中国危重病急救医学
中國危重病急救醫學
중국위중병급구의학
CHINESE CRITICAL CARE MEDICINE
2013年
3期
154-158
,共5页
小窝蛋白-1%内皮型一氧化氮合酶%白细胞介素-1受体相关激酶4%核转录因子-κB%机械通气
小窩蛋白-1%內皮型一氧化氮閤酶%白細胞介素-1受體相關激酶4%覈轉錄因子-κB%機械通氣
소와단백-1%내피형일양화담합매%백세포개소-1수체상관격매4%핵전록인자-κB%궤계통기
Caveolin-1%Endothelial nitric oxide synthase%Interleukin-1 receptor-associated kinase 4%Nuclear factor-κB%Mechanical ventilation
目的 研究小窝蛋白-1(cav-1)及其相关信号链酶在不同潮气量(VT)机械通气大鼠肺组织中的表达变化.方法 按随机数字表法将SD大鼠分为5组,每组8只.对照组(A组)仅切开气管保留自主呼吸；保护性机械通气1h、2h组(B1组、B2组)VT为6 ml/kg;大VT通气1h、2h组(C1组、C2组)VT为30 ml/kg.A组在气管切开即刻,通气组分别于通气1h或2h末处死大鼠,光镜下观察肺组织病理学改变；逆转录-聚合酶链反应(RT-PCR)检测cav-1、eNOS的mRNA表达；免疫组化法检测肺组织中cav-1、内皮型一氧化氮合酶(eNOS)、白细胞介素-1受体相关激酶4(IRAK4)及核转录因子-κB(NF-κB)p65的蛋白表达；比色法测定髓过氧化物酶(MPO)活性；计算肺湿/干质量比值(W/D)；酶联免疫吸附试验(ELISA)测定支气管肺泡灌洗液(BALF)中肿瘤坏死因子-α(TNF-α)含量.结果 A组、B1组、B2组之间肺组织cav-1、eNOS的mRNA表达,cav-1、eNOS、IRAK4、NF-κBp65的蛋白表达,肺W/D比值,MPO和TNF-α水平差异均无统计学意义.与相应B1、B2组比较,C1、C2组cav-1 mRNA(A值比值)和cav-1、IRAK4、NF-κBp65蛋白表达(A值)明显增加(cav-1 mRNA:0.833±0.085比0.384 ±0.011,1.162±0.166比0.388±0.014;cav-1蛋白:0.188±0.011比0.140±0.052,0.210±0.013比0.125±0.014;IRAK4蛋白:0.181 ±0.009比0.150±0.008,0.205±0.085比0.155 ±0.012;NF-κBp65蛋白:0.294±0.011比0.236±0.015,0.304±0.012比0.239±0.005),eNOS的mRNA(A值比值)和蛋白表达(A值)明显减少(eNOS mRNA:0.174±0.016比0.278=0.021,0.107±0.014比0.262±0.045；eNOS蛋白:0.180±0.017比0.211±0.010,0.161±0.016比0.216±0.013),肺W/D比值、MPO活性(U/g)和TNF-α含量(ng/L)明显升高(W/D:5.64±0.42比4.63±0.12,6.73±0.83比4.70±0.15;MPO:1.86±0.26比0.85±0.11,2.14±0.24比0.88±0.18; TNF-α:386.53±29.12比50.57±10.98,455.77±37.78比52.11±9.92),差异均有统计学意义(均P＜0.05).随通气时间延长,C2组各指标较C1组改变更为明显(均P＜0.05).结论 cav-1及其相关信号链酶在机械通气中的表达参与了呼吸机相关性肺损伤.
