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Published on: August 10, 2018
Subtractive gene expression profiling of articular cartilage and mesenchymal stem cells: serpins as
1Division of Experimental Orthopaedics, Orthopaedic Clinic, University of Heidelberg, Germany.
Objective:
Mesenchymal stem cells (MSCs) are a population of cells broadly discussed to support cartilage repair. The differentiation of MSCs into articular chondrocytes is, however, still poorly understood on the molecular level. The aim of this study was to perform an almost genome-wide screen for genes differentially expressed between cartilage and MSCs and to extract new markers useful to define chondrocyte differentiation stages.
Methods:
Gene expression profiles of MSCs (n=8) and articular cartilage from OA patients (n=7) were compared on a 30,000 cDNA-fragment array and differentially expressed genes were extracted by subtraction. Expression of selected genes was assessed during in vitro chondrogenic differentiation of MSCs and during dedifferentiation of expanded chondrocytes using quantitative and semi-quantitative reverse transcriptase-polymerase chain reaction (RT-PCR). Protein secretion was measured by enzyme-linked immunosorbent assay.
Results:
Eighty-seven genes were differentially expressed between MSCs and cartilage with a more than three-fold difference. Sixty-seven of them were higher expressed in cartilage and among them 15 genes were previously not detected in cartilage. Differential expression was confirmed for 69% of 26 reanalysed genes by RT-PCR. The profiles of three unknown transcripts and six protease-related molecules were characterised during differentiation. SERPINA1 and SERPINA3 mRNA expression correlated with chondrogenic differentiation of MSCs and dedifferentiation of chondrocytes, and SERPINA1 protein levels in culture supernatants could be correlated alike.
Conclusions:
cDNA-array analysis identified SERPINA1 and A3 as new differentiation-relevant genes for cartilage. Since SERPINA1 secretion correlated with both chondrogenesis of MSCs and dedifferentiation during chondrocyte expansion, it represents an attractive marker for refinement of chondrocyte differentiation.
Insights
Researchers identified SERPINA1 and SERPINA3 as key genes in cartilage repair and chondrocyte differentiation. SERPINA1 shows promise as a marker for refining chondrocyte differentiation processes.
Area of Science:
- Biochemistry
- Molecular Biology
- Regenerative Medicine
Background:
- Mesenchymal stem cells (MSCs) are crucial for cartilage repair.
- The molecular mechanisms of MSC differentiation into articular chondrocytes are not fully understood.
Purpose of the Study:
- To conduct a near genome-wide screen for genes differentially expressed between MSCs and articular cartilage.
- To identify novel markers for defining chondrocyte differentiation stages.
Main Methods:
- Compared gene expression profiles of MSCs and osteoarthritis (OA) patient cartilage using a cDNA-fragment array.
- Validated gene expression during in vitro chondrogenesis and chondrocyte dedifferentiation using RT-PCR.
- Measured protein secretion via ELISA.
Main Results:
- Identified 87 differentially expressed genes, with 67 higher in cartilage; 15 were novel.
- Confirmed differential expression for 69% of reanalyzed genes.
- SERPINA1 and SERPINA3 mRNA expression correlated with chondrogenic differentiation and dedifferentiation; SERPINA1 protein levels also correlated.
Conclusions:
- cDNA-array analysis revealed SERPINA1 and SERPINA3 as new differentiation-relevant genes for cartilage.
- SERPINA1 secretion is a potential marker for refining chondrocyte differentiation, tracking both MSC chondrogenesis and chondrocyte expansion.
