Modified ain 93g

Male rats weighing 90 to 110 g (CLEA) were used in our experiments. The rat PH model was generated as described previously [12 (link),13 (link)]. In brief, 20 mg/kg of vascular endothelial growth factor inhibitor SU5416 (Abcam) suspended in CMC (0.5% [w/v] carboxymethylcellulose sodium, 0.9% [w/v] sodium chloride, 0.4% [v/v] polysorbate 80, and 0.9% [v/v] benzyl alcohol in deionized water) was subcutaneously injected at Day 0. The rats were housed in a hypoxic chamber (10% O₂) maintained using a hypoxic air generator (TEIJIN) and monitored with an O₂ analyzer (JIKO-255) for 3 weeks, and then re-exposed to normoxia for an additional 3 or 5 weeks. A purified AIN-93G containing 1,000 IU/kg of cholecalciferol (normal diet) and the modified AIN-93G containing 10,000 IU/kg of that (high vitamin D diet) were purchased from Oriental Yeast Co., Ltd (Japan). PH rats were divided into two groups and fed different dietary amounts of cholecalciferol on ad libitum.

This study was carried out in accordance with the recommendations of Regulations for Animal Experiments and Related Activities at Tohoku University, Center for Laboratory Animal Research, Tohoku University. The protocol was approved by the Center for Laboratory Animal Research, Tohoku University. Virgin female C57BL/6N mice (CLEA Japan, Inc., Tokyo, Japan) about 5 weeks old were maintained under controlled lighting (12-h light cycle) and temperature (24°C). These were divided into two diet groups: Normal (N) and Low Protein (LP) by feeding them AIN-93G and modified AIN-93G (Oriental Yeast Co., Ltd., Tokyo, Japan), respectively. They were allowed free access to food and water ad libitum. The LP diet consisted of only 48% of the protein and 94% of the calorie contents of the N diet as suggested in previous studies (Reeves et al., 1993 (link); Ito et al., 2011a (link),b (link)). After a 2-week acclimatization period, the female mice were time mated and inspected for vaginal plugs the following morning. Plug-positive females were then transferred to single cages and fed ad libitum. Food consumption and body weight were recorded every day. There was a total of 25 pregnant mice per diet group.