BACKGROUND Gastric cancer(GC)is a common gastrointestinal malignancy worldwide.Based on cancer-related mortality,the current prevention and treatment strategies for GC still show poor clinical results.Therefore,it is ...BACKGROUND Gastric cancer(GC)is a common gastrointestinal malignancy worldwide.Based on cancer-related mortality,the current prevention and treatment strategies for GC still show poor clinical results.Therefore,it is important to find effective drug treatment targets.AIM To explore the molecular mechanism of 18β-glycyrrhetinic acid(18β-GRA)regulating the miR-345-5p/TGM2 signaling pathway to inhibit the proliferation of GC cells.METHODS CCK-8 assay was used to determine the effect of 18β-GRA on the survival rate of GES-1 cells and AGS and HGC-27 cells.Cell cycle and apoptosis were detected by flow cytometry,cell migration was detected by a wound healing assay,the effect of 18β-GRA on subcutaneous tumor growth in BALB/c nude mice was investigated,and the cell autophagy level was determined by MDC staining.TMT proteomic analysis was used to detect the differentially expressed autophagy-related proteins in GC cells after 18β-GRA intervention,and then the protein-protein interaction was predicted using STRING(https://string-db.org/).MicroRNAs(miRNAs)transcriptome analysis was used to detect the miRNA differential expression profile,and use miRBase(https://www.mirbase/)and TargetScan(https://www.targetscan.org/)to predict the miRNA and complementary binding sites.Quantitative real-time polymerase chain reaction was used to detect the expression level of miRNA in 18β-GRA treated cells,and western blot was used to detect the expression of autophagy related proteins.Finally,the effect of miR-345-5p on GC cells was verified by mir-345-5p overexpression.RESULTS 18β-GRA could inhibit GC cells viability,promote cell apoptosis,block cell cycle,reduce cell wound healing ability,and inhibit the GC cells growth in vivo.MDC staining results showed that 18β-GRA could promote autophagy in GC cells.By TMT proteomic analysis and miRNAs transcriptome analysis,it was concluded that 18β-GRA could down-regulate TGM2 expression and up-regulate miR-345-5p expression in GC cells.Subsequently,we verified that TGM2 is the target of miR-345-5p,and that overexpression of miR-345-5p significantly inhibited the protein expression level of TGM2.Western blot showed that the expression of autophagy-related proteins of TGM2 and p62 was significantly reduced,and LC3II,ULK1 and AMPK expression was significantly increased in GC cells treated with 18β-GRA.Overexpression of miR-345-5p not only inhibited the expression of TGM2,but also inhibited the proliferation of GC cells by promoting cell apoptosis and arresting cell cycle.CONCLUSION 18β-GRA inhibits the proliferation of GC cells and promotes autophagy by regulating the miR-345-5p/TGM2 signaling pathway.展开更多
Nb-RE微合金化TGM2高速钢(%:0.88～0.89C、4.14～4.16Cr、4.73～4.76Mo、6.09～6.12W、1.85～1.86V、0.05～0.10Nb、0.05～0.06RE)由25 t EAF-30 t LF(VD,加Nb-RE)-1 t ESR工艺冶炼。试验结果表明,经Nb-RE微合金化后,TGM2高速钢Φ96 mm...Nb-RE微合金化TGM2高速钢(%:0.88～0.89C、4.14～4.16Cr、4.73～4.76Mo、6.09～6.12W、1.85～1.86V、0.05～0.10Nb、0.05～0.06RE)由25 t EAF-30 t LF(VD,加Nb-RE)-1 t ESR工艺冶炼。试验结果表明,经Nb-RE微合金化后,TGM2高速钢Φ96 mm材淬火晶粒尺寸明显细化,晶粒度由原来未微合金化钢的9.5级提高至10～10.5级;淬、回火后硬度HRC为65.2～65.8,600℃4 h红硬性HRC为62.1～62.3,Nb-RE TGM2钢制成刀具的切削寿命较原TGM2钢提高20%。展开更多
TGM2A-S钢(/%:0.85C、0.27Si、0.24Mn、0.026P、0.007S、3.98Cr、4.76Mo、6.09W、1.83V、0.12Nb、0.03RE)是在高速钢TGM2A的基础上添加微量铌和稀土开发的新型丝锥用高速钢。TGM2A-S钢的生产工艺流程为25 t EAF-30 t LF-VD(微合金化)-铸...TGM2A-S钢(/%:0.85C、0.27Si、0.24Mn、0.026P、0.007S、3.98Cr、4.76Mo、6.09W、1.83V、0.12Nb、0.03RE)是在高速钢TGM2A的基础上添加微量铌和稀土开发的新型丝锥用高速钢。TGM2A-S钢的生产工艺流程为25 t EAF-30 t LF-VD(微合金化)-铸锭(700 kg)二火锻造(85 mm方)-连轧(Φ8 mm)-冷拉(Φ6.6 mm)-加工丝锥(M6)。结果表明,原工艺:3 t中频感应炉-ESR(280 kg锭)-二火锻造(85 mm方)-连轧(Φ8 mm)-冷拉(Φ6.6 mm)-加工丝锥(M6)生产的TGM2A钢中的O和N含量分别为35.4×10^(-6)和123.6×10^(-6),而改进工艺生产的TGM2A-S钢的O和N含量分别为15.7×10^(-6)和87.7×10^(-6)。TGM2A-S钢的丝锥切削寿命较电渣工艺生产的TGM2A钢提高20%;TGM2A-S钢的淬火晶粒为10.5级,电渣工艺生产的TGM2A钢的晶粒度为10级。展开更多
