In order to explore the effect and mechanisms of tumor necrosis factor-α （TNF-α） on the activity of the acyl coenzyme A： cholesteryl acyltransferase （ACAT）, THP-I monocytes were cul- tured and induced to differ...In order to explore the effect and mechanisms of tumor necrosis factor-α （TNF-α） on the activity of the acyl coenzyme A： cholesteryl acyltransferase （ACAT）, THP-I monocytes were cul- tured and induced to differentiate into macrophages with phorbol ester. TNF-α （60 ng/mL） was added at different time points into the macrophage-containing medium and the ACAT enzyme activity was measured by quantifying the incorporation of [1-^14C] oleoyl CoA into cholesteryl esters. The expression of ACAT-1 protein and mRNA was respectively detected by Western blotting and RT-PCR in THP-1 macrophages 24 h after treatment with TNF-α （60 ng/mL）. The results indicated that ACAT activity in THP-I macrophages treated with TNF-α was increased in a time-dependent manner. The expression levels of ACAT-1 protein and mRNA were significantly increased in THP-I macrophages after treatment with TNF-α （P〈0.05）. It was suggested that TNF-α could increase the activity of ACAT in THP-1 macrophages by up-regulating the expression of ACAT-1 gene.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
目的:利用小干扰RNA(si RNA)技术干扰酰基辅酶A-胆固醇酰基转移酶1(ACAT1)基因在人结肠癌HT29细胞中的表达,观察ACAT1基因表达下降对于人结肠癌HT29细胞增殖和侵袭迁移能力的影响,了解ACAT1在结肠癌发生发展中的作用。方法:通过脂质体Hi...目的:利用小干扰RNA(si RNA)技术干扰酰基辅酶A-胆固醇酰基转移酶1(ACAT1)基因在人结肠癌HT29细胞中的表达,观察ACAT1基因表达下降对于人结肠癌HT29细胞增殖和侵袭迁移能力的影响,了解ACAT1在结肠癌发生发展中的作用。方法:通过脂质体Hiper Fect将ACAT1 si RNA转染至人结肠癌HT29细胞中干扰ACAT1基因的表达,利用Real time PCR检测转染前后ACAT1基因表达的变化,利用ACAT1基因干扰前后的HT29细胞,采用CFSE染色法检测HT29细胞增殖能力的变化;Transwell小室法检测HT29细胞侵袭和迁移能力的变化。结果:ACAT1 si RNA转染HT29细胞后ACAT1 m RNA的表达明显下降,ACAT1 si RNA转染后的HT29细胞与转染前比较,增殖及侵袭转移的能力均受到明显抑制,差异有统计学意义(P<0.05)。结论:ACAT1基因干扰使HT29细胞的增殖减慢,侵袭迁移能力降低,为结肠癌治疗提供新的靶点。展开更多
基金This project was supported by a grant from the National Natural Sciences Foundation of China (No. 30170378)
文摘In order to explore the effect and mechanisms of tumor necrosis factor-α （TNF-α） on the activity of the acyl coenzyme A： cholesteryl acyltransferase （ACAT）, THP-I monocytes were cul- tured and induced to differentiate into macrophages with phorbol ester. TNF-α （60 ng/mL） was added at different time points into the macrophage-containing medium and the ACAT enzyme activity was measured by quantifying the incorporation of [1-^14C] oleoyl CoA into cholesteryl esters. The expression of ACAT-1 protein and mRNA was respectively detected by Western blotting and RT-PCR in THP-1 macrophages 24 h after treatment with TNF-α （60 ng/mL）. The results indicated that ACAT activity in THP-I macrophages treated with TNF-α was increased in a time-dependent manner. The expression levels of ACAT-1 protein and mRNA were significantly increased in THP-I macrophages after treatment with TNF-α （P〈0.05）. It was suggested that TNF-α could increase the activity of ACAT in THP-1 macrophages by up-regulating the expression of ACAT-1 gene.
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.
文摘目的:利用小干扰RNA(si RNA)技术干扰酰基辅酶A-胆固醇酰基转移酶1(ACAT1)基因在人结肠癌HT29细胞中的表达,观察ACAT1基因表达下降对于人结肠癌HT29细胞增殖和侵袭迁移能力的影响,了解ACAT1在结肠癌发生发展中的作用。方法:通过脂质体Hiper Fect将ACAT1 si RNA转染至人结肠癌HT29细胞中干扰ACAT1基因的表达,利用Real time PCR检测转染前后ACAT1基因表达的变化,利用ACAT1基因干扰前后的HT29细胞,采用CFSE染色法检测HT29细胞增殖能力的变化;Transwell小室法检测HT29细胞侵袭和迁移能力的变化。结果:ACAT1 si RNA转染HT29细胞后ACAT1 m RNA的表达明显下降,ACAT1 si RNA转染后的HT29细胞与转染前比较,增殖及侵袭转移的能力均受到明显抑制,差异有统计学意义(P<0.05)。结论:ACAT1基因干扰使HT29细胞的增殖减慢,侵袭迁移能力降低,为结肠癌治疗提供新的靶点。
