Medical Radiology and Radiation Safety. 2020. Vol. 65. No. 1. P. 5–12

V.A. Brunchukov, T.A. Astrelina, V.A. Nikitina, I.V. Kobzeva, Yu.B. Suchkova, D.Yu. Usupzhanova, A.A. Rastorgueva, T.V. Karaseva, T.V. Gordeev, O.A. Maxsimova, L.A. Naumova, S.V. Lischuk, E.A. Dubova, K.A. Pavlov, V.A. Brumberg, A.E. Makhova, E.E. Lomonosova, E.I. Dobrovolsskaya, I.M. Barabash, A.Yu. Bushmanov, A.S. Samoilov

Experimental Treatment of Radiation Skin Lesions with Mesenchymal Stem Cells and Their Conditioned Media

A.I. Burnasyan Federal Medical Biophysical Center, Moscow, Russia
E-mail: This email address is being protected from spambots. You need JavaScript enabled to view it.

Abstract

Purpose: To study the regeneration processes in the treatment of radiation skin lesions with the mesenchymal stem cells (MSC) derived from human gingiva and their conditional medium concentrate (CCM) during animal studies.

Material and methods: The study includes 80 white male Wistar rats weighing 210 ± 30 g at the age of 8–12 weeks, randomized into 4 groups (20 animals in each): control group (C), animal did not receive treatment; control with the introduction of the conditional medium concentrate (CCM) three times on days 1, 14 and 21; the introduction of MSC in a dose of 2 million cells per 1 kg three times on days 1, 14 and 21; the introduction of CCM in the estimated dose of 2 million cells per 1 kg three times on days 1, 14 and 21. Radiation burn simulation was performed (using on an X-ray unit at a dose of 110 Gy) and each animal was observed 17 times: at days 1, 7, 14, 21, 28, 35, 42, 49, 56, 63, 70, 77, 84, 91, 98, 105 and 112. Histological (stained with hematoxylin-eosin) and immunohistochemical (CD31, CD68, and VEGF) studies were performed. MSC was cultivated according to the standard procedure up to passages 3–5, the conditioned medium was collected and concentrated 10 times. The MSC immunophenotype (CD34, CD45, CD90, CD105, CD73, HLA-DR) and viability (7-ADD) were determined using flow cytometry.

Results: Under the assessment of the animal skin on the day 7 in the CCM group, the area was significantly larger compared to the C, MSC, CM groups (р ≤ 0.05). In the CM group on the day 14 the area of the open wound surface and ulcers from day 28 to day 42 was significantly less, compared with the C, MSC and CCM groups (р ≤ 0.05). In group C, from 42 to 77 days of observation, an increase in the area of skin ulcers was observed compared with the CM and CCM groups (р ≤ 0.05). On the day 112, healing of skin ulcers in the CM group was observed in 40 %, in the MSC group in 60 %, and only in 20 % of animals in the CCM group, and in the C group it was not registered. Expression of VEGF marker on endothelial cells and stromal cells was observed in groups C and CM on day 28 and in groups MSCs and CCM on day 112. On the 28th day in the MSC group, the average number of vessels (CD31) in the field of view was 6.0, and on day 112 it was 12.75, р ≤ 0.05, in the CCM group – 19.10 and 28.6, respectively, р ≤ 0.05. An increase in the number of macrophages (CD68) was found in group C from 28 to 112 days (11.6 and 24.73, р ≤ 0.05), and in the CM group the decrease was 22.1 and 13.07, respectively, р ≤ 0.05.

Conclusion: Thus, all used treatment modes of radiation skin lesions, including 3-fold administration of CM, MSC and CCM at a dose of 2 million cells per 1 kg, were effective and resulted in a reduction in the damage area, accelerated ulcer healing, and improvement of the regenerative processes. In addition, the use of MSCs led to the improvement of inflammatory processes’ vascularization and reduction in the radiation skin lesions.

Key words: mesenchymal stem cells, radiation skin lesions, conditioned medium, cellular technologies, X-rays, skin

For citation: Brunchukov VA, Astrelina TA, Nikitina VA, Kobzeva IV, Suchkova YuB, Usupzhanova DYu, Rastorgueva AA, Karaseva TV, Gordeev TV, Maxsimova OA, Naumova LA, Lischuk SV, Dubova EA, Pavlov KA, Brumberg VA, Makhova AE, Lomonosova EE, Dobrovolsskaya EI, Barabash IM, Bushmanov AYu, Samoilov AS. Experimental Treatment of Radiation Skin Lesions with Mesenchymal Stem Cells and Their Conditioned Media. Medical Radiology and Radiation Safety. 2020;65(1):5-0. (In Russ.).

