Colitis-associated colorectal cancer(CAC)is a major contributor to cancer-related mortality worldwide.Titanium dioxide(TiO_(2),E171),a widely used food additive,has been insufficiently studied regarding its effects on...Colitis-associated colorectal cancer(CAC)is a major contributor to cancer-related mortality worldwide.Titanium dioxide(TiO_(2),E171),a widely used food additive,has been insufficiently studied regarding its effects on macrophages within colon tumors during CAC development.In this study,CAC mouse models were used to investigate the biological impact of dietary E171 on macrophages in vivo,while lipopolysaccharide(LPS)-stimulated RAW264.7 macrophage cell lines were employed to elucidate the underlying mechanisms in vitro.We found that dietary E171 intake accelerated CAC development,exacerbated inflammatory responses and oxidative stress,and upregulated CAC-associated genes,including S100a8,S100a9,Lcn2,S100a11,Cxcl2,and interleukin-1α(Il-1α).E171 also increased the expression of S100A8,S100A9,NOD-like receptor family pyrin domain-containing 3(NLRP3),and gasdermin-D Nterminal(GSDMD-N)in macrophages within colon tumors.In inflammatory macrophages,E171 exposure enhanced cell viability,increased reactive oxygen species(ROS)levels,and elevated the expression and secretion of S100A8 and S100A9,consistent with in vivo histological observations.Furthermore,E171-induced secretion of S100A8 and S100A9 in macrophages was suppressed by specific inhibitors,including N-acetylcysteine(NAC,ROS inhibitor),MCC950(NLRP3 inhibitor),Z-YVAD-FMK(caspase 1 inhibitor),disulfiram(GSDMD inhibitor),and transfection of NLRP3 small interfering ribonucleic acid(siRNA).These results indicate that dietary E171 promotes CAC development by activating macrophages,with S100A8 and S100A9 serving as key mediators,and the NLRP3/caspase 1/GSDMD pathway acting as a critical mechanism.展开更多
目的:探究牛结核分枝杆菌减毒株卡介苗(bacillus Calmette-Guérin,BCG)感染人单核-巨噬细胞THP-1后,S100钙结合蛋白A4(S100 calcium-binding protein A4,S100A4)通过磷脂酰肌醇3-激酶(phosphatidylinositol 3-kinase,PI3K)/蛋白激...目的:探究牛结核分枝杆菌减毒株卡介苗(bacillus Calmette-Guérin,BCG)感染人单核-巨噬细胞THP-1后,S100钙结合蛋白A4(S100 calcium-binding protein A4,S100A4)通过磷脂酰肌醇3-激酶(phosphatidylinositol 3-kinase,PI3K)/蛋白激酶B(protein kinase B,PKB/AKT)/哺乳动物雷帕霉素靶蛋白(mammalian target of rapamycin,mTOR)信号通路对BCG诱导的巨噬细胞自噬的调控作用。方法:采用S100A4小干扰RNA(S100A4 small interfering RNA,si-S100A4)转染或PI3K抑制剂LY294002预处理THP-1巨噬细胞,并经BCG感染,设置不同处理组和对照组,通过Western blot方法检测PI3K/AKT/mTOR信号通路相关蛋白和自噬相关蛋白[微管相关蛋白1轻链3-II(microtubule-associated protein 1 light chain 3-II,LC3-II)、自噬相关蛋白7(autophagy-related protein 7,Atg7)和beclin-1]的表达,mRFP-GFP-LC3自噬双标腺病毒方法检测自噬流,透射电子显微镜观察细胞中自噬体和自噬溶酶体数量变化。结果:与未感染组相比,BCG感染组中p-PI3K、p-AKT和p-mTOR蛋白水平显著降低(P<0.05)。与BCG感染组相比,si-S100A4+BCG组上述磷酸化蛋白水平显著升高(P<0.05),LC3-II、Atg7和beclin-1蛋白水平显著降低(P<0.01);LY294003+BCG组上述磷酸化蛋白水平显著降低(P<0.01),LC3-II、Atg7和beclin-1蛋白水平显著升高(P<0.05),自噬体和自噬溶酶体数量显著增多(P<0.01)。与si-S100A4+BCG组相比,si-S100A4+LY294002+BCG组中p-PI3K、p-AKT和p-mTOR蛋白水平显著降低(P<0.01),LC3-II、Atg7和beclin-1蛋白水平显著升高(P<0.01),自噬体和自噬溶酶体数量显著增多(P<0.01),胞内菌载量显著减少(P<0.01)。结论:干扰S100A4经由PI3K/AKT/mTOR信号通路对BCG诱导的巨噬细胞自噬具有抑制作用。展开更多
