Unknown,Transcriptomics,Genomics,Proteomics

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Searching genes responsible for developmental origins of health and disease (DOHaD) by comparing adult and fetal mouse undernutrition models

ABSTRACT: A large number of rodent studies have supported the developmental origins of health and disease (DOHaD) hypothesis that interauterine undernutrition (IU) is a risk factor for non-communicable diseases. The effect of IU is considered to be induced thorough epigenetic programming in the fetal tissues. We have recently carried out global transcriptome expression and promoter DNA methylation analyses on the fetal mouse liver following maternal 50% food restriction (FR) during late gestation, and reported a list of fetal liver genes that were transcriptionally or epigenetically regulated by IU (Ogawa et al., 2014, Congenital Anomalies). We have already reported the genes that were regulated oppositely between maternal and fetal livers (Ogawa et al., 2014, Congenital Anomalies). Here, we considered that the fetal liver is a hematopoietic organ, and exposed nine-week-old male mice (C57BL/6J from Japan SLC, Hamamatsu, Japan; n=8 / group) to 50% FR (chow; CE-2, CLEA, Tokyo Japan) for two weeks and carried out global gene expression analysis on the whole blood by a method reported previously (Ogawa et al., 2014, Congenital Anomalies). Male C57BL/6J mice at 9 weeks (Japan SLC, Hamamatsu, Japan ; n=8 / group) were exposed to 50% FR (chow; CE-2, CLEA, Tokyo Japan) for 2 weeks. After 2 weeks, the mice were anesthetized with somunopentylÂ®(Kyoritsu eiyaku, Tokyo) at a dose of 0.5 mL/kg, and blood was taken from the right ventricle of the heart. Blood samples were immediately immersed in liquid nitrogen and stored at -80Â°C. All animal care and experimental procedures were approved by the Institutional Animal Care and Use Committee of Showa University, and caried out at animal facilities in Showa University. DNA Microarray analysis was carried out on 3 pools, respectively. Pool 1 included samples from animal No. 1-8 in each group (control and 50% FR). Pool 2 included samples from animal No. 1-4 in each group. Pool 3 included samples from animal No. 5-8 in each group.

ORGANISM(S): Mus musculus

SUBMITTER: RANDEEP RAKWAL

PROVIDER: E-GEOD-78817 | biostudies-arrayexpress |

REPOSITORIES: biostudies-arrayexpress

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Project description:Non-optimal fetal environments resulting in low birth weight have epidemiologically been associated with adult diseases. In animal models, maternal undernutrition has successfully demonstrated increased risks for adult diseases. In the present study, we treated pregnant mice with 50% food restriction (FR), and performed global gene expression and promotor DNA methylation profiling on the fetal livers. Considering that effects of food restriction is opposite between before and after birth, we further searched genes which are regulated oppositely against adult calorie restriction and commonly against aging. Searched genes were included in groups related to the immune system, obesity and heart disease. Among these genes, trib1 has already been demonstrated to contribute to an increased risk of cardiovascular disease. The present result suggests that trib1 is a target of DOHaD. In addition, lepr was also dow-regulaed by maternal FR, suggested a potential role of this gene in induction of obesity. Promotor DNA methylation profiling as well as gene expression profiling revealed glucocorticoid target genes were regulated by maternal FR, supported previous reports suggesting important role of glucocorticoid in DOHaD. C57BL/6J mice were purchased from Japan SLC Co. Ltd. (Hamamatsu, Japan). The animals were housed at the Animal Institution in Showa University. They were maintained in cages in a ventilated animal room with controlled temperature and relative humidity with a 12 h light: 12 h dark schedule (lights turned on at 8:00 AM) and had access to chow (CE-2, CLEA Japan) and tap water ad libitum. As per the supplier’s (CLEA) information, CE-2 includes 51.4% nitrogen free extracts, 24.9% crude protein, 8.6% moisture, 6.7% crude ash, 4.6% crude fat, and 3.7% crude fiber. The physiological fuel value is 346.7 kcal per 100 g chow. All animal experiments were started after a period of acclimation of at least one week. Pregnant animals were obtained by housing females with males (1-2 females/male). The day the vaginal plug was observed was designated embryonic day 0 (ED0) and gestation day 0 (GD0). Pregnant mice were exposed to 50% food restriction (FR) from GD 10 to 17 and caesarean section was performed between 10:00-12:00 AM on GD18. Amount of CE-2 chow supplied FR group was calculated as 50% of CE-2 consumed by control group each gestation day. Control group was supplied with chow ad libitum. Pregnant mice were sacrificed by cervical dislocation, and the fetuses were taken out and anesthetized on ice cold phosphate buffered saline. The fetuses were dissected under a dissection microscope, and fetal tissues were carefully removed avoiding any other tissues contamination. The liver was collected from six mothers in each group. Fetal liver was collected from two male fetuses from each mother. The liver was also collected from mothers to compare with fetal one in gene expression analysis. The harvested livers were immediately immersed in liquid nitrogen (N2) and stored at -80ºC. In all combinations, total RNA (1000 ng) was labeled with either Cy3 or Cy5 dye using an Agilent Low RNA Input Fluorescent Linear Amplification Kit (Agilent Technologies Inc., CA, USA). Fluorescently labeled targets of control as well as treated samples were hybridized to the same microarray slide with 60-mer probes (4 x 44K (41,090 gene probes), mouse whole genome, Agilent). A flip labelling (dye-swap or reverse labelling with Cy3 and Cy5 dyes) procedure was followed to nullify the dye bias associated with unequal incorporation of the two Cy dyes into cDNA. Hybridization and wash processes were performed according to the manufacturer’s instructions, and hybridized microarrays were scanned using an Agilent Microarray scanner. For the detection of significantly differentially expressed genes between control and exercise samples each slide image was processed by Agilent Feature Extraction software (version 9.5.3.1).

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Searching genes responsible for developmental origins of health and disease (DOHaD) by comparing adult and fetal mouse undernutrition models

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