Gestational Melatonin Supplementation Attenuates Maternal Sleep Deprivation-Induced Steatohepatitis Susceptibility in Offspring

Abstract

Sleep deprivation (SD) is a common issue among pregnant women. Maternal SD led to adverse effects on offspring health such as cognitive impairment through dysregulated metabolic pathways. However, it remains unknown whether maternal SD increases the offspring's susceptibility to nonalcoholic steatohepatitis (NASH) development. Here, we induced maternal SD during pregnancy and observed that maternal SD during pregnancy promoted the development of diet-induced NASH in offspring of both sexes in adulthood, with exacerbation of liver weight gain, hepatic steatosis, fibrosis, and hepatic dysfunction. The primary hepatocytes isolated from SD offspring were also more susceptible to palmitate acid-induced lipotoxic injury. Mechanistically, the detrimental effects of maternal SD were associated with augmented activation of inflammatory and apoptosis pathways in offspring liver tissues, which were attributed to upregulation of the transcription factor nuclear receptor subfamily 4 group A member 3 (NR4A3). The melatonin signalling is reported to be pivotally affected by sleep disturbance both at the circulation and the placenta, and our further analysis revealed that melatonin supplementation during maternal SD normalised NR4A3 expression in offspring liver and alleviated the increased steatohepatitis susceptibility in offspring. Taken together, these results suggest that maternal SD during pregnancy predisposes offspring to NASH development in adulthood via an NR4A3-dependent mechanism, and maternal melatonin supplementation may hold promise for improving liver health in the offspring.

Keywords

inflammation / melatonin / nonalcoholic steatohepatitis / offspring / sleep deprivation

References

[1] V. W. Wong, M. Ekstedt, G. L. Wong, and H. Hagström, “Changing Epidemiology, Global Trends and Implications for Outcomes of NAFLD,” Journal of Hepatology 79, no. 3 (2023): 842–852.

[2] E. Scorletti and R. M. Carr, “A New Perspective on NAFLD: Focusing on Lipid Droplets,” Journal of Hepatology 76, no. 4 (2022): 934–945.

[3] L. Henry, J. Paik, and Z. M. Younossi, “Review Article: the Epidemiologic Burden of Non-Alcoholic Fatty Liver Disease Across the World,” Alimentary Pharmacology & Therapeutics 56, no. 6 (2022): 942–956.

[4] E. Kotsiliti, “NAFLD Incidence Is Increasing,” Nature Reviews Gastroenterology & Hepatology 20, no. 6 (2023): 344.

[5] T. Huby and E. L. Gautier, “Immune Cell-Mediated Features of Non-Alcoholic Steatohepatitis,” Nature Reviews. Immunology 22, no. 7 (2022): 429–443.

[6] Y. Saiman, R. Hooks, and R. M. Carr, “High-Risk Groups for Non-Alcoholic Fatty Liver and Non-Alcoholic Steatohepatitis Development and Progression,” Current Hepatology Reports 19, no. 4 (2020): 412–419.

[7] C. D. Byrne and G. Targher, “NAFLD: a Multisystem Disease,” Journal of Hepatology 62, no. 1 Suppl (2015): S47–S64.

[8] C. D. Byrne, P. N. Newsome, and M. Noureddin, “Why Are There no Strategies for NAFLD,” Journal of Hepatology 76, no. 4 (2022): 763–764.

[9] L. Gao, Y. Zhao, H. Wu, et al., “Polycystic Ovary Syndrome Fuels Cardiovascular Inflammation and Aggravates Ischemic Cardiac Injury,” Circulation 148, no. 24 (2023): 1958–1973.

[10] H. Hagström, T. G. Simon, B. Roelstraete, O. Stephansson, J. Söderling, and J. F. Ludvigsson, “Maternal Obesity Increases the Risk and Severity of NAFLD in Offspring,” Journal of Hepatology 75, no. 5 (2021): 1042–1048.

