镁硒对氟牙症小鼠牙胚釉原蛋白表达的影响

The antagonistic effect of magnesium-selenium against fluorosis on the expressions of amelogenin in dentalgerm of mice incisor

目的通过动物实验研究镁硒氟对氟牙症小鼠釉原蛋白表达的影响，为氟牙症的防治提供科学数据。方法80只雄性SPF级ICR小鼠，按体质量采用随机数字表法分为8组：对照组、加镁组、加硒组、镁硒组、加氟组、镁氟组、硒氟组、镁硒氟组，每组10只。对照组、加镁组、加硒组、镁硒组饮用双蒸水，加氟组、镁氟组、硒氟组、镁硒氟组饮用含氟（F-）50mg／L的双蒸水溶液。对照组和加氟组常规饲料喂养，加镁组和镁氟组用添加硫酸镁（MgsO4·7H2O，162．5ms／ks）的常规饲料喂养，加硒组和硒氟组用添加亚硒酸钠（Na2SeO3·5H2O，2．0mg／kg）的常规饲料喂养，镁硒组和镁硒氟组用添加MgSO4·7HsO（162．5mg／kg）和Na2SeO3·5H2O（2．0mg／kg）的常规饲料喂养。42d后处死小鼠，获取切牙标本。免疫组织化学染色观察釉原蛋白，并以灰度值表示结果，灰度值越大．蛋白表达越少。结果光镜下加氟组成釉细胞扭曲变形，细胞内有空泡；镁硒氟组成釉细胞与对照组接近。加氟组釉基质中釉原蛋白的表达（灰度值：131．03±11．14）明显高于其余各组（对照组：143．44±2．52，加镁组：143．73±12．43；加硒组：148．89±2．85；镁硒组：148．38±7．58；镁氟组：145．90±7．00；硒氟组：148．70±4．90；镁硒氟组：151．89±4．59。P均〈0．05）。加氟组成釉细胞内釉原蛋白的表达（165．49±5．66）明显低于对照组、加镁组、镁氟组、加硒组（151．35±2．52、149．27±11．13、146．21±4．84、150．39±6．65，P均〈0．05），硒氟组成釉细胞内釉原蛋白的表达（165．464-5．81）明显弱于加镁组、镁氟组、加硒组（P均〈0．05）。析因分析显示，镁、硒单独作用对釉原蛋白在基质中的表达有影响（F=4．195、15．009，P均〈0．05），氟与镁的交互作用对釉原蛋白在基质中的表达有影响（F=4．402，P〈0．05），镁、硒、氟三者的交互作用对釉原蛋白在基质中的表达无影响（F=1．561。P〉0．05）。氟、镁、硒单独作用对釉原蛋白在成釉细胞内的表达均有影响（F=18．463、9．372、4．741，P均〈0．05），氟与镁、镁与硒的交互作用对釉原蛋白在成釉细胞内的表达有影响（F=10．351、5．919，P均〈0．05），镁、硒、氟三者的交互作用对釉原蛋白在成釉细胞内的表达无影响（F=1．460，P〉0．05）。结论镁对氟中毒小鼠的釉原蛋白表达有拮抗作用，但尚不能认为硒对氟中毒小鼠的釉原蛋白表达有影响。

Objective To study the antagonistic effects of magnesium-selenium-fluorine preparation on dental fluorosis of mice and its mechanism, and to provide a foundational basis for prevention and control of dental fluorosis. Methods Eighty male SPF ICR mice, were divided into 8 groups according to body weight by random number table method： control group, magnesium group, selenium group, magnesium-selenium group, fluoride group, magnesium-fluorine group, selenium-fluorine group and magnesium-selenium-fluorine group. The control group, magnesium group, selenium group and magnesium-selenium group drank double steamed water, and the other four groups drank 50 mg/L F- double steamed water. The control group and fluoride group fed conventionally. Magnesium group and magnesium-fluorine group fed conventionally by adding MgSO4-7H20 162.5 mg/kg. Selenium group and selenium-fluorine group fed conventionally by adding Na2SeO3·5H2O 2.0 mg/kg. Magnesium-selenium group and magnesium-selenium-fluorine group fed conventionally by adding MgSO4·7H2O 162.5 mg/kg and Na2SeO3·5H2O 2.0 mg/kg. Then incisor specimens were obtained after the mice were put into death after treatment for 42 days. The expression of amelogenin was observed with immunohistochemical staining. And gray value is expressed as a result, the greater the gray value, the less protein expression. Results The results of light microscope showed that ameloblasts in the fluoride group showed disarrangement and even had vacuolar. The change of ameloblasts in the magnesium-selenium-fluorine group had no significant difference with control group. The expressions of amelogenin in enamel matrix of fluoride group （131.03 ± 11.14） was significantly higher than those of others （control group： 143.44 ± 2.52, magnesium group： 143.73 ± 12.43, selenium group： 148.89 ± 2.85, magnesium-selenium group： 148.38 ± 7.58, magnesium-fluorine group： 145.90 ± 7.00, selenium-fluorine group： 148.70 ±4.90, and magnesiumselenium-fluorine group： 151.89 ± 4.59, all P 〈 0.05）. The expression of amelogenin in ameloblast of fluoride group （165.49 ± 5.66） was significantly lower than those of control group, magnesium group, magnesium-fluorine group and selenium group （151.35 ± 2.52, 149.27 ± 11.13, 146.21 ± 4.84 and 150.39 ± 6.65, all P 〈 0.05）. The expression of amelogenin in ameloblast of selenium-fluorine group （165.46 ± 5.81） was significantly lower than those of magnesium group, magnesium-fluorine group and selenium group （all P 〈 0.05）. The results of factorial analysis showed that magnesium and selenium affected the expression of amelogenin in enamel matrix （F = 4.195, 15.009, all P 〈 0.05）, the interaction between fluoride and magnesium had an effect on the expression of amelogenin in enamel matrix （F = 4.402, P 〈 0.05）. Interaction of fluoride, magnesium and selenium had no effect on the expression of amelogenin in enamel matrix （F = 1.561, P 〉 0.05）. The fluoride, magnesium and selenium affected the expression of amelogenin in ameloblast （F = 18.463, 9.372, 4.741, all P 〈 0.05）, the interactions between fluoride and magnesium, magnesium and selenium had effects on the expression of amelogenin in ameloblast （F = 10.351, 5.919, all P 〈 0.05）. Interaction of fluoride, magnesium and selenium had no effect on the expression of amelogenin in ameloblast （F = 1.460, P 〉 0.05）. Conclusions There is an antagonistic effect of magnesium on the expression of enamel protein in fluorosis mice. However, it can not be considered that there is an effect of selenium on the expression of enamel protein in fluorosis mice.