为分析E3泛素连接酶环指蛋白125(ring finger protein 125,RNF125)在肝癌中的表达及临床预后意义,并探究RNF125对肝癌侵袭和转移的影响及其作用机制,通过ENCORI数据库和临床组织样本分析RNF125在肝癌和癌旁组织中的表达量,使用ENCORI数...为分析E3泛素连接酶环指蛋白125(ring finger protein 125,RNF125)在肝癌中的表达及临床预后意义,并探究RNF125对肝癌侵袭和转移的影响及其作用机制,通过ENCORI数据库和临床组织样本分析RNF125在肝癌和癌旁组织中的表达量,使用ENCORI数据库分析RNF125与肝癌预后的关系。构建敲低和过表达RNF125的肝癌细胞系,通过Transwell试验和细胞划痕试验检验RNF125表达水平对肝癌细胞迁移、侵袭能力的影响。通过动物试验验证调控RNF125对体内肿瘤生长情况及对肿瘤免疫微环境的影响,质谱分析寻找RNF125的靶蛋白,qPCR、蛋白质免疫印迹技术、泛素化试验验证RNF125与MYH9之间的关系,通过蛋白质免疫印迹和细胞免疫荧光技术确定调控RNF125与MYH9对EMT标志蛋白的影响。结果表明:RNF125在肝癌组织中表达较低,RNF125的高表达提示良好的预后。RNF125的表达水平与肝癌细胞迁移和侵袭能力呈负相关。过表达RNF125使肿瘤生长延迟并增加免疫细胞的浸润水平。RNF125特异性结合MYH9并促进其泛素化降解。RNF125与上皮细胞标志物的表达水平呈正相关,与间充质细胞标志物呈负相关。过表达MYH9可逆转RNF125对EMT标志蛋白的影响。综上,RNF125在肝癌中表达下调,RNF125的高表达与肝癌患者的良好预后相关,RNF125通过泛素化MYH9来抑制肝癌的EMT过程,从而抑制肝癌的迁移和侵袭。展开更多
目的探究肌球蛋白重链11(myosin heavy chain 11,MYH11)对喉鳞状细胞癌细胞恶性生物学行为的影响及机制。方法qRT-PCR检测MYH11 mRNA在喉鳞状细胞癌组织以及喉鳞状细胞癌细胞中的表达。喉鳞状细胞癌细胞TU686分为si-MYH11组和si-NC组。F...目的探究肌球蛋白重链11(myosin heavy chain 11,MYH11)对喉鳞状细胞癌细胞恶性生物学行为的影响及机制。方法qRT-PCR检测MYH11 mRNA在喉鳞状细胞癌组织以及喉鳞状细胞癌细胞中的表达。喉鳞状细胞癌细胞TU686分为si-MYH11组和si-NC组。FD-LSC-1细胞分为MYH11组和Vector组。CCK8、流式细胞术、细胞划痕、Transwell实验分别用于检测细胞增殖、凋亡、迁移以及侵袭能力,蛋白质印迹检测各组细胞ERK 1/2磷酸化水平及MAPK相对表达量。结果MYH11高表达于喉鳞状细胞癌组织及细胞。si-MYH11组TU686细胞增殖、迁移和侵袭能力显著低于si-NC组(P_(均)<0.05),细胞凋亡显著高于si-NC组(P<0.01),ERK1/2磷酸化水平及MAPK表达显著低于si-NC组(P<0.01)。MYH11组TU686细胞增殖、迁移和侵袭能力显著高于Vector组(P_(均)<0.05),细胞凋亡显著低于Vector组(P<0.01),ERK1/2磷酸化水平及MAPK表达显著高于Vector组(P<0.01)。结论MYH11激活ERK/MAPK信号通路而促进喉鳞状细胞癌细胞的增殖、迁移和侵袭能力。展开更多
为了解猪脂肪组织肌球蛋白重链3(myosin heavy chain 3,MYH3)基因启动子区DNA甲基化及其mRNA表达水平在生长过程中的变化规律,试验采用亚硫酸氢盐测序(BSP)法分析6日龄和8月龄吉林花猪、松辽黑猪、二元猪脂肪组织MYH3基因启动子区DNA甲...为了解猪脂肪组织肌球蛋白重链3(myosin heavy chain 3,MYH3)基因启动子区DNA甲基化及其mRNA表达水平在生长过程中的变化规律,试验采用亚硫酸氢盐测序(BSP)法分析6日龄和8月龄吉林花猪、松辽黑猪、二元猪脂肪组织MYH3基因启动子区DNA甲基化水平,利用实时荧光定量PCR方法分析3个品种猪脂肪的MYH3基因mRNA表达量。结果表明:6日龄吉林花猪、松辽黑猪、二元猪脂肪的MYH3基因启动子区DNA甲基化水平分别为88.4%、89.3%、87.5%,3个品种间DNA甲基化水平差异不显著(P>0.05),8月龄吉林花猪、松辽黑猪、二元猪MYH3基因启动子区DNA甲基化水平分别为77.7%、76.8%、83.9%,吉林花猪和松辽黑猪DNA甲基化水平显著低于二元猪(P<0.05),吉林花猪与松辽黑猪差异不显著(P>0.05);8月龄吉林花猪、松辽黑猪脂肪的MYH3基因启动子区DNA甲基化水平显著低于6日龄(P<0.05),而二元猪DNA甲基化水平在6日龄和8月龄差异不显著(P>0.05)。6日龄时,3个品种猪脂肪MYH3基因的mRNA表达量差异不显著(P>0.05);8月龄时,吉林花猪MYH3基因的mRNA相对表达量显著高于二元猪和松辽黑猪(P<0.05),松辽黑猪显著高于二元猪(P<0.05)。8月龄吉林花猪和松辽黑猪脂肪MYH3基因的mRNA表达量显著高于6日龄(P<0.05),而二元猪MYH3基因的mRNA表达量在6日龄和8月龄差异不显著(P>0.05)。说明随着年龄增长,吉林花猪、松辽黑猪脂肪的MYH3基因启动子区DNA甲基化水平与mRNA表达水平呈负相关,而二元猪无显著变化,地方品种猪在肉质、风味方面优于引进品种猪,推测启动子区DNA甲基化水平与脂肪生成和沉积具有相关性。展开更多
