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Objective: To investigate the effect and possible mechanism of Y-box-binding protein 1 (YB-1) on sorafenib resistance in hepatoma cells. Methods: Lentiviral vectors with YB-1 overexpression and knockdown were constructed, respectively, to stimulate human hepatoma cell lines (HepG2 and Huh7) alone or in combination with sorafenib.The overexpression part of the experiment was divided into four groups: overexpression control group (Lv-NC), YB-1 overexpression group (Lv-YB-1), overexpression control combined with sorafenib resistance group (Lv-NC+sorafenib), YB-1 overexpression combined with sorafenib resistance group (Lv-YB-1 + sorafenib). The knockdown part of the experiment was also divided into four groups: knockdown control group (Lv-shNC), YB-1 knockdown group (Lv-shYB-1), knockdown control combined with sorafenib resistance group (Lv-shNC + sorafenib), YB-1 knockdown combined with sorafenib resistance group (Lv-shYB-1 + sorafenib). The occurrence of cell apoptosis was detected by TUNEL. The protein expression levels of phosphorylated (p)-ERK and ERK, key proteins in the extracellular regulatory protein kinase (ERK) signaling pathway, were detected by Western blot and quantified by ImageJ software. Subcutaneous tumorigenesis experiments were performed in nude mice. The effect of YB-1 on the efficacy of sorafenib was verified in vivo. The comparison between the two sets of data was carried out by an independent sample t-test. One-way ANOVA was used for comparisons between the three groups of data above. Results: Sorafenib had accelerated the occurrence of apoptosis in hepatoma cells, while YB-1 overexpression had inhibited cell apoptosis, and at the same time also inhibited the apoptosis-accelerating impact of sorafenib. On the contrary, YB-1 knockdown accelerated cell apoptosis and amplified the induction effect of sorafenib on apoptosis. Furthermore, sorafenib resistance had down-regulated p-ERK levels (HepG2: Lv-NC 0.685 ± 0.143, Lv-NC + sorafenib 0.315 ± 0.168, P < 0.05; Huh7: Lv-NC 0.576 ± 0.078, Lv-NC + sorafenib 0.150 ± 0.131, P < 0.01), whereas YB-1 overexpression had inhibited sorafenib resistance p-ERK reduction (HepG2: Lv-NC + sorafenib 0.315 ± 0.168, Lv-YB-1 + sorafenib 0.688 ± 0.042, P < 0.05; Huh7: Lv-NC + sorafenib 0.150 ± 0.131, Lv-YB-1 + sorafenib 0.553 ± 0.041, P < 0.05). YB-1 knockdown further increased sorafenib-induced p-ERK downregulation (HepG2: Lv-shNC + sorafenib 0.911 ± 0.252, Lv-shYB-1 + sorafenib 0.500 ± 0.201, P < 0.05; Huh7: Lv-shNC + sorafenib 0.577 ± 0.082, Lv-shYB-1 + sorafenib 0.350 ± 0.143, P < 0.05), which was further verified in naked mice (Lv-shNC + sorafenib 0.812 ± 0.279, Lv-shYB-1 + sorafenib 0.352 ± 0.109, P < 0.05). Conclusion: YB-1 mediates the occurrence of sorafenib resistance via the ERK signaling pathway in hepatoma cells.
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Humans , Animals , Mice , Cell Line, Tumor , Sorafenib/pharmacology , Drug Resistance, Neoplasm , Y-Box-Binding Protein 1/metabolism , Carcinoma, Hepatocellular/metabolism , MAP Kinase Signaling System , Mice, NudeABSTRACT
Chronic liver diseases have various etiologies and often have poor long-term prognosis in clinical practice. Y-box binding protein-1 (YB-1) is a multifunctional protein, and in-depth studies in recent studies have found that it plays a key role in the development and progression of chronic liver diseases such as liver fibrosis and hepatocellular carcinoma (HCC). This article summarizes the role of YB-1 in chronic liver diseases such as liver fibrosis, HCC (proliferation, apoptosis, metastasis, prognosis, and drug resistance), and liver failure, so as to provide a theoretical basis for the diagnosis and treatment of chronic liver diseases.
