论文部分内容阅读
骨髓是间充质干细胞(MSC)的重要来源。研究显示,标准密度梯度离心法分离骨髓MSC的效率不高,骨髓小粒是造成该法低效的原因。通过组织块法分离骨髓小粒,再结合标准法,可从单份骨髓标本分离获得更多MSC,然而这种方法费时费力。本研究探求分离骨髓MSC更简单、更有效的方法。收集7例正常人骨髓标本,低速离心沉降,用改良组织块法培养漂浮在液面上层骨髓小粒中的MSC,用直接铺种法培养沉降在底层的MSC。然后对MSC进行免疫表型和分化能力鉴定。结果表明:在7例标本中,有3例骨髓小粒漂浮在液面上层,其余骨髓细胞包括部分小粒沉降在底层;漂浮小粒中的MSC可用改良组织块法获得,沉降在底层的MSC可用直接铺种法获得。分离的MSC传代3次后不表达CD45、CD34,表达CD105、CD73、CD44、CD90、CD49e,能向软骨和脂肪细胞分化。结论:以直接铺种法为主、改良组织块培养法为辅,可简单、高效地分离人骨髓中MSC。
Bone marrow is an important source of mesenchymal stem cells (MSCs). Studies have shown that standard density gradient centrifugation is not efficient in isolating bone marrow MSCs, and bone marrow pellets are responsible for inefficiencies in this method. Segregation of bone marrow pellets by tissue block, combined with the standard method, allows the isolation of more MSCs from a single bone marrow specimen, however, this method is time-consuming and laborious. This study explored the isolation of bone marrow MSC more simple and more effective method. Seven normal bone marrow samples were collected and centrifuged at low speed. MSCs floating in the supernatant bone marrow granules were cultured by modified tissue block method and cultured in the bottom MSC by direct seeding method. Then, the immune phenotype and differentiation ability of MSCs were identified. The results showed that: in 7 specimens, 3 cases of bone marrow floating in the upper surface of the liquid, the rest of the bone marrow cells, including some of the small particles of sedimentation in the bottom; floating particles in the MSC can be improved tissue block method, settlement in the bottom of the MSC available direct shop Kind of law obtained. Isolated MSCs did not express CD45 and CD34 three times after passage, and expressed CD105, CD73, CD44, CD90 and CD49e, which differentiated into cartilage and adipocytes. Conclusion: The main method of direct seeding, supplemented by improved tissue culture method, can be simple and efficient separation of human bone marrow MSC.