Effect of GHK-D-Ala peptide on innate immunity mechanisms and lipid peroxidation processes in infected wounds

The objective is to study the effects of Gly-His-Lys-D-Ala (GHK-D-Ala) peptide on mechanisms of innate immunity and lipid peroxidation processes in infected wound.

Materials and methods. The experiments were carried out on 80 Wistar male rats. The infected wound was modeled by applying a full-layer wound on the back and performing primary wound surgery with primary suturing 24 hours later. GHK-D-Ala peptide was injected at a dose of 0.5 µg/kg into the wound area every day for 30 days. The phagocytic activity of blood neutrophils was studied in spontaneous and stimulated nitroblue tetrazolium reduction test (NBT-test). The activity of lipid peroxidation (LPO) processes in rats’ blood serum was assessed by the content of malondialdehyde (MDA) and acylhydroperoxides (AHP).

Results. GHK-D-Ala administration had a positive effect on both phagocytosis mechanisms and the level of LPO products in comparison with the control group. There was a significant increase in NST-positive neutrophils in the spontaneous test against peptide administration on days 3, 7, 10, and 30 (p˂0.001) and in the stimulated NST test on days 3, 7, and 10 (p < 0.001). The functional neutrophil reserve had been significantly increased by day 10 (p˂0.05) and had been significantly lower by day 30 (p < 0.05). Phagocytic index and phagocytic number were significantly higher on days 3 and 10 (p < 0.01). The phagocyte activity index significantly increased on days 3 and 10 (p < 0.05-0.001).A significant decrease in the MDA concentration was demonstrated on days 3, 7, 10, and 30 (p<0,001). There was a significant decrease in the content of AGP on days 3, 7, 10 and 30 (p < 0.001).

Conclusion. The corrective effect of Gly-His-Lys-D-Ala peptide on the phagocytic activity of granulocytes and the processes of lipid peroxidation in infected skin wounds was established.

1. Аралова М.В., Глухов А.А., Остроушко А.П. Кинетика раневого процесса при различных методах стимуляции регенерации в ранах. Вестник экспериментальной и клинической хирургии. 2018;11(3):173-178. DOI: 10.18499/2070-478X-2018-11-3-173-178. URL: https://vestnik-surgery.com/index.php/journal/article/view/1209

2. Гомазков О.А. Нейротрофические факторы мозга: справочно-информационное издание. Электронная версия. Разделы 2-3. Москва: EBEWE Pharma, 2004. с. 89-178

3. Медведев А.Н., Маянский А.Н., Чаленко В.В. Способ исследования поглотительной фазы фагоцитоза. Лабораторное дело. 1991;(2):19-20

4. Рагино Ю.И., Душкин М.И. Определение резистентности к окислению гепарин-осажденных b-липопротеинов сыворотки крови у больных ишемической болезнью сердца. Клиническая лабораторная диагностика. 1998;(11):3-5

5. Смахтин М.Ю., Курцева А.А., Чердаков В.Ю. Иммунорегуляторное и гепатотропное действие пептидов Gly-His-Lys, DSLET и АКТГ 4-10. Вестник Уральской медицинской академической науки. 2006;14(3-1):223

6. Морозов А.М., Сергеев А.Н., Сергеев Н.А., Дубатолов Г.А., Рыжова Т.С., Пахомов М.А., Пельтихина О.В. Современные методы стимуляции процесса регенерации послеоперационных ран. Сибирское медицинское обозрение. 2020;(3):54-60. DOI: 10.20333/2500136-2020-3-54-60

7. Хавинсон В.Х. Лекарственные пептидные препараты: прошлое, настоящее, будущее. Клиническая медицина. 2020;98(3):165-177. DOI: 10.30629/0023-2149-2020-98-3-165-177

8. Щербаков В.И. Применение НСТ-теста для оценки чувствительности нейтрофилов к стимуляторам. Лабораторное дело. 1989;(1):30-33

9. Siméon A., Monier F., Emonard H., Gillery P., Birembaut P., Hornebeck W., Maquart F.X. Expression and activation of matrix metalloproteinases in wounds: modulation by the tripeptide-copper complex glycyl-L-histidyl-L-lysine-Cu2+. J Invest Dermatol. 1999;112(6):957-964. DOI: 10.1046/j.1523-1747.1999.00606.x

10. Hur G.H., Han S.C., Ryu A.R., Eom Y., Kim J.W., Lee M.Y. Effect of oligoarginine conjugation on the antiwrinkle activity and transdermal delivery of GHK peptide. J Pept Sci. 2020;26(2):e3234. DOI: 10.1002/psc.3234

11. Ahmed M.R., Basha S.H., Gopinath D., Muthusamy R., Jayakumar R. Initial upregulation of growth factors and inflammatory mediators during nerve regeneration in the presence of cell adhesive peptide-incorporated collagen tubes. J Peripher Nerv Syst. 2005;10(1):17-30. DOI: 10.1111/j.1085-9489.2005.10105.x

12. Kazemi A., Frazier T., Cave M. Micronutrient-related neurologic complications following bariatric surgery. Curr Gastroenterol Rep. 2010;12(4):288-295. DOI: 10.1007/s11894-010-0120-5

13. Mazurowska L., Mojski M. Biological activities of selected peptides: skin penetration ability of copper complexes with peptides. J Cosmet Sci. 2008;59(1): 59-69.

14. Pickart L. The human tri-peptide GHK and tissue remodeling. J Biomater Sci Polym Ed. 2008;19(8): 969-988. DOI: 10.1163/156856208784909435

15. Pickart L., Pickart E. A possible mechanism whereby skin remodeling may suppress cancer metastasis genes. Wound Repair and Regeneration. 2011;19(2):A42-A42.

16. Pickart L., Vasquez-Soltero J.M., Margolina A. The human tripeptide GHK-Cu in prevention of oxidative stress and degenerative conditions of aging: implications for cognitive health. Oxid Med Cell Longev. 2012;(2012):324832. DOI: 10.1155/2012/324832

17. Pilgeram L. Control of fibrinogen biosynthesis: role of the FFA/albumin ratio. Cardiovasc Eng. 2010;10(2):78-83. DOI: 10.1007/s10558-010-9092-1

18. Pickart L., Margolina A. Regenerative and Protective Actions of the GHK-Cu Peptide in the Light of the New Gene Data. Int J Mol Sci. 2018 J;19(7):1987. DOI: 10.3390/ijms19071987

19. Pickart L., Vasquez-Soltero J.M., Margolina A. GHK Peptide as a Natural Modulator of Multiple Cellular Pathways in Skin Regeneration. Biomed Res Int. 2015;2015:648108. DOI: 10.1155/2015/648108

20. Siméon A., Emonard H., Hornebeck W., Maquart F.X. The tripeptide-copper complex glycyl-L-histidyl-L-lysine-Cu2+ stimulates matrix metalloproteinase-2 expression by fibroblast cultures. Life Sci. 2000;67(18):2257-2265. DOI: 10.1016/s0024-3205(00)00803-1

21. Pastar I., Stojadinovic O., Yin N.C., Ramirez H., Nusbaum A.G., Sawaya A., Patel S.B., Khalid L. et al. Epithelialization in Wound Healing: A Comprehensive Review. Adv Wound Care (New Rochelle). 2014;3(7):445-464. DOI: 10.1089/wound.2013.0473

22. Rowan M.P., Cancio L.C., Elster E.A., Burmeister D.M., Rose L.F., Natesan S., Chan R.K., Christy R.J. et al. Burn wound healing and treatment: review and advancements. Crit Care. 2015;(19):243. DOI: 10.1186/s13054-015-0961-2