Xu Zi-xing, Chen Jian-ting, Li Tao, Zha Ding-sheng, Zhang Xin-xin, Jiang Xiao-rui, Xiao Wen-de, Zhu Qing-an
Department of Spinal and Orthopedic Surgery, Nanfang Hospital, Southern Medical University, Guangzhou 510515, China.
Nan Fang Yi Ke Da Xue Xue Bao. 2011 Feb;31(2):289-94.
To study the changes in the biological behavior of bone marrow mesenchymal stem cells (BMSCs) transfected with red fluorescent protein by lentivirus (RFP-BMSCs) seeded on in poly-D, L-lactide acid (PDLLA) scaffolds with bioactive modification by ammonia plasma and Gly-Arg-Gly-Asp-Ser (GRGDS) in vitro.
Circular sheets of PDLLA scaffolds (8 mm in diameter and 1 mm in thickness) were prepared and aminated with PDLLA (group A) or modified with the peptide conjugate A/PDLLA (group PA), with untreated PDLLA as the control (group P). The RFP-BMSCs were seeded on the scaffold materials and their proliferation and metabolic activity were detected using CyQuant NF and Alamar blue staining. The mineralization on the scaffolds was observed using calcein fluorescent dye under a fluorescent microscope. The adhesion and proliferation of RFP-BMSCs were observed by fluorescent microscope, and scanning electron microscope (SEM) was used to confirm the observed adhesion of the seed cells.
The RFP-BMSCs seeded on the 3 scaffolds all showed proliferative activity at different time points after cell seeding, and the cell numbers decreased significantly in the order of PA>A>P (P<0.001). The cell number was significantly greater in group PA than in group A at all the time points except for days 10 (P=0.077) and 12 (P=0.491), and gradually became similar with the passage of time. The metabolic changes of the cells follow a similar pattern of cell proliferation. RFP-BMSCs showed more active proliferation in group A and group PA than in group P. On days 14 and 21, the intensity of green fluorescence decreased in the order of group PA, A and P. The RFP-BMSCs showed better adhesion in group PA than in group A, and the cells in group P appeared more scattered under scanning electron microscope.
Bioactive modification of PDLLA by ammonia treatment and conjugation with GRGDS peptides may promotes the adhesion, proliferation, metabolism and mineralization of RFP-BMSCs seeded on PDLLA scaffolds.
研究慢病毒转染红色荧光蛋白的骨髓间充质干细胞（RFP - BMSCs）接种于经氨等离子体和甘氨酸 - 精氨酸 - 甘氨酸 - 天冬氨酸 - 丝氨酸（GRGDS）进行生物活性修饰的聚 - D，L - 乳酸（PDLLA）支架上后，其生物学行为的变化。
制备直径8mm、厚度1mm的PDLLA支架圆片，用PDLLA进行胺化处理（A组）或用肽偶联物A/PDLLA进行修饰（PA组），未处理的PDLLA作为对照（P组）。将RFP - BMSCs接种于支架材料上，使用CyQuant NF和Alamar蓝染色检测其增殖和代谢活性。在荧光显微镜下用钙黄绿素荧光染料观察支架上的矿化情况。通过荧光显微镜观察RFP - BMSCs的黏附与增殖情况，并用扫描电子显微镜（SEM）确认观察到的种子细胞黏附情况。
接种于3种支架上的RFP - BMSCs在细胞接种后的不同时间点均表现出增殖活性，细胞数量按PA>A>P的顺序显著减少（P<0.001）。除第10天（P = 0.077）和第12天（P = 0.491）外，PA组在所有时间点的细胞数量均显著多于A组，且随着时间推移逐渐趋于相似。细胞的代谢变化遵循与细胞增殖相似的模式。RFP - BMSCs在A组和PA组中的增殖比P组更活跃。在第14天和第21天，绿色荧光强度按PA组、A组和P组的顺序降低。RFP - BMSCs在PA组中的黏附比A组更好，P组细胞在扫描电子显微镜下显得更分散。
氨处理和与GRGDS肽偶联对PDLLA进行生物活性修饰可能促进接种于PDLLA支架上的RFP - BMSCs的黏附、增殖、代谢和矿化。
