不同类型氟骨症大鼠骨组织5-甲基胞嘧啶与5-羟甲基胞嘧啶的表达  

Expression of 5-methylcytosine and 5-hydroxymethylcytosine in bone tissue of rats with different types of skeletal fluorosis

作　　者：周瑞琦 丁宏伟 杨梦兰 罗鹏[1] 潘雪莉[1] Zhou Ruiqi;Ding Hongwei;Yang Menglan;Luo Peng;Pan Xueli(School of Public Health,the Key Laboratory for Environmental Pollution Monitoring and Disease Control,Ministry of Education,Guizhou Medical University,Guiyang 550025,China)

机构地区：[1]贵州医科大学公共卫生与健康学院、环境污染与疾病监控教育部重点实验室,贵阳550025

出　　处：《中华地方病学杂志》2023年第2期92-98,共7页Chinese Journal of Endemiology

基　　金：国家自然科学基金(81960575)。

摘　　要：目的了解不同类型氟骨症大鼠骨组织5-甲基胞嘧啶(5-mC)、5-羟甲基胞嘧啶(5-hmC)水平,并分析其相关性。方法选取4周龄SPF级健康SD大鼠30只,适应性喂养1周后,采用随机数字表法将大鼠按体重(100~120 g)分为对照组(4 ml·kg^(-1)·bw去离子水灌胃+标准维持饲料)、骨硬化组[20 mg·kg^(-1)·bw氟化钠(NaF)灌胃+标准维持饲料]、骨质疏松/骨软化组(20 mg·kg^(-1)·bw NaF灌胃+低钙低蛋白偏食饲料),每组10只,雌雄各半;每周灌胃6 d,实验周期为5个月。实验结束后采集大鼠心脏血、下肢股骨样本。采用酶联免疫吸附试验(ELISA)法检测血清甲基供体S-腺苷甲硫氨酸(SAM)及其代谢产物S-腺苷同型半胱氨酸(SAH)含量,骨组织5-mC、5-hmC水平;蛋白免疫印迹(Western blot)法检测大鼠骨组织DNA甲基转移酶(DNMTs),DNA羟甲基化酶(TETs)蛋白表达情况;并分析各组大鼠血清SAM含量、SAM/SAH比值与骨组织5-mC水平以及骨组织5-mC与5-hmC水平的相关性。结果对照组、骨硬化组、骨质疏松/骨软化组大鼠血清SAM[11.03(7.06,18.63)、3.96(2.32,9.09)、3.91(2.35,4.46)nmol/L]、SAH含量[(4.69±0.55)、(5.41±1.13)、(13.90±1.09)ng/L],SAM/SAH比值[2.58(1.54,4.12)、0.62(0.52,1.69)、0.14(0.13,0.15)]及骨组织5-mC[103.39(97.37,109.35)、52.50(50.19,68.13)、55.03(49.97,59.57)ng/L]、5-hmC水平[(32.61±8.84)、(56.96±8.48)、(20.34±6.22)ng/L]比较,差异均有统计学意义(H/F=12.81、284.24、21.85、19.37、55.23,均P<0.01)。与对照组比较,骨硬化组、骨质疏松/骨软化组血清SAM含量、SAM/SAH比值、骨组织5-mC水平及骨质疏松/骨软化组骨组织5-hmC水平均较低,而骨质疏松/骨软化组血清SAH含量和骨硬化组骨组织5-hmC水平均较高(均P<0.05);与骨硬化组比较,骨质疏松/骨软化组血清SAH含量较高,而SAM/SAH比值、骨组织5-hmC水平均较低(均P<0.05)。Western blot检测结果显示,对照组、骨硬化组、骨质疏松/骨软化组大鼠骨组织DNMT1、DNMT3A、Objective To learn about the levels of 5-methylcytosine(5-mC)and 5-hydroxymethylcytosine(5-hmC)in bone tissue of rats with different types of skeletal fluorosis and analyze their correlation.Methods Thirty 4-week-old SPF grade healthy SD rats were selected.After adaptive feeding for 1 week,the rats were divided into control group(4 ml·kg^(-1)·bw deionized water+standard maintenance diet),osteosclerosis group[20 mg·kg^(-1)·bw sodium fluoride(NaF)+standard maintenance diet],and osteoporosis/osteomalacia group(20 mg·kg^(-1)·bw NaF+low-calcium and low-protein partial diet)according to their body weight(100-120 g)by random number table method,with 10 rats in each group,half male and half female;gavaged 6 days each week and the experimental period was 5 months.At the end of the experiment,samples of rat heart blood and lower limb femur were collected.The contents of serum methyl donor S-adenosylmethionine(SAM)and its metabolite S-adenosylhomocysteine(SAH)in serum,and the levels of 5-mC and 5-hmC in bone tissue were measured by enzyme-linked immunosorbent assay(ELISA).Western blot was used to determine the expression of DNA methyltransferase(DNMTs)and DNA hydroxymethylase(TETs)in bone tissue of rats.The correlation between serum SAM content,SAM/SAH ratio and bone tissue 5-mC level,and between the bone tissue 5-mC level and 5-hmC level was analyzed.Results Serum SAM[11.03(7.06,18.63),3.96(2.32,9.09),3.91(2.35,4.46)nmol/L],SAH content[(4.69±0.55),(5.41±1.13),(13.90±1.09)ng/L],SAM/SAH ratio[2.58(1.54,4.12),0.62(0.52,1.69),0.14(0.13,0.15)]and bone tissue 5-mC[103.39(97.37,109.35),52.50(50.19,68.13),55.03(49.97,59.57)ng/L],5-hmC levels[(32.61±8.84),(56.96±8.48),(20.34±6.22)ng/L]in the control group,osteosclerosis group and osteoporosis/osteomalacia group were compared,and the differences were statistically significant beween three groups(H/F=12.81,284.24,21.85,19.37,55.23,P<0.01).Compared with the control group,the content of SAM,the ratio of SAM/SAH,the level of 5-mC in the osteosclerosis group and osteoporosis/

关 键 词：氟 DNA甲基化 DNA羟甲基化 

分 类 号：R599.1[医药卫生—内科学]