旨在探究叉头盒蛋白O1(fork head box O 1,FoxO1)对高浓度非酯化脂肪酸(non-esterified fatty acids,NEFA)状态下的奶牛子宫内膜上皮细胞凋亡因子表达变化的影响。通过实时荧光定量PCR法及Western blot法评价添加激活剂白藜芦醇(resvera...旨在探究叉头盒蛋白O1(fork head box O 1,FoxO1)对高浓度非酯化脂肪酸(non-esterified fatty acids,NEFA)状态下的奶牛子宫内膜上皮细胞凋亡因子表达变化的影响。通过实时荧光定量PCR法及Western blot法评价添加激活剂白藜芦醇(resveratrol)后沉默信息调节因子(silent information regulator 1,SIRT1)的表达效果,明确奶牛子宫内膜上皮细胞中FoxO1的表达情况。加入Resveratrol和高浓度的NEFA后,检测FoxO1通路关键因子及细胞凋亡相关因子蛋白的表达变化情况。与对照组相比,添加Resveratrol上调SIRT1的蛋白和mRNA表达水平(P<0.01),下调Ac-FoxO1蛋白表达水平(P<0.01);下调Caspase-3蛋白表达水平无显著影响(P>0.05),上调Bcl-2、Bcl-xl的蛋白表达水平(P<0.01),下调Bax和Bax/Bcl-2的蛋白表达水平(P<0.05)。与NEFA组相比,NEFA+Resveratrol组上调IGF-1、SIRT1和p-AKT蛋白表达水平(P<0.01),下调FoxO1、p-FoxO1、Ac-FoxO1、FoxO1/p-FoxO1、AKT和AKT/p-AKT蛋白表达水平(P<0.01),上调Bcl-xl和Bcl-2蛋白表达水平(P<0.01),下调Caspase-3、Bax和Bax/Bcl-2蛋白表达水平(P<0.01)。激光共聚焦检测结果显示,NEFA+Resveratrol组与NEFA组相比,抑制FoxO1的表达水平(P<0.01)。Resveratrol可有效激活SIRT1的表达,进而抑制FoxO1的表达,降低高浓度NEFA状态下奶牛子宫内膜上皮细胞中凋亡因子的表达水平。展开更多
AIM:Regulatory T cells(Tregs)are a specialized subset of CD4^(+)T cells primarily involved in im⁃munosuppressive functions.AMP-activated protein kinase(AMPK)serves as a metabolic sensor that governs the differen⁃tiati...AIM:Regulatory T cells(Tregs)are a specialized subset of CD4^(+)T cells primarily involved in im⁃munosuppressive functions.AMP-activated protein kinase(AMPK)serves as a metabolic sensor that governs the differen⁃tiation,maturation,and immune functions of Tregs through metabolic reprogramming.However,the impact of AMPKα1(the catalytic subunit of AMPK)knockout specifically in Tregs on the host's immune microenvironment remains largely un⁃explored.METHODS:Histological changes in immune organs were assessed using HE staining.The types of immune cells and their relative population percentages in immune organs and blood were quantified through flow cytometry in both AMPKα1flox/flox(AMPKα1^(fl/fl))mice and Treg-specific AMPKα1 knockout mice(AMPKα1^(fl/fl)Foxp3^(cre)mice).RESULTS:Compared to AMPKα1^(fl/fl)mice,the percentage of eosinophils in the bone marrow of AMPKα1^(fl/fl)Foxp3^(cre)mice was significant⁃ly reduced.Additionally,while the thymus of AMPKα1^(fl/fl)Foxp3^(cre)mice exhibited normal structure,both its size and the ratio of thymus weight to body weight were significantly decreased.The knockout of AMPKα1 in Tregs led to a notable reduction in the total percentage of immature double-negative(DN)cells.Consequently,the percentage of CD4^(+)T cells derived from these DN cells also decreased,even though the percentages of DN1 and DN4 cells were higher in the thymus of AMPKα1^(fl/fl)Foxp3^(cre)mice compared to AMPKα1^(fl/fl)mice.Importantly,the proportion of Siglec-F+CD11b^(+)eosinophils in the thymus was significantly lower in AMPKα1^(fl/fl)Foxp3^(cre)mice.Knockout of AMPKα1 in Tregs resulted in a marked increase in the percentage of CD4^(+)T cells in peripheral blood,alongside a decrease in the proportion of mature CD8^(+)T cells.Similarly,the proportion of CD4^(+)T cells in the spleen of AMPKα1^(fl/fl)Foxp3^(cre)mice was elevated compared to AMPKα1^(fl/fl)mice.In