0.79 ± 0.11,P < 0.001)of Δ6D,no such changes have been observed for SCD1 activity index in pioglitazone-treated cells.SCD1 gene expression(+26.4%,P = 0.041)and activity index(+52.8%,P = 0.035)were significantly increased by MEK inhibition in the presence of pioglitazone,as compared with pioglitazone alone and control cells.However,the response of Δ6D expression and activity index to pioglitazone was unaffected by incubation with PD98059.CONCLUSION:PPARγ and ERK1/2 signaling pathway affect differentially and may have inhibitory crosstalk effects on the genes expression of 6D and SCD1,and subsequently on their enzymatic activities.展开更多
目的:探讨抑制素β亚基A反义RNA1(INHBA-AS1)对宫颈癌HeLa细胞EMT和鸟氨酸代谢途径的影响及其机制。方法:体外常规培养HeLa细胞,实验分为10组:对照组、阴性对照(NC)组、sh-INHBA-AS1组、PluriSIn 1[硬脂酰辅酶A去饱和酶(stearyl CoA des...目的:探讨抑制素β亚基A反义RNA1(INHBA-AS1)对宫颈癌HeLa细胞EMT和鸟氨酸代谢途径的影响及其机制。方法:体外常规培养HeLa细胞,实验分为10组:对照组、阴性对照(NC)组、sh-INHBA-AS1组、PluriSIn 1[硬脂酰辅酶A去饱和酶(stearyl CoA desaturase,SCD)抑制剂]组、NC+PluriSIn 1组、sh-INHBA-AS1+PluriSIn 1组、10058-F4(c-Myc抑制剂)组、NC+10058-F4组、sh-INHBA-AS1+10058-F4组、sh-INHBA-AS1+OE-c-Myc组。平板克隆实验检测各组细胞的增殖能力,FCM检测各组细胞的凋亡情况,Transwell小室实验检测各组细胞的侵袭、迁移能力,qPCR法检测各组细胞中INHBA-AS1、c-Myc、SCD和EMT相关基因(N-cadherin、TGF-β、ZEB1)mRNA的表达,WB法检测各组细胞中c-Myc、SCD、EMT相关(N-cadherin、TGF-β、ZEB1)、S-腺苷-甲硫氨酸脱羧酶(SAMDC)和亚精胺/精胺N1-乙酰转移酶(SSAT)蛋白的表达,ELISA检测各组细胞上清液中鸟氨酸脱羧酶(ODC)的含量。结果:敲减INHBA-AS1表达使HeLa细胞的增殖、侵袭和迁移能力显著降低(均P<0.05)而细胞凋亡率显著升高(P<0.05),q PCR、WB法检测结果显示,敲减INHBA-AS1均可显著抑制HeLa细胞中c-Myc、SCD、N-cadherin、TGF-β、ZEB1和SAMDC的表达(均P<0.05),而促进SSAT的表达(P<0.05),并降低HeLa细胞上清液中ODC的含量(P<0.05)。与c-Myc抑制剂和SCD抑制剂单独处理相比,其联合敲减INHBA-AS1后上述作用更加显著(均P<0.05);与sh-INHBA-AS1组相比,进一步过表达c-Myc后HeLa细胞的增殖能力显著升高(P<0.05)、SCD和N-cadherin蛋白表达水平显著升高(P<0.05)、细胞上清液中ODC含量显著升高(P<0.05)。结论:INHBA-AS1可通过c-Myc调控SCD的表达,从而影响HeLa细胞鸟氨酸代谢和EMT进程,进而促进HeLa细胞的增殖、侵袭和迁移能力。展开更多
BACKGROUND Arachidyl amido cholanoic acid(Aramchol)is a potent downregulator of hepatic stearoyl-CoA desaturase 1(SCD1)protein expression that reduces liver triglycerides and fibrosis in animal models of steatohepatit...BACKGROUND Arachidyl amido cholanoic acid(Aramchol)is a potent downregulator of hepatic stearoyl-CoA desaturase 1(SCD1)protein expression that reduces liver triglycerides and fibrosis in animal models of steatohepatitis.In a phase IIb clinical trial in patients with nonalcoholic steatohepatitis(NASH),52 wk of treatment with Aramchol reduced blood levels of glycated hemoglobin A1c,an indicator of glycemic control.AIM To assess lipid and glucose metabolism in mouse hepatocytes and in a NASH mouse model[induced with a 0.1%methionine and choline deficient diet(0.1MCD)]after treatment with Aramchol.METHODS Isolated primary mouse hepatocytes were incubated with 20μmol/L Aramchol or vehicle for 48 h.Subsequently,analyses were performed including Western blot,proteomics by mass spectrometry,and fluxomic analysis with 13C-uniformly labeled glucose.For the in vivo part of the study,male C57BL/6J mice were randomly fed a control or 0.1MCD for 4 wk and received 1 or 5 mg/kg/d Aramchol or vehicle by intragastric gavage for the last 2 wk.Liver metabolomics were assessed using ultra-high-performance liquid chromatography-time of flight-MS for the determination of glucose metabolism-related metabolites.RESULTS Combination of proteomics and Western blot analyses showed increased AMPK activity while the activity of nutrient sensor mTORC1 was decreased by Aramchol in hepatocytes.This translated into changes in the content of their downstream targets including proteins involved in fatty acid(FA)synthesis and oxidation[PACCα/β(S79),SCD1,CPT1A/B,HADHA,and HADHB],oxidative phosphorylation(NDUFA9,NDUFB11,NDUFS1,NDUFV1,ETFDH,and UQCRC2),tricarboxylic acid(TCA)cycle(MDH2,SUCLA2,and