Abstract
Heat stress is exacerbated by global warming and affects human and animal health, leading to heart damage caused by imbalances in reactive oxygen species (ROS) and the antioxidant system, acid-base chemistry, electrolytes and respiratory alkalosis. Vitamin C scavenges excess ROS, and sodium bicarbonate maintains acid-base and electrolyte balance, and alleviates respiratory alkalosis. Herein, we explored the ability of vitamin C alone and in combination with equimolar sodium bicarbonate (Vitamin C-Na) to stimulate endogenous antioxidants and heat shock proteins (HSPs) to relieve heat stress in H9C2 cells. Control, vitamin C (20 μg/ml vitamin C for 16 h) and vitamin C-Na (20 μg/ml vitamin C-Na for 16 h) groups were heat-stressed for 1, 3 or 5 h. Granular and vacuolar degeneration, karyopyknosis and damage to nuclei and mitochondria were clearly reduced in treatment groups, as were apoptosis, lactate dehydrogenase activity and ROS and malondialdehyde levels, while superoxide dismutase activity was increased. Additionally, CRYAB, Hsp27, Hsp60 and Hsp70 mRNA levels were upregulated at 3 h (p < 0.01), and protein levels were increased for CRYAB at 0 h (p < 0.05) and 1 h (p < 0.01), and for Hsp70 at 3 and 5 h (p < 0.01). Thus, pre-treatment with vitamin C or vitamin C-Na might protect H9C2 cells against heat damage by enhancing the antioxidant ability and upregulating CRYAB and Hsp70.
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References
Aditi A, Graham DY (2012) Vitamin C, gastritis, and gastric disease: a historical review and update. Dig Dis Sci 57(10):2504–2515. https://doi.org/10.1007/s10620-012-2203-7
Ahmad G, Agarwal A, Esteves SC, Sharma R, Almasry M, Al-Gonaim A, Alhayaza G, Singh N, Al Kattan L, Sannaa WM, Sabanegh E (2017) Ascorbic acid reduces redox potential in human spermatozoa subjected to heat-induced oxidative stress. Andrologia e12773:1–8. https://doi.org/10.1111/and.12773
Ahmad T, Mushtaq T, Khan MA, Babar ME, Yousaf M, Hasan ZU, Kamran Z (2009) Influence of varying dietary electrolyte balance on broiler performance under tropical summer conditions. J Anim Physiol Anim Nutr 93(5):613–621. https://doi.org/10.1111/j.1439-0396.2008.00840.x
Aitken-Buck HM, Lamberts RR (2017) To the heart of activation heat. J Physiol 595(14):4577–4578. https://doi.org/10.1113/JP274582
Attia YA, Abd El-Hamid Ael H, Abedalla AA, Berika MA, Al-Harthi MA, Kucuk O, Sahin K, Abou-Shehema BM (2016) Laying performance, digestibility and plasma hormones in laying hens exposed to chronic heat stress as affected by betaine, vitamin C, and/or vitamin E supplementation. SpringerPlus 5(1):1619–1631. https://doi.org/10.1186/s40064-016-3304-0
Bae ON, Lim KM, Han JY, Jung BI, Lee JY, Noh JY, Chung SM, Lee MY, Lee JY, Chung JH (2008) U-shaped dose response in vasomotor tone: a mixed result of heterogenic response of multiple cells to xenobiotics. Toxicological sciences : an Official Journal of the Society of Toxicology 103(1):181–190. https://doi.org/10.1093/toxsci/kfn023
Bakthisaran R, Tangirala R, Rao Ch M (2015) Small heat shock proteins: role in cellular functions and pathology. Biochim Biophys Acta 1854(4):291–319. https://doi.org/10.1016/j.bbapap.2014.12.019
Camins C, Diezfernandez C, Prieo P (1999) Cell-surface expression of heat shock proteins in dog neutrophils after oxidative stress. Toxicilogy in Vitro 13(3):437–443. https://doi.org/10.1016/S0887-2333(99)00012-0
Chand N, Naz S, Khan A, Khan S, Khan RU (2014) Performance traits and immune response of broiler chicks treated with zinc and ascorbic acid supplementation during cyclic heat stress. Int J Biometeorol 58(10):2153–2157. https://doi.org/10.1007/s00484-014-0815-7
