中国全科医学
中國全科醫學
중국전과의학
CHINESE GENERAL PRACTICE
2014年
27期
3211-3216
,共6页
戚迪%何婧%叶媛%冯龙华%王导新
慼迪%何婧%葉媛%馮龍華%王導新
척적%하청%협원%풍룡화%왕도신
呼吸窘迫综合征%雌二醇%上皮钠通道%信号传导%PI3K/AKT/SGK1
呼吸窘迫綜閤徵%雌二醇%上皮鈉通道%信號傳導%PI3K/AKT/SGK1
호흡군박종합정%자이순%상피납통도%신호전도%PI3K/AKT/SGK1
Respiratory distress syndrome%Estradiol%Epithelial sodium channel%Signal transduction%PI3K/ AKT/SGK1
目的：探讨17β-雌二醇介导的PI3K/AKT/SGK1信号通路对小鼠急性呼吸窘迫综合征（ ARDS）的保护作用及其对肺泡上皮钠离子通道（ENaC）的调控。方法40只雄性C57B/L6小鼠随机分为对照组（control组）、模型组（ LPS组）、17β-雌二醇治疗组（ E2组）、渥曼青霉素干预组（ wortmannin 组），每组10只。苏木素-伊红（HE）染色观察肺组织病理改变并进行肺损伤评分；血气分析动脉血氧分压（PaO2）变化；肺湿干比（W/D）检测肺水肿的严重程度；BCA法检测肺泡灌洗液（ BALF）中总蛋白含量；酶联免疫吸附试验（ ELIAS）法检测BALF中白介素（ IL）-1β、肿瘤坏死因子α（ TNF-α）水平及髓过氧化物酶（ MPO）活性；反转录聚合酶链式反应（ RT-PCR）法检测各亚基ENaC mRNA表达；Western blotting法检测AKT、SGK1磷酸化水平及各亚基ENaC蛋白表达水平变化。结果小鼠气管内注射脂多糖（LPS）造模后，LPS组肺损伤评分为（4.2±0.2）分，E2组为（2.7±0.5）分，wort-mannin组为（3.9±0.3）分，差异有统计学意义（ F越46.07，P越0.000）；其中E2组较LPS组降低，wortmannin组较E2组升高（P<0.05）。4组不同时间PaO2比较有差异（F越188.148，P越0.000）。4组小鼠肺组织W/D，BALF中总蛋白含量，IL-1β、TNF-α水平，MPO活性，α-、β-、γ-ENaC的mRNA表达水平，α-、β-、γ-ENaC蛋白表达及p-AKT、p-SGK1表达水平比较有差异（P<0.05）；其中与control组比较，LPS组、E2组、wortmannin组W/D、BALF中总蛋白含量、IL-1β、TNF -α水平、MPO 活性升高，α-、β-、γ-ENaC 的 mRNA 表达水平降低（ P <0.05）；与E2组比较，LPS组、wortmannin组W/D、BALF蛋白含量、IL-1β、TNF-α水平、MPO活性升高，α-、β-、γ-ENaC蛋白表达水平及p-AKT、p-SGK1表达水平降低（P<0.05）。结论17β-雌二醇可通过快速激活PI3K/AKT/SGK1信号通路介导的转录后调控机制上调ENaC表达，从而对小鼠ARDS肺水肿发挥保护作用。
目的：探討17β-雌二醇介導的PI3K/AKT/SGK1信號通路對小鼠急性呼吸窘迫綜閤徵（ ARDS）的保護作用及其對肺泡上皮鈉離子通道（ENaC）的調控。方法40隻雄性C57B/L6小鼠隨機分為對照組（control組）、模型組（ LPS組）、17β-雌二醇治療組（ E2組）、渥曼青黴素榦預組（ wortmannin 組），每組10隻。囌木素-伊紅（HE）染色觀察肺組織病理改變併進行肺損傷評分；血氣分析動脈血氧分壓（PaO2）變化；肺濕榦比（W/D）檢測肺水腫的嚴重程度；BCA法檢測肺泡灌洗液（ BALF）中總蛋白含量；酶聯免疫吸附試驗（ ELIAS）法檢測BALF中白介素（ IL）-1β、腫瘤壞死因子α（ TNF-α）水平及髓過氧化物酶（ MPO）活性；反轉錄聚閤酶鏈式反應（ RT-PCR）法檢測各亞基ENaC mRNA錶達；Western blotting法檢測AKT、SGK1燐痠化水平及各亞基ENaC蛋白錶達水平變化。結果小鼠氣管內註射脂多糖（LPS）造模後，LPS組肺損傷評分為（4.2±0.2）分，E2組為（2.7±0.5）分，wort-mannin組為（3.9±0.3）分，差異有統計學意義（ F越46.07，P越0.000）；其中E2組較LPS組降低，wortmannin組較E2組升高（P<0.05）。4組不同時間PaO2比較有差異（F越188.148，P越0.000）。4組小鼠肺組織W/D，BALF中總蛋白含量，IL-1β、TNF-α水平，MPO活性，α-、β-、γ-ENaC的mRNA錶達水平，α-、β-、γ-ENaC蛋白錶達及p-AKT、p-SGK1錶達水平比較有差異（P<0.05）；其中與control組比較，LPS組、E2組、wortmannin組W/D、BALF中總蛋白含量、IL-1β、TNF -α水平、MPO 活性升高，α-、β-、γ-ENaC 的 mRNA 錶達水平降低（ P <0.05）；與E2組比較，LPS組、wortmannin組W/D、BALF蛋白含量、IL-1β、TNF-α水平、MPO活性升高，α-、β-、γ-ENaC蛋白錶達水平及p-AKT、p-SGK1錶達水平降低（P<0.05）。結論17β-雌二醇可通過快速激活PI3K/AKT/SGK1信號通路介導的轉錄後調控機製上調ENaC錶達，從而對小鼠ARDS肺水腫髮揮保護作用。
