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目的探讨对糖尿病(diabetes mellitus,DM)大鼠心室壁注射移植BMSCs后,BMSCs对DM大鼠心功能的影响及其机制。方法取3～4月龄雄性SD大鼠骨髓分离纯化BMSCs,取第5代BMSCs行DAPI标记。另取清洁级近交系SD雄性大鼠30只,体重约250g,随机分为正常对照组(A组)、DM组(B组)、细胞移植组(C组),每组10只。B、C组实验动物以高脂饲料喂养4周、30mg/kg链尿佐菌素腹腔注射,再以高脂饲料喂养4周制备2型DM动物模型。造模成功后B、C组实验动物心室壁分别注射PBS及DAPI标记的第5代BMSCs(1×105个/μL,160μL),继续高脂饲料喂养8周后,TUNEL法检测各组心肌细胞凋亡,多导生理记录仪检测各组大鼠心功能,Western blot和免疫组织化学检测各组心肌APPL1蛋白的表达,硝酸还原酶法测定各组大鼠NO含量。A组大鼠以基础饲料喂养16周后进行以上相关检测。结果A组心肌细胞凋亡率、APPL1含量、左心室内压最高下降速率(leftventricular-dP/dt,LV-dP/dt)、左心室舒张末期压力(leftventricular end-diastolic pressure,LVEDP)、NO含量分别为:6.14%±0.02%、2.79±0.32、(613.27±125.36)mmHg/s(1mmHg=0.133kPa)、(10.06±3.24)mmHg、(91.54±6.15)nmol/mL;与A组比较,B组心肌细胞凋亡率45.71%±0.04%明显增加,APPL1表达1.08±0.24明显减少,LV-dP/dt(437.58±117.58)mmHg/s显著下降,LVEDP(17.89±2.35)mmHg明显上升,NO含量(38.91±8.67)nmol/mL显著下降,差异均有统计学意义(P<0.05);与B组比较,C组大鼠心肌细胞凋亡率27.43%±0.03%明显下降,APPL1表达2.03±0.22显著增加,LV-dP/dt(559.38±97.37)mmHg/s显著增加,LVEDP(12.55±2.87)mmHg显著下降,NO含量(138.79±7.23)nmol/mL增加,差异均有统计学意义(P<0.05)。结论BMSCs移植可改善DM大鼠心脏舒张功能,其机制可能与激活APPL1信号途径,增加NO产生有关。
Objective To investigate the effect and mechanism of BMSCs on cardiac function in DM rats after injection of BMSCs into the ventricular wall of diabetic mellitus (DM) rats. Methods BMSCs were isolated and purified from bone marrow of 3 to 4-month-old male Sprague-Dawley rats. The fifth passage BMSCs were labeled with DAPI. In addition, 30 SD male rats of clean grade inbred line were weighed and divided into normal control group (group A), DM group (group B) and cell transplantation group (group C), with a body weight of about 250g. Groups B and C were fed with high-fat diet for 4 weeks and intraperitoneal injection of 30 mg / kg streptozotocin and then fed with high-fat diet for 4 weeks to prepare type 2 DM animal model. After successful modeling, the fifth generation BMSCs (1 × 105 / μL, 160μL) of PBS and DAPI were injected into the ventricular wall of groups B and C respectively. After continued high-fat diet for 8 weeks, TUNEL method was used to detect the changes of cardiomyocytes The cardiac function of rats in each group was detected by apoptosis and multi-channel physiology recorder. The expression of APPL1 protein in myocardium of each group was detected by Western blot and immunohistochemistry. The content of NO was determined by nitrate reductase method. A group of rats fed with basal diet 16 weeks after the above-mentioned related tests. Results Apoptosis rate, APPL1 content, left ventricular-dP / dt, left ventricular end-diastolic pressure (LVEDP), NO content Respectively, which were respectively 6.14% ± 0.02%, 2.79 ± 0.32, (613.27 ± 125.36) mmHg / s (1mmHg = 0.133kPa), (10.06 ± 3.24) mmHg and (91.54 ± 6.15) nmol / The apoptotic rate of cardiomyocytes was significantly increased at 45.71% ± 0.04%, the expression of APPL1 was significantly decreased at 1.08 ± 0.24, while the LV-dP / dt decreased significantly at 437.58 ± 117.58 mmHg / s, while the LVEDP increased significantly at 17.89 ± 2.35 mmHg. (38.91 ± 8.67) nmol / mL, the differences were statistically significant (P <0.05). Compared with group B, the apoptotic rate of cardiomyocytes in group C was significantly decreased by 27.43% ± 0.03%, and the expression of APPL1 was 2.03 ± 0.22 (559.38 ± 97.37) mmHg / s, and the LVEDP (12.55 ± 2.87) mmHg decreased significantly (P <0.05). The NO content increased significantly (138.79 ± 7.23 nmol / 0.05). Conclusion BMSCs transplantation can improve cardiac diastolic function in DM rats. The mechanism may be related to the activation of APPL1 signaling pathway and the increase of NO production.