CAJ | 학술논문

背景：碱性成纤维细胞生长因子在韧带损伤愈合中具有重要作用，细胞因子转染的骨髓间充质干细胞可作为种子细胞应用于韧带组织工程。 　　目的：探讨人碱性成纤维细胞生长因子基因修饰的骨髓间充质干细胞与韧带成纤维细胞三维共培养后的交互生物学效应。 　　方法：原代培养大鼠骨髓间充质干细胞，传至第3代后分为3组：对照组、Ad-EGFP组及Ad-bFGF组。各组细胞相应处理后分别与韧带成纤维细胞三维共培养，MTS法检测细胞增殖能力，ELISA法检测各组细胞上清液中碱性成纤维细胞生长因子蛋白水平，RT-PCR检测各组两种细胞中Scleraxis、Ⅰ型胶原、Ⅲ型胶原、核心蛋白多糖、软骨低聚物基质蛋白等相关基因mRNA表达量的变化。 　　结果与结论：腺病毒介导的碱性成纤维细胞生长因子可高效转染骨髓间充质干细胞；共培养3，6 d后，与对照组及Ad-EGFP组相比，Ad-bFGF组两种细胞增殖活性增强(P<0.01)；上清液中碱性成纤维细胞生长因子表达明显增高(P<0.01)，韧带成纤维细胞中Ⅰ型胶原、Ⅲ型胶原、核心蛋白多糖、软骨低聚物基质蛋白mRNA表达均降低(P<0.01)，骨髓间充质干细胞中Scleraxis、Ⅰ型胶原、Ⅲ型胶原mRNA表达均明显升高(P<0.01)。以上结果表明碱性成纤维细胞生长因子转染的骨髓间充质干细胞与韧带成纤维细胞三维共培养促进韧带成纤维细胞增殖的同时抑制了其胶原合成能力，促进骨髓间充质干细胞增殖的同时增强了其向韧带成纤维细胞分化的能力。
배경：감성성섬유세포생장인자재인대손상유합중구유중요작용，세포인자전염적골수간충질간세포가작위충자세포응용우인대조직공정。 　　목적：탐토인감성성섬유세포생장인자기인수식적골수간충질간세포여인대성섬유세포삼유공배양후적교호생물학효응。 　　방법：원대배양대서골수간충질간세포，전지제3대후분위3조：대조조、Ad-EGFP조급Ad-bFGF조。각조세포상응처리후분별여인대성섬유세포삼유공배양，MTS법검측세포증식능력，ELISA법검측각조세포상청액중감성성섬유세포생장인자단백수평，RT-PCR검측각조량충세포중Scleraxis、Ⅰ형효원、Ⅲ형효원、핵심단백다당、연골저취물기질단백등상관기인mRNA표체량적변화。 　　결과여결론：선병독개도적감성성섬유세포생장인자가고효전염골수간충질간세포；공배양3，6 d후，여대조조급Ad-EGFP조상비，Ad-bFGF조량충세포증식활성증강(P<0.01)；상청액중감성성섬유세포생장인자표체명현증고(P<0.01)，인대성섬유세포중Ⅰ형효원、Ⅲ형효원、핵심단백다당、연골저취물기질단백mRNA표체균강저(P<0.01)，골수간충질간세포중Scleraxis、Ⅰ형효원、Ⅲ형효원mRNA표체균명현승고(P<0.01)。이상결과표명감성성섬유세포생장인자전염적골수간충질간세포여인대성섬유세포삼유공배양촉진인대성섬유세포증식적동시억제료기효원합성능력，촉진골수간충질간세포증식적동시증강료기향인대성섬유세포분화적능력。
BACKGROUND:Basic fibroblast growth factor (bFGF) plays an important role in the ligament tissue healing process, and bone marrow mesenchymal stem cel s transfected with growth factors can be used as seed cel s in ligament tissue engineering. OBJECTIVE:To observe biological effect of bone marrow mesenchymal stem cel s transfected with recombinant adenovirus vectors carrying bFGF in three-dimensional co-culture with ligament fibroblasts. METHODS:Passage 3 bone marrow mesenchymal stem cel s were divided into three groups:control group, Ad-EGFP group and Ad-bFGF group. Under a phase contrast microscope, the changes in cel morphology were observed and the rate of transfection was analyzed by flow cytometry. Proliferation of bone marrow mesenchymal stem cel s and ligament fibroblasts was analyzed by MTS, the expression of bFGF protein in bone marrow mesenchymal stem cel s was determined by ELISA. Scleraxis, col agen type I, col agen type III, decorin and cartilage oligomeric matrix protein levels were detected in BMSCs and ligament fibroblasts using real-time fluorescent quantitative PCR. RESULTS AND CONCLUSION:Recombinant adenovirus-mediated bFGF gene could transfect bone marrow mesenchymal stem cel s efficiently. After co-culture for 3, 6 days, compared with the control group and Ad-EGFP group, in the Ad-bFGF group, the proliferation ability of bone marrow mesenchymal stem cel s and ligament fibroblasts was enhanced (P<0.01), the expression of bFGF protein in supernatant was obviously higher (P<0.01), the col agen type I, col agen type III, decorin and cartilage oligomeric matrix protein mRNA expression decreased in the ligament fibroblasts (P<0.01), but the mRNA expression of Scleraxis, col agen type I, col agen type III in bone marrow mesenchymal stem cel s increased (P<0.01). The results suggest that the co-culture of Ad-bFGF-transfected bone marrow mesenchymal stem cel s with ligament fibroblasts promotes the proliferation of ligament fibroblasts while decreases the col agen synthesis at the same time, and stimulates the proliferation and differentiation of bone marrow mesenchymal stem cel s into ligament fibrolasts.