[11] X. Dong, D. Lin, J. Sheng, and Y. Xie, “Intrauterine Hyperglycemia Induces Liver Inflammation in Mouse Male Offspring,” International Immunopharmacology 99 (2021): 107974.

[12] C. E. McCurdy, J. M. Bishop, S. M. Williams, et al., “Maternal High-Fat Diet Triggers Lipotoxicity in the Fetal Livers of Nonhuman Primates,” Journal of Clinical Investigation 119, no. 2 (2009): 323–335.

[13] A. Shrestha, S. K. Dellett, J. Yang, U. Sharma, and L. Ramalingam, “Effects of Fish Oil Supplementation on Reducing the Effects of Paternal Obesity and Preventing Fatty Liver in Offspring,” Nutrients 15, no. 24 (2023): 5038.

[14] M. Smyka, K. Kosińska-Kaczyńska, N. Sochacki-Wójcicka, M. Zgliczyńska, and M. Wielgoś, “Sleep Problems in Pregnancy-A Cross-Sectional Study in Over 7000 Pregnant Women in Poland,” International Journal of Environmental Research and Public Health 17, no. 15 (2020): 5306.

[15] R. Manber, D. Steidtmann, A. S. Chambers, W. Ganger, S. Horwitz, and C. D. Connelly, “Factors Associated With Clinically Significant Insomnia Among Pregnant Low-Income Latinas,” Journal of Women's Health (2002) 22, no. 8 (2013): 694–701.

[16] Q. Zhao, C. Peng, X. Wu, Y. Chen, C. Wang, and Z. You, “Maternal Sleep Deprivation Inhibits Hippocampal Neurogenesis Associated With Inflammatory Response in Young Offspring Rats,” Neurobiology of Disease 68 (2014): 57–65.

[17] Y. M. Zhang, R. M. Wei, M. Z. Ni, et al., “An Enriched Environment Improves Maternal Sleep Deprivation-Induced Cognitive Deficits and Synaptic Plasticity via Hippocampal Histone Acetylation,” Brain and Behavior: A Cognitive Neuroscience Perspective 13, no. 6 (2023): e3018.

[18] M. W. Harskamp-van Ginkel, D. Ierodiakonou, K. Margetaki, et al., “Gestational Sleep Deprivation Is Associated With Higher Offspring Body Mass Index and Blood Pressure,” Sleep 43, no. 12 (2020): 1–9.

[19] T. A. Alvarenga, M. F. Aguiar, R. Mazaro-Costa, S. Tufik, and M. L. Andersen, “Effects of Sleep Deprivation During Pregnancy on the Reproductive Capability of the Offspring,” Fertility and Sterility 100, no. 6 (2013): 1752–1757.

[20] K. Choi, Y. J. Lee, S. Park, N. K. Je, and H. S. Suh, “Efficacy of Melatonin for Chronic Insomnia: Systematic Reviews and Meta-Analyses,” Sleep Medicine Reviews 66 (2022): 101692.

[21] Y. L. Tain and C. N. Hsu, “Melatonin Use During Pregnancy and Lactation Complicated by Oxidative Stress: Focus on Offspring's Cardiovascular-Kidney-Metabolic Health in Animal Models,” Antioxidants (Basel) 13, no. 2 (2024): 226.

[22] D. E. Blask, “Melatonin, Sleep Disturbance and Cancer Risk,” Sleep Medicine Reviews 13, no. 4 (2009): 257–264.

[23] J. Li, X. Peng, H. Zhong, et al., “Sleep Deprivation During Pregnancy Leads to Poor Fetal Outcomes in Sprague-Dawley Rats,” Journal of Reproductive Immunology 160 (2023): 104166.

