University of Twente Student Theses


Cartilage repair after joint distraction: intermittent hydrostatic pressure and its influence on mesenchymal stem cells - an in vitro study.

Dijkstra, Koen (2015) Cartilage repair after joint distraction: intermittent hydrostatic pressure and its influence on mesenchymal stem cells - an in vitro study.

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Abstract:Cartilage regeneration is observed after joint distraction in the treatment of severe knee osteoarthritis (Wiegant et al 2013). More than 20%-denuded bone is completely repopulated in 1 year, which cannot be solely the result of matrix synthesis by resident chondrocytes. We hypothesize that MSCs play a trophic role on resident chondrocytes. The intra-articular intermittent hydrostatic pressure (IHP) in the human knee during distraction is thought to induce activation of this trophic effect. The current study evaluates the effect of IHP on in vitro expression of regenerative mediators in hMSCs. Unravelling the fundamental mechanisms behind cartilage repair after joint distraction would have great implications for the treatment of degenerative joint diseases. Materials & methods: The IHP culture setup consists of two vessels, a pressure vessel and a control vessel. IHP between Patm and Patm + 15 kPa, (as found in vivo) is applied to the cells at a frequency of 0.33Hz. The vessels are placed in a regular incubator (T= 37 °C, [O2] = 20%, [CO2] = 5%, RH=95%). Bone marrow derived human MSCs (n=5 donors) monolayer culture was performed in the IHP setup for 6h, 24h and 48h. mRNA expression levels for a set of cartilage-homeostasis related genes were determined, using hypoxanthine phosphoribosyltransferase (HPRT1) as a reference gene. Targets were; fibroblast growth factor 1, 2 and 18 (FGF1, 2, 18), tissue inhibitor of metalloproteinase 1 and 2 (TIMP1,2), transforming growth factor β 1 (TGFβ1), insulin-like growth factor 1 (IGF1) and hepatocyte growth factor (HGF). The comparative Ct method (ΔCt/ΔΔCt) was used to analyse the qPCR data. Articular cartilage explant cultures in hMSC-conditioned medium are performed to determine the potential positive effects of the hMSC secretome. Results: IHP affected gene expression of cartilage-homeostasis related targets with a large inter donor variation. Overall, mild upregulation of FGF2 and TGFβ1 was already seen after 6h under IHP. FGF1 expression shows hardly an effect, and a mild downregulation was seen for FGF18, TIMP1 IGF1 and HGF. TIMP2 shows an interesting shared effect, which is a significant downregulation for all donors. Conclusion: Under IHP a mild early upregulation of FGF2, TGFβ1 and a mild downregulation of FGF18, TIMP1, IGF1 and HGF was seen. TIMP2 is the only target that shows a significant downregulation for all donors. This data suggests a first step in an early regenerative response upon IHP in hMSCs. A first step in isolation of synovial hMSCs was made. Additional experiments are warranted to further elucidate the exact role of IHP on hMCS from different origins.
Item Type:Essay (Master)
Faculty:TNW: Science and Technology
Subject:42 biology, 44 medicine, 50 technical science in general
Programme:Technical Medicine MSc (60033)
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