Ethanol exposure in prenatal and early postnatal induced cardiac injury in rats: involvement of oxidative stress, Hsp70, ERK 1/2, JNK, and apoptosis in a 3-month follow-up study

  • Alireza Shirpoor
  • Reza Gaderi
  • Roya NaderiEmail author
Original Paper


Alcohol exposure during pregnancy induces a wide range of structural and functional abnormalities in the fetal heart. However, the underlying mechanism of this phenomenon is not well known. This study was undertaken to elucidate probable mechanisms of myocardial damage induced by prenatal and early postnatal ethanol treatment. Pregnant Wistar rats received ethanol 4.5 g/kg BW once per day from the seventh day of gestation (GD7) throughout lactation. The oxidative stress injury of the myocardium in pups was evaluated by measuring levels of oxidative stress biomarkers. Histopathological examinations and Western blot were performed to evaluate histological features, apoptosis, and molecular alterations in the myocardial tissue of male pups on the postnatal day 21 (PN-21) and postnatal day 90 (PN-90). The results showed that maternal ethanol consumption caused oxidative stress (impaired total antioxidant capacity and malondialdehyde), histological changes, and apoptosis of the myocardium in the pups on PN-21 and PN-90. At the molecular levels, Western blot analysis revealed that ethanol modulated the protein expression of p-ERK1/2, p-JNK, and Hsp70 in the myocardial tissue of the pups after 21 and 90 days of birth compared with the controls. These findings revealed that maternal ethanol intake induced cardiac toxicity in part, mediated by oxidative stress and apoptosis in the pups. A further mechanism study revealed that ethanol enhanced ERK1/2 and JNK phosphorylation and Hsp70 protein expression.


Ethanol Offspring ERK1/2 JNK Hsp70 Heart 


Funding information

The work was supported by a grant from the Urmia University of Medical Science, Urmia, Iran.

Compliance with ethical standards

All animal procedures were performed in accordance with the National Institutes of Health Guidelines for the Care and Use of Laboratory Animals and approved by the Animal Care Committee, the Urmia University of Medical Sciences.

Conflict of interest

The authors declare that they have no conflict of interest.


