摘要
目的:观察不同浓度富血小板血浆(PRP)对兔骨髓间充质干细胞体外增殖及向软骨细胞分化的影响。方法:将兔骨髓间充质干细胞培养在水凝胶三维环境内,并按培养液的不同分为4组:空白对照组采用DMEM培养液培养,加入不含血小板的血浆;5%PRP组采用含5%PRP的DMEM培养液培养;10%PRP组采用含10%PRP的DMEM培养液培养;20%PRP组采用含20%PRP的DMEM培养液培养。MTT法检测骨髓间充质干细胞的增殖情况,苏木精-伊红(HE)染色实验及Ⅱ型胶原免疫组化染色分析骨髓间充质干细胞向软骨分化情况。结果:空白对照组细胞增殖呈递增趋势,而含富血小板的3组从培养的第3天开始快速增殖,第6天进入增殖平台期。于第3天到第7天,3组含PRP组细胞数多于空白对照组,其中10%PRP组细胞数最多(均P<0.05)。3组含PRP组均能观察到典型的软骨细胞形态,软骨细胞呈椭圆形或卵圆形,成熟的软骨细胞分布于软骨陷窝内,3组PRP组的软骨样细胞数量均多于空白对照组(均P<0.001),20%PRP组的软骨样细胞数量多于5%PRP组(P<0.001),10%PRP组软骨细胞数量多于5%PRP组和20%PRP组(均P<0.001),空白对照组呈阴性表达,含PRP的3组Ⅱ型胶原均呈阳性表达,3组PRP组阳性面积占比均大于空白对照组(均P<0.001),10%PRP组阳性面积占比大于5%及20%PRP组(P<0.001)。结论:PRP对骨髓间充质干细胞体外增殖有促进作用,并能诱导向软骨细胞分化,浓度为10%PRP组作用较明显。
Objective:To observe the effect of different concentrations of platelet-rich plasma on the proliferation and differentiation of rabbit bone marrow mesenchymal stem cells into chondrocytes in vitro.Methods:The third generation rabbit bone marrow mesenchymal stem cells were cultured in a three-dimensional hydrogel environment,and divided into 4 groups according to different culture medium:Blank control group,in which the cells cultured in DMEM medium with platelet-free plasma;5%platelet rich plasma(PRP)group,10%PRP group,and 20%PRP group,in which the cells culture in DMEM medium containing 5%PRP,10%PRP,and 20%PRP,respectively.The proliferation of cells was detected by MTT assay.The differentiation of bone marrow mesenchymal stem cells into cartilage was analyzed by HE staining and typeⅡcollagen immunohistochemical staining.Results:The proliferation of cells in the blank control group was increased,whereas the proliferation was rapidly increased from the third day of culture and entered the proliferation plateau on the sixth day in PRP groups.On day 3 to day 7,the number of cells in the PRP group was more than that in the blank control group,10%PRP group had the largest number of cells(P<0.05).Typical chondrocyte morphology could be observed in the three groups containing PRP.The chondrocytes were oval,mature chondrocytes were distributed in the cartilaginous recess,and the number of chondrocytes in the PRP group was higher than that in the blank control group(P<0.001).The number of chondrocytes in 20%PRP group was more than 5%PRP group(P<0.001),and the number of capillary cells in the 10%PRP group was more than other 3 groups(P<0.001).The blank control group was negative,and the three groups which containing PRP with typeⅡcollagen showed positive(P<0.001).The proportion of positive area in the three PRP groups was greater than that in the blank control group(P<0.001),and the proportion of positive area in the 10%PRP group was greater than 5%PRP group and 20%PRP group(P<0.001).Conclusion:The platelet-rich plasma
作者
林春博
姚军
农桔安
方徳鹏
吴立蔚
杨渊
余雪
Lin Chunbo;Yao Jun;Nong Ju’an;Fang Depeng;Wu Liwei;Yang Yuan;Yu Xue(Guangxi Medical University Affiliated Langdong Hospital,Guangxi Medical University Kaiyuan Langdong Hospital,Nanning 530028,China;The First Affiliated Hospital of Guangxi Medical University,Nanning,530021,China;The Third Affiliated Hospital of Guangxi Medical University,Nanning 530031,China;Guangxi Medical University Affiliated Tumor Hospital,Nanning 530021,China;Guangxi Medical University,Nanning 530021,China;The People's Hospital of Guilin,Guilin 541002,China)
出处
《广西医科大学学报》
CAS
2020年第12期2147-2152,共6页
Journal of Guangxi Medical University
基金
国家自然科学基金资助项目(No.81760390)
广西医科大学青年科学基金资助项目(No.GXMUYSF201717)
广西壮族自治区卫生和计划生育委员会自筹科研课题资助项目(No.Z20181037)
广西医药卫生科研课题资助项目(No.Z20180803)。
关键词
富血小板血浆
骨髓间充质干细胞
软骨分化
HE染色
Ⅱ型胶原
platelet-rich plasma
bone marrow mesenchymal stem cells
cartilage differentiation
HE staining
typeⅡcollagen
