| Melamine is typical three triazine heterocyclic compound.It is a kind of chemical raw material which is widely used in plastic and fertilizer industry.Cyanuric acid is a product of melamine hydrolysis by continuous deamination.Researches of melamine and its combination with cyanuric acid were mainly focused on kidney toxicity,liver toxicity,reproductive toxicity and genetic toxicity on organs and tissues.However,there were only a few studies of their toxic effects on the immune system.In addition,studies on natural antidotes to the toxicity of melamine and cyanuric acid had become a research hotspot recently.The purpose of this study was to investigate the effects of melamine and its combination with cyanuric acid on the immune fuction of male mice,and explore the effects of tea polyphenols,Vitamin C and Vitamin E as natural antidotes.TEST ONE The regulative effect of tea polyphenols,Vitamin C and Vitamin E on effect of melamine and its combination with cyanuric acid on the B cells of mice.Methods:56 male Kunming mice were randomly divided into seven groups:melamine group(30mg.kg-1),melamine+tea polyphenols group(30+400mg.kg-1),melamine+Vitamin C and Vitamin E group(30+215/286mg.kg-1),melamin+cyanuric acid combination group(15+15mg.kg-1),combination+tea polyphenols group(15+15+400mg.kg-1),combination+Vitamin C and Vitamin E group(15+15+215/286 mg.kg-1)and control group.All intragastric administrations were made every other day.After 28 days,the mices were injected with 1%sheep red blood cells to stimulate the body's immune response.Then blood samples were taken from the eyeballs of mice,and all mice were sacrificed by cervical dislocation.The expressive rates of surface marker CD138+ of plasma cells in the peripheral blood and peripheral blood samples were detected respectly by flow cytometer and automatic hematology analyzer.The spleen tissues were collected immediately under sterile conditions.RT-PCR method was used to detect the relative expression level of Blimp-1 mRNA and the relative expression of this protein by Western Blot.Result:? Compared with the control group,the number of white blood cells was significantly increased in the combination group(P<0.05),combination+tea polyphenols group and combination+Vitamin C and Vitamin E group.However,the number of white blood cells in the peripheral blood of mice from combination+tea polyphenols group was significantly lower than the combination group(P<0.05).?From the number of lymphocytes of mice,combination group and combination+Vitamin C and Vitamin E group were significantly increased compared with the control group(P<0.05),while there was no significant difference in combination+tea polyphenols group(P>0.05).? Compared with control group,the expression rate of CD 13 8+ very significantly reduced in the melamine group and the combination group of melamine and cyanuric acid(P<0.01).However,there were no significant differences in the expression rates of CD 13 8+ in melamine+tea polyphenols,melamine+Vitamin C and Vitamin E group,combination+tea polyphenols,combination+Vitamin C and Vitamin E(P>0.05).?From the relative expression of Blimp-1 protein,melamine+cyanuric acid combination group compared with control group decreased highly significantly(P<0.01).Other experimental group compared with the control group had no significant difference(P>0.05).Compared with the control group,the relative expression of Blimp-1 mRNA did not change significantly in each group(P>0.05).The results show that these substances could hinder the maturation and differentiation of B lymphocytes by reducing the expression of Blimp-1 protein,and tea polyphenols,Vitamin C and Vitamin E could regulate this inhibition to a certain extent.TEST TWO The regulative effect of tea polyphenols,Vitamin C and Vitamin E on effect of melamine and its combination with cyanuric acid on the T cells of mice.Methods:The treatments of experimental animals were the same as test one.The flow cytometry was used to detect the CD3+,CD4+,CD8+ on the T cell surface markers and CD8a+CD44hiCD62Llow on the CTL cell surface markers in mice.RT-PCR and Western Blot methods were used to detect the relative gene expression of Notch-1 and the relative expression of this protein.Result:?The ratio of CD4+/CD8+ in the combination group of melamin+cyanuric acid was significantly lower than the control group(P<0.05).However,compared with the control group,CD4+/CD8+ ratios were no significangt difference in the combination+tea polyphenols group with combination+Vitamin C and Vitamin E group of mice(P>0.05).The decreasion of the ratio implies that the immune function of the body may be inhaibited to some extent.The CD4+/CD8+ ratio was restored by using the tea polyphenols,Vitamin C and Vitamin E.?There were no significant defference in the expression rate of CTL cells in the peripheral blood of the experimental group compared with the control group(P>0.05),but the values were lower than control group.?Compared with the control group,the combination group of melamin+cyanuric acid was very significantly increased in the Notch-1 gene relative expression(P<0.01),and there were no significant differences in the combination+tea polyphenols group with combination+Vitamin C and Vitamin E group(P>0.05).Compared with the combination group,the relative expression level of Notch-1 mRNA was significantly lower in the combination+tea polyphenols group(P<0.05).The results show that these substances could regulate the maturation and differentiation of T lymphocytes by increasing the relative expression of Notch-1 mRNA.But melamine and its combination with cyanuric acid effect on the body,Vitamin C and Vitamin E was significantly higher than the control group(P<0.05),while other groups was not significantly difference(P>0.05).This is not consistent with the results of mRNA,the reason for further study.The above studies indicate that tea polyphenols,Vitamin C and Vitamin E have a certain regulatory effects on the toxicity of these substances. |
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