CM-Dil体外标记人脐带间充质干细胞传代示踪的可行性

背景：掌握人脐带间充质干细胞的移植示踪方法是研究其生物学特性的关键。 目的：观察用CM-Dil标记人脐带间充质干细胞及在体外传代示踪的可行性。 方法：采用酶消化法体外分离培养人脐带间充质干细胞，通过流式细胞仪检测细胞免疫表型和细胞周期、体外成脂成骨诱导鉴定该细胞。将第5代细胞用CM-Dil标记，并将细胞传代，荧光显微镜观察体外标记情况。 结果与结论：第3代人脐带间充质干细胞强表达CD44，CD29，低表达CD106，不表达CD34、CD40；有80%以上的细胞处在G0/G1期，成脂成骨诱导后，油红O染色和碱性磷酸酶染色分别阳性。CM-Dil标记人脐带间充质干细胞细胞标记率达90%以上，体外传代后荧光强度逐渐减退，传8代后，荧光基本消失。说明人脐带间充质干细胞增殖、分化能力强，CM-Dil标记细胞示踪方法简单易行。

Feasibility of in vitro CM-Dil-labeled human umbilical cord mesenchymal stem cells passage tracing

BACKGROUND: The key to study biological characteristics of human umbilical cord mesenchymal stem cells (hUCMSCs) is to master their tracing method of hUCMSCs transplantation. OBJECTIVE: To analyze the tracing feasibility of CM-Dil labeled hUCMSCs in vitro. METHODS: hUCMSCs were isolated and cultured in vitro after enzyme digestion. Immunophenotype and cell cycle were analyzed by flow cytometry, as well as induction of the adipogenic, osteogenic differentiation of hUCMSCs were identified in vitro. The fifth passage of cells were labeled with CM-Dil, passaged and observed by fluorescence microscope in vitro. RESULTS AND CONCLUSION: hUCMSCs at passage 3 were strongly positive for CD44 and CD29, weakly positive for CD106, but negative for CD34 and CD40. Cells in G0/G1 phase accounted for more than 80%. Following adipogenic and osteogenic induction, oil red and alkaline phosphatase staining were positive respectively. The efficiency of CM-DiI labeling hUCMSCs was above 90%. Fluorescence intensity gradually decreased in vitro. After passaged for 8 generations, it nearly quenched. These indicated that hUCMSCs have great potential to proliferation and differentiation. The tracing method of CM-DiI labeling hUCMSCs is effective and easy.