共查询到20条相似文献，搜索用时 206 毫秒
1.
AIMTo investigate the role of soluble Klotho protein in THP-1-derived foam cell formation. METHODSTHP-1 monocytes were induced into macrophages by treatment with 160 nmol/L phorbol myristate acetate for 48 h, and then were divided into 6 groups: negative control group (THP-1-derived macrophages), positive control group [THP-1-derived foam cells induced by oxidized low-density lipoprotein (ox-LDL) for 48 h], and 25, 50, 100 and 200 μg/L soluble Klotho protein groups (THP-1-derived macrophages pretreated with soluble Klotho protein at the indicat?ed concentraions for 2 h and then induced by ox-LDL for 48 h). Lipid droplets in cytoplasm were observed by oil red O staining. The cholesterol outflow rate was detected by scintillation counting technique. The content of intracellular total cholesterol, free cholesterol and cholesterol ester was detected by enzyme fluorescence analysis. The expression of acyl-coenzyme A:cholesterol acyltransferase 1 （ACAT1） and ATP-binding cassette transport?er A1 (ABCA1) at mRNA and protein levels was determined by RT-qPCR and Western blot, respectively. RESULTSOil red O staining and lipid mass quantification showed that THP-1-derived foam cell formation was dose-dependently suppressed by soluble Klotho protein. The cholesterol efflux rate of THP-1-derived foam cells was increased by soluble Klotho protein in a dose-dependent manner (P <0.05). In addition, soluble Klotho protein decreased the expression of ACAT1 and increased the expression of ABCA1 in a dose-dependent manner (P <0.05). CONCLUSION The soluble Klotho protein inhibits THP-1-derived foam cell formation in a dose-dependent manner by down-regulating the expression of ACAT1 and up-regulating the expression of ABCA1. 相似文献
2.
YUAN Zhong-hua LIU Xiao-hui WANG Zhong-qun JIA Wei NIU Xi-lin HUANG An-fei LIU Lu-shan YI Guang-hui WANG Zuo REN Zhong TANG Chao-ke TIAN Guo-ping YANG Yong-zong 《园艺学报》2008,24(5):833-842
AIM: To investigate the mechanism of adipophilin accumulating cellular cholesteryl ester in THP-1 macrophages. METHODS: New Zealand rabbit atherosclerotic model was made with high cholesterol diet for 12 weeks. The expressions of adipophilin and PKCα were determined by Western blotting and immunochemical staining in aortic arteries. Cholesteryl ester-loading cells (CE-loading cells) were made from THP-1 macrophages incubated with oxidized low density lipoprotein. In CE-loading cells, expressions of adipophilin and ACAT1 were analyzed by RT-PCR, and the activity of PKC was determined by PepTag assay and spectrophotometry. When the CE-loading cells were incubated with PKC activator PMA and inhibitor calphostin C, expression of adipophilin was observed with RT-PCR, and cellular lipid was measured with oil red O staining and HPLC. The pcDNA3.1-HA-adi vector was transfected to THP-1 macrophage for making adipophilin over expression cells. After the CE-loading adipophilin over expression cells were incubated with or without ACAT inhibitor, the ACAT1 expression and cellular cholesteryl ester were analyzed. RESULTS: Compared with control, both adipophilin and PKCα expression increased in aortic arteries of atherosclerotic animal. In CE-loading THP-1 macrophages, adipophilin and ACAT1 highly were expressed and PKC activity was augmented also. PMA enhanced the high expression of adipophilin and cellular cholesteryl ester in CE-loading THP-1 macrophages, but calphostin C inhibited the effect. ACAT1 expression and cellular cholesteryl ester increased in adipophilin over expression cells, the effect was impaired by incubating with ACAT inhibitor. CONCLUSION: The results suggest that adipophilin increases ACAT1 activity through enhancing PKC activity, resulting in cellular cholesteryl ester accumulation in THP-1 macrophages. 相似文献
3.
