中华急诊医学杂志
中華急診醫學雜誌
중화급진의학잡지
CHINESE JOURNAL OF EMERGENCY MEDICINE
2010年
5期
482-487
,共6页
周红梅%尤文军%张晓进%丁正年%程蕴琳%刘莉
週紅梅%尤文軍%張曉進%丁正年%程蘊琳%劉莉
주홍매%우문군%장효진%정정년%정온림%류리
内毒素血症%心功能不全%热休克蛋白27%P13K/Akt信号通路%炎症%转基因鼠%细胞凋亡%组织学
內毒素血癥%心功能不全%熱休剋蛋白27%P13K/Akt信號通路%炎癥%轉基因鼠%細胞凋亡%組織學
내독소혈증%심공능불전%열휴극단백27%P13K/Akt신호통로%염증%전기인서%세포조망%조직학
Endotoxemia%Cardiac dysfunction%Heat shock protein 27%PI3K/Akt signaling pathway%Inflammation%Transgenic mice%Apoptosis%Histology
目的 探讨Hsp27的保护作用是否与激活P13K/Akt信号通路、减轻炎症反应有关.方法 (1)诱导小鼠内毒素血症心肌特异性高表达Hsp27转基因鼠(Hsp 27 Tg)和野生型对照鼠(WT)均腹腔注射内毒素(LPS,10ms/ks);(2)心功能测定 LPS注射后6 h,以心脏超声评价,n:6;(3)P13K/Akt信号通路活性测定 LPS注射后1 h收集心脏,以Western blot法测定Akt及Gsk-3(的磷酸化水平,n=4;(4)NF-B炎症通路活性检测 LPS注射后1 h以Western blot测定IκBα水平,n=4;同时在大鼠心肌细胞中进行相同实验,n:3;(5)心肌细胞凋亡LPS注射后24 h,以TUNEL法检测,,n=4.结果 (1)Hsp27显著改善LPS所致的心功能不全.与基础值相比,LPS处理使WT,Hsp27 Tg均出现心功能不全,但Hsp 27 Tg的心功能不全程度显著轻于WT(P<0.01或P<0.05).(2)Hsp27显著抑制LPS所致的IκBα降解.与WT比较,高表达Hsp27显著减轻了LPS诱导的小鼠心肌IκBα降解[(41.43±24.10)%vs.(72.92±9.20)%,P<0.05],培养的大鼠心肌细胞实验结果与之类似.(3)Hsp27可增强激活P13K/Akt信号通路.LPS处理后,WT和Hsp27Tg鼠心肌组织中磷酸化Akt水平分别为(3.11±0.83)和(5.13±0.73),磷酸化GSK-30水平分别为(3.19±1.04)和(5.71±1.20).与WT比较,Hsp27Tg心肌组织中的磷酸化Akt和GSK-3β水平均显著提高(P<0.05).类似结果在体外培养的细胞实验中也被证实.(4)Hsp27显著抑制LPS所致的心肌细胞凋亡.LPS处理后24 h,WT和Hsp27Tg心肌细胞凋亡率分别为(6.46±1.74)%和(2.88±0.91)%,与WT比较,心肌细胞凋亡在Hsp27Tg中被显著减轻(P<0.01).结论 高表达Hsp27对小鼠内毒素血症心功能不全有显著改善作用,其保护作用与激活P13K/Akt信号通路、抑制NFκB;依赖性炎症反应有关.
目的 探討Hsp27的保護作用是否與激活P13K/Akt信號通路、減輕炎癥反應有關.方法 (1)誘導小鼠內毒素血癥心肌特異性高錶達Hsp27轉基因鼠(Hsp 27 Tg)和野生型對照鼠(WT)均腹腔註射內毒素(LPS,10ms/ks);(2)心功能測定 LPS註射後6 h,以心髒超聲評價,n:6;(3)P13K/Akt信號通路活性測定 LPS註射後1 h收集心髒,以Western blot法測定Akt及Gsk-3(的燐痠化水平,n=4;(4)NF-B炎癥通路活性檢測 LPS註射後1 h以Western blot測定IκBα水平,n=4;同時在大鼠心肌細胞中進行相同實驗,n:3;(5)心肌細胞凋亡LPS註射後24 h,以TUNEL法檢測,,n=4.結果 (1)Hsp27顯著改善LPS所緻的心功能不全.與基礎值相比,LPS處理使WT,Hsp27 Tg均齣現心功能不全,但Hsp 27 Tg的心功能不全程度顯著輕于WT(P<0.01或P<0.05).(2)Hsp27顯著抑製LPS所緻的IκBα降解.與WT比較,高錶達Hsp27顯著減輕瞭LPS誘導的小鼠心肌IκBα降解[(41.43±24.10)%vs.(72.92±9.20)%,P<0.05],培養的大鼠心肌細胞實驗結果與之類似.(3)Hsp27可增彊激活P13K/Akt信號通路.LPS處理後,WT和Hsp27Tg鼠心肌組織中燐痠化Akt水平分彆為(3.11±0.83)和(5.13±0.73),燐痠化GSK-30水平分彆為(3.19±1.04)和(5.71±1.20).與WT比較,Hsp27Tg心肌組織中的燐痠化Akt和GSK-3β水平均顯著提高(P<0.05).類似結果在體外培養的細胞實驗中也被證實.(4)Hsp27顯著抑製LPS所緻的心肌細胞凋亡.LPS處理後24 h,WT和Hsp27Tg心肌細胞凋亡率分彆為(6.46±1.74)%和(2.88±0.91)%,與WT比較,心肌細胞凋亡在Hsp27Tg中被顯著減輕(P<0.01).結論 高錶達Hsp27對小鼠內毒素血癥心功能不全有顯著改善作用,其保護作用與激活P13K/Akt信號通路、抑製NFκB;依賴性炎癥反應有關.
