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Preparation of Decellularized Tissue as Dual Cell Carrier Systems: A Step Towards Facilitating Re-epithelization and Cell Encapsulation for Tracheal Reconstruction

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Abstract

Surgical treatment of tracheal diseases, trauma, and congenital stenosis has shown success through tracheal reconstruction coupled with palliative care. However, challenges in surgical-based tracheal repairs have prompted the exploration of alternative approaches for tracheal replacement. Tissue-based treatments, involving the cultivation of patient cells on a network of extracellular matrix (ECM) from donor tissue, hold promise for restoring tracheal structure and function without eliciting an immune reaction. In this study, we utilized decellularized canine tracheas as tissue models to develop two types of cell carriers: a decellularized scaffold and a hydrogel. Our hypothesis posits that both carriers, containing essential biochemical niches provided by ECM components, facilitate cell attachment without inducing cytotoxicity. Canine tracheas underwent vacuum-assisted decellularization (VAD), and the ECM-rich hydrogel was prepared through peptic digestion of the decellularized trachea. The decellularized canine trachea exhibited a significant reduction in DNA content and major histocompatibility complex class II, while preserving crucial ECM components such as collagen, glycosaminoglycan, laminin, and fibronectin. Scanning electron microscope and fluorescent microscope images revealed a fibrous ECM network on the luminal side of the cell-free trachea, supporting epithelial cell attachment. Moreover, the ECM-rich hydrogel exhibited excellent viability for human mesenchymal stem cells encapsulated for 3 days, indicating the potential of cell-laden hydrogel in promoting the development of cartilage rings of the trachea. This study underscores the versatility of the trachea in producing two distinct cell carriers—decellularized scaffold and hydrogel—both containing the native biochemical niche essential for tracheal tissue engineering applications.

Graphical Abstract

Canine tracheae were harvested and subjected to decellularization. The resulting decellularized tracheae were used to prepare two different cell-carrier systems: scaffold and hydrogel. The scaffold was utilized for seeding human bronchial airway epithelium cells (HBEpCs) on the luminal surface, while the hydrogel was used for encapsulating hMSCs and seeding human umbilical vein endothelial cells (HUVECs). The extracellular matrix-rich cell scaffolds provided a biological niche for re-epithelial lining. hMSCs and HUVECs showed remarkable viability in and on the hydrogel, respectively. However, the formation of capillary-like structures by HUVECs was not detected.

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References

  1. Dhasmana, A., A. Singh, and S. Rawal. Biomedical grafts for tracheal tissue repairing and regeneration “Tracheal tissue engineering: an overview.” J. Tissue Eng. Regen. Med. 14:653–672, 2020.

    Article  CAS  PubMed  Google Scholar 

  2. Furlow, P. W. M., and J. Douglas. Surgical anatomy of the trachea. Ann. Cardiothorac. Surg. 7:255–260, 2018.

    Article  PubMed  PubMed Central  Google Scholar 

  3. Soriano, L., T. Khalid, D. Whelan, et al. Development and clinical translation of tubular constructs for tracheal tissue engineering: a review. Eur. Respir. Rev. 30:210154, 2021.

    Article  PubMed  PubMed Central  Google Scholar 

  4. Etienne, H., D. Fabre, A. G. Caro, et al. Tracheal replacement. Eur. Respir. J. 51:1702211, 2018.

    Article  PubMed  Google Scholar 

  5. Bogan, S. L., G. Z. Teoh, and M. A. Birchall. Tissue engineered airways: a prospects article. J. Cell. Biochem. 117:1497–1505, 2016.

    Article  CAS  PubMed  Google Scholar 

  6. Crowley, C., M. Birchall, and A. M. Seifalian. Trachea transplantation: from laboratory to patient. J. Tissue Eng. Regen. Med. 9:357–367, 2015.

    Article  PubMed  Google Scholar 

  7. Torre, M., P. Varela, D. Romero, and E. Leopold. Complex tracheal resection in children: myths, reality and lessons learned. In Semin Pediatr Surg. Elsevier, 2021, p. 151059.

  8. Fishman, J. M., K. Wiles, M. W. Lowdell, et al. Airway tissue engineering: an update. Expert Opin. Biol. Ther. 14:1477–1491, 2014.

