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  本文選題：脂肪源間充質(zhì)干細(xì)胞 + 成骨誘導(dǎo)�。� 參考：《新疆醫(yī)科大學(xué)》2013年碩士論文

【摘要】：目的：探討小鼠脂肪源間充質(zhì)干細(xì)胞對造血干/祖細(xì)胞體外擴(kuò)增的影響并對其機(jī)制進(jìn)行初步研究。方法：無菌條件下獲取C57BL/6小鼠的脂肪源干細(xì)胞（ADSCs），培養(yǎng)至P3代：①細(xì)胞形態(tài)學(xué)分析；②流式細(xì)胞術(shù)檢測細(xì)胞表面免疫表型；③脂肪源干細(xì)胞定向成骨、成脂細(xì)胞誘導(dǎo)。將小鼠成纖維細(xì)胞、骨髓間充質(zhì)干細(xì)胞、成骨誘導(dǎo)前后的脂肪源干細(xì)胞經(jīng)絲裂霉素C處理后作為滋養(yǎng)層，以骨髓單個核細(xì)胞（BMNCs）作為長期培養(yǎng)起始細(xì)胞（LTC-IC），對其進(jìn)行35天長期體外共培養(yǎng)，將本實驗分4組：A組：BMNCs加小鼠成纖維細(xì)胞，，B組：BMNCs加骨髓間充質(zhì)干細(xì)胞（BMSCs），C組：BMNCs加ADSCs，D組：BMNCs加成骨誘導(dǎo)后的脂肪源干細(xì)胞（osteo-ADSCs）；分別于0d、14d、28d、35d行甲基纖維素14d短期處理分析造血干/祖細(xì)胞集落增殖狀態(tài)；同時應(yīng)用酶聯(lián)免疫吸附法（ELISA）檢測上清液中的基質(zhì)細(xì)胞衍生因子-1（SDF-1）、N-鈣黏素（N-cadherin）、β-連環(huán)蛋白（β-catenin）、Jagged1蛋白（JAG1）的表達(dá)情況。結(jié)果：1. ADSCs的生物學(xué)分析：ADSCs在體外呈梭形生長并可穩(wěn)定傳代；成骨、成脂誘導(dǎo)均為陽性；流式結(jié)果分析：CD29高表達(dá)（95.37±0.95)％；CD34低表達(dá)（8.7±0.4）％。2.細(xì)胞體外二維共培養(yǎng)后對造血干/祖細(xì)胞（HSPCs）擴(kuò)增的影響：①不同時間點造血干/祖細(xì)胞（HSPCs）形成的集落數(shù)不同，且成骨誘導(dǎo)后的脂肪源干細(xì)胞更具有造血支持作用；②SDF-1、N-cadherin、β-catenin、JAG1的表達(dá)情況在不同時間點不同，SDF-1、β-catenin的表達(dá)結(jié)果顯示：C、D組表達(dá)明顯高于A、B兩組（P 0.05）；且C組與D組比較有統(tǒng)計學(xué)意義（P0.05）；而N-cadherin、JAG1在各組間比較無統(tǒng)計學(xué)意義（P＞0.05）。結(jié)論： C57BL/6小鼠的脂肪源干細(xì)胞及成骨誘導(dǎo)后的脂肪源干細(xì)胞在體外可有效地刺激造血。
[Abstract]:Aim: to investigate the effect of adipose derived mesenchymal stem cells (MSCs) on hematopoietic stem / progenitor cell expansion in vitro and its mechanism. Methods: adipose derived stem cells from C57BL/6 mice were obtained under aseptic condition and cultured to P3: 1 cell morphology. Flow cytometry was used to detect the surface immunophenotype of adipose derived stem cells to orient osteogenesis and adipogenic cells to induce adipogenic cells. Mouse fibroblasts, bone marrow mesenchymal stem cells and adipose stem cells before and after osteogenesis were treated with mitomycin C as trophoblast layer. Bone marrow mononuclear cells (BMNCs) were cocultured in vitro for 35 days with bone marrow mononuclear cells (BMNCs) as the starting cells for 35 days. The experiment was divided into 4 groups: group A, w: BMNCs plus mouse fibroblast cells, group B: BMNCs + bone marrow mesenchymal stem cells, group C: BMNCs + ADSCsC, group C: BMNCs added to adipose derived stem cells induced by bone osteo-ADSCsC, bone derived stem cells osteo-ADSCs1 were added to group C, respectively, on day 14, 14 days, 28 days after 35 days of methylcellulose treatment, hematopoiesis was analyzed. Stem / progenitor cell colony proliferation; At the same time, the expression of stromal cell derived factor-1, SDF-1, N-cadherin, 尾-catenin (尾-catenin1 protein) in the supernatant was detected by enzyme linked immunosorbent assay (Elisa). The result is 1: 1. The biological analysis of ADSCs showed fusiform growth and stable passage in vitro, osteogenesis and adipogenic induction were positive, flow analysis showed that the high expression of CD29 was 95.37 鹵0.95% and the low expression of CD34 was 8.7 鹵0.40.0.2. Effect of two dimensional co-culture of cells on the expansion of hematopoietic stem / progenitor cells HSPCsThe colony number of hematopoietic stem / progenitor cells was different at different time points, and adipose derived stem cells induced by osteogenesis had more hematopoietic support. 2the expression of SDF-1N-cadherin, 尾 -cateninine JAG1 was different at different time points. The expression of 尾 -catenin in group B was significantly higher than that in group Agna B (P 0.05), and that in group C was significantly higher than that in group D (P 0.05), but the expression of N-cadherin in JAG1 was not significantly higher than that in group A (P > 0.05). Conclusion: adipose stem cells of C57BL/6 mice and adipose stem cells induced by osteogenesis can stimulate hematopoiesis effectively in vitro.
【學(xué)位授予單位】：新疆醫(yī)科大學(xué)
【學(xué)位級別】：碩士
【學(xué)位授予年份】：2013
【分類號】：R329.2

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