肺疾病,慢性阻塞性%过氧化物酶体增殖物激活受体%罗格列酮%NF-κB%肿瘤坏死因子α
肺疾病,慢性阻塞性%過氧化物酶體增殖物激活受體%囉格列酮%NF-κB%腫瘤壞死因子α
폐질병,만성조새성%과양화물매체증식물격활수체%라격렬동%NF-κB%종류배사인자α
Pulmonary disease,chronic obstructive%Peroxisome proliferator-activated receptors%Rosiglitazone%NF-kappa B%Tumor necrosis factor-alpha
目的 探讨罗格列酮对COPD患者过氧化物酶体增殖物激活受体-γ(PPAR-γ)、核因子-κB及肿瘤坏死因子-α(TNF-α)的影响.方法 2010年4-11月,选择COPD急性加重期患者30例,其中男22例,女8例,年龄54～87岁,平均(72±9)岁；健康体检者(对照组)24例,其中男18例,女6例,年龄52 ～80岁,平均(69±10)岁.抽取受试者空腹肘静脉血10 ml,分离培养外周血单个核细胞(PBMCs).将30例COPD患者的PBMCs均分为3份,分别根据不同的药物干预分为非干预组、罗格列酮干预组(罗格列酮组)和罗格列酮联合GW9662干预组(联合干预组),对照组PBMCs不加任何药物干预.采用实时荧光定量PCR检测PBMCs中PPAR-γ和核因子-κB的mRNA表达,细胞免疫荧光法结合激光扫描共聚焦显微镜检测PPAR-γ和核因子-κB蛋白表达及核转位,酶联免疫吸附试验检测细胞培养上清液中TNF-α含量.多组间比较采用方差分析,组间两两比较采用LSD-t检验,相关性分析采用Pearson检验.结果 mRNA(2-ΔΔCt值)和蛋白表达(荧光强度值):非干预组PPAR-γ(0.52±0.10和55±11)低于对照组(1和85±9),核因子-κB(1.69±0.07和145±17)高于对照组(1和118±7)；罗格列酮组PPAR-γ(4.47±0.11和204±12)高于非干预组,核因子-κB(0.33±0.04和59±14)低于非干预组；联合干预组PPAR-γ(2.25±0.31和142±23)低于罗格列酮组,高于非干预组,核因子-κB(0.64±0.02和90±10)高于罗格列酮组,低于非干预组；组间比较差异均有统计学意义(F值为29.21～567.42,均P＜0.01).TNF-α浓度(μg/L):非干预组(96.2±1.4)高于对照组(85.3±1.0),罗格列酮组(63.0±2.5)低于非干预组,联合干预组(83.3±1.9)高于罗格列酮组,低于非干预组,组间比较差异有统计学意义(F值为293.72,P＜0.01).非干预组PPAR-γ蛋白主要位于细胞质,核因子-kB蛋白主要位于细胞核；对照组PPAR-γ和核因子-κB蛋白在PBMCs细胞质和细胞核中均有表达；罗格列酮组PPAR-γ蛋白转位于细胞核,核因子-kB蛋白转位于细胞质；联合干预组PPAR-γ蛋白转回细胞质,核因子-κB蛋白部分转移至细胞核.相关分析结果表明,PPAR-γ蛋白表达与核因子-κB蛋白表达及TNF-α浓度均呈负相关(r值分别为-0.935和-0.924,均P＜0.01),核因子-κB蛋白表达与TNF-α的浓度呈正相关(r=0.846,P＜0.01).结论 COPD的炎症可能与PPAR-γ表达及活性不足有关,罗格列酮能通过上调PPAR-γ表达和活性抑制核因子-kB,进而抑制TNF-α分泌,在COPD炎症中起着重要作用.