    Article  CAS  PubMed  Google Scholar 

  9. Edward, C., and L. W. Richard. Surgical stapling device–tissue interactions: what surgeons need to know to improve patient outcomes. Med. Dev. (Auckl). 7:305–318, 2014.

    Google Scholar 

  10. Stefan, W., and E. Weam. Management of tracheobronchial injuries. J. Thorac. Dis. 12:6143–6151, 2020.

    Article  Google Scholar 

  11. Jin, S., and I. Sugitani. Narrative review of management of thyroid surgery complications. Gland Surg. 10:1135–1146, 2021.

    Article  PubMed  PubMed Central  Google Scholar 

  12. Kang, Y., C. Wang, Y. Qiao, et al. Tissue-engineered trachea consisting of electrospun patterned sc-PLA/GO-g-IL fibrous membranes with antibacterial property and 3D-printed skeletons with elasticity. Biomacromolecules. 20:1765–1776, 2019.

    Article  CAS  PubMed  Google Scholar 

  13. Wu, T., H. Zheng, J. Chen, et al. Application of a bilayer tubular scaffold based on electrospun poly (l-lactide-co-caprolactone)/collagen fibers and yarns for tracheal tissue engineering. J. Mater. Chem. B. 5:139–150, 2017.

    Article  CAS  PubMed  Google Scholar 

  14. Okamoto, T., Y. Yamamoto, M. Gotoh, et al. Cartilage regeneration using slow release of bone morphogenetic protein-2 from a gelatin sponge to treat experimental canine tracheomalacia: a preliminary report. ASAIO J. 49:63–69, 2003.

    Article  CAS  PubMed  Google Scholar 

  15. Yamamoto, Y., T. Okamoto, M. Goto, et al. Experimental study of bone morphogenetic proteins-2 slow release from an artificial trachea made of biodegradable materials: evaluation of stenting time. ASAIO J. 49:533–536, 2003.

    Article  CAS  PubMed  Google Scholar 

  16. Hsieh, C.-T., C.-Y. Liao, N.-T. Dai, et al. 3D printing of tubular scaffolds with elasticity and complex structure from multiple waterborne polyurethanes for tracheal tissue engineering. Appl. Mater. Today. 12:330–341, 2018.

    Article  Google Scholar 

  17. Liao, J., Q. Guo, B. Xu, and X. Li. Overview of decellularized materials for tissue repair and organ replacement. In: Decellularized Materials, 2021, pp. 1–67.

  18. Abdul-Al, M., G. K. Kyeremeh, M. Saeinasab, et al. Stem cell niche microenvironment: review. Bioengineering (Basel). 8:108, 2021.

    Article  CAS  PubMed  Google Scholar 

  19. Tong, Z., L. Jin, J. M. Oliveira, et al. Adaptable hydrogel with reversible linkages for regenerative medicine: dynamic mechanical microenvironment for cells. Bioact Mater. 6:1375–1387, 2021.

    CAS  PubMed  Google Scholar 

  20. Barbulescu, G. I., F. M. Bojin, V. L. Ordodi, et al. Decellularized extracellular matrix scaffolds for cardiovascular tissue engineering: current techniques and challenges. Int. J. Mol. Sci. 23:13040, 2022.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  21. Zhang, W., A. Du, S. Liu, et al. Research progress in decellularized extracellular matrix-derived hydrogels. Regen. Ther. 18:88–96, 2021.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  22. Zhu, D., Z. Jiang, N. Li, et al. Insights into the use of genetically modified decellularized biomaterials for tissue engineering and regenerative medicine. Adv. Drug Deliv. Rev. 188:114413, 2022.

    Article  CAS  PubMed  Google Scholar 

  23. Mendibil, U., R. Ruiz-Hernandez, S. Retegi-Carrion, et al. Tissue-specific decellularization methods: rationale and strategies to achieve regenerative compounds. Int. J. Mol. Sci. 21:5447, 2020.

    Article  PubMed  PubMed Central  Google Scholar 

  24. Brown, M., J. Li, C. Moraes, et al. Decellularized extracellular matrix: New promising and challenging biomaterials for regenerative medicine. Biomaterials. 289:121786, 2022.

