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AIM: To investigate the effect of miR-195-5p on ox-LDL-induced vascular endothelial cell injury and its mechanism. METHODS: The expression of miR-195-5p in HUVECs cells induced by ox-LDL was detected by RT-PCR. Cell proliferation, the expression of oxidative stress associated factors (MDA and LDH) and apoptosis were detected by CCK-8, ELISA and flow cytometry, respectively. The potential targets of miR-195-5p were determined by dual luciferase reporter assay. The expression of target protein in ox-LDL-induced HUVECs cells and the relationship between miR-195-5p and FBXW7 expression were detected by Western blotting. RESULTS: The expression of miR-195-5p in ox-LDL-induced HUVECs cells was significantly higher than that in normal HUVECs cells. Subsequently, miR-195-5p silencing in ox-LDL-induced HUVECs cells effectively promoted the proliferation of HUVECs cells, whereas suppressed the expression of oxidative stress associated factors (MDA and LDH) and apoptosis level. Luciferase reporter assay and Western blot results showed that miR-195-5p could directly target FBXW7 protein gene and negatively regulate its expression. Finally, miR-195-5p modulated the proliferation, oxidative stress and apoptosis of HUVECs cells induced by ox-LDL by regulating the expression of FBXW7. CONCLUSION: miR-195-5p is highly expressed in HUVECs cells induced by ox-LDL. In addition, miR-195-5p can aggravate ox-LDL-induced cytotoxicity, oxidative stress and apoptosis of HUVECs cells by inhibiting the expression of FBXW7.
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Objective To investigate the effect of FBXW7 on the expression and localization of HSF1 in colorectal cancer cells. Methods The expression levels of HSF1 and pHSF1Ser326 protein in FBXW7 deletion (KO) and wild-type (WT) FBXW7-expressing counterpart colorectal cancer cells were detected by Western blot. The nucleoprotein expression and localization of pHSF1Ser326 in heat-shocked or recovery stage cells were observed by Western blot and immunofluorescence method. Results The HSF1 expression level in DLD1 cells transfected with FBXW7α was decreased significantly (P < 0.01). The expression levels of HSF1 and pHSF1Ser326 protein in FBXW7 KO cells were higher than those in WT cells (all P < 0.05). HSF1 and pHSF1Ser326 in FBXW7 KO cells were mainly expressed in nucleus and weakly expressed in cytoplasm. After warm stimulation, the expression of HSF1 and pHSF1Ser326 in WT cells recovered to the unstimulated level, while the expression of HSF1 and pHSF1Ser326 in FBXW7 KO cells were higher in the nucleus (all P < 0.01). Conclusion Loss of FBXW7 could affect the nuclear HSF1 recovery after warm stimulation. It may be associated with FBXW7 deletion inhibiting the degradation of nuclear HSF1.
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Objective:To investigate the effects of Notch1 signaling on histone acetylation of Foxp3 gene and its roles in regulating regulatory T (Treg) cells in children with acute B-cell precursor lymphoblastic leukemia (BCP-ALL).Methods:Blood samples were collected form 38 children with BCP-ALL before treatment and 15 age-matched healthy children (control group). Flow cytometry was performed to detect the proportion of peripheral blood CD4 + CD25 hiFoxp3 + Treg cells and the expression of Foxp3, cytotoxic lymphocyte antigen 4 (CTLA4), glucocorticoid-induced tumor necrosis factor receptor (GITR), CD39 and Notch1 at protein level. Histone 4 acetylation (H4Ac) at Foxp3 gene promoter and the binding abilities of Foxp3 gene promoter to NICD1 and p300 in CD4 + T cells were measured by chromatin immunoprecipitation. Quantitative real-time PCR was performed to detect the expression of Foxp3, presenilin 1 (PSEN1), mastermind-like transcriptional coactivator 1 (MAML1), SKI-interacting protein (SKIP), F-box and WD40 domain protein 7 (FBXW7), glycogen synthase kinase-3 beta (GSK3β) and IKAROS at mRNA level in CD4 + T cells. The concentrations of TGF-β and IL-10 in plasma were evaluated by ELISA. Results:(1) The proportion of peripheral blood CD4 + CD25 hiFoxp3 + Treg cells, the expression of differentiation- and function-associated molecules (Foxp3, CTLA4, GITR and CD39) and the concentrations of TGF-β and IL-10 in plasma were higher in the BCP-ALL group than in the control group ( P<0.05). (2) In children with acute BCP-ALL, H4Ac at Foxp3 promoter and the binding abilities of Foxp3 gene promoter to NICD1 and p300 were significantly increased as compared with those in control group( P<0.05). The binding abilities of Foxp3 gene promoter to NICD1 and p300 were positively correlated with the expression of Foxp3 at mRNA level ( r=0.58 and 0.46, both P<0.05). After competitive inhibition, the three aforementioned indexes in the acute BCP-ALL group were significantly lower than those in untreated group ( P<0.05); the binding ability of Foxp3 gene promoter to NICD1 in the control group was also significantly lower than that in untreated control group ( P<0.05), but no statistical differences in the other two indexes were found between the control groups with or without treatment ( P>0.05). ⑶ Compared with the control group, the expression of Notch1, PSEN1, MAML1 and SKIP in CD4 + T cells were elevated significantly ( P<0.05), while the transcription level of negative regulatory factor FBXW7 was decreased remarkably in children with acute BCP-ALL ( P<0.05). No statistical differences in the expression of GSK3β or IKAROS were found between the two groups ( P>0.05). Conclusions:Overactivation of Notch1 signaling caused by low expression of FBXW7 might be the key factor resulting in histone 4 hyperacetylation at foxp3 gene promoter and Treg cell dysfunction in children with acute BCP-ALL.
