Analyzing the hippocampal gene network to determine the effects of coral calcium hydride on antioxidant ability in rat brain

In our previous study, we reported that a diet rich in antioxidants such as coral calcium hydride (CCH) increased the endogenous antioxidant ability in the hippocampus of rats. We conducted this study to test the hypothesis that diet supplementation with CCH would change the gene expression in rats and to understand how CCH enhances antioxidant ability. We used a DNA array to compare the expression levels in the hippocampus of rats fed with CCH for 2 weeks with those of rats fed a normal diet. Immune response-related genes were down-regulated, while nuclear respiratory factor 2 and aldehyde dehydrogenase 3A were up-regulated. Our findings about the changes in the mRNA levels of these genes well explain the physiological finding of enhanced antioxidant ability in rat brain. CCH was obtained from ICB, Ltd., Sendai, Japan, and coral calcium (CC) was purchased from Coralbio, Okinawa, Japan. Male Wistar rats were acquired from Kyudo, Co., Ltd. and maintained at the Experimental Animal Center of the University of Miyazaki at a controlled ambient temperature of 23 ± 1 °C and 50 ± 10% relative humidity. The Committee for Ethics on Animal Experiments, Faculty of Medicine, University of Miyazaki, Japan, reviewed and approved the experimental design. Six-week-old male Wistar rats (n = 8) were assigned to 2 groups: standard diet-fed group (CE-2, Clea Japan, Inc., Tokyo, Japan) and CCH-fed group. The CCH diet was standard CE-2 feed supplemented with 0.1% CCH powder. Inhibition of accelerated aging and an increase in the in vivo antioxidant ability was observed in SAM/P-8 mice fed a diet supplemented with 0.1% CCH. In accordance with these reports, the CCH concentration used in our study was set at 0.1%.The animals were killed by cervical dislocation at the age of 8 weeks. They were decapitated and the hippocampi were removed and rapidly frozen in liquid nitrogen. The hippocampi were then homogenized with a conventional rotor-stator homogenizer. Total RNA was then extracted from the tissues by using the RNeasy Lipid Tissue Mini Kit (Qiagen, Valencia, CA).