Huang Feng, Xu Chang, Li Bin, Wang Shaosong, Du Xin, Guo Xiao, Rong Peijing
Beijing Hospital of TCM, Capital Medical University, Beijing 100010, China; Institution of Acupuncture and Moxibustion, China Academy of Chinese Medical Sciences, Beijing 100700.
Beijing Shijitan Hospital, Capital Medical University.
Zhongguo Zhen Jiu. 2018 Sep 12;38(9):969-77. doi: 10.13703/j.0255-2930.2018.09.017.
To observe the effect of acupoint injection of bone mesenchymal stem cells (BMSCs) combined with Chinese herbs of benefiting for activating blood circulation for capillary density and arterioles density in skeletal muscle in ischemic hind limb of diabetes mellitus (DM) rats.
A total of 80 rats were randomized into a normal sham operation group (10 rats) and a model group (70 rats). Disposable intraperitoneal injection of streptozotocin (STZ, 50.0 mg/kg) was used to establish DM model, and the rats in the model group were randomized into 7 subgroups, 10 rats in each one. The subgroups were the DM sham operation group, DM ischemic group, Chinese herb group (intragastric herbs of benefiting for activating blood circulation), local injection group (BMSCs local injection), local injection + Chinese herb group (BMSCs local injection combined with intragastric herbs of benefiting for activating blood circulation), acupoint injection group (BMSCs acupoint injection), acupoint injection + Chinese herb group (BMSCs acupoint injection combined with intragastric herbs of benefiting for activating blood circulation). The local injection was phosphate buffer (PBS) injection at the equidistant 5 points along the line between the ischemic tissue and the normal tissue a time. The acupoints were "Sanyinjiao" (SP 6), "Zhaohai" (KI 6), "Huantiao" (GB 30), "Housanli" (ST 36) and "Yanglingquan" (GB 34). 100 μL BMSCs with 1×10/mL was totally injected at the above acupoints for one rat, 20 μL an acupoint. 1.5 kg/L Chinese herbs were applied by intragastric administration, including 120 g Radix Astragali, 120 g Codonopsis, 48 g Radix Glycyrrhiza, 120 g Angelica sinensis, 120 g Blood Rattan, 48 g Achyranthes bidentata. Intragastric distilled water was used in the other non-Chinese herb groups. The expressions of α-smooth muscle actin (α-actin), latelet endothelial cell adhesion molecule (CD31) and von willebrand factor (vWF) in the skeletal muscle were detected with immunohistochemical SP two-step method.
Twenty-one days after intervention, the expressions of α-actin and CD31 on the operation hind limb were higher than those on the healthy hind limb in all the groups, except the Chinese herb group (<0.05<0.01). The vWF expressions on the operation side were lower than those on the healthy side in the Chinese herb group, the local injection group, the local injection + Chinese herb group and the acupoint injection + Chinese herb group (<0.05, <0.01). The α-actin expression on the operation side in the acupoint injection + Chinese herb group was higher than those in the normal sham operation group, DM sham operation group, the DM ischemic group and the local injection group (<0.05, <0.01). The CD31 expressions in the acupoint injection group, the acupoint injection + Chinese herb group, local injection + Chinese herb group were higher than those in the normal sham operation group, DM sham operation group and DM ischemic group (<0.05, <0.01). The CD31 expression in the acupoint injection + Chinese herb group was higher than those in the Chinese herb group and the local injection group (both <0.05). The vWF expressions in the local injection + Chinese herb group, the acupoint injection group and the acupoint injection + Chinese herb group lower than those in the DM sham operation group and the DM ischemic group (<0.05, <0.01).
schemia increases the expressions of the vascular density related factors of α-actin and CD31. It is more obvious for the increasing expressions of α-actin and CD31, and decreasing expression of vWF with the interventions of simple BMSCs injection and simple Chinese herbs of benefiting for activating blood circulation, especially with the combination of the above tow methods. It is indicated that acupoint injection of BMSCs combined with Chinese herbs of benefiting for activating blood circulation can improve the angiogenesis of ischemic tissue.
观察穴位注射骨髓间充质干细胞（BMSCs）联合活血化瘀中药对糖尿病（DM）大鼠缺血后肢骨骼肌毛细血管密度和微动脉密度的影响。
将80只大鼠随机分为正常假手术组（10只）和模型组（70只）。采用一次性腹腔注射链脲佐菌素（STZ，50.0mg/kg）建立DM模型，将模型组大鼠随机分为7个亚组，每组10只。亚组分别为DM假手术组、DM缺血组、中药组（灌胃活血化瘀中药）、局部注射组（BMSCs局部注射）、局部注射+中药组（BMSCs局部注射联合灌胃活血化瘀中药）、穴位注射组（BMSCs穴位注射）、穴位注射+中药组（BMSCs穴位注射联合灌胃活血化瘀中药）。局部注射为每次在缺血组织与正常组织连线等距离的5个点注射磷酸盐缓冲液（PBS）。穴位为“三阴交”（SP 6）、“照海”（KI 6）、“环跳”（GB 30）、“后三里”（ST 36）和“阳陵泉”（GB 34）。每只大鼠在上述穴位共注射100μL浓度为1×10/mL的BMSCs，每个穴位注射20μL。采用灌胃法给予1.5kg/L中药，包括黄芪120g、党参120g、甘草48g、当归120g、鸡血藤120g、牛膝48g。其他非中药组灌胃蒸馏水。采用免疫组织化学SP两步法检测骨骼肌中α-平滑肌肌动蛋白（α-actin）、血小板内皮细胞黏附分子（CD31）和血管性血友病因子（vWF）的表达。
干预21天后，除中药组外，所有组手术侧后肢α-actin和CD31的表达均高于健康侧后肢（<0.05<0.01）。中药组、局部注射组、局部注射+中药组和穴位注射+中药组手术侧vWF的表达低于健康侧（<0.05，<0.01）。穴位注射+中药组手术侧α-actin的表达高于正常假手术组、DM假手术组、DM缺血组和局部注射组（<0.05，<0.01）。穴位注射组、穴位注射+中药组、局部注射+中药组CD31的表达高于正常假手术组、DM假手术组和DM缺血组（<0.05，<0.01）。穴位注射+中药组CD31的表达高于中药组和局部注射组（均<0.05）。局部注射+中药组、穴位注射组和穴位注射+中药组vWF的表达低于DM假手术组和DM缺血组（<0.05，<0.01）。
缺血增加了α-actin和CD31等血管密度相关因子的表达。单纯BMSCs注射和单纯活血化瘀中药干预后，α-actin和CD31表达增加、vWF表达降低更明显，尤其是上述两种方法联合应用时。表明穴位注射BMSCs联合活血化瘀中药可改善缺血组织的血管生成。