[24] Y. M. Zhang, R. M. Wei, Z. Y. Li, et al., “Melatonin Improves Maternal Sleep Deprivation-Induced Learning and Memory Impairment, Inflammation, and Synaptic Dysfunction in Murine Male Adult Offspring,” Brain and Behavior: A Cognitive Neuroscience Perspective 14, no. 5 (2024): e3515.

[25] K. Wang, J. Liu, J. Xie, et al., “Sleep Deprivation From Mid-Gestation Leads to Impaired of Motor Coordination in Young Offspring Mice With Microglia Activation in the Cerebellar Vermis,” Sleep Medicine 115 (2024): 193–201.

[26] W. Liang, A. L. Menke, A. Driessen, et al., “Establishment of a General NAFLD Scoring System for Rodent Models and Comparison to Human Liver Pathology,” PLoS One 9, no. 12 (2014): e115922.

[27] G. Yu, “Thirteen Years of clusterProfiler,” Innovation (Camb) 5, no. 6 (2024): 100722.

[28] Q. Fan, M. Chen, L. Zuo, et al., “Myocardial Ablation of G Protein-Coupled Receptor Kinase 2 (GRK2) Decreases Ischemia/Reperfusion Injury Through an Anti-Intrinsic Apoptotic Pathway,” PLoS One 8, no. 6 (2013): e66234.

[29] A. H. Yang, M. A. Tincopa, F. Tavaglione, et al., “Prevalence of Steatotic Liver Disease, Advanced Fibrosis and Cirrhosis Among Community-Dwelling Overweight and Obese Individuals in the USA,” Gut 73, no. 12 (2024): 2045–2053.

[30] P. S. Dalvi, S. Yang, N. Swain, et al., “Long-Term Metabolic Effects of Malnutrition: Liver Steatosis and Insulin Resistance Following Early-Life Protein Restriction,” PLoS One 13, no. 7 (2018): e0199916.

[31] J. D. Ly, D. R. Grubb, and A. Lawen, “The mitochondrial membrane potential (deltapsi(m)) in apoptosis; an update,” Apoptosis 8, no. 2 (2003): 115–128.

[32] M. Nagaoka, T. Yashiro, Y. Uchida, et al., “The Orphan Nuclear Receptor NR4A3 Is Involved in the Function of Dendritic Cells,” Journal of Immunology 199, no. 8 (2017): 2958–2967.

[33] J. Martínez-González, L. Cañes, J. Alonso, et al., “NR4A3: A Key Nuclear Receptor in Vascular Biology, Cardiovascular Remodeling, and Beyond,” International Journal of Molecular Sciences 22, no. 21 (2021): 11371.

[34] Z. Fang, H. G. Kim, M. Huang, et al., “Sestrin Proteins Protect Against Lipotoxicity-Induced Oxidative Stress in the Liver via Suppression of C-Jun N-Terminal Kinases,” Cellular and Molecular Gastroenterology and Hepatology 12, no. 3 (2021): 921–942.

[35] A. Deutsch, B. Rinner, M. Pichler, et al., “NR4A3 Suppresses Lymphomagenesis Through Induction of Proapoptotic Genes,” Cancer Research 77, no. 9 (2017): 2375–2386.

[36] Y. Zhao, L. Gao, J. Chen, et al., “Remote Limb Ischemic Conditioning Alleviates Steatohepatitis via Extracellular Vesicle-Mediated Muscle-Liver Crosstalk,” Cell Metabolism 37, no. 4 (2025): 886–902.e7.

[37] E. Kaye, E. Marques, J. Agudelo Areiza, S. Modaresi, and A. Slitt, “Exposure to a PFOA, PFOS and PFHxS Mixture During Gestation and Lactation Alters the Liver Proteome in Offspring of CD-1 Mice,” Toxics 12, no. 5 (2024): 348.

[38] Y. Q. Su, Y. Lin, S. J. Huang, et al., “Pyroptosis Is Involved in Maternal Nicotine Exposure-Induced Metabolic Associated Fatty Liver Disease Progression in Offspring Mice,” Molecular Reproduction and Development 91, no. 8 (2024): e23719.

