Identification of Subpopulations With Characteristics of Mesench
Identification of Subpopulations With Characteristics of Mesench
We first identified and isolated cellular subpopulations with characteristics of mesenchymal progenitor cells (MPCs) in osteoarthritic cartilage using fluorescence-activated cell sorting (FACS). Cells from osteoarthritic cartilage were enzymatically isolated and analyzed directly or after culture expansion over several passages by FACS using various combinations of surface markers that have been identified on human MPCs (CD9, CD44, CD54, CD90, CD166). Culture expanded cells combined and the subpopulation derived from initially sorted CD9, CD90, CD166 cells were tested for their osteogenic, adipogenic and chondrogenic potential using established differentiation protocols. The differentiation was analyzed by immunohistochemistry and by RT-PCR for the expression of lineage related marker genes. Using FACS analysis we found that various triple combinations of CD9, CD44, CD54, CD90 and CD166 positive cells within osteoarthritic cartilage account for 212% of the total population. After adhesion and cultivation their relative amount was markedly higher, with levels between 24% and 48%. Culture expanded cells combined and the initially sorted CD9/CD90/CD166 triple positive subpopulation had multipotency for chondrogenic, osteogenic and adipogenic differentiation. In conclusion, human osteoarthritic cartilage contains cells with characteristics of MPCs. Their relative enrichment during in vitro cultivation and the ability of cell sorting to obtain more homogeneous populations offer interesting perspectives for future studies on the activation of regenerative processes within osteoarthritic joints.
Mesenchymal progenitor cells (MPCs) from bone marrow are able to differentiate in various types of connective tissue, including cartilage, bone and adipose tissue. This led to more precise characterization of these cells by analysis of cell surface markers and differentiation related gene expression. In parallel, it was recognized that MPCs not only reside in bone marrow but also in various other connective tissues, such as periost, and adipose and muscle tissue. Cells within the joint that are capable of differentiating into chondrocytes, osteoblasts and adipocytes were recently described in synovia, patellar fat pad and articular cartilage.
In the present study we purified progenitor-like cells from the cartilage of human osteoarthritic joints and showed that these cells are capable of proliferation and osteogenic, adipogenic and chondrogenic lineage progression. Those cells could be distinguished from articular chondrocytes by simultaneous staining with several triple combinations of cell surface antigens. We used these marker sets for quantification of MPCs by flow cytometric analysis in the original cell population and after in vitro cultivation. Finally, we sorted these cells according to the expression of triplicate surface markers and demonstrated that this subpopulation is capable of osteogenic, adipogenic and chondrogenic differentiation. These findings should provide a basis for identification of MPCs in articular cartilage and for studies of their roles in joint physiology and disease, as well as in induction of regenerative processes within osteoarthritic joints.
We first identified and isolated cellular subpopulations with characteristics of mesenchymal progenitor cells (MPCs) in osteoarthritic cartilage using fluorescence-activated cell sorting (FACS). Cells from osteoarthritic cartilage were enzymatically isolated and analyzed directly or after culture expansion over several passages by FACS using various combinations of surface markers that have been identified on human MPCs (CD9, CD44, CD54, CD90, CD166). Culture expanded cells combined and the subpopulation derived from initially sorted CD9, CD90, CD166 cells were tested for their osteogenic, adipogenic and chondrogenic potential using established differentiation protocols. The differentiation was analyzed by immunohistochemistry and by RT-PCR for the expression of lineage related marker genes. Using FACS analysis we found that various triple combinations of CD9, CD44, CD54, CD90 and CD166 positive cells within osteoarthritic cartilage account for 212% of the total population. After adhesion and cultivation their relative amount was markedly higher, with levels between 24% and 48%. Culture expanded cells combined and the initially sorted CD9/CD90/CD166 triple positive subpopulation had multipotency for chondrogenic, osteogenic and adipogenic differentiation. In conclusion, human osteoarthritic cartilage contains cells with characteristics of MPCs. Their relative enrichment during in vitro cultivation and the ability of cell sorting to obtain more homogeneous populations offer interesting perspectives for future studies on the activation of regenerative processes within osteoarthritic joints.
Mesenchymal progenitor cells (MPCs) from bone marrow are able to differentiate in various types of connective tissue, including cartilage, bone and adipose tissue. This led to more precise characterization of these cells by analysis of cell surface markers and differentiation related gene expression. In parallel, it was recognized that MPCs not only reside in bone marrow but also in various other connective tissues, such as periost, and adipose and muscle tissue. Cells within the joint that are capable of differentiating into chondrocytes, osteoblasts and adipocytes were recently described in synovia, patellar fat pad and articular cartilage.
In the present study we purified progenitor-like cells from the cartilage of human osteoarthritic joints and showed that these cells are capable of proliferation and osteogenic, adipogenic and chondrogenic lineage progression. Those cells could be distinguished from articular chondrocytes by simultaneous staining with several triple combinations of cell surface antigens. We used these marker sets for quantification of MPCs by flow cytometric analysis in the original cell population and after in vitro cultivation. Finally, we sorted these cells according to the expression of triplicate surface markers and demonstrated that this subpopulation is capable of osteogenic, adipogenic and chondrogenic differentiation. These findings should provide a basis for identification of MPCs in articular cartilage and for studies of their roles in joint physiology and disease, as well as in induction of regenerative processes within osteoarthritic joints.
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