AIM:To explore the role of a previously-found MYH9 tail domain mutation(p.E1384Q)in the pathogenesis of congenital cataract.METHODS:The cell experiments were conducted in vitro.Wild-type(WT)MYH9 and p.E1384Q mutant fr...AIM:To explore the role of a previously-found MYH9 tail domain mutation(p.E1384Q)in the pathogenesis of congenital cataract.METHODS:The cell experiments were conducted in vitro.Wild-type(WT)MYH9 and p.E1384Q mutant fragments were constructed,which was then transiently transfected into Hek293T cell lines.Western blotting and quantitative real time polymerase chain reaction(qRT-PCR)were used to analyze the protein and mRNA level of non-muscle myosin IIA(NM IIA)and F-actin in transfected cells,and fluorescence microscopy was applied to explore the subcellular localization of NM IIA and F-actin.Cell counting kit-8(CCK8),woundhealing and double staining flow cytometry assays were performed to evaluate the proliferation,migration and apoptosis function of transfected cells,respectively.Transmission electron microscope was conducted to observe the alteration of organelle structure.RESULTS:The transiently-transfected WT and p.E1384Q mutant Hek293T cell lines was constructed.Western blot demonstrated that,comparing with MYH9WT group,the relative protein amount of NM IIA and F-actin significantly decreased in MYH9E1384Q cells(P<0.001).qRT-PCR analysis revealed that the relative mRNA amount of NM IIA and F-actin also significantly reduced in MYH9E1384Q cells when compared with MYH9WT.The immunofluorescence microscopy showed that the fluorescence signal of NM IIA and F-actin significantly decreased in E1384Q cells.The diffuse cytoplasmic distribution of NM IIA in MYH9WT was changed to be clumped distribution,presenting a“speckled”pattern characterized by aggregates of small size in MYH9E1384Q.Functional study revealed that the E1384Q mutation significantly inhibited cell proliferation(P=0.003)and migration(P<0.001),and promoted apoptosis(P<0.001).Electron microscope showed that the mutation remarkably decreased the number of mitochondria(P<0.001)and changed the phenotype of mitochondria.CONCLUSION:The missense gene mutation in MYH9(p.E1384Q)causing congenital cataract results in decreased amount and altered subcellular distribution of NM IIA and F-actin,accompanied by decreased cell proliferation and migration,promotes apoptosis and mitochondrial alteration.展开更多
BACKGROUND Chronic intestinal pseudo-obstruction(CIPO)is a rare and debilitating disorder,characterized by severe impairments in gastrointestinal motility.The affected sites include the enteric/intrinsic autonomic ner...BACKGROUND Chronic intestinal pseudo-obstruction(CIPO)is a rare and debilitating disorder,characterized by severe impairments in gastrointestinal motility.The affected sites include the enteric/intrinsic autonomic nerves(neuropathy),intestinal smooth muscle cells(myopathy),and interstitial cells of Cajal(mesenchymopathy).The etiology can be genetic,idiopathic,or acquired.Owing to its nonspecific clinical presentation and lack of definitive diagnostic