[39] R. Wang, W. Feng, Y. Wang, Y. Jiang, Y. Lin, and X. Chen, “Maternal Obstructive Sleep Apnea Aggravates Metabolic Dysfunction-Associated Fatty Liver Disease via HMGB1-TLR4 Signaling-Mediated Endoplasmic Reticulum Stress in Male Offspring Rats,” Biochimica et Biophysica Acta - Molecular Basis of Disease 1870, no. 1 (2024): 166889.

[40] Z. Long, J. Fan, G. Wu, et al., “Gestational Bisphenol A Exposure Induces Fatty Liver Development in Male Offspring Mice Through the Inhibition of HNF1b and Upregulation of PPARγ,” Cell Biology and Toxicology 37, no. 1 (2021): 65–84.

[41] R. Lin, D. Wu, F. J. Wu, et al., “Non-Alcoholic Fatty Liver Disease Induced by Perinatal Exposure to Bisphenol a Is Associated With Activated mTOR and TLR4/NF-κB Signaling Pathways in Offspring Rats,” Frontiers in Endocrinology (Lausanne) 10 (2019): 620.

[42] D. E. Brumbaugh, P. Tearse, M. Cree-Green, et al., “Intrahepatic Fat Is Increased in the Neonatal Offspring of Obese Women With Gestational Diabetes,” Journal of Pediatrics 162, no. 5 (2013): 930-6.e1.

[43] A. Mouralidarane, J. Soeda, C. Visconti-Pugmire, et al., “Maternal Obesity Programs Offspring Nonalcoholic Fatty Liver Disease by Innate Immune Dysfunction in Mice,” Hepatology 58, no. 1 (2013): 128–138.

[44] A. R. Purcell, N. Rodrigo, Q. Cao, et al., “Maternal Weight Intervention in the Perinatal Period Improves Liver Health in the Offspring of Mothers With Obesity,” Nutrients 16, no. 1 (2023): 109.

[45] H. A. Paul, K. H. Collins, A. C. Nicolucci, et al., “Maternal Prebiotic Supplementation Reduces Fatty Liver Development in Offspring Through Altered Microbial and Metabolomic Profiles in Rats,” FASEB Journal 33, no. 4 (2019): 5153–5167.

[46] M. G. Pruis, A. Lendvai, V. W. Bloks, et al., “Maternal Western Diet Primes Non-Alcoholic Fatty Liver Disease in Adult Mouse Offspring,” Acta Physiologica (Oxford) 210, no. 1 (2014): 215–227.

[47] L. T. Nguyen, H. Chen, A. Zaky, C. Pollock, and S. Saad, “SIRT1 Overexpression Attenuates Offspring Metabolic and Liver Disorders as a Result of Maternal High-Fat Feeding,” Journal of Physiology 597, no. 2 (2019): 467–480.

[48] H. Peng, J. Li, H. Xu, et al., “Offspring NAFLD Liver Phospholipid Profiles Are Differentially Programmed by Maternal High-Fat Diet and Maternal One Carbon Supplement,” Journal of Nutritional Biochemistry 111 (2023): 109187.

[49] Y. Guo, P. Zhou, L. Qiao, H. Guan, J. Gou, and X. Liu, “Maternal Protein Deficiency Impairs Peroxisome Biogenesis and Leads to Oxidative Stress and Ferroptosis in Liver of Fetal Growth Restriction Offspring,” Journal of Nutritional Biochemistry 121 (2023): 109432.

[50] P. Li, K. Yan, X. Chang, et al., “Sex-Specific Maternal Calcium Requirements for the Prevention of Nonalcoholic Fatty Liver Disease by Altering the Intestinal Microbiota and Lipid Metabolism in the High-Fat-Diet-Fed Offspring Mice,” Gut Microbes 11, no. 6 (2020): 1590–1607.

