鸡骨髓源造血干细胞的体外培养及分化诱导  

Culture and Differentiation Induction of Chicken Bone Marrow Derived Hematopoietic Stem Cell in vitro

作　　者：何书海[1,2] 张晒 焦凤超 位格格[1] 王旭 黄守孝 董建国 HE Shuhai;ZHANG Shai;JIAO Fengchao;WEI Gege;WANG Xu;HUANG Shouxiao;DONG Jianguo(College of Animal Science and Technology,Xinyang University of Agriculture and Forestry,Xinyang,He′nan 464000;Henan Provincial Waterfowl Resources Development and Utilization and Disease Control Engineering Technology Research Center,Xinyang,He′nan 464000)

机构地区：[1]信阳农林学院动物科技学院,河南信阳464000 [2]河南省水禽资源开发利用与疫病防控工程技术研究中心,河南信阳464000

出　　处：《中国家禽》2023年第10期6-11,共6页China Poultry

基　　金：河南省重点研发与推广专项(科技攻关)(232102111043);信阳农林学院家禽重大疫病防控科技创新团队(2022CXTD06)。

摘　　要：为获取鸡骨髓源造血干细胞(HSC)和建立其体外培养方法,试验从鸡长骨中获取骨髓单个核细胞,流式细胞术分选出CD34^(+)HSC后,在体外进行培养和诱导分化,进一步观察CD34^(+)HSC的形态、生长状态、集落形成能力和分化能力。结果显示:IMDM培养基中添加50 ng/mL干细胞生长因子、30 ng/mL Flt-3配体、10μg/mL白细胞介素-3、50 ng/mL白细胞介素-6以及20%鸡血清可维持鸡CD34^(+)HSC的短期体外培养,最长可存活10 d。将上述培养体系加入到甲基纤维素半固体培养基中,可使鸡CD34^(+)HSC形成典型的干细胞克隆集落;在上述培养体系中加入50 ng/mL血小板生成素和80 ng/mL的粒细胞集落刺激因子,可诱导鸡CD34^(+)HSC向粒系细胞分化。瑞氏-姬姆萨染色结果显示,鸡CD34^(+)HSC的胞核较大,呈蓝紫色,核染色质细而分散,胞浆呈浅蓝色,均匀无颗粒。研究提示:在培养体系中添加鸡造血细胞生长因子,可实现鸡骨髓源CD34+HSC的体外扩增和定向诱导分化。To obtain chicken bone marrow derived hematopoietic stem cell(HSC)and establish their culture methods in vi⁃tro,bone marrow mononuclear cells were obtained from chicken long bones,and CD34^(+)HSC were separated by flow cytometer.These cells were cultured and induced to differentiate in vitro,and the morphology,growth state,colony forming ability and differ⁃entiation ability of CD34^(+)HSC were studied.The results showed that the IMDM medium with 50 ng/mL stem cell growth factor,30 ng/mL Flt-3 ligand,10μg/mL interleukin 3,50 ng/mL interleukin 6 and 20%chicken serum could ensure the survival and cloning of chicken CD34^(+)HSC in vitro.The above culture system was able to maintain the short-term culture of chicken CD34^(+)HSC,which could survive for up to 10 days.The methyl cellulose semi-solid medium with above culture system could make chicken CD34^(+)HSC form a typical stem cell clone colony.Adding 50 ng/mL thrombopoietin and 80 ng/mL granulocyte colony stimulating factor to the above culture system could induce chicken CD34^(+)HSC to differentiate into granulocytes.The results of Wright′s-Giemsa staining showed that the nucleus of chicken CD34^(+)HSC was large and blue purple,the nuclear chromatin was fine and scattered,the cytoplasm was light blue,and there were no granules.The results suggested that adding growth factors of chicken hematopoietic to the culture system could realize in vitro expansion and directional differentiation of chicken bone mar⁃row derived CD34+HSC.

关 键 词：鸡 骨髓 造血干细胞 分离培养 分化诱导 

分 类 号：S831.2[农业科学—畜牧学]