Phlebotomy-induced anemia alters hippocampal neurochemistry in neonatal mice

Abstract

Background Phlebotomy-induced anemia (PIA) is common in preterm infants. The hippocampus undergoes rapid differentiation during late fetal/early neonatal life and relies on adequate oxygen and iron to support oxidative metabolism necessary for development. Anemia shortchanges these two critical substrates, potentially altering hippocampal development and function. Methods PIA (hematocrit <25%) was induced in neonatal mice pups from postnatal day (P)3 to P14. Neurochemical concentrations in the hippocampus were determined using in vivo 1H NMR spectroscopy at 9.4T and compared with control animals at P14. Gene expression was assessed using qRT-PCR. Results PIA decreased brain iron concentration, increased hippocampal lactate and creatine concentrations, and decreased phosphoethanolamine (PE) concentration and the phosphocreatine/creatine ratio. Hippocampal transferrin receptor (Tfrc) gene expression was increased, while the expression of calcium/calmodulin-dependent protein kinase type II alpha (CamKIIα) was decreased in PIA mice. Conclusion This clinically relevant model of neonatal anemia alters hippocampal energy and phospholipid metabolism and gene expression during a critical developmental period. Low target hematocrits for preterm neonates in the NICU may have potential adverse neural implications.

Anemia induces gut inflammation and injury in an animal model of preterm infants

“…However, as many gut resident immune cells, including macrophages, do not circulate, sampling preterm infants for gut resident macrophage number and function, in addition to direct evaluation of intestine barrier activity during the development of anemia is not currently feasible. As a result, we decided to complement this study with a previously validated pre‐clinical model of PIA to directly evaluate the impact of anemia on intestine immunity and barrier function . However, even in this model, significant barriers exist, as the size and overall cellular availability of individual mouse pups limit the scope of analysis that can be accomplished in this setting.…”

“…Indeed, sample volume limitations in both clinical and animal studies of neonatal immune function have likely contributed to challenges associated with conducting studies that produce a level of understanding in neonates comparable to that obtained from similarly conducted studies in adult mice and patients clinically. Importantly, the rate of anemia employed in this study was designed to match the rate observed in many patients and does not induce detectable changes in heart rate or the development of acidosis that would be expected to occur in the setting of significant hemodynamic alterations/shock . While nearly all pups reached our threshold of severe anemia (25% Hct) on P7, followed by analysis of the potential impact of anemia the following day, future studies will be necessary to examine the effects of extended anemia exposure and threshold variations.…”

“…As a result, we decided to complement this study with a previously validated pre-clinical model of PIA to directly evaluate the impact of anemia on intestine immunity and barrier function. 19,20 However, even in this model, significant barriers exist, as the size and overall cellular availability of individual mouse pups limit the scope of analysis that can be accomplished in this setting. Indeed, sample volume limitations in both clinical and animal studies of neonatal immune function have likely contributed to challenges associated with conducting studies that produce a level of understanding in neonates comparable to that obtained from similarly conducted studies in adult mice and patients clinically.…”

“…Importantly, the rate of anemia employed in this study was designed to match the rate observed in many patients and does not induce detectable changes in heart rate or the development of acidosis that would be expected to occur in the setting of significant hemodynamic alterations/ shock. 19,20 While nearly all pups reached our threshold of severe anemia (25% Hct) on P7, followed by analysis of the potential impact of anemia the following day, future studies will be necessary to examine the effects of extended anemia exposure and threshold variations. Furthermore, it is not clear whether the lack of ability of anemia to induce NEC in this model accurately reflects what actually occurs clinically.…”

“…Anemia was induced by twice-daily phlebotomy (5.25 μL/g), from day three after birth (postnatal day 3 (P3)) to obtain and maintain the desired hematocrit of <25% as outlined previously. 19,20 Graduated glass microhematocrit tubes were used to measure blood draws and hematocrit. Control pups were pricked through the neck scruff without bleeding.…”