341 - Prophylactic and treatment EPO downregulate pro-inflammatory genes upregulated by phlebotomy-induced anemia in neonatal mice

Poster Number: 341
Publication Number: 341.122

Garima Singh, University of Minnesota Medical School, Minneapolis, MN, United States; Michael K. Georgieff, University of Minnesota, Minneapolis, MN, United States; Tate A. Gisslen, University of Minnesota, Edina, MN, United States

Background: Anemia in hospitalized preterm infants, often exacerbated by repeated phlebotomy, is common and associated with long-term neurodevelopmental deficits. Phlebotomy-induced anemia (PIA) in a preclinical mouse model causes dose-dependent upregulation of pro-inflammatory genes in the hippocampus, a region critical to normal learning and memory development. Treatment and prevention of anemia with recombinant erythropoietin (EPO) is frequently used for preterm infants. How prophylactic and treatment EPO affects inflammatory gene expression upregulated by PIA in the hippocampus is not known.

Objective: Determine the effect of prophylactic and treatment EPO on hippocampal pro-inflammatory gene expression in neonatal mice at 18% PIA.

Design/Methods: Mice were phlebotomized by twice-daily facial venipuncture starting at postnatal day (P) 3 to target hematocrits (hct) of 18%. Thereafter, pups were phlebotomized once daily to maintain hematocrit levels until P14. A subset of pups received prophylactic EPO starting on P3 while a second subset of pups was treated with rescue EPO once pups reached a hct of 18%. Hippocampal tissue was collected at P14 to prepare RNA and cDNA. Gene expression of was determined by PCR (n=10-12/group) assayed in duplicate and normalized against the reference gene S18.

Results: Expression of genes associated with inflammation, TNFa, CCL2, CXCL10, and TLR4 were upregulated in the hippocampus by 18% PIA. These genes plus Stat6 and TLR4 we downregulated in comparison to 18% PIA to levels similar to controls when given either prophylactic or rescue EPO. The NFkB gene RelA and AIF were downregulated compared to 18% PIA by prophylactic EPO only. IL-6 expression was upregulated after prophylactic and treatment EPO compared to controls and upregulated after treatment EPO compared to both 18% PIA and prophylactic EPO groups.Conclusion(s): Prophylactic or treatment EPO normalizes expression of hippocampal pro-inflammatory genes that were upregulated by 18% PIA. Anemia and its treatment with EPO both affect immune regulation in the hippocampus.