Skip to main content

Thank you for visiting nature.com. You are using a browser version with limited support for CSS. To obtain the best experience, we recommend you use a more up to date browser (or turn off compatibility mode in Internet Explorer). In the meantime, to ensure continued support, we are displaying the site without styles and JavaScript.

  • Basic Science Article
  • Published:

Fetal anemia and elevated hepcidin in a mouse model of fetal alcohol spectrum disorder

Pediatric Research volume 94, pages 503–511 (2023)Cite this article

Abstract

Introduction

Prenatal alcohol exposure (PAE) impairs offspring growth and cognition, and this is worsened by concurrent iron deficiency. Alcohol disrupts fetal iron metabolism and produces functional iron deficiency, even when maternal iron status is adequate. We used a mouse model of moderate PAE to investigate the mechanisms underlying this dysregulated iron status.

Methods

C57BL/6J female mice received 3 g/kg alcohol daily from embryonic day (E) 8.5–17.5 and were assessed at E17.5.

Results

Alcohol reduced fetal hemoglobin, hematocrit, and red blood cell counts, despite elevated erythropoietin production. Alcohol suppressed maternal hepcidin expression and the upstream iron-sensing BMP/SMAD pathway, consistent with its effects in the nonpregnant state. In contrast, alcohol elevated fetal hepcidin, although this was not accompanied by an upregulation of the BMP/SMAD or proinflammatory IL-6/STAT3 pathways. Fetal expression of hepatic genes contributing to hemoglobin synthesis and iron metabolism were unaffected by alcohol, whereas those affecting ribosome biogenesis were suppressed, suggesting a novel candidate effector for this fetal anemia.

Conclusion

These data confirm and extend prior observations that PAE disrupts maternal and fetal iron metabolism and impairs the fetus’s ability to regulate iron status. We propose this dysregulation increases gestational iron needs and represents a conserved response to PAE.

Impact

  • Prenatal alcohol exposure causes a functional iron deficiency in a model that also impairs cognition in later life.

  • Prenatal alcohol exposure causes fetal anemia.

  • This fetal anemia is accompanied by elevated hepcidin and erythropoietin.

  • Findings are consistent with prior observations that prenatal alcohol exposure increases maternal–fetal iron requirements during pregnancy.

This is a preview of subscription content, access via your institution

Access options

Buy this article

  • Purchase on Springer Link
  • Instant access to full article PDF
Buy now

Prices may be subject to local taxes which are calculated during checkout

Fig. 1: Alcohol exposure increased maternal and fetal erythropoietin (Epo) expression.
Fig. 2: Alcohol exposure trended to reduce hepcidin in the mother and increased hepcidin in the fetus.
Fig. 3: Alcohol exposure did not create a proinflammatory environment or alter indicators of the IL-6/JAK/STAT3 pathway.
Fig. 4: Alcohol exposure reduced the activity of the maternal BMP6/SMAD pathway and minimally affected the fetal BMP6/SMAD pathway.

Similar content being viewed by others

Data availability

Data from this study are available upon reasonable request for non-commercial purposes.

Materials availability

Materials from this study are available upon reasonable request for non-commercial purposes.

References

  1. Wozniak, J. R., Riley, E. P. & Charness, M. E. Clinical presentation, diagnosis, and management of fetal alcohol spectrum disorder. Lancet Neurol. 18, 760–770 (2019).

    PubMed  PubMed Central  Google Scholar 

  2. Young, J. K., Giesbrecth, H. E., Eskin, M. N., Aliani, M. & Suh, M. Nutrition implications for fetal alcohol spectrum disorder. Adv. Nutr. 5, 675–692 (2014).

    CAS  PubMed  PubMed Central  Google Scholar 

  3. Huebner, S. M., Blohowiak, S. E., Kling, P. J. & Smith, S. M. Prenatal alcohol exposure alters fetal iron distribution and elevates hepatic hepcidin in a rat model of fetal alcohol spectrum disorders. J. Nutr. 146, 1180–1188 (2016).

    CAS  PubMed  PubMed Central  Google Scholar 

  4. Huebner, S. M. et al. Dietary iron fortification normalizes fetal hematology, hepcidin, and iron distribution in a rat model of prenatal alcohol exposure. Alcohol Clin. Exp. Res. 42, 1022–1033 (2018).

    CAS  PubMed  PubMed Central  Google Scholar 

  5. Carter, R. C. et al. Prenatal alcohol-related alterations in maternal, placental, neonatal, and infant iron homeostasis. Am. J. Clin. Nutr. 114, 1107–1122 (2021).