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Objective To explore the correlation between Y-box binding protein-1 ( YB-1) and P-glycoprotein ( P-gp) in drug-resistant hepatocellular carcinoma ( HCC) Bel-7402/ADM cells, and speculate the related mechanism of drug resistance.Methods Bel-7402/ADM cells were developed by concentration gradient escalation and intermittent administration of large dose. The levels of YB-1 mRNA and MDR1 mRNA were detected by means of RT-PCR.Western blotting was used to detect the protein expression of YB-1 and P-gp in the Bel-7402 cells and doxorubicin resistant Bel-7402 ( Bel-7402/ADM) cells. Bel-7402/ADM cells were transfected with small interfering RNA ( siRNA) targeting human YB-1. Expression levels of YB-1 and MDR1 mRNA and protein were detected by means of RT-PCR and Western blotting. Results IC50 values of ADM on hepatoma carcinoma cells Bel-7402 and Bel-7402/ADM were (2.23±0.07) and (7.02±0.03) μmol?L-1. The mRNA expression levels of MDR1 and YB-1 were all significantly higher in Bel-7402/ADM cells than in Bel-7402 ( P<0.01) . The mRNA expression levels of MDR1 and YB-1 in Bel-7402/ADM cells transfected with YB-1 siRNA were reduced significantly (P<0.01). The protein levels of YB-1 and MDR1 in Bel-7402/ADM cells transfected with YB-1 siRNA were reduced significantly ( P<0. 05 ) . Conclusion These results suggest that the high expression level of YB-1 is probably correlated with multidrug resistance in HCC Bel-7402/ADM cells.
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Objective To explore the correlation between Y-box binding protein-1 ( YB-1) and P-glycoprotein ( P-gp) in drug-resistant hepatocellular carcinoma ( HCC) Bel-7402/ADM cells, and speculate the related mechanism of drug resistance.Methods Bel-7402/ADM cells were developed by concentration gradient escalation and intermittent administration of large dose. The levels of YB-1 mRNA and MDR1 mRNA were detected by means of RT-PCR.Western blotting was used to detect the protein expression of YB-1 and P-gp in the Bel-7402 cells and doxorubicin resistant Bel-7402 ( Bel-7402/ADM) cells. Bel-7402/ADM cells were transfected with small interfering RNA ( siRNA) targeting human YB-1. Expression levels of YB-1 and MDR1 mRNA and protein were detected by means of RT-PCR and Western blotting. Results IC50 values of ADM on hepatoma carcinoma cells Bel-7402 and Bel-7402/ADM were (2.23±0.07) and (7.02±0.03) μmol?L-1. The mRNA expression levels of MDR1 and YB-1 were all significantly higher in Bel-7402/ADM cells than in Bel-7402 ( P<0.01) . The mRNA expression levels of MDR1 and YB-1 in Bel-7402/ADM cells transfected with YB-1 siRNA were reduced significantly (P<0.01). The protein levels of YB-1 and MDR1 in Bel-7402/ADM cells transfected with YB-1 siRNA were reduced significantly ( P<0. 05 ) . Conclusion These results suggest that the high expression level of YB-1 is probably correlated with multidrug resistance in HCC Bel-7402/ADM cells.
