【摘要】：目的 研究骨反應(yīng)形態(tài)蛋白（BMP-2）與軟骨細(xì)胞共同誘導(dǎo)骨髓基質(zhì)干細(xì)胞（BMSCs）向軟骨細(xì)胞分化是否有協(xié)同作用,并檢測出BMP-2對BMSCs和軟骨細(xì)胞共培養(yǎng)誘導(dǎo)分化為軟骨細(xì)胞的最佳濃度。 方法 將體外分別培養(yǎng)擴(kuò)增兔BMSCs和軟骨細(xì)胞按相同的比例混合（7：3），，均以5.0×107/ml的細(xì)胞中濃度接種混合培養(yǎng)。根據(jù)不同的細(xì)胞因子BMP-2的濃度分為以下幾組，5ng/ml,10ng/ml,20ng/ml,30ng/ml,40ng/ml，50ng/ml以單獨(dú)培養(yǎng)軟骨細(xì)胞作為陽性對照組，以單獨(dú)培養(yǎng)BMSCs作為陰性對照組，以混合培養(yǎng)后不加BMP-2細(xì)胞因子作為0ng/ml組。加入生長因子培養(yǎng)24h、48h、72h后以四唑鹽（MTT）比色法測細(xì)胞相對數(shù)(OD值)，然后分別計(jì)算BMP-2在不同濃度時(shí)于490nm處的OD值的平均值，并轉(zhuǎn)換為細(xì)胞相對數(shù)，得出細(xì)胞因子BMP-2與軟骨細(xì)胞共同作用于骨髓基質(zhì)干細(xì)胞向軟骨細(xì)胞分化和增殖有協(xié)同作用。然后用免疫細(xì)胞化學(xué)染色法確定骨髓基質(zhì)干細(xì)胞分化后的細(xì)胞有Ⅱ型膠原蛋白表達(dá)為軟骨細(xì)胞。通過對GAG蛋白定量的檢測確定BMP-2與軟骨細(xì)胞共同作用于骨髓基質(zhì)干細(xì)胞向軟骨細(xì)胞分化時(shí)的最佳細(xì)胞因子濃度。 結(jié)果 陰性對照組與0ng/ml組之間OD值的單因素比較P＜0.05，有統(tǒng)計(jì)學(xué)意義。這說明軟骨細(xì)胞單獨(dú)誘導(dǎo)骨髓基質(zhì)干細(xì)胞能引起細(xì)胞數(shù)量的增加，而免疫組化可以證明增加的細(xì)胞能分泌II型膠原蛋白，為軟骨細(xì)胞。當(dāng)BMP-2的濃度在5ng/ml至50ng/ml范圍內(nèi)時(shí)測得的OD值與0ng/ml組的比較也均有統(tǒng)計(jì)學(xué)意義，因而兩種因素共同作用時(shí)骨髓基質(zhì)干細(xì)胞向軟骨細(xì)胞分化顯著提高，并以20ng/ml處理后OD值最高。 結(jié)論 細(xì)胞因子BMP-2與軟骨細(xì)胞共同作用于骨髓基質(zhì)干細(xì)胞對于其向軟骨細(xì)胞分化有協(xié)同作用，且BMP-2與軟骨細(xì)胞共同作用于骨髓基質(zhì)干細(xì)胞向軟骨細(xì)胞分化的最佳作用濃度為20ng/ml。
[Abstract]:Objective to investigate the synergistic effect of bone reactive morphogenetic protein (BMP-2) and chondrocytes on the differentiation of bone marrow stromal stem cells (BMSCs) into chondrocytes. The optimal concentration of BMP-2 on chondrocytes induced by co-culture of BMSCs and chondrocytes was determined. Methods the cultured rabbit BMSCs and chondrocytes were mixed in the same proportion (7:3) in vitro, and cultured in 5. 0 脳 107/ml medium. According to the concentration of different cytokine BMP-2, they were divided into the following groups: 5 ng / ml, 10 ng / ml, 20 ng / ml, 30 ng / ml, 40 ng / ml, 50 ng / ml, respectively, and 50 ng / ml of chondrocytes as the positive control group and BMSCs as the negative control group. The 0ng/ml group was treated with mixed culture without BMP-2 cytokines. Growth factor was added to culture for 24 h, 48 h, 72 h later, the relative cell number (OD) was measured by (MTT) colorimetry, and then the average OD value of BMP-2 at 490nm at different concentrations was calculated and converted to cell relative number. It is concluded that cytokine BMP-2 and chondrocytes have synergistic effects on differentiation and proliferation of bone marrow stromal cells into chondrocytes. Then immunocytochemical staining was used to determine that type 鈪

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