中华实验外科杂志
中華實驗外科雜誌
중화실험외과잡지
CHINESE JOURNAL OF EXPERIMENTAL SURGERY
2015年
4期
805-807
,共3页
王伟涛%曹胜利%方红波%郭文治%李捷%阎冰%张水军
王偉濤%曹勝利%方紅波%郭文治%李捷%閻冰%張水軍
왕위도%조성리%방홍파%곽문치%리첩%염빙%장수군
供体心脏%脑死亡%脱噬作用%c-Jun氨基末端激酶抑制剂
供體心髒%腦死亡%脫噬作用%c-Jun氨基末耑激酶抑製劑
공체심장%뇌사망%탈서작용%c-Jun안기말단격매억제제
Donor heart%Brain death%Apoptosis%c-Jun N-terminal kinase inhibitor
目的 观察c-Jun氨基末端激酶(JNK)信号传导通路特异阻断剂SP600125对脑死亡状态下大鼠心肌细胞凋亡的保护作用.方法 将20只健康雄性大鼠随机分为4组,每组5只.空白对照(Sham)组:仅给予颅内置管,不加压诱导脑死亡;脑死亡(BD)组:仅诱导大鼠脑死亡并维持脑死亡6 h;SP600125组:在诱导脑死亡前1h腹腔注射SP600125(10 mg/kg),并维持脑死亡6h;二甲基亚砜(DMSO)组:在诱导脑死亡前1h腹腔注射DMSO(10 mg/kg),并维持脑死亡6h.应用实时定量聚合酶链反应(Real-time PCR)和Western blot检测各组心肌细胞中半胱氨酸天冬氨酸特异性蛋白酶-3(Caspase-3)与细胞色素C(Cyt-C)mRNA和蛋白的表达,并用原位缺口末端标记法(TUNEL)染色比较各组心肌细胞凋亡.结果 (1)Real-time PCR结果显示BD组Caspase-3mRNA和Cyt-C mRNA表达(2.37 ±0.12、2.03±0.08)显著高于Sham组(P ＜0.05);BD组Caspase-3 mRNA和Cyt-C mRNA表达与DMSO组(2.38±0.11、2.06±0.09)接近(P＞0.05);SP600125组Caspase-3mRNA和Cyt-C mRNA表达(1.21±0.05、1.23±0.24)显著低于BD组(P＜0.05).(2)Western blot结果显示,BD组Caspae-3和Cyt-C表达(145.63±7.21、73.67±7.35)比Sham组(38.86±8.95、18.74±8.77)显著增多(P＜0.05);BD组Caspase-3和Cyt-C表达与DMSO组(146.56±6.87、72.45±6.84)接近(P ＞0.05);SP600125组Caspase-3和Cyt-C表达(39.45±5.73、20.34±5.69)显著低于BD组(P＜0.05).(3) TUNEL法结果显示,BD组心肌细胞凋亡率[(26.39±4.99)％]比Sham组[(1.41±0.35)％]显著增加(P＜0.05),DMSO组[(26.28±4.92)％]与Sham组凋亡率接近(P＞0.05),SP600125组[(7.63±3.09)％]比BD组显著降低(P＜0.05).结论 SP600125通过特异性地抑制JNK活性显著地减轻脑死亡状态下大鼠心肌细胞凋亡.
目的 觀察c-Jun氨基末耑激酶(JNK)信號傳導通路特異阻斷劑SP600125對腦死亡狀態下大鼠心肌細胞凋亡的保護作用.方法 將20隻健康雄性大鼠隨機分為4組,每組5隻.空白對照(Sham)組:僅給予顱內置管,不加壓誘導腦死亡;腦死亡(BD)組:僅誘導大鼠腦死亡併維持腦死亡6 h;SP600125組:在誘導腦死亡前1h腹腔註射SP600125(10 mg/kg),併維持腦死亡6h;二甲基亞砜(DMSO)組:在誘導腦死亡前1h腹腔註射DMSO(10 mg/kg),併維持腦死亡6h.應用實時定量聚閤酶鏈反應(Real-time PCR)和Western blot檢測各組心肌細胞中半胱氨痠天鼕氨痠特異性蛋白酶-3(Caspase-3)與細胞色素C(Cyt-C)mRNA和蛋白的錶達,併用原位缺口末耑標記法(TUNEL)染色比較各組心肌細胞凋亡.結果 (1)Real-time PCR結果顯示BD組Caspase-3mRNA和Cyt-C mRNA錶達(2.37 ±0.12、2.03±0.08)顯著高于Sham組(P ＜0.05);BD組Caspase-3 mRNA和Cyt-C mRNA錶達與DMSO組(2.38±0.11、2.06±0.09)接近(P＞0.05);SP600125組Caspase-3mRNA和Cyt-C mRNA錶達(1.21±0.05、1.23±0.24)顯著低于BD組(P＜0.05).(2)Western blot結果顯示,BD組Caspae-3和Cyt-C錶達(145.63±7.21、73.67±7.35)比Sham組(38.86±8.95、18.74±8.77)顯著增多(P＜0.05);BD組Caspase-3和Cyt-C錶達與DMSO組(146.56±6.87、72.45±6.84)接近(P ＞0.05);SP600125組Caspase-3和Cyt-C錶達(39.45±5.73、20.34±5.69)顯著低于BD組(P＜0.05).(3) TUNEL法結果顯示,BD組心肌細胞凋亡率[(26.39±4.99)％]比Sham組[(1.41±0.35)％]顯著增加(P＜0.05),DMSO組[(26.28±4.92)％]與Sham組凋亡率接近(P＞0.05),SP600125組[(7.63±3.09)％]比BD組顯著降低(P＜0.05).結論 SP600125通過特異性地抑製JNK活性顯著地減輕腦死亡狀態下大鼠心肌細胞凋亡.