    PubMed  PubMed Central  Google Scholar 

  6. Georgieff, M. K. Iron deficiency in pregnancy. Am. J. Obstet. Gynecol. 223, 516–524 (2020).

    CAS  PubMed  PubMed Central  Google Scholar 

  7. Katsarou, A. & Pantopoulos, K. Basics and principles of cellular and systemic iron homeostasis. Mol. Asp. Med. 75, 100866 (2020).

    CAS  Google Scholar 

  8. Harrison-Findik, D. D. et al. Iron-mediated regulation of liver hepcidin expression in rats and mice is abolished by alcohol. Hepatology 46, 1979–1985 (2007).

    CAS  PubMed  Google Scholar 

  9. Ioannou, G. N., Dominitz, J. A., Weiss, N. S., Heagerty, P. J. & Kowdley, K. V. The effect of alcohol consumption on the prevalence of iron overload, iron deficiency, and iron deficiency anemia. Gastroenterology 126, 1293–1301 (2004).

    CAS  PubMed  Google Scholar 

  10. Costa-Matos, L. et al. Liver hepcidin mRNA expression is inappropriately low in alcoholic patients compared with healthy controls. Eur. J. Gastroenterol. Hepatol. 24, 1158–1165 (2012).

    CAS  PubMed  Google Scholar 

  11. Harrison-Findik, D. D. Is the iron regulatory hormone hepcidin a risk factor for alcoholic liver disease? World J. Gastroenterol. 15, 1186–1193 (2009).

    CAS  PubMed  PubMed Central  Google Scholar 

  12. Sangkhae, V. & Nemeth, E. Regulation of the iron homeostatic hormone hepcidin. Adv. Nutr. Int. Rev. J. 8, 126–136 (2017).

    CAS  Google Scholar 

  13. Wang, C. Y., Meynard, D. & Lin, H. Y. The role of TMPRSS6/matriptase-2 in iron regulation and anemia. Front. Pharmacol. 5, 114 (2014).

    PubMed  PubMed Central  Google Scholar 

  14. Xiao, X., Alfaro-Magallanes, V. M. & Babitt, J. L. Bone morphogenic proteins in iron homeostasis. Bone 138, 115495 (2020).

    CAS  PubMed  PubMed Central  Google Scholar 

  15. Canali, S., Wang, C., Zumbrennen-Bullough, K. B., Bayer, A. & Babitt, J. L. Bone morphogenetic protein 2 controls iron homeostasis in mice independent of Bmp6. Blood 92, 1204–1213 (2017).

    CAS  Google Scholar 

  16. Carow, B. & Rottenberg, M. E. SOCS3, a major regulator of infection and inflammation. Front Immunol. 5, 1–13 (2014).

    CAS  Google Scholar 

  17. Huang, H., Constante, M., Layoun, A. & Santos, M. M. Contribution of STAT3 and SMAD4 pathways to the regulation of hepcidin by opposing stimuli. Blood 113, 3593–3599 (2009).

    CAS  PubMed  Google Scholar 

  18. Saini, N. et al. Alcohol’s dysregulation of maternal–fetal IL-6 and p-STAT3 is a function of maternal iron status. Alcohol Clin. Exp. Res 43, 2332–2343 (2009).

    Google Scholar 

  19. Mooney, S. M., Pjetri, E., Friday, W. B. & Smith, S. M. Growth and behavioral differences in a C57BL/6J mouse model of prenatal alcohol exposure. Alcohol 97, 51–57 (2021).

    CAS  PubMed  PubMed Central  Google Scholar 

  20. Virdee, M. S. et al. An enriched biosignature of gut microbiota-derived metabolites characterizes maternal plasma in a mouse model of fetal alcohol spectrum disorder. Sci. Rep. 11, 1–16 (2021).

    Google Scholar 

  21. Smith, S. M. et al. Aging-related behavior, adiposity, and glucose impairments and their association following prenatal alcohol exposure in the C57BL/6J mouse. Nutrients 14, 1438 (2022).

    CAS  PubMed  PubMed Central  Google Scholar 

  22. Saini, N. et al. Untargeted metabolome analysis reveals reductions in maternal hepatic glucose and amino acid content that correlate with fetal organ weights in a mouse model of fetal alcohol spectrum disorders. Nutrients 14, 1096 (2022).

    CAS  PubMed  PubMed Central  Google Scholar 

  23. Walter, K. R., Ricketts, D. K., Presswood, B. H., Smith, S. M. & Mooney, S. M. Prenatal alcohol exposure causes persistent microglial activation and age- and sex- specific effects on cognition and metabolic outcomes in an Alzheimer’s Disease mouse model. Am. J. Drug Alcohol Abus. 4, 1–19 (2022).