  1. Adickes ED, Mollner TJ, Lockwood SK (1990) Ethanol induced morphologic alterations during growth and maturation of cardiac myocytes. Alcohol Clin Exp Res 14:827–831CrossRefGoogle Scholar
  2. Aroor AR, Shukla SD (2004) MAP kinase signaling in diverse effects of ethanol. Life Sci 74:2339–2364CrossRefGoogle Scholar
  3. Cesconetto PA, Andrade CM, Cattani D, Domingues JT, Parisotto EB, Filho DW, Zamoner A (2016) Maternal exposure to ethanol during pregnancy and lactation affects glutamatergic system and induces oxidative stress in offspring hippocampus. Alcohol Clin Exp Res 40:52–61. CrossRefGoogle Scholar
  4. Chong KY, Lai CC, Lille S, Chang C, Su CY (1998) Stable overexpression of the constitutive form of heat shock protein 70 confers oxidative protection. J Mol Cell Cardiol 30:599–608. CrossRefGoogle Scholar
  5. Coll TA, Chaufan G, Perez-Tito L, Ventureira MR, Sobarzo CMA, Rios de Molina MDC, Cebral E (2017) Oxidative stress and cellular and tissue damage in organogenic outbred mouse embryos after moderate perigestational alcohol intake. Mol Reprod Dev 84:1086–1099. CrossRefGoogle Scholar
  6. Cui SZ, Wang SJ, Li J, Xie GQ, Zhou R, Chen L, Yuan XR (2011) Alteration of synaptic plasticity in rat dorsal striatum induced by chronic ethanol intake and withdrawal via ERK pathway. Acta Pharmacol Sin 32:175–181. CrossRefGoogle Scholar
  7. Daft PA, Johnston MC, Sulik KK (1986) Abnormal heart and great vessel development following acute ethanol exposure in mice. Teratology 33:93–104. CrossRefGoogle Scholar
  8. Denny L, Coles S, Blitz R (2017) Fetal alcohol syndrome and fetal alcohol spectrum disorders. Am Fam Physician 96:515–522Google Scholar
  9. El-Mas MM, Abdel-Rahman AA (2015) Estrogen modulation of the ethanol-evoked myocardial oxidative stress and dysfunction via DAPK3/Akt/ERK activation in male rats. Toxicol Appl Pharmacol 287:284–292. CrossRefGoogle Scholar
  10. Gallo S, Sala V, Gatti S, Crepaldi T (2015) Cellular and molecular mechanisms of HGF/Met in the cardiovascular system. Clin Sci (Lond) 129:1173–1193. CrossRefGoogle Scholar
  11. Gao G, Jiang S, Ge L, Zhang S, Zhai C, Chen W, Sui S (2018) Atorvastatin improves doxorubicin-induced cardiac dysfunction by modulating Hsp70, Akt and MAPK signalling pathways. J Cardiovasc Pharmacol 73:223–231. CrossRefGoogle Scholar
  12. Goh JM, Bensley JG, Kenna K, Sozo F, Bocking AD, Brien J, Walker D, Harding R, Black MJ (2011) Alcohol exposure during late gestation adversely affects myocardial development with implications for postnatal cardiac function. Am J Physiol Heart Circ Physiol 300:H645–H651. CrossRefGoogle Scholar
  13. Jones KL, Smith DW (1973) Recognition of the fetal alcohol syndrome in early infancy. Lancet 302:999–1001CrossRefGoogle Scholar
  14. Junior MDF, Cavalcante KVN, Ferreira LA, Lopes PR, Pontes CNR, Bessa ASM, Neves AR, Francisco FA, Pedrino GR, Xavier CH, Mathias PCF, Castro CH, Gomes RM (2019) Postnatal early overfeeding induces cardiovascular dysfunction by oxidative stress in adult male Wistar rats. Life Sci 226:173–184. CrossRefGoogle Scholar
  15. Kalluri HS, Ticku MK (2002) Ethanol-mediated inhibition of mitogen-activated protein kinase phosphorylation in mouse brain. Eur J Pharmacol 439:53–58CrossRefGoogle Scholar
  16. Liu X, Zhang C, Zhang C, Li J, Guo W, Yan D, Yang C, Zhao J, Xia T, Wang Y, Xu R, Wu X, Shi J (2016) Heat shock protein 70 inhibits cardiomyocyte necroptosis through repressing autophagy in myocardial ischemia/reperfusion injury. In Vitro Cell Dev Biol Anim 52:690–698. CrossRefGoogle Scholar
  17. Lowry OH, Rosebrough NJ, Farr AL, Randall RJ (1951) Protein measurement with the Folin phenol reagent. J Biol Chem 193:265–275Google Scholar
  18. Lu Z, Xu S (2006) ERK1/2 MAP kinases in cell survival and apoptosis. IUBMB Life 58:621–631. CrossRefGoogle Scholar
  19. Mestril R, Chi SH, Sayen MR, O'Reilly K, Dillmann WH (1994) Expression of inducible stress protein 70 in rat heart myogenic cells confers protection against simulated ischemia-induced injury. J Clin Invest 93:759–767. CrossRefGoogle Scholar
  20. Mohaddes G, Abdolalizadeh J, Babri S, Hossienzadeh F (2017) Ghrelin ameliorates blood-brain barrier disruption during systemic hypoxia. Exp Physiol 102:376–382. CrossRefGoogle Scholar
  21. Muniz JJ, Leite LN, Lacchini R, Tanus-Santos JE, Tirapelli CR (2018) Dysregulated mitogen-activated protein kinase and matrix metalloproteinase in ethanol-induced cavernosal dysfunction. Can J Physiol Pharmacol 96:266–274. CrossRefGoogle Scholar
