CCK correlates with the generation and progression of pancreatic cancer. The research aims to construct eukaryotic expression plasmid pIRES2-EGFP/CCK (CCK pDNA) and transiently express it in COS-7 cells. Total RNA was...CCK correlates with the generation and progression of pancreatic cancer. The research aims to construct eukaryotic expression plasmid pIRES2-EGFP/CCK (CCK pDNA) and transiently express it in COS-7 cells. Total RNA was extracted from porcine intestinal mucosa. RT-PCR was used to amplify the aimed segments CCKcDNA which was then digested with EcoR1 and BamH1 and inserted into a eukaryotic expression plasmid pIRES2-EGFP to construct CCK pDNA. The con- structed plasmid was transfected into COS-7 cells by lepofectamineTM2000-mediated transfer method. The expression of CCK in transfected COS-7 cells was detected 24, 48 and 72 h post-transfection with fluorescence microscopy and the expression level of CCK mRNA in transfected COS-7 cells was assayed by using RT-PCR. The results showed CCK pDNA was successfully constructed and expressed transiently in COS-7 cells. Green fluorescent protein could be detected in the COS-7 cells transfected with porcine CCK pDNA 24 h post-transfection. At 48th h post-transfection, the number of positive cells was increased significantly and much brighter green fluorescence could be detected. And 72 h post-transfection, the green fluorescence of positive cells became even stronger, while no green fluorescence was detected in the control group. The expression of CCK mRNA in the cells was detectable by using RT-PCR. In COS-7 cells transfected with CCK pDNA a high level of porcine CCK mRNA was detected while no expression of porcine CCKmRNA was found in the cells trans- fected with null plasmid. It was concluded CCK pDNA was expressed successfully in COS-7 cells, which lays a foundation for further research on the relationship between CCK and tumor.展开更多
We evaluated whether the dodecylamine derivative of hydroxocobalamin acts as a potent inhibitor of cobalamin-dependent enzymes in an African green monkey kidney cell, COS-7. When the dodecylamine derivative (1.0 μmol...We evaluated whether the dodecylamine derivative of hydroxocobalamin acts as a potent inhibitor of cobalamin-dependent enzymes in an African green monkey kidney cell, COS-7. When the dodecylamine derivative (1.0 μmol/L) did not show any cytotoxicity in the cultured cells, the derivative could not affect methylmalonyl-CoA mutase (holo-enzyme) activity, but significantly inhibit methionine synthase (holo-enzyme) activity in the cell homogenates of COS-7 grown in 1.0 μmol/L hydroxocobalamin-supplemented medium. An immunoblot analysis indicated that the dodecylamine derivative could not decrease the protein level of methionine synthase, but significantly inhibit the enzyme activity.展开更多
To establish a cell line for stable expression of human beta-defensin 3 (hBD3). Methods Full length cDNA of hBD3 was isolated from previously constructed pGEM-hBD3 and then inserted into pcDNA3. The recombinant vector...To establish a cell line for stable expression of human beta-defensin 3 (hBD3). Methods Full length cDNA of hBD3 was isolated from previously constructed pGEM-hBD3 and then inserted into pcDNA3. The recombinant vector identified carrying hBD3 with right direction was introduced into COS-7 cells by Lipofe-ctamine. Cell clones survived in G418-rich medium and with stable expression of hBD3 in both mRNA and protein levels were identified by RT-PCR and Western blot respectively. Genomic integration of the hBD3 gene with the COS-7 cells was confirmed by Southern dot blot and primary analysis. The antimicrobial activity of the secreted hBD3 was also evaluated. Results COS-7 cells transfected with pcDNA3-hBD3 expressed hBD3 stably in mRNA and protein level. Southern dot blot analysis showed successful integration of the hBD3 gene into the genome of COS-7 cell and the hBD-3 protein secreted into the culture medium showed antimicrobial activity. Conclusion We successfully established a hBD3-expressing cell line.展开更多