    Article  CAS  PubMed  Google Scholar 

  25. Ohata, K., and H. C. Ott. Human-scale lung regeneration based on decellularized matrix scaffolds as a biologic platform. Surg. Today. 50:633–643, 2020.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  26. Badileanu, A., C. Mora-Navarro, A. M. Gracioso Martins, et al. Fast automated approach for the derivation of acellular extracellular matrix scaffolds from porcine soft tissues. ACS Biomater. Sci. Eng. 6:4200–4213, 2020.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  27. Ye, C., J. Chen, Y. Qu, et al. Naringin in the repair of knee cartilage injury via the TGF-β/ALK5/Smad2/3 signal transduction pathway combined with an acellular dermal matrix. JOT. 32:1–11, 2022.

    PubMed  Google Scholar 

  28. Sensi, F., E. D’angelo, A. Biccari, et al. Establishment of a human 3D pancreatic adenocarcinoma model based on a patient-derived extracellular matrix scaffold. Transl. Res. 253:57–67, 2022.

    Article  PubMed  Google Scholar 

  29. Biehl, A., A. M. G. Martins, Z. G. Davis, et al. Towards a standardized multi-tissue decellularization protocol for the derivation of extracellular matrix materials. Biomater Sci. 11:641–654, 2023.

    Article  CAS  PubMed  Google Scholar 

  30. Frey, A., L. P. Lunding, J. C. Ehlers, et al. More than just a barrier: the immune functions of the airway epithelium in asthma pathogenesis. Front. Immunol. 11:761, 2020.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  31. Hansson, G. C. Mucus and mucins in diseases of the intestinal and respiratory tracts. J. Intern. Med. 285:479–490, 2019.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  32. Zhang, H., W. Fu, and Z. Xu. Re-epithelialization: a key element in tracheal tissue engineering. Regen. Med. 10:1005–1023, 2015.

    Article  CAS  PubMed  Google Scholar 

  33. Minnich, D. J., and D. J. Mathisen. Anatomy of the trachea, carina, and bronchi. Thorac. Surg. Clin. 17:571–585, 2007.

    Article  PubMed  Google Scholar 

  34. Elliott, M. J., P. De Coppi, S. Speggiorin, et al. Stem-cell-based, tissue engineered tracheal replacement in a child: a 2-year follow-up study. Lancet. 380:994–1000, 2012.

    Article  PubMed  PubMed Central  Google Scholar 

  35. Elliott, M. J., C. R. Butler, A. Varanou-Jenkins, et al. Tracheal replacement therapy with a stem cell-seeded graft: lessons from compassionate use application of a GMP-compliant tissue-engineered medicine. Stem Cells Transl. Med. 6:1458–1464, 2017.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  36. Hamilton, N., M. Kanani, D. Roebuck, et al. Tissue-engineered tracheal replacement in a child: a 4-year follow-up study. Am. J. Transplant. 15:2750–2757, 2015.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  37. Baiguera, S., C. Del Gaudio, E. Kuevda, et al. Dynamic decellularization and cross-linking of rat tracheal matrix. Biomaterials. 35:6344–6350, 2014.

    Article  CAS  PubMed  Google Scholar 

  38. Shin, Y. S., J. W. Choi, J.-K. Park, et al. Tissue-engineered tracheal reconstruction using mesenchymal stem cells seeded on a porcine cartilage powder scaffold. Ann. Biomed. Eng. 43:1003–1013, 2015.

    Article  PubMed  Google Scholar 

  39. Sun, F., Y. Jiang, Y. Xu, et al. Genipin cross-linked decellularized tracheal tubular matrix for tracheal tissue engineering applications. Sci. Rep. 6:1–12, 2016.

    Google Scholar 

  40. Giraldo-Gomez, D. M., S. J. García-López, L. Tamay-de-Dios, et al. Fast cyclical-decellularized trachea as a natural 3D scaffold for organ engineering. Mater. Sci. Eng. C. 105:110142, 2019.

    Article  CAS  Google Scholar 

  41. Lee, J. S., J. Park, D.-A. Shin, et al. Characterization of the biomechanical properties of canine trachea using a customized 3D-printed apparatus. Auris Nasus Larynx. 46:407–416, 2019.

    Article  PubMed  Google Scholar 

  42. Batioglu-Karaaltin, A., M. V. Karaaltin, E. Ovali, et al. In vivo tissue-engineered allogenic trachea transplantation in rabbits: a preliminary report. Stem Cell Rev. Rep. 11:347–356, 2015.