  22. Mustroph J, Lebek S, Maier LS, Neef S (2018) Mechanisms of cardiac ethanol toxicity and novel treatment options. Pharmacol Ther 197:1–10. CrossRefGoogle Scholar
  23. Niehaus WG Jr, Samuelsson B (1968) Formation of malonaldehyde from phospholipid arachidonate during microsomal lipid peroxidation. Eur J Biochem 6:126–130CrossRefGoogle Scholar
  24. Nogales F, Ojeda ML, Jotty K, Murillo ML, Carreras O (2017) Maternal ethanol consumption reduces Se antioxidant function in placenta and liver of embryos and breastfeeding pups. Life Sci 190:1–6. CrossRefGoogle Scholar
  25. Portbury AL, Ronnebaum SM, Zungu M, Patterson C, Willis MS (2012) Back to your heart: ubiquitin proteasome system-regulated signal transduction. J Mol Cell Cardiol 52:526–537. CrossRefGoogle Scholar
  26. Ren J, Wold LE, Natavio M, Ren BH, Hannigan JH, Brown RA (2002) Influence of prenatal alcohol exposure on myocardial contractile function in adult rat hearts: role of intracellular calcium and apoptosis. Alcohol Alcohol 37:30–37. CrossRefGoogle Scholar
  27. Ronchi R, Marano L, Braidotti P, Bianciardi P, Calamia M, Fiorentini C, Samaja M (2004) Effects of broad band electromagnetic fields on HSP70 expression and ischemia-reperfusion in rat hearts. Life Sci 75:1925–1936. CrossRefGoogle Scholar
  28. Rose BA, Force T, Wang Y (2010) Mitogen-activated protein kinase signaling in the heart: angels versus demons in a heart-breaking tale. Physiol Rev 90:1507–1546. CrossRefGoogle Scholar
  29. Sadeghzadeh M, Shirpoor A, Khalaji N, Naderi R, Samadi M, Rasmi Y (2018) The effect of chronic ethanol consumption on sexual motivation and behavior of adult male Wistar rats in the copulatory phase. Addict Health 10:190–197. Google Scholar
  30. Sadeghzadeh M, Shirpoor A, Naderi R, Kheradmand F, Gharalari FH, Samadi M, Khalaji N, Gharaaghaji R (2019) Long-term ethanol consumption promotes changes in beta-defensin isoform gene expression and induces structural changes and oxidative DNA damage to the epididymis of rats. Mol Reprod Dev.
  31. Sanna PP, Simpson C, Lutjens R, Koob G (2002) ERK regulation in chronic ethanol exposure and withdrawal. Brain Res 948:186–191CrossRefGoogle Scholar
  32. Sato M, Maulik N, Das DK (2002) Cardioprotection with alcohol: role of both alcohol and polyphenolic antioxidants. Ann N Y Acad Sci 957:122–135CrossRefGoogle Scholar
  33. Seleverstov O, Tobiasz A, Jackson JS, Sullivan R, Ma D, Sullivan JP, Davison S, Akkhawattanangkul Y, Tate DL, Costello T, Barnett S, Li W, Mari G, Dopico AM, Bukiya AN (2017) Maternal alcohol exposure during mid-pregnancy dilates fetal cerebral arteries via endocannabinoid receptors. Alcohol 61:51–61. CrossRefGoogle Scholar
  34. Shen HM, Liu ZG (2006) JNK signaling pathway is a key modulator in cell death mediated by reactive oxygen and nitrogen species. Free Radic Biol Med 40:928–939. CrossRefGoogle Scholar
  35. Shirpoor A, Norouzi L, Khadem-Ansari MH, Ilkhanizadeh B, Karimipour M (2014) The protective effect of vitamin E on morphological and biochemical alteration induced by pre and postnatal ethanol administration in the testis of male rat offspring: a three months follow-up study. J Reprod Infertil 15:134–141Google Scholar
  36. Shirpoor A, Nemati S, Ansari MH, Ilkhanizadeh B (2015) The protective effect of vitamin E against prenatal and early postnatal ethanol treatment-induced heart abnormality in rats: a 3-month follow-up study. Int Immunopharmacol 26:72–79. CrossRefGoogle Scholar
  37. Singh SP, Patel DG, Snyder AK (1976) Adverse reversible effects of chronic ethanol intake on carbohydrate metabolism. Proc Soc Exp Biol Med 152:449–454CrossRefGoogle Scholar
  38. Steiner JL, Lang CH (2017) Etiology of alcoholic cardiomyopathy: mitochondria, oxidative stress and apoptosis. Int J Biochem Cell Biol 89:125–135. CrossRefGoogle Scholar
  39. Su CY, Chong KY, Owen OE, Dillmann WH, Chang C, Lai CC (1998) Constitutive and inducible hsp70s are involved in oxidative resistance evoked by heat shock or ethanol. J Mol Cell Cardiol 30:587–598. CrossRefGoogle Scholar
  40. Sun H-Y, Wang N-P, Halkos M, Kerendi F, Kin H, Guyton RA, Vinten-Johansen J, Zhao Z-Q (2006) Postconditioning attenuates cardiomyocyte apoptosis via inhibition of JNK and p38 mitogen-activated protein kinase signaling pathways. Apoptosis 11:1583–1593CrossRefGoogle Scholar
