Mesenchymal stem cell application improves tendon healing via anti-apoptotic effect (Animal study)

Abstract

Objective: The aim of this study was to determine the effects of mesenchymal stem cell (MSC) application and the possible pathways of MSC’s effects on tendon strength and healing after tendon repair.

Methods: The study included 40 Wistar albino rats. Mesenchymal stem cells were obtained from the femurs and tibias of 6 rats. Achilles tendons of the remaining 34 rats were cut and repaired with open surgical procedures. Rats were divided into 2 groups. Percutaneous MSCs were applied to the study group (n=17) and physiological serum only was applied to the control group (n=17) at the 4th week. Rats were sacrificed using the cervical dislocation method under ether anesthesia at the 12th week and samples were analyzed by histological and immunohistochemical methods. For biomechanical analysis, a traction force was applied at 10 mm/min and load to failure was recorded for each specimen in Newtons.

Results: Histologically, there was no significant difference between groups (p>0.05). In the immunohistochemical studies, MSCs were located more intensively at the repair zone. Apoptosis was minimally present in the study group and was clearly increased in the control group. Increase in tendon strength was significantly higher in the study group than in the control group at the 12th week (p<0.05).

Conclusion: The application of MSCs to decrease re-ruptures has a positive effect on tendon strength, probably due to their anti-apoptotic effects. Mesenchymal stem cell application can be used percutaneously and is effective in clinical practice in the late stages of tendon healing.

Keywords

• Apoptosis
• mesenchymal stem cell
• tendon healing

Mezenkimal kök hücrelerin anti-apoptotik etkileri ve tendon iyileşmesi (Hayvan çalışması)

Öz

Amaç: Bu çalışmada tendon onarımı sonrasında, mezenkimal kök hücre (MKH) uygulamasının tendon gücü ve iyileşmesi üzerine etkileri ve olası etki mekanizmaları araştırıldı.

Çalışma planı: Çalışmaya alınan 40 adet Wistar albino sıçanın 6’sının femur ve tibiaları mezenkimal kök hücre elde etmek için kullanıldı. Kalan 34 sıçanda açık olarak Aşil tendon kesilip onarıldı. Sıçanlar daha sonra 2 gruba ayrıldı. Dördüncü haftada çalışma grubuna (n=17) perkütan MKH uygulaması yapılırken, kontrol grubuna (n=17) sadece serum fizyolojik verildi. Sıçanlardan, 12. haftada eter anestezisi altında servikal dislokasyonla sakrifiye edilerek alınan örnekler, histolojik ve immünohistokimyasal olarak değerlendirildi. Biyomekanik analizde 10 mm/dk ile traksiyon kuvveti uygulandı ve kopma gücü her tendon için Newton cinsinden kaydedildi.

Bulgular: Histolojik olarak gruplar arasında anlamlı fark bulunamadı (p>0.05). Yapılan immünohistokimyasal çalışmalarda ise verilen MKH’lerin daha yoğun olarak onarım sahasına yerleştiği, kontrol grubunda belirgin olarak artmış apoptozun çalışma grubunda çok düşük olduğu görüldü. Tendon gücü açısından ise 12. haftada, mezenkimal kök hücre verilen çalışma grubunun kontrol grubuna göre istatistiksel olarak anlamlı derecede daha üstün olduğu bulundu (p<0.05).

Çıkarımlar: Nüks yırtıklarını azaltmak için başvurulan MKH uygulaması, muhtemel anti-apoptotik etkileri sayesinde tendonun gücünü arttırmaktadır. Mezenkimal kök hücre uygulamaları tendon iyileşmesinin geç dönemlerinde perkütan olarak etkili bir şekilde kullanılabilir.

Anahtar Kelimeler

• Apoptoz; mezenkimal kök hücre; tendon iyileşmesi.