[51] H. Liu, B. He, W. Hu, et al., “Prenatal Dexamethasone Exposure Induces Nonalcoholic Fatty Liver Disease in Male Rat Offspring via the miR-122/YY1/ACE2-MAS1 Pathway,” Biochemical Pharmacology 185 (2021): 114420.

[52] C. D. Byrne and G. Targher, “What's New in NAFLD Pathogenesis, Biomarkers and Treatment,” Nature Reviews. Gastroenterology & Hepatology 17, no. 2 (2020): 70–71.

[53] F. Heinecke, M. B. Mazzucco, D. Fornes, S. Roberti, A. Jawerbaum, and V. White, “The Offspring From Rats Fed a Fatty Diet Display Impairments in the Activation of Liver Peroxisome Proliferator Activated Receptor Alpha and Features of Fatty Liver Disease,” Molecular and Cellular Endocrinology 511 (2020): 110818.

[54] N. da Silva Lima, M. F. Fondevila, E. Nóvoa, et al., “Inhibition of ATG3 Ameliorates Liver Steatosis by Increasing Mitochondrial Function,” Journal of Hepatology 76, no. 1 (2022): 11–24.

[55] S. Sutti and E. Albano, “Adaptive Immunity: An Emerging Player in the Progression of NAFLD,” Nature Reviews. Gastroenterology & Hepatology 17, no. 2 (2020): 81–92.

[56] E. J. Park, J. H. Lee, G. Y. Yu, et al., “Dietary and Genetic Obesity Promote Liver Inflammation and Tumorigenesis by Enhancing IL-6 and TNF Expression,” Cell 140, no. 2 (2010): 197–208.

[57] M. A. Febbraio, S. Reibe, S. Shalapour, G. J. Ooi, M. J. Watt, and M. Karin, “Preclinical Models for Studying NASH-Driven HCC: How Useful Are They,” Cell Metabolism 29, no. 1 (2019): 18–26.

[58] X. Wang, Q. He, C. Zhou, et al., “Prolonged Hypernutrition Impairs TREM2-Dependent Efferocytosis to License Chronic Liver Inflammation and NASH Development,” Immunity 56, no. 1 (2023): 58–77.e11.

[59] Q. Zhao, X. Xie, Y. Fan, et al., “Phenotypic Dysregulation of Microglial Activation in Young Offspring Rats With Maternal Sleep Deprivation-Induced Cognitive Impairment,” Scientific Reports 5 (2015): 9513.

[60] J. Lv, S. Xu, C. Meng, et al., “Ferroptosis Participated in Hippocampal Neuroinflammation Damage of in Offspring Rats After Maternal Sleep Deprivation,” Journal of Neuroimmunology 375 (2023): 578021.

[61] A. M. Baratta, N. R. Kanyuch, C. A. Cole, H. Valafar, J. Deslauriers, and A. Pocivavsek, “Acute Sleep Deprivation During Pregnancy in Rats: Rapid Elevation of Placental and Fetal Inflammation and Kynurenic Acid,” Neurobiology of Stress 12 (2020): 100204.

[62] J. Vierstra, J. Lazar, R. Sandstrom, et al., “Global Reference Mapping of Human Transcription Factor Footprints,” Nature 583, no. 7818 (2020): 729–736.

[63] Z. Wang, G. Benoit, J. Liu, et al., “Structure and Function of Nurr1 Identifies a Class of Ligand-Independent Nuclear Receptors,” Nature 423, no. 6939 (2003): 555–560.

[64] T. Sekiya, “Comparison Between Nr4a Transcription Factor Regulation and Function in Lymphoid and Tumor Treg Cells,” Frontiers in Immunology 13 (2022): 866339.

[65] A. F. Close, N. Dadheech, H. Lemieux, Q. Wang, and J. Buteau, “Disruption of Beta-Cell Mitochondrial Networks by the Orphan Nuclear Receptor Nor1/Nr4a3,” Cells 9, no. 1 (2020): 168.