DOI: 10.12737/1024-6177-2020-65-1-5-12

Список литературы / References

1. Радиационная медицина. Руководство для врачей-исследователей и организаторов здравоохранения. Под ред. Л.А. Ильина. М.: ИздАТ. 2001;(2):432 [Radiation Medicine. A Guide for Medical Researchers and Health Care Organizers. Ed. LA Ilyin. Moscow: Izdat. 2001;(2):432 (In Russ.)].
2. Howpel JW, Coggle JE, Wells J, et al. The acute effects of different energy beta-emiters on pig and mouse skin. Drit J Radiobiology. 1968; (190):47-51.
3. Осанов ДП. Дозиметрия и радиационная биофизика кожи. М.: Энергоатомиздат. 1983. 152 с. [Osanov DP. Dosimetry and Radiation Biophysics of the Skin. M.: Energoatomizdat. 1983:152 (In Russ.)].
4. Zheng K, Wu W, Jang S, et al. Bone marrow mesenchymal stem cell implantation for the treatment of radioactivity‑induced acute skin damage in rats. Molecular Medicine Reports. 2015;12:7065-71.
5. Мороз ББ, Онищенко НА, Лебедев ВГ, и др. Влияние мультипотентных мезенхимальных стромальных клеток костного мозга на течение местных лучевых поражений у крыс после локального β-облучения. Радиац. биол. Радиоэкология. 2009;49(6):688-93. [Moroz BB, Onishchenko NA, Lebedev VG, et al. Influence of multipotent mesenchymal bone marrow stromal cells on local radiation injury in rats after local β-irradiation. Radiation Biology. Radioecology. 2009;49(6):688-93 (In Russ.)].
6. Котенко КВ, Еремин ИИ, Мороз ББ, и др. Клеточные технологии в лечении радиационных ожогов: опыт ФМБЦ им. А.И. Бурназяна. Клеточная трансплантология и тканевая инженерия. 2012;7(2):97-102. [Kotenko KV, Eremin LI, Moroz BB, et al. Cell technologies in the treatment of radiation burns: experience of the Burnasyan Federal Medical Biophysical Centre. Cell Transplantation and Tissue Engineering. 2012;7(2):97-102 (In Russ.)].
7. Темнов АА, Астрелина ТА, Рогов КА, и др. Исследование влияния факторов кондиционированной среды, полученной при культивировании мезенхимальных стволовых клеток костного мозга, на течение тяжелых местных лучевых поражений кожи у крыс. Медицинская радиология и радиационная безопасность. 2018;63(1):35-9. [Temnov AA, Astrelina TA, Rogov КА, et al. Investigation of the influence of the conditioning medium factors obtained during the cultivation of bone marrow mesenchymal stem cells on the course of severe local radiation injuries of skin in rats. Medical Radiology and Radiation Safety. 2018;63(1):35-9. (In Russ.)].
8. Kim HS, Choi DY, Yun SJ, et al. Proteomic analysis of microvesicles derived from human mesenchymal stem cells. J Proteome Res. 2012;11(2):839-49.
9. Daltro PS, Barreto BC, Silva PG, et al. Therapy with mesenchymal stromal cells or conditioned medium reverse cardiac alterations in a high-fat diet-induced obesity model. The International Society for Cellular Therapy Position Statement Cytotherapy. 2017;19:1176-88.
10. Gnecchi M, Zhang Z, Ni A, Dzau VJ. Paracrine mechanisms in adult stem cells signaling and therapy. Circ Res. 2008;103:1204-19.
11. Lee C, Shim S, Jang H, et al. Human umbilical cord blood derived mesenchymal stromal cells and small intestinal submucosa hydrogel composite promotes combined radiation-wound healing of mice. The International Society for Cellular Therapy Position Statement Cytotherapy. 2017;19(9):1048-59.
12. Öksüz S, Şahin Alagöz M, Karagöz H, et al. Comparison of treatments with local mesenchymal stem cells and mesenchymal stem cells with increased vascular endothelial growth factor fxpression on irradiation injury of expanded skin. Ann Plast Surg. 2015;75:219-30.
13. Mitrano TI, Grob MS, Carrión F, et al. Culture and Characterization of Mesenchymal Stem Cells From Human Gingival Tissue. Journal of Periodontology. 2010;81:917-25.
14. Zhang QZ, Nguyen AL, Yu WH, Le AD. Human oral mucosa and gingiva: a unique reservoir for mesenchymal stem cells. J Dent Res. 2012;91(11):1011-8.
15. Duan HG, Ji F, Zheng CQ, et al. Conditioned medium from umbilical cord mesenchymal stem cells improves nasal mucosa damage by radiation. Biotechnol Lett. 2018;40(6):999-1007.
16. Dominici M, Le Blanc K, Mueller I, Slaper-Cortenbach I, Marini F, Krause D, et al. Minimal criteria for defining multipotent mesenchymal stromal cells. The International Society for Cellular Therapy Position Statement. Cytotherapy. 2006;8:315-7.

PDF (RUS) Full-text article (in Russian)

Conflict of interest. The authors declare no conflict of interest.

Financing. The study was financially supported by the Federal Target Program “Ensuring Nuclear and Radiation Safety for 2016–2020. and for the period until 2030”. A series of publications is planned based on the results of the studies.

Contribution: Article was prepared with equal participation of the authors.

Compliance with the rules of bioethics. The study was performed in accordance with the ethical standards for the treatment of animals adopted by the European Convention for the protection of vertebrates used for research and other scientific purposes.

Article received: 12.10.2018. Accepted for publication: 11.12.2019