SUCLG2),and ribosome(P-p70S6K[T389]and P-S6[S235/S236]).Flux experiments with 13Cuniformely labeled glucose showed that TCA cycle cataplerosis was reduced by Aramchol in hepatocytes,as indicated by the increase in the number of rounds that malate remained in the TCA cycle.Finally,liver metabolomic analysis showed that glucose homeostasis was improved by Aramchol in 0.1MCD fed mice in a dose-dependent manner,showing normalization of glucose,G6P,F6P,UDP-glucose,and Rbl5P/Xyl5P.CONCLUSION Aramchol exerts its effect on glucose and lipid metabolism in NASH through activation of AMPK and inhibition of mTORC1,which in turn activate FAβ-oxidation and oxidative phosphorylation.展开更多
Dietary macronutrients and micronutrients play important roles in human health.On the other hand,the excessive energy derived from food is stored in the form of triacylglycerol.A variety of dietary and hormonal factor...Dietary macronutrients and micronutrients play important roles in human health.On the other hand,the excessive energy derived from food is stored in the form of triacylglycerol.A variety of dietary and hormonal factors affect this process through the regulation of the activities and expression levels of those key player enzymes involved in fatty acid biosynthesis such as acetyl-CoA carboxylase,fatty acid synthase,fatty acid elongases,and desaturases.As a micronutrient,vitamin A is essential for the health of humans.Recently,vitamin A has been shown to play a role in the regulation of glucose and lipid metabolism.This review summarizes recent research progresses about the roles of vitamin A in fatty acid synthesis.It focuses on the effects of vitamin A on the activities and expression levels of mRNA and proteins of key enzymes for fatty acid synthesis in vitro and in vivo.It appears that vitamin A status and its signaling pathway regulate the expression levels of enzymes involved in fatty acid synthesis.Future research directions are also discussed.展开更多
基金Supported by A Grant from the Drug Applied Research Center of Tabriz University of Medical Sciences,to Darabi M,Research Projects numbers 89/102 and 90/73
文摘AIM:To investigate the effect of MEK/ERK1/2 pathway on peroxisome proliferator-activated receptors(PPARγ)agonist-induced alterations in Δ6-desaturase(Δ6D)and stearoyl-CoA desaturase 1(SCD1)in hepatocellular carcinoma cell line HepG2.METHODS:HepG2 cells cultured in RPMI-1640 were exposed to the commonly used ERK1/2 pathway inhibitor PD98059 and PPARγ agonist,pioglitazone.Total RNA was isolated and reverse transcribed from treated cells.Changes in gene expression and metabolites ratio,as activity index for Δ6D and SCD1,were then determined using reverse transcriptionpolymerase chain reaction and gas liquid chromatography,respectively.RESULTS:The expression of both Δ6D(P = 0.03)and SCD1(P = 0.01)increased following PD98059 treatment,with a higher impact on SCD1(24.5%vs 62.5%).Although pioglitazone increased the mRNA level(1.47 ± 0.10 vs 0.88 ± 0.02,P = 0.006)and activity index(1.40 ± 0.07 vs 0.79 ± 0.11,P < 0.001)of Δ6D,no such changes have been observed for SCD1 activity index in pioglitazone-treated cells.SCD1 gene expression(+26.4%,P = 0.041)and activity index(+52.8%,P = 0.035)were significantly increased by MEK inhibition in the presence of pioglitazone,as compared with pioglitazone alone and control cells.However,the response of Δ6D expression and activity index to pioglitazone was unaffected by incubation with PD98059.CONCLUSION:PPARγ and ERK1/2 signaling pathway affect differentially and may have inhibitory crosstalk effects on the genes expression of 6D and SCD1,and subsequently on their enzymatic activities.