contrast,the proportion of neutrophils significantly decreased,while mononuclear cell proportions increased in the spleen of AMPKα1^(fl/fl)Foxp3^(cre)mice.In lymph nodes,the medullary boundaries in AMPKα1^(fl/fl)Foxp3^(cre)mice were blurred,and the lymphoid follicles were missing,a feature not observed in AMPKα1^(fl/fl)mice.Furthermore,the knockout of AMPKα1 in Tregs reduced the CD3^(+)T cell population,particularly the CD8^(+)T cell population,in lymph nodes.Although the mature Treg cell population was significantly lower in AMPKα1^(fl/fl)Foxp3^(cre)mice,the percentage of CD4^(+)T cells was markedly in⁃creased.In contrast,there was no statistically significant difference in granulocyte populations between AMPKα1^(fl/fl)Foxp3^(cre)and AMPKα1^(fl/fl)mice.CONCLUSION:The populations of mature Tregs,CD8^(+)T cells and eosinophils in various im⁃mune organs were significantly altered in mice with Treg-specific AMPKα1 knockout,suggesting a potential remodeling of the host immune microenvironment in response to inflammatory stimuli.展开更多
文摘旨在探究叉头盒蛋白O1(fork head box O 1,FoxO1)对高浓度非酯化脂肪酸(non-esterified fatty acids,NEFA)状态下的奶牛子宫内膜上皮细胞凋亡因子表达变化的影响。通过实时荧光定量PCR法及Western blot法评价添加激活剂白藜芦醇(resveratrol)后沉默信息调节因子(silent information regulator 1,SIRT1)的表达效果,明确奶牛子宫内膜上皮细胞中FoxO1的表达情况。加入Resveratrol和高浓度的NEFA后,检测FoxO1通路关键因子及细胞凋亡相关因子蛋白的表达变化情况。与对照组相比,添加Resveratrol上调SIRT1的蛋白和mRNA表达水平(P<0.01),下调Ac-FoxO1蛋白表达水平(P<0.01);下调Caspase-3蛋白表达水平无显著影响(P>0.05),上调Bcl-2、Bcl-xl的蛋白表达水平(P<0.01),下调Bax和Bax/Bcl-2的蛋白表达水平(P<0.05)。与NEFA组相比,NEFA+Resveratrol组上调IGF-1、SIRT1和p-AKT蛋白表达水平(P<0.01),下调FoxO1、p-FoxO1、Ac-FoxO1、FoxO1/p-FoxO1、AKT和AKT/p-AKT蛋白表达水平(P<0.01),上调Bcl-xl和Bcl-2蛋白表达水平(P<0.01),下调Caspase-3、Bax和Bax/Bcl-2蛋白表达水平(P<0.01)。激光共聚焦检测结果显示,NEFA+Resveratrol组与NEFA组相比,抑制FoxO1的表达水平(P<0.01)。Resveratrol可有效激活SIRT1的表达,进而抑制FoxO1的表达,降低高浓度NEFA状态下奶牛子宫内膜上皮细胞中凋亡因子的表达水平。
基金Supported by the National Natural Science Foundation of China(No.81800423)the Guangdong Medical Science and Technology Research project(No.B2022102)。
文摘AIM:Regulatory T cells(Tregs)are a specialized subset of CD4^(+)T cells primarily involved in im⁃munosuppressive functions.AMP-activated protein kinase(AMPK)serves as a metabolic sensor that governs the differen⁃tiation,maturation,and immune functions of Tregs through metabolic reprogramming.However,the impact of AMPKα1(the catalytic subunit of AMPK)knockout specifically in Tregs on the host's immune microenvironment remains largely un⁃explored.METHODS:Histological changes in immune organs were assessed using HE staining.The types of immune cells and their relative population percentages in immune organs and blood were quantified through flow cytometry in both AMPKα1flox/flox(AMPKα1^(fl/fl))mice and Treg-specific AMPKα1 knockout mice(AMPKα1^(fl/fl)Foxp3^(cre)mice).RESULTS:Compared to AMPKα1^(fl/fl)mice,the percentage of eosinophils in the bone marrow of AMPKα1^(fl/fl)Foxp3^(cre)mice was significant⁃ly reduced.Additionally,while the thymus of AMPKα1^(fl/fl)Foxp3^(cre)mice exhibited normal structure,both its size and the ratio of thymus weight to