本文選題：脂肪干細胞 + TGF-β1　； 參考：《中南大學》2010年碩士論文

【摘要】： 目的本文研究大鼠脂肪干細胞(adipose-derived stem cells ADSCs)在體外分離培養(yǎng)、誘導分化成軟骨細胞的能力,初步觀察不同濃度的TGF-β1對其增殖、分化成軟骨細胞的影響,為軟骨組織工程及臨床應用提供科學依據。 方法從3周齡SD大鼠腹股溝脂肪墊分離出脂肪組織,通過Ⅰ型膠原酶消化法分離干細胞,接種于含有胎牛血清的DMEM培養(yǎng)液中分離傳代。每日以倒置顯微鏡觀察細胞形態(tài)及生長情況。取第三代脂肪干細胞分成5組,包含1個對照組(培養(yǎng)液為人脂肪干細胞完全培養(yǎng)基：高糖DMEM、10%胎牛血清、50nmol/L抗壞血酸、6.25mg/L胰島素、100U/ml青霉素、100μg/ml鏈霉素、100nmol/L地塞米松,不含TGF-β1)和4個TGF-β1濃度組(濃度分別是1、10、40、80μg/L加人脂肪干細胞完全培養(yǎng)基)。采用MTT比色法進行分化和增殖活性比較,Ⅱ型膠原免疫細胞化學染色和細胞甲苯胺藍染色檢測成軟骨細胞分化情況。 結果原代細胞接種4-6小時后即可見沉降貼壁,7-8天左右可見大量增殖,按一定方向性排列呈漩渦狀或束狀,傳代后細胞增殖迅速,生長穩(wěn)定。成軟骨誘導后行細胞甲苯胺藍染色示：10μg/LTGF-β1濃度組陽性,40μg/LTGF-β1濃度組和80μg/LTGF-β1濃度染色較淡,染色細胞少,0μg/LTGF-β1濃度組和1μg/LTGF-β1濃度組為陰性。Ⅱ型膠原免疫組織化學染色顯示：10μg/L TGF-β1濃度組陽性；40μg/LTGF-β1濃度組和80μg/LTGF-β1濃度組為弱陽性；0μg/LTGF-β1濃度組和1μg/LTGF-β1濃度組為陰性。MTT實驗顯示10μg/L的TGF-β1可以促進細胞增殖(P0.05)。 結論1.大鼠脂肪干細胞能向成軟骨細胞誘導分化；2.10μg/L的TGF-β1是脂肪干細胞增殖、分化成軟骨細胞的最佳濃度；3.0、1、40、80μg/L的TGF-β1均不利脂肪干細胞增殖及軟骨細胞形成。
[Abstract]:Objective to study the ability of adipose-derived stem cells ADSCs isolated from rat adipose stem cells to induce the differentiation of chondrocytes in vitro, and to observe the effects of different concentrations of TGF- 尾 1 on the proliferation and differentiation of chondrocytes. To provide scientific basis for cartilage tissue engineering and clinical application. Methods Adipose tissue was isolated from 3 week-old SD rats with inguinal fat pad. Stem cells were isolated by type I collagenase digestion and cultured 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 5 groups, including one control group (human adipose stem cell complete medium: high glucose DMEM10% fetal bovine serum, 50nmol / L ascorbic acid 6.25mg / L insulin / 100U / ml penicillin 100 渭 g/ml streptomycin 100 nmol / L dexamethasone, 100 渭 mol / L ascorbic acid / L ascorbic acid, 100 渭 g/ml streptomycin 100 渭 mol / L dexamethasone. TGF- 尾 _ 1 and 4 TGF- 尾 _ 1 groups were added to human adipose stem cell culture medium with 40 渭 g / L TGF- 尾 _ (1) and 40 渭 g / L TGF- 尾 _ (1), respectively. The differentiation and proliferation activity of chondroblasts were compared by MTT colorimetry. The differentiation of chondroblasts was detected by type 鈪，

本文編號：2047581