Fetal and neonatal iron deficiency but not copper deficiency increases vascular complexity in the developing rat brain

Abstract

Objectives: Anemia caused by nutritional deficiencies, such as iron and copper deficiencies, is a global health problem. Iron and copper deficiencies have their most profound effect on the developing fetus/infant, leading to brain development deficits and poor cognitive outcomes. Tissue iron depletion or chronic anemia can induce cellular hypoxic signaling. In mice, chronic hypoxia induces a compensatory increase in brain blood vessel outgrowth. We hypothesized that developmental anemia, due to iron or copper deficiencies, induces angiogenesis/vasculogenesis in the neonatal brain.

Methods: To test our hypothesis, three independent experiments were performed where pregnant rats were fed iron- or copper-deficient diets from gestational day 2 through mid-lactation. Effects on the neonatal brain vasculature were determined using quantitative real-time polymerase chain reaction to assess mRNA levels of angiogenesis/vasculogenesis-associated genes and GLUT1 immunohistochemistry to assess brain blood vessel density and complexity.

Results: Iron deficiency, but not copper deficiency, increased mRNA expression of brain endothelial cell- and angiogenesis/vasculogenesis-associated genes (i.e. Glut1, Vwf, Vegfa, Ang2, Cxcl12, and Flk1) in the neonatal brain, suggesting increased cerebrovascular density. Iron deficiency also increased hippocampal and cerebral cortical blood vessel branching by 62 and 78%, respectively.

Discussion: This study demonstrates increased blood vessel complexity in the neonatal iron-deficient brain, which is likely due to elevated angiogenic/vasculogenic signaling. At least initially, this is probably an adaptive response to maintain metabolic substrate homeostasis in the developing iron-deficient brain. However, this may also contribute to long-term neurodevelopmental deficits.

Keywords:

• Iron deficiency
• Copper deficiency
• Anemia
• Brain
• Development
• Neovasculogenesis
• Angiogenesis
• Blood–brain barrier

Acknowledgments

We thank the members of the Anderson and Prohaska labs for their invaluable assistance with tissue collection and selected assays. In particular we would like to thank Ariel Johnson, Margaret Broderius, and Kevin Viken. The Duluth Medical Research Institute core facilities were utilized for qPCR experiments. The Duluth Imaging Center was used for confocal microscopy. T.W.B. received financial support from the UM Lyle and Sharon Bighley Graduate Fellowship and the UM Doctoral Dissertation Fellowship. S.S. received financial support from the Pathways to Advanced Degrees in Life Sciences program. Grants supporting this research included NIH 5R03HD055423-02.

Disclaimer statements

Contributors

All authors contributed to the analysis and interpretation of data presented in this manuscript. TWB, JRP, MKG and GWA designed the study. TWB, SS, and TAN performed the associated work and TWB, JRP, MKG and GWA drafted and revised the manuscript.

Funding

National Institutes of Health.

Conflicts of interest

The authors have no conflicts of interest to declare.

Ethics approval

All animal studies were conducted in accordance with the principles and procedures outlined in the NIH guide for the Care and Use of Laboratory Animals. The local Institutional Animal Care and Use Committee approved these procedures.