不同浓度庆大霉素对大鼠脂肪干细胞体外软骨形成能力影响的研究
发布时间：2018-10-31 13:35
【摘要】： 目的本文研究大鼠脂肪干细胞(adipose-derived stem cells ADSCs)在体外分离培养、诱导分化为成软骨的能力,并初步观察不同浓度(0μg／m、10μg／ml、50μg／ml、100μg／ml、200μg／ml)庆大霉素对其增殖、分化成软骨的影响。 方法从3周龄SD大鼠腹股沟脂肪垫分离出脂肪组织,通过Ⅰ型胶原酶消化法分离干细胞,型胶原酶消化法分离干细胞,接种于含有胎牛血清的DMEM培养液中分离传代。每日以倒置显微镜观察细胞形态及生长情况。取第三代脂肪干细胞分五组,包含一个对照组(培养液为人脂肪干细胞完全培养基：高糖DMEM、10%胎牛血清、50nmol／L抗坏血酸、6.25mg／L胰岛素、100U／ml青霉素、100μg／ml链霉素、100nmol／L地塞米松,10μg／L转化生长因子β1,不含庆大霉素)和四个含庆大霉素浓度组(分别是10μg／ml、50μg／ml、100μg／ml、200μg／ml加人脂肪干细胞完全培养基)。采用MTT比色法进行增殖活性比较,Ⅱ型胶原免疫细胞化学染色和细胞甲苯胺蓝染色检测成软骨分化情况。 结果1.体外培养的脂肪间充质干细胞呈扁平的梭形成纤维样细胞生长,且细胞形态均一,传代稳定。2.MTT实验显示庆大霉素在100μg／ml、200μg／ml抑制细胞增殖,其余各浓度组对细胞增殖无明显影响3.诱导后细胞甲苯胺蓝染色显示100μg／ml组及200μg／ml组细胞细胞染色较浅,大部分细胞失染,余各组阳性表达：Ⅱ型胶原免疫组织化学染色显示100μg／ml组及200μg／ml组细胞弱阳性表达,余各组阳性表达。 结论1大鼠脂肪干细胞易取材,培养传代后增殖迅速,生长稳定。2大鼠脂肪干细胞能向软骨细胞诱导分化。3庆大霉素在一定浓度(10μg／ml、50μg／ml)下对脂肪干细胞的增殖和软骨分化没有影响,但高于此浓度(100μg／ml,200μg／ml)则抑制脂肪干细胞的增殖和软骨分化能力。
[Abstract]:Objective to study the ability of rat adipose stem cells (adipose-derived stem cells ADSCs) to differentiate into chondrocytes in vitro, and to observe the effects of different concentrations (0 渭 g / m ~ (10) 渭 g / ml ~ (10) 渭 g 路ml ~ (-1) of 50 渭 g / ml / ml ~ (100 渭 g / ml) on the differentiation of rat adipose stem cells into cartilage. Effects of gentamicin on proliferation and differentiation of cartilage. Methods Adipose tissue was isolated from the inguinal fat pad of 3-week-old SD rats. Stem cells were isolated by type I collagenase digestion. The stem cells were isolated and subcultured in DMEM medium containing fetal bovine serum. The morphology and growth of cells were observed by inverted microscope every day. The third generation of adipose stem cells were divided into five groups, which consisted of a control group (human adipose stem cell culture medium: high glucose DMEM,10% fetal bovine serum, 50nmol/L ascorbic acid, 6.25mg/L insulin, 100U/ml penicillin). 100 渭 g/ml streptomycin, 100nmol/L dexamethasone, 10 渭 g / L transforming growth factor 尾 1 (without gentamicin) and four gentamicin containing groups (100 渭 g / ml 100 渭 g 路ml / ml 100 渭 g / ml + 100 渭 g / ml 100 渭 g/ml) and 4 gentam The proliferative activity was compared by MTT colorimetry. The differentiation of chondrocytes was detected by type 鈪，
本文编号：2302383
[Abstract]:Objective to study the ability of rat adipose stem cells (adipose-derived stem cells ADSCs) to differentiate into chondrocytes in vitro, and to observe the effects of different concentrations (0 渭 g / m ~ (10) 渭 g / ml ~ (10) 渭 g 路ml ~ (-1) of 50 渭 g / ml / ml ~ (100 渭 g / ml) on the differentiation of rat adipose stem cells into cartilage. Effects of gentamicin on proliferation and differentiation of cartilage. Methods Adipose tissue was isolated from the inguinal fat pad of 3-week-old SD rats. Stem cells were isolated by type I collagenase digestion. The stem cells were isolated and subcultured in DMEM medium containing fetal bovine serum. The morphology and growth of cells were observed by inverted microscope every day. The third generation of adipose stem cells were divided into five groups, which consisted of a control group (human adipose stem cell culture medium: high glucose DMEM,10% fetal bovine serum, 50nmol/L ascorbic acid, 6.25mg/L insulin, 100U/ml penicillin). 100 渭 g/ml streptomycin, 100nmol/L dexamethasone, 10 渭 g / L transforming growth factor 尾 1 (without gentamicin) and four gentamicin containing groups (100 渭 g / ml 100 渭 g 路ml / ml 100 渭 g / ml + 100 渭 g / ml 100 渭 g/ml) and 4 gentam The proliferative activity was compared by MTT colorimetry. The differentiation of chondrocytes was detected by type 鈪，
本文编号：2302383
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