Experimental study on the causes of spontaneous osteogenesis of Masquelet induced membrane_Chinese Journal of Reparative and Reconstructive Surgery

Authors：

YIN Qudong ¹ , MAO Dong ² ,  RUI Yongjun ¹

1. Department of Orthopaedics, Wuxi No.9 People’s Hospital Affiliated to Suzhou University, Suzhou Jiangsu, 214062, P. R. China;
2. Orthopedic Research Institute, Wuxi No.9 People’s Hospital Affiliated to Suzhou University, Suzhou Jiangsu, 214062, P. R. China;

Corresponding author：

RUI Yongjun, Email: ruiyongjun_md@163.com

Keywords：

Bone defect; Masquelet technology; induced membrane; osteogenic activity; spontaneous osteogenesis; rat

DOI：

10.7507/1002-1892.202403021

Video：

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Objective To investigate the causes of spontaneous osteogenesis of Masquelet induced membrane. Methods Forty-two male Sprague-Dawley rats aged 7-9 weeks were selected to establish a critical-sized bone defect of the right middle femur model. Then the rats were randomly divided into 4 groups, with 12 rats in groups A-C and 6 rats in group D. The bone defects in groups A-C were filled with vancomycin-loaded polymethyl methacrylate bone cement spacers. Then the Kirschner wires were used for intramedullary fixation in groups A and B, and the bone cement was used to connect the bone cement spacers and the bone ends in group B. The steel plate was used to fixation in group C. The bone defect in group D was only fixed with steel plate as a blank control group. The general condition was observed after operation. At 5 weeks after operation, 6 rats in groups A-C were selected for STRO-1 immunohistochemistry to observe the content of mesenchyme stem cells (MSCs) in the induced membrane (STRO-1⁺ cells). At 12 weeks after operation, the remaining rats in groups A-D were taken for X-ray observation, gross observation, and histological observation (HE, safranin O-green staining) to observe the effect of inducing spontaneous osteogenesis of the membrane. Results All rats in the 4 groups survived until the completion of the experiment. At 5 weeks after operation, the immunohistochemical staining showed that group B was negative, while the contents of MSCs in the induced membrane in groups A and C were 14.20%±1.92% and 5.00%±0.71%, respectively, with a significant difference (P<0.05). At 12 weeks after operation, group A showed significant new bone growth towards the center of the bone defect at the osteotomy site, with an average length of 3.1 mm on one side. Histological observation revealed the presence of bone and cartilage lesions, fibers, and a small amount of neovascularization in the induced membrane. Group C only had a small amount of new bone at the bone end, while groups B and D did not have any new bone, but bone resorption or atrophy at the bone end and a small amount of neovascularization were noted in the induced membrane. Group D showed collagen fiber proliferation and a small amount of neovascularization. Conclusion Although the induced membrane of Masquelet technology has osteogenesis, the key factor for the spontaneous osteogenesis of the induced membrane is the bone marrow overflow from the bone marrow cavity providing MSCs. The presence of bone and cartilage in the new bone indicates the spontaneous osteogenesis of the induced membrane belongs to endochondral ossification.