body weight were significantly decreased.The knockout of AMPKα1 in Tregs led to a notable reduction in the total percentage of immature double-negative(DN)cells.Consequently,the percentage of CD4^(+)T cells derived from these DN cells also decreased,even though the percentages of DN1 and DN4 cells were higher in the thymus of AMPKα1^(fl/fl)Foxp3^(cre)mice compared to AMPKα1^(fl/fl)mice.Importantly,the proportion of Siglec-F+CD11b^(+)eosinophils in the thymus was significantly lower in AMPKα1^(fl/fl)Foxp3^(cre)mice.Knockout of AMPKα1 in Tregs resulted in a marked increase in the percentage of CD4^(+)T cells in peripheral blood,alongside a decrease in the proportion of mature CD8^(+)T cells.Similarly,the proportion of CD4^(+)T cells in the spleen of AMPKα1^(fl/fl)Foxp3^(cre)mice was elevated compared to AMPKα1^(fl/fl)mice.In contrast,the proportion of neutrophils significantly decreased,while mononuclear cell proportions increased in the spleen of AMPKα1^(fl/fl)Foxp3^(cre)mice.In lymph nodes,the medullary boundaries in AMPKα1^(fl/fl)Foxp3^(cre)mice were blurred,and the lymphoid follicles were missing,a feature not observed in AMPKα1^(fl/fl)mice.Furthermore,the knockout of AMPKα1 in Tregs reduced the CD3^(+)T cell population,particularly the CD8^(+)T cell population,in lymph nodes.Although the mature Treg cell population was significantly lower in AMPKα1^(fl/fl)Foxp3^(cre)mice,the percentage of CD4^(+)T cells was markedly in⁃creased.In contrast,there was no statistically significant difference in granulocyte populations between AMPKα1^(fl/fl)Foxp3^(cre)and AMPKα1^(fl/fl)mice.CONCLUSION:The populations of mature Tregs,CD8^(+)T cells and eosinophils in various im⁃mune organs were significantly altered in mice with Treg-specific AMPKα1 knockout,suggesting a potential remodeling of the host immune microenvironment in response to inflammatory stimuli.
基金supported by the National High-Tech R&D Program of China(“863”Program)(2009AA22704)the National Natural Science Foundation of China(30873089,81173129)+3 种基金the Program for Changjiang Scholars and Innovative Research Team in University(IRT0946)the Open Foundation of Innovative Platform in University of Hunan Province of China(10K078)the Science and Technology Plan Key Grant of Hunan Province of China(2009TP40682)the Fundamental Research Funds for the Central Universities(201023100001)
文摘目的:在细胞水平研究烟酰胺单核苷酸(nicotinamide mononucleotide,NMN)对胰岛素分泌的调节作用及其对与胰岛素分泌相关的重要转录因子胰十二指肠同源盒基因(pancreatic and duodenalhomeobox-1,PDX-1)和分叉头框家族转录因子1(forkhead box-containing protein O-1,FoxO1)基因表达的影响。方法:采用大鼠胰岛素ELISA试剂盒检测RIN-m5f细胞胰岛素分泌水平。用Real-time PCR检测RIN-m5f细胞PDX-1和FoxO1的mRNA表达水平。用Western印迹检测RIN-m5f细胞PDX-1蛋白表达水平。结果:用瑞格列奈10 nmol/L+NMN 100μmol/L处理RIN-m5f细胞48 h,与空白对照及DMSO对照组相比,胰岛素分泌量均显著增高(P<0.05);与NMN 50μmol/L组比较,胰岛素分泌量的增高也有统计学意义(P<0.05)。10,50和100μmol/L的NMN作用RIN-m5f细胞36 h,PDX-1的mRNA表达量均上调(依次为P<0.05,P<0.01,P<0.001)。100μmol/L剂量组与10μmol/L和50μmol/L剂量组比较差异也有统计学意义(P<0.001)。50,100和200μmol/L的NMN作用RIN-m5f细胞36或48 h,PDX-1的蛋白表达量与对照组比较差异无统计学意义(P>0.05)。结论:NMN可以调控RIN-m5f细胞中胰岛素的分泌及PDX-1的mRNA表达水平。