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Objective To investigate the expression and significance of FBXW7 and ENO1 in ovarian serous adenocarcinoma of different grades. Methods Immunohistochemistry was used to study the expression of FBXW7 and ENO1 in 60 cases of ovarian serous adenocarcinoma. The relationship between FBXW7 and ENO1 proteins and the prognosis of ovarian serous adenocarcinoma was analyzed. Results The positive rate of FBXW7 expression was 22. 5％ ( 9/40) in 40 cases of ovarian high grade adenocarcinoma and 10 cases in 15 cases of normal oviduct. The positive rate of FBXW7 expression was 66. 7％ ( 10/15) ,and the difference was statistically significant ( P=0. 003) . The expression of FBXW7 in 20 cases of low grade adenocarcinoma was 5 cases,and the positive rate was 25. 0％( 5/20) . In 15 cases of normal ovarian tissue,9 cases were positive,and the positive rate was 60. 0％( 9/15) . The difference was statistically significant ( P=0. 04) . The expression of ENO1 protein was 27 in 40 cases of high grade adenocarcinoma,and the positive rate of expression was 67. 5％( 27/40) . 5 cases were positive in 15 normal fallopian tubes, and the positive rate was 33. 3％( 5/15 ) . The difference was statistically significant ( P = 0. 024 ) . The expression of ENO1 protein was 15 in 20 cases of low grade adenocarcinoma,and the positive rate of expression was 75. 0％( 15/20) . In 15 cases of normal ovarian tissue,4 cases were positive, and the positive rate was 26. 7％( 4/15 ) . The difference was statistically significant ( P=0. 006) . There was no correlation between the low expression of FBXW7 and the high expression of ENO1 in high grade ovarian adenocarcinoma ( P= 0. 199 ) , but there was a significant correlation between the low expression of FBXW7 and the high expression of ENO1 in low grade ovarian adenocarcinoma ( P<0. 05) . In low grade serous adenocarcinoma,the 5-year survival rates were 44. 4％ and 32. 1％ respectively,with no significant difference ( P = 0. 052 ) . In ovarian high-grade serous adenocarcinoma, the 5-year survival rates of high-expression group and low-expression group were 20. 0％ and 7. 7％, respectively, with no significant difference ( P=0. 097) . In low grade ovarian serous adenocarcinoma,the 5-year survival rate was 7. 4％ in high expression group and 50. 0％ in low expression group ( P=0. 023) . The 5-year survival rates of ENO1 in high-grade serous adenocarcinoma were 0％ and 40. 0％ in high-expression group and low-expression group respectively ( P=0. 001) . Conclusion The low expression of FBXW7 in ovarian adenocarcinoma suggests that FBXW7 may be a tumor suppressor gene in ovarian serous adenocarcinoma, and ENO1 may be an oncogene in ovarian adenocarcinoma. The high expression of FBXW7 in serous adenocarcinoma indicates that ENO1 may be an oncogene,and the survival rate of FBXW7 in serous adenocarcinoma is higher than that in low expression group. The survival rate of the high-expression group was lower than that of the low-expression group. Therefore, they may become a new diagnostic index and therapeutic target for ovarian serous adenocarcinoma.
