ABSTRACT
Objective:To clone and sequence the full-length of human PPAR? cDNA in order to further construct the eukaryotic expression vector carrying hPPAR? gene.Methods:hPPAR? cDNA was cloned from HepG2 cells total RNA by RT-PCR.The PCR product recovered from gel were ligated with pMD19-T vector and transformed into DH5? competent cell.The integrity and fidelity of hPPAR? cDNA sequence inserted in T vector were verified by BamH I and Sal I double excising and DNA sequencing assays.Results:The positive clone T vector plasmid containing correct sequence of hPPAR? cDNA were verified by enzyme digestion as well as sequence analysis and was named as pMD19-hPPAR?-T vector.The sequence of inserted hPPAR? cDNA was in accordance with the corresponding sequence in GeneBank database(AY919140).Conclusion:hPPAR? gene is successfully cloned and the pMD19-hPPAR?-T intermediate vector is constructed,which provide a good basis for further constructing the eukaryotic expression vector carrying hPPAR? gene and studying the receptor's function.
ABSTRACT
Objective To investigate the protective effect of peroxisome proliferator-activated receptor ?(PPAR?) activator 15-deoxyprostaglandin J2(15d-PGJ2) in rat hepatic ischemia-reperfusion injury and its mechanism.Methods The models of 70% warm ischemia-reperfusion injury were established in SD rats,rats were randomly divided into 4 groups: sham operation group,ischemia-reperfusion group,15d-PGJ2 group and 15d-PGJ2+GW9662 group.After reperfusion,serum AST and ALT levels were determined;the liver tissues were removed for measurement of activity of NF-?B and myeloperoxidase(MPO),TNF-? content and expression of ICAM-1.Results Compared with sham operation group,the serum levels of ALT and AST,and the activities of MPO and NF-?B,TNF-? content and expression of ICAM-1 in ischemia-reperfusion group,15d-PGJ2 group and 15d-PGJ2+GW9662 group were greatly improved(P
ABSTRACT
Objective:To study the inhibitory effects of peroxisome proliferator-activated receptor ? activator Fenofibrate on the angiotensin Ⅱ(Ang Ⅱ)-induced cardiac hypertrophy in vitro. Methods: A model of hypertrophy of neonatal rat cardiac myocytes was established with Ang Ⅱ stimulation.With the aid of Leca Qwin Image software,the surface areas of cardiac myocytes were analyzed.The mRNA expression of ?、?-myosin heavy chains(?-MHC、?-MHC) and PPAR? was measured by reverse transcription-polymerase chain reaction(RT-PCR),and the cultured myocyte viability was estimated by MTT assay. Results:Fenofibrate pretreatment 24 h prior to Ang ⅡSignificantly reduced Ang Ⅱ-induced cardiac hypertrophy,inhibited the effect of AngⅡ on the cardiac myocyte viability and increased expression of?/?-MHC mRNA and PPAR? mRNA in a dose-dependent manner.In contrast,Fenofibrate had no significant effect on Ang Ⅱ treated cardiac myocytes when Fenofibrate treatment was concomitant with Ang Ⅱ. Conclusion: PPAR?-dependent pathway was involved in the inhibition of cardiac hypertrophy,but chronic treatment was needed.
ABSTRACT
Aim To study the effect of Curcumin on the apoptosis of hepatic stellate cells (HSC), and the correlation between the effect and peroxisome proliferator-activated receptor ?(PPAR?) signal.Methods The HSC was isolated from normal SD rats through in situ perfusion of liver with protease E and density-gradient centrifugation with Nycodenz.The subcultured cells were treated with corresponding compounds. Cell apoptosis was detected by Hoechst 33258 staining. PPAR? subcellular distribution was detected by immunofluorescent staining. Total RNA, total protein and nuclear protein were extracted respectively, target gene and protein levels were determined by semi-quantitative RT-PCR or Western blot.Results There was nearly no apoptosis in activated HSC. Curcumin treatment induced the apoptosis of HSC, enhancing PPAR? nuclear translocation/redistribution.At the transcription and translation level,curcumin upregulated nuclear PPAR? expression, inhibited anti-apoptotic Bcl-2 expression, and promoted pro-apoptotic Bax expression; but all these effects could be reversed by PPAR? antagonist GW9662.Conclusions Curcumin induces HSC apoptosis by enhancing PPAR? expression and nuclear translocation/redistribution.
