6-羟基多巴帕金森病大鼠双侧纹状体提取液对骨髓基质细胞的诱导作用

目的 评价6-羟基多巴(6-OHDA)帕金森病(PD)大鼠纹状体提取液在体外对骨髓基质细胞(BMSCs)的诱导分化作用.方法 建立单侧(右侧)PD大鼠模型,分别配制成毁损侧纹状体诱导液(L-SC)和未毁损侧纹状体诱导液(I-SC).三代BMSCs培养至第3天进行试验.诱导组分别取L-SC和I-SC培养BMSCs,相差显微镜下观察细胞生长状态和细胞形态变化;48 h后收集细胞,免疫细胞化学染色检测细胞表面神经标记物巢蛋白(nestin)、神经特异性烯醇化酶(NSE)、胶质纤维酸性蛋白(GFAP)和酪氨酸羟化酶(TH),计数阳性率.另取部分BMSCs分别在含血清培养液(Ser-C)和无血清培养液(F-SerC)中培养,设为对照组.结果 镜下观察,L-SC组和I-SC组BMSCs部分脱落,仍然贴壁的细胞失去原有外观,有的细胞具有突起样结构.Ser-C组BMSCs生长良好,F-SerC组中的BMSCs则不能继续贴壁生长.免疫细胞化学染色显示,Ser-C组仅GFAP有阳性表达.诱导后,L-SC诱导组nestin和GFAP阳性率明显升高,I-SC诱导组NSE和GFAP阳性率明显升高,其中GFAP阳性率与Ser-C组比较差异均有统计学意义(P＜0.05),L-SC诱导组与I-SC诱导组之间GFAP阳性率差异无统计学意义(P＞0.05).各组均无TH表达.结论 PD大鼠纹状体提取液可以引起BMSCs形态学改变.与Ser-C相比,诱导后GFAP阳性细胞显著增多,同时出现nestin和NSE阳性细胞.

Lesion striatal extracts from 6-hydroxydopamine-induced PD rats induce the differentiation of BMSCs

Objective To explore the possibility of BMSCs differentiating into neurons under thc condition of in vitro culture by striatal extracts (SEs) from 6-hydroxydopamine (6-OHDA)-induced PD rats. Methods PD rat models were established by stereotaxic injection of 6-OHDA into the right medial forebrain bundle. Bilateral striatum was dissected; lesion striatal extracts (L-SEs) and intact striatal extracts (I-SEs) were prepared. The in vitro isolated and expanded third-passage BMSCs were induced by the 2 striatal extracts, respectively. Cells were observed under phase contrast microscope. The differentiated cells were detected and measured by immunocytochemical stain with nestin, neuron special enolase (NSE), glial fibrillary acidic protein (GFAP) and tyroxine hydroxylase (TH). Besides that, some other BMSCs were cultured in the serumal culture solution (Ser-C) and free serum culture solution (F-SerC) as controls. Results Morphological changes of BMSCs after treatment could be observed under contrast phase microscope. Some cells even had enations and some cells in the L-SEs and I-SEs inducement groups were out of shape. Cells in the Ser-C treatment group grew well, while cells in F-SerC treatment group could not grow well. Immunohistochemical staining indicated that GFAP positive cells expressed in the Ser-C treatment group; the GFAP and nestin positive cells in the L-SEs inducement group were increased and the GFAP and NSE positive cells in the L-SEs inducement group were increased as compared with those in the Ser-C treatment group; furthermore, the percentages of GFAP positive cells in the L-SEs and I-SEs inducement groups (53.09±32.27%, 54.60±33.14%) were significantly different with those in the Ser-C treatment group (15.05±3.92%, P＜0.05) and those were not obviously different between the L-SEs and I-SEs inducement groups (P＞0.05). No TH positive cells existed in all the groups. Conclusion SEs from 6-OHDA-induced PD rats can lead to morphological changes of BMSCs, and some cells have enations. A certain number of cells express neural special proteins after inducemet, including GFAP positive cells and some nestin/NSE positive cells.