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骨向分化诱导来自脐带血细胞刺激与 BMP-2 和 BMP-6。

Chondrogenesis from umbilical cord blood cells stimulated with BMP-2 and BMP-6.

机构信息

Laboratory of Molecular Biology of Cartilage, Division of Rheumatology, Department of Clinical Medicine, State University of Campinas, UNICAMP, R. Vital Brasil, 50, Prédio FCM 08, Campus Universitário Barão Geraldo, Campinas, SP, Brazil.

出版信息

Rheumatol Int. 2013 Jan;33(1):121-8. doi: 10.1007/s00296-011-2328-6. Epub 2012 Jan 12.

Abstract

Umbilical cord blood contains undifferentiated mesenchymal stem cells (MSCs) with chondrogenic potential that may be used for the repair of joint damage. The role of growth factors during the process of chondrogenesis is still not entirely understood. The objective of this study was to evaluate the formation of chondrocytes, cartilaginous matrix and type II collagen from human umbilical cord blood stem cells exposed to two different growth factors, BMP-6 and BMP-2, while being cultured as a micromass or a monolayer. Umbilical cord blood was obtained from full-term deliveries, and then, mononuclear cells were separated and cultured for expansion. Afterward, these cells were evaluated by flow cytometry using antibodies specific for MSCs and induced to chondrogenic differentiation in micromass and monolayer cultures supplemented with BMP-2 and BMP-6. Cellular phenotype was evaluated after 7, 14 and 21 days by RT-PCR and Western blot analysis to identify the type II collagen and aggrecan. The expanded cells displayed surface antigens characteristic of mesenchymal progenitor cells and were negative for hematopoietic differentiation antigens. Type II collagen and aggrecan mRNAs were expressed from day 14 in cells stimulated with BMP-2 or BMP-6. Type II collagen was demonstrated by Western blotting in both groups, and the greatest expression was observed 21 days after the cells were stimulated with BMP-2 cultured in micromass. BMP-2 in micromass culture was more efficient to induce the chondrogenesis.

摘要

脐带血中含有未分化的间充质干细胞(MSCs),具有成软骨潜力,可用于修复关节损伤。在软骨生成过程中,生长因子的作用尚不完全清楚。本研究的目的是评估人脐带血干细胞在培养为微团或单层时暴露于两种不同的生长因子 BMP-6 和 BMP-2 后形成软骨细胞、软骨基质和 II 型胶原的情况。从足月分娩中获取脐带血,然后分离单核细胞并进行培养以扩增。之后,使用针对 MSC 的特异性抗体通过流式细胞术评估这些细胞,并在补充有 BMP-2 和 BMP-6 的微团和单层培养物中诱导其向软骨分化。在第 7、14 和 21 天通过 RT-PCR 和 Western blot 分析评估细胞表型,以鉴定 II 型胶原和聚集蛋白聚糖。扩增的细胞表现出间充质祖细胞的表面抗原特征,并且对造血分化抗原呈阴性。在受到 BMP-2 或 BMP-6 刺激的细胞中,从第 14 天开始表达 II 型胶原和聚集蛋白聚糖 mRNAs。在两组中均通过 Western blot 证明了 II 型胶原的表达,并且在受到 BMP-2 刺激并在微团中培养的细胞中,21 天后观察到最大表达。在微团培养物中 BMP-2 更有效地诱导软骨生成。

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