| Objective Ulcerative colitis(UC) is chronic, repeated relapsing colorectal mucosa continuity inflammatory disease, morbidity and mortality worldwide increased year by year, and of all ages have the disease. Treatment methods include hormones, immunosuppressive agents, biological agents and surgical treatment, but long-term efficacy and quality of life are not ideal, the fundamental reason lies in the pathogenesis is not clear. Currently, UC is considered a multifactorial, heterogeneous, multi-gene genetic diseases, a variety of factors including the result of the interaction of environmental, genetic, infection, immunity, intestinal microbes. In recent years, we found Epigenetics is the bridge between environmental and genetic factors, in which DNA methylation is an important epigenetic modification, it is possible to adjust the normal development of mammalian cells, differentiation, gene expression and chromatin structure. DNA methylation require methyl donor, including folic acid, choline, B vitamins and methionine. DNA hypermethylation is associated with gene silencing, and gene expression is active while DNA hypomethylation. It is found that increased expression of IFN-? is associated with reduced levels of gene methylation in intestinal T lymphocytes of UC patients, and IFN-? gene(IFNG) level of methylation in the regulation of mucosal cytokine secretion plays an important role. Evidence that during mammals development, environmental factors during pregnancy and the neonatal period result in permanent change in phenotype maybe by epigenetic modification. Thus, changing maternal dietary factors may involve in UC development by influencing DNA methylation. In this study, BALC/c mice for the study, female by administration of a methyl donor deficient diet, to build offspring mice colitis model, and folate, vitamin B12 and homocysteine levels of peripheral blood serum of offspring mice were detected. The expression of IFN-? in colonic mucosa of offspring mice was examined, and DNA methylation level in IFNG promoter was examined, to explore whether the maternal methyl donor deficiency promote the pathogenesis and development of UC by affecting DNA methylation.Methods 1 Animal model: One month before pregnancy, 7-week-old BALB/c adult female mice were fed with either standard diet(C, n=3) or methyl donor deficient diet(D, that is, without folic acid, choline, vitamin B12 and methionine, n=4), assigned diet was constantly maintained until the weaning of offspring, i.e. postnatal day 21; the pups were fed with the same diet as their mother until killing. After 23 days of age, Colitis was induced by administration of 2.5% dextran sulfate sodium(DSS) dissolved in water as drinking of model groups for five days. Offspring mice were grouped as follows: C/DSS- group, D/DSS- group, C/DSS+ group, D/DSS+ group, six mice in each group.2 Disease activity index of colitis(DAI): offspring mice were observed daily physical condition, body weight, stool, blood in the stool and degree of perianal erosion. The colitis score was calculated by assigning scores of these parameters resulting in the DAI. The score ranges from 0 to 4(total score), which represents the sum of scores for weight loss, stool consistency and rectal bleeding divided by three.3 The enzyme-linked immunosorbent assay(ELISA) was used to detect the folate, vitamin B12 and homocysteine(Hcy) levels of peripheral blood serum of offspring mice.4 Immunohistochemical staining method was used to detect the expression levels of the IFN-? in colon mucosa of offspring mice.5 Genomic DNA of colon tissue samples were extracted, and after bisulfite conversion using Sequenom Mass Array methylation testing methods detected methylation levels of Cp G island in IFNG promoter.Results1 Confirmation of methyl donor deficiency: compared with the standard diet group(C/DSS- and C/DSS+ group), methyl donor deficient diet significantly decreased the serum concentration of both folic acid(8.87±1.11 vs 11.34±0.31 nmol/L, respectively, P<0.01) and vitamin B12(409.2±56.27 vs 676.1±51.66 ng/L, respectively, P<0.01); and was accompanied by an increase in the serum concentration of homocysteine(8.45±0.35 vs 6.77±0.36 ?mol/L, respectively, P<0.01) in D/DSS- and D/DSS+ group.2 Impact of methyl donor deficiency on the DAI and histological scores: Maximum severity of colonic inflammation was reached at 5 days initiation after DSS treatment. The DAI was higher in the C/DSS+ group than in the C/DSS- group(P<0.05). Methyl donor deficient diet further aggravated the severity of colitis induced by DSS, as reflected by the dramatically higher DAI in the D/DSS+ group compared with C/DSS+(P<0.01). The DAI was no statistical difference in C/DSS- group and D/DSS- group. histology score was higher in D/DSS- group than C/DSS- group, As D/DSS+ group was significantly higher than C/DSS+ group, thus also showed methyl donor deficient diet further aggravated the severity of experimental colitis.3 Expression level of IFN-? in colonic mucosa tissue: Compared with C/DSS- group and D/DSS- group, the expression level of IFN-? were significantly higher in C/DSS+ group and D/DSS+ group(c2=14.875, P<0.01) and especially D/DSS+ group increased more significantly than C/DSS+ group(P<0.01).4 Correlation of IFN-? expression levels and DAI: Expression level of IFN-? was positively correlated with DAI in C/DSS+ group and D/DSS+ group(r=0.853 and 0.840, both P<0.05).5 Methylation level of IFNG promoter Cp G islands: methylation level of same Cp G sites between the four samples found no significant differences, and total methylation levels of all Cp G sites between the four groups also found no significant differences.Conclusions1 Maternal methyl donor deficient diet aggravated experimental colitis in offspring mice.2 Methyl donor deficient diet can cause increased expression of IFN-?, which is related with experimental colitis.3 Methyl donor deficient diet caused increased expression of IFN-? and aggravated experimental colitis, not by hypomethylation of IFNG promoter Cp G islands. |
PDF Full Text Request