【目的】肉牛肌内脂肪沉积与牛肉的风味、多汁性和嫩度密切相关。脂肪沉积过程表现为脂肪细胞的增殖(数量增多)和分化(脂质生成),受到了多基因协同调控。前人研究发现,小鼠中Snail1可以参与肌肉发育和脂质稳态调控,但其在牛脂肪生成过...【目的】肉牛肌内脂肪沉积与牛肉的风味、多汁性和嫩度密切相关。脂肪沉积过程表现为脂肪细胞的增殖(数量增多)和分化(脂质生成),受到了多基因协同调控。前人研究发现,小鼠中Snail1可以参与肌肉发育和脂质稳态调控,但其在牛脂肪生成过程中的作用仍未知,有待进一步研究。【方法】以秦川牛为研究对象,克隆得到Snail1 CDS区序列,构建Snail1时空表达谱,运用生物信息学软件对其功能结构及靶基因进行预测。进一步,通过RNAi干扰结合CCK8、EdU、细胞流式及实时荧光定量PCR等方法探究Snail1对牛脂肪细胞增殖的影响。【结果】秦川牛Snail1与NCBI公布序列相比存在2处碱基同义突变,其在秦川牛新生牛肺、肾周脂肪、小肠呈现较高丰度表达;而在成年牛中,Snail1在肾周脂肪组织中的表达量最高,背最长肌中的表达量次之,肺脏组织中的表达量最低。生物信息学分析发现,Snail1启动子区存在1个651 bp CpG岛及C/EBP、PPARα等与脂肪生成相关的转录因子结合位点。CKⅠ(Ser92/96)、CKⅡ(Ser25/119,Thr89)、CDK1(Ser13/104/112/119/143/183/214/221)、CDK5(Ser105/107)等多个细胞周期相关激酶可能参与了Snail1蛋白的磷酸化修饰。通过对牛已注释基因启动子区提取、靶基因预测及KEGG动态网络构建发现,成脂相关的MAPK、PI3K-Akt、mTOR等信号通路为Snail1参与脂肪生成相关的潜在节点信号通路。进一步,通过RNAi干扰试验对其功能研究表明,Snail1下调促进了牛前体脂肪细胞的增殖,增加了复制期阳性细胞的比例(P<0.01)且促进了G1/S细胞周期转换。RT-qPCR和Western-blot检测表明,干扰Snail1显著上调了促增殖调控基因CCNB1、CCND2、CDK2、CDK4(P<0.05)和蛋白的表达。【结论】Snail1在新生牛肾周脂肪及成年牛肾周脂肪和背最长肌中表达量相对较高。干扰Snail1促进了牛前体脂肪细胞的增殖、G1/S细胞周期转变和CCNB1、CCND2、CDK2、CDK4等增殖相关基因表达;CKⅠ、CKⅡ、CDK1/5等多个细胞周期相关激酶可能通过磷酸化修饰Snail1蛋白进而参与细胞增殖调控,而MAPK、PI3K-Akt、mTOR等为Snail1影响牛脂肪细胞增殖潜在的关键节点通路。研究结果为进一步探究Snail1参与牛脂肪生成作用机制奠定了基础。展开更多
Background The intestinal epithelium performs essential physiological functions,such as nutrient absorption,and acts as a barrier to prevent the entry of harmful substances.Mycotoxins are prevalent contaminants found ...Background The intestinal epithelium performs essential physiological functions,such as nutrient absorption,and acts as a barrier to prevent the entry of harmful substances.Mycotoxins are prevalent contaminants found in ani-mal feed that exert harmful effects on the health of livestock.Zearalenone(ZEA)is produced by the Fusarium genus and induces gastrointestinal dysfunction and disrupts the health and immune system of animals.Here,we evaluated the molecular mechanisms that regulate the effects of ZEA on the porcine intestinal epithelium.Results Treatment of IPEC-J2 cells with ZEA decreased the expression of E-cadherin and increased the expression of Snai1 and Vimentin,which induced Snail1-mediated epithelial-to-mesenchymal transition(EMT).In addition,ZEA induces Snail-mediated EMT through the activation of TGF-βsignaling.The treatment of IPEC-J2 cells with atractyle-nolideⅢ,which were exposed to ZEA,alleviated EMT.Conclusions Our findings provide insights into the molecular mechanisms of ZEA toxicity in porcine intestinal epi-thelial cells and ways to mitigate it.展开更多
Objective This study aimed to investigate the role of the long noncoding RNA(lncRNA)maternally expressed gene 3(MEG3)in the epithelial-mesenchymal transition(EMT)of bladder cancer cells and the potential mechanisms.Me...Objective This study aimed to investigate the role of the long noncoding RNA(lncRNA)maternally expressed gene 3(MEG3)in the epithelial-mesenchymal transition(EMT)of bladder cancer cells and the potential mechanisms.Methods Cell invasion,migration,and wound healing assays were conducted to assess the effects of MEG3 on the invasive and migratory capabilities of bladder cancer cells.The expression levels of E-cadherin were measured using Western blotting,RT-qPCR,and dual luciferase reporter assays.RNA immunoprecipitation and pull-down assays were performed to investigate the interactions between MEG3 and its downstream targets.Results MEG3 suppressed the invasion and migration of bladder cancer cells and modulated the transcription of E-cadherin.The binding of MEG3 to the zinc finger region of the transcription factor Snail prevented its ability to transcriptionally repress E-cadherin.Additionally,MEG3 suppressed the phosphorylation of extracellular regulated protein kinase(ERK),c-Jun N-terminal kinase(JNK),and P38,thereby decreasing the expression of Snail and stimulating the expression of E-cadherin.Conclusion MEG3 plays a vital role in suppressing the EMT in bladder cancer cells,indicating its potential as a promising therapeutic target for the treatment of bladder cancer.展开更多