methods,misdiagnosis of CIPO is common.CASE SUMMARY This case involved an older male with insidious onset in adolescence who presented with postprandial bloating,intermittent diarrhea,and weight loss.During the disease course,the patient experienced two episodes of intestinal obstruction.Imaging revealed multisegmental digestive tract abnormalities(gastric emptying disorder,significant duodenal dilatation,and segmental jejunal dilatation).Whole-exome sequencing revealed a rare MYH11 mutation[NM_0010-40113.2:C.5819del(p.Pro1940HisfsTer91)],confirming hereditary myopathic CIPO.CONCLUSION This report adds to our current understanding of CIPO etiology by reinforcing the role of MYH11 variants in the pathogenesis of the CIPO phenotype.展开更多
BACKGROUND Hypertrophic cardiomyopathy(HCM)is one of the most prevalent inherited myocardial disorders and is charac-terized by considerable genetic and phenotypic heterogeneity.A subset of patients with HCM progress ...BACKGROUND Hypertrophic cardiomyopathy(HCM)is one of the most prevalent inherited myocardial disorders and is charac-terized by considerable genetic and phenotypic heterogeneity.A subset of patients with HCM progress to a dilated phase of HCM(DPHCM),which is associated with a poor prognosis;however,the underlying pathogenesis remains inadequately understood.CASE SUMMARY In this study,we present a case involving a pedigree with familial DPHCM and conduct a retrospective review of patients with DPHCM with identified gene mutations.Through panel sequencing targeting the coding regions of 312 genes associated with inherited cardiomyopathy,a heterozygous missense mutation(c.746G>A,p.Arg249Glu)in the MYH7 gene was identified in the proband(III-5).Sanger sequencing subsequently confirmed this pathogenic mutation in three additional family members(II-4,III-4,and IV-3).A total of 26 well-documented patients with DPHCM were identified in the literature.Patients with DPHCM are commonly middle-aged and male.The mean age of patients with DPHCM was 53.43±12.79 years.Heart failure,dyspnoea,and atrial fibrillation were the most prevalent symptoms observed,accompanied by an average left ventricular end-diastolic size of 58.62 mm.CONCLUSION Our findings corroborate the pathogenicity of the MYH7(c.746G>A,p.Arg249Glu)mutation for DPHCM and suggest that the Arg249Gln mutation may be responsible for high mortality.展开更多