目的 探討囉格列酮對COPD患者過氧化物酶體增殖物激活受體-γ(PPAR-γ)、覈因子-κB及腫瘤壞死因子-α(TNF-α)的影響.方法 2010年4-11月,選擇COPD急性加重期患者30例,其中男22例,女8例,年齡54～87歲,平均(72±9)歲；健康體檢者(對照組)24例,其中男18例,女6例,年齡52 ～80歲,平均(69±10)歲.抽取受試者空腹肘靜脈血10 ml,分離培養外週血單箇覈細胞(PBMCs).將30例COPD患者的PBMCs均分為3份,分彆根據不同的藥物榦預分為非榦預組、囉格列酮榦預組(囉格列酮組)和囉格列酮聯閤GW9662榦預組(聯閤榦預組),對照組PBMCs不加任何藥物榦預.採用實時熒光定量PCR檢測PBMCs中PPAR-γ和覈因子-κB的mRNA錶達,細胞免疫熒光法結閤激光掃描共聚焦顯微鏡檢測PPAR-γ和覈因子-κB蛋白錶達及覈轉位,酶聯免疫吸附試驗檢測細胞培養上清液中TNF-α含量.多組間比較採用方差分析,組間兩兩比較採用LSD-t檢驗,相關性分析採用Pearson檢驗.結果 mRNA(2-ΔΔCt值)和蛋白錶達(熒光彊度值):非榦預組PPAR-γ(0.52±0.10和55±11)低于對照組(1和85±9),覈因子-κB(1.69±0.07和145±17)高于對照組(1和118±7)；囉格列酮組PPAR-γ(4.47±0.11和204±12)高于非榦預組,覈因子-κB(0.33±0.04和59±14)低于非榦預組；聯閤榦預組PPAR-γ(2.25±0.31和142±23)低于囉格列酮組,高于非榦預組,覈因子-κB(0.64±0.02和90±10)高于囉格列酮組,低于非榦預組；組間比較差異均有統計學意義(F值為29.21～567.42,均P＜0.01).TNF-α濃度(μg/L):非榦預組(96.2±1.4)高于對照組(85.3±1.0),囉格列酮組(63.0±2.5)低于非榦預組,聯閤榦預組(83.3±1.9)高于囉格列酮組,低于非榦預組,組間比較差異有統計學意義(F值為293.72,P＜0.01).非榦預組PPAR-γ蛋白主要位于細胞質,覈因子-kB蛋白主要位于細胞覈；對照組PPAR-γ和覈因子-κB蛋白在PBMCs細胞質和細胞覈中均有錶達；囉格列酮組PPAR-γ蛋白轉位于細胞覈,覈因子-kB蛋白轉位于細胞質；聯閤榦預組PPAR-γ蛋白轉迴細胞質,覈因子-κB蛋白部分轉移至細胞覈.相關分析結果錶明,PPAR-γ蛋白錶達與覈因子-κB蛋白錶達及TNF-α濃度均呈負相關(r值分彆為-0.935和-0.924,均P＜0.01),覈因子-κB蛋白錶達與TNF-α的濃度呈正相關(r=0.846,P＜0.01).結論 COPD的炎癥可能與PPAR-γ錶達及活性不足有關,囉格列酮能通過上調PPAR-γ錶達和活性抑製覈因子-kB,進而抑製TNF-α分泌,在COPD炎癥中起著重要作用.
목적 탐토라격렬동대COPD환자과양화물매체증식물격활수체-γ(PPAR-γ)、핵인자-κB급종류배사인자-α(TNF-α)적영향.방법 2010년4-11월,선택COPD급성가중기환자30례,기중남22례,녀8례,년령54～87세,평균(72±9)세；건강체검자(대조조)24례,기중남18례,녀6례,년령52 ～80세,평균(69±10)세.추취수시자공복주정맥혈10 ml,분리배양외주혈단개핵세포(PBMCs).장30례COPD환자적PBMCs균분위3빈,분별근거불동적약물간예분위비간예조、라격렬동간예조(라격렬동조)화라격렬동연합GW9662간예조(연합간예조),대조조PBMCs불가임하약물간예.채용실시형광정량PCR검측PBMCs중PPAR-γ화핵인자-κB적mRNA표체,세포면역형광법결합격광소묘공취초현미경검측PPAR-γ화핵인자-κB단백표체급핵전위,매련면역흡부시험검측세포배양상청액중TNF-α함량.다조간비교채용방차분석,조간량량비교채용LSD-t검험,상관성분석채용Pearson검험.결과 mRNA(2-ΔΔCt치)화단백표체(형광강도치):비간예조PPAR-γ(0.52±0.10화55±11)저우대조조(1화85±9),핵인자-κB(1.69±0.07화145±17)고우대조조(1화118±7)；라격렬동조PPAR-γ(4.47±0.11화204±12)고우비간예조,핵인자-κB(0.33±0.04화59±14)저우비간예조；연합간예조PPAR-γ(2.25±0.31화142±23)저우라격렬동조,고우비간예조,핵인자-κB(0.64±0.02화90±10)고우라격렬동조,저우비간예조；조간비교차이균유통계학의의(F치위29.21～567.42,균P＜0.01).TNF-α농도(μg/L):비간예조(96.2±1.4)고우대조조(85.3±1.0),라격렬동조(63.0±2.5)저우비간예조,연합간예조(83.3±1.9)고우라격렬동조,저우비간예조,조간비교차이유통계학의의(F치위293.72,P＜0.01).비간예조PPAR-γ단백주요위우세포질,핵인자-kB단백주요위우세포핵；대조조PPAR-γ화핵인자-κB단백재PBMCs세포질화세포핵중균유표체；라격렬동조PPAR-γ단백전위우세포핵,핵인자-kB단백전위우세포질；연합간예조PPAR-γ단백전회세포질,핵인자-κB단백부분전이지세포핵.상관분석결과표명,PPAR-γ단백표체여핵인자-κB단백표체급TNF-α농도균정부상관(r치분별위-0.935화-0.924,균P＜0.01),핵인자-κB단백표체여TNF-α적농도정정상관(r=0.846,P＜0.01).결론 COPD적염증가능여PPAR-γ표체급활성불족유관,라격렬동능통과상조PPAR-γ표체화활성억제핵인자-kB,진이억제TNF-α분비,재COPD염증중기착중요작용.