    Google Scholar 

  24. Reeves, P. G., Neilson, F. H. & Fahey, G. C. Jr AIN-93 purified diets for laboratory rodents: final report of the American Institute of Nutrition ad hoc writing committee on the reformulation of the AIN-76A rodent diet. J. Nutr. 123, 1939–1951 (1993).

    CAS  PubMed  Google Scholar 

  25. Bustin, S. A. et al. The MIQE Guidelines: minimum information for publication of quantitative real-time PCR experiments. Clin. Chem. 55, 1–12 (2009).

    Google Scholar 

  26. Kwan, S. T. C. et al. Maternal iron nutriture modulates placental development in a rat model of fetal alcohol spectrum disorder. Alcohol 84, 57–66 (2020).

    CAS  PubMed  Google Scholar 

  27. Livak, K. J. & Schmittgen, T. D. Analysis of relative gene expression data using real-time quantitative PCR and the 2−ΔΔCT method. Methods 25, 402–408 (2001).

    CAS  PubMed  Google Scholar 

  28. Berres, M. E., Garic, A., Flentke, G. R. & Smith, S. M. Transcriptome profiling identifies ribosome biogenesis as a target of alcohol teratogenicity and vulnerability during early embryogenesis. PLoS ONE 12, e0169351 (2017).

    PubMed  PubMed Central  Google Scholar 

  29. Garic, A., Berres, M. E. & Smith, S. M. High-throughput transcriptome sequencing identifies candidate genetic modifiers of vulnerability to fetal alcohol spectrum disorders. Alcohol Clin. Exp. Res. 38, 1874–1882 (2014).

    CAS  PubMed  PubMed Central  Google Scholar 

  30. Anders, S. & Huber, W. Differential expression analysis for sequence count data. Genome Biol. 11, R106 (2010).

    CAS  PubMed  PubMed Central  Google Scholar 

  31. Langmead, B. & Salzberg, S. L. Fast gapped-read alignment with Bowtie 2. Nat. Methods 9, 357–359 (2012).

    CAS  PubMed  PubMed Central  Google Scholar 

  32. Liao, Y., Smyth, G. K. & Shi, W. featureCounts: an efficient general-purpose program for assigning sequence reads to genomic features. Bioinformatics 30, 923–930 (2014).

    CAS  PubMed  Google Scholar 

  33. Love, M. I., Huber, W. & Anders, S. Moderated estimation of fold change and dispersion for RNA-seq data with DESeq2. Genome Biol. 15, 1–21 (2014).

    Google Scholar 

  34. Lombardero, M., Kovacs, K. & Scheithauer, B. W. Erythropoietin: a hormone with multiple functions. Pathobiology 78, 41–56 (2011).

    CAS  PubMed  Google Scholar 

  35. Yang, A. et al. Expression of hepcidin and ferroportin in the placenta, and ferritin and transferrin receptor 1 levels in maternal and umbilical cord blood in pregnant women with and without gestational diabetes. Int. J. Environ. Res. Public Health 13, 766 (2016).

    PubMed  PubMed Central  Google Scholar 

  36. Cardaropoli, S., Todros, T., Nuzzo, A. M. & Rolfo, A. Maternal serum levels and placental expression of hepcidin in preeclampsia. Pregnancy Hypertens. 11, 47–53 (2018).

    PubMed  Google Scholar 

  37. Meynard, D. et al. Lack of the bone morphogenetic protein BMP6 induces massive iron overload. Nat. Genet. 41, 478–481 (2009).

    CAS  PubMed  Google Scholar 

  38. Helfrich, K. K. et al. Gestational iron supplementation improves fetal outcomes in a rat model of prenatal alcohol exposure. Nutrients 14, 1653 (2022).

    CAS  PubMed  PubMed Central  Google Scholar 

  39. Kampen, K. R., Sulima, S. O., Vereecke, S., De & Keersmaecker, K. Hallmarks of ribosomopathies. Nucleic Acids Res. 48, 1013–1028 (2020).

    CAS  PubMed  Google Scholar 

  40. Flentke, G. R., Baulch, J. W., Berres, M. E., Garic, A. & Smith, S. M. Alcohol-mediated calcium signals dysregulate pro-survival Snai2/PUMA/Bcl2 networks to promote p53-mediated apoptosis in avian neural crest progenitors. Birth Defects Res. 111, 686–699 (2019).