Chrisostomos P, Barry P, Shahzad C, Giuliano S, Ronan O, John EL, Smark R, Peter WP, Laurence HP (2000) The ATPase cycle of Hsp90 drives a molecular 'clamp' via transient dimerization of the N-terminal domains. EMBO J 19:4383–4392
Collier RJ, Collier JL, Rhoads RP, Baumgard LH (2008) Invited review: genes involved in the bovine heat stress response. J Dairy Sci 91(2):445–454. https://doi.org/10.3168/jds.2007-0540
Crandall, C.G., Wilson, T.E., 2014. Human cardiovascular responses to passive heat stress. 17-43
Cui J, Sinoway LI (2014) Cardiovascular responses to heat stress in chronic heart failure. Current Heart Failure Reports 11(2):139–145. https://doi.org/10.1007/s11897-014-0191-y
Cumming KT, Raastad T, Holden G, Bastani NE, Schneeberger D, Paronetto MP, Mercatelli N, Ostgaard HN, Ugelstad I, Caporossi D, Blomhoff R, Paulsen G (2014) Effects of vitamin C and E supplementation on endogenous antioxidant systems and heat shock proteins in response to endurance training. Physiological Reports 2:1–14
Dangi SS, Gupta M, Dangi SK, Chouhan VS, Maurya VP, Kumar P, Singh G, Sarkar M (2015) Expression of HSPs: an adaptive mechanism during long-term heat stress in goats (Capra Hircus). Int J Biometeorol 59(8):1095–1106. https://doi.org/10.1007/s00484-014-0922-5
Dokladny K, Moseley PL, Ma TY (2006) Physiologically relevant increase in temperature causes an increase in intestinal epithelial tight junction permeability. Am J Physiol Gastrointest Liver Physiol 290(2):G204–G212. https://doi.org/10.1152/ajpgi.00401.2005
Ferreira IB, Matos Junior JB, Sgavioli S, Vicentini TI, Morita VS, Boleli IC (2015) Vitamin C prevents the effects of high rearing temperatures on the quality of broiler thigh meat1. Poult Sci 94(5):841–851. https://doi.org/10.3382/ps/pev058
Garrido C, Gurbuxani S, Ravagnan L, Kroemer G (2001) Heat shock proteins: endogenous modulators of apoptotic cell death. Biochem Biophys Res Commun 286(3):433–442. https://doi.org/10.1006/bbrc.2001.5427
Grizzle J, Iheanacho M, Saxton A, Broaden J (1992) Nutritional and environmental factors involved in egg shell quality of laying hens. Br Poult Sci 33(4):781–794. https://doi.org/10.1080/00071669208417520
Imik H, Ozlu H, Gumus R, Atasever MA, Urcar S, Atasever M (2012) Effects of ascorbic acid and alpha-lipoic acid on performance and meat quality of broilers subjected to heat stress. Br Poult Sci 53(6):800–808. https://doi.org/10.1080/00071668.2012.740615
Jaiswal SK, Gupta VK, Ansari MD, Siddiqi NJ, Sharma B (2017) Vitamin C acts as a hepatoprotectant in carbofuran treated rat liver slices in vitro. Toxicol Rep 4:265–273. https://doi.org/10.1016/j.toxrep.2017.06.001
Jang IS, Ko YH, Moon YS, Sohn SH (2014) Effects of vitamin C or E on the pro-inflammatory cytokines, heat shock protein 70 and antioxidant status in broiler chicks under summer conditions. Asian Australas J Anim Sci 27(5):749–756. https://doi.org/10.5713/ajas.2013.13852
Kazim S, Osman K, Nurhan S, Mustafa S (2002) Effects of vitamin C and vitamin E on lipid peroxidation status. Serum Hormone, Metabolite, and Mineral Concentrations of Japanese Quails Reared under Heat Stress Int J Vitam 72:91–100
Khassaf M, Mcardle A, Esanu C, Vasilaki A, Mcardle F, Griffiths RD, Brodie DA, Jackson MJ (2003) Effect of vitamin C supplements on antioxidant defence and stress proteins in human lymphocytes and skeletal muscle. J Physiol 549(2):645–652. https://doi.org/10.1113/jphysiol.2003.040303