ABSTRACT
Objective To study the regulatory ability of peroxisome proliferator-activated receptor ?(PPAR?) ligands to the inflammatory response in human gallbladder epithelial cells. Methods Culture human gallbladder epithelial cells and identify them . Cells were treated for 24 hours with 0, 10 ?mol/L, 20 ?mol/L, 30 ?mol/L, 50 ?mol/L and 100 ?mol/L of Ciglitazone during cellular growth peak(5th day), then stimulated them with hIL-1? 5 ng/ml for 2 hours and measured the concentration of IL-6、IL-8 and TNF-? in cellular supernatants by riadioimmunoassay. Results Contrasted with control group, the expression of IL-6 and IL-8 in each test group were inhibited ((P
ABSTRACT
Objective To investigate the significance of expression of peroxisome proliferator activated receptor ?(PPAR?) and retinoid X receptor ?(RXR?) in chronic gastritis, gastric mucosal dysplasia and gastric carcinoma and to identify the correlation between PPAR? and RXR?. Methods Avidin biotin peroxidase complex immunohistochemical methods were adopted to examine the expression of PPAR? and RXR? in 53 patients with gastric carcinoma, and 18 with gastric mucosal dysplasia, 31 with chronic non atrophic gastritis and 30 with chronic atrophic gastritis were served as controls. Results The positive rates of PPAR? and RXR? were 41.5% and 54.7% in gastric carcinoma respectively, 27.8% and 38.9% in gastric mucosal dysplasia, 10.0% and 20.0% in chronic atrophic gastritis, 6.5% and 16.1% in chronic non atrophic gastritis. From chronic non atrophic gastritis, chronic atrophic gastritis to gastric carcinoma, expressions of PPAR? and RXR? showed an ascending tendency. Compared with those in chronic gastritis, expressions of PPAR? and RXR? in gastric mucosal dysplasia and gastric carcinoma were significantly enhanced ( P0.05). There was a significant correlation between expressions of PPAR? and RXR? in gastric carcinoma ( r =0.54). Conclusion PPAR? and RXR? protein overexpression is a relatively early event in gastric carcinogenesis, and it may play both an independent and synergetic role in progression of gastric carcionma.
ABSTRACT
Objective To investigate the effects of valsartan on the expression of acyl-coenzyme A: cholesterol acyltransferase-1(ACAT-1) and peroxisome proliferator activated receptor-gamma(PPAR-?) in atherosclerotic plaques on rabbit aortic wall.Methods Twenty-four male Japanese white rabbits were randomly assigned into three groups(8 each): control group,valsartan group and high cholesterol feeding group.All rabbits were fed according to the experimental protocol for 12 weeks.Blood samples were taken from vein for measurement of serum lipids.The ratio of intima/media thickness of the aorta was measured.ACAT-1 mRNA/protein and PPAR-? mRNA/protein were determined by reverse transcription polymerase chain reaction(RT-PCR) and Western blotting,respectively.Results After 12 weeks,the levels of serum total cholesterol(TC),triglyeride(TG) and low density lipoprotein-cholesterol(LDL-C) in valsartan group and cholesterol group were significantly higher than those in control group(P0.05).The intima thickness and the ratio of intima/media in carotid arteries in cholesterol group were significantly higher than those in control group and valsartan group(P
ABSTRACT
Objective To study the relationship between the expression of peroxisome proliferator-activated receptor? (PPAR?) mRNA and the severity of coronary lesions in coronary heart disease(CHD) patients. Methods 153 patients with CHD who had undergone coronary angiography were admitted, the expression of PPAR? mRNA in peripheral lymphocytes was assessed using RT-PCR. The severity of coronary artery lesions was analyzed, and the lesions of coronary artery were delineated. Results Compared with control, in CHD patients IRI BMI TC LDL-C were elevated and HDL-C decreased. Significant decrease in PPAR? mRNA was observed in CHD patients (0.40?0.12 vs 0.62?0.13, P
ABSTRACT
AIM: To investigate the effects of rosiglitazone(ROSI),an agonist of peroxisome proliferator-activated receptor ?(PPAR?),on the lung expression of intercellular adhesion molecule-1(ICAM-1) and cytokine-induced neutrophil chemoattractant(CINC) in rats with acute lung injury. METHODS: Thirty-six male Wistar rats were randomly divided into six groups: control group,ROSI group,GW9662(a PPAR? antagonist) group,lipopolysaccharide(LPS,6 mg/kg,iv) group,ROSI-LPS group(0.3 mg/kg ROSI iv 30 min prior to LPS) and GW9662-ROSI-LPS group(0.3 mg/kg GW9662,iv,20 min before ROSI).Four hours after LPS injection,wet/dry weight(W/D) ratio,myeloperoxidase (MPO) activity,malondialdehyde(MDA) and CINC-1 concentrations were assayed in the lung tissues.Immunohistochemical analysis of ICAM-1 expression was also studied.RESULTS: Pretreatment with ROSI significantly attenuated LPS-induced increases in W/D ratio,MPO activity,MDA and CINC-1 concentrations as well as ICAM-1 expression in the lung tissues.The specific PPAR? antagonist GW9662 antagonized the effects of ROSI.CONCLUSION: Pretreatment with ROSI reduces LPS-induced lung injury in rats.The mechanism involves inhibition of the lung expression of ICAM-1 and CINC-1 by the activation of PPAR?.