목적 관찰c-Jun안기말단격매(JNK)신호전도통로특이조단제SP600125대뇌사망상태하대서심기세포조망적보호작용.방법 장20지건강웅성대서수궤분위4조,매조5지.공백대조(Sham)조:부급여로내치관,불가압유도뇌사망;뇌사망(BD)조:부유도대서뇌사망병유지뇌사망6 h;SP600125조:재유도뇌사망전1h복강주사SP600125(10 mg/kg),병유지뇌사망6h;이갑기아풍(DMSO)조:재유도뇌사망전1h복강주사DMSO(10 mg/kg),병유지뇌사망6h.응용실시정량취합매련반응(Real-time PCR)화Western blot검측각조심기세포중반광안산천동안산특이성단백매-3(Caspase-3)여세포색소C(Cyt-C)mRNA화단백적표체,병용원위결구말단표기법(TUNEL)염색비교각조심기세포조망.결과 (1)Real-time PCR결과현시BD조Caspase-3mRNA화Cyt-C mRNA표체(2.37 ±0.12、2.03±0.08)현저고우Sham조(P ＜0.05);BD조Caspase-3 mRNA화Cyt-C mRNA표체여DMSO조(2.38±0.11、2.06±0.09)접근(P＞0.05);SP600125조Caspase-3mRNA화Cyt-C mRNA표체(1.21±0.05、1.23±0.24)현저저우BD조(P＜0.05).(2)Western blot결과현시,BD조Caspae-3화Cyt-C표체(145.63±7.21、73.67±7.35)비Sham조(38.86±8.95、18.74±8.77)현저증다(P＜0.05);BD조Caspase-3화Cyt-C표체여DMSO조(146.56±6.87、72.45±6.84)접근(P ＞0.05);SP600125조Caspase-3화Cyt-C표체(39.45±5.73、20.34±5.69)현저저우BD조(P＜0.05).(3) TUNEL법결과현시,BD조심기세포조망솔[(26.39±4.99)％]비Sham조[(1.41±0.35)％]현저증가(P＜0.05),DMSO조[(26.28±4.92)％]여Sham조조망솔접근(P＞0.05),SP600125조[(7.63±3.09)％]비BD조현저강저(P＜0.05).결론 SP600125통과특이성지억제JNK활성현저지감경뇌사망상태하대서심기세포조망.
Objective To investigate the protective role of SP600125,a selective c-Jun N-terminal kinase inhibitor,in the apoptosis of cardiomyocytes in brain death rats.Methods Twenty Sprague-Dawley (SD) rats were randomly divided into:Sham group (n =5),only given intracranial insertion of Fogarty tube after anesthesia; brain-death group (n =5),given brain death for 6 h ; SP600125 group (n =5),given peritoneal injection of SP6600125 (10 mg/kg) 1 h before brain death for 6 h; and DMSO group (n =5),given peritoneal injection of placebo solution DMSO (10 mg/kg) 1 h before brain death for 6 h.At 6 h,the apical myocardial samples were obtained for detection of Cysteinyl aspartate-specific protease (Caspase)-3 and cytochrome-C (Cyt-C) mRNA and protein expression by real-time quantitative polymerase chain reaction (real-time PCR) and Western blotting respectively.Apoptosis of cardiac muscle cells was determined by terminal deoxynucleotidyl transferase dUTP nick end labelling (TUNEL) assay.Results (1) Real-time PCR showed that the mRNA expression of Caspase-3 and Cyt-C mRNA in brain death group was (2.37 ±0.12,2.03 ±0.08) times higher than that in sham group (P ＜0.05).There was no significant difference in the mRNA expression of Caspase-3 and Cyt-C between brain death group and DMSO group (2.38 ± 0.11 and 2.06 ± 0.09) (P ＞ 0.05).The mRNA expression of Caspase-3 and Cyt-C in SP600125 group (1.21 ±0.05 and 1.23 ±0.24) was significantly lower than that in brain death group (2.38 ± 0.11 and 2.06 ± 0.09) (P ＜ 0.05).(2) Western blotting revealed that the expression of Caspase-3 and Cyt-C in brain death group (145.63 ± 7.21 and 73.67 ± 7.35) was increased significantly as compared with that in Sham group (38.86 ± 8.95 and 18.74 ± 8.77,P ＜ 0.05).The expression of Caspase-3 and Cyt-C in brain death group was similar to that in DMSO group (146.56 t 6.87 and 72.45 ±6.84,P＞0.05),and that in SP600125 group (39.45 ±5.73 and 20.34 ±5.69) was decreased significantly as compared with that in brain death group (P ＜0.05).(3) TUNEL figured out that the apoptosis rate of cardiomyocytes in brain death group was (26.39 ± 4.99) ％,significantly higher than in Sham group [(1.41 ± 0.35) ％,P ＜ 0.05],that in brain death group was similar to that in DMSO group [(26.28 ± 4.92) ％,P ＞ 0.05],and that in SP600125 group [(7.63 ± 3.09) ％] was significantly reduced as compared with that in brain death group (P ＜ 0.05).Conclusion SP600125 played protective roles in the apoptosis of cardiomyocytes in brain death rats through specifically inhibiting the activity of JNK signaling pathway.