目的 研究小窩蛋白-1(cav-1)及其相關信號鏈酶在不同潮氣量(VT)機械通氣大鼠肺組織中的錶達變化.方法 按隨機數字錶法將SD大鼠分為5組,每組8隻.對照組(A組)僅切開氣管保留自主呼吸；保護性機械通氣1h、2h組(B1組、B2組)VT為6 ml/kg;大VT通氣1h、2h組(C1組、C2組)VT為30 ml/kg.A組在氣管切開即刻,通氣組分彆于通氣1h或2h末處死大鼠,光鏡下觀察肺組織病理學改變；逆轉錄-聚閤酶鏈反應(RT-PCR)檢測cav-1、eNOS的mRNA錶達；免疫組化法檢測肺組織中cav-1、內皮型一氧化氮閤酶(eNOS)、白細胞介素-1受體相關激酶4(IRAK4)及覈轉錄因子-κB(NF-κB)p65的蛋白錶達；比色法測定髓過氧化物酶(MPO)活性；計算肺濕/榦質量比值(W/D)；酶聯免疫吸附試驗(ELISA)測定支氣管肺泡灌洗液(BALF)中腫瘤壞死因子-α(TNF-α)含量.結果 A組、B1組、B2組之間肺組織cav-1、eNOS的mRNA錶達,cav-1、eNOS、IRAK4、NF-κBp65的蛋白錶達,肺W/D比值,MPO和TNF-α水平差異均無統計學意義.與相應B1、B2組比較,C1、C2組cav-1 mRNA(A值比值)和cav-1、IRAK4、NF-κBp65蛋白錶達(A值)明顯增加(cav-1 mRNA:0.833±0.085比0.384 ±0.011,1.162±0.166比0.388±0.014;cav-1蛋白:0.188±0.011比0.140±0.052,0.210±0.013比0.125±0.014;IRAK4蛋白:0.181 ±0.009比0.150±0.008,0.205±0.085比0.155 ±0.012;NF-κBp65蛋白:0.294±0.011比0.236±0.015,0.304±0.012比0.239±0.005),eNOS的mRNA(A值比值)和蛋白錶達(A值)明顯減少(eNOS mRNA:0.174±0.016比0.278=0.021,0.107±0.014比0.262±0.045；eNOS蛋白:0.180±0.017比0.211±0.010,0.161±0.016比0.216±0.013),肺W/D比值、MPO活性(U/g)和TNF-α含量(ng/L)明顯升高(W/D:5.64±0.42比4.63±0.12,6.73±0.83比4.70±0.15;MPO:1.86±0.26比0.85±0.11,2.14±0.24比0.88±0.18; TNF-α:386.53±29.12比50.57±10.98,455.77±37.78比52.11±9.92),差異均有統計學意義(均P＜0.05).隨通氣時間延長,C2組各指標較C1組改變更為明顯(均P＜0.05).結論 cav-1及其相關信號鏈酶在機械通氣中的錶達參與瞭呼吸機相關性肺損傷.
목적 연구소와단백-1(cav-1)급기상관신호련매재불동조기량(VT)궤계통기대서폐조직중적표체변화.방법 안수궤수자표법장SD대서분위5조,매조8지.대조조(A조)부절개기관보류자주호흡；보호성궤계통기1h、2h조(B1조、B2조)VT위6 ml/kg;대VT통기1h、2h조(C1조、C2조)VT위30 ml/kg.A조재기관절개즉각,통기조분별우통기1h혹2h말처사대서,광경하관찰폐조직병이학개변；역전록-취합매련반응(RT-PCR)검측cav-1、eNOS적mRNA표체；면역조화법검측폐조직중cav-1、내피형일양화담합매(eNOS)、백세포개소-1수체상관격매4(IRAK4)급핵전록인자-κB(NF-κB)p65적단백표체；비색법측정수과양화물매(MPO)활성；계산폐습/간질량비치(W/D)；매련면역흡부시험(ELISA)측정지기관폐포관세액(BALF)중종류배사인자-α(TNF-α)함량.결과 A조、B1조、B2조지간폐조직cav-1、eNOS적mRNA표체,cav-1、eNOS、IRAK4、NF-κBp65적단백표체,폐W/D비치,MPO화TNF-α수평차이균무통계학의의.여상응B1、B2조비교,C1、C2조cav-1 mRNA(A치비치)화cav-1、IRAK4、NF-κBp65단백표체(A치)명현증가(cav-1 mRNA:0.833±0.085비0.384 ±0.011,1.162±0.166비0.388±0.014;cav-1단백:0.188±0.011비0.140±0.052,0.210±0.013비0.125±0.014;IRAK4단백:0.181 ±0.009비0.150±0.008,0.205±0.085비0.155 ±0.012;NF-κBp65단백:0.294±0.011비0.236±0.015,0.304±0.012비0.239±0.005),eNOS적mRNA(A치비치)화단백표체(A치)명현감소(eNOS mRNA:0.174±0.016비0.278=0.021,0.107±0.014비0.262±0.045；eNOS단백:0.180±0.017비0.211±0.010,0.161±0.016비0.216±0.013),폐W/D비치、MPO활성(U/g)화TNF-α함량(ng/L)명현승고(W/D:5.64±0.42비4.63±0.12,6.73±0.83비4.70±0.15;MPO:1.86±0.26비0.85±0.11,2.14±0.24비0.88±0.18; TNF-α:386.53±29.12비50.57±10.98,455.77±37.78비52.11±9.92),차이균유통계학의의(균P＜0.05).수통기시간연장,C2조각지표교C1조개변경위명현(균P＜0.05).결론 cav-1급기상관신호련매재궤계통기중적표체삼여료호흡궤상관성폐손상.