Background:Previous studies have revealed the critical role of transglutaminase 2(TGM2)as a potential therapeutic target in cancers,but the oncogenic roles and underlying mechanisms of TGM2 in gastric cancer(GC)are no...Background:Previous studies have revealed the critical role of transglutaminase 2(TGM2)as a potential therapeutic target in cancers,but the oncogenic roles and underlying mechanisms of TGM2 in gastric cancer(GC)are not fully understood.In this study,we examined the role and potential mechanism of TGM2 in GC.Methods:Western blotting,immunohistochemistry,CCK8,colony formation and transwell assays were used to measure TGM2 expression in the GC cells and tissues and to examine the in vitro role of TGM2 in GC.Xenograft and in vivo metastasis experiments were performed to examine the in vivo role of TGM2 in GC.Gene set enrichment analysis,quantitative PCR and western blotting were conducted to screen for potential TGM2 targets involved in GC.Gain/loss-offunction and rescue experiments were conducted to detect the biological roles of STAT1 in GC cells in the context of TGM2.Co-immunoprecipitation,mass spectrometry,quantitative PCR and western blotting were conducted to identify STAT1-interacting proteins and elucidate their regulatory mechanisms.Mutations in TGM2 and two molecules(ZM39923 and A23187)were used to identify the enzymatic activity of TGM2 involved in the malignant progression of GC and elucidate the underlying mechanism.Results:In this study,we demonstrated elevated TGM2 expression in the GC tissues,which closely related to pathological grade,and predicted poor survival in patients with GC.TGM2 overexpression or knockdown promoted(and inhibited)cell proliferation,migration,and invasion,which were reversed by STAT1 knockdown or overexpression.Further studies showed that TGM2 promoted GC progression by inhibiting STAT1 ubiquitination/degradation.Then,tripartite motif-containing protein 21(TRIM21)was identified as a ubiquitin E3 ligase of STAT1 in GC.TGM2 maintained STAT1 stability by facilitating the dissociation of TRIM21 and STAT1 with GTP-binding enzymatic activity.A23187 abolished the role of TGM2 in STAT1 and reversed the pro-tumor role of TGM2 in vitro and in vivo.Conclusions:This study revealed a critical role and regulatory mechanism of TGM2 on STAT1 in GC and highlighted the potential of TGM2 as a therapeutic target,which elucidates the development of medicine or strategies by regulating the GTP-binding activity of TGM2 in GC.展开更多
We have studied the expression of a subset of genes encoding important tumor growth related factors in U87 glioma cells with IRE1 (inositol requiring enzyme-1) knockdown as well as their hypoxic regulation. It was sho...We have studied the expression of a subset of genes encoding important tumor growth related factors in U87 glioma cells with IRE1 (inositol requiring enzyme-1) knockdown as well as their hypoxic regulation. It was shown that the expression levels of activating transcription factor 6 (ATF6), clusterin (CLU), adhesion G protein-coupled receptor E5 (ADGRE5), transglutaminase?2, C polypeptide (TGM2), leukemia inhibitory factor (LIF), phosphoserine aminotransferase 1 (PSAT1), glyoxalase I (GLO1) and tetraspanin 13 (TSPAN13) are significantly down-regulated in glioma cells with the knockdown of IRE1 signaling enzyme. It was also shown that in glioma cells subjected to hypoxia, the expression levels of PSAT1, TSPAN13, EIF2AK3, and TGM2 genes were up-regulated, whereas the expression of ATF6 gene was down-regulated. At the same time, the expression levels of LIF, CLU, and ADGRE5 genes did not change in response to hypoxic treatment.?Furthermore, inhibition of IRE1, a key effector of an unfolded protein response pathway, modified the effect of hypoxia on the expression of most studied genes. Present study demonstrates that IRE1 knockdown down-regulated the expression of most studied genes and modified their hypoxic regulation and that these changes possibly contributed to the suppression of glioma growth in cells without IRE1 signaling enzyme function.展开更多
基金Supported by the Ningxia Natural Science Foundation,No.2022AAC03144.
