Background:Periodontitis is characterized by alveolar bone resorption,aggravated by osteoblast dysfunction,and associated with intracellular oxidative stress linked to the nuclear factor erythroid 2-related factor 2(N...Background:Periodontitis is characterized by alveolar bone resorption,aggravated by osteoblast dysfunction,and associated with intracellular oxidative stress linked to the nuclear factor erythroid 2-related factor 2(NRF2)level.We evaluated the molecular mechanism of periodontitis onset and development and the role of NRF2 in osteogenic differentiation.Methods:Primary murine mandibular osteoblasts were extracted and exposed to Porphyromonas gingivalis lipopolysaccharide(Pg-LPS)or other stimuli.Reactive oxygen species(ROS)and 5,5',6,6'-tetrachloro-1,1',3,3'-tetraethylbenzimidazolylcarbocyanine iodide(JC-1)staining were used to detect intracellular oxidative stress.Alkaline phosphatase staining and alizarin red S staining were used to detect the osteogenic differentiation of osteoblasts.Immunofluorescence and western blotting were used to determine the changes in the mitogen-activated protein kinase(MAPK)pathway and related molecule activities.Immunofluorescence colocalization and co-immunoprecipitation were performed to examine the nuclear translocation of NRF2 and its interaction with dual-specific phosphatase 1(DUSP1)in cells.Results:Ligated tissue samples showed higher alveolar bone resorption rate and lower NRF2 level than healthy periodontal tissue samples.Pg-LPS increased intracellular oxidative stress levels and inhibited osteogenic differentiation,whereas changes in NRF2 expression were correlated with changes in the oxidative stress and osteogenesis rate.NRF2 promoted the dephosphorylation of the MAPK pathway by nuclear translocation and the upregulation of DUSP1 expression,thus enhancing the osteogenic differentiation capacity of mandibular osteoblasts.The interaction between NRF2 and DUSP1 was observed.Conclusions:NRF2 and its nuclear translocation can regulate the osteogenic differentiation of mandibular osteoblasts under Pg-LPS conditions by interacting with DUSP1 in a process linked to the MAPK pathway.These findings form the basis of periodontitis treatment.展开更多
目的:在肝癌的治疗中,化学疗法是重要的治疗手段,尤其在结合外科手术切除的联合治疗中,化疗被广泛的应用于临床。而体内试验证实,化疗药物诱导的细胞衰老在治疗肿瘤的过程中,普遍存在并发挥重要作用,我们着重探讨在药物诱导的人肝癌细...目的:在肝癌的治疗中,化学疗法是重要的治疗手段,尤其在结合外科手术切除的联合治疗中,化疗被广泛的应用于临床。而体内试验证实,化疗药物诱导的细胞衰老在治疗肿瘤的过程中,普遍存在并发挥重要作用,我们着重探讨在药物诱导的人肝癌细胞衰老中,细胞内重要信号途径的变化,发现并利用对细胞衰老有重要意义的蛋白,增强化学治疗肿瘤的效果。在本课题中,我们利用化疗药物阿霉素(doxorubicin)和蛋白酶体抑制剂MG132诱导的人衰老肝癌细胞模型,对MAPK信号途径的负调控因子DUSP家族的表达变化进行检测,筛选表达水平发生显著变化的双特异性磷酸酶(DUSP)家族成员,并初步探讨其在细胞衰老中的作用。方法:利用低剂量的阿霉素和蛋白酶体抑制剂MG132诱导人肝癌细胞衰老;通过Real time RT-PCR和Western blotting检测DUSP家族基因mRNA和蛋白质水平的变化;利用siRNA干扰技术敲降DUSP1,检测药物诱导肝癌细胞衰老水平的变化。结果:经低剂量的阿霉素和蛋白酶体抑制剂短时间处理后,人肝癌细胞系Huh7和SMMC-7721细胞发生明显的细胞衰老;在诱导的衰老细胞中,DUSP1mRNA和蛋白质水平呈显著升高。在敲降DUSP1后,MG132诱导的细胞衰老得到显著的缓解。结论:DUSP1在药物诱导的人肝癌细胞衰老中具有重要的作用,为研究临床治疗中化疗药物引起肿瘤细胞衰老的具体机制带来启发。展开更多
Background:Previously,phosphoinositide-3-kinase regulatory subunit 1(PIK3R1)and dual specificity phosphatase 1(DUSP1)were identified as promising candidate genes for milk production traits due to their being different...Background:Previously,phosphoinositide-3-kinase regulatory subunit 1(PIK3R1)and dual specificity phosphatase 1(DUSP1)were identified as promising candidate genes for milk production traits due to their being differentially expressed between the dry period and the peak of lactation in livers of dairy cows.Hence,in this study,the single nucleotide polymorphisms(SNPs)of PIK3R1 and DUSP1 genes were identified and their genetic associations with milk yield,fat yield,fat percentage,protein yield,and protein percentage,were investigated using 1067 Chinese Holstein cows from 40 sire families.Results:By re-sequencing the entire coding region and 2000 bp of the 5′and 3′flanking