本文選題：種子細胞 + 脂肪間充質(zhì)干細胞��； 參考：《山西醫(yī)科大學》2011年碩士論文

【摘要】：目的:觀察誘導時機的不同對大鼠脂肪間充質(zhì)干細胞成骨分化的影響。 方法:脂肪間充質(zhì)干細胞(ADSCs)來源于SD大鼠,雌雄不分,體重在150-200g(由山西醫(yī)科大學實驗動物中心提供)。取其腹股溝處脂肪墊,分離、純化、培養(yǎng),傳至第三代,將細胞分為誘導組和非誘導組。誘導組又分為,在細胞接種同時(A組),50%-60%融合時(B組),85%-95%融合時(C組)分別加入同一濃度的由地塞米松、維生素C和β-甘油磷酸鈉組成的等量誘導劑誘導。非誘導組的A、B、C組由普通培養(yǎng)基培養(yǎng)不加入誘導劑。在誘導后的7,14,21,28 d進行鈣鈷染色、茜素紅染色、堿性磷酸酶活性的檢測來判斷各組細胞的成骨能力,并且定期在倒置顯微鏡下觀察細胞形態(tài)變化及生長狀況。 結(jié)果: 1.相同初始接種密度,誘導組細胞生長速度略慢于同期非誘導組。 2.經(jīng)鈣鈷染色后,誘導組大部分染色的細胞呈陽性反應,細胞胞漿中可見淺棕色至棕黑色的細胞顆粒。相同初始接種密度,同期比較,干預時融合度越高陽性細胞越多,以細胞達85%-95%融合時誘導組出現(xiàn)結(jié)節(jié)數(shù)量最多、體積最大;非誘導組未見片狀強陽性細胞及明顯結(jié)節(jié)形成。 3.茜素紅染色后,誘導組從第7d開始可見少量散在紅色結(jié)節(jié),到28d呈現(xiàn)大片紅色致密結(jié)節(jié)。同期相比,以誘導組細胞達85%-95%融合時形成結(jié)節(jié)早,且結(jié)節(jié)數(shù)量多、體積大,非誘導組未見明顯陽性結(jié)節(jié)形成。 4.細細胞內(nèi)堿性磷酸酶活性檢測表明:誘導組與同期非誘導組相比細胞內(nèi)堿性磷酸酶活性明顯提高(P0.05),表現(xiàn)細胞成骨分化特點。誘導組組內(nèi)比較,第7天A組與B組無明顯差別(P0.05),與C組比較有明顯差別(P0.01),C組高;第14與21天A、B、C組間比較均有差異(P0.01),C組B組A組;第28天A組與B、C組比較有差異(P0.05),A組低,B組與C組比較無明顯差異(P0.05)。非誘導組內(nèi)各組比較無明顯差異(P0.05)。 結(jié)論: 1.ADSCs在體外一定條件下經(jīng)過誘導劑誘導后,可向成骨細胞方向分化。 2.初始接種密度相同的情況下,細胞間相互接觸程度越高,成骨能力越強。
[Abstract]:Aim: to observe the effect of different induction time on osteogenic differentiation of adipose mesenchymal stem cells in rats. Methods: adipose mesenchymal stem cells (ADSCs) were derived from SD rats, male and female, weight 150-200g (provided by Experimental Animal Center, Shanxi Medical University). The cells were separated, purified, cultured and transferred to the third generation. The cells were divided into induction group and non-induced group. The induction group was also divided into two groups: group A and group B were induced by dexamethasone, vitamin C and sodium 尾 -glycerophosphate at the same concentration. No inducer was added to the normal culture medium in AZB group (non-induction group). Calcium cobalt staining, alizarin red staining and alkaline phosphatase activity were used to determine the osteogenic ability of the cells at 28 days after induction, and the morphological changes and growth status of the cells were observed under inverted microscope. Results: 1. At the same initial inoculation density, the cell growth rate of the induction group was slightly slower than that of the non-induction group. 2. After calcium and cobalt staining, most of the cells in the induction group showed positive reaction, and light brown and black cell granules were found in the cytoplasm of the cells. At the same initial inoculation density, at the same time, the higher the fusion degree, the more positive cells were in the intervention, and the number and volume of the nodules in the induction group were the largest when the cells reached 85% -95%, but no flake strong positive cells and obvious nodule formation were found in the non-induction group. 3. After alizarin red staining, a small number of red nodules were seen in the induction group from the 7th day, and a large number of red dense nodules appeared on the 28th day. At the same time, 85% to 95% of the cells in the induction group formed the nodules early, and the number and volume of the nodules were large. No positive nodules were found in the non-induced group. 4. The detection of alkaline phosphatase activity in fine cells showed that the activity of alkaline phosphatase in the induction group was significantly higher than that in the non-induction group at the same time, showing the characteristics of osteogenic differentiation. There was no significant difference between group A and group B on the 7th day, but there was a significant difference between group C and group C, and there was significant difference between group A and group B on the 14th and 21st days. On the 28th day, there was no significant difference between group A and group B (P 0.05). There was no significant difference between the groups in the non-induction group (P 0. 05). Conclusion: 1.ADSCs can differentiate into osteoblasts after induced by inducer under certain conditions in vitro. 2. Under the same initial inoculation density, the higher the intercellular contact, the stronger the osteogenic ability.
【學位授予單位】：山西醫(yī)科大學
【學位級別】：碩士
【學位授予年份】：2011
【分類號】：R329

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