本文選題：骨髓間充質(zhì)干細(xì)胞 + 增殖��； 參考：《大連醫(yī)科大學(xué)》2012年碩士論文

【摘要】：目的：本研究以SD大鼠為動(dòng)物模型，研究大鼠BMSCs體外分離培養(yǎng)、鑒定和增殖，評(píng)價(jià)不同濃度的LPS對(duì)BMSCs增殖能力的影響；并探討通過增殖的BMSCs對(duì)ALI修復(fù)的可行性。 方法： 1.原代BMSCs的分離培養(yǎng)、誘導(dǎo)分化鑒定、流式細(xì)胞鑒定，并觀察不同代的BMSCs的生長狀況。給予不同濃度的LPS與BMSCs共培養(yǎng)，利用MTT法判斷BMSCs增殖時(shí)LPS的最適濃度。 2.隨機(jī)從28只雌性大鼠中取24只，經(jīng)腹腔注入0.5ml濃度為5mg/kg的LPS建立ALI，造模0.5h后，將其隨機(jī)分成3組：治療組A（8只，經(jīng)尾靜脈注入濃度為1×104/ml的BMSCs，體積為0.5ml），治療組B（8只，，經(jīng)尾靜脈注入預(yù)先用濃度為0.01μg/mlLPS共培養(yǎng)的BMSCs1×104/ml，體積為0.5ml），損傷組（8只，經(jīng)尾靜脈注入0.5mlPBS），對(duì)照組（4只，經(jīng)尾靜脈注入等量的PBS1ml）。24h后檢測各組大鼠的血?dú)庵笜?biāo)、肺濕干比、病理變化、及支氣管肺泡灌洗液中TNF-α和IL-10的變化。 結(jié)果： 1.分離培養(yǎng)的BMSCs呈現(xiàn)長梭形，緊密排列似漩渦狀。經(jīng)誘導(dǎo)液誘導(dǎo)后，能分化成脂肪細(xì)胞和成骨細(xì)胞。流式鑒定表面抗原CD29和CD90表達(dá)陽性，CD45和CD11表達(dá)陰性，符合骨髓間充質(zhì)干細(xì)胞特性。MTT法顯示第3代和第5代相比第7代的BMSCs增殖較旺盛。濃度為0.01μg/ml的LPS對(duì)BMSCs增殖作用最顯著。 2.成功造模24h后，損傷組明顯出現(xiàn)低氧、二氧化碳潴留和酸中毒，肺濕干比值較正常高，病理切片示肺泡間隔斷裂增寬，組織疏松，并有炎性細(xì)胞侵潤。BALF中的TNF-α較對(duì)照組明顯增高、IL-10較對(duì)照組明顯降低。治療組血?dú)夥治鲋笜?biāo)趨向正常，肺濕干比趨向正常，病理切片肺泡結(jié)構(gòu)相對(duì)均勻完整，肺泡間隔增寬不明顯，炎性細(xì)胞侵潤明顯減少，BALF中的TNF-α較損傷組明顯降低、IL-10較損傷組明顯增高，而且治療組B比治療組A效果佳。 結(jié)論： 1.BMSCs經(jīng)誘導(dǎo)可分化成脂肪細(xì)胞、成骨細(xì)胞。 2.本實(shí)驗(yàn)用濃度為0.01μg/ml的LPS刺激BMSCs后其增殖能力增強(qiáng)。 3.本實(shí)驗(yàn)經(jīng)腹腔注入濃度5mg/kg的LPS可建立ALI模型。 4.注入增殖的BMSCs后可改善病理學(xué)變化，有助于促炎因子TNF-α的降低和抗炎因子IL-10的增高。
[Abstract]:Aim: to study the effects of different concentrations of LPS on the proliferation of BMSCs and to explore the feasibility of ALI repair by proliferating BMSCs in this study, using SD rats as an animal model to study the isolation, culture, identification and proliferation of rat BMSCs in vitro, and to evaluate the effect of different concentrations of LPS on the proliferation of BMSCs. Methods: 1. Isolation and culture of primary BMSCs, identification of induced differentiation, flow cytometry, and observation of the growth status of BMSCs in different generations. Different concentrations of LPS and BMSCs were co-cultured and the optimum concentration of LPS was determined by MTT method. 2. Twenty-four female rats were randomly divided into three groups after intraperitoneal injection of LPS with 0.5ml concentration of 5mg/kg for 0.5 h. The rats in the treatment group were randomly divided into three groups: 8 rats in the treatment group were injected with 1 脳 104/ml BMSCs through the caudal vein, 8 rats in the treatment group were injected with BMS at the concentration of 1 脳 104/ml, and 8 rats in the treatment group were injected with LPS at the concentration of 1 脳 104/ml. The rats in the injury group were injected with 0. 01 渭 g/mlLPS co-cultured BMSCs1 脳 104 / ml (0.5 ml / ml), 8 rats in the injury group and 4 rats in the control group were injected through the tail vein. The blood gas index, lung wet / dry ratio and pathological changes were measured after the same amount of PBS1ml).24h was injected into the tail vein. The changes of TNF- 偽 and IL-10 in bronchoalveolar lavage fluid. Results: 1. The isolated BMSCs showed a long fusiform shape, closely arranged like a whirlpool. After induction, adipocytes and osteoblasts could be differentiated into adipocytes and osteoblasts. Flow cytometry showed that the positive expression of CD29 and CD90 was negative, which was consistent with the characteristics of bone marrow mesenchymal stem cells. The proliferation of BMSCs in the third and fifth generation was stronger than that in the seventh generation. LPS at the concentration of 0. 01 渭 g/ml had the most significant effect on the proliferation of BMSCs. 2. 24 hours after successful modeling, hypoxia, carbon dioxide retention, acidosis, wet / dry ratio of lung were higher than normal in the injury group. Pathological sections showed that the alveolar septum was broken and widened, and the tissue was loose. TNF- 偽 in BALF with inflammatory cell infiltration was significantly higher than that in control group and IL-10 was significantly lower than that in control group. In the treatment group, the indexes of blood gas analysis tended to be normal, the wet / dry lung ratio tended to be normal, the alveolar structure was relatively uniform and intact, and the alveolar septum was not widened obviously in the treatment group. TNF- 偽 in BALF was significantly reduced by inflammatory cells, and IL-10 was significantly increased in BALF than in injury group, and the effect of treatment group B was better than that of treatment group A. Conclusion: 1.BMSCs can be induced to differentiate into adipocytes and osteoblasts. 2. In this experiment, the proliferation ability of BMSCs was enhanced by LPS at a concentration of 0. 01 渭 g/ml. 3. In this experiment, ALI model could be established by intraperitoneal injection of LPS with concentration of 5mg/kg. 4. Injection of proliferative BMSCs can improve pathological changes and contribute to the decrease of pro-inflammatory factor TNF- 偽 and the increase of anti-inflammatory factor IL-10.
【學(xué)位授予單位】：大連醫(yī)科大學(xué)
【學(xué)位級(jí)別】：碩士
【學(xué)位授予年份】：2012
【分類號(hào)】：R563

【參考文獻(xiàn)】

相關(guān)期刊論文 前1條

1 ;Upregulated functional expression of Toll like receptor 4 in mesenchymal stem cells induced by lipopolysaccharide[J];Chinese Medical Journal;2007年19期

本文編號(hào)：1991041