References

1. Hoffmann A, Gross G. Tendon and ligament engineering in the adult organism: mesenchymal stem cells and genetherapeutic approaches. Int Orthop 2007;31:791-7.
2. Lui PP, Cheuk YC, Hung LK, Fu SC, Chan KM. Increased apoptosis at the late stage of tendon healing. Wound Repair Regen 2007;15:702-7.
3. Steiner M. Biomechanics of tendon healing. J Biomech 1982;15:951-8.
4. Karaoz E, Aksoy A, Ayhan S, Sariboyaci AE, Kaymaz F, Kasap M. Characterization of mesenchymal stem cells from rat bone marrow: ultrastructural properties, differentiation potential and immunophenotypic markers. Histochem Cell Biol 2009;132:533-46.
5. Kiliç E, Ceyhan T, Cetinkaya DU. Evaluation of differentiation potential of human bone marrow-derived mesenchymal stromal cells to cartilage and bone cells. [Article in Turkish] Acta Orthop Traumatol Turc 2007;41:295-301.
6. Adas G, Arikan S, Karatepe O, Kemik O, Ayhan S, Karaoz E, et al. Mesenchymal stem cells improve the healing of ischemic colonic anastomoses (experimental study). Langenbecks Arch Surg 2011;396:115-26.
7. Saygi B, Yildirim Y, Cabukoğlu C, Kara H, Ramadan SS, Esemenli T. The effect of dehydration and irrigation on the healing of Achilles tendon: an experimental study. Ulus Travma Acil Cerrahi Derg 2008;14:103-9.
8. Uysal CA, Tobita M, Hyakusoku H, Mizuno H. Adiposederived stem cells enhance primary tendon repair: biomechanical and immunohistochemical evaluation. J Plast Reconstr Aesthet Surg 2012;65:1712-9.

9. Xu YX, Chen L, Wang R, Hou WK, Lin P, Sun L, et al. Mesenchymal stem cell therapy for diabetes through paracrine mechanisms. Med Hypotheses 2008;71:390-3.
10. Ichim TE, Alexandrescu DT, Solano F, Lara F, Campion Rde N, Paris E, et al. Mesenchymal stem cells as antiinflammatories: implications for treatment of Duchenne muscular dystrophy. Cell Immunol 2010;260:75-82.
11. Gnecchi M, He H, Noiseux N, Liang OD, Zhang L, Morello F, et al. Evidence supporting paracrine hypothesis for Akt-modified mesenchymal stem cell-mediated cardiac protection and functional improvement. FASEB J 2006;20:661-9.
12. Karaoz E, Genç ZS, Demircan PÇ, Aksoy A, Duruksu G. Protection of rat pancreatic islet function and viability by coculture with rat bone marrow-derived mesenchymal stem cells. Cell Death Dis 2010;1:e36.
13. Mae T, Shino K, Maeda A, Toritsuka Y, Horibe S, Ochi T. Effect of gamma irradiation on remodeling process of tendon allograft. Clin Orthop Relat Res 2003;414:305-14.
14. Aoki M, Ogiwara N, Ohta T, Nabeta Y. Early active motion and weightbearing after cross-stitch achilles tendon repair. Am J Sports Med 1998;26:794-800.
15. Cretnik A, Kosanović M, Smrkolj V. Percutaneous suturing of the ruptured Achilles tendon under local anesthesia. J Foot Ankle Surg 2004;43:72-81.
16. Gelberman RH, Nunley JA 2nd, Osterman AL, Breen TF, Dimick MP, Woo SL. Influences of the protected passive mobilization interval on flexor tendon healing. A prospective randomized clinical study. Clin Orthop Relat Res 1991;264:189-96.
17. Cetti R, Henriksen LO, Jacobsen KS. A new treatment of ruptured Achilles tendons. A prospective randomized study. Clin Orthop Relat Res 1994;308:155-65.
18. Möller M, Movin T, Granhed H, Lind K, Faxén E, Karlsson J. Acute rupture of tendon Achillis. A prospective randomised study of comparison between surgical and nonsurgical treatment. J Bone Joint Surg Br 2001;83:843-8.
19. Awad HA, Boivin GP, Dressler MR, Smith FN, Young RG, Butler DL. Repair of patellar tendon injuries using a cell-collagen composite. J Orthop Res 2003;21:420-31.
20. Chong AK, Ang AD, Goh JC, Hui JH, Lim AY, Lee EH, et al. Bone marrow-derived mesenchymal stem cells influence early tendon-healing in a rabbit achilles tendon model. J Bone Joint Surg Am 2007;89:74-81.
21. Young RG, Butler DL, Weber W, Caplan AI, Gordon SL, Fink DJ. Use of mesenchymal stem cells in a collagen matrix for Achilles tendon repair. J Orthop Res 1998;16:406
22. Hapa O, Cakici H, Gideroğlu K, Ozturan K, Kükner A, Buğdayci G. The effect of ethanol intake on tendon healing: a histological and biomechanical study in a rat model. Arch Orthop Trauma Surg 2009;129:1721-6.
23. Caplan AI. Review: mesenchymal stem cells: cell-based reconstructive therapy in orthopedics. Tissue Eng 2005;11:1198-211.