【摘要】：目的探討大鼠骨髓間充質(zhì)干細(xì)胞(BMSCs)異體移植對(duì)小鼠實(shí)驗(yàn)性自身免疫性腦脊髓炎(EAE)的治療作用。方法全骨髓貼壁培養(yǎng)法獲得大鼠BMSCs,流式細(xì)胞術(shù)檢測(cè)細(xì)胞免疫表型,并誘導(dǎo)成骨方向分化。取雌性C57BL/6小鼠,隨機(jī)分為3組:正常對(duì)照組、PBS組和大鼠BMSCs組,用髓鞘少突膠質(zhì)細(xì)胞糖蛋白(MOG)35~55聯(lián)合完全弗氏佐劑誘導(dǎo)建立EAE模型。小鼠免疫后38d和48d腹腔注射PBS或大鼠BMSCs進(jìn)行治療,神經(jīng)功能評(píng)分觀察各組小鼠神經(jīng)功能變化。二次治療12d后取各組小鼠脊髓、脾臟和外周血。HE染色及Luxol fast blue染色觀察脊髓炎性細(xì)胞浸潤及髓鞘脫失情況;脾臟制成單細(xì)胞懸液,細(xì)胞CFSE標(biāo)記后10mg/L刀豆球蛋白(Con A)和MOG_(35~55)刺激培養(yǎng)3d,觀察脾細(xì)胞增殖情況。ELISA檢測(cè)大鼠BMSCs移植后外周血細(xì)胞因子干擾素γ(IFN-γ)、白細(xì)胞介素17(IL-17)含量。結(jié)果流式細(xì)胞術(shù)顯示,大鼠BMSCs第3代(P3)細(xì)胞表達(dá)抗原CD29、CD90、CD106,不表達(dá)CD45。體外誘導(dǎo)其能向成骨分化。小鼠發(fā)病后,大鼠BMSCs組小鼠神經(jīng)功能缺損癥狀較PBS組減輕,評(píng)分降低。HE染色和Luxol fast blue染色結(jié)果顯示,大鼠BMSCs組脊髓炎細(xì)胞浸潤和脫髓鞘比同時(shí)間點(diǎn)PBS組減輕(P0.05)。Con A和MOG_(35~55)刺激培養(yǎng)后,PBS組和大鼠BMSCs組脾細(xì)胞增殖增加,而大鼠BMSCs組又較PBS組降低。與PBS組相比,大鼠BMSCs組血漿細(xì)胞因子IFN-γ、IL-17含量降低(P0.05)。結(jié)論全骨髓貼壁培養(yǎng)法能有效分離純化BMSCs,大鼠BMSCs異體移植對(duì)小鼠EAE模型有治療作用。
[Abstract]:Objective To study the effect of bone marrow mesenchymal stem cells (BMSCs) on experimental autoimmune encephalomyelitis (EAE) in mice. Methods BMSCs and flow cytometry were used to detect the cell-immune phenotype and to induce the differentiation of the osteogenesis. The female C57BL/6 mice were randomly divided into three groups: the normal control group, the PBS group and the rat BMSCs group, and the EAE model was established by the combination of the myelin oligodendrocyte glycoprotein (MOG)35-55 and the complete Freund's adjuvant. After the mice were immunized for 38d and 48d, the rats were injected with PBS or rat BMSCs for treatment, and the neurological function scores of the mice were observed to observe the changes of the neurological function in each group. The spinal cord, spleen and peripheral blood of each group were taken after 12 days of secondary treatment. The proliferation of spleen cells was observed by HE staining and Luxol fast blue staining to observe the infiltration of myelitis and the depigmentation of the pulp. The spleen was made into single cell suspension, and the cell CFSE was labeled with 10 mg/ L of concanavalin (Con A) and MOG _ (35-55). The levels of IL-17 and IL-17 in peripheral blood of rat BMSCs were detected by ELISA. Results Flow cytometry showed that the expression of CD29, CD90 and CD106 in the third generation (P3) cells of rat BMSCs did not express CD45. In vitro induce it to differentiate into osteogenesis. After the onset of the mouse, the neurological deficit in the BMSCs group was reduced in the group of PBS and the score was decreased. The results of HE staining and Luxol fast blue staining showed that the proliferation and proliferation of the spleen cells in the PBS group and the rat BMSCs group increased after the rats BMSCs group and the rats BMSCs group were stimulated and cultured, and the BMSCs group in the rat was lower than the PBS group. Compared with the PBS group, the content of IFN-1 and IL-17 in the rat BMSCs group decreased (P0.05). Conclusion The whole bone marrow adherent culture method can effectively separate and purify BMSCs, and the rat BMSCs allografts have a therapeutic effect on the mouse EAE model.
【作者單位】： 貴州醫(yī)科大學(xué)基礎(chǔ)醫(yī)學(xué)院組織學(xué)與胚胎學(xué)教研室;貴州醫(yī)科大學(xué)貴州省細(xì)胞工程生物醫(yī)藥技術(shù)國家地方聯(lián)合工程實(shí)驗(yàn)室貴州省再生醫(yī)學(xué)重點(diǎn)實(shí)驗(yàn)室;
【基金】：貴州省教育廳基金項(xiàng)目[黔教合KY字2016(071)] 貴州省科技廳優(yōu)秀青年科技人才培養(yǎng)對(duì)象專項(xiàng)基金[黔科合人字(2015)07號(hào)] 貴州省科技合作項(xiàng)目聯(lián)合基金[黔科合LH字(2016)7363]
【分類號(hào)】：R457.7;R744.5

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