  41. Sun J, Chen X, Chen H, Ma Z, Zhou J (2015a) Maternal alcohol consumption before and during pregnancy and the risks of congenital heart defects in offspring: a systematic review and meta-analysis. Congenit Heart Dis 10:E216–E224. CrossRefGoogle Scholar
  42. Sun L, Fan H, Yang L, Shi L, Liu Y (2015b) Tyrosol prevents ischemia/reperfusion-induced cardiac injury in H9c2 cells: involvement of ROS, Hsp70, JNK and ERK, and apoptosis. Molecules 20:3758–3775. CrossRefGoogle Scholar
  43. Sun F, Zuo YZ, Ge J, Xia J, Li XN, Lin J, Zhang C, Xu HL, Li JL (2018) Transport stress induces heart damage in newly hatched chicks via blocking the cytoprotective heat shock response and augmenting nitric oxide production. Poult Sci 97:2638–2646. CrossRefGoogle Scholar
  44. Tobiasz AM, Duncan JR, Bursac Z, Sullivan RD, Tate DL, Dopico AM, Bukiya AN, Mari G (2018) The effect of prenatal alcohol exposure on fetal growth and cardiovascular parameters in a baboon model of pregnancy. Reprod Sci 25:1116–1123. CrossRefGoogle Scholar
  45. Wan C, Chen Y, Yin P, Han D, Xu X, He S, Liu M, Hou X, Liu F, Xu J (2016) Transport stress induces apoptosis in rat myocardial tissue via activation of the mitogen-activated protein kinase signaling pathways. Heart Vessel 31:212–221. CrossRefGoogle Scholar
  46. Wang L, Zhang TP, Zhang Y, Bi HL, Guan XM, Wang HX, Wang X, Du J, Xia YL, Li HH (2016) Protection against doxorubicin-induced myocardial dysfunction in mice by cardiac-specific expression of carboxyl terminus of hsp70-interacting protein. Sci Rep 6:28399.
  47. Wang P, Luo Q, Qiao H, Ding H, Cao Y, Yu J, Liu R, Zhang Q, Zhu H, Qu L (2017) The neuroprotective effects of carvacrol on ethanol-induced hippocampal neurons impairment via the antioxidative and antiapoptotic pathways. Oxidative Med Cell Longev 2017:4079425–4079417. Google Scholar
  48. Wang H, Li XN, Li PC, Liu W, Du ZH, Li JL (2019) Modulation of heat-shock response is associated with Di (2-ethylhexyl) phthalate (DEHP)-induced cardiotoxicity in quail (Coturnix japonica). Chemosphere 214:812–820. CrossRefGoogle Scholar
  49. Webster WS, Germain MA, Lipson A, Walsh D (1984) Alcohol and congenital heart defects: an experimental study in mice. Cardiovasc Res 18:335–338CrossRefGoogle Scholar
  50. Williamson CL, Dabkowski ER, Dillmann WH, Hollander JM (2008) Mitochondria protection from hypoxia/reoxygenation injury with mitochondria heat shock protein 70 overexpression. Am J Physiol Heart Circ Physiol 294:H249–H256. CrossRefGoogle Scholar
  51. Wu Y, Reece EA, Zhong J, Dong D, Shen WB, Harman CR, Yang P (2016) Type 2 diabetes mellitus induces congenital heart defects in murine embryos by increasing oxidative stress, endoplasmic reticulum stress, and apoptosis. Am J Obstet Gynecol 215:366.e361–366.e310. CrossRefGoogle Scholar
  52. Yan X, Pan B, Lv T, Liu L, Zhu J, Shen W, Huang X, Tian J (2017) Inhibition of histone acetylation by curcumin reduces alcohol-induced fetal cardiac apoptosis. J Biomed Sci 24(1):1. CrossRefGoogle Scholar
  53. Yao YW, Zhang GH, Zhang YY, Li WD, Wang CH, Yin CY, Zhang FM (2011) Lipopolysaccharide pretreatment protects against ischemia/reperfusion injury via increase of HSP70 and inhibition of NF-kappaB. Cell Stress Chaperones 16:287–296. CrossRefGoogle Scholar
  54. Zhao B, Wang Y, Li Y, Qiao X, Yan P, Zhu Y, Lai J (2015) Differential phosphorylation of NMDAR1-CaMKII-MAPKs in the rat nucleus accumbens following chronic ethanol exposure. Neurosci Lett 597:60–65. CrossRefGoogle Scholar
  55. Zhao Y, Fan JH, Luo Y, Talukder M, Li XN, Zuo YZ, Li JL (2019) Di-(2-ethylhexyl) phthalate (DEHP)-induced hepatotoxicity in quail (Coturnix japonica) via suppression of the heat shock response. Chemosphere 228:685–693. CrossRefGoogle Scholar
  56. Zhu Z, Huang Y, Lv L, Tao Y, Shao M, Zhao C, Xue M, Sun J, Niu C, Wang Y, Kim S, Cong W, Mao W, Jin L (2018) Acute ethanol exposure-induced autophagy-mediated cardiac injury via activation of the ROS-JNK-Bcl-2 pathway. J Cell Physiol 233:924–935. CrossRefGoogle Scholar

Copyright information

© Cell Stress Society International 2019

Authors and Affiliations

  1. 1.Nephrology and Kidney Transplant Research CenterUrmia University of Medical SciencesUrmiaIran
  2. 2.Department of Physiology, Faculty of MedicineUrmia University of Medical SciencesUrmiaIran

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