基金National Institutes of Health Grant,No.R01CA172086Plan Nacional of I+D,No.SAF2017-88041-R+5 种基金Ministerio de Economía y Competitividad de España,No.SAF2017-87301-RAsociación Española contra el Cáncer,No.AECC17/302Ayudas Fundación BBVA a equipos de Investigación Científica 2018Fondo Europeo de Desarrollo Regional,Ministerio de Economia y Competitividad de España,No.PGC2018-099857-BI00Basque Government Grants,No.IT1264-19Ministerio de Economia y Competitividad de España for the Severo Ochoa Excellence Accreditation,No.SEV-2016-0644.The funders had no role in study design,data collection and analysis,decision to publish,or preparation of the manuscript.
文摘BACKGROUND Arachidyl amido cholanoic acid(Aramchol)is a potent downregulator of hepatic stearoyl-CoA desaturase 1(SCD1)protein expression that reduces liver triglycerides and fibrosis in animal models of steatohepatitis.In a phase IIb clinical trial in patients with nonalcoholic steatohepatitis(NASH),52 wk of treatment with Aramchol reduced blood levels of glycated hemoglobin A1c,an indicator of glycemic control.AIM To assess lipid and glucose metabolism in mouse hepatocytes and in a NASH mouse model[induced with a 0.1%methionine and choline deficient diet(0.1MCD)]after treatment with Aramchol.METHODS Isolated primary mouse hepatocytes were incubated with 20μmol/L Aramchol or vehicle for 48 h.Subsequently,analyses were performed including Western blot,proteomics by mass spectrometry,and fluxomic analysis with 13C-uniformly labeled glucose.For the in vivo part of the study,male C57BL/6J mice were randomly fed a control or 0.1MCD for 4 wk and received 1 or 5 mg/kg/d Aramchol or vehicle by intragastric gavage for the last 2 wk.Liver metabolomics were assessed using ultra-high-performance liquid chromatography-time of flight-MS for the determination of glucose metabolism-related metabolites.RESULTS Combination of proteomics and Western blot analyses showed increased AMPK activity while the activity of nutrient sensor mTORC1 was decreased by Aramchol in hepatocytes.This translated into changes in the content of their downstream targets including proteins involved in fatty acid(FA)synthesis and oxidation[PACCα/β(S79),SCD1,CPT1A/B,HADHA,and HADHB],oxidative phosphorylation(NDUFA9,NDUFB11,NDUFS1,NDUFV1,ETFDH,and UQCRC2),tricarboxylic acid(TCA)cycle(MDH2,SUCLA2,and SUCLG2),and ribosome(P-p70S6K[T389]and P-S6[S235/S236]).Flux experiments with 13Cuniformely labeled glucose showed that TCA cycle cataplerosis was reduced by Aramchol in hepatocytes,as indicated by the increase in the number of rounds that malate remained in the TCA cycle.Finally,liver metabolomic analysis showed that glucose homeostasis was improved by Aramchol in 0.1MCD fed mice in a dose-dependent manner,showing normalization of glucose,G6P,F6P,UDP-glucose,and Rbl5P/Xyl5P.CONCLUSION Aramchol exerts its effect on glucose and lipid metabolism in NASH through activation of AMPK and inhibition of mTORC1,which in turn activate FAβ-oxidation and oxidative phosphorylation.
基金Supported by the Financial Support of the Overseas Training Program for Outstanding Young and Middle-Aged Teachers in Universities in Jiangsu Province,China(to Yang FC).
文摘Dietary macronutrients and micronutrients play important roles in human health.On the other hand,the excessive energy derived from food is stored in the form of triacylglycerol.A variety of dietary and hormonal factors affect this process through the regulation of the activities and expression levels of those key player enzymes involved in fatty acid biosynthesis such as acetyl-CoA carboxylase,fatty acid synthase,fatty acid elongases,and desaturases.As a micronutrient,vitamin A is essential for the health of humans.Recently,vitamin A has been shown to play a role in the regulation of glucose and lipid metabolism.This review summarizes recent research progresses about the roles of vitamin A in fatty acid synthesis.It focuses on the effects of vitamin A on the activities and expression levels of mRNA and proteins of key enzymes for fatty acid synthesis in vitro and in vivo.It appears that vitamin A status and its signaling pathway regulate the expression levels of enzymes involved in fatty acid synthesis.Future research directions are also discussed.
