Summary: To investigate the effects of leptin on expression of acyl-coenzymeA:cholesterol acyltransferases-1 (ACAT-1) in monocyte-macrophage differentiation, human monocytic cells (THP-1) were cultured in RPMI 1640 an...Summary: To investigate the effects of leptin on expression of acyl-coenzymeA:cholesterol acyltransferases-1 (ACAT-1) in monocyte-macrophage differentiation, human monocytic cells (THP-1) were cultured in RPMI 1640 and made to differentiate into macrophages under the incubation with phorbol myristate acetate (PMA) for 48 h. The cells were divided into 4 groups according to different intervention factors as follows: MCs cultured in RPMI1640 medium with 10 % FBS for 48 h served as MC group (control group), MCs cultured in medium with serum-free RPMI1640 containing 5 % BSA, 100 nmol/L PMA for 48 h as MP group, MCs cultured in RPMI1640 medium with 10 % FBS, 10 μmol/ml leptin for 48 h as leptin-MC group, and MCs cultured in medium with serum-free RPMI1640 containing 5 % BSA, 100 nmol/L PMA, and 10 μmol/ml leptin for 48 h as leptin-MP group. Immunocytochemistry, reverse transcription polymerase chain reaction (RT-PCR) and Western blot were performed, respectively, to observe the effects of leptin on expression of ACAT-1 in the monocyte-macrophage differentiation. Our results showed that expression of ACAT-1 protein and mRNA in MP-group is two times that in MC-group (P<0.05), and the expression of ACAT-1 protein and mRNA increased by up to 4 folds in leptin-MP group as compared with that of MC group (P<0.01). Thus, our results support the idea that expression of ACAT-1 increases more in cultured human macrophages than in monocytes, and leptin can significantly promote ACAT-1 expression. It was concluded that high expression of ACAT-1 may accelerate the development of human atherogenesis,and leptin might participate in atherogenesis by increasing expression of ACAT-1.展开更多
AIM To examine the role that enzyme Acyl-CoA:diacylglycerol acyltransferase-1(DGAT1) plays in postprandial gut peptide secretion and signaling.METHODS The standard experimental paradigm utilized to evaluate the incret...AIM To examine the role that enzyme Acyl-CoA:diacylglycerol acyltransferase-1(DGAT1) plays in postprandial gut peptide secretion and signaling.METHODS The standard experimental paradigm utilized to evaluate the incretin response was a lipid challenge.Following a lipid challenge,plasma was collected via cardiac puncture at each time point from a cohort of 5-8 mice per group from baseline at time zero to 10 h.Incretin hormones [glucagon like peptide-1(GLP-1),peptide tyrosine-tyrosine(PYY) and glucose dependent insulinotropic polypeptide(GIP)] were then quantitated.The impact of pharmacological inhibition of DGAT1 on the incretin effect was evaluated in WT mice.Additionally,a comparison of loss of DGAT1 function either by genetic ablation or pharmacological inhibition.To further elucidate the pathways and mechanisms involved in the incretin response to DGAT1 inhibition,other interventions [inhibitors of dipeptidyl peptidase-IV(sitagliptin),pancreatic lipase(Orlistat),GPR119 knockout mice] were evaluated.RESULTS DGAT1 deficient mice and wildtype C57/BL6J mice werelipid challenged and levels of both active and total GLP-1 in the plasma were increased.This response was further augmented with DGAT1 inhibitor PF-04620110 treated wildtype mice.Furthermore,PF-04620110 was able to dose responsively increase GLP-1 and PYY,but blunt GIP at all doses of PF-04620110 during lipid challenge.Combination treatment of PF-04620110 and Sitagliptin in wildtype mice during a lipid challenge synergistically enhanced postprandial levels of active GLP-1.In contrast,in a combination study with Orlistat,the ability of PF-04620110 to elicit an enhanced incretin response was abrogated.To further explore this observation,GPR119 knockout mice were evaluated.In response to a lipid challenge,GPR119 knockout mice exhibited no increase in active or total GLP-1 and PYY.However,PF-04620110 was able to increase total GLP-1 and PYY in GPR119 knockout mice as compared to vehicle treated wildtype mice.CONCLUSION Collectively,these data provide some insight into the mechanism by which inhibition of DGAT1 enhances intestinal hormone release.展开更多
Objective:To investigate the effects of rosiglitazone,a synthetic ligand of peroxisome proliferators-activated receptor gamma(PPARγ),on the expression of acyl-coenzyme A:cholesterol acyltransferase-1(ACAT-1) in phorb...Objective:To investigate the effects of rosiglitazone,a synthetic ligand of peroxisome proliferators-activated receptor gamma(PPARγ),on the expression of acyl-coenzyme A:cholesterol acyltransferase-1(ACAT-1) in phorbol myristate acetate(PMA)-pretreated THP-1 cells after the inducement of advanced glycation end products(AGEs).Methods:After THP-1 cells were cultured in the presence of 0.1 μmol/L PMA for 72 h to induce phagocytic differentiation,the obtained THP-1 macrophages were treated with rosiglitazone for 4 h at different concentrations(1,5 or 10 μmol/L) and then exposed to AGEs-modified bovine serum albumin(AGEs-BSA) for 24 h at a concentration of 200 mg/L.Reverse transcription polymerase chain reaction(RT-PCR) and Western blot analysis were performed to detect the mRNA and protein expressions of ACAT-1 respectively.Results:Administration of AGEs-BSA(200 mg/L) into the THP-1 macrophages resulted in up-regulation of ACAT-1 at mRNA and protein levels when compared with the expressions in macrophages incubated with serum-free RPMI1640.Pretreatment of rosiglitazone inhibited significantly the increased expression of ACAT-1 induced by AGEs-BSA in a concentration-dependent manner.Conclusion:PPARγ activation by rosiglitazone down-regulates ACAT-1 expression induced by AGEs in THP-1 macrophages,which might provide a new way for treating atherogenesis in diabetic patients.展开更多
