| Budget Amount *help |
¥14,400,000 (Direct Cost: ¥14,400,000)
Fiscal Year 2006: ¥1,300,000 (Direct Cost: ¥1,300,000)
Fiscal Year 2005: ¥1,900,000 (Direct Cost: ¥1,900,000)
Fiscal Year 2004: ¥11,200,000 (Direct Cost: ¥11,200,000)
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| Research Abstract |
Back ground: Helper T cells can be classified into Th1 and Th2 subtype. The former cells have been shown to contribute to cellular immunity, the latter to humoral immunity. Based on the theory, impairment of balancing Th1 and Th2 cells is thought to cause immune diseases, including allergic diseases. Th1 and Th2 cells are polarized from naive T cells by getting in contact with mature dendritic cells (DCs), which are antigen presenting cells resident in front line of immunity. Only DCs can activate naive T cells. When DCs secrete IL-12, naive T cells become Th1 cells, while when they secrete IL-4, they become Th2 cells. Therefore, it is very important what kind of cytokines DCs secrete.
It has been previously demonstrated that DCs generated from code blood secrete less amounts of IL-12 compared to those from adult peripheral blood. The authors suggested that this is correlated with the immune status of infants, who have weak cellular immunity and develop allergic diseases easily.
Objectiv
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e : We hypothesized that less secreted IL-12 by DCs from both allergy group and cord blood would be due to methylation of the promoter region of IL-12 gene, and that allergic status might be attributed to delay of demethylation of the region. To elucidate it, we examined the methylation status in the promoter region of IL-12 gene of monocytes, which are precursors of DCs, prepared from peripheral blood from healthy adults (HA) and allergic adults (AA) and from cord blood from normal infants (NI) group.
Methods : Monocytes were purified by using MACS system with an antibody for CD 14 from each of three groups. Genomic DNAs were extracted. Methylation status in the promoter region of IL-12p35 gene was investigated with bisulphate-sequencing analysis method. Real time RT-PCR reactions were performed to determine IL-12p35 mRNA level in cultured and LPS stimulated monocytes of peripheral blood from HA and AA group.
Results : Of 7 CpGs at -388, -385, -375,-352,-331, -326, -322 position prior to start codon, there were significant differences in methylated ratio among three groups. Contrary to our expectations, methylated ratios were significantly lower in AA and NI groups than in HA group. There was no difference in IL-12p35 mRNA level between HA and AA groups.
Conclusion : These results suggest that hypo-methylation in the promoter region might be associated with suppression of gene expression of IL-12p35 in allergic patients and normal infants. Less
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