vtioВестник трансплантологии и искусственных органовRussian Journal of Transplantology and Artificial Organs1995-1191Academician V.I.Shumakov National Medical Research Center of Transplantology and Artificial Organs", Ministry of Health of the Russian Federation10.15825/1995-1191-2022-4-109-117vtio-1555Research ArticleРегенеративная медицина и клеточные технологииRegenerative Medicine and Cell TechnologiesВлияние биоматриксов на жизнеспособность и инсулинпродуцирующую функцию островков Лангерганса человека in vitroIn vitro effect of bioscaffolds on viability and insulin‑producing function of human islets of LangerhansПономареваА. С.PonomarevaA. S.

Пономарева Анна Сергеевна

123182, Москва, ул. Щукинская, д. 1

Тел.: (499) 196-26-61, (926) 585-23-73

Anna Ponomareva

1, Shchukinskaya str., Moscow, 123182, Russian Federation

Phone: (499) 196-26-61, (926) 585-23-73

a.s.ponomareva@gmail.comБарановаН. В.BaranovaN. V.

Москва

Moscow

МилосердовИ. А.MiloserdovI. A.

Москва

Moscow

СевастьяновВ. И.SevastianovV. I.

Москва

Moscow

ФГБУ «Национальный медицинский исследовательский центр трансплантологии и искусственных органов имени академика В.И. Шумакова» Минздрава РоссииРоссияShumakov National Medical Research Center of Transplantology and Artificial OrgansRussian Federation202229082022244109117Copyright © Пономарева А.С., Баранова Н.В., Милосердов И.А., Севастьянов В.И., 20222022Пономарева А.С., Баранова Н.В., Милосердов И.А., Севастьянов В.И.Ponomareva A.S., Baranova N.V., Miloserdov I.A., Sevastianov V.I.Данная работа распространяется под лицензией Creative Commons Attribution 4.0.This work is licensed under a Creative Commons Attribution 4.0 License.https://journal.transpl.ru/vtio/article/view/1555

Культивирование островков Лангерганса с биоматриксами – миметиками внеклеточного матрикса (ВКМ) может обеспечивать характерное для островков нативное микроокружение, что является одним из основных условий создания тканевого эквивалента поджелудочной железы (ПЖ).

Цель работы

Цель работы: сравнение секреторной способности жизнеспособных панкреатических островков человека в монокультуре (контрольная группа) и культивированных в присутствии двух биоматриксов: биополимерного коллагенсодержащего гидрогелевого матрикса (опытная группа 1) и тканеспецифического матрикса из децеллюляризованной ПЖ посмертного донора (опытная группа 2).

Материалы и методы

Материалы и методы. Островки Лангерганса выделяли из хвостовой части ПЖ по методике с использованием коллагеназы. Жизнеспособность культивированных островков определяли методом флуоресцентного окрашивания витальным красителем, секреторную способность – методом иммуноферментного анализа (ИФА).

Результаты

Результаты. Панкреатические островки, культивированные с биоматриксами, не проявляли признаков деградации и фрагментации и оставались жизнеспособными в течение всего срока наблюдения (7 суток). В монокультуре островков на этом сроке происходили значительные деструктивные изменения. Базальная концентрация инсулина в опытных группах 1 и 2 на первые сутки культивирования повышалась на 18,8 и 39,5% по сравнению с контрольной группой, на четвертые сутки инкубации – на 72,8 и 102,7%, на седьмые сутки – на 146,4 и 174,6% соответственно. Уровень секреции инсулина островков с тканеспецифическим матриксом был на 17,4% выше, чем при культивировании с биополимерным коллагенсодержащим матриксом.

Заключение

Заключение. Биополимерный и тканеспецифический миметики ВКМ способствуют не только сохранению жизнеспособности изолированных островков Лангерганса, но и поддержанию их инсулинпродуцирующей функции в течение 7 суток на более высоком уровне по сравнению с монокультурой. В условиях проведенных экспериментов выявлено незначительное потенциальное преимущество применения тканеспецифического матрикса по сравнению с биополимерным матриксом для создания тканевого эквивалента поджелудочной железы.

The culture of islets of Langerhans with bioscaffolds – extracellular matrix (ECM) mimetics – can provide a native microenvironment suitable for islets. This is one of the main conditions for creating a pancreatic tissue equivalent.