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@#[Abstract] Objective: To investigate the impact of FBXW7 gene mutation on cell proliferation, apoptosis, migration, and invasion processes of colorectal cancer HCT-116 cell line. Methods: Recombinant plasmids carrying wild-type and mutant-type FBXW7 SNP were constructed and transfected into HCT-116 cell line; the FBXW7 protein expression level in HCT-116 strains after transfection was detected by Western blotting. Subsequently, cell proliferation capacity was tested by CCK-8 assay; tumor cell colony formation ability was tested by HTCA; cell apoptosis function was tested by FCM; cell migration and invasion were tested by scratch assay and Transwell assay, respectively. Results: Higher HBXW7 protein expression level was detected in HCT-116 strain transfected with wild-type HBXW7 in comparison to the control group (strains transfected with mutant-type HBXW7), negative-control (strains transfected with empty plasmids), and blank-control (strains untransfected) (all P<0.05). Compared with the other groups, strains transfected with wildtype HBXW7 exhibited significantly reduced proliferation, colony formation, migration and invasion ability (all P<0.05), but obviously increased apoptosis rate (P<0.05). Conclusion： ：FBXW7 gene mutation can down-regulate its protein expression, and further promote the proliferation, migration and invasion as well as inhibit the apoptosis of HCT-116 cells.
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As a tumor suppressor gene,FBXW7 is a member of the F-box protein family,and FBXW7 can regulates cell growth and cycle progression.FBXW7 has more than 20 substrates,most of which are cancer proteins,and a few are tumor suppressor factors,which are highly expressed in most tumors.FBXW7's gene deficiency or deletion can cause chromosome instability and lead to tumorigenesis.In this paper,the molecular structure,functional characteristics and the mechanism of action in gynecological malignant tumors of ovarian cancer are reviewed.
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Objective To investigate the expression of FBXW7 during the development of Notch1induced murine leukemia.Methods Notch1 over-expressing murine model of T-cell acute lymphoblastic leukemia was used to study the expression of FBXW7 by real-time PCR methods.Bone marrow mononuclear cells (BMNC) were isolated on different days after transplantation and CD45.2+ GFP+ leukemia cells were sorted by flow cytometry at late stage.The expression changes of FBXW7 were tested by real-time PCR.Results The mouse bone marrow cells both from leukemia and control groups expressed FBXW7.Different expression patterns of FBXW7 were observed during the development of leukemia. The expression of FBXW7 was gradually increased in control group, whereas the expression level of FBXW7 in leukemia group was decreased steadily and reached one-sixth of that in control group on 12th day.Furthermore,lower expression level of FBXW7 was observed in CD45+.2 GFP+ leukemia cells.Conclusion Decreased expression of FBXW7 is observed in Notch1-induced mouse leukemia model,suggesting that the abnormal ubiquitin degradation pathway mediated by FBXW7 might contribute to the leukemogenesis in Notch1-induced murine leukemia model.
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Objective: To study the expression of F-box/WD repeat-containing protein 7(FBXW7), fatty acid synthase (FAS) and minichromosome maintenance protein7 (MCM7) in colorectal carcinoma and the related clinical significance. Methods: Immunohistochemistry method was used to examine the expression of FBXW7, FAS and MCM7 in the colorectal carcinoma, colorectal adenoma and para-carcinoma normal mucosa tissues. Results: The positive rates of FBXW7 in the colorectal carcinoma, adenoma and normal tissues were 58.2%, 75.0%, and 88.9%, respectively, with significant difference found between the colorectal carcinoma and normal mucosa tissues (P<0.05); and FBXW7 expression was significantly correlated with the differentiation degree(χ2=10.516, P=0.001), lymphatic metastasis (χ2=4.489, P=0.034) and the tumor size(χ2=9.974, P=0.002). The positive rates of FAS in the colorectal carcinoma, adenoma and normal tissue were 94.3%, 75.0%, and 55.6%, respectively, with significant difference found between the colorectal carcinoma and normal mucosa tissues (P<0.01); and FAS expression was significantly correlated with the patient age(χ2=7.643, P=0.006). The positive rates of MCM7 in the colorectal carcinoma, adenoma and normal tissue were 95.8%, 66.7%, and 22.2%, respectively, with the difference being significant between the three groups (P<0.01). FBXW7 expression was negatively correlated with that of FAS and MCM7(r= -0.276, P=0.008; r= -0.443, P=0.000), and FAS expression was positively correlated with MCM7 expression(r=0.228, P=0.024). Conclusion: FBXW7, FAS and MCM7 might be new markers for early diagnosis and prognosis prediction of colorectal carcinoma; they may also serve as new therapeutic targets for colorectal carcinoma.