医学研究生学报
醫學研究生學報
의학연구생학보
JOURNAL OF MEDICAL POSTGRADUATE
2015年
2期
123-126
,共4页
肖容容%高景红%范越%周露%施睿臻%李庆平
肖容容%高景紅%範越%週露%施睿臻%李慶平
초용용%고경홍%범월%주로%시예진%리경평
肾素血管经张素系统%骨髓间充质干细胞%缺氧/无血清
腎素血管經張素繫統%骨髓間充質榦細胞%缺氧/無血清
신소혈관경장소계통%골수간충질간세포%결양/무혈청
Renin-angiotensin system%Bone marrow mesenchymal stem cell%Hypoxia/serum deprivation
目的：肾素-血管紧张素系统（ renin-angiotensin system ， RAS）参与缺氧心肌损伤过程。文中旨在观察骨髓间充质干细胞（mesenchymal stem cells， MSCs）自身主要反应元件血管紧张素受体1（angiotensin II type 1 receptor， AT1-R）、AT2-R和血管紧张素转换酶（ angiotensin-converting enzyme ， ACE）在缺氧条件下的表达情况。方法分离培养大鼠的MSCs，用倒置相差显微镜观察细胞形态，应用CD29、CD11b／c抗体进行细胞鉴定。实验分为对照组和Hypoxia／SD组：对照组用含10％FBS的L-DMEM培养MSCs，Hypoxia／SD组采用缺氧联合无血清处理24 h诱导缺氧损伤模型。2组分别用锥虫蓝染色、CCK-8检测及流式细胞术，测定细胞活力和凋亡率。应用Real-time Quantitative PCR法检测细胞内AT1-R、AT2-R和ACE的mRNA表达水平；用Western blot检测细胞内AT1-R、AT2-R和ACE蛋白表达的变化。结果 MSCs 特异性抗原CD29阳性率＞97％，非特异性抗原CD11b／c阳性率＜1％，分离培养成功；与对照组相比，Hypoxia／SD组24 h可显著增加MSCs凋亡率[（6．73±0．78）％vs （19．93±4．92）％， P＜0．01]，降低细胞活力[（78．49±4．94）％ vs （37．33±2．91）％，P＜0．01]。 Hypoxia／SD环境下MSCs自身的RAS主要反应元件AT1-R、AT2-R及ACE的mRNA和蛋白表达量均增加。结论推断缺氧导致MSCs自身RAS激活，从而提高MSCs对缺氧损伤区的心肌修复能力。
目的：腎素-血管緊張素繫統（ renin-angiotensin system ， RAS）參與缺氧心肌損傷過程。文中旨在觀察骨髓間充質榦細胞（mesenchymal stem cells， MSCs）自身主要反應元件血管緊張素受體1（angiotensin II type 1 receptor， AT1-R）、AT2-R和血管緊張素轉換酶（ angiotensin-converting enzyme ， ACE）在缺氧條件下的錶達情況。方法分離培養大鼠的MSCs，用倒置相差顯微鏡觀察細胞形態，應用CD29、CD11b／c抗體進行細胞鑒定。實驗分為對照組和Hypoxia／SD組：對照組用含10％FBS的L-DMEM培養MSCs，Hypoxia／SD組採用缺氧聯閤無血清處理24 h誘導缺氧損傷模型。2組分彆用錐蟲藍染色、CCK-8檢測及流式細胞術，測定細胞活力和凋亡率。應用Real-time Quantitative PCR法檢測細胞內AT1-R、AT2-R和ACE的mRNA錶達水平；用Western blot檢測細胞內AT1-R、AT2-R和ACE蛋白錶達的變化。結果 MSCs 特異性抗原CD29暘性率＞97％，非特異性抗原CD11b／c暘性率＜1％，分離培養成功；與對照組相比，Hypoxia／SD組24 h可顯著增加MSCs凋亡率[（6．73±0．78）％vs （19．93±4．92）％， P＜0．01]，降低細胞活力[（78．49±4．94）％ vs （37．33±2．91）％，P＜0．01]。 Hypoxia／SD環境下MSCs自身的RAS主要反應元件AT1-R、AT2-R及ACE的mRNA和蛋白錶達量均增加。結論推斷缺氧導緻MSCs自身RAS激活，從而提高MSCs對缺氧損傷區的心肌脩複能力。
목적：신소-혈관긴장소계통（ renin-angiotensin system ， RAS）삼여결양심기손상과정。문중지재관찰골수간충질간세포（mesenchymal stem cells， MSCs）자신주요반응원건혈관긴장소수체1（angiotensin II type 1 receptor， AT1-R）、AT2-R화혈관긴장소전환매（ angiotensin-converting enzyme ， ACE）재결양조건하적표체정황。방법분리배양대서적MSCs，용도치상차현미경관찰세포형태，응용CD29、CD11b／c항체진행세포감정。실험분위대조조화Hypoxia／SD조：대조조용함10％FBS적L-DMEM배양MSCs，Hypoxia／SD조채용결양연합무혈청처리24 h유도결양손상모형。2조분별용추충람염색、CCK-8검측급류식세포술，측정세포활력화조망솔。응용Real-time Quantitative PCR법검측세포내AT1-R、AT2-R화ACE적mRNA표체수평；용Western blot검측세포내AT1-R、AT2-R화ACE단백표체적변화。결과 MSCs 특이성항원CD29양성솔＞97％，비특이성항원CD11b／c양성솔＜1％，분리배양성공；여대조조상비，Hypoxia／SD조24 h가현저증가MSCs조망솔[（6．73±0．78）％vs （19．93±4．92）％， P＜0．01]，강저세포활력[（78．49±4．94）％ vs （37．33±2．91）％，P＜0．01]。 Hypoxia／SD배경하MSCs자신적RAS주요반응원건AT1-R、AT2-R급ACE적mRNA화단백표체량균증가。결론추단결양도치MSCs자신RAS격활，종이제고MSCs대결양손상구적심기수복능력。
Objective The renin-angiotensin system ( RAS) is involved in myocardial anoxic injury .This study aimed to in-vestigate the expressions of AT 1-R, AT2-R, and angiotensin-converting enzyme ( ACE ) in bone marrow mesenchymal stem cells (MSCs) under hypoxia. Methods Rat MSCs were isolated, cultured, and identified with CD29 and CD11b/c antibodies.The is-chemic injury model was established by exposing the MSCs to hypoxia and serum deprivation ( Hypoxia/SD) for 24 hours, while the control cells were cultured in L-DMEM with 10%FBS.The vitality and apoptosis of the cells were detected by trypan blue staining , CCK8 assay, and Annexin V-FITC staining.The mRNA and protein expressions of AT 1-R, AT2-R, and ACE were determined by real-time quantitative PCR and Western blot , respectively. Results The positive rate of CD29 was >97%and that of CD11b/c was <1% in the MSCs.Compared with the control group, Hypoxia/SD significantly increased the rate of cell apoptosis ([6.73 ±0.78]%vs [19.93 ±4.92]%, P<0.01), decreased the rate of cell viability ([78.49 ±4.94]%vs [37.33 ±2.91]%, P<0.01), and up-regulated the mRNA and protein expressions of AT 1-R, AT2-R, and ACE. Conclusion Hypoxia/SD activates the RAS in MSCs and improves the protective function of the cells against myocardial anoxic injury .
