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Hydrogen-rich saline attenuates skin ischemia/reperfusion induced apoptosis via regulating Bax/Bcl-2 ratio and ASK-1/JNK pathway

Published:March 21, 2015DOI:https://doi.org/10.1016/j.bjps.2015.03.001

      Summary

      Introduction

      Many pathways have been reported involving the effect of hydrogen-rich saline on protecting skin flap partial necrosis induced by the inflammation of ischemia/reperfusion injury. This study focused on the influence of hydrogen-rich saline treatment on apoptosis pathway of ASK-1/JNK and Bcl-2/Bax radio in I/R injury of skin flaps.

      Methods

      Adult male Sprague–Dawley rats were divided into three groups. Group 1 was sham surgery group, Group 2 and 3 were ischemia/reperfusion surgery treated with physiological saline and hydrogen-rich saline respectively. Blood perfusion of flap was measured by Laser doppler flowmeters. Hematoxylin and eosin staining was used to observe morphological changes. Early apoptosis in skin flap was observed through TUNEL staining and presented as the percentage of TUNEL-positive cells of total cells. pASK-1, pJNK, Bcl-2 and Bax were examined by immunodetection. In addition Bcl-2, Bax and caspase-3 were detected by qPCR. Caspase-3 activity was also measured.

      Results

      Compared to the Group 2, tissues from the group 3 were observed with a high expression of Bcl-2 and a low expression of pASK-1, pJNK, and Bax, a larger survival area and a high level of blood perfusion. Hydrogen-rich saline ameliorated inflammatory infiltration and decreased cell apoptosis.

      Conclusion

      The results indicate that hydrogen-rich saline could ameliorate ischemia/reperfusion injury and improve flap survival rate by inhibiting the apoptosis factor and, at the same time, promoting the expression of anti-apoptosis factor.

