Objective:To discuss the abnormal expression of Wnt inhibitory factor(WIFI) in hepatocellular carcinoma cells and its regulating effect on the hepatocellular carcinoma invasion and metastasis factors of tissue inhibit...Objective:To discuss the abnormal expression of Wnt inhibitory factor(WIFI) in hepatocellular carcinoma cells and its regulating effect on the hepatocellular carcinoma invasion and metastasis factors of tissue inhibitor of matrix metalloproteinases-3(TIMP-3)and caveolin-1.Methods:RT-PCR and Western blot were employed to detect the expression of WIF1 in six hepatocellular carcinoma eell lines of HepG2,Hep3 B,Huh7,PLC/PRF/5.SMMC-7721 and MHCC97 and the immortalized human liver cell line THLE-3.Besides,Lipofectamine 2000 was employed to transfect the eukaryotic expression vector pcDNA3.lWIF1 and blank plasmid pcDNA3.1 into hepatocellular carcinoma cell lines.Transwell assay was used to detect the effect of WIF1 on the invasion ability of hepatocellular carcinoma cells;Western blot was used to detect the effect of WIF1 on the expression of TIMP-3 and caveolin-1in hepatocellular carcinoma cells,it also discussed the effect on the expression of β-catenin.Results:The expression of WIF1 in hepatocellular carcinoma cell lines was lower than that in the normal liver cell lines(P<0.01);while there was basically no expression of WIF1 in the human highly metastatic cell line MHCC-97 and moderate expression in HepG2 and SMMC-7721.Therefore,HepG2 and SMMC-7721 were chosen as the further experimental cell lines.After transfecting the eukaryotic expression vector peDNA3.1-WEFl and blank plasmid pcDNA3.1 into hepatocellular carcinoma eell lines,compared with(he blank plasmid group,the cell viability and invasion ability in the WIF1 group were all reduced(P<0.01),the expression of TIMP-3,caveolin-1 and mRNA were all down-regulated(P<0.01),and the expression of β-catenin was decreased(P<0.01).Conclusions:Because of down-regulation or missing of expression of WIFI in hepatocellular carcinoma cell lines,the up-regulation of WIFI expression can significantly inhibit the invasion and metastasis of HepG2 and SMMC-7721 of hepatocellular carcinoma cell lines,which are related to the up-regulated expression of TIMP-3 and down-regulated expression of caveolin-1 and may be realized through the Wnt/β-catenin signaling pathway.展开更多
Background:Osteoporosis arises mainly from an imbalance in bone remodeling,characterized by the impaired ability of osteoblasts(OBs)to form new bone.A sig-nificant challenge remains in identifying and validating key g...Background:Osteoporosis arises mainly from an imbalance in bone remodeling,characterized by the impaired ability of osteoblasts(OBs)to form new bone.A sig-nificant challenge remains in identifying and validating key genes involved in os-teogenic differentiation to develop effective treatments.Methods:The regulatory role of WIF1 in osteogenic differentiation stages was in-vestigated using Western blot and quantitative polymerase chain reaction assays to measure the expression levels of osteogenic-related genes in MC3T3-E1 cells.Mineralization of OBs was assessed through alizarin red S(ARS)and alkaline phos-phatase(ALP)staining assays.To explore the relationship with mitophagy,RNA se-quencing was performed to examine the effects of Wif1 overexpression on genes re-lated to osteogenic differentiation and mitophagy processes.Additionally,histological staining and micro-computed tomography were conducted on ovar-iectomized(OVX)mice treated with Wif1-overexpressing OB-derived extracellular vesicles(EVs)to evaluate their impact on bone loss and osteogenic differentiation in bone marrow stromal cells(BMSCs).Results:Wif1 was identified as a crucial marker for late-stage osteogenic differentia-tion,playing a role in regulating mitophagy to release functional EVs.In vitro ana-lyses showed that Wif1 overexpression accelerated osteogenic differentiation by ac-tivating genes related to late-stage osteogenic differentiation and mitophagy processes.In vivo experiments demonstrated that administering Wif1-overexpressing OB-derived EVs to OVX mice partially reversed bone loss and countered the sup-pression of osteogenic differentiation in BMSCs.Conclusions:Our findings shed light on the molecular mechanism of Wif1 in osteogenic differentiation and suggested the therapeutic efficacy of Wif1-overexpressing OB-derived EVs in osteoporosis.展开更多
