【摘要】：目的本實驗旨在通過研究G-CSF動員后分離、采集自體外周血干細(xì)胞經(jīng)皮經(jīng)腔冠脈內(nèi)移植聯(lián)合EPO治療豬急性心肌梗死,觀察其對心肌細(xì)胞病理學(xué)改變及細(xì)胞凋亡的影響。方法健康小型豬30只,隨機分為5組,每組6只。動物單純干細(xì)胞移植組(SC)、單純促紅細(xì)胞生成素組(EPO)、EPO聯(lián)合SC組(EPO+SC)、AMI對照組(control),假手術(shù)組(sham)。其中4組采用經(jīng)皮穿刺球囊封堵冠狀動脈左前降支的方法建立豬急性心肌梗死模型,1組行假手術(shù)。SC組建模后1h給予G-CSF連用7天,建模第8天采用血細(xì)胞分離機單采PBSC,通過IRA通道回輸PBSC。EPO+SC組建模后1h給予G-CSF連用7天,rHu-EPO連續(xù)5天,并于建模后第8天以相同方法采集、標(biāo)記、回輸PBSC。EPO組建模后給予rHu-EPO注射連續(xù)5天,于第8天經(jīng)IRA回輸與SC組相同容量的生理鹽水。Control組及sham組建模后僅經(jīng)IRA回輸相同容量的生理鹽水。采用蘇木精-依紅(HE)染色觀察心肌組織形態(tài)；TUNEL染色方法檢測心肌細(xì)胞凋亡指數(shù)；Western-blot方法檢測心肌梗死區(qū)域心肌組織中凋亡蛋白Bax、Caspase-3,抗凋亡蛋白Bcl-2的表達(dá)量。結(jié)果建模過程中因麻醉過量、室顫各死亡1只實驗動物,剩余30只成功建模。HE染色：sham組可見心肌細(xì)胞排列整齊,心肌細(xì)胞結(jié)構(gòu)完整；control組可見梗死區(qū)大片心肌細(xì)胞連續(xù)性壞死,梗死邊緣區(qū)存在大量炎癥細(xì)胞浸潤及纖維組織增生；EPO組與SC組心肌梗死區(qū)心肌組織相似,纖維化程度及炎性細(xì)胞浸潤情況較輕,期間散在分布正常心肌細(xì)胞；EPO+SC組心梗部位心肌組織可見更多正常心肌細(xì)胞,纖維化程度輕,邊緣少量炎性細(xì)胞浸潤。TUNEL結(jié)果：EPO+SC組凋亡指數(shù)顯著低于對照組(37.93±12.60%vs75.70±15.50%,p0.01),EPO組凋亡指數(shù)與EPO+SC組相比較高(54.08±16.06%vs37.93±12.60%,p=0.047),EPO組與SC組凋亡指數(shù)均低于對照組(p0.01)Western blot結(jié)果：EPO+SC組Bcl-2蛋白表達(dá)顯著高于EPO組、Control組及sham組(p0.01),Bax及Caspase-3蛋白表達(dá)量均減低(P0.05)。結(jié)論自體外周血干細(xì)胞移植聯(lián)合給予重組人促紅細(xì)胞生成素在急性心肌梗死的治療中可減輕梗死區(qū)域心肌的病理學(xué)改變、抑制梗死區(qū)心肌細(xì)胞的凋亡,其機制可能是通過上調(diào)抗凋亡蛋白、下調(diào)凋亡蛋白的表達(dá)來實現(xiàn)的。
[Abstract]:Objective to investigate the effects of percutaneous transcatheter coronary transplantation of autologous peripheral blood stem cells (PBSCs) and EPO on myocardial cell pathological changes and apoptosis in porcine acute myocardial infarction (AMI) after G-CSF mobilization. Methods 30 healthy miniature pigs were randomly divided into 5 groups, 6 in each group. Simple stem cell transplantation group (SC), erythropoietin group (EPO) EPO + SC (EPO SC) control group (control), sham-operation group (sham). The left anterior descending coronary artery was occluded by percutaneous balloon in 4 groups. The model of acute myocardial infarction in pigs was established. Group 1 was sham-operated. Group SC was given G-CSF for 7 days after modeling. On the 8th day of modeling, the blood cell separator was used to collect PBSCs, and the PBSC.EPO SC group was given G-CSF for 7 days for 5 days after modeling, and collected and labeled by the same method on the 8th day after modeling. PBSC.EPO group was injected with rHu-EPO for 5 days after modeling. On the 8th day, normal saline of the same volume as SC group and sham group were injected with IRA for 8 days. Only saline of the same volume was injected through IRA after modeling in sham group. The expression of apoptosis protein (Baxton-Caspase-3) and anti-apoptotic protein (Bcl-2) in myocardial tissue of myocardial infarction area was detected by using hematoxylin and ired (HE) staining. Results during the course of modeling, one experimental animal died of ventricular fibrillation due to excessive anesthesia. The remaining 30 rats were successfully modeled. The myocardial cells were arranged neatly and the structure of myocardial cells was intact in the group of HE staining. Continuous necrosis of myocardial cells in infarct area was observed in control group. There were a large number of inflammatory cell infiltration and fibrous tissue proliferation in the margin of infarction. The degree of fibrosis and infiltration of inflammatory cells were less in control group than in SC group. In the EPO SC group, more normal cardiomyocytes were found in the myocardial tissue of the EPO SC group, and the degree of fibrosis was slight. The apoptotic index of EPO group was significantly lower than that of control group (37.93 鹵12.60%vs75.70 鹵15.50, p0.01). The apoptotic index of EPO group was significantly higher than that of EPO SC group (54.08 鹵16.06%vs37.93 鹵12.60p 0.047). The apoptotic index of EPO group and SC group were significantly lower than that of control group (p0.01) Western blot result). The expression of Bax and Caspase-3 in EPO group were significantly lower than those in control group and sham group (p0.01) (P0.05). Conclusion Autologous peripheral blood stem cell transplantation combined with recombinant human erythropoietin in the treatment of acute myocardial infarction can attenuate the pathological changes of myocardium and inhibit the apoptosis of myocardial cells in infarcted area. The mechanism may be by upregulating anti-apoptotic protein and down-regulating the expression of apoptotic protein.
【學(xué)位授予單位】：青島大學(xué)
【學(xué)位級別】：碩士
【學(xué)位授予年份】：2013
【分類號】：R542.22;R457.7

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