Background:Colorectal cancer(CRC)is common and deadly,often leading to metastasis,challenging treatment,and poor outcomes.Understanding its molecular basis is crucial for developing effective therapies.Aims:This study...Background:Colorectal cancer(CRC)is common and deadly,often leading to metastasis,challenging treatment,and poor outcomes.Understanding its molecular basis is crucial for developing effective therapies.Aims:This study aimed to investigate the role of Myosin Heavy Chain 11(MYH11)in CRC progression,especially its effects on epithelial-mesenchymal transition(EMT)and cell behavior,and to explore its potential regulation by the EMT transcription factor zinc finger E-box binding homeobox 1(ZEB1).Methods:Differential expression analysis was performed in the GSE123390 and TCGA-READ datasets,and 317 intersection genes were identified.The hub gene MYH11 was identified based on Protein-protein interaction(PPI)analysis and expression validation.The effects of MYH11 and the EMT transcription factor(ZEB1)on the behavior of CRC cells were investigated in vitro.Results:Bioinformatics research revealed that MYH11 was considerably downregulated in CRC samples as compared to normal samples.Overexpression of MYH11 inhibited the proliferation,migration,and invasion of CRC cells.Western blotting(WB)testing showed that MYH11 overexpression inhibited EMT by elevating E-cadherin levels while suppressing ZEB1,vimentin,and N-cadherin expressions.By contrast,overexpression of ZEB1 promoted EMT and enhanced migration,invasion,and proliferation of CRC cells.The negative impacts of MYH11 affecting EMT markers and cell behaviors were partially mitigated by co-overexpression of MYH11 and ZEB1,indicating that MYH11 regulates EMT and CRC progression through ZEB1.Conclusion:Our study shows MYH11 curbs CRC growth by blocking EMT and invasion,but ZEB1 overexpression reduces this effect.It uncovers key CRC pathways and suggests MYH11’s therapeutic potential.展开更多
文摘为分析E3泛素连接酶环指蛋白125(ring finger protein 125,RNF125)在肝癌中的表达及临床预后意义,并探究RNF125对肝癌侵袭和转移的影响及其作用机制,通过ENCORI数据库和临床组织样本分析RNF125在肝癌和癌旁组织中的表达量,使用ENCORI数据库分析RNF125与肝癌预后的关系。构建敲低和过表达RNF125的肝癌细胞系,通过Transwell试验和细胞划痕试验检验RNF125表达水平对肝癌细胞迁移、侵袭能力的影响。通过动物试验验证调控RNF125对体内肿瘤生长情况及对肿瘤免疫微环境的影响,质谱分析寻找RNF125的靶蛋白,qPCR、蛋白质免疫印迹技术、泛素化试验验证RNF125与MYH9之间的关系,通过蛋白质免疫印迹和细胞免疫荧光技术确定调控RNF125与MYH9对EMT标志蛋白的影响。结果表明:RNF125在肝癌组织中表达较低,RNF125的高表达提示良好的预后。RNF125的表达水平与肝癌细胞迁移和侵袭能力呈负相关。过表达RNF125使肿瘤生长延迟并增加免疫细胞的浸润水平。RNF125特异性结合MYH9并促进其泛素化降解。RNF125与上皮细胞标志物的表达水平呈正相关,与间充质细胞标志物呈负相关。过表达MYH9可逆转RNF125对EMT标志蛋白的影响。综上,RNF125在肝癌中表达下调,RNF125的高表达与肝癌患者的良好预后相关,RNF125通过泛素化MYH9来抑制肝癌的EMT过程,从而抑制肝癌的迁移和侵袭。
文摘为了解猪脂肪组织肌球蛋白重链3(myosin heavy chain 3,MYH3)基因启动子区DNA甲基化及其mRNA表达水平在生长过程中的变化规律,试验采用亚硫酸氢盐测序(BSP)法分析6日龄和8月龄吉林花猪、松辽黑猪、二元猪脂肪组织MYH3基因启动子区DNA甲基化水平,利用实时荧光定量PCR方法分析3个品种猪脂肪的MYH3基因mRNA表达量。结果表明:6日龄吉林花猪、松辽黑猪、二元猪脂肪的MYH3基因启动子区DNA甲基化水平分别为88.4%、89.3%、87.5%,3个品种间DNA甲基化水平差异不显著(P>0.05),8月龄吉林花猪、松辽黑猪、二元猪MYH3基因启动子区DNA甲基化水平分别为77.7%、76.8%、83.9%,吉林花猪和松辽黑猪DNA甲基化水平显著低于二元猪(P<0.05),吉林花猪与松辽黑猪差异不显著(P>0.05);8月龄吉林花猪、松辽黑猪脂肪的MYH3基因启动子区DNA甲基化水平显著低于6日龄(P<0.05),而二元猪DNA甲基化水平在6日龄和8月龄差异不显著(P>0.05)。6日龄时,3个品种猪脂肪MYH3基因的mRNA表达量差异不显著(P>0.05);8月龄时,吉林花猪MYH3基因的mRNA相对表达量显著高于二元猪和松辽黑猪(P<0.05),松辽黑猪显著高于二元猪(P<0.05)。8月龄吉林花猪和松辽黑猪脂肪MYH3基因的mRNA表达量显著高于6日龄(P<0.05),而二元猪MYH3基因的mRNA表达量在6日龄和8月龄差异不显著(P>0.05)。说明随着年龄增长,吉林花猪、松辽黑猪脂肪的MYH3基因启动子区DNA甲基化水平与mRNA表达水平呈负相关,而二元猪无显著变化,地方品种猪在肉质、风味方面优于引进品种猪,推测启动子区DNA甲基化水平与脂肪生成和沉积具有相关性。
