Transcriptome Profiling Analysis of Kidneys and Brain in Dehydrated Infant Mice

Abstract

Water is an essential nutrient as a structural and functional component in the body. To maintain a healthy life optimal hydration status is essential and suffi-cient water intake is recommended. Compared to adults, infants have higher body water composition ratio, reaching 75%, and immature fluid regulatory system, which leads to higher risk of dehydration. In spite of the importance of water, there is few evidence underlying the impacts of dehydration on physical and cognitive development. We, therefore, investigated the molecular mecha-nisms by which dehydration draws physiological changes in infancy. 3-week-old infant mice that just finished weaning were provided limited access to a water bottle for fifteen minutes per day for one week (RES 1W) and four weeks (RES 4W) to induce dehydration status while the control group con-sumed water ad libitum with free access to the water bottle for a week (CON 1W) and 4 weeks (CON 4W). Compared with control mice, mice in the RES group consumed 2.4 times less water. The four-week dehydration resulted in increased plasma osmolality and the transcript level of vasopressin in the brain compared to the control group. Transcriptome analysis was conducted to un-derstand physiological changes in the kidney induced by dehydration. We found that structural and functional kidney development was still ongoing in early life by analyzing the renal transcriptional networks of infant (4 weeks old

CON 1W) and juvenile (7 weeks old

CON 4W) mice. Kidneys in 4 week and 7 week old mice showed significantly distinctive functional gene networks. Gene sets related to cell cycle regulators, fetal kidney patterning molecules and immature glomerular barrier integrity were upregulated in infantile kidneys while heightened expressions of genes associated with ion transport and drug metabolism were observed in juvenile kidneys. Western blot analysis was con-ducted to validate the protein levels of representative molecules in morpholog-ical and functional maturation in the kidney. GPC3 that mediates ureteric bud branching in the metanephros was slightly higher in infantile kidneys at the protein level compared to the juvenile kidneys. CYP4A that is one of the most critical proteins in xenobiotics and lipid metabolism had increased protein lev-els in juvenile kidneys with normal renal development. Dehydration during infancy suppressed renal growth by interrupting the SHH signaling pathway, which targets cell cycle regulators. Importantly, disruption of the developmen-tal program ultimately led to long-term alterations in renal filtration functions by causing a decline in glomerular filtration barrier integrity. We also investi-gated whether dehydration affects cognitive function. Dehydration from infan-cy to juvenile resulted in impairment of learning ability. In the Barnes maze test, dehydrated mice (RES 4W) showed 4 times longer latency to find a target hole after 4 days of training compared to control mice (CON 4W). Brain weight was determined at the end of experiment and there was no significant difference between control and dehydrated mice. A series of neurotrophic factor was ex-amined for their expression and we found only BDNF expression was signifi-cantly lower in RES 4W than CON 4W. The transcript level of BDNF only in hippocampus of RES 4W was significantly different from CON 4W. Next-generation RNA sequencing analysis in the brain revealed that CREB and its downstream target genes related to memory consolidation were significantly dysregulated by dehydration. These results suggest that sufficient water intake is required for normal renal growth and learning development during infancy. This study provides a meaningful perspective of pediatric dehydration with a molecular and physiological explanation of why infants are more vulnerable to dehydration than adults. Furthermore, these findings suggest new insights into the systemic effects of dehydration on development and may provide possible markers for clinical application in pediatric dehydration.