      Keywords

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      References

        • Harder Y.
        • Amon M.
        • Laschke M.W.
        • et al.
        An old dream revitalised: preconditioning strategies to protect surgical flaps from critical ischaemia and ischaemia-reperfusion injury.
        J Plast Reconstr Aesthet Surg. 2008; 61: 503-511
        • Schoenberg M.H.
        • Beger H.G.
        Reperfusion injury after intestinal ischemia.
        Crit Care Med. 1993; 21: 1376-1386
        • Ozmen S.
        • Ayhan S.
        • Demir Y.
        • et al.
        Impact of gradual blood flow increase on ischaemia-reperfusion injury in the rat cremaster microcirculation model.
        J Plast Reconstr Aesthet Surg. 2008; 61: 939-948
        • Aydogan H.
        • Gurlek A.
        • Parlakpinar H.
        • et al.
        Beneficial effects of caffeic acid phenethyl ester (CAPE) on the ischaemia-reperfusion injury in rat skin flaps.
        J Plast Reconstr Aesthet Surg. 2007; 60: 563-568
        • Cetin C.
        • Kose A.A.
        • Aral E.
        • et al.
        Protective effect of fucoidin (a neutrophil rolling inhibitor) on ischemia reperfusion injury: experimental study in rat epigastric island flaps.
        Ann Plast Surg. 2001; 47: 540-546
        • Burns A.T.
        • Davies D.R.
        • McLaren A.J.
        • et al.
        Apoptosis in ischemia/reperfusion injury of human renal allografts.
        Transplantation. 1998; 66: 872-876
        • Ohsawa I.
        • Ishikawa M.
        • Takahashi K.
        • et al.
        Hydrogen acts as a therapeutic antioxidant by selectively reducing cytotoxic oxygen radicals.
        Nat Med. 2007; 13: 688-694
        • Fukuda K.
        • Asoh S.
        • Ishikawa M.
        • et al.
        Inhalation of hydrogen gas suppresses hepatic injury caused by ischemia/reperfusion through reducing oxidative stress.
        Biochem Res Commun. 2007; 361: 670-674
        • Hayashida K.
        • Sano M.
        • Ohsawa I.
        • et al.
        Inhalation of hydrogen gas reduces infarct size in the rat model of myocardial ischemia-reperfusion injury.
        Biochem Res Commun. 2008; 373: 30-35
        • Buchholz B.M.
        • Kaczorowski D.J.
        • Sugimoto R.
        • et al.
        Hydrogen inhalation ameliorates oxidative stress in transplantation induced intestinal graft injury.
        Am J Transpl. 2008; 8: 2015-2024
        • Cai J.
        • Kang Z.
        • Liu K.
        • et al.
        Neuroprotective effects of hydrogen saline in neonatal hypoxia-ischemia rat model.
        Brain Res. 2009; 1256: 129-137
        • Zhou L.
        • Wang X.
        • Xue W.
        • et al.
        Beneficial effects of hydrogen-rich saline against spinal cord ischemia-reperfusion injury in rabbits.
        Brain Res. 2013; 1517: 150-160
        • Jiang D.
        • Wu D.
        • Zhang Y.
        • et al.
        Protective effects of hydrogen rich saline solution on experimental testicular ischemia-reperfusion injury in rats.
        J Urol. 2012; 187: 2249-2253
        • Shi J.
        • Yao F.
        • Zhong C.
        • et al.
        Hydrogen saline is protective for acute lung ischaemia/reperfusion injuries in rats.
        Heart Lung Circ. 2012; 21: 556-563
        • Chen H.
        • Sun Y.P.
        • Hu P.F.
        • et al.
        The effects of hydrogen-rich saline on the contractile and structural changes of intestine induced by ischemia-reperfusion in rats.
        J Surg Res. 2011; 167: 316-322
        • Zhang Y.
        • Sun Q.
        • He B.
        • et al.
        Anti-inflammatory effect of hydrogen-rich saline in a rat model of regional myocardial ischemia and reperfusion.
        Int J Cardiol. 2011; 148: 91-95
        • Kuntscher M.V.
        • Schirmbeck E.U.
        • Menke H.
        • et al.
        Ischemic preconditioning by brief extremity ischemia before flap ischemia in a rat model.
        Plast Reconstr Surg. 2002; 109: 2398-2404
        • Dole M.
        • Wilson F.R.
        • Fife W.P.
        Hyperbaric hydrogen therapy: a possible treatment for cancer.
        Science. 1975; 190: 152-154
        • Gharib B.
        • Hanna S.
        • Abdallahi O.M.
        • et al.
        Anti-inflammatory properties of molecular hydrogen: investigation on parasite-induced liver inflammation.
        C R Acad Sci III. 2001; 324: 719-724
        • Liu Y.
        • Yang L.
        • Tao K.
        • et al.
        Protective effects of hydrogen enriched saline on liver ischemia reperfusion injury by reducing oxidative stress and HMGB1 release.
        BMC Gastroenterol. 2014; 14: 12
        • Ning Y.
        • Shang Y.
        • Huang H.
        • et al.
        Attenuation of cigarette smoke-induced airway mucus production by hydrogen-rich saline in rats.
        PLoS One. 2013; 8: e83429
        • Oharazawa H.
        • Igarashi T.
        • Yokota T.
        • et al.