基金Supported by Beijing Municipal Natural Science Foundation(No.7202229No.7242168)China Primary Health Care Foundation(No.MTP2022C025).
文摘AIM:To explore the role of a previously-found MYH9 tail domain mutation(p.E1384Q)in the pathogenesis of congenital cataract.METHODS:The cell experiments were conducted in vitro.Wild-type(WT)MYH9 and p.E1384Q mutant fragments were constructed,which was then transiently transfected into Hek293T cell lines.Western blotting and quantitative real time polymerase chain reaction(qRT-PCR)were used to analyze the protein and mRNA level of non-muscle myosin IIA(NM IIA)and F-actin in transfected cells,and fluorescence microscopy was applied to explore the subcellular localization of NM IIA and F-actin.Cell counting kit-8(CCK8),woundhealing and double staining flow cytometry assays were performed to evaluate the proliferation,migration and apoptosis function of transfected cells,respectively.Transmission electron microscope was conducted to observe the alteration of organelle structure.RESULTS:The transiently-transfected WT and p.E1384Q mutant Hek293T cell lines was constructed.Western blot demonstrated that,comparing with MYH9WT group,the relative protein amount of NM IIA and F-actin significantly decreased in MYH9E1384Q cells(P<0.001).qRT-PCR analysis revealed that the relative mRNA amount of NM IIA and F-actin also significantly reduced in MYH9E1384Q cells when compared with MYH9WT.The immunofluorescence microscopy showed that the fluorescence signal of NM IIA and F-actin significantly decreased in E1384Q cells.The diffuse cytoplasmic distribution of NM IIA in MYH9WT was changed to be clumped distribution,presenting a“speckled”pattern characterized by aggregates of small size in MYH9E1384Q.Functional study revealed that the E1384Q mutation significantly inhibited cell proliferation(P=0.003)and migration(P<0.001),and promoted apoptosis(P<0.001).Electron microscope showed that the mutation remarkably decreased the number of mitochondria(P<0.001)and changed the phenotype of mitochondria.CONCLUSION:The missense gene mutation in MYH9(p.E1384Q)causing congenital cataract results in decreased amount and altered subcellular distribution of NM IIA and F-actin,accompanied by decreased cell proliferation and migration,promotes apoptosis and mitochondrial alteration.
基金Supported by The National High Level Hospital Clinical Research Funding,No.2022-PUMCH-B-129.