Background:Vascular smooth muscle cells(VSMCs)undergo a conversion from a contractile phenotype to a proliferative synthetic phenotype,contributing to the pathogenesis of cardiovascular diseases.Semaphorin 7A(SEMA7A)i...Background:Vascular smooth muscle cells(VSMCs)undergo a conversion from a contractile phenotype to a proliferative synthetic phenotype,contributing to the pathogenesis of cardiovascular diseases.Semaphorin 7A(SEMA7A)is a glycosylphosphatidylinositol-anchored membrane protein that plays an important role in vascular homeostasis by regulating endothelial cell behaviors.However,the expression and role of SEMA7A in VSMCs remain unclear.Methods:In this study,we screened for VSMC-regulating genes in publicly available datasets and analyzed the expression of SEMA7A in human coronary artery smooth muscle cells(hCASMCs)treated with platelet-derived growth factor-BB(PDGF-BB).The effects of SEMA7A overexpression and knockdown on hCASMC proliferation and migration were examined.The signaling pathways involved in the action of SEMA7A in hCASMCs were determined.Results:Bioinformatic analysis showed that SEMA7A was significantly dysregulated in VSMCs treated with oxidized low-density lipoprotein or overexpressing progerin,a pro-atherogenic gene.The PDGF-BB stimulation led to a concentration-and time-dependent induction of SEMA7A.Depletion of SEMA7A attenuated PDGF-BB-induced hCASMC proliferation and migration.Conversely,overexpression of SEMA7A enhanced hCASMC proliferation and migration.Mechanistically,SEMA7A stimulated the activation of theβ-catenin pathway and upregulated c-Myc,CCND1,and MMP7.Knockdown ofβ-catenin impaired SEMA7A-induced hCASMC proliferation and migration.Conclusions:SEMA7A triggers phenotype switching in VSMCs through theβ-catenin signaling pathway and may serve as a potential therapeutic target for cardiovascular diseases.展开更多
Objective:To explore the regulatory mechanism of transient receptor potential melastatin-7(TRPM7)in high glucose-induced renal tubular epithelial cell injury.Methods:The expression of TRPM7 in the serum of diabetic ne...Objective:To explore the regulatory mechanism of transient receptor potential melastatin-7(TRPM7)in high glucose-induced renal tubular epithelial cell injury.Methods:The expression of TRPM7 in the serum of diabetic nephropathy patients and high glucose-induced HK-2 cells was detected by RT-qPCR.Then,the TRPM7 interference vector was constructed,and the downstream high mobility group box 1(HMGB1)/Toll-like receptor 4(TLR4)signaling pathway proteins were detected.Next,in addition to interference with TRPM7 expression,overexpression of HMGB1 in high glucose-induced HK-2 cells was performed.Cell activity,apoptosis,oxidative stress levels,and inflammation levels were determined by CCK8,TUNEL,Western blotting,immunofluorescence and related kits.Results:TRPM7 expression was upregulated in the serum of diabetic nephropathy patients and high glucose-induced HK-2 cells.Interference with TRPM7 reduced cell damage,epithelial-mesenchymal transition,oxidative stress,and inflammatory response in high glucose-induced HK-2 cells via inhibiting the HMGB1/TLR4 signaling pathway.However,the effects induced by TRPM7 silencing were abrogated by HMGB1 overexpression.Conclusions:Decreased TRPM7 alleviates high glucose-induced renal tubular epithelial cell injury by inhibiting the HMGB1/TLR4 signaling pathway.Further animal experiments and clinical trials are warranted to verify its effect.展开更多
