北京鸭肝间充质干细胞分离、培养及分化潜能研究

doi:10.16431/j.cnki.1671-7236.2019.10.009

摘要/Abstract

摘要： 试验旨在研究北京鸭肝间充质干细胞（liver mesenchymal stem cells，LMSCs）的分离、培养、鉴定及多向分化潜能等生物学特性。取孵化19日龄健康鸭胚通过酶消法分离出北京鸭LMSCs，进行体外培养，通过免疫荧光、RT-PCR和流式细胞术等技术对北京鸭LMSCs特异性标记物（CD29、CD44、CD71、CD73、CD90、CD105、CD133及CD166）进行检测，因北京鸭LMSCs分离过程中会存在少量造血干细胞，所以对造血干细胞特异性标记物CD31、CD34进行检测，同时对细胞进行成脂和成骨的分化诱导，检测其多向分化潜能。结果显示，通过酶消法获得的北京鸭LMSCs具有间充质干细胞特异性标记物，造血干细胞特异性标记物为阴性表达，证明其为肝间充质干细胞，且向成脂肪细胞和成骨细胞分化诱导。RT-PCR检测结果发现，成脂分化诱导中，过氧化物酶体增殖因子活化受体-γ（peroxisome proliferator-activated receptor γ，PPAR-γ）、脂蛋白酶（lipoprotein lipase，LPL）呈阳性表达；成骨分化诱导中，Ⅰ型胶原蛋白（Collagen Ⅰ）和Runt相关转录因子2（Runt-related transcription factor 2，RUNX2）呈阳性表达。北京鸭LMSCs较强的自我增殖能力可使畜禽遗传资源有无限利用的可能性，多向分化潜能强则说明重编程的水平较低，对畜禽种质资源的复原有巨大的优势，为组织再生研究提供了一定的理论基础，对于畜禽资源的开发与利用打开了一个更为宽广的平台。

关键词: 北京鸭; 间充质干细胞; 生物学特性; 诱导分化; 肝脏

Abstract: The aim of this study was to study the isolatation,culture,identification and multi-lineage differentiation potential of liver mesenchymal stem cells (LMSCs) in Beijing duck.LMSCs were isolated from 19-day-old healthy Beijing duck embryos by enzymatic digestion and cultured in vitro.The specific markers (CD29,CD44,CD71,CD73,CD90,CD105,CD133 and CD166) of LMSCs were detected by immunofluorescence,RT-PCR and flow cytometry.Because the LMSCs were mixed with some hematopoietic stem cells,CD31 and CD34 which were the specific markers of hematopoietic stem cells were detected.Furthermore,LMSCs were induced to differentiate into adipogenic and osteogenic cells to detect their multi-lineage differentiation potential.The results showed that LMSCs obtained by enzymatic digestion expressed specific markers of mesenchymal stem cells,and the specific markers of hematopoietic stem cells were negative.It was proved that LMSCs were hepatic mesenchymal stem cells,and differentiated into adipocytes and osteoblasts.RT-PCR detection results revealed that adipogenic cells expressed peroxisome proliferator-activated receptor γ (PPAR-γ) and lipoprotein lipase (LPL),osteogenic cells expressed Collogen Ⅰ and Runt-related transcription factor 2 (RUNX2).The strong self-proliferation ability of Beijing duck LMSCs made it possible to make unlimited use of livestock and poultry genetic resources.The strong multi-directional differentiation potential indicated that the level of reprogramming was low,and it had great advantages for the recovery of livestock and poultry germplasm resources.It provided a theoretical basis for tissue regeneration research and opened up a broader platform for the development and utilization of livestock and poultry resources.

Key words: Beijing duck; mesenchymal stem cells; biological characteristics; induced differentiation; liver

中图分类号:

S852.16⁺5

李婷婷, 许龙, 梁峻. 北京鸭肝间充质干细胞分离、培养及分化潜能研究[J]. 中国畜牧兽医, 2019, 46(10): 2882-2889.

LI Tingting, XU Long, LIANG Jun. Study on Isolation,Culture and Differentiation Potential of Liver Mesenchymal Stem Cells in Beijing Duck[J]. China Animal Husbandry and Veterinary Medicine, 2019, 46(10): 2882-2889.

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