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Increased Exhaustion of the Subchondral Bone-Derived Mesenchymal Stem/ Stromal Cells in Primary Versus Dysplastic Osteoarthritis

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Abstract

Mesenchymal stem/ stromal cell (MSC) exhaustion has been suggested to be a hallmark of aging. Osteoarthritis has a complex etiology that comprises several factors. Dysplasia has been shown to be an individual risk factor for osteoarthritis. Subchondral bone changes are often the first detectable alterations in osteoarthritis. In this study, we aimed to determine whether skeletal MSCs are differentially affected in patients with primary versus dysplastic osteoarthritis. Patients undergoing hip arthroplasty due to primary osteoarthritis (n = 11) and osteoarthritis with hip dysplasia (n = 10) were included in the study. Femoral head subchondral bone was used for isolation of MSCs. The cells were compared using detailed ex-vivo and in-vitro analyses, which included immunophenotyping, colony-forming-unit fibroblast assay, growth kinetics, senescence, multilineage potential, immunophenotyping, and MSC marker-gene expression profiling. Isolated cells from primary osteoarthritis patients showed decreased viability in comparison with those from dysplasia patients, with similar mesenchymal fractions (i.e., CD45/ CD19/ CD14/ CD34-negative cells). In-vitro expanded MSCs from primary osteoarthritis patients showed reduced osteogenic and chondrogenic potential in comparison with dysplasia patients. There were no differences in clonogenicity, growth kinetics, senescence, adipogenic potential, and immunophenotype between these groups. Gene expression profiling showed well-known marker of bone marrow MSCs, the leptin receptor, to be significantly lower for primary osteoarthritis patients. Our study shows that the pathology of primary osteoarthritis is accompanied by bone MSC exhaustion, while biomechanical dysfunction associated with hip dysplasia can induce secondary osteoarthritis without this MSC impairment. Our study suggests that subchondral bone MSC exhaustion is implicated in the pathology of primary osteoarthritis.

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Acknowledgements

The authors thank Gorana Furlanič and Helena Poniž for their valuable assistance at Valdoltra Orthopaedic Hospital, Laura Poženel, Tina Levstek, Sandra Mramor, Iva Blaževič, Kyriaki Hadjianastasi, Lina Andersson, Matjaž Jeras and Majda Sirnik for their much appreciated technical and general help. The authors acknowledge Chris Berrie for scientific English editing of the manuscript.

Funding

This study was funded by the Slovenian Research Agency, P3–298 Research Programme and J3–7245 research project and partially funded by the ARTE Project EU INTERREG ITALIA SLOVENIA 2014 2020 and J3–1749 research project funded by the Slovenian Research Agency.

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Authors and Affiliations

  1. University of Ljubljana, Faculty of Pharmacy, Chair of Clinical Biochemistry, Askerceva 7, 1000, Ljubljana, Slovenia

    Klemen Čamernik, Janja Marc & Janja Zupan

  2. Valdoltra Orthopaedic Hospital, Jadranska 31, 6280, Ankaran, Slovenia

    Anže Mihelič, Rene Mihalič & Rihard Trebše

  3. Ludwig Boltzmann Institute for Experimental and Clinical Traumatology, AUVA Research Center, Austrian Cluster for Tissue Regeneration, Donaueschingenstrasse 13, A-1200, Vienna, Austria

    Darja Marolt Presen

  4. Department of Endocrinology, Diabetes and Metabolic Diseases, University Medical Centre, Zaloška cesta 2, 1000, Ljubljana, Slovenia

    Andrej Janež

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  1. Klemen Čamernik
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Contributions

All authors contributed to the study conception and design. Material preparation, data collection and analysis were performed by Klemen Čamernik, Anže Mihelič, Rene Mihalič, Rihard Trebše, Darja Marolt Presen and Janja Zupan. The first draft of the manuscript was written by Klemen Čamernik and Janja Zupan, and all of the authors commented on previous versions of the manuscript. All authors have read and approved the final manuscript.

Corresponding author

Correspondence to Janja Zupan.

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Conflict of Interests

R. Trebše declares payment for development of educational presentations for Zimmer, Medacta, De Puy, Link and Hareus, and R. Mihalič declares consultancy for Medacta International. The rest of the authors declare that they have no conflicts of interest regarding this study.

Ethics Approval and Consent to Participate

Approval for this study was obtained from the National Medical Ethics Committee of the Republic of Slovenia (reference numbers: 0120–523/2016–2, KME 45/10/16). Written informed consent was obtained from all of the donors included in this study.

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ESM 1

Selection of the best reference gene for qPCR for adipogenesis and osteogenesis. NormFinder [22] was used to select the most stable reference gene between the treated cells and the controls for adipogenesis and osteogenesis, in a subset of 32 samples. The best combination of the three genes was GAPDH, RPL13A and RPLP0. Shown is the expression of the osteogenic genes (COL1A1, OC, parathyroid hormone 1 receptor [PTHR1], RUNX2) in another subset of 10 pairs of MSC samples treated with osteogenic media. There was no difference in the normalization of these genes to all three reference genes (GAPDH, RPL13A and RPLP0) and to GAPDH only (two-way ANOVA with Bonferroni correction: p = 0.3003 for COL1A1; p > 0.9999 for OC, PTHR1 and RUNX2). (PNG 242 kb)

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Čamernik, K., Mihelič, A., Mihalič, R. et al. Increased Exhaustion of the Subchondral Bone-Derived Mesenchymal Stem/ Stromal Cells in Primary Versus Dysplastic Osteoarthritis. Stem Cell Rev and Rep 16, 742–754 (2020). https://doi.org/10.1007/s12015-020-09964-x

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