Objective:Through integrated bioinformatics analysis,the goal of this work was to find new,characterised N7-methylguanosine modification-related long non-coding RNAs(m7G-lncRNAs)that might be used to predict the progn...Objective:Through integrated bioinformatics analysis,the goal of this work was to find new,characterised N7-methylguanosine modification-related long non-coding RNAs(m7G-lncRNAs)that might be used to predict the prognosis of laryngeal squamous cell carcinoma(LSCC).Methods:The clinical data and LSCC gene expression data for the current investigation were initially retrieved from the TCGA database&sanitised.Then,using co-expression analysis of m7G-associated mRNAs&lncRNAs&differential expression analysis(DEA)among LSCC&normal sample categories,we discovered lncRNAs that were connected to m7G.The prognosis prediction model was built for the training category using univariate&multivariate COX regression&LASSO regression analyses,&the model’s efficacy was checked against the test category data.In addition,we conducted DEA of prognostic m7G-lncRNAs among LSCC&normal sample categories&compiled a list of co-expression networks&the structure of prognosis m7G-lncRNAs.To compare the prognoses for individuals with LSCC in the high-&low-risk categories in the prognosis prediction model,survival and risk assessments were also carried out.Finally,we created a nomogram to accurately forecast the outcomes of LSCC patients&created receiver operating characteristic(ROC)curves to assess the prognosis prediction model’s predictive capability.Results:Using co-expression network analysis&differential expression analysis,we discovered 774 m7G-lncRNAs and 551 DEm7G-lncRNAs,respectively.We then constructed a prognosis prediction model for six m7G-lncRNAs(FLG−AS1,RHOA−IT1,AC020913.3,AC027307.2,AC010973.2 and AC010789.1),identified 32 DEPm7G-lncRNAs,analyzed the correlation between 32 DEPm7G-lncRNAs and 13 DEPm7G-mRNAs,and performed survival analyses and risk analyses of the prognosis prediction model to assess the prognostic performance of LSCC patients.By displaying ROC curves and a nomogram,we finally checked the prognosis prediction model's accuracy.Conclusion:By creating novel predictive lncRNA signatures for clinical diagnosis&therapy,our findings will contribute to understanding the pathogenetic process of LSCC.展开更多
AIM:To investigate the expression and effect of histone deacetylase 7(HDAC7)in human retinal microvascular endothelial cells(HRMECs)under high glucose condition and related mechanism,and the expression of HDAC7 in the...AIM:To investigate the expression and effect of histone deacetylase 7(HDAC7)in human retinal microvascular endothelial cells(HRMECs)under high glucose condition and related mechanism,and the expression of HDAC7 in the retinal tissue in diabetic rats.METHODS:The expression of HDAC7 in HRMECs under high glucose and the retinal tissue from normal or diabetic rats were detected with immunohistochemistry and Western blot.LV-shHDAC7 HRMECs were used to study the effect of HDAC7 on cell activities.Cell count kit-8(CCK-8),5-ethynyl2’-deoxyuridine(EdU),flow cytometry,scratch test,Transwell test and tube formation assay were used to examine the ability of cell proliferation,migration,and angiogenesis.Finally,a preliminary exploration of its mechanism was performed by Western blot.RESULTS:The expression of HDAC7 was both upregulated in retinal tissues of diabetic rats and high glucosetreated HRMECs.Down-regulation of HDAC7 expression significantly reduced the ability of proliferation,migration,and tube formation,and reversed the high glucose-induced high expression of CDK1/Cyclin B1 and vascular endothelial growth factor in high glucose-treated HRMECs.CONCLUSION:High glucose can up-regulate the expression of HDAC7 in HRMECs.Down-regulation of HDAC7 can inhibit HRMECs activities.HDAC7 is proposed to be involved in pathogenesis of diabetic retinopathy and a therapeutic target.展开更多