文摘BACKGROUND Gastric cancer(GC)is a common gastrointestinal malignancy worldwide.Based on cancer-related mortality,the current prevention and treatment strategies for GC still show poor clinical results.Therefore,it is important to find effective drug treatment targets.AIM To explore the molecular mechanism of 18β-glycyrrhetinic acid(18β-GRA)regulating the miR-345-5p/TGM2 signaling pathway to inhibit the proliferation of GC cells.METHODS CCK-8 assay was used to determine the effect of 18β-GRA on the survival rate of GES-1 cells and AGS and HGC-27 cells.Cell cycle and apoptosis were detected by flow cytometry,cell migration was detected by a wound healing assay,the effect of 18β-GRA on subcutaneous tumor growth in BALB/c nude mice was investigated,and the cell autophagy level was determined by MDC staining.TMT proteomic analysis was used to detect the differentially expressed autophagy-related proteins in GC cells after 18β-GRA intervention,and then the protein-protein interaction was predicted using STRING(https://string-db.org/).MicroRNAs(miRNAs)transcriptome analysis was used to detect the miRNA differential expression profile,and use miRBase(https://www.mirbase/)and TargetScan(https://www.targetscan.org/)to predict the miRNA and complementary binding sites.Quantitative real-time polymerase chain reaction was used to detect the expression level of miRNA in 18β-GRA treated cells,and western blot was used to detect the expression of autophagy related proteins.Finally,the effect of miR-345-5p on GC cells was verified by mir-345-5p overexpression.RESULTS 18β-GRA could inhibit GC cells viability,promote cell apoptosis,block cell cycle,reduce cell wound healing ability,and inhibit the GC cells growth in vivo.MDC staining results showed that 18β-GRA could promote autophagy in GC cells.By TMT proteomic analysis and miRNAs transcriptome analysis,it was concluded that 18β-GRA could down-regulate TGM2 expression and up-regulate miR-345-5p expression in GC cells.Subsequently,we verified that TGM2 is the target of miR-345-5p,and that overexpression of miR-345-5p significantly inhibited the protein expression level of TGM2.Western blot showed that the expression of autophagy-related proteins of TGM2 and p62 was significantly reduced,and LC3II,ULK1 and AMPK expression was significantly increased in GC cells treated with 18β-GRA.Overexpression of miR-345-5p not only inhibited the expression of TGM2,but also inhibited the proliferation of GC cells by promoting cell apoptosis and arresting cell cycle.CONCLUSION 18β-GRA inhibits the proliferation of GC cells and promotes autophagy by regulating the miR-345-5p/TGM2 signaling pathway.
