蛛网膜下腔移植骨髓源干细胞与中药联合干预对缺血再灌注大鼠脑组织中胶质源性神经营养因子的影响

Effect of subarachnoid transplantation of bone marrow origin stem cell united with TCMon the gilial cell-line derived neurotrophic factor in brain of cerebal ischemai rats

目的观察骨髓间充质千细胞（bonemarrowmesenchymalstemcells，BMSC）或骨髓单个核细胞（bonemarrowmononuclearcells，BMNC）蛛网膜下腔移植、以及联合中药蝮龙抗栓丸干预对缺血再灌注（middlecerebralarterialocclusion，MCAO）大鼠脑组织中胶质源性神经营养因子（gilialcell．1inederivedneurotrophicfactor，GDNF）含量的影响。方法分离培养骨髓源干细胞BMNCs、BMSCs，将Wistar大鼠140只随机分为7组：正常组、假手术组、缺血再灌注模型组、BMNC组、中药联合BMNC组、BMSC组、中药联合BMSC组。除正常组、假手术组外，其余5组建立MCAO大鼠模型，并在模型建立24h后，应用蛛网膜下腔注射法进行干预：缺血再灌注模型组组注射100ul0．01M的PBS溶液；BMNC组、中药联合BMNC组移植100μl的BMNCs悬液2．0×107个；BMSC组、中药联合BMSC组移植100glBMSCs悬液2．0×107个；中药联合BMNC组、中药联合BMSC组移植当日开始配合中药蝮龙抗栓丸灌胃治疗。移植后4d、28d应用定量酶联免疫吸附测定法（ELISA）检测脑组织内GDNF的含量。结果28d时，与正常组（53．46±3．91）pg／ml比较，缺血再灌注模型组GDNF含量升高[（62．60±4．05）pg／ml，P〈0．05）]；与缺血再灌注模型组比较，在移植后4d、28dBMNC组（194．21±39．56）pg／ml、（67．70±4．73）pg／m1]、BMSC组[（169．83±28．84）pg／ml、（82．66±32．23）pg／m1]，28d时BMSC组升高明显（P〈0．05）。与单纯移植BMNC组、BMSC组比较，4d、28d中药联合BMNC组[（560．61±194．84）pg／ml、（265．83±93．58）pg／m1]、中药联合BMSC组[（370．93±46．19）pg／ml、（247．34±98．02）pg／m1]GDNF含量均升高（P均〈O．05）；且4d时中药联合BMNC组升高明显（P〈0．05）。结论蛛网膜下腔移植BMSC、BMNC可上调MCAO大鼠脑组织中GDNF水平，中药联合BMNC组、中药联合BMSC可提高GDNF水平，中药联合治疗比单纯BMNC或BMSC移植有一定优势。

Objective Observe the influence on the Gilial cell-line derived neurotrophic factor （GDNF） after treatment stroke rats with Bone marrow mesenchymal stem cells （BMSCs）or Bone marrow mononuelear cells （BMNCs）combined with traditional Chinese medicine （TCM）. Methods Separating and cultivating BMSCs and BMNCs were. 140 Wistar rats were divided into 7 groups randomly, normal group, pretended surgery group, model group, BMNC group, BMNC＋TCM group, BMSC group, BMSC＋TCM group. The other five groups were performed for 2 hours middle cerebral arterial occlusion （MCAO） except normal group and pretended surgery group. Intervention methods in each group after 24 hours of MCAO： model group： Subarachnoid injection of 100 μl 0.01M PBS, BMNC group and BMNC＋TCM group, Subarachnoidinjection of 2 × 107 BMNCS, BMSC group and BMSCWTCM group, Subarachnoid injection of 2 × 107 BMSCS, BMNC ＋TCM group and BMSC ＋TCM group were treated united with TCM on the transplantation day （po.Qd.）. GDNF level in all groups＇ rat brain were analyzed by Enzyme-linked immuno sorbent assay （ELISA） method at the 4th day and the 28th day after transplantation. Results The GDNF level of model group [ （62.60± 4.05） pg/ml ] is higher than normal group＇s [- （53.46± 3.91） pg/ml ] at the 28th day （P〈 0.05）. The GDNF levels of BMNC group [ （194.21±39.56）pg/ml, （67.70±4.73）pg/ml] and BMSC group [ （169.83±28.84）pg/ml, （82.66±32.23）pg/ml] are higher than model group＇s at the 4th and 28th day（P〈0.05）. The GDNF level of BMSC group is higher than BMNC group＇s at the 28th day（P〈0.05）. The GDNF levels of group BMNC＋ TCM group[- （560.61 ± 194.84） pg/ml, （265.83 ± 93.58） pg/ml and BMSC ＋ TCM group[- （370.93 ± 46.19） pg/ml, （247.34± 98.02）pg/ml] are higher significantly than BMNC group＇s or BMSC group＇s at the 4th and 28th day（P〈0.05）. At the 4th day the GDNF level of the BMNC＋TCM group is higher than BMSC＋TCM group＇s （P〈0.05）. Conclusion Subarachnoid transplantation of BMNCs or BMSCs will increase the GDNF level in brain of MCAO rats. The transplantantion combined with TCM can inprove the capability of the enhance. That reflect the advantage of transplantantion bone marrow origin stem cell united with TCM.