【摘要】： 目的: 探討軟骨細胞和骨髓間充質干細胞(BMSCs)混合培養(yǎng)對構建組織工程軟骨的影響,并確定兩者的最佳比例。 方法: 取一月齡新西蘭兔六只,雌雄不限。體外分離培養(yǎng)擴增兔關節(jié)軟骨細胞和BMSCs。收獲第一代末的軟骨細胞和BMSCs,按不同比例(軟骨細胞/BMSCs:4/1,2/1,1/1,1/2,1/4,單純軟骨細胞)混合培養(yǎng)一代。以4×107/ml的細胞終濃度接種40μl于聚乳酸聚羥基乙酸聚合物支架( PLGA:4mm×4mm×2mm)上靜態(tài)培養(yǎng)2天,移入周期性壓力場下培養(yǎng)(壓力:0～200kPa ,頻率:0.1 Hz,時間:8h/d)三周。三周后觀察各組組織工程軟骨的大小形態(tài)質地光澤和組織彈性;HE染色法觀察組織工程軟骨細胞增殖及分布;甲苯胺藍染色法觀察氨基糖胺聚糖(GAG)的分泌及分布狀況,并用1,9二甲基亞甲藍法定量檢測GAG的含量;采用Ⅱ型膠原免疫組織化學法觀察Ⅱ型膠原的分泌及分布,并用image-proplus6.0圖象分析系統(tǒng)對Ⅱ型膠原染色面積行半定量分析;比色法測定組織工程軟骨的DNA含量觀察組織工程軟骨的細胞增殖情況。 結果: 軟骨細胞和BMSCs混合培養(yǎng)組與單純軟骨細胞組相比,體積較大,表面光滑,有彈性,有光澤。組織學顯示混合培養(yǎng)組結構致密,細胞外基質分布更均勻,其中2/1組可見軟骨陷窩�；旌吓囵B(yǎng)組的Ⅱ型膠原染色面積、GAG含量、DNA含量高于單純軟骨細胞組,其中2/1組含量最高,差異有統(tǒng)計學意義(P0.05)。 結論: 軟骨細胞和BMSCs混合培養(yǎng)能顯著提高組織工程軟骨的質量,其中軟骨細胞和BMSCs的濃度比為2/1時效果最好。
[Abstract]:Aim: to investigate the effect of co-culture of chondrocytes and bone marrow mesenchymal stem cells (BMSCs) on the construction of tissue engineered cartilage and determine the optimal ratio of the two. Methods: six New Zealand rabbits (male and female) of one month old were selected. Isolation and amplification of rabbit articular chondrocytes and BMSCs. in vitro The first generation of chondrocytes and BMSCs, were harvested and cultured in different proportion (chondrocytes / BMSCs:4/1,2/1,1/1,1/2,1/4, pure chondrocytes). The cells were inoculated with 40 渭 l of 4 脳 107/ml cells in PLGA:4mm 脳 4mm 脳 2mm for 2 days, then cultured under cyclic pressure (pressure: 0~200kPa, frequency: 0. 1 Hz, time: 8h/d) for three weeks. The size of tissue engineered cartilage in each group was observed three weeks later. Form? Texture? HE staining was used to observe the proliferation and distribution of tissue engineered chondrocytes, toluidine blue staining was used to observe the secretion and distribution of aminoglycosaminoglycan (GAG), and the content of GAG was detected by 1- (9) dimethylmethylene blue method. The secretion and distribution of type 鈪，

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