Objective To investigate the expression of caveolin-1 (cav-1) and its downstream signal under mechanical ventilation with different tidal volumes (VT) in lung tissue of rats.Methods Forty healthy male Sprague-Dawley (SD) rats were randomly assigned into five groups (each n=8).The rats in control group (group A) remained to have spontaneous breathing but underwent tracheostomy only.The rats in protective ventilation group underwent protective ventilation for 1 hour or 2 hours (group B1,B2),with VT set at 6 ml/kg.The rats in high Vr ventilation group were given mechanical ventilation for 1 hour or 2 hours (group C1,C2),with VT set at 30 ml/kg.After incision of trachea in group A,and mechanical ventilation was given for 1 hour or 2 hours in ventilation groups.Rats of group A were sacrificed immediately.Rats of other groups were sacrificed 1 hour or 2 hours after mechanical ventilation.Specimens of lung tissues and bronchoalveolar lavage fluid (BALF) were harvested.Lung pathological changes were observed with optical microscope.The expression levels of cav-1 mRNA and eNOS mRNA in lung tissue were measured by reverse transcription-polymerase chain reaction (RT-PCR).The protein levels of car-1,endothelial nitric oxide synthase (eNOS),interleukin-1 receptor-associated kinase 4 (IRAK4) and nuclear factor-κB (NF-κB) p65 in lung tissues were assayed with immunohistochemistry staining.Lung myeloperoxidase (MPO) activity was measured by colorimetric analysis,and wet/dry weight ratio (W/D) was calculated.The levels of tumor necrosis factor-α (TNF-α) in BALF was measured by enzyme-linked immunosorbent assay (ELISA).Results No statistical significance in the mRNA expression of cav-1 and eNOS,the protein expression of cav-1,eNOS,IRAK4,NF-κBp65,as well as W/D ratio,MPO and TNF-α in BALF was found among group A,group B1 and group B2.The expression of car-1 mRNA (A value ratio) and cav-1,IRAK4,NF-κBp65 protein (A value) were significantly up-regulated (car-1 mRNA:0.833 ± 0.085 vs.0.384 ± 0.011,1.162 ± 0.166 vs.0.388 ± 0.014; cav-1 protein:0.188 ± 0.011 vs.0.140 ± 0.052,0.210 ± 0.013 vs.0.125 ± 0.014; IRAK4 protein:0.181 ± 0.009 vs.0.150 ± 0.008,0.205 ± 0.085 vs.0.155 ± 0.012;NF-κBp65 protein:0.294 ± 0.011 vs.0.236 ± 0.015,0.304 ± 0.012 vs.0.239 ± 0.005),the expression of eNOS mRNA (A value ratio) and protein (A value) was significantly down-regulated (eNOS mRNA:0.174 ± 0.016 vs.0.278 ± 0.021,0.107 ± 0.014 vs.0.262 ± 0.045; eNOS protein:0.180 ± 0.017 vs.0.211 ± 0.010,0.161 ± 0.016 vs.0.216 ± 0.013),while W/D ratio,MPO (U/g) and TNF-α (ng/L) in BALF were significantly increased (W/D:5.64 ± 0.42 vs.4.63 ±0.12,6.73 ±0.83 vs.4.70 ±0.15; MPO:1.86 ±0.26 vs.0.85 ±0.11,2.14 ±0.24 vs.0.88 ±0.18;TNF-α:386.53 ± 29.12 vs.50.57 ± 10.98,455.77 ± 37.78 vs.52.11 ± 9.92) in group C1 and group C2 compared with those in group B1 and group B2 (all P＜0.05).With prolongation of time of mechanical ventilation,changes in those parameters were more obvious in group C2 as compared with group C1 (all P＜0.05).Conclusion Cav-1 and the activation of downstream signals in lung tissue participate in the development of the ventilator-induced lung injury.
