基质细胞衍生因子-1/CXC族细胞因子受体4轴在缺氧预处理骨髓间充质干细胞移植促进Sprague-Dawley大鼠急性心肌梗死心脏功能恢复中的作用

Exp ression of stromal cell-derived factor 1 and CXC chemokine receptor 4 in acute infarct my ocardium of Sprague-Dawley rats after implantation of hypoxia-preconditioned bone marrow mesenchymal stem cells

目的观察缺氧预处理的同种异体骨髓间充质干细胞(MSCs)移植入受体后,基质细胞衍生因子(SDF)-1/CXC族细胞因子受体(CXCR)4轴在促进Sprague-Dawley(SD)大鼠急性心肌梗死(AMI)心脏功能恢复中的作用。方法对SD大鼠的MSCs进行体外培养、鉴定,并行缺氧预处理。128只SD大鼠被随机分入MSCs移植组(MSCs组)、缺氧预处理MSCs移植组(预处理MSCs组)、AMI组、假手术组,每组32只。MSCs组、预处理MSCs组和AMI组大鼠均予冠状动脉左前降支结扎造成AMI 10min后,预处理MSCs组大鼠在梗死区域、梗死区边缘局部共注射5×105/\mL经缺氧预处理的MSCs,MSCs组大鼠注射5×105/\mL MSCs,AMI组大鼠注射相同容量的0.9%氯化钠溶液。假手术组大鼠仅行开胸处理。造模前和造模后28d采用超声心动图测量左心室舒张末期内径(LVDd)、左心室收缩末期内径(LVDs)、左心室射血分数(LVEF)3项反映心脏功能的指标。分别于造模后1、7、14和28d每组各处死8只大鼠取其心脏组织行免疫组织化学、常规病理染色,计算心肌梗死面积百分比、梗死区心室壁厚度和心室肌内微血管密度(MVD)。采用实时定量PCR和免疫激光共聚焦定性、定量测定各组心肌组织内SDF-1和CXCR4的表达。结果造模后28d时超声心动图检测结果显示,预处理MSCs组、MSCs组、AMI组的LVDd和LVDs均显著长于假手术组(P值均<0.05),LVEF均显著低于假手术组(P值均<0.05);预处理MSCs组、MSCs组的LVDd和LVDs均显著短于AMI组(P值均<0.05),LVEF均显著高于AMI组(P值均<0.05);预处理MSCs组的LVDd和LVDs均显著短于MSCs组(P值均<0.05),LVEF均显著高于MSCs组(P<0.05)。造模后28d,预处理MSCs组、MSCs组的心肌梗死面积百分比均显著小于AMI组(P值均<0.05),预处理MSCs组显著小于MSCs组(P<0.05);预处理MSCs组、MSCs组左心室前壁厚度均显著厚于AMI组(P值均<0.05),预处理MSCs组显著厚于MSCs组(P<0.05)。造模后28d,预处理MSCs组、MSCs组、AMI组心肌梗死区的MVD定量和AMI组梗死边缘区的MVD定量均显著低于假手术组(P值均<0.05),预处理MSCs组、MSCs组梗死边缘区的MVD定量均显著高于假手术组(P值均<0.05),预处理MSCs组、MSCs组心肌梗死区和梗死边缘区的MVD定量均显著高于AMI组(P值均<0.05),预处理MSCs组心肌梗死区和梗死边缘区的MVD定量均显著高于MSCs组(P值均<0.05)。造模后1、7、14和28d,预处理MSCs组、MSCs组和AMI组的SDF-1和CXCR4蛋白荧光强度均显著高于假手术组(P值均<0.05),预处理MSCs组、MSCs组均显著高于AMI组(P值均<0.05),预处理MSCs组均显著高于MSCs组(P值均<0.05)。结论SD大鼠AMI区局部注射缺氧预处理的MSCs或MSCs后可使SDF-1和CRCR4在受损组织局部表达上调,同时心功能得到改善。SDF-1/CRCR4轴在心肌梗死区局部表达上调可能是缺氧预处理MSCs或MSCs移植后改善心脏功能的重要途径之一。