文摘【目的】肉牛肌内脂肪沉积与牛肉的风味、多汁性和嫩度密切相关。脂肪沉积过程表现为脂肪细胞的增殖(数量增多)和分化(脂质生成),受到了多基因协同调控。前人研究发现,小鼠中Snail1可以参与肌肉发育和脂质稳态调控,但其在牛脂肪生成过程中的作用仍未知,有待进一步研究。【方法】以秦川牛为研究对象,克隆得到Snail1 CDS区序列,构建Snail1时空表达谱,运用生物信息学软件对其功能结构及靶基因进行预测。进一步,通过RNAi干扰结合CCK8、EdU、细胞流式及实时荧光定量PCR等方法探究Snail1对牛脂肪细胞增殖的影响。【结果】秦川牛Snail1与NCBI公布序列相比存在2处碱基同义突变,其在秦川牛新生牛肺、肾周脂肪、小肠呈现较高丰度表达;而在成年牛中,Snail1在肾周脂肪组织中的表达量最高,背最长肌中的表达量次之,肺脏组织中的表达量最低。生物信息学分析发现,Snail1启动子区存在1个651 bp CpG岛及C/EBP、PPARα等与脂肪生成相关的转录因子结合位点。CKⅠ(Ser92/96)、CKⅡ(Ser25/119,Thr89)、CDK1(Ser13/104/112/119/143/183/214/221)、CDK5(Ser105/107)等多个细胞周期相关激酶可能参与了Snail1蛋白的磷酸化修饰。通过对牛已注释基因启动子区提取、靶基因预测及KEGG动态网络构建发现,成脂相关的MAPK、PI3K-Akt、mTOR等信号通路为Snail1参与脂肪生成相关的潜在节点信号通路。进一步,通过RNAi干扰试验对其功能研究表明,Snail1下调促进了牛前体脂肪细胞的增殖,增加了复制期阳性细胞的比例(P<0.01)且促进了G1/S细胞周期转换。RT-qPCR和Western-blot检测表明,干扰Snail1显著上调了促增殖调控基因CCNB1、CCND2、CDK2、CDK4(P<0.05)和蛋白的表达。【结论】Snail1在新生牛肾周脂肪及成年牛肾周脂肪和背最长肌中表达量相对较高。干扰Snail1促进了牛前体脂肪细胞的增殖、G1/S细胞周期转变和CCNB1、CCND2、CDK2、CDK4等增殖相关基因表达;CKⅠ、CKⅡ、CDK1/5等多个细胞周期相关激酶可能通过磷酸化修饰Snail1蛋白进而参与细胞增殖调控,而MAPK、PI3K-Akt、mTOR等为Snail1影响牛脂肪细胞增殖潜在的关键节点通路。研究结果为进一步探究Snail1参与牛脂肪生成作用机制奠定了基础。
基金supported by the Basic Science Research Program through the National Research Foundation of Korea(NRF)funded by the Ministry of Education(2022R1I1A3070740)。
文摘Background The intestinal epithelium performs essential physiological functions,such as nutrient absorption,and acts as a barrier to prevent the entry of harmful substances.Mycotoxins are prevalent contaminants found in ani-mal feed that exert harmful effects on the health of livestock.Zearalenone(ZEA)is produced by the Fusarium genus and induces gastrointestinal dysfunction and disrupts the health and immune system of animals.Here,we evaluated the molecular mechanisms that regulate the effects of ZEA on the porcine intestinal epithelium.Results Treatment of IPEC-J2 cells with ZEA decreased the expression of E-cadherin and increased the expression of Snai1 and Vimentin,which induced Snail1-mediated epithelial-to-mesenchymal transition(EMT).In addition,ZEA induces Snail-mediated EMT through the activation of TGF-βsignaling.The treatment of IPEC-J2 cells with atractyle-nolideⅢ,which were exposed to ZEA,alleviated EMT.Conclusions Our findings provide insights into the molecular mechanisms of ZEA toxicity in porcine intestinal epi-thelial cells and ways to mitigate it.
基金supported by the National Natural Science Foundation of China(Nos.82273443,81602234 and 81802538)the Natural Science Foundation of Hubei Province(Nos.2017CFB637 and 2023AFB1041).
文摘Objective This study aimed to investigate the role of the long noncoding RNA(lncRNA)maternally expressed gene 3(MEG3)in the epithelial-mesenchymal transition(EMT)of bladder cancer cells and the potential mechanisms.Methods Cell invasion,migration,and wound healing assays were conducted to assess the effects of MEG3 on the invasive and migratory capabilities of bladder cancer cells.The expression levels of E-cadherin were measured using Western blotting,RT-qPCR,and dual luciferase reporter assays.RNA immunoprecipitation and pull-down assays were performed to investigate the interactions between MEG3 and its downstream targets.Results MEG3 suppressed the invasion and migration of bladder cancer cells and modulated the transcription of E-cadherin.The binding of MEG3 to the zinc finger region of the transcription factor Snail prevented its ability to transcriptionally repress E-cadherin.Additionally,MEG3 suppressed the phosphorylation of extracellular regulated protein kinase(ERK),c-Jun N-terminal kinase(JNK),and P38,thereby decreasing the expression of Snail and stimulating the expression of E-cadherin.Conclusion MEG3 plays a vital role in suppressing the EMT in bladder cancer cells,indicating its potential as a promising therapeutic target for the treatment of bladder cancer.