Objective

Objective: to compare the secretory capacity of viable human pancreatic islets in monoculture (control group) and cultured in the presence of two bioscaffolds: biopolymer collagen-based hydrogel scaffold (experimental group 1) and tissue-specific scaffold from decellularized deceased donor pancreas (experimental group 2).

Materials and methods

Materials and methods. Islets of Langerhans were isolated from the caudal pancreas using a collagenase technique. The viability of cultured islets was accessed by vital fluorescence staining, while secretory capacity was evaluated by enzyme-linked immunosorbent assay (ELISA).

Results

Results. Pancreatic islets cultured with bioscaffolds showed no signs of degradation and fragmentation, they remained viable throughout the entire period of observation (7 days). The monoculture of islets showed significant destructive changes during this period. Basal insulin levels in experimental groups 1 and 2 increased by 18.8% and 39.5% on day 1 of culture compared to the control group, by 72.8% and 102.7% on day 4 of incubation, and by 146.4% and 174.6% on day 7, respectively. The insulin secretion level of islets with tissue-specific scaffolds was 17.4% higher than that when cultured with biopolymer collagen-based scaffolds.

Conclusion

Conclusion. Biopolymer and tissue-specific ECM mimetics contribute not only to preservation of the viability of isolated islets of Langerhans but also maintain their insulin secretion capacity for 7 days at a higher level in comparison with monoculture. The experiments revealed that the use of a tissue-specific scaffold for the creation of a pancreatic tissue equivalent has slight potential advantage over biopolymer scaffold.