AIM:To study the action of ATP binding cassette transporter(ABC) A 1 on cholesterol efflux in THP-1 macrophage-derived foam cells.METHODS:After exposure of the cultured THP-1 macrophage-derived foam cells to 22(R)-hydroxycholesterol and 4, 4'-diisothiocyanostilbene-2, 2'-disulfonic acid (DIDS) at different concentration for 24 hours, cholesterol efflux and ABCA1 mRNA level were determined by FJ-2107P type liquid scintillator and reverse trancriptase-polymerase chaim reaction(RT-PCR), respectively.RESULTS:Oxidized LDL promoted cholesterol efflux in THP-1 macrophages and 22(R)-hydroxycholesterol increased cholesterol efflux in THP-1 macrophage-derived foam cells in a dose-dependent manner and DIDS inhibited cholesterol efflux in THP-1 macrophage-derived foam cells in a dose-dependent manner. Exposure of the cultured THP-1 macrophage-derived foam cells to 22(R)-hydroxycholesterol and DIDS at different concentration for 24 hours, resulted in increase and decrease in the expression of ABCA1 mRNA in THP-1 macrophage-derived foam cells in a dose-dependent manner, respectively.CONCLUSION:ABCA1 playes an important role in cholesterol efflux in THP-1 macrophage-derived foam cells. 相似文献
4.
AIM: To investigate the effect of homocysteine (Hcy) on expression of interleukin-8 (IL-8) mRNA and protein in THP-1-derived macrophages (THP-1 macrophages). METHODS: Cultured THP-1 monocytes were induced to macrophages by 0.1 μmol/L PMA treatment for 72 hours, then the differentiated THP-1 macrophages were incubated with homocysteine (0.01 mmol/L-0.20 mmol/L) for 24 hours, or with 0.10 mmol/L Hcy for various time up to 48 hours. IL-8 protein in THP-1 supernatants was measured by ELISA, and IL-8 mRNA expression was detected by semiquantitive RT-PCR. RESULTS: Compared with control, Hcy significantly increased the expression of IL-8 protein in a concentration-dependent manner. 0.05 mmol/L, 0.10 mmol/L and 0.20 mmol/L Hcy increased IL-8 production by 1.28 fold, 1.32 fold and 1.55 fold, respectively (P＜0.01). IL-8 production were elevated significantly 3 h after treatment with 0.10 mmol/L Hcy. In addition, Hcy also increased IL-8 mRNA expression in a concentration-and time-dependent manner. CONCLUSION: Hcy may contribute to atherogenesis by inducing IL-8 expression and secretion in THP-1 macrophages. 相似文献
5.
6.
7.
AIM: To study the effect of acyl coenzyme A: cholesteryl acyltransferase 1 (ACAT1) antisense oligonucleotides on the formation of foam cells (FC). METHODS: THP-1 cells were cultured and differentiated into macrophages (MP) by phorbol myristate acetate (PMA). Over-expressing ACAT1 gene THP-1 cells were constructed. The ACAT1 antisense and missense oligonucleotides conducted by LipofectamineTM 2000 were incubated with above cells. Ac-LDL was added 6 h later and incubated for 24 h. The expression of ACAT1 protein was detected by Western blotting. The ACAT activity was measured by quantifying the incorporation of ［1-14C］ oleoyl CoA into cholesteryl esters. The formation of foam cells was detected by oil red O staining. RESULTS: The ACAT1 antisense oligonucleotides inhibited the activity of ACAT in macrophages and over-expressing ACAT1 gene THP-1 cells. It also inhibited the formation of foam cell in macrophages and over-expressing ACAT1 gene THP-1 cells with lipid loading. The missense oligonucleotides did not show the inhibitory effects. CONCLUSION: The ACAT1 antisense oligonucleotides inhibit the activity of ACAT and the formation of foam cells. 相似文献
8.
AIM:To investigate the effect of insulin on ox-LDL transferring the THP-1 cells to foam cells and influencing the LPL mRNA expression in THP-1 cells.METHODS:THP-1 cells were incubated with 50 mg/Lox-LDL and insulin at concentrations of 10 mU/L, 100 mU/L, 1 000 mU/L and 10 000 mU/L, respectively. The expression of LPL mRNA in cells was detected by RT-PCR. Lipoprotein lipase of THP-1 cells was presented by no-specific lipase staining. THP-1 cells were stained with oil red O. Accumulation of total cholesterol (TC) in THP-1 cells was determined with oxidase assay.RESULTS:In 100 mU/L、1 000 mU/L、10 000 mU/L insulin groups, LPL mRNA expression increased 2 times, the average cell perilength was longer, the percentage of positive oil red O staining cells was significant higher, the content of cholesterol in THP-1 cells was higher than in ox-LDL control (P＜0.05).CONCLUSION:Insulin accelerates transferring of THP-1 cells to foam cell with exposed to ox-LDL because LPL mRNA expression increased in the cells. 相似文献
9.