文摘BACKGROUND Chronic intestinal pseudo-obstruction(CIPO)is a rare and debilitating disorder,characterized by severe impairments in gastrointestinal motility.The affected sites include the enteric/intrinsic autonomic nerves(neuropathy),intestinal smooth muscle cells(myopathy),and interstitial cells of Cajal(mesenchymopathy).The etiology can be genetic,idiopathic,or acquired.Owing to its nonspecific clinical presentation and lack of definitive diagnostic methods,misdiagnosis of CIPO is common.CASE SUMMARY This case involved an older male with insidious onset in adolescence who presented with postprandial bloating,intermittent diarrhea,and weight loss.During the disease course,the patient experienced two episodes of intestinal obstruction.Imaging revealed multisegmental digestive tract abnormalities(gastric emptying disorder,significant duodenal dilatation,and segmental jejunal dilatation).Whole-exome sequencing revealed a rare MYH11 mutation[NM_0010-40113.2:C.5819del(p.Pro1940HisfsTer91)],confirming hereditary myopathic CIPO.CONCLUSION This report adds to our current understanding of CIPO etiology by reinforcing the role of MYH11 variants in the pathogenesis of the CIPO phenotype.
基金Supported by National Natural Science Foundation of China,No.81770379.
文摘BACKGROUND Hypertrophic cardiomyopathy(HCM)is one of the most prevalent inherited myocardial disorders and is charac-terized by considerable genetic and phenotypic heterogeneity.A subset of patients with HCM progress to a dilated phase of HCM(DPHCM),which is associated with a poor prognosis;however,the underlying pathogenesis remains inadequately understood.CASE SUMMARY In this study,we present a case involving a pedigree with familial DPHCM and conduct a retrospective review of patients with DPHCM with identified gene mutations.Through panel sequencing targeting the coding regions of 312 genes associated with inherited cardiomyopathy,a heterozygous missense mutation(c.746G>A,p.Arg249Glu)in the MYH7 gene was identified in the proband(III-5).Sanger sequencing subsequently confirmed this pathogenic mutation in three additional family members(II-4,III-4,and IV-3).A total of 26 well-documented patients with DPHCM were identified in the literature.Patients with DPHCM are commonly middle-aged and male.The mean age of patients with DPHCM was 53.43±12.79 years.Heart failure,dyspnoea,and atrial fibrillation were the most prevalent symptoms observed,accompanied by an average left ventricular end-diastolic size of 58.62 mm.CONCLUSION Our findings corroborate the pathogenicity of the MYH7(c.746G>A,p.Arg249Glu)mutation for DPHCM and suggest that the Arg249Gln mutation may be responsible for high mortality.
基金funded by Outstanding Leaders Training Programof Pudong Health Commission of Shanghai(No.PWR12023-03)In-house Project of Shanghai Pudong NewArea People’sHospital(No.E24-02).
文摘Background:Colorectal cancer(CRC)is common and deadly,often leading to metastasis,challenging treatment,and poor outcomes.Understanding its molecular basis is crucial for developing effective therapies.Aims:This study aimed to investigate the role of Myosin Heavy Chain 11(MYH11)in CRC progression,especially its effects on epithelial-mesenchymal transition(EMT)and cell behavior,and to explore its potential regulation by the EMT transcription factor zinc finger E-box binding homeobox 1(ZEB1).Methods:Differential expression analysis was performed in the GSE123390 and TCGA-READ datasets,and 317 intersection genes were identified.The hub gene MYH11 was identified based on Protein-protein interaction(PPI)analysis and expression validation.The effects of MYH11 and the EMT transcription factor(ZEB1)on the behavior of CRC cells were investigated in vitro.Results:Bioinformatics research revealed that MYH11 was considerably downregulated in CRC samples as compared to normal samples.Overexpression of MYH11 inhibited the proliferation,migration,and invasion of CRC cells.Western blotting(WB)testing showed that MYH11 overexpression inhibited EMT by elevating E-cadherin levels while suppressing ZEB1,vimentin,and N-cadherin expressions.By contrast,overexpression of ZEB1 promoted EMT and enhanced migration,invasion,and proliferation of CRC cells.The negative impacts of MYH11 affecting EMT markers and cell behaviors were partially mitigated by co-overexpression of MYH11 and ZEB1,indicating that MYH11 regulates EMT and CRC progression through ZEB1.Conclusion:Our study shows MYH11 curbs CRC growth by blocking EMT and invasion,but ZEB1 overexpression reduces this effect.It uncovers key CRC pathways and suggests MYH11’s therapeutic potential.
