Effects Of Iodine On Methylation Of Thyroid BRAF Gene In Rats

Objective To study the effects of different iodine nutritional status on methylation of rat thyroid sarcoma virogena B1(BRAF)gene,and to explore whether iodine affects the expression of BRAF through BRAF gene methylation.Methods Forty female spf-grade SD rats were randomly divided into 5 groups,respectively called low iodine group(LI,n=8),moderate iodine group(NI,n=8),5 times high iodine group(5HI,n=8),10 times high iodine group(10HI,n=8),and 20 times high iodine group(20HI,n=8).The model was made by drinking deionized water with different iodine contents and feeding low-iodine feed.Collected 24 hours urine of rats during determination of urine iodine,put to death in rats after 3 months,to return blood specimens and thyroid tissue specimens,chemiluminescence method is used to determine three iodine rats serum free thyroid original glycine(FT3)and free thyroxine(FT4),thyroid stimulating hormone(TSH)level,the application of curing sequencing PCR(Bisulfite sequencing PCR,BSP)gene promoter region CPG island methylation analysis,determination of BRAF protein expression using immunohistochemical techniques.Results(1)The results of 24-hour urinary iodine level of rats showed that the median of urinary iodine in the 5 groups was positively correlated with the iodine intake,suggesting that the model was successful.However,compared with the normal iodine group,the median of urinary iodine in the 5,10 and 20 times high iodine groups did not reach the corresponding multiple difference.(2)The results of thyroid hormone level measurement showed that among the 5 groups of rats,serum FT3 was the highest in the normal iodine group,but the serum FT3 concentration was lower in the low iodine group and the high iodine group,and the difference was statistically significant(P < 0.05).However,there was no significant difference in serum FT3 level between the low-iodine group and the high-iodine group.Compared with the appropriate iodine group,serum FT4 was slightly higher in the low-iodine group and the high-iodine group,but the difference was not statistically significant(P > 0.05).There was no significant difference in serum TSH between the low iodine group,the normal iodine group and the high iodine group(P > 0.05).(3)The methylation results showed that the BRAF methylation rate of rats in the five gr-oups was not statistically significant(P >0.05).(4)Immunohistochemical results showed that the differences in AREA,cumulative optical density(IOD)and optical density between the five groups showed that the expression of BRAF in thyroid tissues of the low-iodine group,the normal-iodine group and the high-iodine group increased gradually,and the differences were statistically significant(P < 0.05).By pair-wise comparison(LSD method),thyroid BRAF expression was lower in the low-iodine group and higher in the high-iodine group than in the normal iodine group,and the difference was statistically significant(P < 0.05).Compared with the group with 5 times higher iodine content,the expression in the group with 10 times higher iodine content and the group with 20 times higher iodine content was higher than that in the group with 5 times higher iodine content,and the difference was statistically significant(P < 0.05).Conclusion(1)Iodine deficiency or excess iodine can cause iodine nutritional disorders,and different iodine intake can affect thyroid hormone levels in rats.(2)Iodine affects the expression of thyroid BRAF protein in rats,but has no significant effect on the methylation of BRAF gene,suggesting that iodine may affect the expressi-on of BRAF protein by other means.