DiI荧光标记的软骨细胞与支架材料复合后的观察

Observation of the DiI labeled chondrocytes combined with scaffold by fluorescent microscope

目的:通过DiI荧光染料标记观察软骨细胞在聚乳酸/乙醇酸共聚物支架(PLGA)体外和体内培养环境中的生长状态,为研究组织工程化软骨探索一种理想的软骨细胞示踪方法。方法:体外分离培养大鼠剑突软骨细胞,应用DiI标记软骨细胞,通过MTT法测定细胞的增殖活性。DiI荧光标记的软骨细胞种植于PLGA支架上分两组:一组体外培养1周,另一组体外培养1周后,再植入同系大鼠大网膜体内培养1周。分别取出细胞-支架复合物,在荧光显微镜下观察体外和体内条件下软骨细胞的荧光表达情况。结果:应用DiI标记不影响软骨细胞的增殖,MTT测定结果显示标记组和对照组的A值未见显著性差异(P>0.05)。标记后的软骨细胞显示环状红色荧光,胞核未着色。体外培养和体内培养的软骨细胞-支架复合物,均可在荧光显微镜下观察到红色荧光表达。结论:DiI荧光染料能够有效标记软骨细胞,标记的细胞-支架复合物可直接在荧光显微镜下进行观察,可作为体外和体内构建组织工程软骨的较好的示踪方法。

Aim ： To find out a ideal cell labeling method for the study of tissue engineering cartilage, through observing the growth state of chondroeytes labeled by DiI and seeded on DegraPol scaffold in vitro and in vivo. Methods： Chondrocytes isolated from rat xyphoid was labeled by Dil, MTT test was used to determine the proliferation status of chondrocytes. Chondrocytes labeled by DiI were seeded onto PLAG scaffold, cultivated in vitro 1 week and cultivated in vivo 1 week after 1 week in vitro culture, respectively. The growth state of the DiI labeled cells by fluorescent microscope. Results： The fluoresce dye DiI showed no effect on the living status of chondrocytes. MTr test showed there was no significant difference between the proliferation of DiI labeled and unmarked chondrocytes. The ehondroeytes labeled by DiI were observed clearly by fluorescent microscope. Conclusion： The chondrocytes labeled by DiI are well compatible with PLGA scaffold and observed continuously by fluorescent microscope. DiI label can be used as a good tracing method to observe tissue engineering cartilage.