文摘Nb-RE微合金化TGM2高速钢(%:0.88～0.89C、4.14～4.16Cr、4.73～4.76Mo、6.09～6.12W、1.85～1.86V、0.05～0.10Nb、0.05～0.06RE)由25 t EAF-30 t LF(VD,加Nb-RE)-1 t ESR工艺冶炼。试验结果表明,经Nb-RE微合金化后,TGM2高速钢Φ96 mm材淬火晶粒尺寸明显细化,晶粒度由原来未微合金化钢的9.5级提高至10～10.5级;淬、回火后硬度HRC为65.2～65.8,600℃4 h红硬性HRC为62.1～62.3,Nb-RE TGM2钢制成刀具的切削寿命较原TGM2钢提高20%。
基金the National Natural Science Foundation of China(81802996,81871946,and 82072708)Special Foundation for National Science and Technology Basic Research Program of China(2019FY101104)+2 种基金the Program for Development of Innovative Research Team in the First Affiliated Hospital of NJMUthe Priority Academic ProgramDevelopment of JiangsuHigher Education Institutions(JX10231801)Jiangsu Key Medical Discipline(General Surgery)(ZDXKA2016005),Jiangsu Key Lab of Cancer Biomarkers,Prevention and Treatment,Collaborative Innovation Center for Cancer Personalized Medicine,Nanjing Medical University.
文摘Background:Previous studies have revealed the critical role of transglutaminase 2(TGM2)as a potential therapeutic target in cancers,but the oncogenic roles and underlying mechanisms of TGM2 in gastric cancer(GC)are not fully understood.In this study,we examined the role and potential mechanism of TGM2 in GC.Methods:Western blotting,immunohistochemistry,CCK8,colony formation and transwell assays were used to measure TGM2 expression in the GC cells and tissues and to examine the in vitro role of TGM2 in GC.Xenograft and in vivo metastasis experiments were performed to examine the in vivo role of TGM2 in GC.Gene set enrichment analysis,quantitative PCR and western blotting were conducted to screen for potential TGM2 targets involved in GC.Gain/loss-offunction and rescue experiments were conducted to detect the biological roles of STAT1 in GC cells in the context of TGM2.Co-immunoprecipitation,mass spectrometry,quantitative PCR and western blotting were conducted to identify STAT1-interacting proteins and elucidate their regulatory mechanisms.Mutations in TGM2 and two molecules(ZM39923 and A23187)were used to identify the enzymatic activity of TGM2 involved in the malignant progression of GC and elucidate the underlying mechanism.Results:In this study,we demonstrated elevated TGM2 expression in the GC tissues,which closely related to pathological grade,and predicted poor survival in patients with GC.TGM2 overexpression or knockdown promoted(and inhibited)cell proliferation,migration,and invasion,which were reversed by STAT1 knockdown or overexpression.Further studies showed that TGM2 promoted GC progression by inhibiting STAT1 ubiquitination/degradation.Then,tripartite motif-containing protein 21(TRIM21)was identified as a ubiquitin E3 ligase of STAT1 in GC.TGM2 maintained STAT1 stability by facilitating the dissociation of TRIM21 and STAT1 with GTP-binding enzymatic activity.A23187 abolished the role of TGM2 in STAT1 and reversed the pro-tumor role of TGM2 in vitro and in vivo.Conclusions:This study revealed a critical role and regulatory mechanism of TGM2 on STAT1 in GC and highlighted the potential of TGM2 as a therapeutic target,which elucidates the development of medicine or strategies by regulating the GTP-binding activity of TGM2 in GC.
文摘We have studied the expression of a subset of genes encoding important tumor growth related factors in U87 glioma cells with IRE1 (inositol requiring enzyme-1) knockdown as well as their hypoxic regulation. It was shown that the expression levels of activating transcription factor 6 (ATF6), clusterin (CLU), adhesion G protein-coupled receptor E5 (ADGRE5), transglutaminase?2, C polypeptide (TGM2), leukemia inhibitory factor (LIF), phosphoserine aminotransferase 1 (PSAT1), glyoxalase I (GLO1) and tetraspanin 13 (TSPAN13) are significantly down-regulated in glioma cells with the knockdown of IRE1 signaling enzyme. It was also shown that in glioma cells subjected to hypoxia, the expression levels of PSAT1, TSPAN13, EIF2AK3, and TGM2 genes were up-regulated, whereas the expression of ATF6 gene was down-regulated. At the same time, the expression levels of LIF, CLU, and ADGRE5 genes did not change in response to hypoxic treatment.?Furthermore, inhibition of IRE1, a key effector of an unfolded protein response pathway, modified the effect of hypoxia on the expression of most studied genes. Present study demonstrates that IRE1 knockdown down-regulated the expression of most studied genes and modified their hypoxic regulation and that these changes possibly contributed to the suppression of glioma growth in cells without IRE1 signaling enzyme function.