AIM: To investigate the regulation of ghrelin on the expression of ATP-binding cassette transporter A1 and G1 (ABCA1/ABCG1)during the foam cell formation. METHODS: The human monocytic leukemia cell line (THP-1)was chosen in our study. The differentiation of THP-1 cells into macrophages was induced by using phorbol myristate acetate (PMA). Macrophages were then incubated with oxidized LDL (ox-LDL)to generate foam cells. Ghrelin of different concentrations were treated at different time points during foam cell formation. The ABCA1/ABCG1 protein and mRNA levels were detected by Western blotting and RT-PCR. The effect of variance of cholesterol content was measured by zymochemistry via-fluorospectrophotometer. RESULTS: Ghrelin reduced the content of lipid droplet in foam cells, and increased the efflux of intracellular cholesterol significantly. Ghrelin increased ABCA1 protein mass and mRNA level in dose-dependent manner. The changes of ABCG1 protein and mRNA level were the same as ABCA1. CONCLUSION: Ghrelin interfere atherosclerosis by up-regulating the expression of ABCA1 and ABCG1. 相似文献
10.
AIM: To establish the THP-1-derived foam cell formation and to evaluate the effects of angiotensin-(1-7) and MDL (an inhibitor of adenylate cyclase) on the expression of ATP-binding cassete transporter A1(ABCA1) and the content of cholesterol. METHODS: THP-1-derived macrophages were treated with oxidized low-density lipoprotein(ox-LDL) to develop into foam cells. The foam cells were divided into 4 groups: control group, MDL group, Ang-(1-7) group and MDL+Ang-(1-7) group. At 24 h after treatment, the content of cAMP was measured by ELISA. The mRNA and protein levels of ABCA1 were determined by real-time RT-PCR and Western blotting, respectively. The content of cholesterol was detected by high performance liquid chromatography. RESULTS: The cAMP, the mRNA and protein levels of ABCA1 in Ang-(1-7) group were significantly higher, and the content of cholesterol was significantly lower than those in control group (P<0.05). On the contrary, the cAMP, the mRNA and protein levels of ABCA1 in MDL group were significantly lower and the content of cholesterol was significantly higher than those in control group (P<0.05). The results in MDL+Ang-(1-7) group were between Ang-(1-7) group and control group. CONCLUSION: Ang-(1-7) inhibits the formation of foam cells by promoting the expression of ABCA1 and decreasing the content of cholesterol. MDL partly antagonizes the effect of Ang-(1-7) by inhibiting the adenylate cyclase and decreasing the content of cAMP. 相似文献
11.
AIM: To investigate the expressions of peroxisome proliferator-activated receptor γ (PPAR γ) and acyl-coenzyme A: cholesterol acyltransferase-1 (ACAT1), and to discuss the mechanisms and pathways of Chlamydia pneumoniae (C.pn)-induced macrophage foam cell formation. METHODS: THP-1-derived macrophages were incubated for 48 h with or without C.pn (1×105 to 1×106 IFU) and/or rosiglitazone (1 to 20 μmol/L), a specific PPAR γ agonist. Lipid droplets in cytoplasm were observed by oil red O staining. The contents of intracellular cholesterol ester were detected by enzyme-fluorescence. PPAR γ, ACAT1 mRNA and protein expressions were determined by RT-PCR and Western blotting, respectively. RESULTS: THP-1-derived macrophages infected with C.pn at concentration of 5×105 and 1×106 IFU resulted in the large accumulation of lipid droplets and the ratio of cholesteryl ester (CE) to total cholesterol (TC) was much higher than 50%when co-incubated with low density lipoprotein (LDL). C.pn up-regulated the expressions of ACAT1 mRNA and protein, and down-regulated the expressions of PPAR γ mRNA and protein in a concentration-dependent manner (P<0.05). Rosiglitazone (10, 20 μmol/L) markedly suppressed the accumulation of lipid droplets and CE by C.pn. Moreover, rosiglitazone inhibited the up-regulation of ACAT1 mRNA and protein expression by C.pn infection in a concentration-dependent manner (P<0.05). CONCLUSION: C.pn induces macrophage foam cell formation by up-regulating ACAT1 expression via PPARγ pathway, which may provide new evidences for the development and progression of atherosclerosis initiated by C.pn infection. 相似文献
12.