1.	Lu W, Zhao R, Fan X, et al. Time-varying characteristics of the induced membrane and its effects on bone defect repair. Injury, 2023, 54(2): 318-328.
2.	殷渠东, 孙振中, 顾三军. 应用Masquelet技术修复骨缺损的研究进展. 中国修复重建外科杂志, 2013, 27(10): 1273-1276.
3.	Klein C, Monet M, Barbier V, et al. The Masquelet technique: Current concepts, animal models, and perspectives. J Tissue Eng Regen Med, 2020, 14(9): 1349-1359.
4.	Mathieu L, Murison JC, de Rousiers A, et al. The Masquelet technique: can disposable polypropylene syringes be an alternative to standard PMMA spacers? A rat bone defect model. Clin Orthop Relat Res, 2021, 479(12): 2737-2751.
5.	Luo J, Bo F, Wang J, et al. Application of the induced membrane technique of tibia using extracorporeal vs. intracorporeal formation of a cement spacer: a retrospective study. BMC Musculoskelet Disord, 2022, 23(1): 460. doi: 10.1186/s12891-022-05355-0.
6.	Wang JB, Yin QD, Gu SJ, et al. Induced membrane technique in the treatment of infectious bone defect: A clinical analysis. Orthop Traumatol Surg Res, 2019, 105(3): 535-539.
7.	Pereira R, Perry WC, Crisologo PA, et al. Membrane-induced technique for the management of combined soft tissue and osseous defects. Clin Podiatr Med Surg, 2021, 38(1): 99-110.
8.	Klaue K, Knothe U, Anton C, et al. Bone regeneration in long-bone defects: tissue compartmentalisation? In vivo study on bone defects in sheep. Injury, 2009, 40 Suppl 4: S95-S102.
9.	Gruber HE, Gettys FK, Montijo HE, et al. Genomewide molecular and biologic characterization of biomembrane formation adjacent to a methacrylate spacer in the rat femoral segmental defect model. J Orthop Trauma, 2013, 27(5): 290-297.
10.	Yin Q, Chen X, Dai B, et al. Varying degrees of spontaneous osteogenesis of Masquelet's induced membrane: experimental and clinical observations. BMC Musculoskeletal Disorders, 2023, 24(1): 384. doi: 10.1186/s12891-023-06498-4.
11.	Wang K, Gao F, Zhang Y, et al. Comparison of osteogenic activity from different parts of induced membrane in the Masquelet technique. Injury, 2023, 54(11): 111022. doi: 10.1016/j.injury.2023.111022.
12.	Henrich D, Seebach C, Nau C, et al. Establishment and characterization of the Masquelet induced membrane technique in a rat femur critical-sized defect model. J Tissue Eng Regen Med, 2016, 10(10): E382-E396.
13.	Pélissier P, Lefevre Y, Delmond S, et al. Influences of induced membranes on heterotopic bone formation within an osteo-inductive complex. Experimental study in rabbits. Ann Chir Plast Esthet, 2009, 54(1): 16-20.
14.	Caballé-Serrano J, Munar-Frau A, Ortiz-Puigpelat O, et al. On the search of the ideal barrier membrane for guided bone regeneration. J Clin Exp Dent, 2018, 10(5): e477-e483.
15.	Alford AI, Nicolaou D, Hake M, et al. Masquelet's induced membrane technique: Review of current concepts and future directions. J Orthop Res, 2021, 39(4): 707-718.
16.	Tanner MC, Boxriker S, Haubruck P, et al. Expression of VEGF in peripheral serum is a possible prognostic factor in bone-regeneration via Masquelet-technique—A pilot study. J Clin Med, 2021, 10(4): 776. doi: 10.3390/jcm10040776.
17.	Gindraux F, Loisel F, Bourgeois M, et al. Induced membrane maintains its osteogenic properties even when the second stage of Masquelet's technique is performed later. Eur J Trauma Emerg Surg, 2020, 46(2): 301-312.
18.	Fenelon M, Etchebarne M, Siadous R, et al. Comparison of amniotic membrane versus the induced membrane for bone regeneration in long bone segmental defects using calcium phosphate cement loaded with BMP-2. Mater Sci Eng C Mater Biol Appl, 2021, 124: 112032. doi: 10.1016/j.msec.2021.112032.
19.	薛英, 刘强. 膜引导性骨再生研究进展. 国际骨科学杂志, 2006, 27(2): 110-112.
20.	Miska M, Schmidmaier G. Diamond concept for treatment of nonunions and bone defects. Unfallchirurg, 2020, 123(9): 679-686.
21.	张浩, 卢世壁, 王继芳. 引导性骨再生模型(GBR)成骨特点及骨髓的作用. 中华骨科杂志, 1998, 18(9): 540-543.
22.	芮菲, 卜凡玉, 张樱严, 等. Masquelet技术治疗骨缺损时体内制作与体外制作骨水泥间隔的疗效比较. 中华创伤骨科杂志, 2021, 23(8): 674-680.