Objective Pseudogenes are initially regarded as nonfunctional genomic sequences,but some pseudogenes regulate tumor initiation and progression by interacting with other genes to modulate their transcriptional activiti...Objective Pseudogenes are initially regarded as nonfunctional genomic sequences,but some pseudogenes regulate tumor initiation and progression by interacting with other genes to modulate their transcriptional activities.Olfactory receptor family 7 subfamily E member 47 pseudogene(OR7E47P)is expressed broadly in lung tissues and has been identified as a positive regulator in the tumor microenvironment(TME)of lung adenocarcinoma(LUAD).This study aimed to elucidate the correlation between OR7E47P and tumor immunity in lung squamous cell carcinoma(LUSC).Methods Clinical and molecular information from The Cancer Genome Atlas(TCGA)LUSC cohort was used to identify OR7E47P-related immune genes(ORIGs)by weighted gene correlation network analysis(WGCNA).Based on the ORIGs,2 OR7E47P clusters were identified using non-negative matrix factorization(NMF)clustering,and the stability of the clustering was tested by an extreme gradient boosting classifier(XGBoost).LASSO-Cox and stepwise regressions were applied to further select prognostic ORIGs and to construct a predictive model(ORPScore)for immunotherapy.The Botling cohorts and 8 immunotherapy cohorts(the Samstein,Braun,Jung,Gide,IMvigor210,Lauss,Van Allen,and Cho cohorts)were included as independent validation cohorts.Results OR7E47P expression was positively correlated with immune cell infiltration and enrichment of immune-related pathways in LUSC.A total of 57 ORIGs were identified to classify the patients into 2 OR7E47P clusters(Cluster 1 and Cluster 2)with distinct immune,mutation,and stromal programs.Compared to Cluster 1,Cluster 2 had more infiltration by immune and stromal cells,lower mutation rates of driver genes,and higher expression of immune-related proteins.The clustering performed well in the internal and 5 external validation cohorts.Based on the 7 ORIGs(HOPX,STX2,WFS,DUSP22,SLFN13,GGCT,and CCSER2),the ORPScore was constructed to predict the prognosis and the treatment response.In addition,the ORPScore was a better prognostic factor and correlated positively with the immunotherapeutic response in cancer patients.The area under the curve values ranged from 0.584 to 0.805 in the 6 independent immunotherapy cohorts.Conclusion Our study suggests a significant correlation between OR7E47P and TME modulation in LUSC.ORIGs can be applied to molecularly stratify patients,and the ORPScore may serve as a biomarker for clinical decision-making regarding individualized prognostication and immunotherapy.展开更多
Objective:To determine the destructive ability of oxocrebanine,an anti-breast cancer active compound obtained from Stephania hainanensis H.S.Lo et Y.Tsoong,on microtubule network,and investigate the effect of oxocreba...Objective:To determine the destructive ability of oxocrebanine,an anti-breast cancer active compound obtained from Stephania hainanensis H.S.Lo et Y.Tsoong,on microtubule network,and investigate the effect of oxocrebanine on microtubule network homeostasis at both molecular and cellular levels.Methods:the EBI site competition method and molecular docking method were used to determine the occupation of the microtubule site of oxocrebanine.Western Blot was used to detect the effect of oxocrebanine on microtubule-associated proteins including STAT3,PAK1,CAMK4,and PKA.Results:The results of EBI site competition assay showed that the binding of EBI toβ-Tubulin covalent fusions produced adducts that appeared in regions of lower molecular weight thanβ-tubulin(ctrl 2).Molecular docking results showed that oxocrebanine could occupy the colchicine site of microtubule proteins.As revealed by Western Blot,the expression of STAT3 protein was decreased after MCF-7 cells have been treated with low,medium,and high concentration of oxocrebanine or the positive drug taxol for 48 h(P<0.01).The expression levels of PAK1 and Camk4 proteins aslo showed significant reductions(P<0.05,or P<0.01).Oxocrebanine also decreased the PKA protein in MCF-7 cells compared to the control group(P<0.01).Conclusions:Oxocrebanine,a ligand that binds at the colchicine site of tubulin,perturbs tubulin polymerization and causes mitosis in MCF-7 cells,thus leading to MCF-7 cell death.Oxocrebanine may promote microtubule dynamics through stathmin by inhibiting the expression levels of STAT3,PAK1,Camk4,and PKA proteins in MCF-7 cells.Oxocrebanine interfers with spindle formation,and ultimately causes mitotic catastrophe in MCF-7 cells.展开更多