Objective To explore the effects of rosiglitazone on peroxisome proliferator activated receptor-γ (PPAR-γ),nuclear factor-κB and tumor necrosis factor-α (TNF-t) in patients with chronic obstructive pulmonary disease (COPD).Methods From Apr.2010 to Nov.2010,30 patients with acute exacerbations of COPD,22 males and 8 females,age 54 - 87 ( mean 72 ± 9) years and 24 healthy controls,18 males and 6 females,age 52 - 80 ( mean 69 ± 10 ) years were included.The peripheral blood mononuclear cells (PBMCs) were isolated from venous blood and then cultured.On the basis of the treatment given,the PBMCs of COPD patients were divided into 3 groups:non-treatment group,rosiglitazone treatment group (rosiglitazone group) and rosiglitazone and GW9662 treatment group (combined treatment group).Cells from the healthy controls (control group) did not receive any drug treatment.The mRNA expression of PPAR-γ and NF-κB was measured with real-time PCR.The protein expression and nuclear translocation of PPAR-γ and NF-κB were detected using immunofluorescence with laser scanning confocal microscopy.The TNF-α level in culture supernatant was measured with ELISA.One-way ANOVA and LSD-t test and the Pearson correlation coefficient were used for statistical analysis.Results The mRNA and protein levels of PPAR-γwere lower in the non-treatment group (0.52 ± 0.10,55 ± 11 ) than those in the control group (1,85 ±9),while the levels of NF-κB mRNA and protein were higher in the non-treatment group ( 1.69 ±0.07,145 ± 17) than those in the control group ( 1,118 ±7).The mRNA and protein levels of PPAR-γin the rosiglitazone group (4.47 ± 0.11,204 ± 12 ) were significantly increased compared with the non-treatment group,while the mRNA and protein levels of NF-κB (0.33 ± 0.04,59 ± 14 ) were remarkably decreased compared with the non-treatment group.The mRNA and protein levels of PPAR-γ (2.25 ±0.31,142 ± 23 ) were significantly decreased in the combined treatment group compared to the rosiglitazone group,but higher compared with the non-treatment group,while the mRNA and protein levels of NF-κB (0.64 ± 0.02,90 ± 10) were increased compared with the rosiglitazone group,but decreased compared to the non-treatment group ( F =29.21 - 567.42,all P ＜ 0.01 ).The TNF-α level was significantly higher in the non-treatment group (96.2 ± 1.4) μg/L than that in the control group ( 85.3 ± 1.0) μg/L.The TNF-α level in the rosiglitazone group (63.0 ± 2.5) μg/L was remarkably decreased compared with the non-treatment group,while that in the combined treatment group (83.3 ± 1.9) μg/L was increased compared with the rosiglitazone group,but decreased compared to the non-treatment group ( F =293.72,P ＜ 0.01 ).The proteins of PPAR-γ and NF-κB were respectively located in cytoplasm and in nucleus in the non-treatment group,meanwhile they were located in both cytoplasm and nucleus in the control group.PPAR-γ protein was translocated from cytoplasm into nucleus and NF-κB protein was transiocated from nucleus into cytoplasm in the rosiglitazone group.In the combined treatment group,PPAR-γ protein translocated from nucleus into cytoplasm and NF-κB protein partly translocated from cytoplasm into nucleus.By linear correlation analysis,PPAR-γ protein was negatively correlated with NF-κB protein and TNF-α level ( r =- 0.935,- 0.924,all P ＜ 0.01 ),while NF-κB protein was positively correlated to TNF-α level (r =0.846,P ＜0.01 ).Conclusions The expression and activity of PPAR-γ were decreased in COPD patients.PPAR-γ agonist rosiglitazone inhibited inflammation in COPD through upregulating the expression and activity of PPAR-γ and inhibition of NF-κB and TNF-α.It suggests that PPAR-γ may play an important role in the inflammation of COPD.