基金supported by the National Natural Science Foundation of China(Nos.81974441 and 82203619)the Science and Technology Planning Project of Shenzhen Municipality(Nos.JCYJ20190814105619048 and JCYJ20220530154202005)。
文摘Colitis-associated colorectal cancer(CAC)is a major contributor to cancer-related mortality worldwide.Titanium dioxide(TiO_(2),E171),a widely used food additive,has been insufficiently studied regarding its effects on macrophages within colon tumors during CAC development.In this study,CAC mouse models were used to investigate the biological impact of dietary E171 on macrophages in vivo,while lipopolysaccharide(LPS)-stimulated RAW264.7 macrophage cell lines were employed to elucidate the underlying mechanisms in vitro.We found that dietary E171 intake accelerated CAC development,exacerbated inflammatory responses and oxidative stress,and upregulated CAC-associated genes,including S100a8,S100a9,Lcn2,S100a11,Cxcl2,and interleukin-1α(Il-1α).E171 also increased the expression of S100A8,S100A9,NOD-like receptor family pyrin domain-containing 3(NLRP3),and gasdermin-D Nterminal(GSDMD-N)in macrophages within colon tumors.In inflammatory macrophages,E171 exposure enhanced cell viability,increased reactive oxygen species(ROS)levels,and elevated the expression and secretion of S100A8 and S100A9,consistent with in vivo histological observations.Furthermore,E171-induced secretion of S100A8 and S100A9 in macrophages was suppressed by specific inhibitors,including N-acetylcysteine(NAC,ROS inhibitor),MCC950(NLRP3 inhibitor),Z-YVAD-FMK(caspase 1 inhibitor),disulfiram(GSDMD inhibitor),and transfection of NLRP3 small interfering ribonucleic acid(siRNA).These results indicate that dietary E171 promotes CAC development by activating macrophages,with S100A8 and S100A9 serving as key mediators,and the NLRP3/caspase 1/GSDMD pathway acting as a critical mechanism.
文摘目的:探究牛结核分枝杆菌减毒株卡介苗(bacillus Calmette-Guérin,BCG)感染人单核-巨噬细胞THP-1后,S100钙结合蛋白A4(S100 calcium-binding protein A4,S100A4)通过磷脂酰肌醇3-激酶(phosphatidylinositol 3-kinase,PI3K)/蛋白激酶B(protein kinase B,PKB/AKT)/哺乳动物雷帕霉素靶蛋白(mammalian target of rapamycin,mTOR)信号通路对BCG诱导的巨噬细胞自噬的调控作用。方法:采用S100A4小干扰RNA(S100A4 small interfering RNA,si-S100A4)转染或PI3K抑制剂LY294002预处理THP-1巨噬细胞,并经BCG感染,设置不同处理组和对照组,通过Western blot方法检测PI3K/AKT/mTOR信号通路相关蛋白和自噬相关蛋白[微管相关蛋白1轻链3-II(microtubule-associated protein 1 light chain 3-II,LC3-II)、自噬相关蛋白7(autophagy-related protein 7,Atg7)和beclin-1]的表达,mRFP-GFP-LC3自噬双标腺病毒方法检测自噬流,透射电子显微镜观察细胞中自噬体和自噬溶酶体数量变化。结果:与未感染组相比,BCG感染组中p-PI3K、p-AKT和p-mTOR蛋白水平显著降低(P<0.05)。与BCG感染组相比,si-S100A4+BCG组上述磷酸化蛋白水平显著升高(P<0.05),LC3-II、Atg7和beclin-1蛋白水平显著降低(P<0.01);LY294003+BCG组上述磷酸化蛋白水平显著降低(P<0.01),LC3-II、Atg7和beclin-1蛋白水平显著升高(P<0.05),自噬体和自噬溶酶体数量显著增多(P<0.01)。与si-S100A4+BCG组相比,si-S100A4+LY294002+BCG组中p-PI3K、p-AKT和p-mTOR蛋白水平显著降低(P<0.01),LC3-II、Atg7和beclin-1蛋白水平显著升高(P<0.01),自噬体和自噬溶酶体数量显著增多(P<0.01),胞内菌载量显著减少(P<0.01)。结论:干扰S100A4经由PI3K/AKT/mTOR信号通路对BCG诱导的巨噬细胞自噬具有抑制作用。