基金a grant from National Natu-ral Sciences Foundation of China (No. 30170917)
文摘CCK correlates with the generation and progression of pancreatic cancer. The research aims to construct eukaryotic expression plasmid pIRES2-EGFP/CCK (CCK pDNA) and transiently express it in COS-7 cells. Total RNA was extracted from porcine intestinal mucosa. RT-PCR was used to amplify the aimed segments CCKcDNA which was then digested with EcoR1 and BamH1 and inserted into a eukaryotic expression plasmid pIRES2-EGFP to construct CCK pDNA. The con- structed plasmid was transfected into COS-7 cells by lepofectamineTM2000-mediated transfer method. The expression of CCK in transfected COS-7 cells was detected 24, 48 and 72 h post-transfection with fluorescence microscopy and the expression level of CCK mRNA in transfected COS-7 cells was assayed by using RT-PCR. The results showed CCK pDNA was successfully constructed and expressed transiently in COS-7 cells. Green fluorescent protein could be detected in the COS-7 cells transfected with porcine CCK pDNA 24 h post-transfection. At 48th h post-transfection, the number of positive cells was increased significantly and much brighter green fluorescence could be detected. And 72 h post-transfection, the green fluorescence of positive cells became even stronger, while no green fluorescence was detected in the control group. The expression of CCK mRNA in the cells was detectable by using RT-PCR. In COS-7 cells transfected with CCK pDNA a high level of porcine CCK mRNA was detected while no expression of porcine CCKmRNA was found in the cells trans- fected with null plasmid. It was concluded CCK pDNA was expressed successfully in COS-7 cells, which lays a foundation for further research on the relationship between CCK and tumor.
文摘We evaluated whether the dodecylamine derivative of hydroxocobalamin acts as a potent inhibitor of cobalamin-dependent enzymes in an African green monkey kidney cell, COS-7. When the dodecylamine derivative (1.0 μmol/L) did not show any cytotoxicity in the cultured cells, the derivative could not affect methylmalonyl-CoA mutase (holo-enzyme) activity, but significantly inhibit methionine synthase (holo-enzyme) activity in the cell homogenates of COS-7 grown in 1.0 μmol/L hydroxocobalamin-supplemented medium. An immunoblot analysis indicated that the dodecylamine derivative could not decrease the protein level of methionine synthase, but significantly inhibit the enzyme activity.
文摘To establish a cell line for stable expression of human beta-defensin 3 (hBD3). Methods Full length cDNA of hBD3 was isolated from previously constructed pGEM-hBD3 and then inserted into pcDNA3. The recombinant vector identified carrying hBD3 with right direction was introduced into COS-7 cells by Lipofe-ctamine. Cell clones survived in G418-rich medium and with stable expression of hBD3 in both mRNA and protein levels were identified by RT-PCR and Western blot respectively. Genomic integration of the hBD3 gene with the COS-7 cells was confirmed by Southern dot blot and primary analysis. The antimicrobial activity of the secreted hBD3 was also evaluated. Results COS-7 cells transfected with pcDNA3-hBD3 expressed hBD3 stably in mRNA and protein level. Southern dot blot analysis showed successful integration of the hBD3 gene into the genome of COS-7 cell and the hBD-3 protein secreted into the culture medium showed antimicrobial activity. Conclusion We successfully established a hBD3-expressing cell line.
基金Acknowledgment This work was supported by grants from the National Natural Science Foundation (30671563, 30700597) and Key Project of Science and Technology of Gansu Province (0801NKDA034) and by gifts from the Development Planning Project of Science and Technology of Lanzhou (07-2-12, 2008- l- 167).
