title: Topical Administration of Oxygenated Hemoglobin Improved Wound Healing in an Ischemic Rabbit Ear Model.
abstract: BACKGROUND
  Localized oxygen deficiency plays a central role in the pathogenesis of chronic wounds; thus, rectifying localized ischemia with oxygen therapy has been postulated to be an integral aspect of the management of chronic wounds. The efficacy of a novel approach for oxygen therapy on chronic wound healing was evaluated.
  
  
  METHODS
  Oxygen was delivered to ischemic wounds by means of the topical application of oxygenated, chemically modified bovine hemoglobin (IKOR 2084) in a validated rabbit ear ischemic wound model. The wound healing was evaluated histologically by measuring epithelial gap and neo-granulation tissue area. In situ expression of endothelial cells (CD31) and proliferative cells (Ki-67) was examined by immunohistochemistry analysis. The mRNA of vascular endothelial growth factor, endothelial nitric oxide synthase, and matrix metalloproteinase-9 was quantified by real-time reverse-transcriptase polymerase chain reaction. The collagen was detected by Sirius red staining.
  
  
  RESULTS
  In comparison with topical application of saline, the administration of oxygenated IKOR 2084 increases wound reepithelialization and formation of neo-granulation tissue in a dose-dependent manner, and cellular proliferation (Ki-67). Conversely, the administration of deoxygenated IKOR 2084 aggravated the ischemic wound healing process. Moreover, the topical administration of oxygenated IKOR 2084 induces angiogenesis as evidenced by concomitant increases in CD31 protein and vascular endothelial growth factor and endothelial nitric oxide synthase mRNA expression in treated wounds. Oxygenated IKOR 2084 administration also increased collagen deposition in wounds, with decreases in the expression of matrix metalloproteinase-9 mRNA.
  
