Osteogenic human MSC-derived extracellular vesicles regulate MSC activity and osteogenic differentiation and promote bone regeneration in a rat calvarial defect model

Niyaz Al-Sharabi; Samih Mohamed-Ahmed; Siddharth Shanbhag; Carina Kampleitner; Rammah Elnour; Shuntaro Yamada; Neha Rana; Even Birkeland; Stefan Tangl; Reinhard Gruber; Kamal Mustafa

doi:10.1186/s13287-024-03639-x

Osteogenic human MSC-derived extracellular vesicles regulate MSC activity and osteogenic differentiation and promote bone regeneration in a rat calvarial defect model

Stem Cell Res Ther. 2024 Feb 7;15(1):33. doi: 10.1186/s13287-024-03639-x.

Authors

Niyaz Al-Sharabi¹, Samih Mohamed-Ahmed², Siddharth Shanbhag^{2

3}, Carina Kampleitner^{4

5

6}, Rammah Elnour⁷, Shuntaro Yamada², Neha Rana², Even Birkeland⁸, Stefan Tangl^{4

6}, Reinhard Gruber^{6

9

10}, Kamal Mustafa²

Affiliations

¹ Department of Clinical Dentistry, Faculty of Medicine, Center for Translational Oral Research (TOR), University of Bergen, 5009, Bergen, Norway. N.Al-Sharabi@uib.no.
² Department of Clinical Dentistry, Faculty of Medicine, Center for Translational Oral Research (TOR), University of Bergen, 5009, Bergen, Norway.
³ Department of Immunology and Transfusion Medicine, Haukeland University Hospital, 5021, Bergen, Norway.
⁴ Karl Donath Laboratory for Hard Tissue and Biomaterial Research, University Clinic of Dentistry, Medical University of Vienna, 1090, Vienna, Austria.
⁵ Ludwig Boltzmann Institute for Traumatology, The Research Center in Cooperation with AUVA, 1200, Vienna, Austria.
⁶ Austrian Cluster for Tissue Regeneration, 1200, Vienna, Austria.
⁷ Department of Clinical Medicine, Faculty of Medicine, University of Bergen, 5009, Bergen, Norway.
⁸ The Proteomics Facility of the University of Bergen (PROBE), University of Bergen, 5021, Bergen, Norway.
⁹ Department of Oral Biology, University Clinic of Dentistry, Medical University of Vienna, 1090, Vienna, Austria.
¹⁰ Department of Periodontology, School of Dental Medicine, University of Bern, 3010, Bern, Switzerland.

Abstract

Background: There is growing evidence that extracellular vesicles (EVs) play a crucial role in the paracrine mechanisms of transplanted human mesenchymal stem cells (hMSCs). Little is known, however, about the influence of microenvironmental stimuli on the osteogenic effects of EVs. This study aimed to investigate the properties and functions of EVs derived from undifferentiated hMSC (Naïve-EVs) and hMSC during the early stage of osteogenesis (Osteo-EVs). A further aim was to assess the osteoinductive potential of Osteo-EVs for bone regeneration in rat calvarial defects.

Methods: EVs from both groups were isolated using size-exclusion chromatography and characterized by size distribution, morphology, flow cytometry analysis and proteome profiling. The effects of EVs (10 µg/ml) on the proliferation, migration, and osteogenic differentiation of cultured hMSC were evaluated. Osteo-EVs (50 µg) or serum-free medium (SFM, control) were combined with collagen membrane scaffold (MEM) to repair critical-sized calvarial bone defects in male Lewis rats and the efficacy was assessed using µCT, histology and histomorphometry.

Results: Although Osteo- and Naïve-EVs have similar characteristics, proteomic analysis revealed an enrichment of bone-related proteins in Osteo-EVs. Both groups enhance cultured hMSC proliferation and migration, but Osteo-EVs demonstrate greater efficacy in promoting in vitro osteogenic differentiation, as evidenced by increased expression of osteogenesis-related genes, and higher calcium deposition. In rat calvarial defects, MEM with Osteo-EVs led to greater and more consistent bone regeneration than MEM loaded with SFM.

Conclusions: This study discloses differences in the protein profile and functional effects of EVs obtained from naïve hMSC and hMSC during the early stage of osteogenesis, using different methods. The significant protein profile and cellular function of EVs derived from hMSC during the early stage of osteogenesis were further verified by a calvarial bone defect model, emphasizing the importance of using differentiated MSC to produce EVs for bone therapeutics.

Keywords: Bone regeneration; Extracellular vesicles; Mesenchymal stem cells; Naïve-EVs; Osteo-EVs; Rat calvarial defects.

MeSH terms

Animals
Bone Regeneration / physiology
Cell Differentiation
Extracellular Vesicles* / metabolism
Humans
Male
Mesenchymal Stem Cells* / metabolism
Osteogenesis / genetics
Proteomics
Rats
Rats, Inbred Lew

Abstract

MeSH terms

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