    CAS  PubMed  PubMed Central  Google Scholar 

  41. Halmesmaki, E., Teramo, K. A., Widness, J. A., Clemsons, G. K. & Ylikorkala, O. Maternal alcohol abuse is associated with elevated fetal erythropoietin levels. Obs. Gynecol. 76, 219–222 (1990).

    CAS  Google Scholar 

  42. Davis, M. & Clarke, S. Influence of microRNA on the maintenance of human iron metabolism. Nutrients 5, 2611–2628 (2013).

    CAS  PubMed  PubMed Central  Google Scholar 

  43. Mahnke, A. H., Salem, N. A., Tseng, A. M., Chung, D. D. & Miranda, R. C. Nonprotein-coding RNAs in fetal alcohol spectrum disorders. Prog. Mol. Biol. Transl. Sci. 157, 299–342 (2018).

    CAS  PubMed  Google Scholar 

  44. Huebner, S. M., Tran, T. D., Rufer, E. S., Crump, P. M. & Smith, S. M. Maternal iron deficiency worsens the associative learning deficits and hippocampal and cerebellar losses in a rat model of fetal alcohol spectrum disorders. Alcohol Clin. Exp. Res. 39, 2097–2107 (2015).

    CAS  PubMed  PubMed Central  Google Scholar 

  45. Rufer, E. S. et al. Adequacy of maternal iron status protects against behavioral, neuroanatomical, and growth deficits in fetal alcohol spectrum disorders. PLoS ONE 7, e47199 (2012).

    Google Scholar 

Download references

Acknowledgements

We thank Brandon Presswood and Dane Ricketts for technical support, and Pamela Kling for helpful suggestions during this study.

Funding

This study was supported by R01 AA022999 and R01 AA011085 to S.M.S., T32 DK007686 to K.K.H., K99 AA028291 to N.S., F32 AA028684 to O.C.R., and F32 AA027121 to S.T.C.K.

Author information

Authors and Affiliations

  1. Nutrition Research Institute, University of North Carolina at Chapel Hill, Kannapolis, NC, 28081, USA

    Kaylee K. Helfrich, Nipun Saini, Sze Ting (Cecilia) Kwan, Olivia C. Rivera, Sandra M. Mooney & Susan M. Smith

  2. Department of Nutrition, University of North Carolina at Chapel Hill, Kannapolis, NC, 28081, USA

    Kaylee K. Helfrich, Sandra M. Mooney & Susan M. Smith

Authors
  1. Kaylee K. Helfrich
    View author publications

    You can also search for this author in PubMed Google Scholar

  2. Nipun Saini
    View author publications

    You can also search for this author in PubMed Google Scholar

  3. Sze Ting (Cecilia) Kwan
    View author publications

    You can also search for this author in PubMed Google Scholar

  4. Olivia C. Rivera
    View author publications

    You can also search for this author in PubMed Google Scholar

  5. Sandra M. Mooney
    View author publications

    You can also search for this author in PubMed Google Scholar

  6. Susan M. Smith
    View author publications

    You can also search for this author in PubMed Google Scholar

Contributions

Conception and design: K.K.H., N.S., S.M.M., and S.M.S.; data acquisition: K.K.H., N.S., S.T.C.K., O.C.R.; data analysis and interpretation: K.K.H., N.S., S.M.M., S.M.S.; manuscript drafting: K.K.H. and S.M.S. All authors read and approved the manuscript.

Corresponding author

Correspondence to Susan M. Smith.

Ethics declarations

Competing interests

The authors declare no competing interests.

Ethics approval and consent to participate

Patient consent is not relevant to this study.

Additional information

Publisher’s note Springer Nature remains neutral with regard to jurisdictional claims in published maps and institutional affiliations.

Supplementary Information

Rights and permissions

Springer Nature or its licensor (e.g. a society or other partner) holds exclusive rights to this article under a publishing agreement with the author(s) or other rightsholder(s); author self-archiving of the accepted manuscript version of this article is solely governed by the terms of such publishing agreement and applicable law.

Reprints and permissions

About this article

Check for updates. Verify currency and authenticity via CrossMark

Cite this article

Helfrich, K.K., Saini, N., Kwan, S.T.(. et al. Fetal anemia and elevated hepcidin in a mouse model of fetal alcohol spectrum disorder. Pediatr Res 94, 503–511 (2023). https://doi.org/10.1038/s41390-023-02469-6

Download citation

  • Received:

  • Revised:

  • Accepted:

  • Published:

  • Issue Date:

  • DOI: https://doi.org/10.1038/s41390-023-02469-6

This article is cited by

Search

Advanced search

Quick links