A. El Haj (Birmingham, GB)
University of Birmingham Healthcare Technology InstitutePresenter Of 2 Presentations
18.2.10 - Mechano-ion-channel activation of TRPV4 promotes cartilage differentiation of MSCs and chondroprogenitors
Abstract
Purpose
The absence of a long term treatment for OA has led to the development of new approaches for early intervention. Mechanical regulators, such as TRPV4, play a key role in OA and are potential targets for early treatments. We demonstrate a new technology to remotely mechano-activate ion channels such as TRPV4 to control MSc and chondrogenic differentiation and maturation.
Methods and Materials
MScs from bone marrow and other sources cultured as pellets (4x105 cells) and micromass cultures (5x104 cells). In some cases, cells were previously transfected with GFP- collagen type 2 reporters. Cells were labelled with Nanomag (250 nm) with carboxylic coating and linked to the specific antibody for each group: TRPV4 and RGD. Cells were cultured for 21 days in chondrogenic, chondrogenic without TGF-B3 and basal media.The tagged cells were submitted to a 1h daily cyclical magnetic field controlled by a MICA bioreactor (MICA Biosystems Ltd) or static. Chondrogenic differentiation was monitored with temporal key markers.
Results
In response to the mechano-activation of TRPV4, we observed an increase in Collagen 2 and enhanced early expression onset in transfected MSCs at 14 days in culture. Histological and molecular analysis demonstrated a similar pattern for Collagen 2 and aggrecan up-regulation in MSCs. Enlarged pellet size was observed in the TRPV4 tagged chondroprogenitors compared to controls with an increase in uniform expression patterns (Figure 1). Elevated proliferation levels were found in the activated ion channel group. Further work underway includes ex vivo models to demonstrate remote control.
Conclusion
The development of allogeneic injectable solutions for early OA therapy could provide solutions which would improve long lasting motility with ageing. MICA approaches using MNPs tagged to receptor targets has the potential to generate these new therapies. The use of ion channels such as TRPV4 as targets for enhancing chondrogenesis is being explored further in ex vivo and animal models.
18.3.2 - In vivo MRI cell tracking of autologous mesenchymal stem cells in an ovine osteochondral defect model.
Abstract
Purpose
We aim to devise and implement a SPION (superparamagnetic iron oxide nanoparticle) based technique to non-invasively monitor the intra-articular delivery and bio-distribution of stem cells within a knee joint by MR-imaging. Osteochondral injuries represent a significant clinical problem requiring cell-based therapies to restore function of the damaged joint. Pre-clinical studies are fundamental in translating such therapies, however, technologies to non-invasively assess in vivo cell fate are currently limited. We implement this technique to monitor and compare in vivo cell fate in a pre-clinical ovine model of a chronic and an acute osteochondral injury.
Methods and Materials
Autologous ovine MSCs were isolated, expanded and labelled with Nanomag using the cell-penetrating peptide, P218R. In vitro and ex vivo MRI detection thresholds and cellular viability, proliferation and differentiation were determined prior to in vivo studies. A single 8mm diameter osteochondral defect was created in the medial femoral condyle in the left knee joint of each sheep with the contralateral joint serving as the control. 10x106 Nanomag labelled-MSCs were delivered by intra-articular injection at 1 week or 4.5 weeks post injury. Sheep were sacrificed 7 days post implantation and immediately MR imaged using an ESAOTE 0.2T and Siemens 3T scanner and prior to histological assessement.
Results
In vitro MRI data demonstrated significant increase in MRI contrast as a result of P218R:Nanomag with no impairment of cell function. MRI images revealed evidence of implanted cells within the synovial joint of the injured leg of the chronic model only with no signs of cell localisation to the defect site in either model. This was validated histologically determining the location of implanted cells in the synovium with evidence of engulfment of Nanomag-labelled cells by leukocytes.
Conclusion
We demonstrates the potential of Nanomag and P218R as an effective means of imaging and tracking cells within the knee.