树突细胞迁移与糖尿病大鼠主动脉内膜增生关系的研究

A study on association of migration of dendritic cells and aortal intima hyperplasia in diabetic rats

目的 探讨糖尿病大鼠树突细胞（DCs）迁移与主动脉内膜增生的关系. 方法 将20 只SD 大鼠 按随机数字表法分为正常对照组（6只）、糖尿病组（7只）和糖尿病合并高脂饮食组（7只）。采用腹腔注射链脲 佐菌素及高脂饮食喂养8 周复制糖尿病大鼠内膜增生模型。骨髓法制备大鼠DCs，应用羟基荧光素二酯酸盐琥 珀酰亚胺酯（CFSE）标记后回输大鼠体内，分别于注射CFSE 即刻和1、3、5、7 d 取大鼠外周血；7 d 处死大鼠，取 其胸主动脉标本，并制备单细胞悬液；采用流式细胞术检测CFSE 标记DCs 的比例，光镜下观察各组大鼠主动 脉内膜增生情况. 结果 注射即刻正常对照组、糖尿病组、糖尿病合并高脂饮食组外周血CFSE 标记的DCs 分 布比例比较差异无统计学意义（均P ＞0.05），糖尿病组、糖尿病合并高脂饮食组于注射1 d 开始显著下降，7 d 时 达谷值，与正常对照组比较差异有统计学意义〔（0.67±0.15）%、（0.43±0.24）% 比（1.32±0.60）%，均P ＜0.05〕。 与正常对照组比较，糖尿病组、糖尿病合并高脂饮食组主动脉单细胞悬液DCs 比例明显增加〔（2.01±0.32）%、 （1.59±0.52）%比（0.73±0.49）%，均P ＜0.01〕，管腔面积明显减小（×104 μm2 ：2.81±0.58、2.56±0.25比4.37±0.72）， 新生内膜面积明显增大（×104 μm2 ：4.77±1.13、4.58±1.30 比3.05±1.03），内膜增生百分比增加〔（62.6±6.7）%、 （63.2±7.4）% 比（40.7±10.4）%〕，差异均有统计学意义（P ＜0.05 或P ＜0.01）。光镜下可见糖尿病组和糖尿病合 并高脂饮食组主动脉中膜、内膜均明显增厚，并伴大量细胞外基质形成，血管平滑肌呈环形增生，管腔明显狭窄.结论 糖尿病大鼠及糖尿病合并高脂饮食喂养大鼠的CFSE 标记DCs 从外周血向主动脉壁内迁移加速，与主动 脉内膜增生具有相关性；DCs 的迁移可能在主动脉内膜增生中发挥重要作用.

Objective To study the relationship between aortal intima hyperplasia and the migration of dendritic cells （DCs）in diabetic rat models. Methods Twenty Sprague-Dawley （SD） rats were divided into three groups randomly ： controls （n = 6）, diabetic rats （n =7） and diabetic rats plus high-fat feeding （n =7）. The intima hyperplasia rat models with diabetes mellitus were acquired by means of an intra-peritoneal injection of streptozoein plus high-fat feeding for 8 weeks. Bone marrow-derived DCs were marked with earboxyfluoreseein diaeetate succinimidyl ester （CFSE）, and were injected back into the rat models. The blood samples were collected immediately and on the 1st, 3rd, 5th and 7th day after injection. The rats were saerifieed on the 7th day and cell suspensions from the artery wall were acquired. The proportions of CFSE marked DCs were assayed by flow eytometry, and the proliferation of aortal intima were observed by light microscope. Results There were no significant differences in the ratio of DCs with CFSE in peripheral blood among controls, diabetic rats and diabetic rats plus high-fat feeding groups （all P〉0.05）immediately after injection of the marked cells. The ratios of DCs with CFSE in peripheral blood were significantly lowered on the 1st day after injection, and the valley points were reaehed on the 7th day both in groups of diabetic rats and diabetic rats plus high-fat feeding compared with the ratio of the control group （0.67±0.15） %, （0.43±0.24）% vs. （1.32±0.60）%, both P〈0.05]. The ratios of DCs with CFSE in aortic single cell suspensions were significantly increased both in diabetic rats and diabetie rats plus high-fat feeding compared with the ratio in controls [ （2.01 ±0.32） %, （1.59±0.52） % vs. （0.73±0.49） %, both P〈0.01 ] ; while the lumen areas were significantly decreased （ × 10^4 μm2 ： 2.81±0.58, 2.56± 0.25 vs. 4.37±0.72）, neointimal areas were significantly increased （ × 10^4 μm2 ：4.77±1.13, 4.58±1.30 vs. 3.05±1.03）, and the percentages of intimal hyperplasia were increased [ （62.6±6.7） %, （63.2±7.4） %, vs. （40.7± 10.4） %] in the two diabetic groups, the differences from control group being statistieally significant （P〈0.05 or P〈0.01）. Under the light microscope, the aortal tuniea media and intima in diabetic rats in groups with or without high-fat feeding were thickened significantly with proliferated smooth muscle cells lined up in order circularly and a large amount of extraeellular matrix formation. And the lumens were obviously narrowed in groups of diabetic rats with or without high-fat feeding when compared with the lumen in controls. Conclusion The accelerated migration of DCs marked with CFSE from peripheral blood to aortal wall in diabetic rats with or without high-fat feeding possesses the relationship with aortal intima hyperplasia, suggesting the migration play an important role in the course of aortal intima hyperplasia.