Kishore A, Sodhi M, Kumari P, Mohanty AK, Sadana DK, Kapila N, Khate K, Shandilya U, Kataria RS, Mukesh M (2014) Peripheral blood mononuclear cells: a potential cellular system to understand differential heat shock response across native cattle (Bos indicus), exotic cattle (Bos taurus), and riverine buffaloes (Bubalus bubalis) of India. Cell Stress Chaperones 19(5):613–621. https://doi.org/10.1007/s12192-013-0486-z
Levine M, Padayatty SJ, Espey MG (2011) Vitamin C: a concentration-function approach yields pharmacology and therapeutic discoveries. Adv Nutr 2(2):78–88. https://doi.org/10.3945/an.110.000109
Lin H, Decuypere E, Buyse J (2006) Acute heat stress induces oxidative stress in broiler chickens. Comp Biochem Physiol A Mol Integr Physiol 144(1):11–17. https://doi.org/10.1016/j.cbpa.2006.01.032
Liu F, Cottrell JJ, Furness JB, Rivera LR, Kelly FW, Wijesiriwardana U, Pustovit RV, Fothergill LJ, Bravo DM, Celi P, Leury BJ, Gabler NK, Dunshea FR (2016) Selenium and vitamin E together improve intestinal epithelial barrier function and alleviate oxidative stress in heat-stressed pigs. Exp Physiol 101(7):801–810. https://doi.org/10.1113/EP085746
Mujahid A (2011) Nutritional strategies to maintain efficiency and production of chickens under high environmental temperature. J Poult Sci 48(3):145–154. https://doi.org/10.2141/jpsa.010115
Mahmoud KZ, Edens FW, Eisen EJ, Havenstein GB (2003) Effect of ascorbic acid and acute heat exposure on heat shock protein 70 expression by young white leghorn chickens. Comparative biochemistry and physiology Toxicology & pharmacology : CBP 136(4):329–335. https://doi.org/10.1016/j.cca.2003.10.006
Mahmoud KZ, Edens FW, Eisen EJ, Havenstein GB (2004) Ascorbic acid decreases heat shock protein 70 and plasma corticosterone response in broilers (Gallus gallus domesticus) subjected to cyclic heat stress. Comp Biochem Physiol B: Biochem Mol Biol 137(1):35–42. https://doi.org/10.1016/j.cbpc.2003.09.013
Malcolm JJ, Sergio P, Juan B, Richard B, Harald C, Ruan ME, Jacoba F, Helen RG (2002) Antioxidants, reactive oxygen and nitrogen species, gene induction and mitochondrial function. Mol Asp Med 23:209–285
Matsumura T, Matsumoto H, Hayakawa Y (2017) Heat stress hardening of oriental armyworms is induced by a transient elevation of reactive oxygen species during sublethal stress. Arch Insect Biochem Physiol 96:1–10
Morrison D, Hughes J, Della Gatta PA, Mason S, Lamon S, Russell AP, Wadley GD (2015) Vitamin C and E supplementation prevents some of the cellular adaptations to endurance-training in humans. Free Radic Biol Med 89:852–862. https://doi.org/10.1016/j.freeradbiomed.2015.10.412
Niu ZY, Liu FZ, Yan QL, Li WC (2009) Effects of different levels of vitamin E on growth performance and immune responses of broilers under heat stress. Poult Sci 88(10):2101–2107. https://doi.org/10.3382/ps.2009-00220
Padayatty SJ, Levine M (2016) Vitamin C: the known and the unknown and goldilocks. Oral Dis 22(6):463–493. https://doi.org/10.1111/odi.12446
Panda AK, Ramarao SV, Raju MV, Chatterjee RN (2008) Effect of dietary supplementation with vitamins E and C on production performance, immune responses and antioxidant status of white leghorn layers under tropical summer conditions. Br Poult Sci 49(5):592–599. https://doi.org/10.1080/00071660802337233
Rafiee F, Mazhari M, Ghoreishi M, Esmaeilipour O (2016) Effect of lemon verbena powder and vitamin C on performance and immunity of heat-stressed broilers. J Anim Physiol Anim Nutr 100(5):807–812. https://doi.org/10.1111/jpn.12457
Redmond SB, Tell RM, Coble D, Mueller C, Palic D, Andreasen CB, Lamont SJ (2010) Differential splenic cytokine responses to dietary immune modulation by diverse chicken lines. Poult Sci 89(8):1635–1641. https://doi.org/10.3382/ps.2010-00846
