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. 2024 May 20;15(1):144.
doi: 10.1186/s13287-024-03755-8.

Synergistic effect of umbilical cord extracellular vesicles and rhBMP-2 to enhance the regeneration of a metaphyseal femoral defect in osteoporotic rats

Amelie Deluca  1   2 , Andrea Wagner  3   4 , Patrick Heimel  4   5   6 , Christian Deininger  3   7 , Florian Wichlas  7 , Heinz Redl  4   5 , Eva Rohde  8   9 , Herbert Tempfer  3   4 , Mario Gimona  8   10 , Andreas Traweger  3   4
Affiliations

Synergistic effect of umbilical cord extracellular vesicles and rhBMP-2 to enhance the regeneration of a metaphyseal femoral defect in osteoporotic rats

Amelie Deluca et al. Stem Cell Res Ther. .

Abstract

Background: The aim of this study was to evaluate potential synergistic effects of a single, local application of human umbilical cord MSC-derived sEVs in combination with a low dose of recombinant human rhBMP-2 to promote the regeneration of a metaphyseal femoral defect in an osteoporotic rat model.

Methods: 6 weeks after induction of osteoporosis by bilateral ventral ovariectomy and administration of a special diet, a total of 64 rats underwent a distal femoral metaphyseal osteotomy using a manual Gigli wire saw. Defects were stabilized with an adapted Y-shaped mini-locking plate and were subsequently treated with alginate only, or alginate loaded with hUC-MSC-sEVs (2 × 109), rhBMP-2 (1.5 µg), or a combination of sEVs and rhBMP-2 (n = 16 for each group). 6 weeks post-surgery, femora were evaluated by µCT, descriptive histology, and biomechanical testing.

Results: Native radiographs and µCT analysis confirmed superior bony union with callus formation after treatment with hUC-MSC-sEVs in combination with a low dose of rhBMP-2. This finding was further substantiated by histology, showing robust defect consolidation 6 weeks after treatment. Torsion testing of the explanted femora revealed increased stiffness after application of both, rhBMP-2 alone, or in combination with sEVs, whereas torque was only significantly increased after treatment with rhBMP-2 together with sEVs.

Conclusion: The present study demonstrates that the co-application of hUC-MSC-sEVs can improve the efficacy of rhBMP-2 to promote the regeneration of osteoporotic bone defects.

Keywords: BMP-2; Drug delivery; osteoporosis; Exosomes; Extracellular vesicles; Metaphyseal defect.

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Conflict of interest statement

The authors declare no competing interests. (1) Title of the approved project: “Regeneration of a metaphyseal defect in osteoporotic rats by implantation of exosomal vesicles”; (2) Name of the institutional approval committee: Federal Ministry of Education, Science and Research – V/3b (Animal testing and genetic engineering); (3) Approval Number: 2020 − 0.547.757; (4) Date of approval: 24. 09. 2020 The collection of human umbilical cord for isolation of mesenchyma stromal cells and enrichment of extracellular vesicles has been approved by the Ethics Commission for the State of Salzburg and informed consent was obtained from all donors (1). Title of the approved project: “Characterization of somatic cells and stem cells from human umbilical cord tissue”; (2) Name of the institutional approval committee: Ethics Commission for the State of Salzburg; (3) Approval Number: 415-E/1547/2-2012; (4) Date of approval: 05.11.2012.

Figures

Fig. 1
Fig. 1
Overview of groups according to the applied biomaterials and used follow-up procedures including native radiographs, histology, micro computed tomography (µCT), analysis, and torsional testing. Wks = weeks
Fig. 2
Fig. 2
MASCSPlex profiling of hUC-MSC-sEV preparation. Bar graph shows the robust expression of the sEV marker proteins CD9, CD29, CD63, and CD81 (black bars). In addition, the MSC-EV-specific markers CD44 and MSCP/NG2 were clearly present (blue bars)
Fig. 3
Fig. 3
: Radiological follow-up 1-, 2-, 4- and 6-weeks post-surgery after distal femoral plate fixation. The defect was treated with either alginate only (AH), alginate with huUC-MSC-sEVs (AH + sEVs), alginate with rhBMP-2 (AH + rhBMP-2) or alginate with a combination of BMP-2 + sEVs (AH + rhBMP-2 + sEVs)
Fig. 4
Fig. 4
: Histological evaluation 4 and 6 weeks after treatment of Masson Goldner trichrome-stained FFPE sections. Dashed lines indicate the defect area. Scale bar = 100 μm
Fig. 5
Fig. 5
µCT images of the defect area 6 weeks after treatment. (A) Representative sagittal µCT sections of the defect area. (B) Quantitative analysis of BV/TV for all 4 treatment groups, revealing no significant difference between the newly formed bone volume
Fig. 6
Fig. 6
Biomechanical analysis 6 weeks post-OP. (A) Torque and (B) torsional stiffness values determined for femora treated with alginate (AH), sEVs alone, rhBMP-2 alone, or rhBMP-2 in combination with sEVs. The red line indicates mean values determined for healthy femora, with ± 1SD indicated (grey area). * = p < 0.05; ** = p < 0.01; *** = p < 0.005
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