文摘Summary: To investigate the effects of leptin on expression of acyl-coenzymeA:cholesterol acyltransferases-1 (ACAT-1) in monocyte-macrophage differentiation, human monocytic cells (THP-1) were cultured in RPMI 1640 and made to differentiate into macrophages under the incubation with phorbol myristate acetate (PMA) for 48 h. The cells were divided into 4 groups according to different intervention factors as follows: MCs cultured in RPMI1640 medium with 10 % FBS for 48 h served as MC group (control group), MCs cultured in medium with serum-free RPMI1640 containing 5 % BSA, 100 nmol/L PMA for 48 h as MP group, MCs cultured in RPMI1640 medium with 10 % FBS, 10 μmol/ml leptin for 48 h as leptin-MC group, and MCs cultured in medium with serum-free RPMI1640 containing 5 % BSA, 100 nmol/L PMA, and 10 μmol/ml leptin for 48 h as leptin-MP group. Immunocytochemistry, reverse transcription polymerase chain reaction (RT-PCR) and Western blot were performed, respectively, to observe the effects of leptin on expression of ACAT-1 in the monocyte-macrophage differentiation. Our results showed that expression of ACAT-1 protein and mRNA in MP-group is two times that in MC-group (P<0.05), and the expression of ACAT-1 protein and mRNA increased by up to 4 folds in leptin-MP group as compared with that of MC group (P<0.01). Thus, our results support the idea that expression of ACAT-1 increases more in cultured human macrophages than in monocytes, and leptin can significantly promote ACAT-1 expression. It was concluded that high expression of ACAT-1 may accelerate the development of human atherogenesis,and leptin might participate in atherogenesis by increasing expression of ACAT-1.
文摘AIM To examine the role that enzyme Acyl-CoA:diacylglycerol acyltransferase-1(DGAT1) plays in postprandial gut peptide secretion and signaling.METHODS The standard experimental paradigm utilized to evaluate the incretin response was a lipid challenge.Following a lipid challenge,plasma was collected via cardiac puncture at each time point from a cohort of 5-8 mice per group from baseline at time zero to 10 h.Incretin hormones [glucagon like peptide-1(GLP-1),peptide tyrosine-tyrosine(PYY) and glucose dependent insulinotropic polypeptide(GIP)] were then quantitated.The impact of pharmacological inhibition of DGAT1 on the incretin effect was evaluated in WT mice.Additionally,a comparison of loss of DGAT1 function either by genetic ablation or pharmacological inhibition.To further elucidate the pathways and mechanisms involved in the incretin response to DGAT1 inhibition,other interventions [inhibitors of dipeptidyl peptidase-IV(sitagliptin),pancreatic lipase(Orlistat),GPR119 knockout mice] were evaluated.RESULTS DGAT1 deficient mice and wildtype C57/BL6J mice werelipid challenged and levels of both active and total GLP-1 in the plasma were increased.This response was further augmented with DGAT1 inhibitor PF-04620110 treated wildtype mice.Furthermore,PF-04620110 was able to dose responsively increase GLP-1 and PYY,but blunt GIP at all doses of PF-04620110 during lipid challenge.Combination treatment of PF-04620110 and Sitagliptin in wildtype mice during a lipid challenge synergistically enhanced postprandial levels of active GLP-1.In contrast,in a combination study with Orlistat,the ability of PF-04620110 to elicit an enhanced incretin response was abrogated.To further explore this observation,GPR119 knockout mice were evaluated.In response to a lipid challenge,GPR119 knockout mice exhibited no increase in active or total GLP-1 and PYY.However,PF-04620110 was able to increase total GLP-1 and PYY in GPR119 knockout mice as compared to vehicle treated wildtype mice.CONCLUSION Collectively,these data provide some insight into the mechanism by which inhibition of DGAT1 enhances intestinal hormone release.
文摘Objective:To investigate the effects of rosiglitazone,a synthetic ligand of peroxisome proliferators-activated receptor gamma(PPARγ),on the expression of acyl-coenzyme A:cholesterol acyltransferase-1(ACAT-1) in phorbol myristate acetate(PMA)-pretreated THP-1 cells after the inducement of advanced glycation end products(AGEs).Methods:After THP-1 cells were cultured in the presence of 0.1 μmol/L PMA for 72 h to induce phagocytic differentiation,the obtained THP-1 macrophages were treated with rosiglitazone for 4 h at different concentrations(1,5 or 10 μmol/L) and then exposed to AGEs-modified bovine serum albumin(AGEs-BSA) for 24 h at a concentration of 200 mg/L.Reverse transcription polymerase chain reaction(RT-PCR) and Western blot analysis were performed to detect the mRNA and protein expressions of ACAT-1 respectively.Results:Administration of AGEs-BSA(200 mg/L) into the THP-1 macrophages resulted in up-regulation of ACAT-1 at mRNA and protein levels when compared with the expressions in macrophages incubated with serum-free RPMI1640.Pretreatment of rosiglitazone inhibited significantly the increased expression of ACAT-1 induced by AGEs-BSA in a concentration-dependent manner.Conclusion:PPARγ activation by rosiglitazone down-regulates ACAT-1 expression induced by AGEs in THP-1 macrophages,which might provide a new way for treating atherogenesis in diabetic patients.