AIM: This study was designed to investigate the apoptotic effect of doxycycline in THP-1 cells.METHODS: After differentiated by PMA, THP-1 cells were treated with doxycycline at different concentrations ranging from zero to 80 mg/L. The morphological changes of THP-1 cells were observed under light microscope. MTT assay were used to examine the effects of doxycycline on proliferation of THP-1 cells. Apoptotic THP-1 cells were measured by Annexin-V flow cytometry analysis and TdT-mediated dUTP nick end labeling assay.RESULTS: Treated with a certain concentration of doxycycline, differentiated THP-1 cells contracted and turn round, a number of cells were dead. MTT assay and positive Annexin V-FITC on cell membrane and TUNEL assay showed that doxycycline induced apoptosis in THP-1 cells in a dose-dependent manner.CONCLUSION: Doxycycline induces apoptosis in THP-1 cells in a dose-dependent manner. 相似文献
13.
14.
15.
ZHANG Ju GUAN Heng-yun ZHANG Peng-ju CHEN Wei-wen LIU Wen-wen ZHAO Jian HU Xiao-yan JIANG An-li 《园艺学报》2009,25(8):1495-1500
AIM: To construct a recombinant eukaryotic expression vector, pSilencer 4.1-let-7a1 and to express it in lung cancer A549 cells for detecting its effect on the proliferation of A549 cells. METHODS: The pre-let-7a1 sequence was amplified by RT-PCR using RNA from human lung cancer A549 cells, and then inserted into pSilencer 4.1-CMV neo vector to generate pSilencer 4.1-let-7a1 which was transfected into lung cancer A549 cells. The expression of miRNA let-7a1 was verified by RT-PCR. Its activity in A549 cells was determined by luciferase reporter assay after cotransfection of let-7a1 target sequence-reporter gene plasmid with pMIR-report let-7a1T, which was constructed by inserting let-7a1 target sequence into the luciferase reporter 3’UTR of pMIR-report luciferase vector. The effect of pSilencer 4.1-let-7a1 transfection on A549 cell proliferation was detected by MTT method. RESULTS: The sequences of cloned pre-let-7a1 were correct. RT-PCR results indicated that pSilencer 4.1-let-7a1 was effectively expressed in the transfected A549 cells. The relative luciferase activity was decreased significantly after A549 cells were co-transfected with pSilencer 4.1-let-7a1 and pMIR-report let-7a1T, indicating that let-7a1 was expressed effectively and had biologic activity in A549 cells that were transfected with pSilencer 4.1-let-7a1. MTT results showed that miRNA let-7a1 gene overexpression in A549 inhibited cell proliferation. CONCLUSION: The eukaryotic expression vector pSilencer 4.1-let-7a1 is successfully constructed and effectively expresses in A549 cell. The overexpression of miRNAlet-7a1 gene inhibits lung cancer A549 cell proliferation. 相似文献
16.
AIM: To investigate whether homocysteine (HCY) induce the expression of macrophage inflammatory protein-1α(MIP-1α)in cultured THP-1 monocytes. METHODS: After exposure of THP-1 monocytes to HCY at increasing concentrations (0.05,0.1 and 0.2 mmol/L) for 8 h, or at 0.1 mmol/L of HCY for different incubation times (4, 8 and 16 h), the expressions of MIP-1α mRNA and protein were determined by RT-PCR and immunocytochemistry, respectively. RESULTS: RT-PCR showed that the expression of MIP-1α mRNA increased with the concentrations of HCY compared with the control group. Meanwhile, after the treatment of 0.1 mmol/L HCY to the cells for different times, the MIP-1α mRNA expression increased at 4 h, peaked at 8 h, and then decreased at 16 h. The authenticity of RT-PCR products was confirmed by DNA sequencing. Image analysis of Immunocytochemistry assay showed the expression of MIP-1α protein in experimental groups increased in a dose- and time-dependent manner(P＜0.01). CONCLUSIONS: HCY induced monocytes to express MIP-1α mRNA and protein. 相似文献
17.