基金supported by the Basic Research Program of Shanxi Province(Free Exploration)of China(20210302124416)Science and Technology Grant for Selected Returned Chinese Scholars of Shanxi Province of China(20220043)Four“Batches”Innovation Project of Invigorating Medical through Science and Technology of Shanxi Province of China(2022XM08).
文摘Background:Vascular smooth muscle cells(VSMCs)undergo a conversion from a contractile phenotype to a proliferative synthetic phenotype,contributing to the pathogenesis of cardiovascular diseases.Semaphorin 7A(SEMA7A)is a glycosylphosphatidylinositol-anchored membrane protein that plays an important role in vascular homeostasis by regulating endothelial cell behaviors.However,the expression and role of SEMA7A in VSMCs remain unclear.Methods:In this study,we screened for VSMC-regulating genes in publicly available datasets and analyzed the expression of SEMA7A in human coronary artery smooth muscle cells(hCASMCs)treated with platelet-derived growth factor-BB(PDGF-BB).The effects of SEMA7A overexpression and knockdown on hCASMC proliferation and migration were examined.The signaling pathways involved in the action of SEMA7A in hCASMCs were determined.Results:Bioinformatic analysis showed that SEMA7A was significantly dysregulated in VSMCs treated with oxidized low-density lipoprotein or overexpressing progerin,a pro-atherogenic gene.The PDGF-BB stimulation led to a concentration-and time-dependent induction of SEMA7A.Depletion of SEMA7A attenuated PDGF-BB-induced hCASMC proliferation and migration.Conversely,overexpression of SEMA7A enhanced hCASMC proliferation and migration.Mechanistically,SEMA7A stimulated the activation of theβ-catenin pathway and upregulated c-Myc,CCND1,and MMP7.Knockdown ofβ-catenin impaired SEMA7A-induced hCASMC proliferation and migration.Conclusions:SEMA7A triggers phenotype switching in VSMCs through theβ-catenin signaling pathway and may serve as a potential therapeutic target for cardiovascular diseases.
文摘Objective:To explore the regulatory mechanism of transient receptor potential melastatin-7(TRPM7)in high glucose-induced renal tubular epithelial cell injury.Methods:The expression of TRPM7 in the serum of diabetic nephropathy patients and high glucose-induced HK-2 cells was detected by RT-qPCR.Then,the TRPM7 interference vector was constructed,and the downstream high mobility group box 1(HMGB1)/Toll-like receptor 4(TLR4)signaling pathway proteins were detected.Next,in addition to interference with TRPM7 expression,overexpression of HMGB1 in high glucose-induced HK-2 cells was performed.Cell activity,apoptosis,oxidative stress levels,and inflammation levels were determined by CCK8,TUNEL,Western blotting,immunofluorescence and related kits.Results:TRPM7 expression was upregulated in the serum of diabetic nephropathy patients and high glucose-induced HK-2 cells.Interference with TRPM7 reduced cell damage,epithelial-mesenchymal transition,oxidative stress,and inflammatory response in high glucose-induced HK-2 cells via inhibiting the HMGB1/TLR4 signaling pathway.However,the effects induced by TRPM7 silencing were abrogated by HMGB1 overexpression.Conclusions:Decreased TRPM7 alleviates high glucose-induced renal tubular epithelial cell injury by inhibiting the HMGB1/TLR4 signaling pathway.Further animal experiments and clinical trials are warranted to verify its effect.
基金supported by a grant Hebei Provincial Health Commission project from the Foundation of Basic Research(No.20191843).