Objective To study the expression of stromal cell-derived factor （SDF） 1 and CXC chemokine receptor （CXCR） 4 in the acute myocardial infarction （AMI） of Sprague-Dawley （SD） rats after implantation of mesenchymal stem cells （MSCs） with hypoxic preconditioning. Methods A total of 128 SD rats were randomly divided into four groups （ n = 32） ： MSC group, hypoxic preconditioning MSC group （HPMSC group）, AMI group and sham operation group. MSCs were cultured in vitro and underwent hypoxic preconditioning before transplantation. The left anterior descending coronary arteries were ligated to create AMI in all rats except those in the sham operation group. Ten minutes later, 5 × 105/μL HPMSCs, 5 × 105/μL MSCs and isovolume of normal saline were injected into the area of acute infarct myocardium in HPMSC group, MSC group and AMI group, respectively. In the sham operation group, only left anterior thoracotomy was performed through 3, 4 intercostal region. Cardiac function indices, such as left ventricular end diastolic dimension （LVDd）, left ventricular end- systolic dimension （LVDs） and left ventricular ejection fraction （LVEF） were measured by ultrasoundcardiogram on day 28 before and after modeling. Eight rats were sacrificed on Day 1, 7, 14 and 28 after modeling in each group, respectively, and the heart was harvested for the examination of immunohistochemistry and pathology and the detection of infarct size, thickness of infarct ventricular wall and microvessel density （MVD） of ventricular myocardium. The expression of SDF-1 and CXCR4 in the area of acute myocardial infarction were analyzed by real- time polymerase chain reaction （PCR） and immune laser copolymerization. Results The results of ultrasoundcardiogram indicated that, on Day 28 after AMI, LVDd and LVDs in MSC group, HPMSC group and AMI group were significantly increased while LVEF were significantly decreased as compared with that in sham operation group （all P〈0.05）r LVDd and LVDs in MSC group and HPMSC group were significantly decreased while LVEF were significantly increased as compared with those in AMI group （both P〈0.05） ; LVDd and LVDs in HPMSC group were significantly decreased while LVEF was significantly increased as compared with that in MSC group （all P〈0.05）. The myocardial infarct size of MSC and HPMSC groups on Day 28 after AMI was significantly smaller than that that in AMI group （both P 〈 0. 05）, and the myocardial infarct size of HPMSC group was significantly smaller than that in MSC group （P〈0.05）. The left ventricular anterior wall in MSC and HPMSC groups was significantly thicker than that in AMI group （both P〈0.05）; and the left ventricular anterior wall in HPMSC group was significantly thicker than that in MSC group （P〈0.05）. The MVD in the infarct area in MSC, HPMSC and AMI groups and the MVD in the marginal zone of infarction in AMI group were significantly less than that in sham operation group （all P〈0.05）. The MVD in the marginal zone of infarction in MSC and HPMSC groups were significantly more than that in sham operation group （both P〈0.05）. The MVD in both the infarct area and the marginal zone in MSC and HPMSC groups were significantly more than that in AMI group （all P〈0.05）. The MVD in both the infarct area and the marginal zone in HPMSC group were also significantly more than that in MSC group （both P〈0.05）. On Day 1, 7, 14 and 28 after modeling, protein levels of SDF-1 and CXCR4 in MSC, HPMSC and AMI groups were significantly higher than those in sham operation group （all P〈0.05） protein levels of SDF-1 and CXCR4 in MSC and HPMSC groups were significantly higher than those in AMI group （all P〈0.05） protein levels of SDF-1 and CXCR4 in HPMSC group were significantly higher than those in MSC （all P〈0. 05）. Conclusion After implantation of HPMSCs or MSCs in SD rats with AMI, the expression of SDF-1 and CXCR4 can be upregulated and cardiac function will improve, which indicates that the upregulation of SDF-1 and CXCR4 is one of the most important pathway to improve cardiac function in AMI rats after implantation of HPMSCs or MSCs.