Global comparison of the colonic gene expression profiles between 14-month-old senescenceaccelerated mouse (SAM)-P6 mice and SAM-R1 mice, a wild-type control, was conducted with an oligonucleotide microarray containing more than 5,000 mouse genes. Eight genes were upregulated more than two-fold and 94 genes were downregulated more than two-fold in SAM-P6 mice. The three cell defense genes intelectin1 (Itln1), trefoil factor 3 (intestinal) (Tff3) and “deleted in malignant brain tumors 1” (Dmbt1) were among those extensively downregulated. Quantitative RT-PCR analysis confirmed that Itln1 mRNA was almost undetectable in SAM-P6 colon, whereas it was readily detected in SAM-R1 colon. Colonic expression of both Tff3 and Dmbt1 mRNA was also substantially decreased, to one third and two thirds of the levels in SAM-R1 mice, respectively. A 14 kDa Tff3 dimer was detected by Western blotting in the colon of all three SAM-R1 mice, but was not present in three SAM-P6 mice. No upregulation of 3 cell defense genes was detected in 3-month-old SAM-R1 as well as SAM-P6 mice. These results suggest that a diminution of the intestinal trefoil factor system may be involved in the acceleration of aging in SAM-P6 mice.

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Intracellular hydrodynamics is of considerable importance to regulate cellular functions. Using Raman microspectroscopy and imaging, we visualized the time-dependent changes of water concentration in single human acute promyelocytic leukemia HL-60 cells, and also showed the labelfree biomedical imaging of water molecules, lipids and proteins for analyzing molecular distribution. Taken together, our results demonstrated that we can distinguish between intracellular water and other major cell components, and that when the physiological status of cells changes with time, the Raman spectral intensities for intracellular water concentration also changes. Therefore, this biospectroscopic analysis of intracellular hydrodynamics can provide biomarkers to examine activities in dynamic living systems, and has significant potential in determining cell specificity for novel therapeutic approaches.

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Rat hepatoma H4IIE cells were stimulated with dexamethasone and dibutyryl cAMP to increase gene expressions of gluconeogenic enzymes, glucose-6-phosphatase (G6Pase) and phosphoenolpyruvate carboxykinase (PEPCK). Inclusion of catechin-rich green tea beverage (GTB) in the culture medium reduced the up-regulation of these genes as well as that of hepatocyte nuclear factor 4 alpha (HNF4α) gene. GTB was fractionated into chloroform-soluble (Fraction I), ethyl acetatesoluble (Fraction II), methanol-soluble (Fraction III) and residual (Fraction IV) fractions. Fractions II and III containing catechins caused an attenuation of the up-regulated expression of these genes as well as the down-regulation of HNF4α gene expression. Fraction IV had a synergistic effect on the up-regulation by dexamethasone/dibutyryl cAMP of the PEPCK gene expression and upregulated HNF4α gene expression. These results suggest that GTB down-regulated the expression of the HNF4α gene to cause the down-regulated gene expression of gluconeogenic enzymes. One reason why GTB did not down-regulate hepatic PEPCK gene expression in previous animal experiments may be that the component(s) acting to up-regulate PEPCK gene expression was more effective in vivo than in cultured cells.

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Lactic acid bacteria, either alive or dead, can improve villous atrophy caused by weaning in both piglets and mice. In this experiment, we tried to detect the molecules involved in this phenomenon with a real-time RT-PCR array approach. Weaning pups of mice were administered either a suspension of an Enterococcus faecalis EC-12 dried cell preparation (EC-12) or saline for 11 consecutive days after weaning. The jejunal and ileal villous heights were measured histologically, and the expression levels of 86 genes were analyzed for the jejunal and ileal epithelial cells and the lamina propria (LP). EC-12 induced significantly higher villous height in the jejunum and the ileum. Interleukin (IL)-6, fibroblast growth factor (FGF)-7, -10, and -22, and the platelet-derived growth factor (PDGF) β in the jejunal and the ileal LP were the most enhanced genes by EC-12. The possible role of these molecules in the improvement of villous atrophy is discussed.

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We reported previously that the forced expression of the chemokine BRAK/CXCL14 in head and neck squamous cell carcinoma cells decreased the rate of tumor formation and size of tumor xenografts in athymic nude mice and SCID mice. In order to clarify the expression of BRAK/CXCL14 affected either the settlement of carcinoma cells in host tissues in vivo or proliferation of the colonized carcinoma cells or both, we prepared oral floor carcinoma-derived HSC-2 cells in which BRAK/CXCL14 expression was induced upon doxycycline treatment. Then 30 nude mice were separated into 3 groups composed of 10 mice per group: Group I, the control, in which the engineered cells were directly xenografted onto the back of the mice; Group II, the cells were xenografted and then the mice were treated with doxycycline; and Group III, the cells were pretreated with doxycycline during culture, and the host mice were also treated with the drug before and after xenografting. The effects of BRAK/CXCL14 expression were examined by measuring the tumor size. The order of the size of tumor xenografts was Group I > II > III, even though the growth rate of the engineered cells was the same whether or not the cells were cultured in the presence of the drug. In addition, the size of tumors was significantly down-regulated after xenografting the doxycycline-pretreated cells in Group III. These data indicate that BRAK/CXCL14 expression in oral floor carcinoma cells reduced both the rate of settlement and the proliferation of the cells in vivo after settlement of the cells.

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DW/J-grt is a congenital hypothyroid mouse model that is characterized by growth retardation, significantly lowered T3 and T4 levels, and severe thyroid hypoplasia related to TSH hyporesponsiveness. Previously, we identified the point mutation of the Tpst2 gene in DW/J-grt mice that causes a decrease in the enzymatic activity, and demonstrated that the Tpst2 transgene rescues the mutant phenotypes both in vitro and in vivo. The severity of hypothyroidism is highly variable indicating the influence of modifier genes in humans. In this study, to identify the modifier/resistant gene(s) to hypothyroidism, we produced congenic strains carrying this Tpst2^grt mutation on the C57BL/6J and 129/SvJcl (129) genetic backgrounds and analyzed growth rate and thyroid function. Interestingly, the 129 congenic mice exhibited normal growth and thyroid function. The result suggests that 129 strain has the modifier(s) of attenuation of hypothyroidism. Therefore, the identification of the modifier loci of 129 mice will provide important new information about the gene(s) related to congenital hypothyroidism.

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