Ruell PA, Thompson MW, Hoffman KM (2009) Heat shock proteins as an aid in the treatment and diagnosis of heat stroke. J Therm Biol 34(1):1–7. https://doi.org/10.1016/j.jtherbio.2008.09.004
Sahin N, Onderci M, Sahin K, Gursu MF, Smith MO (2004) Ascorbic acid and melatonin reduce heat-induced performance inhibition and oxidative stress in Japanese quails. Br Poult Sci 45(1):116–122. https://doi.org/10.1080/00071660410001668941
Sahin N, Tuzcu M, Orhan C, Onderci M, Eroksuz Y, Sahin K (2009) The effects of vitamin C and E supplementation on heat shock protein 70 response of ovary and brain in heat-stressed quail. Br Poult Sci 50(2):259–265. https://doi.org/10.1080/00071660902758981
Sharma S, Ramesh K, Hyder I, Uniyal S, Yadav VP, Panda RP, Maurya VP, Singh G, Kumar P, Mitra A, Sarkar M (2013) Effect of melatonin administration on thyroid hormones, cortisol and expression profile of heat shock proteins in goats (Capra hircus) exposed to heat stress. Small Rumin Res 112(1-3):216–223. https://doi.org/10.1016/j.smallrumres.2012.12.008
Shashidharamurthy R, Koteiche HA, Dong J, Mchaourab HS (2005) Mechanism of chaperone function in small heat shock proteins: dissociation of the HSP27 oligomer is required for recognition and binding of destabilized T4 lysozyme. J Biol Chem 280(7):5281–5289. https://doi.org/10.1074/jbc.M407236200
Sinkalu VO, Ayo JO (2016) Combined effects of retinol, ascorbic acid and alpha-tocopherol on diurnal variations in rectal temperature of Black Harco pullets subjected to heat stress. Int J Biometeorol. https://doi.org/10.1007/s00484-00016-01157-00484
Song E, Tang S, Xu J, Yin B, Bao E, Hartung J (2016) Lenti-siRNA Hsp60 promote bax in mitochondria and induces apoptosis during heat stress. Biochem Biophys Res Commun 481(1-2):125–131. https://doi.org/10.1016/j.bbrc.2016.10.153
Sottile ML, Nadin SB (2017) Heat shock proteins and DNA repair mechanisms: an updated overview. Cell Stress Chaperones. https://doi.org/10.1007/s00484-00016-01157-00484
Szczubial M (2015) Effect of supplementation with vitamins E, C and beta-carotene on antioxidative/oxidative status parameters in sows during the postpartum period. Pol J Vet Sci 18(2):299–305. https://doi.org/10.1515/pjvs-2015-0039
Tan GY, Yang L, Fu YQ, Feng JH, Zhang MH (2010) Effects of different acute high ambient temperatures on function of hepatic mitochondrial respiration, antioxidative enzymes, and oxidative injury in broiler chickens. Poult Sci 89(1):115–122. https://doi.org/10.3382/ps.2009-00318
Tang S, Chen H, Cheng Y, Nasir MA, Kemper N, Bao E (2016a) Expression profiles of heat shock protein 27 and alphaB-crystallin and their effects on heat-stressed rat myocardial cells in vitro and in vivo. Mol Med Rep 13(2):1633–1638. https://doi.org/10.3892/mmr.2015.4693
Tang S, Yin B, Song E, Chen H, Cheng Y, Zhang X, Bao E, Hartung J (2016b) Aspirin upregulates alphaB-crystallin to protect the myocardium against heat stress in broiler chickens. Sci Rep 6(1):37273. https://doi.org/10.1038/srep37273
Torki M, Zangeneh S, Habibian M (2014) Performance, egg quality traits, and serum metabolite concentrations of laying hens affected by dietary supplemental chromium picolinate and vitamin C under a heat-stress condition. Biol Trace Elem Res 157(2):120–129. https://doi.org/10.1007/s12011-013-9872-8
Tower J (2011) Heat shock proteins and Drosophila aging. Exp Gerontol 46(5):355–362. https://doi.org/10.1016/j.exger.2010.09.002
Wang M, Sun GB, Du YY, Tian Y, Liao P, Liu XS, Ye JX, Sun XB (2017) Myricitrin protects cardiomyocytes from hypoxia/reoxygenation injury: involvement of heat shock protein 90. Front Pharmacol 8:353. https://doi.org/10.3389/fphar.2017.00353