[66] Y. Zhao, D. A. Howatt, F. Gizard, et al., “Deficiency of the NR4A Orphan Nuclear Receptor NOR1 Decreases Monocyte Adhesion and Atherosclerosis,” Circulation Research 107, no. 4 (2010): 501–511.

[67] L. Pei, A. Castrillo, and P. Tontonoz, “Regulation of Macrophage Inflammatory Gene Expression by the Orphan Nuclear Receptor Nur77,” Molecular Endocrinology 20, no. 4 (2006): 786–794.

[68] L. Cañes, I. Martí-Pàmies, C. Ballester-Servera, et al., “High NOR-1 (Neuron-Derived Orphan Receptor 1) Expression Strengthens the Vascular Wall Response to Angiotensin II Leading to Aneurysm Formation in Mice,” Hypertension 77, no. 2 (2021): 557–570.

[69] H. Qing, K. L. Jones, E. B. Heywood, H. Lu, A. Daugherty, and D. Bruemmer, “Deletion of the NR4A Nuclear Receptor NOR1 in Hematopoietic Stem Cells Reduces Inflammation but Not Abdominal Aortic Aneurysm Formation,” BMC Cardiovascular Disorders 17, no. 1 (2017): 271.

[70] N. E. Ilott, T. Schneider, J. Mill, et al., “Long-Term Effects of Gestational Nicotine Exposure and Food-Restriction on Gene Expression in the Striatum of Adolescent Rats,” PLoS One 9, no. 2 (2014): e88896.

[71] R. J. Reiter, D. X. Tan, A. Korkmaz, and S. A. Rosales-Corral, “Melatonin and Stable Circadian Rhythms Optimize Maternal, Placental and Fetal Physiology,” Human Reproduction Update 20, no. 2 (2014): 293–307.

[72] Y. L. Tain, L. T. Huang, C. N. Hsu, and C. T. Lee, “Melatonin Therapy Prevents Programmed Hypertension and Nitric Oxide Deficiency in Offspring Exposed to Maternal Caloric Restriction,” Oxidative Medicine and Cellular Longevity 2014 (2014): 283180.

[73] Y. L. Tain, C. T. Lee, J. Y. Chan, and C. N. Hsu, “Maternal Melatonin or N-Acetylcysteine Therapy Regulates Hydrogen Sulfide-Generating Pathway and Renal Transcriptome to Prevent Prenatal N(G)-Nitro-L-Arginine-Methyl Ester (L-NAME)-Induced Fetal Programming of Hypertension in Adult Male Offspring,” American Journal of Obstetrics and Gynecology 215, no. 5 (2016): 636.e1–636.e72.

[74] C. Lapa Neto, I. de Melo, P. Alpiovezza, et al., “Melatonin Associated With a High-Fat Diet During Pregnancy and Lactation Prevents Liver Changes in the Offspring,” General and Comparative Endocrinology 343 (2023): 114357.

[75] M. Ajackson, B. A. Nagagata, I. A. Marcondes-de-Castro, C. A. Mandarim-de-Lacerda, and M. B. Aguila, “Adult Mice Offspring of Obese Mothers Supplemented With Melatonin Show Lessened Liver Steatosis, Inflammation, Oxidative Stress, and Endoplasmic Reticulum Stress,” Life Sciences 312 (2023): 121253.

[76] A. S. McElhinny, F. C. Davis, and C. M. Warner, “The Effect of Melatonin on Cleavage Rate of C57BL/6 and CBA/ca Preimplantation Embryos Cultured in Vitro,” Journal of Pineal Research 21, no. 1 (1996): 44–48.

RIGHTS & PERMISSIONS

2025 The Author(s). Cell Proliferation published by Beijing Institute for Stem Cell and Regenerative Medicine and John Wiley & Sons Ltd.