        Protection of the retina by rapid diffusion of hydrogen: administration of hydrogen-loaded eye drops in retinal ischemia-reperfusion injury.
        Invest Ophthalmol Vis Sci. 2010; 51: 487-492
        • Chen H.
        • Xing B.
        • Liu X.
        • et al.
        Ozone oxidative preconditioning inhibits inflammation and apoptosis in a rat model of renal ischemia/reperfusion injury.
        Eur J Pharmacol. 2008; 581: 306-314
        • Guo C.
        • Liang F.
        • Shah M.W.
        • et al.
        Hydrogen sulfide protected gastric epithelial cell from ischemia/reperfusion injury by Keap1 s-sulfhydration, MAPK dependent anti-apoptosis and NF-kappaB dependent anti-inflammation pathway.
        Eur J Pharmacol. 2014; 725: 70-78
        • Saeki K.
        • Kobayashi N.
        • Inazawa Y.
        • et al.
        Oxidation-triggered c-Jun N-terminal kinase (JNK) and p38 mitogen-activated protein (MAP) kinase pathways for apoptosis in human leukaemic cells stimulated by epigallocatechin-3-gallate (EGCG): a distinct pathway from those of chemically induced and receptor-mediated apoptosis.
        Biochem J. 2002; 368: 705-720
        • Van Laethem A.
        • Nys K.
        • Van Kelst S.
        • et al.
        Apoptosis signal regulating kinase-1 connects reactive oxygen species to p38 MAPK-induced mitochondrial apoptosis in UVB-irradiated human keratinocytes.
        Free Radic Biol Med. 2006; 41: 1361-1371
        • Fujino G.
        • Noguchi T.
        • Matsuzawa A.
        • et al.
        Thioredoxin and TRAF family proteins regulate reactive oxygen species-dependent activation of ASK1 through reciprocal modulation of the N-terminal homophilic interaction of ASK1.
        Mol Cell Biol. 2007; 27: 8152-8163
        • Xue X.
        • Piao J.H.
        • Nakajima A.
        • et al.
        Tumor necrosis factor alpha (TNFalpha) induces the unfolded protein response (UPR) in a reactive oxygen species (ROS)-dependent fashion, and the UPR counteracts ROS accumulation by TNFalpha.
        J Biol Chem. 2005; 280: 33917-33925
        • Schulze-Osthoff K.
        • Bakker A.C.
        • Vanhaesebroeck B.
        • et al.
        Cytotoxic activity of tumor necrosis factor is mediated by early damage of mitochondrial functions. Evidence for the involvement of mitochondrial radical generation.
        J Biol Chem. 1992; 267: 5317-5323
        • Goossens V.
        • Grooten J.
        • De Vos K.
        • et al.
        Direct evidence for tumor necrosis factor-induced mitochondrial reactive oxygen intermediates and their involvement in cytotoxicity.
        Proc Natl Acad Sci U S A. 1995; 92: 8115-8119
        • Fernandez-Checa J.C.
        • Kaplowitz N.
        • Garcia-Ruiz C.
        • et al.
        GSH transport in mitochondria: defense against TNF-induced oxidative stress and alcohol-induced defect.
        Am J Physiol. 1997; 273: 7-17
        • Corda S.
        • Laplace C.
        • Vicaut E.
        • et al.
        Rapid reactive oxygen species production by mitochondria in endothelial cells exposed to tumor necrosis factor-alpha is mediated by ceramide.
        Am J Respir Cell Mol Biol. 2001; 24: 762-768
        • Ling Z.
        • You-bin W.
        • Shi-rui Q.
        • et al.
        Protective effect of hydrogen-rich saline on ischemia/reperfusion injury in rat skin flap.
        J Zhejiang Univ Sci B. 2013; 14: 382-391
        • Kroemer G.
        • Dallaporta B.
        • Resche-Rigon M.
        The mitochondrial death/life regulator in apoptosis and necrosis.
        Annu Rev Physiol. 1998; 60: 619-642
        • Sidoti-de F.C.
        • Rincheval V.
        • Risler Y.
        • et al.
        TNF-alpha activates at least two apoptotic signaling cascades.
        Oncogene. 1998; 17: 1639-1651
        • Fleury C.
        • Mignotte B.
        • Vayssiere J.L.
        Mitochondrial reactive oxygen species in cell death signaling.
        Biochimie. 2002; 84: 131-141
        • Belka C.
        • Budach W.
        Anti-apoptotic Bcl-2 proteins: structure, function and relevance for radiation biology.
        Int J Radiat Biol. 2002; 78: 643-658
        • Lee S.H.
        • Park S.W.
        • Pyo C.W.
        • et al.
        Requirement of the JNK-associated Bcl-2 pathway for human lactoferrin-induced apoptosis in the Jurkat leukemia T cell line.
        Biochimie. 2009; 91: 102-108
        • Qiu X.
        • Li H.
        • Tang H.
        • et al.
        Hydrogen inhalation ameliorates lipopolysaccharide-induced acute lung injury in mice.
        IntImmunopharmacol. 2011; 11: 2130-2137
        • Huang C.S.
        • Kawamura T.
        • Peng X.
        • et al.
        Hydrogen inhalation reduced epithelial apoptosis in ventilator-induced lung injury via a mechanism involving nuclear factor-kappa B activation.
        Biochem Res Commun. 2011; 408: 253-258
        • Lalier L.
        • Cartron P.F.
        • Juin P.
        • et al.
        Bax activation and mitochondrial insertion during apoptosis.
        Apoptosis. 2007; 12: 887-896