Westman J, Drieu K, Sharma H (2000) Antioxidant compounds EGB-761 and BN-52021 attenuate heat shock protein (HSP72 kD) response, edema and cell changes following hyperthermic brain injury. Amino Acids 19(1):339–350. https://doi.org/10.1007/s007260070065
Xu J, Tang S, Song E, Yin B, Bao E (2017a) Inhibition of heat shock protein 70 intensifies heat-stressed damage and apoptosis of chicken primary myocardial cells in vitro. Mol Med Rep 15(5):2881–2889. https://doi.org/10.3892/mmr.2017.6337
Xu J, Tang S, Song E, Yin B, Wu D, Bao E (2017b) Hsp70 expression induced by co-enzyme Q10 protected chicken myocardial cells from damage and apoptosis under in vitro heat stress. Poult Sci 96(5):1426–1437. https://doi.org/10.3382/ps/pew402
Xu J, Tang S, Yin B, Sun J, Song E, Bao E (2017c) Co-enzyme Q10 and acetyl salicylic acid enhance Hsp70 expression in primary chicken myocardial cells to protect the cells during heat stress. Mol Cell Biochem 435(1-2):73–86. https://doi.org/10.1007/s11010-11017-13058-11011
Yörük MA, Gül M, Hayirli A, Laçin E (2004) Laying performance and egg quality of hens supplemented with Humate and sodium bicarbonate during the late laying period. J Appl Anim Res 26(1):17–21. https://doi.org/10.1080/09712119.2004.9706498
Yamagishi N, Ishihara K, Saito Y, Hatayama T (2003) Hsp105 but not Hsp70 family proteins suppress the aggregation of heat-denatured protein in the presence of ADP. FEBS Lett 555(2):390–396. https://doi.org/10.1016/S0014-5793(03)01292-4
Yuan Y, Pan S, Yang S-L, Liu Y-L, Xu Q-M (2017) Antioxidant and cardioprotective effects of Ilex cornuta on myocardial ischemia injury. Chin J Nat Med 15(2):94–104. https://doi.org/10.1016/S1875-5364(17)30025-0
Zanini C, Giribaldi G, Mandili G, Carta F, Crescenzio N, Bisaro B, Doria A, Foglia L, Di Montezemolo LC, Timeus F, Turrini F (2007) Inhibition of heat shock proteins (HSP) expression by quercetin and differential doxorubicin sensitization in neuroblastoma and Ewing's sarcoma cell lines. J Neurochem 103(4):1344–1354. https://doi.org/10.1111/j.1471-4159.2007.04835.x
Zhou H, Malik MA, Arab A, Hill MT, Shikanov A (2015) Hydrogel based 3-dimensional (3D) system for toxicity and high-throughput (HTP) analysis for cultured murine ovarian follicles. PLoS One 10(10):e0140205. https://doi.org/10.1371/journal.pone.0140205
Zulkifli I, Al-Aqil A, Omar AR, Sazili AQ, Rajion MA (2009) Crating and heat stress influence blood parameters and heat shock protein 70 expression in broiler chickens showing short or long tonic immobility reactions. Poult Sci 88(3):471–476. https://doi.org/10.3382/ps.2008-00287
Acknowledgements
The current study was supported by grants from the National Natural Science Foundation of China (grant no. 31672520), the Fundamental Research Funds for the Central Universities (grant no. KJQN201709), the National Natural Science Foundation of China (grant no. 31602027), the National Natural Science Foundation of China (grant no. 31372403), Jiangsu Natural Science Foundation of China (grant no. BK20160732), China Postdoctoral Science Foundation (2016M591860) and the Priority Academic Program Development of Jiangsu Higher Education Institutions, Graduate Research and Innovation Projects in Jiangsu Province.
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Yin, B., Tang, S., Sun, J. et al. Vitamin C and sodium bicarbonate enhance the antioxidant ability of H9C2 cells and induce HSPs to relieve heat stress. Cell Stress and Chaperones 23, 735–748 (2018). https://doi.org/10.1007/s12192-018-0885-2
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DOI: https://doi.org/10.1007/s12192-018-0885-2