    Article  CAS  PubMed  Google Scholar 

  43. Butler, C. R., R. E. Hynds, C. Crowley, et al. Vacuum-assisted decellularization: an accelerated protocol to generate tissue-engineered human tracheal scaffolds. Biomaterials. 124:95–105, 2017.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  44. Lange, P., K. Greco, L. Partington, et al. Pilot study of a novel vacuum-assisted method for decellularization of tracheae for clinical tissue engineering applications. J. Tissue Eng. Regen. Med. 11:800–811, 2017.

    Article  CAS  PubMed  Google Scholar 

  45. Luo, Y., and L. Ma. Bioprosthetic heart valves with reduced immunogenic residuals using vacuum-assisted decellularization treatment. Biomed. Mater. 15:065012, 2020.

    Article  CAS  PubMed  Google Scholar 

  46. Pouliot, R. A., B. M. Young, P. A. Link, et al. Porcine lung-derived extracellular matrix hydrogel properties are dependent on pepsin digestion time. Tissue Eng. C. 26:332–346, 2020.

    Article  CAS  Google Scholar 

  47. Visscher, D. O., H. Lee, P. P. van Zuijlen, et al. A photo-crosslinkable cartilage-derived extracellular matrix bioink for auricular cartilage tissue engineering. Acta Biomater. 121:193–203, 2021.

    Article  CAS  PubMed  Google Scholar 

  48. Yodmuang, S., S. L. McNamara, A. B. Nover, et al. Silk microfiber-reinforced silk hydrogel composites for functional cartilage tissue repair. Acta Biomater. 11:27–36, 2015.

    Article  CAS  PubMed  Google Scholar 

  49. López-Martínez, S., H. Campo, L. de Miguel-Gómez, et al. A natural xenogeneic endometrial extracellular matrix hydrogel toward improving current human in vitro models and future in vivo applications. Front. Bioeng. Biotechnol. 9:156, 2021.

    Article  Google Scholar 

  50. Kang, H.-W., S. J. Lee, I. K. Ko, et al. A 3D bioprinting system to produce human-scale tissue constructs with structural integrity. Nat. Biotechnol. 34:312–319, 2016.

    Article  CAS  PubMed  Google Scholar 

  51. Matias, G. S., A. C. Carreira, V. F. Batista, et al. Ionic detergent under pressure-vacuum as an innovative strategy to generate canine tracheal scaffold for organ engineering. Cells Tissues Organs. 212:535–545, 2022.

    Article  PubMed  Google Scholar 

  52. Ravindra, A., W. D’Angelo, L. Zhang, et al. Human bronchial epithelial cell growth on homologous versus heterologous tissue extracellular matrix. J. Surg. Res. 263:215–223, 2021.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  53. Saldin, L. T., M. C. Cramer, S. S. Velankar, et al. Extracellular matrix hydrogels from decellularized tissues: structure and function. J Acta Biomater. 49:1–15, 2017.

    Article  CAS  Google Scholar 

  54. Tsou, Y.-H., J. Khoneisser, P.-C. Huang, and X. Xu. Hydrogel as a bioactive material to regulate stem cell fate. Bioactive Mater. 1:39–55, 2016.

    Article  Google Scholar 

  55. Balestrini, J. L., A. L. Gard, K. A. Gerhold, et al. Comparative biology of decellularized lung matrix: implications of species mismatch in regenerative medicine. Biomaterials. 102:220–230, 2016.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  56. Zhong, Y., W. Yang, Z. Yin Pan, et al. In vivo transplantation of stem cells with a genipin linked scaffold for tracheal construction. J. Biomater. Appl. 34:47–60, 2019.

    Article  CAS  PubMed  Google Scholar 

  57. Kokubun, K., D. Pankajakshan, M. J. Kim, and D. K. Agrawal. Differentiation of porcine mesenchymal stem cells into epithelial cells as a potential therapeutic application to facilitate epithelial regeneration. J. Tissue Eng. Regen. Med. 10:E73–E83, 2016.

    Article  CAS  PubMed  Google Scholar 

  58. Kobayashi, K., T. Suzuki, Y. Nomoto, et al. A tissue-engineered trachea derived from a framed collagen scaffold, gingival fibroblasts and adipose-derived stem cells. Biomaterials. 31:4855–4863, 2010.