LEONG Man-cheong LI Lian-xi YANG Zhi-hong ZHANG Shuo HE Min JIN Hui-ming HU Ren-ming 《园艺学报》2008,24(5):852-855
AIM: To evaluate the expression of TNF-related apoptosis inducing ligand receptor 4 (TRAIL-R4) of THP-1 cells and human aorta smooth muscle cells under high glucose intervention. METHODS: Monocytic cell line THP-1 was incubated with PMA to induce to mature macrophage, Adhesion molecules CD11b and CD11c were assessed by FACS. TRAIL-R4 levels in THP-1 cells treated with different glucose concentrations were determined by Western blotting. The changes of TRAIL-R4 protein expressions were observed at different time points in human aorta smooth muscle cells. Western blotting was employed to evaluate TRAIL-R4 levels after the intervention of PKC activator. RESULTS: Incubation with 160 nmol/L PMA induced mature macrophages. TRAIL-R4 expression was up-regulated after incubation with 20 mmol/L glucose in macrophages. TRAIL-R4 was elevated in a time course manner under high glucose level in human aorta smooth muscle cells. Moreover, activation of PKC induced TRAIL-R4 expressions. CONCLUSION: Up-regulated TRAIL-R4 protein levels induced by high glucose levels might inhibit apoptosis of monocytes and smooth muscle cells and contribute to the progression of atherosclerosis. 相似文献
18.
AIM: To investigate the effects of ghrelin on the expression of acyl coenzyme A:cholesterol acyltransferases-1 (ACAT-1) during the formation of foam cells. METHODS: The human monocytic leukemia cell line THP-1 was used in the study. The differentiation of THP-1 cells into macrophages was induced by phorbol 12-myristate 13-acetate (PMA). Macrophages were incubated with oxidized LDL (ox-LDL) to generate foam cells. Ghrelin of different concentrations were used during the formation of foam cells. The ACAT-1 protein and mRNA levels were detected by Western blotting and RT-PCR. The variance of cholesterol content was measured by zymochemistry via-fluorospectrophotometer. RESULTS: Ghrelin reduced the content of cholesterol ester in foam cells obviously. ACAT-1 protein and mRNA levels were also decreased. Ghrelin reduced ACAT-1 protein mass and mRNA level in a dose-dependent manner. CONCLUSION: Ghrelin might retard the formation of atherosclerosis via down-regulating the expression of ACAT-1. 相似文献
19.
20.
ZHANG Ju GUAN Heng-yun ZHANG Peng-ju CHEN Wei-wen CHEN Zhao-bo NI Na-na ZHU Wen-jie WANG Kun HU Xiao-yan JIANG An-li 《园艺学报》2010,26(9):1764-1768
AIM: To investigate the effect of NKX3.1 on the Dicer1 gene expression.METHODS: The NKX3.1 eukaryotic expression plasmid was transfected into PC3 cells. The stable clones were isolated using cloning cylinders and grew continuously under G418 selection. The gene expression profile in PC3 (+) cells induced by NKX3.1 was analyzed by cDNA microarray. The effect of NKX3.1 on the Dicer1 expression was further investigated by RT-PCR and Western blotting in PC3 and PC3 (+) cells according to the results of gene chip. To determine if the increase in Dicer1 promotes the mature of microRNA, the pMIR-report luciferase expression plasmid of miRNA let-7a1 target sequence (pMIR-report-let7a1T) was constructed and transfected into PC3 and PC3 (+) cells. The effect of the miRNA let-7a-1 on its target sequence was determined by luciferase reporter assay.RESULTS: The result of gene chip showed that the expression level of Dicer1 gene was higher in PC3 (+) cells than that in PC3 cells. The results of RT-PCR and Western blotting indicated that the expression of Dicer1 gene was much higher in PC3 (+) cells than that in PC3 cells. The relative luciferase activity was much lower in PC3 (+) cells than that in PC3 cells when the cells were transfected with the pMIR-report-let7a1T vector.CONCLUSION: Up-regulation of Dicer1 expression induced by NKX3.1 promotes the mature and functions of microRNAs in prostate cancer PC3 cells. 相似文献