小肠结肠炎,坏死性%胞间信号肽类和蛋白质类%凋亡蛋白酶活化因子1%细胞凋亡%线粒体%大鼠
小腸結腸炎,壞死性%胞間信號肽類和蛋白質類%凋亡蛋白酶活化因子1%細胞凋亡%線粒體%大鼠
소장결장염,배사성%포간신호태류화단백질류%조망단백매활화인자1%세포조망%선립체%대서
Enterocolitis,necrotizing%Intercellular signaling peptides and proteins%Apoptotic protease-activating factor 1%Apoptosis%Mitochondria%Rats
目的 探讨肝素结合性表皮生长因子样生长因子(heparin-binding epidermal growth factor-like growth factor,HB-EGF)在新生儿坏死性小肠结肠炎(neonatal necrotizing enterocolitis,NEC)新生大鼠模型线粒体途径细胞凋亡中的作用. 方法 新生无特定病原体Sprague-Dawley大鼠随机分为3组,每组10只.NEC模型组:采用代乳品人工喂养,并给予100％氮气缺氧90 s,4℃冷刺激10 min,每天2次,连续3 d；HB-EGF干预组:在NEC模型组基础上予HB-EGF灌胃,每次800 μg/kg,每天4次,连续3d.正常对照组:鼠乳喂养3d,不予任何干预.大鼠生后72h禁食12h后处死.取回肠末端组织,HE染色观察病理改变并评分；电镜下观察线粒体超微结构变化；免疫组织化学方法检测细胞色素C含量；Western印迹技术检测凋亡诱导因子(apoptosis inducing factor,AIF)及凋亡肽酶激活因子-1(apoptotic protease activating factor-1,APAF-1)的表达.组间差异比较采用单因素方差分析,两两比较采用q检验,P＜0.05为差异有统计学意义. 结果 (1)HB-EGF干预组NEC发生率为2/10,低于模型组(9/10),差异有统计学意义(x2=7.27,P＜0.01)；对照组未发生NEC.(2)NEC模型组线粒体在肠上皮细胞及肌层细胞中存在明显肿胀,基质内有多数电子透亮区,线粒体超微结构严重损伤.HB-EGF干预组有少量线粒体肿胀,损伤较NEC模型组减轻.(3)NEC模型组回肠组织细胞色素C表达较对照组增强,差异有统计学意义(0.030±0.018与0.002±0.001,q=6.15,P＜0.01),HB-EGF干预组回肠组织细胞色素C表达(0.014±0.018)较NEC模型组减弱,差异有统计学意义(q=3.53,P＜0.05).NEC模型组的APAF-1表达较对照组增强(1.364±0.299与0.215±0.033,q=15.31,P＜0.05),AIF表达也增强(0.181±0.050与0.127±0.045,q=3.71,P＜0.05)；与NEC模型组比较,HB-EGF干预组APAF-1的表达(0.455±0.123)减低(q=4.04,P＜0.05),AIF的表达(0.289±0.045)则明显增强(q=7.32,P＜0.05). 结论 HB-EGF能降低新生大鼠NEC发生率,其机制之一可能是通过下调APAF-1表达而减少新生大鼠线粒体途径细胞凋亡.
目的 探討肝素結閤性錶皮生長因子樣生長因子(heparin-binding epidermal growth factor-like growth factor,HB-EGF)在新生兒壞死性小腸結腸炎(neonatal necrotizing enterocolitis,NEC)新生大鼠模型線粒體途徑細胞凋亡中的作用. 方法 新生無特定病原體Sprague-Dawley大鼠隨機分為3組,每組10隻.NEC模型組:採用代乳品人工餵養,併給予100％氮氣缺氧90 s,4℃冷刺激10 min,每天2次,連續3 d；HB-EGF榦預組:在NEC模型組基礎上予HB-EGF灌胃,每次800 μg/kg,每天4次,連續3d.正常對照組:鼠乳餵養3d,不予任何榦預.大鼠生後72h禁食12h後處死.取迴腸末耑組織,HE染色觀察病理改變併評分；電鏡下觀察線粒體超微結構變化；免疫組織化學方法檢測細胞色素C含量；Western印跡技術檢測凋亡誘導因子(apoptosis inducing factor,AIF)及凋亡肽酶激活因子-1(apoptotic protease activating factor-1,APAF-1)的錶達.組間差異比較採用單因素方差分析,兩兩比較採用q檢驗,P＜0.05為差異有統計學意義. 結果 (1)HB-EGF榦預組NEC髮生率為2/10,低于模型組(9/10),差異有統計學意義(x2=7.27,P＜0.01)；對照組未髮生NEC.(2)NEC模型組線粒體在腸上皮細胞及肌層細胞中存在明顯腫脹,基質內有多數電子透亮區,線粒體超微結構嚴重損傷.HB-EGF榦預組有少量線粒體腫脹,損傷較NEC模型組減輕.(3)NEC模型組迴腸組織細胞色素C錶達較對照組增彊,差異有統計學意義(0.030±0.018與0.002±0.001,q=6.15,P＜0.01),HB-EGF榦預組迴腸組織細胞色素C錶達(0.014±0.018)較NEC模型組減弱,差異有統計學意義(q=3.53,P＜0.05).NEC模型組的APAF-1錶達較對照組增彊(1.364±0.299與0.215±0.033,q=15.31,P＜0.05),AIF錶達也增彊(0.181±0.050與0.127±0.045,q=3.71,P＜0.05)；與NEC模型組比較,HB-EGF榦預組APAF-1的錶達(0.455±0.123)減低(q=4.04,P＜0.05),AIF的錶達(0.289±0.045)則明顯增彊(q=7.32,P＜0.05). 結論 HB-EGF能降低新生大鼠NEC髮生率,其機製之一可能是通過下調APAF-1錶達而減少新生大鼠線粒體途徑細胞凋亡.
