摘要
为明确DNA甲基化是否参与LPS诱导的奶牛乳腺炎,本试验研究了DNA甲基化抑制剂对LPS诱导的奶牛乳腺上皮细胞炎症因子表达的影响。首先用浓度为0μg/mL、1μg/mL、10μg/mL和100μg/mL的LPS分别处理奶牛乳腺上皮细胞6 h、12 h、24 h,荧光定量PCR(qRT-PCR)检测白介素-6(IL-6)、牛防御素(BNBD-5)和肿瘤坏死因子(TNF-α)的mRNA表达水平,ELISA法测TNF-α的浓度,随后用浓度为1.0μmol/L、2.5 umol/L和5.0 umol/L的DNA甲基化转移酶抑制剂(5-Aza-2′-deoxycytidine,5-AZA)分别处理奶牛乳腺上皮细胞24 h、48 h、72 h,qRT-PCR检测TNF-α的mRNA表达水平。结果显示LPS处理后,IL-6、BNBD-5、TNF-α的mRNA表达水平和TNF-α的蛋白水平显著上升;5-AZA处理后的TNF-α的表达量相对于LPS处理组显著上升。表明DNA甲基化参与LPS诱导的奶牛乳腺上皮细胞TNF-α的表达上调。
This experiment’s aim was whether DNA methylation involve in the bovine mastitis by LPS, and DNA methyltransferase influence the expression of TNF-α by LPS in bovine mammary epithelial cells(BMEC). BMEC were treated with 0 μg/mL、 1 μg/mL、10 μg/mL and 100 μg/mL LPS and different time like 6 h、12 h and 24 h, we could get the mRNA expression of IL-6、BNBD-5 and TNF-α by the qRT-PCR and the protein expression level of TNF-α by the ELISA;BMEC were treated with 1.0 umol/L、2.5 μmol/L and 5.0 μmol/L 5-AZA and different time like 24 h、48 h and 72 h, we could get the expression of TNF-α by the qRT-PCR. The results showed that the expression of IL-6、BNBD-5 and TNF-α had significant increased after the cells treated by LPS and the model was founded;the expression of TNF-α had significant increased after the cells treated by the 5-AZA. In conclusion, the 5-AZA had the influences on the expression of TNF-α on the BMEC.
作者
安冬
王军
刘红羽
燕晓晓
吕文发
AN Dong;WANG Jun;LIU Hong-yu;YAN Xiao-xiao;LYU Wen-fa(College of Animal Science and Technology,Jilin Agricultural University,Changchun 130118,China)
出处
《中国兽医杂志》
CAS
北大核心
2018年第12期30-32,36,共4页
Chinese Journal of Veterinary Medicine
基金
吉林省重点科技攻关项目(20150204074NY)
