Ischemic Non-Healing Skin Wound Model: Cell Death and Wound Healing Mechanisms

The aim of the study was to develop an ischemic skin flap model in laboratory mice with the description of necrotic and apoptotic processes in time course, and demonstrate the regenerative characteristics of non-healing skin wound under the conditions of a formed flap.

Materials and Methods. The study involved 20 BALB/c mice. The first experimental stage describes an ischemic flap on day 1, 2, 3, and 5 after flap formation using histology, immunofluorescence, morphometry, and TUNEL. At the second stage a non-healing wound was modeled on the flap, and its pathogenesis was histologically compared with an acute wound.

Results. A formed flap showed such histological ischemic features as blood-filled vessels, micro blood clots in blood vessels, coagulation necrosis, migration of inflammatory cells, as well as extravasation, true skin edema and hyperproliferation of epithelial cells. Maximum pathological changes were found on the third postoperative day, and by day 5 the wound partially healed. Morphometry demonstrated a significant increase in blood-filled vessels and the tendency for epithelial layer reduction in a skin flap of experimental animals compared to the control ones. Apoptotic cells were observed on days 1, 2, 3, and 5 of a formed flap. Some areas showed cell clusters around hair follicles. No apoptotic cells were revealed in control animals.

By the moment of complete regeneration of acute wounds, non-healing wounds exhibited no epithelialization and a formed mature cicatrix that enables to interpret the difference in wound healing as significant.

Conclusion. Ischemia causes major pathological skin changes, as well as results in delayed wound healing. A developed non-healing wound model can be used to study wound healing mechanisms and the ways to influence them.

1. Das S., Baker A. Biomaterials and nanotherapeutics for enhancing skin wound healing. Front Bioeng Biotechnol 2016; 4: 82, https://doi.org/10.3389/fbioe.2016.00082.
2. Zhou K., Ma Y., Brogan M.S. Chronic and non-healing wounds: the story of vascular endothelial growth factor. Med Hypotheses 2015; 85(4): 399–404, https://doi.org/10.1016/j.mehy.2015.06.017.
3. Demidova-Rice T., Hamblin M., Herman I. Acute and impaired wound healing: pathophysiology and current methods for drug delivery, part 2: role of growth factors in normal and pathological wound healing: therapeutic potential and methods of delivery. Adv Skin Wound Care 2012; 25(8): 349–370, https://doi.org/10.1097/01.asw.0000418541.31366.a3.
4. Zhang Q., Chang Q., Cox R.A., Gong X., Gould L.J. Hyperbaric oxygen attenuates apoptosis and decreases inflammation in an ischemic wound model. J InvestDermatol 2008; 128(8): 2102–2112, https://doi.org/10.1038/jid.2008.53.
5. Gould L., Leong M., Sonstein J., Wilson S. Optimization and validation of an ischemic wound model. Wound Repair Regen 2005; 13(6): 576–582, https://doi.org/10.1111/j.1524-475x.2005.00080.x.
6. Lee D., Hong H., Roh H., Lee W. The effect of polydeoxyribonucleotide on ischemic rat skin flap survival. Ann Plast Surg 2015; 75(1): 84–90, https://doi.org/10.1097/sap.0000000000000053.
7. Weinlich R., Oberst A., Beere H., Green D. Necroptosis in development, inflammation and disease. Nat Rev Mol Cell Biol 2016; 18(2): 127–136, https://doi.org/10.1038/nrm.2016.149.
8. Caccamo A., Branca C., Piras I.S., Ferreira E., Huentelman M.J., Liang W.S., Readhead B., Dudley J.T., Spangenberg E.E., Green K., Belfiore R. Necroptosis activation in Alzheimer’s disease. Nat Neurosci 2017; 20(9): 1236–1246, https://doi.org/10.1038/nn.4608.
9. Tsai T.C., Tung Y.T., Kuo Y.H., Liao J.W., Tsai H.C., Chong K.Y., Chen H.L., Chen C.M. Anti-inflammatory effects of Antrodia camphorata, a herbal medicine, in a mouse skin ischemia model. J Ethnopharmacol 2015; 159: 113–121, https://doi.org/10.1016/j.jep.2014.11.015.
10. Atlas and synopsis of lever’s histopathology of the skin. Edited by Elder D.E., Elenitsas R., Rubin A.I., Iofredda M., Miller J., Miller O.F. Lippincott Williams & Wilkins; 2013.
11. Rah D., Min H., Kim Y., Cheon Y. Effect of platelet-rich plasma on ischemia-reperfusion injury in a skin flap mouse model. Int J Med Sci 2017; 14(9): 829–839, https://doi.org/10.7150/ijms.19573.
12. Rany iranevayainfektsiya [Wounds and wound infection]. Pod. red. Kuzinа M.I., Kostyuchenok B.M. [Kuzin M.I., Kostyuchenok B.M. (editors)]. Moscow: Meditsina; 1990.
13. Moor A.N., Tummel E., Prather J.L., Jung M., Lopez J.J., Connors S., Gould L.J. Consequences of age on ischemic wound healing in rats: altered antioxidant activity and delayed wound closure. Age 2014; 36(2): 733–748, https://doi.org/10.1007/s11357-014-9617-4.

Morgun Е.I., Rogovaya О.S., Vorotelyak Е.А. Ischemic Non-Healing Skin Wound Model: Cell Death and Wound Healing Mechanisms. Sovremennye tehnologii v medicine 2018; 10(4): 69, https://doi.org/10.17691/stm2018.10.4.08