목적 탐토간소결합성표피생장인자양생장인자(heparin-binding epidermal growth factor-like growth factor,HB-EGF)재신생인배사성소장결장염(neonatal necrotizing enterocolitis,NEC)신생대서모형선립체도경세포조망중적작용. 방법 신생무특정병원체Sprague-Dawley대서수궤분위3조,매조10지.NEC모형조:채용대유품인공위양,병급여100％담기결양90 s,4℃랭자격10 min,매천2차,련속3 d；HB-EGF간예조:재NEC모형조기출상여HB-EGF관위,매차800 μg/kg,매천4차,련속3d.정상대조조:서유위양3d,불여임하간예.대서생후72h금식12h후처사.취회장말단조직,HE염색관찰병리개변병평분；전경하관찰선립체초미결구변화；면역조직화학방법검측세포색소C함량；Western인적기술검측조망유도인자(apoptosis inducing factor,AIF)급조망태매격활인자-1(apoptotic protease activating factor-1,APAF-1)적표체.조간차이비교채용단인소방차분석,량량비교채용q검험,P＜0.05위차이유통계학의의. 결과 (1)HB-EGF간예조NEC발생솔위2/10,저우모형조(9/10),차이유통계학의의(x2=7.27,P＜0.01)；대조조미발생NEC.(2)NEC모형조선립체재장상피세포급기층세포중존재명현종창,기질내유다수전자투량구,선립체초미결구엄중손상.HB-EGF간예조유소량선립체종창,손상교NEC모형조감경.(3)NEC모형조회장조직세포색소C표체교대조조증강,차이유통계학의의(0.030±0.018여0.002±0.001,q=6.15,P＜0.01),HB-EGF간예조회장조직세포색소C표체(0.014±0.018)교NEC모형조감약,차이유통계학의의(q=3.53,P＜0.05).NEC모형조적APAF-1표체교대조조증강(1.364±0.299여0.215±0.033,q=15.31,P＜0.05),AIF표체야증강(0.181±0.050여0.127±0.045,q=3.71,P＜0.05)；여NEC모형조비교,HB-EGF간예조APAF-1적표체(0.455±0.123)감저(q=4.04,P＜0.05),AIF적표체(0.289±0.045)칙명현증강(q=7.32,P＜0.05). 결론 HB-EGF능강저신생대서NEC발생솔,기궤제지일가능시통과하조APAF-1표체이감소신생대서선립체도경세포조망.
Objective To investigate the effect of heparin-binding epidermal growth factor-like growth factor (HB-EGF) on mitochondrial pathway of apoptosis in rats with neonatal necrotizing enterocolitis (NEC).Methods Sprague-Dawley neonatal rats were randomly divided into three groups with ten in each.NEC group rats were formula fed,and hypoxia exposed by 100％ N2 for 90 s and cold stress at 4 ℃ for 10 min twice a day for three days.Additionally,rats in HB-EGF group received HB-EGF 800μg/kg by gavage four times a day for three days.Rats in control group were given breast milk feeding for three days without any interventions.Seventy-two hours after born,all neonatal rats were sacrificed after fasting for 12 h,from which the terminal ileum was removed.HE-staining was done for histologic evaluation.Mitochondrial ultrastructure was observed under electron microscopy.Cytochrome C was detected by immunohistochemical analysis and apoptosis inducing factor (AIF) and apoptotic protease activating factor-1 (APAF-1) were measured by Western blot.Analysis of variance and q-test were used to compare the difference among groups.Results (1) The incidence of NEC in HB-EGF group was lower than that in NEC group (2/10 vs 9/10,x2 =7.27,P＜0.01).(2) In NEC group,mitochondria in epithelial cells and muscle cells of intestine were significantly swelling,appearing many electron-lucent zones in matrix.Ultrastructure of mitochondria were severely damaged.In HB-EGF group,mitochondria were less swelling and showed milder damage than those in NEC group.(3) The expression of cytochrome C in ileal tissue in NEC group was higher than that in control group (0.030±0.018 vs 0.002±0.001,q=6.15,P＜0.01).The expression of cytoehrome C in ileal tissue in HB-EGF group was lower than that in NEC group (0.014±0.018 vs 0.030±0.018,q=3.53,P＜0.05).The expression of APAF-1 and AIF in NEC group was higher than those in control group (1.364±0.299 vs 0.215±0.033,q=15.31,P＜0.05;0.181±0.050 vs0.127±0.045,q=3.71,P＜0.05).Compared to NEC group,the expression of APAF-1 was lower (0.455±0.123 vs 1.364±0.299,q=4.04,P＜0.05) and the expression of AIF was higher (0.289±0.045 vs 0.181±0.050,q=7.32,P＜0.05) in HB-EGF group.Conclusions HB-EGF could reduce the incidence of NEC in neonatal rats by inhibiting the mitochondrial pathway related apoptosis through down regulation of APAF-1.
