【摘要】：目的培養(yǎng)人臍帶間充質(zhì)干細胞并分析細胞的生物安全性，從而為臨床應(yīng)用提供依據(jù)。方法剝除新鮮臍帶血管，取華通式膠，采用組織貼壁法培養(yǎng)細胞，將不同代數(shù)的細胞（P1～P9）采用形態(tài)學(xué)、表面標志物測定、分化能力檢測及增殖能力等手段進行鑒定，對P3～P6細胞進行病原學(xué)、核型、急慢性毒性、過敏及致瘤性的研究，以明確臍帶間充質(zhì)干細胞的安全性。結(jié)果人臍帶間充質(zhì)干細胞呈梭型流水狀生長，并可在體外穩(wěn)定擴增，細胞增殖呈‘S’型曲線；流式檢測陽性標志CD90、CD44、CD73表達量均在90%以上，陰性標志表達量在P3～P6表達量較低；所培養(yǎng)細胞可以分化為成脂細胞、成骨細胞以及成軟骨細胞。通過對細胞病原學(xué)檢測，未發(fā)現(xiàn)細菌、真菌及其他病原微生物；采用染色體G顯帶法對P3～P6臍帶間充質(zhì)干細胞進行染色核型分析，細胞在傳代過程中未發(fā)現(xiàn)染色體畸變；通過動物實驗觀察臍帶間充質(zhì)干細胞的安全性，注射人臍帶間充質(zhì)干細胞后未出現(xiàn)急性中毒，慢性毒性實驗顯示各組織器官未出現(xiàn)病理改變，致瘤實驗在注射部位未發(fā)現(xiàn)腫瘤細胞，過敏實驗未發(fā)現(xiàn)實驗動物出現(xiàn)皮膚局部及全身的過敏反應(yīng)。結(jié)論人臍帶可以分離培養(yǎng)間充質(zhì)干細胞，間充質(zhì)干細胞可在體外穩(wěn)定擴增培養(yǎng)，臍帶來源的間充質(zhì)干細胞具有生物安全性，可為今后臨床治療提供選擇。 目的探索經(jīng)鼻粘膜移植不同類型干細胞治療大鼠腦白質(zhì)損傷方法的可行性。方法培養(yǎng)鼠源少突膠質(zhì)前體細胞、鼠源骨髓間充質(zhì)干細胞、人臍帶間充質(zhì)干細胞。7日齡的SD大鼠制作腦白質(zhì)損傷模型，共40只，隨機分為四組，每組10只：（Ⅰ）鼠源少突膠質(zhì)前體細胞移植組、（Ⅱ）鼠源骨髓間充質(zhì)干細胞移植組、（Ⅲ）人臍帶間充質(zhì)干細胞移植組、（Ⅳ）對照組。10日齡模型大鼠經(jīng)鼻粘膜進行細胞移植，移植前30min經(jīng)鼻黏膜滴入透明質(zhì)酸酶200U，細胞移植量5×105/20μl，于移植后24h、48h、72h以及7d取腦組織固定脫水，進行冰凍切片，觀察細胞移植后的遷移情況。結(jié)果所培養(yǎng)的三種細胞生長良好，大鼠腦白質(zhì)損傷模型患側(cè)MBP表達量明顯減少，，經(jīng)鼻粘膜細胞移植后，模型鼠損傷部位可以發(fā)現(xiàn)標記的鼠源少突膠質(zhì)前體細胞和鼠源骨髓間充質(zhì)干細胞，并隨著時間的延長遷移至損傷部位的細胞數(shù)量逐漸增多，損傷側(cè)未發(fā)現(xiàn)移植的人臍帶間充質(zhì)干細胞。結(jié)論鼠源少突膠質(zhì)前體細胞和鼠源骨髓間充質(zhì)干細胞具有向損傷部位遷移的能力。經(jīng)鼻粘膜細胞移植治療腦白質(zhì)損傷是一種簡便、無創(chuàng)、可行的治療方法。
[Abstract]:Objective to culture human umbilical cord mesenchymal stem cells and analyze their biological safety. Methods the fresh umbilical cord blood tube was stripped, the Huatong glue was taken, and the cells were cultured by tissue adherent method. The cells of different generations (P1~P9) were identified by means of morphology, surface marker measurement, differentiation ability test and proliferation ability. The etiology, karyotype, acute and chronic toxicity, allergy and tumorigenicity of P3~P6 cells were studied to determine the safety of umbilical cord mesenchymal stem cells. Results the mesenchymal stem cells of human umbilical cord were spindle-like and could be expanded stably in vitro. The proliferation of human umbilical cord mesenchymal stem cells was'S' curve. The CD90,CD44,CD73 expression of positive markers was above 90%, and the expression of negative markers was lower in P3~P6. The cultured cells could differentiate into adipoblasts, osteoblasts and chondroblasts. No bacteria, fungi and other pathogenic microorganisms were found in the cell pathogeny, and chromosome aberration was not found in the passage of P3~P6 umbilical cord mesenchymal stem cells by chromosome G banding method. The safety of umbilical cord mesenchymal stem cells was observed by animal experiments. There was no acute poisoning after injection of human umbilical cord mesenchymal stem cells. Chronic toxicity test showed no pathological changes in tissues and organs. Tumor cells were not found in the injection site in tumorigenic test, and allergic reaction in skin and whole body was not found in allergic test. Conclusion Mesenchymal stem cells can be isolated from human umbilical cord and cultured stably in vitro. Mesenchymal stem cells derived from umbilical cord have biological safety and can provide a choice for clinical treatment in the future. Objective to explore the feasibility of different types of stem cells transplanted through nasal mucosa in the treatment of brain white matter injury in rats. Methods Rat oligodendrocyte precursor cells, mouse bone marrow mesenchymal stem cells and human umbilical cord mesenchymal stem cells were cultured. The white matter injury model was made in 7-day-old SD rats. 40 rats were randomly divided into four groups. There were 10 rats in each group: (I) rodent oligodendrocyte precursor cell transplantation group, (鈪

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