    Article  CAS  PubMed  Google Scholar 

  59. Hulmes, D. Collagen diversity, synthesis and assembly. In: Collagen, edited by P. Fratzl. Boston: Springer, 2008, pp. 15–47.

    Chapter  Google Scholar 

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Acknowledgements

The authors would like to gratitude to Mrs. Athitaya Rungwong from the Department of Anatomy, Faculty of Medicine, Chulalongkorn University, and Mr. Kittipot Kongsonthana from the Department of Anatomy, Faculty of Veterinary Science, Chulalongkorn University for invaluable advice on histological work and training.

Funding

This work was supported by Ratchadapiseksompotch Fund, Chulalongkorn University [Grant Number CU_GR_62_76_31_06 to C.S.]; PMUC 2566 [Grant Number C10F640050 to S.Y.]; a research assistant from the Graduate school, Chulalongkorn University [Grant Number GCUGE1725622004D to P.S.].

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Authors and Affiliations

  1. Medical Sciences Program, Faculty of Medicine, Chulalongkorn University, Pathumwan, Bangkok, 10330, Thailand

    Pensuda Sompunga

  2. Veterinary Stem Cell and Bioengineering Innovation Center (VSCBIC), Faculty of Veterinary Science, Chulalongkorn University, Bangkok, 10330, Thailand

    Watchareewan Rodprasert & Chenphop Sawangmake

  3. Veterinary Stem Cell and Bioengineering Research Unit, Faculty of Veterinary Science, Chulalongkorn University, Bangkok, 10330, Thailand

    Watchareewan Rodprasert & Chenphop Sawangmake

  4. Department of Anatomy, Faculty of Veterinary Science, Chulalongkorn University, Pathumwan, Bangkok, 10330, Thailand

    Sayamon Srisuwatanasagul

  5. Department of Pathology, Faculty of Veterinary Science, Chulalongkorn University, Pathumwan, Bangkok, 10330, Thailand

    Somporn Techangamsuwan

  6. Biology Program, Faculty of Science, Buriram Rajabhat University, Muang, Buriram, 31000, Thailand

    Sirinee Jirajessada

  7. Department of Biochemistry, Faculty of Science, Mahidol University, Bangkok, 10400, Thailand

    Rattanavinan Hanchaina & Thaned Kangsamaksin

  8. Research Affairs, Faculty of Medicine, Chulalongkorn University, Ananda Mahidol Building, 1873 Rama 4 Rd, Pathumwan, Bangkok, 10330, Thailand

    Supansa Yodmuang

  9. Center of Excellence in Biomaterial Engineering for Medical and Health, Chulalongkorn University, Pathumwan, Bangkok, 10330, Thailand

    Supansa Yodmuang

  10. Clinical Excellence Center for Advanced Therapy Medicinal Products, King Chulalongkorn Memorial Hospital, Pathumwan, Bangkok, 10330, Thailand

    Supansa Yodmuang

  11. Avatar Biotech for Oral Health & Healthy Longevity Research Unit, Chulalongkorn University, Pathumwan, Bangkok, 10330, Thailand

    Supansa Yodmuang

  12. Department of Pharmacology, Faculty of Veterinary Science, Chulalongkorn University, Bangkok, 10330, Thailand

    Chenphop Sawangmake

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  1. Pensuda Sompunga
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Correspondence to Supansa Yodmuang or Chenphop Sawangmake.

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The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper.

Ethical Approval

Canine tracheae for decellularization were harvested under sterile conditions from canine cadaver, donated for anatomy class of the Faculty of Veterinary science, Chulalongkorn University, Thailand. Animal surgery and husbandry were performed by the Thailand guidelines on the use of experimental animals (ANIMALS FOR SCIENTIFIC PURPOSES ACT, B.E. 2558 (A.D. 2015). This study was approved by the Institutional Animal Care and Use Committee (IACUC) of Chulalongkorn University (No. 2231001).

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Sompunga, P., Rodprasert, W., Srisuwatanasagul, S. et al. Preparation of Decellularized Tissue as Dual Cell Carrier Systems: A Step Towards Facilitating Re-epithelization and Cell Encapsulation for Tracheal Reconstruction. Ann Biomed Eng 52, 1222–1239 (2024). https://doi.org/10.1007/s10439-024-03448-6

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