DOS BULLETIN - Dansk Ortopædisk Selskab
DOS BULLETIN - Dansk Ortopædisk Selskab DOS BULLETIN - Dansk Ortopædisk Selskab
2010-378_DOS nr. 3 2010 29/09/10 10:08 Side 124 Different responses to hypoxia of mesenchymal stem cells compared to chondrocytes Casper Bindzus Foldager, Anna Bay Nielsen, Bjørn Borsøe Christensen, Moustapha Kassem, Cody Bünger, Martin Lind Orthopaedic Research Lab, Aarhus University Hospital; Department of Endocrinology and Metabolism, University Hospital of Odense, Sports Trauma Clinic, Aarhus University Hospital Background: Articular cartilage is a specialized tissue in which the native chondrocytes are exposed to low oxygen tensions. This hypoxic environment is essential for the chondrogenic viability. Hypoxia inducible factors (HIFs) are believed to play an essential role in the adaption to this environment. The most commonly used treatment for cartilage defects is microfracture, which relies on mesenchymal stem cells (MSCs) to differentiate into chondrocytes for subsequent cartilage production. Purpose: The aim of this study is to investigate the response of hypoxia on normal and differentiating MSCs compared to native chondrocytes. Methods: Three cell cultures were used: 1. Human chondrocytes; 2. Immortalized (TERT-) MSCs in chondrogenic differentiation medium (diff); 3. TERT-MSCs in standard medium(std). The three groups were divided into three oxygen tensions: 21%, 15%, and 5%. They were cultured in groups for 1, 2, 7, and 14 days. Quantitative gene expression analysis was obtained using qRT-PCR and protein synthesis was measured with ELISA. Statistics were performed with ANOVA with repeated measures. Findings: Chondrocytes and TERT-MSCs(diff) did not show an increase in HIF-1alpha and HIF- 2aplha with lowering of oxygen from 21% to 5%. However, TERT-MSCs(std) showed a significant 3-fold increase. The expression of VEGF in chondrocytes showed 5-fold increase while TERT-MSC(diff) showed a 8- fold increase and TERT-MSCs a 12-fold increase. Conclusion: Vascularization of regenerating cartilage is a main contributor to the generation of fibrous tissue and failed cartilage formation. This is the first study to describe the unfortunate response of stem cells compared to chondrocytes, which might contribute significantly to the understanding of impaired regeneration of cartilage using microfracture. 124
2010-378_DOS nr. 3 2010 29/09/10 10:08 Side 125 Bioreactor activated graft material for early implant fixation in bone Susan Snoek Henriksen, Ming Ding, Søren Overgaard Orthopaedic Research Unit, Dept. of Orthopaedic Surgery and Traumatology, Odense University Hospital Background: Combined incubation of a composite scaffold with bone marrow stromal cells in a perfusion bioreactor, could make up a novel hybrid graft material with optimal properties for early fixation of bone to implant. Purpose: The aim of this study was to create a bioreactor activated graft (BAG) material, which can induce early fixation of bone to implant, similar to that of allograft. Methods: Mononuclear cells were isolated from the bone marrow of 8 female sheep, and injected into a perfusion bioreactor with poly-lactic acid (PLA 12%) coated scaffold granules (HA/â-TCP). Hybrid material was incubated in the bioreactor for 2 weeks. Porous titanium alloy implants (diameter=length=10mm) were inserted bilaterally in each of the distal femurs of the sheep; thus 4 implants in each sheep. The concentric gap (2 mm) surrounding the implant was filled with 1) BAG (autogenous), 2) granules, 3) granules+bone marrow aspirate (BMA, autologous) or 4) allograft. The sheep were euthanized after 6 weeks. Push-out tests were performed to assess implant fixation. Findings: Results were assessed by One-way ANOVA. P-values less than 0.05 were considered significant. No significant differences regarding failure energy (kJ/m2, p=0.070) could be seen between the groups. Shear stiffness (MPa) was significantly higher for the allograft group, p=0.002. The BAG showed up to 1.5 and 2.4 folds better properties regarding shear stiffness than the granules alone or the granules+BMA respectively. The BAG also had up to 1.8 and 5.3 fold higher failure energy than the granules alone or the granules+BMA respectively. Conclusion: The present study is the first to show an effect of bioreactor activated bone substitute on mechanical fixation. However the mechanical fixation did not reach that of allograft. Probably, an extended observation time could increase the mechanical fixation of implants treated with the bioreactor activated graft. 125
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2010-378_<strong>DOS</strong> nr. 3 2010 29/09/10 10:08 Side 124<br />
Different responses to hypoxia of mesenchymal stem<br />
cells compared to chondrocytes<br />
Casper Bindzus Foldager, Anna Bay Nielsen, Bjørn Borsøe<br />
Christensen, Moustapha Kassem, Cody Bünger, Martin Lind<br />
Orthopaedic Research Lab, Aarhus University Hospital; Department of<br />
Endocrinology and Metabolism, University Hospital of Odense,<br />
Sports Trauma Clinic, Aarhus University Hospital<br />
Background: Articular cartilage is a specialized tissue in which the<br />
native chondrocytes are exposed to low oxygen tensions. This hypoxic<br />
environment is essential for the chondrogenic viability. Hypoxia<br />
inducible factors (HIFs) are believed to play an essential role in the adaption<br />
to this environment. The most commonly used treatment for cartilage<br />
defects is microfracture, which relies on mesenchymal stem cells<br />
(MSCs) to differentiate into chondrocytes for subsequent cartilage production.<br />
Purpose: The aim of this study is to investigate the response of hypoxia<br />
on normal and differentiating MSCs compared to native chondrocytes.<br />
Methods: Three cell cultures were used: 1. Human chondrocytes; 2.<br />
Immortalized (TERT-) MSCs in chondrogenic differentiation medium<br />
(diff); 3. TERT-MSCs in standard medium(std). The three groups were<br />
divided into three oxygen tensions: 21%, 15%, and 5%. They were cultured<br />
in groups for 1, 2, 7, and 14 days. Quantitative gene expression<br />
analysis was obtained using qRT-PCR and protein synthesis was measured<br />
with ELISA. Statistics were performed with ANOVA with repeated<br />
measures.<br />
Findings: Chondrocytes and TERT-MSCs(diff) did not show an increase<br />
in HIF-1alpha and HIF- 2aplha with lowering of oxygen from 21% to<br />
5%. However, TERT-MSCs(std) showed a significant 3-fold increase.<br />
The expression of VEGF in chondrocytes showed 5-fold increase while<br />
TERT-MSC(diff) showed a 8- fold increase and TERT-MSCs a 12-fold<br />
increase.<br />
Conclusion: Vascularization of regenerating cartilage is a main contributor<br />
to the generation of fibrous tissue and failed cartilage formation.<br />
This is the first study to describe the unfortunate response of stem cells<br />
compared to chondrocytes, which might contribute significantly to the<br />
understanding of impaired regeneration of cartilage using microfracture.<br />
124