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Objective To investigate the clinicopathologic significance of Y-box binding protein-1 (YB-1)expression in cervical cancer and its correlation with epithelial-mesenchymal transition. Methods A series of 202 samples,including 50 cases of normal cervical tissues,100 cases of cervical intraepithelial neo-plasia(CIN)and 52 cases of squamous cell carcinoma(SCC),were examined YB-1 and E-cadherin by immu-nohistochemical staining. Results The E-cadherin cell membrane immunoreactivity for normal/ CINⅠ,CINⅡ-Ⅲ,and SCC tissues were 100% ,64% and 3. 85% ,respectively( χ2 = 40. 909;χ2 = 119. 088;χ2 =25. 274;P < 0. 05). The negative and aberrant expression of E-cadherin was higher in metastatic SCC (100. 0% ,20 / 20)than that in non-metastatic SCC(68. 75% ,22 / 32)(χ2 = 5. 857,P = 0. 016). 94. 23%(49 / 52)cases of SCC exhibited strong YB-1 cytoplasmic immunoreactivity. The positive rates in normal/ CINⅠand CINⅡ-Ⅲ were 0(0 / 100)and 10. 00%(5 / 50)(χ2 = 72. 591;χ2 = 139. 059;χ2 = 5. 857;P < 0. 05). The cytoplasmic expression of YB-1 was higher in metastatic SCC(100. 0% ,20 / 20)than that in non-metastatic SCC(59. 38% ,19 / 32)(χ2 = 10. 833,P = 0. 001). The rates were 60. 71%(17 / 28)and 91. 67%(22 /24)in early stage SCC and late stage SCC(χ2 = 6. 603,P = 0. 01). Conclusion YB-1 over-expression is as-sociated with the malignant transformation of cervical epithelium,stage progression and metastasis of cervical cancer. The up-regulation of YB-1 is also associated with the down-regulation of E-cadherin,and it may predict the malignant transformation of CIN and distal metastasis of cervical cancer.
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Objective To detect the expressions of Y-box-binding protein-1(YB-1)and epithelial-mesenchymal transition(EMT)markers(E-cadherin and N-cadherin)in colorectal cancer(CRC),to analyze the relationship between the expression of YB-1 and clinicopathological parameters,to evaluate the correlations among YB-1,E-cadherin and N-cadherin. Methods The expressions of YB-1,E-cadherin and N-cadherin in 120 primary CRC tumors and corresponding normal tissues were detected by western blot and immunohistochem-istry and the results were analyzed. Results The expressions of YB-1,E-cadherin and N-cadherin in tumors were significantly different from those in corresponding normal tissues(χ2 = 47. 373,P ﹤ 0. 05;χ2 = 83. 145, P ﹤ 0. 05;χ2 = 41. 832,P ﹤ 0. 05). The expression of YB-1 in tumors was associated significantly with tumor differentiation,tumor invasion,lymph node metastasis and distance metastasis(χ2 = 8. 077,P = 0. 008;χ2 =8. 178,P = 0. 006;χ2 = 15. 152,P ﹤ 0. 001;χ2 = 7. 368,P = 0. 011). It was negatively correlated with E-cadherin expression(r = - 0. 238,P = 0. 009),but positively correlated with N-cadherin expression(r =0. 361,P ﹤ 0. 001). Conclusion YB-1 may promote the occurrence and development of CRC by participating in EMT program.
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Objective To investigate the effect of gene silencing of Y-box binding protein-1(YB-1) by RNA interference on the proliferation and migration in prostate cancer cells lines PC-3 cells .Methods YB-1 siRNAs(pGenesil-1-YB-1-1 and pGenesil-1-YB-1-2) were synthesized and transfected into cloned into the the PC-3 cells by liposome .The expressions of YB-1 were measure by RT-PCR and Western blotting .The proliferation and migration were respectively detected by MTT and Transwell method . Results ThemRNA and protein expressions of YB-1 were significantly decreased by pGenesil-1-YB-1-1 and pGenesil-1-YB-1-2 (P<0 .05) ,compared with the control group ,the inhibition ratio of mRNA expression was 36 .23% and 39 .42% respectively and the inhibition ratio of protein expression was 41 .56% and 55 .33% respectively .The proliferation and migration were significantly decreased by pGenesil-1-YB-1-1 and pGenesil-1-YB-1-2(P<0 .05) .Conclusion YB-1 gene silencing by RNA interference inhibits the proliferation and migration in prostate cancer cells lines PC-3 cells .
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In the process of epithelial-mesenchymal transition (EMT),cell-cell adherence is disrupted,apico-basal polarity is lost,the ability of anti-apoptosis,migration and invasion is acquired.Y-box-binding protein 1 (YB-1) is a member of the cold-shock protein superfamily,containing a structurally and functionally conserved cold shock domain.Studies indicate that YB-1 can promote the occurrence and development of tumors by regulating EMT.In the process of EMT mediated by YB-1,various transcription factors and signal pathways play important roles.