發(fā)布時間：2018-10-08 16:55

【摘要】：目的：探討在體外條件下不同濃度的利福噴丁溶液對兔脂肪干細胞增殖活性的影響。方法：本試驗取健康新西蘭大白兔(雌雄不限)，體重2.5-3.0kg，由新疆醫(yī)科大學(xué)實驗動物中心提供。用3％戊巴比妥鈉(1ml/kg)或速眠新(0.3ml/kg)麻醉后，8％硫化鈉水溶液術(shù)區(qū)脫毛備皮，消毒，取肩胛間區(qū)皮下脂肪，無菌切取皮下脂肪。放入裝有10ml PBS的培養(yǎng)皿中，帶回實驗室。用I型膠原酶將細胞消化后，接種在DMEM生長液中進行培養(yǎng)。在倒置顯微鏡下觀察細胞生長態(tài)勢以及形狀。待細胞增殖到一定程度時進行傳代，取第三代細胞接種后加入成骨誘導(dǎo)劑誘導(dǎo)成骨，于成骨誘導(dǎo)第17天鏡下觀察有鈣結(jié)節(jié)形成，茜素紅染色結(jié)果陽性，，證實所取細胞為兔脂肪干細胞，然后再取第三代兔脂肪干細胞分別設(shè)3組不同利福噴丁濃度(根據(jù)每組利福噴丁濃度不同，分為10μg/ml組、20μg/ml組、40μg/ml組)及空白對照組進行細胞培養(yǎng)，采用CCK-8比色法進行增殖分化活性比較。用堿性磷酸酶及礦化結(jié)節(jié)染色鑒定成骨細胞分化能力。結(jié)果：堿性磷酸酶及礦化結(jié)節(jié)染色均為陽性。與空白對照組相比：利福噴丁溶液濃度為10μg/ml、20μg/ml、40μg/ml時細胞增殖活性在統(tǒng)計學(xué)上無差異(P＞0.05)；在顯微鏡下可觀察到利福噴丁溶液濃度為40μg/ml時兔脂肪干細胞形態(tài)發(fā)生明顯變化。結(jié)論：從兔脂肪組織中分離、培養(yǎng)出的脂肪干細胞(ADSCs)在具有在體外誘導(dǎo)成為成骨細胞(OB)的分化潛能。利福噴丁溶液濃度為10μg/ml、20μg/ml、40μg/ml時細胞增殖活性在統(tǒng)計學(xué)上無差異(P＞0.05)；在顯微鏡下可觀察到利福噴丁溶液濃度為40μg/ml時兔脂肪干細胞形態(tài)發(fā)生明顯變化。
[Abstract]:Aim: to investigate the effect of rifapentine solution at different concentrations on the proliferation of rabbit adipose stem cells in vitro. Methods: the healthy New Zealand white rabbits (male and female), weighing 2.5-3.0 kg, were provided by Experimental Animal Center of Xinjiang Medical University. After anaesthesia with 3% pentobarbital sodium (1ml/kg) or 0.3ml/kg, 8% sodium sulphide solution was used to remove hair and skin, sterilize, take subcutaneous fat in interscapular area, and cut subcutaneous fat aseptically. Put it in a petri dish containing 10ml PBS and bring it back to the lab. Cells were digested with type I collagenase and cultured in DMEM growth liquid. Cell growth and shape were observed under inverted microscope. After the third generation cells were inoculated with osteogenic inducer to induce osteogenesis, calcium nodules were observed under the microscope on the 17th day of osteogenesis induction, and alizarin red staining was positive. The cells were confirmed to be rabbit adipose stem cells, and then the third generation of rabbit adipose stem cells were divided into three groups with different concentrations of rifapentine (10 渭 g/ml group, 20 渭 g/ml group, 40 渭 g/ml group) and blank control group, respectively. The activity of proliferation and differentiation was compared by CCK-8 colorimetry. The differentiation ability of osteoblasts was evaluated by alkaline phosphatase and mineralized nodule staining. Results: alkaline phosphatase and mineralized nodules were positive. When the concentration of rifapentine solution was 10 渭 g / ml or 20 渭 g / ml or 40 渭 g/ml, the proliferative activity of rabbit adipose stem cells had no statistical difference (P > 0. 05), and the morphological changes of adipose stem cells were observed under microscope when the concentration of rifapentine solution was 40 渭 g/ml. Conclusion: adipose stem cell (ADSCs) isolated from rabbit adipose tissue has the potential to differentiate into osteoblast (OB) in vitro. When the concentration of rifapentine solution was 10 渭 g / ml ~ (20 渭 g / ml), the proliferative activity of rabbit adipose stem cells had no statistical difference (P > 0. 05), and the morphological changes of adipose stem cells were observed under microscope when the concentration of rifapentine solution was 40 渭 g/ml.
【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】：2012
【分類號】：R329

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