CAJ | 학술논문

目的观察生理浓度丁酸钠作用Caco-2细胞肠屏障时JNK细胞信号通路的作用及与AMPK通路的关系.方法建立Caco-2细胞单层肠屏障模型,分成不加药物的对照组(Con组)、仅添加2 mmol/L丁酸钠的生理浓度丁酸钠组(2BA组)、仅加入JNK特异性抑制剂的20μmol/LSP600125组(SP组)、20 μmol/L SP600125+生理浓度丁酸钠组(Mix组).检测各组处理后24、48、72 h跨上皮细胞电阻(TER)、CCK-8法吸光度及72 h菊粉-FITC透过率、流式细胞仪检测细胞凋亡水平、吖啶橙(AO)/碘化丙啶(PI)双染法荧光显微镜检测凋亡率、Western blot法检测JNK、AMPK、ACC等蛋白的表达.结果 2BA组TER在药物作用后24、48、72 h均明显高于Con组,以72 h时最显著,此时2BA组为(509.64±12.62)Ω·cm2,而Con组为(459.30±7.62)Ω·cm2 (P＜0.05);Mix组TER为(261.01±9.22)Ω·cm2较2BA组(509.64±12.62)Ω·cm2低.Mix组72 h CCK-8吸光度为0.47±0.13、72 h菊粉-FITC为(7.89±0.80)cm/h、流式细胞仪凋亡水平为(61.23±1.54)％及AO/PI双染荧光显微镜凋亡结果为9.7±1.3,均比2BA组的0.98±0.08、(4.05±1.38)cm/h、(15.90±5.95)％、5.5±1.3高(P＜0.05).2BA组p-AMPK、p-ACC水平较Con组和Mix组上调(P＜0.05).Mix组p-JNK水平与2BA组相比显著下调(P＜().01).结论生理浓度丁酸钠可增强肠屏障功能,而JNK特异性抑制剂可破坏这一作用,提示JNK在生理浓度丁酸钠增强肠屏障功能起协同作用,这一作用可能与AMPK活化有关.
목적 관찰생리농도정산납작용Caco-2세포장병장시JNK세포신호통로적작용급여AMPK통로적관계.방법 건립Caco-2세포단층장병장모형,분성불가약물적대조조(Con조)、부첨가2 mmol/L정산납적생리농도정산납조(2BA조)、부가입JNK특이성억제제적20μmol/LSP600125조(SP조)、20 μmol/L SP600125+생리농도정산납조(Mix조).검측각조처리후24、48、72 h과상피세포전조(TER)、CCK-8법흡광도급72 h국분-FITC투과솔、류식세포의검측세포조망수평、아정등(AO)/전화병정(PI)쌍염법형광현미경검측조망솔、Western blot법검측JNK、AMPK、ACC등단백적표체.결과 2BA조TER재약물작용후24、48、72 h균명현고우Con조,이72 h시최현저,차시2BA조위(509.64±12.62)Ω·cm2,이Con조위(459.30±7.62)Ω·cm2 (P＜0.05);Mix조TER위(261.01±9.22)Ω·cm2교2BA조(509.64±12.62)Ω·cm2저.Mix조72 h CCK-8흡광도위0.47±0.13、72 h국분-FITC위(7.89±0.80)cm/h、류식세포의조망수평위(61.23±1.54)％급AO/PI쌍염형광현미경조망결과위9.7±1.3,균비2BA조적0.98±0.08、(4.05±1.38)cm/h、(15.90±5.95)％、5.5±1.3고(P＜0.05).2BA조p-AMPK、p-ACC수평교Con조화Mix조상조(P＜0.05).Mix조p-JNK수평여2BA조상비현저하조(P＜().01).결론 생리농도정산납가증강장병장공능,이JNK특이성억제제가파배저일작용,제시JNK재생리농도정산납증강장병장공능기협동작용,저일작용가능여AMPK활화유관.
Objective To explore the impact of JNK under concentrations of sodium butyrate on the Caco-2 cell intestinal barrier model and the relationship with AMP-activated protein kinase (AMPK).Methods A monolayer model of intestinal barrier was established by culturing Caco-2 cell.There were four experimental groups of control (Con),20 mol/L SP600125 (SP),physiological concentrations of sodium butyrate (2BA) and 20 μmol/L SP600125 + 2 mmol/L sodium butyrate (mix).We determined the levels of transepithelial electrical resistance (TER) at 24 h,48h,72 h and the permeability of inulin-FITC,Caco-2 cells vigor by CCK-8 assay,AO/PI fluorescent dye,apoptosis by flow cytometer and the expression of JNK,AMPK and ACC at 72 h.SP600125 was administered 1h before sodium butyrate.Results 2BA group's TER was higher than that of Con group (P＜0.05) while that of mix group was lower than 2BA group (P＜0.05).The absorbance,permeability of inulin-FITC,apoptotic cells of AO/PI fluorescent dye and apoptotic level of mix group were all higher than those of 2BA group (P＜0.05).In contrast to Con and mix groups,the expressions of p-AMPK and p-ACC of 2BA group were higher (P＜0.05).p-JNK of mix group was lower than that of 2BA group (P＜0.01).Conclusions Physiological concentration of sodium butyrate promotes intestinal barrier function while JNK may act synergistically in this process.And it is probably related with the activation of AMPK signal pathway.