Artikel
RNA-Seq based identification of factors involved in knee joint regeneration after induced osteoarthritis in Notophthalmus viridescens
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Veröffentlicht: | 12. September 2014 |
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Background: In mammals, tissue damage often results in fibrotic scar formation, which in turn can significantly impair organ function. In contrast to mammals, many urodele amphibians including the red-spotted newt Notophthalmus viridescens are able to restore damaged organs or even lost limbs or tails. In our working group, the newt has been established as a model organism to study endogenous knee joint regeneration in adult vertebrates. After osteoarthritis (OA) induction, newts display similar symptoms of disease, including joint instability and luxation. In contrast to mammals, newts were able to restore their joint function within 3 months. To evaluate the molecular pathways involved during the regenerative process, an RNA-Seq analysis was performed.
Methods: Intra-articular injection of collagenase, mono-iodoacetate or surgical manipulation of articular cartilage was performed to induce OA in newts. To elucidate gene expression changes on the transcriptional level during regeneration, a quantitative transcriptome analysis after artificially induced knee joint damage using Illumina® next-generation sequencing (NGS) was performed. All OA models were used for comparative evaluation of the respective molecular pathways. Currently, candidate genes are being selected using public available databases, and pathway analyses of the gained data have being performed. Deregulation of selected candidate genes have been found on the mRNA level (Real Time PCR) and protein level (immunohistochemistry).
Results: Several matricellular proteins including tenascin-C (TN-C), CCN family members and their related pathways were found to be differentially regulated during the regenerative process in all three OA models. Deregulation of these candidates was verified on the mRNA level by Real Time PCR in OA vs. control knees of the newts. The spatiotemporal expression pattern during newt knee joint regeneration of these candidates will further be investigated on the protein level.
Conclusion: Extracellular matrix proteins are known to provide conclusive cues during regenerative or repair processes. Identification of novel proteins and evaluation of their impact on cellular behaviour including their effects on cellular adhesion or migration can help to understand the pathways involved guiding regeneration.