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. 2023 Mar 7:11:1110703.
doi: 10.3389/fbioe.2023.1110703. eCollection 2023.

Rat bone marrow mesenchymal stem cells induced by rrPDGF-BB promotes bone regeneration during distraction osteogenesis

Shuo Wu  1 , Lijie Zhang  2 , Ruidan Zhang  3 , Kang Yang  4 , Qin Wei  5 , Qiyu Jia  1 , Jian Guo  1 , Chuang Ma  1
Affiliations

Rat bone marrow mesenchymal stem cells induced by rrPDGF-BB promotes bone regeneration during distraction osteogenesis

Shuo Wu et al. Front Bioeng Biotechnol. .

Abstract

Background: In the clinical treatment of large bone defects, distraction osteogenesis can be used. However, some patients may suffer from poor bone regeneration, or even delayed healing or non-union. Problems with the aggregation and proliferation of primary osteoblasts, or problems with the differentiation of primary osteoblasts will lead to poor bone regeneration. Therefore, supplementing exogenous primary osteoblasts and growth factors when using distraction osteogenesis may be a treatment plan with great potential. Methods: Bone marrow mesenchymal stem cells (BMSCs) were extracted from rats and cultured. Subsequently, Recombinant Rat Platelet-derived Growth Factor BB (rrPDGF-BB) was used to induce bone marrow mesenchymal stem cells. At the same time, male adult rats were selected to make the right femoral distraction osteogenesis model. During the mineralization period, phosphate buffer salt solution (control group), non-induction bone marrow mesenchymal stem cells (group 1) and recombinant rat platelet-derived growth factor BB intervened bone marrow mesenchymal stem cells (group 2) were injected into the distraction areas of each group. Then, the experimental results were evaluated with imaging and histology. Statistical analysis of the data showed that the difference was statistically significant if p < 0.05. Results: After intervention with recombinant rat platelet-derived growth factor BB on bone marrow mesenchymal stem cells, the cell morphology changed into a thin strip. After the cells were injected in the mineralization period, the samples showed that the callus in group 2 had greater hardness and the color close to the normal bone tissue; X-ray examination showed that there were more new callus in the distraction space of group 2; Micro-CT examination showed that there were more new bone tissues in group 2; Micro-CT data at week eight showed that the tissue volume, bone volume, percent bone volume, bone trabecular thickness, bone trabecular number and bone mineral density in group 2 were the largest, and the bone trabecular separation in group 2 was the smallest. There was a statistical difference between the groups (p < 0.05); HE staining confirmed that group 2 formed more blood vessels and chondrocytes earlier than the control group. At 8 weeks, the bone marrow cavity of group 2 was obvious, and some of them had been fused. Conclusion: The study confirmed that injecting bone marrow mesenchymal stem cellsBB into the distraction space of rats can promote the formation of new bone in the distraction area and promote the healing of distraction osteogenesis.

Keywords: BMSCs; PDGF-BB; distraction osteogenesis (DO); femur; osteogenesis; rat.

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Conflict of interest statement

The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest.

Figures

FIGURE 1
FIGURE 1
Design drawing of external fixation device and Kirschner wire. (A-C) Design drawing of external fixation device. (D) Threaded Kirschner wire matched with external fixation device.
FIGURE 2
FIGURE 2
Schematic diagram of the operation of rat right femoral distraction osteogenesis model. (A-C) The surgical procedure. (D) X-ray examination after operation.
FIGURE 3
FIGURE 3
Diagram of the process of distraction osteogenesis in rats. (A) Before distraction the femur (black arrow shows osteotomy area). (B) After distraction the femur (red arrow shows the distraction area).
FIGURE 4
FIGURE 4
Post-operative picture of the rat and schematic diagram of injected cells. (A) Post-operative photos of rats with external fixation devices. (B) Schematic diagram of injected of cells into the distraction osteogenesis area of rat femur.
FIGURE 5
FIGURE 5
Photos of BMSCs in SD rats (X50). (A) Photos of primary cells. (B) Photos of the third generation cells.
FIGURE 6
FIGURE 6
Flow cytometry analysis of rat BMSCs. (A) Expression of CD29. (B) Expression of CD45. (C) Expression of CD34.
FIGURE 7
FIGURE 7
Photos of rrPDGF-BB induced BMSCs (X50). (A) One days after intervention. (B) Three days after intervention.
FIGURE 8
FIGURE 8
X-ray photos of femoral samples at different time points.
FIGURE 9
FIGURE 9
Micro-CT images of femurs in each group at the 8th week.
FIGURE 10
FIGURE 10
Micro-CT parameters of rat femora at week 8. (A) Micro-CT analyses of tissue volume (TV) in each group. (B) Micro-CT analyses of bone volume (BV) in each group. (C) Micro-CT analyses of percent bone volume (BV/TV) in each group. (D) Micro-CT analyses of trabecular thickness (Tb.Th) in each group. (E) Micro-CT analyses of trabecular number (Tb.N) in each group. (F) Micro-CT analyses of trabecular separation (Tb.Sp) in each group. (G) Micro-CT analyses of bone mineral density (BMD) in each group. ***p-value <0.001. **p-value <0.01. *p-value <0.05.
FIGURE 11
FIGURE 11
General picture of rat femur.
FIGURE 12
FIGURE 12
HE staining photos of femoral samples at different time points (X50).
See this image and copyright information in PMC

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