文摘Objective:Through integrated bioinformatics analysis,the goal of this work was to find new,characterised N7-methylguanosine modification-related long non-coding RNAs(m7G-lncRNAs)that might be used to predict the prognosis of laryngeal squamous cell carcinoma(LSCC).Methods:The clinical data and LSCC gene expression data for the current investigation were initially retrieved from the TCGA database&sanitised.Then,using co-expression analysis of m7G-associated mRNAs&lncRNAs&differential expression analysis(DEA)among LSCC&normal sample categories,we discovered lncRNAs that were connected to m7G.The prognosis prediction model was built for the training category using univariate&multivariate COX regression&LASSO regression analyses,&the model’s efficacy was checked against the test category data.In addition,we conducted DEA of prognostic m7G-lncRNAs among LSCC&normal sample categories&compiled a list of co-expression networks&the structure of prognosis m7G-lncRNAs.To compare the prognoses for individuals with LSCC in the high-&low-risk categories in the prognosis prediction model,survival and risk assessments were also carried out.Finally,we created a nomogram to accurately forecast the outcomes of LSCC patients&created receiver operating characteristic(ROC)curves to assess the prognosis prediction model’s predictive capability.Results:Using co-expression network analysis&differential expression analysis,we discovered 774 m7G-lncRNAs and 551 DEm7G-lncRNAs,respectively.We then constructed a prognosis prediction model for six m7G-lncRNAs(FLG−AS1,RHOA−IT1,AC020913.3,AC027307.2,AC010973.2 and AC010789.1),identified 32 DEPm7G-lncRNAs,analyzed the correlation between 32 DEPm7G-lncRNAs and 13 DEPm7G-mRNAs,and performed survival analyses and risk analyses of the prognosis prediction model to assess the prognostic performance of LSCC patients.By displaying ROC curves and a nomogram,we finally checked the prognosis prediction model's accuracy.Conclusion:By creating novel predictive lncRNA signatures for clinical diagnosis&therapy,our findings will contribute to understanding the pathogenetic process of LSCC.
基金Supported by the Shaanxi Province Traditional Chinese Medicine Project(No.SZY-KJCYC-2023-028)。
文摘AIM:To investigate the expression and effect of histone deacetylase 7(HDAC7)in human retinal microvascular endothelial cells(HRMECs)under high glucose condition and related mechanism,and the expression of HDAC7 in the retinal tissue in diabetic rats.METHODS:The expression of HDAC7 in HRMECs under high glucose and the retinal tissue from normal or diabetic rats were detected with immunohistochemistry and Western blot.LV-shHDAC7 HRMECs were used to study the effect of HDAC7 on cell activities.Cell count kit-8(CCK-8),5-ethynyl2’-deoxyuridine(EdU),flow cytometry,scratch test,Transwell test and tube formation assay were used to examine the ability of cell proliferation,migration,and angiogenesis.Finally,a preliminary exploration of its mechanism was performed by Western blot.RESULTS:The expression of HDAC7 was both upregulated in retinal tissues of diabetic rats and high glucosetreated HRMECs.Down-regulation of HDAC7 expression significantly reduced the ability of proliferation,migration,and tube formation,and reversed the high glucose-induced high expression of CDK1/Cyclin B1 and vascular endothelial growth factor in high glucose-treated HRMECs.CONCLUSION:High glucose can up-regulate the expression of HDAC7 in HRMECs.Down-regulation of HDAC7 can inhibit HRMECs activities.HDAC7 is proposed to be involved in pathogenesis of diabetic retinopathy and a therapeutic target.
基金the Wuhan University Medical Faculty Innovation Seed Fund Cultivation Project(No.TFZZ2018025)the Chen Xiao-ping Foundation for the Development of Science and Technology of Hubei Province(No.CXPJJH12000001-2020313)the National Natural Science Foundation of China(No.81670123 and No.81670144).