목적 탐토Hsp27적보호작용시부여격활P13K/Akt신호통로、감경염증반응유관.방법 (1)유도소서내독소혈증심기특이성고표체Hsp27전기인서(Hsp 27 Tg)화야생형대조서(WT)균복강주사내독소(LPS,10ms/ks);(2)심공능측정 LPS주사후6 h,이심장초성평개,n:6;(3)P13K/Akt신호통로활성측정 LPS주사후1 h수집심장,이Western blot법측정Akt급Gsk-3(적린산화수평,n=4;(4)NF-B염증통로활성검측 LPS주사후1 h이Western blot측정IκBα수평,n=4;동시재대서심기세포중진행상동실험,n:3;(5)심기세포조망LPS주사후24 h,이TUNEL법검측,,n=4.결과 (1)Hsp27현저개선LPS소치적심공능불전.여기출치상비,LPS처리사WT,Hsp27 Tg균출현심공능불전,단Hsp 27 Tg적심공능불전정도현저경우WT(P<0.01혹P<0.05).(2)Hsp27현저억제LPS소치적IκBα강해.여WT비교,고표체Hsp27현저감경료LPS유도적소서심기IκBα강해[(41.43±24.10)%vs.(72.92±9.20)%,P<0.05],배양적대서심기세포실험결과여지유사.(3)Hsp27가증강격활P13K/Akt신호통로.LPS처리후,WT화Hsp27Tg서심기조직중린산화Akt수평분별위(3.11±0.83)화(5.13±0.73),린산화GSK-30수평분별위(3.19±1.04)화(5.71±1.20).여WT비교,Hsp27Tg심기조직중적린산화Akt화GSK-3β수평균현저제고(P<0.05).유사결과재체외배양적세포실험중야피증실.(4)Hsp27현저억제LPS소치적심기세포조망.LPS처리후24 h,WT화Hsp27Tg심기세포조망솔분별위(6.46±1.74)%화(2.88±0.91)%,여WT비교,심기세포조망재Hsp27Tg중피현저감경(P<0.01).결론 고표체Hsp27대소서내독소혈증심공능불전유현저개선작용,기보호작용여격활P13K/Akt신호통로、억제NFκB;의뢰성염증반응유관.
Objective To investigate the cardiac protection of Hsp27 against endotoxic cardiac depression mediated by activation of PI3K/Akt pathway and the suppression of NFκB-mediated inflammatory response in mice. Method (1) Transgenic mice with cardiac specific overexpression of Hsp27 (Hsp27 Tg) and wild littermate controls (WT) were given 10 mg/kg LPS injected intraperitoneally to induce endotoxemia, (2) The cardiac function measurement in mice was performed by using echocardiography 6 hours after LPS treatment (n = 6), (3) The activity of PBK/Akt pathway was evaluated by Western blot for [hosphor-Akt (p-Akt) and phosphor-Gsk-3β (p-Gsk-3β) one hour after LPS administration ( n = 4)], (4) Activity of inflammatory response was evaluated by protein degradation of IκBα (n = 4), (5) The apoptosis of myocardial cells was determined by TUNEL assay on the paraffin section of cardiac tissue 24 hours after LPS exposure (n = 4). Results (1) Hsp27 attenuated cardiac dysfunction significantly following LPS treatment. Compared with the primary value, LPS induced the depression of cardiac function both in WT rats and Hsp27Tg rats. However, the cardiac dysfunction was attenuated significantly in Hsp27Tg rats compared with that in WT rats ( P < 0.01 or 0.05) . (2) Hsp27 attenuated IκBα degradation after LPS administration. Compared with the primary value, LPS led to LκBα degradation by (72.92 + 9.20) % in WT rats and by (41.43 + 24.10) % in Hsp27Tg rats. The overexpression of Hsp27 lessened the IκBα degradation significantly (P < 0.05). The similar results were obtained in rat myocardial cell culture of experiments. (3) Hsp27 enhanced the activation of PI3K/Akt signaling following LPS exposure. One hour after LPS administration, the relative levels of p-Akt and p-GSK-30 were (3.11 + 0.83) and (3.19 + 1.04), respectively in WT rats, and (5.13 + 0.73) and (5.71 + 1.20) in Hsp27Tg rats, respectively. Compared with WT rats, the levels of p-Akt and p-GSK-3β were significantly higher in Hsp27Tg rats (P < 0.05). (4) The Hsp27 lessened LPS-induced the apopto-sis of myocardial cells. Twenty-four hours after LPS treatment, the percentages of myocardial cell apoptosis were (6.46+ 1.74)% in WT rats and (2.88 + 0.91)% in Hsp27Tg rats. Compared with WT rats, LPS-induced apoptosis in myocardial cells was significantly decreased in Hsp27Tg rats (P < 0.01). Conclusions The overexpression ofHsp27 attenuates cardiac dysfunction significantly during endotoxemia, and the mechanisms may be attributed to the activation of PDK/Akt signaling pathway.