regions of the two genes,one SNP in the 5′untranslated region(UTR),three in the 3′UTR,and two in the 3′flanking region of PIK3R1 were identified,and one in the 5′flanking region,one in the 3′UTR,and two in the 3′flanking region of DUSP1 were found.Subsequent single-locus association analyses showed that five SNPs in PIK3 R1,rs42590258,rs210389799,rs208819656,rs41255622,rs133655926,and rs211408208,and four SNPs in DUSP1,rs207593520,rs208460068,rs209154772,and rs210000760,were significantly associated with milk,fat and protein yields in the first or second lactation(P values≤0.0001 and 0.0461).In addition,by the Haploview 4.2 software,the six and four SNPs in PIK3R1 and DUSP1 respectively formed one haplotype block,and the haplotype-based association analyses showed significant associations between their haplotype combinations and the milk traits in both two lactations(P values≤0.0001 and 0.0364).One SNP,rs207593520(T/G),was predicted to alter the transcription factor binding sites(TFBSs)in the 5′flanking region of DUSP1.Further,the dual-luciferase assay showed that the transcription activity of allele T in rs207593520 was significantly higher than that of allele G,suggesting the activation of transcriptional activity of DUSP1 gene by allele T of rs207593520.Thus,the rs207593520 SNP was highlighted as a potential causal mutation that should be further verified.Conclusions:We demonstrated novel and significant genetic effects of the PIK3R1 and DUSP1 genes on milk production traits in dairy cows,and our findings provide information for use in dairy cattle breeding.展开更多
基金supported by the National Natural Science Foundation of China(Nos.82170954 and 82301066)the Medical Science and Technology Project of Zhejiang Province of China(No.2024KY1064)the Zhejiang Provincial Natural Science Foundation of China(No.LQN25H140005)。
文摘Background:Periodontitis is characterized by alveolar bone resorption,aggravated by osteoblast dysfunction,and associated with intracellular oxidative stress linked to the nuclear factor erythroid 2-related factor 2(NRF2)level.We evaluated the molecular mechanism of periodontitis onset and development and the role of NRF2 in osteogenic differentiation.Methods:Primary murine mandibular osteoblasts were extracted and exposed to Porphyromonas gingivalis lipopolysaccharide(Pg-LPS)or other stimuli.Reactive oxygen species(ROS)and 5,5',6,6'-tetrachloro-1,1',3,3'-tetraethylbenzimidazolylcarbocyanine iodide(JC-1)staining were used to detect intracellular oxidative stress.Alkaline phosphatase staining and alizarin red S staining were used to detect the osteogenic differentiation of osteoblasts.Immunofluorescence and western blotting were used to determine the changes in the mitogen-activated protein kinase(MAPK)pathway and related molecule activities.Immunofluorescence colocalization and co-immunoprecipitation were performed to examine the nuclear translocation of NRF2 and its interaction with dual-specific phosphatase 1(DUSP1)in cells.Results:Ligated tissue samples showed higher alveolar bone resorption rate and lower NRF2 level than healthy periodontal tissue samples.Pg-LPS increased intracellular oxidative stress levels and inhibited osteogenic differentiation,whereas changes in NRF2 expression were correlated with changes in the oxidative stress and osteogenesis rate.NRF2 promoted the dephosphorylation of the MAPK pathway by nuclear translocation and the upregulation of DUSP1 expression,thus enhancing the osteogenic differentiation capacity of mandibular osteoblasts.The interaction between NRF2 and DUSP1 was observed.Conclusions:NRF2 and its nuclear translocation can regulate the osteogenic differentiation of mandibular osteoblasts under Pg-LPS conditions by interacting with DUSP1 in a process linked to the MAPK pathway.These findings form the basis of periodontitis treatment.
