CAJ | 학술논문

目的观察脱氧胆酸(DCA)对AR42J胰腺腺泡细胞的损伤作用并探讨其对核转录因子(TF)活性的影响。方法应用噻唑蓝(MTT)比色法检测DCA作用下细胞存活率改变,流式细胞术AV/PI双染法检测细胞的凋亡/坏死率。细胞经0.4mmoL/L DCA分别作用15 min、30 min、4h后收集培液上清,收集细胞并提取细胞质和细胞核蛋白,分别检测培液上清和胞质淀粉酶的活性,利用Luminex检测细胞核TF的DNA结合活性。结果 DCA对AR42J胰腺腺泡细胞的损伤作用呈浓度和时间依赖性,对细胞质内和培液中的淀粉酶水平无明显影响。在检测的40种TF活性变化中,DCA诱导ATF2、AR33、STAT5、NFAT、FKHR和NKX-2.5这6种TF活性明显升高,而RUNX/AML、NF-Y、MEF2和E2F1这4种TF活性则明显下降,其余30种TF活性无明显变化。结论 DCA对腺泡细胞的损伤作用主要表现为凋亡和坏死,对细胞内酶的合成和分泌功能没有明显影响。DCA诱导细胞核TF活性的变化,可能是其诱导细胞损伤的分子生物学基础。
목적 관찰탈양담산(DCA)대AR42J이선선포세포적손상작용병탐토기대핵전록인자(TF)활성적영향。방법 응용새서람(MTT)비색법검측DCA작용하세포존활솔개변,류식세포술AV/PI쌍염법검측세포적조망/배사솔。세포경0.4mmoL/L DCA분별작용15 min、30 min、4h후수집배액상청,수집세포병제취세포질화세포핵단백,분별검측배액상청화포질정분매적활성,이용Luminex검측세포핵TF적DNA결합활성。결과 DCA대AR42J이선선포세포적손상작용정농도화시간의뢰성,대세포질내화배액중적정분매수평무명현영향。재검측적40충TF활성변화중,DCA유도ATF2、AR33、STAT5、NFAT、FKHR화NKX-2.5저6충TF활성명현승고,이RUNX/AML、NF-Y、MEF2화E2F1저4충TF활성칙명현하강,기여30충TF활성무명현변화。결론 DCA대선포세포적손상작용주요표현위조망화배사,대세포내매적합성화분비공능몰유명현영향。DCA유도세포핵TF활성적변화,가능시기유도세포손상적분자생물학기출。
Objective To study the cytotoxic effect of desoxycholic acid (DCA) on pancreatic acinar cells AR42J, its impact on the synthesis and secretion function of amylase, and the influence on the activity of nuclear transcription factor (TF). MethodsThe cytotoxic effect of DCS was detected in rat AR42J cells by using methyl thiazol tetrazolium (MTT) assay. The rate of apoptosis or necrosis was determined by flow cytometry. After the cells were incubated with DCA (0. 4 mmol/L) for 15 min, 30 min, or 4 h, the medium was collected to detect the activity of amylase. The cytoplamic protein was extracted to detect the activity of amylase, and nuclear protein was extracted to detect the DNA binding activity of 40 TFs by Luminex. Results DCA exerted cytotoxic effects on AR42J cells in a time-and dose-dependent manner, and induced cell apoptosis and necrosis. DCA had no significant influence on the amylase synthesis and secretion function of acinar cells. For the DNA binding activity of the 40 nuclear TFs, 0. 4 mmol/L DCA up-regulated the activity of ATF2, AR33, STAT5, NFAT, FKHR, and NKX-2.5, but down-regulated the activity of RUNX/AML, NF-Y, MEF2, and E2F1. Conclusion DCA can cause AR42J pancreatic acinar cell damage by apoptosis and necrosis in a dose-and time-dependent manner. The alteration of DNA binding activity of TFs induced by DCA may be the molecular mechanism of cell damage.