Development of Biomarkers for Use in Prediction of Equine MSC Cultures with High Chondrogenic Potency

Presented by: Hamed Alizadeh

 

Authors: Hamed Alizadeh-1, Stewart Russell-2, Pratibha Potla-3, Starlee Lively-3, Sarah Lepage-1, S. Amanda Ali-3, Mohit Kapoor-3, Thomas G. Koch-1

Affiliations: 1-Department of Biomedical Sciences, OVC, University of Guelph, ON, Canada; 2-Create Fertility Centre, Toronto, ON, Canada 3-Krembil Research Institute, University Health Network, Toronto, ON, Canada

Introduction: Multipotent Mesenchymal Stromal Cells (MSCs) are a heterogeneous population of cells with varying chondrogenic potency. Biomarkers predicting the chondrogenic potential of MSCs would allow for more time- and cost-effective identification of MSC cultures suitable for cartilage repair strategies compared to current post-chondrogenic induction determination of potency. MicroRNAs (miRNAs) are involved in the regulation of many cell functions and may be useful biomarkers to determine the chondrogenic potential of undifferentiated MSCs.

Hypothesis / Objectives: We hypothesized that equine cord blood-derived (eCB) MSC cultures with variable chondrogenic potency are associated with differential global expression of miRNAs. The objective of this study was to determine if miRNAs were differently expressed between eCB-MSC cultures with knowing high and low chondrogenic potency.

Materials and Methods: To assess chondrogenic potential of eCB-MSCs, the neocartilages were initially evaluated histologically and biochemically. Subsequently, global miRNA expression profiles of the eCB-MSC cultures [9 high and 9 low chondrogenic potential (HCP and LCP)] before the onset of chondrogenic differentiation were performed via next generation sequencing (NGS). A total of 1257 miRNAs were included in the analysis.

Results: We found 9 miRNAs that were differentially expressed (adjusted p-value <0.05) between HCP and LCP. A few miRNAs were of interest, miR-485-5p has been found previously to inhibit chondrogenesis through targeting SOX9, and we have identified this miRNA as 3.6-fold downregulated in our HCP CB-MSCs. Similarly, miR-1247-5p was downregulated in HCP CB-MSC, and has been demonstrated to target SOX9, decreasing chondrogenic potential.

Conclusions: Our findings suggest an association between miRNA expression and chondrogenic differentiation potential. We anticipate that further empirical studies help us to determine candidate miRNAs as chondrogenic predictive markers of undifferentiated eCB-MSCs.

Acknowledgements, Funding, and Conflicts of Interest: This project was supported by the Equine Guelph and Partners (T.G.K.), NSERC-DG (T.G.K.), CFI-LOF (T.G.K.), MRI-ERA (T.G.K.) and the Dean’s Office, Ontario Veterinary College (H.A., PhD scholarship), and partial PhD stipend support from the Department of Biomedical Sciences Graduate Growth Fund, Ontario Veterinary College (H.A.).

H. Alizadeh and co-authors (Stewart Russell, Pratibha Potla, Starlee Lively, Sarah Lepage, S. Amanda Ali, Mohit Kapoor) declare no competing financial interests. T.G. Koch acts in a volunteer capacity as non-executive Director, Scientific Affairs (ex officio) of eQcell therapies Inc., Aurora, ON, Canada, a company for which Dr. Koch’s research laboratory provides equine stem cell isolation and storage services. Dr. Koch holds a minor non-controlling share in eQcell therapies Inc.