поджелудочная железакультивирование островков Лангергансаинсулинпродуцирующая функциятканеспецифический матриксбиополимерный матрикpancreasculture of the islets of Langerhansinsulin-producing functiontissue-specific scaffoldbiopolymer scaffoldReferencesKumar N, Joisher H, Ganguly A. Polymeric scaffolds for pancreatic tissue engineering: a review. Rev Diabet Stud. 2018; 14 (4): 334–353.Kumar N, Joisher H, Ganguly A. Polymeric scaffolds for pancreatic tissue engineering: a review. Rev Diabet Stud. 2018; 14 (4): 334–353.Creusot RJ, Battaglia M, Roncarolo MG et al. Concise review: cell-based therapies and other non-traditional approacher for type 1 diabetes. Stem Cells. 2016; 34 (4):809–819.Creusot RJ, Battaglia M, Roncarolo MG et al. Concise review: cell-based therapies and other non-traditional approacher for type 1 diabetes. Stem Cells. 2016; 34 (4):809–819.Gururaj Setty S, Crasto W, Jarvis J et al. New insulins and newer insulin regimens: a review of their role in improving glycaemic control in patients with diabetes. Postgrad Med J. 2016; 92 (1085): 152–164.Gururaj Setty S, Crasto W, Jarvis J et al. New insulins and newer insulin regimens: a review of their role in improving glycaemic control in patients with diabetes. Postgrad Med J. 2016; 92 (1085): 152–164.Gan MJ, Albanese-O’Neill A, Haller MJ. Type 1 diabetes: current concepts in epidemiology, pathophysiology, clinical care, and research. Curr Probl Pediatr Adolesc Health Care. 2012; 42 (10): 269–291.Gan MJ, Albanese-O’Neill A, Haller MJ. Type 1 diabetes: current concepts in epidemiology, pathophysiology, clinical care, and research. Curr Probl Pediatr Adolesc Health Care. 2012; 42 (10): 269–291.Mannucci E, Monami M, Dicembrini I et al. Achieving HbA1c targets in clinical trials and in the real world: a systematic review and meta-analysis. J Endocrinol Invest. 2014; 37 (5): 477–495.Mannucci E, Monami M, Dicembrini I et al. Achieving HbA1c targets in clinical trials and in the real world: a systematic review and meta-analysis. J Endocrinol Invest. 2014; 37 (5): 477–495.Bottino R, Knoll MF, Knoll CA et al. The future of islet transplantation is now. Front Med (Lausanne). 2018; 5: 202.Bottino R, Knoll MF, Knoll CA et al. The future of islet transplantation is now. Front Med (Lausanne). 2018; 5: 202.Shapiro AM, Pokrywczynska M, Ricordi C. Clinical pancreatic islet transplantation. Nat Rev Endocrinol. 2017; 13 (5): 268–277.Shapiro AM, Pokrywczynska M, Ricordi C. Clinical pancreatic islet transplantation. Nat Rev Endocrinol. 2017; 13 (5): 268–277.Maffi P, Secchi A. Clinical results of islet transplantation. Pharmacol Res. 2015; 98: 86–91.Maffi P, Secchi A. Clinical results of islet transplantation. Pharmacol Res. 2015; 98: 86–91.Ehlers MR. Strategies for clinical trials in type 1 diabetes. J Autoimmun. 2016; 71: 88–96.Ehlers MR. Strategies for clinical trials in type 1 diabetes. J Autoimmun. 2016; 71: 88–96.Bosco D, Armanet M, Morel P et al. Unique arrangement of alpha- and beta-cells in human islets of Langerhans. Diabetes. 2010; 59 (5): 1202–1210.Bosco D, Armanet M, Morel P et al. Unique arrangement of alpha- and beta-cells in human islets of Langerhans. Diabetes. 2010; 59 (5): 1202–1210.Amer LD, Mahoney MJ and Bryant SJ. Tissue engineering approaches to cell-based type 1 diabetes therapy. Tissue engineering. 2014; 20 (5): 455–467.Amer LD, Mahoney MJ and Bryant SJ. Tissue engineering approaches to cell-based type 1 diabetes therapy. Tissue engineering. 2014; 20 (5): 455–467.Riopel M, Wang К. Collagen matrix support of pancreatic islet survival and function. Frontiers in Bioscience. 2014; 19: 77–90. doi: 10.2741/4196.Riopel M, Wang К. Collagen matrix support of pancreatic islet survival and function. Frontiers in Bioscience. 2014; 19: 77–90. doi: 10.2741/4196.Пономарева АС, Кирсанова ЛА, Баранова НВ, Бубенцова ГН, Милосердов ИА, Волкова ЕА, Севастьянов ВИ. Методика выделения жизнеспособных островков Лангерганса из фрагмента хвостовой части поджелудочной железы человека. Вестник трансплантологии и искусственных органов. 2018; 20 (4): 76–82.Ponomareva AS, Kirsanova LA, Baranova NV, Bubentsova GN, Miloserdov IA, Volkova EA, Sevastianov VI. A technique for separating viable islets of Langerhans from a fragment of human pancreatic tail. Russian Journal of Transplantology and Artificial Organs. 2018; 20 (4): 76–82. doi.org/10.15825/1995-1191-2018-4-76-82.Llacua LA, Faas MM, de Vos P. Extracellular matrix molecules and their potential contribution to the function of transplanted pancreatic islets. Diabetologia. 2018; 61 (6): 1261–1272.Llacua LA, Faas MM, de Vos P. Extracellular matrix molecules and their potential contribution to the function of transplanted pancreatic islets. Diabetologia. 2018; 61 (6): 1261–1272.