  
  CONCLUSIONS
  This study suggests that the topical application of oxygenated IKOR 2084 ameliorates the reparative progress of ischemic wounds through enhanced angiogenesis, cellular proliferation, and collagen deposition.
authors.affiliation: Chicago, Ill.; and Aberdeen, S.D. From the Division of Plastic Surgery, Northwestern University Feinberg School of Medicine; and Clinical Research Division, IKOR Life Sciences, Inc.
authors.forename: Ping
authors.initials: P
authors.lastname: Xie
authors.forename: Shengxian
authors.initials: S
authors.lastname: Jia
authors.forename: Ross
authors.initials: R
authors.lastname: Tye
authors.forename: Wei
authors.initials: W
authors.lastname: Xu
authors.forename: Aimei
authors.initials: A
authors.lastname: Zhong
authors.forename: Seok J
authors.initials: SJ
authors.lastname: Hong
authors.forename: Robert D
authors.initials: RD
authors.lastname: Galiano
authors.forename: Thomas A
authors.initials: TA
authors.lastname: Mustoe
mesh_terms: D000287:Administration, Topical; D000818:Animals; D002417:Cattle; D049109:Cell Proliferation; D004195:Disease Models, Animal; D004423:Ear / Q000098:blood supply*; D005260:Female; D006454:Hemoglobins / Q000008:administration & dosage*; D011817:Rabbits; D015427:Reperfusion Injury / Q000188:drug therapy* / Q000378:metabolism / Q000473:pathology; D016896:Treatment Outcome; D014945:Wound Healing / Q000187:drug effects*
chemical_list: D006454:Hemoglobins
references.citation: Gurtner GC, Werner S, Barrandon Y, Longaker MT. Wound repair and regeneration. Nature. 2008;453:314–321
references.citation: Valacchi G, Zanardi I, Sticozzi C, Bocci V, Travagli V. Emerging topics in cutaneous wound repair. Ann N Y Acad Sci. 2012;1259:136–144
references.citation: Ahn ST, Mustoe TA. Effects of ischemia on ulcer wound healing: A new model in the rabbit ear. Ann Plast Surg. 1990;24:17–23
references.citation: Kranke P, Bennett MH, Martyn-St James M, Schnabel A, Debus SE. Hyperbaric oxygen therapy for chronic wounds. Cochrane Database Syst Rev. 2012;4:CD004123
references.citation: Oztürk F, Ermertcan AT, Inanir I. Hyperbaric oxygen therapy for the management of chronic wounds. Cutan Ocul Toxicol. 2013;32:72–77
references.citation: Brimson CH, Nigam Y. The role of oxygen-associated therapies for the healing of chronic wounds, particularly in patients with diabetes. J Eur Acad Dermatol Venereol. 2013;27:411–418
references.citation: Howard MA, Asmis R, Evans KK, Mustoe TA. Oxygen and wound care: A review of current therapeutic modalities and future direction. Wound Repair Regen. 2013;21:503–511
references.citation: Said HK, Hijjawi J, Roy N, Mogford J, Mustoe T. Transdermal sustained-delivery oxygen improves epithelial healing in a rabbit ear wound model. Arch Surg. 2005;140:998–1004
references.citation: Woo KY, Coutts PM, Sibbald RG. Continuous topical oxygen for the treatment of chronic wounds: A pilot study. Adv Skin Wound Care. 2012;25:543–547
references.citation: Davis SC, Cazzaniga AL, Ricotti C, et al. Topical oxygen emulsion: A novel wound therapy. Arch Dermatol. 2007;143:1252–1256
references.citation: Plock JA, Tromp AE, Contaldo C, et al. Hemoglobin vesicles reduce hypoxia-related inflammation in critically ischemic hamster flap tissue. Crit Care Med. 2007;35:899–905
references.citation: Plock JA, Rafatmehr N, Sinovcic D, et al. Hemoglobin vesicles improve wound healing and tissue survival in critically ischemic skin in mice. Am J Physiol Heart Circ Physiol. 2009;297:H905–H910
references.citation: Steinberg JP, Hong SJ, Geringer MR, Galiano RD, Mustoe TA. Equivalent effects of topically-delivered adipose-derived stem cells and dermal fibroblasts in the ischemic rabbit ear model for chronic wounds. Aesthet Surg J. 2012;32:504–519
references.citation: Lokmic Z, Musyoka J, Hewitson TD, Darby IA. Hypoxia and hypoxia signaling in tissue repair and fibrosis. Int Rev Cell Mol Biol. 2012;296:139–185
references.citation: Mustoe TA, Pierce GF, Morishima C, Deuel TF. Growth factor-induced acceleration of tissue repair through direct and inductive activities in a rabbit dermal ulcer model. J Clin Invest. 1991;87:694–703
references.citation: Zhao LL, Davidson JD, Wee SC, Roth SI, Mustoe TA. Effect of hyperbaric oxygen and growth factors on rabbit ear ischemic ulcers. Arch Surg. 1994;129:1043–1049
references.citation: Deshmukh R, Trivedi V. Methemoglobin exposure produces toxicological effects in macrophages due to multiple ROS spike induced apoptosis. Toxicol In Vitro. 2013;27:16–23
references.citation: Tonnesen MG, Feng X, Clark RA. Angiogenesis in wound healing. J Investig Dermatol Symp Proc. 2000;5:40–46
references.citation: Gordillo GM, Roy S, Khanna S, et al. Topical oxygen therapy induces vascular endothelial growth factor expression and improves closure of clinically presented chronic wounds. Clin Exp Pharmacol Physiol. 2008;35:957–964
references.citation: Fukumura D, Gohongi T, Kadambi A, et al. Predominant role of endothelial nitric oxide synthase in vascular endothelial growth factor-induced angiogenesis and vascular permeability. Proc Natl Acad Sci USA. 2001;98:2604–2609
references.citation: Zhang Q, Gould LJ. Hyperbaric oxygen reduces matrix metalloproteinases in ischemic wounds through a redox-dependent mechanism. J Invest Dermatol. 2014;134:237–246
references.citation: Siddiqui A, Galiano RD, Connors D, Gruskin E, Wu L, Mustoe TA. Differential effects of oxygen on human dermal fibroblasts: Acute versus chronic hypoxia. Wound Repair Regen. 1996;4:211–218
references.citation: Biswas S, Roy S, Banerjee J, et al. Hypoxia inducible microRNA 210 attenuates keratinocyte proliferation and impairs closure in a murine model of ischemic wounds. Proc Natl Acad Sci USA. 2010;107:6976–6981
references.citation: Brismar K, Lind F, Kratz G. Dose-dependent hyperbaric oxygen stimulation of human fibroblast proliferation. Wound Repair Regen. 1997;5:147–150
references.citation: Squires JE. Artificial blood. Science. 2002;295:1002–1005