文摘Objective Pseudogenes are initially regarded as nonfunctional genomic sequences,but some pseudogenes regulate tumor initiation and progression by interacting with other genes to modulate their transcriptional activities.Olfactory receptor family 7 subfamily E member 47 pseudogene(OR7E47P)is expressed broadly in lung tissues and has been identified as a positive regulator in the tumor microenvironment(TME)of lung adenocarcinoma(LUAD).This study aimed to elucidate the correlation between OR7E47P and tumor immunity in lung squamous cell carcinoma(LUSC).Methods Clinical and molecular information from The Cancer Genome Atlas(TCGA)LUSC cohort was used to identify OR7E47P-related immune genes(ORIGs)by weighted gene correlation network analysis(WGCNA).Based on the ORIGs,2 OR7E47P clusters were identified using non-negative matrix factorization(NMF)clustering,and the stability of the clustering was tested by an extreme gradient boosting classifier(XGBoost).LASSO-Cox and stepwise regressions were applied to further select prognostic ORIGs and to construct a predictive model(ORPScore)for immunotherapy.The Botling cohorts and 8 immunotherapy cohorts(the Samstein,Braun,Jung,Gide,IMvigor210,Lauss,Van Allen,and Cho cohorts)were included as independent validation cohorts.Results OR7E47P expression was positively correlated with immune cell infiltration and enrichment of immune-related pathways in LUSC.A total of 57 ORIGs were identified to classify the patients into 2 OR7E47P clusters(Cluster 1 and Cluster 2)with distinct immune,mutation,and stromal programs.Compared to Cluster 1,Cluster 2 had more infiltration by immune and stromal cells,lower mutation rates of driver genes,and higher expression of immune-related proteins.The clustering performed well in the internal and 5 external validation cohorts.Based on the 7 ORIGs(HOPX,STX2,WFS,DUSP22,SLFN13,GGCT,and CCSER2),the ORPScore was constructed to predict the prognosis and the treatment response.In addition,the ORPScore was a better prognostic factor and correlated positively with the immunotherapeutic response in cancer patients.The area under the curve values ranged from 0.584 to 0.805 in the 6 independent immunotherapy cohorts.Conclusion Our study suggests a significant correlation between OR7E47P and TME modulation in LUSC.ORIGs can be applied to molecularly stratify patients,and the ORPScore may serve as a biomarker for clinical decision-making regarding individualized prognostication and immunotherapy.
基金Natural Science Foundation of Hainan Province(No.820RC776)。
文摘Objective:To determine the destructive ability of oxocrebanine,an anti-breast cancer active compound obtained from Stephania hainanensis H.S.Lo et Y.Tsoong,on microtubule network,and investigate the effect of oxocrebanine on microtubule network homeostasis at both molecular and cellular levels.Methods:the EBI site competition method and molecular docking method were used to determine the occupation of the microtubule site of oxocrebanine.Western Blot was used to detect the effect of oxocrebanine on microtubule-associated proteins including STAT3,PAK1,CAMK4,and PKA.Results:The results of EBI site competition assay showed that the binding of EBI toβ-Tubulin covalent fusions produced adducts that appeared in regions of lower molecular weight thanβ-tubulin(ctrl 2).Molecular docking results showed that oxocrebanine could occupy the colchicine site of microtubule proteins.As revealed by Western Blot,the expression of STAT3 protein was decreased after MCF-7 cells have been treated with low,medium,and high concentration of oxocrebanine or the positive drug taxol for 48 h(P<0.01).The expression levels of PAK1 and Camk4 proteins aslo showed significant reductions(P<0.05,or P<0.01).Oxocrebanine also decreased the PKA protein in MCF-7 cells compared to the control group(P<0.01).Conclusions:Oxocrebanine,a ligand that binds at the colchicine site of tubulin,perturbs tubulin polymerization and causes mitosis in MCF-7 cells,thus leading to MCF-7 cell death.Oxocrebanine may promote microtubule dynamics through stathmin by inhibiting the expression levels of STAT3,PAK1,Camk4,and PKA proteins in MCF-7 cells.Oxocrebanine interfers with spindle formation,and ultimately causes mitotic catastrophe in MCF-7 cells.