Севастьянов ВИ, Перова НВ. Инъекционный гетерогенный биополимерный гидрогель для заместительной и регенеративной хирургии и способ его получения. Патент на изобретение РФ № 2433828. 2011.Sevastianov VI. Perova NV. Injectable heterogeneous biopolymer hydrogel for replacement and regenerative surgery and the method of its preparation. [Patent for invention of the Russian Federation No. 2433828]. 2011.Goh SK, Bertera S, Olsen P et al. Perfusion-decellularized pancreas as a natural 3D scaffold for pancreatic tissue and whole organ engineering. Biomaterials. 2013; 34 (28): 6760–6772.Goh SK, Bertera S, Olsen P et al. Perfusion-decellularized pancreas as a natural 3D scaffold for pancreatic tissue and whole organ engineering. Biomaterials. 2013; 34 (28): 6760–6772.Napierala H, Hillebrandt K-H, Haep N, Tang P, Tintemann M, Gassner J et al. Engineering an endocrine neopancreas by repopulation of a decellularized rat pancreas with islets of Langerhans. Sci Rep. 2017; 2 (7): 41777. doi: 10.1038/srep41777.Napierala H, Hillebrandt K-H, Haep N, Tang P, Tintemann M, Gassner J et al. Engineering an endocrine neopancreas by repopulation of a decellularized rat pancreas with islets of Langerhans. Sci Rep. 2017; 2 (7): 41777. doi: 10.1038/srep41777.Sevastianov VI, Baranova NV, Kirsanova LA, Ponomareva AS, Basok YuB, Nemets EA, Gautier SV. Comparative analysis of the influence of extracellular matrix biomimetics on the viability and insulin-producing function of isolated pancreatic islets. J Gene Engg Bio Res. 2021; 3 (2): 17–25.Sevastianov VI, Baranova NV, Kirsanova LA, Ponomareva AS, Basok YuB, Nemets EA, Gautier SV. Comparative analysis of the influence of extracellular matrix biomimetics on the viability and insulin-producing function of isolated pancreatic islets. J Gene Engg Bio Res. 2021; 3 (2): 17–25.Rana D, Zreigat H, Benkirane-Jessel N et al. Development of decellularized scaffolds for stem cell-driven tissue engineering. J Tissue Eng Regen Med. 2017; 11 (4): 942–965.Rana D, Zreigat H, Benkirane-Jessel N et al. Development of decellularized scaffolds for stem cell-driven tissue engineering. J Tissue Eng Regen Med. 2017; 11 (4): 942–965.Shirakigawa N, Ijima H. Decellularized tissue engineering. Advances in Biomaterials for Biomedical Applications. Springer Nature Singapore Pte Ltd. 2017; 66: 185–226.Shirakigawa N, Ijima H. Decellularized tissue engineering. Advances in Biomaterials for Biomedical Applications. Springer Nature Singapore Pte Ltd. 2017; 66: 185–226.Пономарева АС, Кирсанова ЛА, Баранова НВ, Сургученко ВА, Бубенцова ГН, Басок ЮБ и др. Децеллюляризация фрагмента донорской поджелудочной железы для получения тканеспецифического матрикса. Вестник трансплантологии и искусственных органов. 2020; 22 (1): 123–133.Ponomareva AS, Kirsanova LA, Baranova NV, Surguchenko VA, Bubentsova GN, Basok YuB et al. Decellularization of donor pancreatic fragment to obtain a tissue-specific matrix scaffold. Russian Journal of Transplantology and Artificial Organs. 2020; 22 (1): 123–133. doi: 10.15825/1995-1191-2020-1-123-133.Sackett SD, Tremmel DM, Ma F et al. Extracellular matrix scaffold and hydrogel derived from decellularized and delipidized human pancreas. Sci Rep. 2018; 8 (1): 10452.Sackett SD, Tremmel DM, Ma F et al. Extracellular matrix scaffold and hydrogel derived from decellularized and delipidized human pancreas. Sci Rep. 2018; 8 (1): 10452.Guruswamy Damodaran R, Vermette P. Decellularized pancreas as a native extracellular matrix scaffold for pancreatic islet seeding and culture. J Tissue Eng Regen Med. 2018; 12 (5): 1230–1237.Guruswamy Damodaran R, Vermette P. Decellularized pancreas as a native extracellular matrix scaffold for pancreatic islet seeding and culture. J Tissue Eng Regen Med. 2018; 12 (5): 1230–1237.Salvatori M, Katari R, Patel T, Peloso A, Mugweru J, Owusu K, Orlando G. Extracellular matrix scaffold technology for bioartificial pancreas engineering: state of the art and future challenges. J DiabSci Technol. 2014; 8 (1): 159–169. doi: 10.1177/1932296813519558.Salvatori M, Katari R, Patel T, Peloso A, Mugweru J, Owusu K, Orlando G. Extracellular matrix scaffold technology for bioartificial pancreas engineering: state of the art and future challenges. J DiabSci Technol. 2014; 8 (1): 159–169. doi: 10.1177/1932296813519558.Mirmalek-Sani S-H, Orlando G, McQuilling J, Pareta R, Mack D, Salvatori M et al. Porcine pancreas extracellular matrix as a platform endocrine pancreas bioengineering. Biomaterials. 2013; 34 (22): 5488–5495. doi: 10.1016/j.biomaterials.2013.03.054.Mirmalek-Sani S-H, Orlando G, McQuilling J, Pareta R, Mack D, Salvatori M et al. Porcine pancreas extracellular matrix as a platform endocrine pancreas bioengineering. Biomaterials. 2013; 34 (22): 5488–5495. doi: 10.1016/j.biomaterials.2013.03.054.

The authors declare that there are no conflicts of interest present.