文摘目的:在肝癌的治疗中,化学疗法是重要的治疗手段,尤其在结合外科手术切除的联合治疗中,化疗被广泛的应用于临床。而体内试验证实,化疗药物诱导的细胞衰老在治疗肿瘤的过程中,普遍存在并发挥重要作用,我们着重探讨在药物诱导的人肝癌细胞衰老中,细胞内重要信号途径的变化,发现并利用对细胞衰老有重要意义的蛋白,增强化学治疗肿瘤的效果。在本课题中,我们利用化疗药物阿霉素(doxorubicin)和蛋白酶体抑制剂MG132诱导的人衰老肝癌细胞模型,对MAPK信号途径的负调控因子DUSP家族的表达变化进行检测,筛选表达水平发生显著变化的双特异性磷酸酶(DUSP)家族成员,并初步探讨其在细胞衰老中的作用。方法:利用低剂量的阿霉素和蛋白酶体抑制剂MG132诱导人肝癌细胞衰老;通过Real time RT-PCR和Western blotting检测DUSP家族基因mRNA和蛋白质水平的变化;利用siRNA干扰技术敲降DUSP1,检测药物诱导肝癌细胞衰老水平的变化。结果:经低剂量的阿霉素和蛋白酶体抑制剂短时间处理后,人肝癌细胞系Huh7和SMMC-7721细胞发生明显的细胞衰老;在诱导的衰老细胞中,DUSP1mRNA和蛋白质水平呈显著升高。在敲降DUSP1后,MG132诱导的细胞衰老得到显著的缓解。结论:DUSP1在药物诱导的人肝癌细胞衰老中具有重要的作用,为研究临床治疗中化疗药物引起肿瘤细胞衰老的具体机制带来启发。
基金financially supported by the National Natural Science Foundation of China(31872330,31802041)Beijing Dairy Industry Innovation Team(BAIC06–2018/2019)+3 种基金Beijing Science and Technology Program(D171100002417001)National Science and Technology Programs of China(2013AA102504)earmarked fund for Modern Agro-industry Technology Research System(CARS-36)the Program for Changjiang Scholar and Innovation Research Team in University(IRT_15R62).
文摘Background:Previously,phosphoinositide-3-kinase regulatory subunit 1(PIK3R1)and dual specificity phosphatase 1(DUSP1)were identified as promising candidate genes for milk production traits due to their being differentially expressed between the dry period and the peak of lactation in livers of dairy cows.Hence,in this study,the single nucleotide polymorphisms(SNPs)of PIK3R1 and DUSP1 genes were identified and their genetic associations with milk yield,fat yield,fat percentage,protein yield,and protein percentage,were investigated using 1067 Chinese Holstein cows from 40 sire families.Results:By re-sequencing the entire coding region and 2000 bp of the 5′and 3′flanking regions of the two genes,one SNP in the 5′untranslated region(UTR),three in the 3′UTR,and two in the 3′flanking region of PIK3R1 were identified,and one in the 5′flanking region,one in the 3′UTR,and two in the 3′flanking region of DUSP1 were found.Subsequent single-locus association analyses showed that five SNPs in PIK3 R1,rs42590258,rs210389799,rs208819656,rs41255622,rs133655926,and rs211408208,and four SNPs in DUSP1,rs207593520,rs208460068,rs209154772,and rs210000760,were significantly associated with milk,fat and protein yields in the first or second lactation(P values≤0.0001 and 0.0461).In addition,by the Haploview 4.2 software,the six and four SNPs in PIK3R1 and DUSP1 respectively formed one haplotype block,and the haplotype-based association analyses showed significant associations between their haplotype combinations and the milk traits in both two lactations(P values≤0.0001 and 0.0364).One SNP,rs207593520(T/G),was predicted to alter the transcription factor binding sites(TFBSs)in the 5′flanking region of DUSP1.Further,the dual-luciferase assay showed that the transcription activity of allele T in rs207593520 was significantly higher than that of allele G,suggesting the activation of transcriptional activity of DUSP1 gene by allele T of rs207593520.Thus,the rs207593520 SNP was highlighted as a potential causal mutation that should be further verified.Conclusions:We demonstrated novel and significant genetic effects of the PIK3R1 and DUSP1 genes on milk production traits in dairy cows,and our findings provide information for use in dairy cattle breeding.