목적：탐토17β-자이순개도적PI3K/AKT/SGK1신호통로대소서급성호흡군박종합정（ ARDS）적보호작용급기대폐포상피납리자통도（ENaC）적조공。방법40지웅성C57B/L6소서수궤분위대조조（control조）、모형조（ LPS조）、17β-자이순치료조（ E2조）、악만청매소간예조（ wortmannin 조），매조10지。소목소-이홍（HE）염색관찰폐조직병리개변병진행폐손상평분；혈기분석동맥혈양분압（PaO2）변화；폐습간비（W/D）검측폐수종적엄중정도；BCA법검측폐포관세액（ BALF）중총단백함량；매련면역흡부시험（ ELIAS）법검측BALF중백개소（ IL）-1β、종류배사인자α（ TNF-α）수평급수과양화물매（ MPO）활성；반전록취합매련식반응（ RT-PCR）법검측각아기ENaC mRNA표체；Western blotting법검측AKT、SGK1린산화수평급각아기ENaC단백표체수평변화。결과소서기관내주사지다당（LPS）조모후，LPS조폐손상평분위（4.2±0.2）분，E2조위（2.7±0.5）분，wort-mannin조위（3.9±0.3）분，차이유통계학의의（ F월46.07，P월0.000）；기중E2조교LPS조강저，wortmannin조교E2조승고（P<0.05）。4조불동시간PaO2비교유차이（F월188.148，P월0.000）。4조소서폐조직W/D，BALF중총단백함량，IL-1β、TNF-α수평，MPO활성，α-、β-、γ-ENaC적mRNA표체수평，α-、β-、γ-ENaC단백표체급p-AKT、p-SGK1표체수평비교유차이（P<0.05）；기중여control조비교，LPS조、E2조、wortmannin조W/D、BALF중총단백함량、IL-1β、TNF -α수평、MPO 활성승고，α-、β-、γ-ENaC 적 mRNA 표체수평강저（ P <0.05）；여E2조비교，LPS조、wortmannin조W/D、BALF단백함량、IL-1β、TNF-α수평、MPO활성승고，α-、β-、γ-ENaC단백표체수평급p-AKT、p-SGK1표체수평강저（P<0.05）。결론17β-자이순가통과쾌속격활PI3K/AKT/SGK1신호통로개도적전록후조공궤제상조ENaC표체，종이대소서ARDS폐수종발휘보호작용。
Objective Toinvestigatetheeffectandmechanismof17β-estradiol-mediatedPI3K/AKT/SGK1path-wayonepithelialsodiumchannel(ENaC)regulationinmicewithARDS.Methods 40maleC57B/L6micewererandomlydi-vided into control group,LPS group,E2 group and wortmannin group,with each group 10 mice. The pathological changes in mice lung tissue were evaluated by HE staining;Arterial oxygen tension( PaO2 )changes were measured by the blood gas analy-zer;The lung permeability was measured by W/D;The levels of total protein in BALF were measured by BCA ( bicinchoninic acid);Interleukin-1β( IL-1β),tumor necrosis factor-α( TNF-α)and myeloperoxidase( MPO)activities in the bron-choalveolar lavage fluid ( BALF ) were measured by ELISA;The mRNA transcription levels of ENaC were analyzed by RT-PCR;The protein expression levels of α-,β-,γ-ENaC,p-AKT and p-SGK1 were analyzed by Western blotting method.Results AftermodelingbyinjectingLPS,thelunginjuryscorewas(4.2±0.2)inLPSgroup,(2.7±0.5)inE2 group and(3. 9±0. 3)in wortmannin group,showing statistically significant differences(F=46. 07,P=0. 000). Among the groups,E2 group was lower than LPS group and wortmannin group was higher than E2 group(P<0. 05). The PaO2 among the four groups at different time showed statistically significant differences(F=188. 148,P=0. 000). The W/D,BALF proteinlevel,IL-1β,TNF-α,MPO activity,α-,β-,γ-ENaC mRNA transcription level,α-,β-,γ-ENaC expression and p-AKT and p-SGK1 expression levels showed statistically significant differences(P<0. 05). Compared with the control group,the W/D,BALF protein level,IL-1β,TNF-αlevel and MPO activity in E2 group,LPS group and wortmannin group were increased,while the α-,β-,γ-ENaC mRNA transcription level were decreased(P<0. 05). Compared with the E2 group,the W/D,BALF protein level,IL -1β and TNF -α levels,MPO activity,α -,β -,γ -ENaC expression and p-AKTandp-SGK1expressionlevelsweredecreased(P<0.05).℅onclusion 17β-estradiolcanup-regulateENaCvia the post-transcription mechanism involving rapid activation of PI3K/AKT/SGK1 pathway,thus playing a protective role in alve-olar fluid clearance in mice with ARDS.