基金supported by National Science&Technology Pillar Program during the 12th Five-year Plan Period under the research and development of new trisacryl gelatin microspheres(No.2012BAH5B06)
文摘Objective:To discuss the abnormal expression of Wnt inhibitory factor(WIFI) in hepatocellular carcinoma cells and its regulating effect on the hepatocellular carcinoma invasion and metastasis factors of tissue inhibitor of matrix metalloproteinases-3(TIMP-3)and caveolin-1.Methods:RT-PCR and Western blot were employed to detect the expression of WIF1 in six hepatocellular carcinoma eell lines of HepG2,Hep3 B,Huh7,PLC/PRF/5.SMMC-7721 and MHCC97 and the immortalized human liver cell line THLE-3.Besides,Lipofectamine 2000 was employed to transfect the eukaryotic expression vector pcDNA3.lWIF1 and blank plasmid pcDNA3.1 into hepatocellular carcinoma cell lines.Transwell assay was used to detect the effect of WIF1 on the invasion ability of hepatocellular carcinoma cells;Western blot was used to detect the effect of WIF1 on the expression of TIMP-3 and caveolin-1in hepatocellular carcinoma cells,it also discussed the effect on the expression of β-catenin.Results:The expression of WIF1 in hepatocellular carcinoma cell lines was lower than that in the normal liver cell lines(P<0.01);while there was basically no expression of WIF1 in the human highly metastatic cell line MHCC-97 and moderate expression in HepG2 and SMMC-7721.Therefore,HepG2 and SMMC-7721 were chosen as the further experimental cell lines.After transfecting the eukaryotic expression vector peDNA3.1-WEFl and blank plasmid pcDNA3.1 into hepatocellular carcinoma eell lines,compared with(he blank plasmid group,the cell viability and invasion ability in the WIF1 group were all reduced(P<0.01),the expression of TIMP-3,caveolin-1 and mRNA were all down-regulated(P<0.01),and the expression of β-catenin was decreased(P<0.01).Conclusions:Because of down-regulation or missing of expression of WIFI in hepatocellular carcinoma cell lines,the up-regulation of WIFI expression can significantly inhibit the invasion and metastasis of HepG2 and SMMC-7721 of hepatocellular carcinoma cell lines,which are related to the up-regulated expression of TIMP-3 and down-regulated expression of caveolin-1 and may be realized through the Wnt/β-catenin signaling pathway.
基金supported by the Major Research Plan of the National Natural Science Foundation of China(92249303)the Key Project of the National Natural Science Foundation of China(82230071).
文摘Background:Osteoporosis arises mainly from an imbalance in bone remodeling,characterized by the impaired ability of osteoblasts(OBs)to form new bone.A sig-nificant challenge remains in identifying and validating key genes involved in os-teogenic differentiation to develop effective treatments.Methods:The regulatory role of WIF1 in osteogenic differentiation stages was in-vestigated using Western blot and quantitative polymerase chain reaction assays to measure the expression levels of osteogenic-related genes in MC3T3-E1 cells.Mineralization of OBs was assessed through alizarin red S(ARS)and alkaline phos-phatase(ALP)staining assays.To explore the relationship with mitophagy,RNA se-quencing was performed to examine the effects of Wif1 overexpression on genes re-lated to osteogenic differentiation and mitophagy processes.Additionally,histological staining and micro-computed tomography were conducted on ovar-iectomized(OVX)mice treated with Wif1-overexpressing OB-derived extracellular vesicles(EVs)to evaluate their impact on bone loss and osteogenic differentiation in bone marrow stromal cells(BMSCs).Results:Wif1 was identified as a crucial marker for late-stage osteogenic differentia-tion,playing a role in regulating mitophagy to release functional EVs.In vitro ana-lyses showed that Wif1 overexpression accelerated osteogenic differentiation by ac-tivating genes related to late-stage osteogenic differentiation and mitophagy processes.In vivo experiments demonstrated that administering Wif1-overexpressing OB-derived EVs to OVX mice partially reversed bone loss and countered the sup-pression of osteogenic differentiation in BMSCs.Conclusions:Our findings shed light on the molecular mechanism of Wif1 in osteogenic differentiation and suggested the therapeutic efficacy of Wif1-overexpressing OB-derived EVs in osteoporosis.