1. Lu W, Zhao R, Fan X, et al. Time-varying characteristics of the induced membrane and its effects on bone defect repair. Injury, 2023, 54(2): 318-328.
2. 殷渠东, 孙振中, 顾三军. 应用Masquelet技术修复骨缺损的研究进展. 中国修复重建外科杂志, 2013, 27(10): 1273-1276.
3. Klein C, Monet M, Barbier V, et al. The Masquelet technique: Current concepts, animal models, and perspectives. J Tissue Eng Regen Med, 2020, 14(9): 1349-1359.
4. Mathieu L, Murison JC, de Rousiers A, et al. The Masquelet technique: can disposable polypropylene syringes be an alternative to standard PMMA spacers? A rat bone defect model. Clin Orthop Relat Res, 2021, 479(12): 2737-2751.
5. Luo J, Bo F, Wang J, et al. Application of the induced membrane technique of tibia using extracorporeal vs. intracorporeal formation of a cement spacer: a retrospective study. BMC Musculoskelet Disord, 2022, 23(1): 460. doi: 10.1186/s12891-022-05355-0.
6. Wang JB, Yin QD, Gu SJ, et al. Induced membrane technique in the treatment of infectious bone defect: A clinical analysis. Orthop Traumatol Surg Res, 2019, 105(3): 535-539.
7. Pereira R, Perry WC, Crisologo PA, et al. Membrane-induced technique for the management of combined soft tissue and osseous defects. Clin Podiatr Med Surg, 2021, 38(1): 99-110.
8. Klaue K, Knothe U, Anton C, et al. Bone regeneration in long-bone defects: tissue compartmentalisation? In vivo study on bone defects in sheep. Injury, 2009, 40 Suppl 4: S95-S102.
9. Gruber HE, Gettys FK, Montijo HE, et al. Genomewide molecular and biologic characterization of biomembrane formation adjacent to a methacrylate spacer in the rat femoral segmental defect model. J Orthop Trauma, 2013, 27(5): 290-297.
10. Yin Q, Chen X, Dai B, et al. Varying degrees of spontaneous osteogenesis of Masquelet's induced membrane: experimental and clinical observations. BMC Musculoskeletal Disorders, 2023, 24(1): 384. doi: 10.1186/s12891-023-06498-4.
11. Wang K, Gao F, Zhang Y, et al. Comparison of osteogenic activity from different parts of induced membrane in the Masquelet technique. Injury, 2023, 54(11): 111022. doi: 10.1016/j.injury.2023.111022.
12. Henrich D, Seebach C, Nau C, et al. Establishment and characterization of the Masquelet induced membrane technique in a rat femur critical-sized defect model. J Tissue Eng Regen Med, 2016, 10(10): E382-E396.
13. Pélissier P, Lefevre Y, Delmond S, et al. Influences of induced membranes on heterotopic bone formation within an osteo-inductive complex. Experimental study in rabbits. Ann Chir Plast Esthet, 2009, 54(1): 16-20.
14. Caballé-Serrano J, Munar-Frau A, Ortiz-Puigpelat O, et al. On the search of the ideal barrier membrane for guided bone regeneration. J Clin Exp Dent, 2018, 10(5): e477-e483.
15. Alford AI, Nicolaou D, Hake M, et al. Masquelet's induced membrane technique: Review of current concepts and future directions. J Orthop Res, 2021, 39(4): 707-718.
16. Tanner MC, Boxriker S, Haubruck P, et al. Expression of VEGF in peripheral serum is a possible prognostic factor in bone-regeneration via Masquelet-technique—A pilot study. J Clin Med, 2021, 10(4): 776. doi: 10.3390/jcm10040776.
17. Gindraux F, Loisel F, Bourgeois M, et al. Induced membrane maintains its osteogenic properties even when the second stage of Masquelet's technique is performed later. Eur J Trauma Emerg Surg, 2020, 46(2): 301-312.
18. Fenelon M, Etchebarne M, Siadous R, et al. Comparison of amniotic membrane versus the induced membrane for bone regeneration in long bone segmental defects using calcium phosphate cement loaded with BMP-2. Mater Sci Eng C Mater Biol Appl, 2021, 124: 112032. doi: 10.1016/j.msec.2021.112032.
19. 薛英, 刘强. 膜引导性骨再生研究进展. 国际骨科学杂志, 2006, 27(2): 110-112.
20. Miska M, Schmidmaier G. Diamond concept for treatment of nonunions and bone defects. Unfallchirurg, 2020, 123(9): 679-686.
21. 张浩, 卢世壁, 王继芳. 引导性骨再生模型(GBR)成骨特点及骨髓的作用. 中华骨科杂志, 1998, 18(9): 540-543.
22. 芮菲, 卜凡玉, 张樱严, 等. Masquelet技术治疗骨缺损时体内制作与体外制作骨水泥间隔的疗效比较. 中华创伤骨科杂志, 2021, 23(8): 674-680.

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