ABSTRACT
Objective To study the role of PPARa gene in human hepatocellular carcinoma. Methods Transmission elctron microscopy was used to study the morphologic differences between the MDR cell line HepG2/ADM and its mother cell line HepG2;Real-time RT-PCR and Western blotting were used to check the expression of PPARa in mRNA level and protein level in the two cell lines respectively. Results Transmission elctron microscopy showed that the HepG2/ADM cells had more viliformed maculas on their nuclear memberanes, more vacuoles in their cytoplasmas, more rough endoplastic reticulums; the PPARa gene was down regulated in the HepG2/ADM cells. Conclusion PPARa is related with the multidrug resistance phenomenon in the HepG2/ADM cells. The down regulation of PPARa may be one of the mechanisms conferring the forming of MDR in tumor cells.
ABSTRACT
Objective:To investigate the effect of advanced glycosylation end-products(AGEs) on the expression of peroxisome proliferator-activated receptor-?(PPAR?) in THP-1 macrophages and explore the function of AGEs in atherosclerosis.Methods: Differentiation of THP-1 cell was induced by PMA.The THP-1 macrophages were exposed to AGE-modified bovine serum albumin(AGE-BSA) in different concentration or the same concentration for different time.Expression of PPAR? mRNA and protein in THP-1 macrophages was measured by RT-PCR and immunocytochemical method.Results:RT-PCR showed that expression of PPAR? mRNA was 1.235?0.044,0.752?0.055,0.494?0.026,0.277?0.025 in 50,100,200 and 400?g/ml AGE-BSA groups.PPAR? mRNA expression in THP-1 macrophages following 200?g/ml AGE-BSA for 12,24,36,48h was 1.260?0.043,0.467?0.033,0.360?0.012.Western-Blot showed that after exposure of THP-1 macrophages to 50,100,200and 400mg/L AGE-BSA,the average integrated optical density values of PPAR? protein expression were 36.460?0.625,24.561?0.636,19.326?0.803,12.715?0.752 respetively,significantly lower than that in BSA group(P
ABSTRACT
Objective:To explore the effects of the nuclear receptor peroxisome proliferator-activated receptor (PPAR)? activators on tumor necrosis factor ? (TNF-?) expression in neonatal rat cardiac myocytes.Methods:Primary culture of cardiac myocytes was prepared from 1～3 days old Spraque-Dawley rats.Cardiac myocytes were pretreated with rosiglitazone at different concentrations and then stimulated with angiotensin Ⅱ(1 ?mol/L).ELISA and reverse transcripition-polymerase chain reaction(RT-PCR) were used to examine TNF-? in cultured supernatants and TNF-? mRNA in cardiac myocytes,respectively.Results:Pretreatment of cardiac myocytes with rosiglitazone inhibited the increase of TNF-? and TNF-? mRNA induced by angiotensinⅡ in a concentration-dependent manner.Conclusion:Rosiglitazone inhibits the expression of TNF-? in cardiac myocytes,which provides a new way for treating chronic heart failure.
ABSTRACT
Objective:To observe the protective effect of peroxisome proliferator-activated receptor ?(PPAR?) on TNF?-induced injury of the inner medullary collecting duct(IMCD) cells.Methods: Cultured IMCD cells were randomly divided into blank control,TNF? stimulation,PPAR? transfection(pPPAR?-control) and PPAR? transfection + TNF? stimulation groups.Stimulation was induced with 50 ng/ml TNF? for 24 h and mouse wild-type PPAR? plasmid was used to transfect IMCD cells.Cell supernatant MCP-1 or TGF?_1 was detected by ELISA method.Results: IMCD cells transfected with wild-type PPAR? plasmid had high expression of PPAR? mRNA and protein.The contents of MCP-1 and TGF?_1 in the supernatant were similar in blank control group and pPPAR?-control group.Compared with blank control group,TNF? stimulation group had decreased contents of MCP-1 and TGF?_1in the supernatant(P
ABSTRACT
Objective: To investigate the effects of green tea polysaccharides (TPs) on glucose metabolism and the activity of peroxisome proliferator-activated receptor gamma (PPAR-?) in KKAy type 2 diabetic mice. Methods: Glucose tolerance test, fasting and postprandial glucose, gluconeogenesis, and insulin sensitivity were investigated in type 2 diabetic mice with orally administered TPs at the dose of 500mg/kg for 4-10 w. Effect of TPs on activity of PPAR-? was tested in vitro. Results: TPs could not only improve glucose tolerance, but also reduce fasting and postprandial blood glucose. In addition, TPs could inhibit gluconeogenesis and enhance insulin sensitivity in KKAy diabetic mice. TPs had also an effect of activating of PPAR-? with dose-response. Conclusion: TPs have beneficial effect of lowering blood glucose in KKAy type 2 diabetic mice, which may be induced by enhancing insulin sensitivity by activating of PPAR-?.