Objective To explore the roles of transcription factor E26 transformation-specific 1(ETS1)and F-box protein 45(FBXO45)in invasion and metastasis in hepatocellular carcinoma cells and the potential molecular mechanism.Methods Jaspar,hTFtarget and Cistrome transcription factor database prediction websites were used to predict the transcription factors of FBXO45.According to the intersection of the predicted results of each database,the expression of FBXO45 was detected after the candidate transcription factors were knockdown in HCCLM3 and Huh7 liver cancer cells,respectively.The most significant influence on FBXO45 expression was selected for further analysis,and chromatin immunoprecipitation assay(ChIP)was used to verify the binding to the FBXO45 promoter.Finally,the potential transcription factor of FBXO45 was identified.The effect of ETS1 overexpression on invasion and migration in HCCLM3 and Huh7 cells was detected by Transwell assay,and the expression levels of epithelial-mesenchymal transition(EMT)pathway proteins were detected by Western blot assay.The effects of FBXO45 knockdown on the invasion and migration under the condition of overexpression of ETS1 were also studied.Results Intersection of FBXO45 transcription factors identified 3 candidate transcription factors,ETS1,SPI1 and YY1.When the 3 transcription factors were knocked down in HCCLM3 and Huh7 cells,respectively,ETS1 knockdown significantly reduced the expression of FBXO45.According to the analysis of The Cancer Genome Atlas(TCGA)data and the Gene Expression Omnibus(GEO)data,the expression levels of ETS1 and FBXO45 were significantly positively correlated(R=0.31,P<0.000 1;R=0.40,P=0.021 9).ChIP suggested that ETS1 could specifically bind to FBXO45 promoter sequence to regulate its expression,confirming that ETS1 was a potential transcription factor of FBXO45.After overexpression of ETS1 in HCCLM3 and Huh7 cells,the invasion and migration abilities of cells were significantly enhanced,and the expression of N-cadherin and Snail was up-regulated(P<0.01).In addition,in the case of ETS1 overexpression,FBXO45 knockdown significantly inhibited the invasion and migration(P<0.01).Conclusion ETS1 activates the transcription of FBXO45 and leads its high expression,which enhances the invasion and migration of HCC cells via EMT pathway and promotes the progression of hepatocellular carcinoma.

Objective:To investigate the expression of nucleoporin 43 (NUP43) in hepato-cellular carcinoma tissues and its impact on prognosis of patients and proliferation and migration of hepatocellular carcinoma cells.Methods:The retrospective cohort study and experi-mental study were conducted. The clinicopathological data of 102 hepatocellular carcinoma patients who were admitted to The First Affiliated Hospital of Army Medical University from January 2008 to December 2012 were collected. There were 83 males and 19 females, aged 56(range, 19-87)years. The expression of NUP43 in hepatocellular carcinoma tissues was analyzed by immunohistochemical staining. HepG2 and SK-HEP-1 hepatocellular carcinoma cells were cultured in vitro. The Western blot was used to verify the effects of Flag-NUP43 overexpression plasmid in transfected cells. The CCK-8 and cell migration experiments were used to analyze the effect of NUP43 overexpression on HepG2 and SK-HEP-1 hepa-tocellular carcinoma cells. Measurement data of normal distribution were expressed as Mean± SD, and comparison between groups was conducted using the independent sample t test. Measurement data of skewed distribution were represented as M(range). Count data were expressed as absolute numbers, and comparisons between groups was conducted using the paired chi-square test. The Kaplan-Meier method was used to calculate survival time, and Log-rank test was used for survival analysis. The R 4.2.1 software was used to draw survival curves. The COX proportional hazards regre-ssion model was used for univariate and multivariate analyses. Results:(1) Expression of NUP43 in hepatocellular carcinoma and adjacent tissues, and analysis of clinicopathological characteristics of patients with high and low expression of NUP43. Results of immunohistochemical staining showed that NUP43 was mainly expressed in the cytoplasm and nuclear membrane of cells. Of 102 hepatocellular carcinoma tissue samples, there were 49 samples with low expression of NUP43 and 53 samples with high expression of NUP43. Of 102 hepatocellular carcinoma adjacent tissue samples, there were 80 samples with low expression of NUP43 and 22 samples with high expression of NUP43. There was a significant difference in the expression of NUP43 between hepatocellular carcinoma and adjacent tissues ( χ2=16.505, P<0.05). Of 102 hepatocellular carcinoma tissue samples, there were significant differences in tumor diameter, pathological grading, and intrahepatic metastasis between the patients with low expression of NUP43 and the patients with high expression of NUP43 ( χ2=5.104, 23.217, 4.169, P<0.05). (2) Survival of hepatocellular carcinoma patients and prognostic factors analysis. The follow-up time of 102 hepatocellular carcinoma patients was 17.9(range, 0.1-107.9)months. The postoperative 1-, 3-, and 5-year overall survival rates were 79.59%, 53.06% and 34.69% for the patients with low expression of NUP43, versus 52.83%, 18.87%, and 9.43% for the patients with high expre-ssion of NUP43, showing a significant difference between them ( χ2=27.071, P<0.05). Results of multi-variate analysis showed that gender, NUP43 expression, TNM staging, and pathological grading were independent influencing factors for postoperative survival in patients with hepatocellular carcinoma ( hazard ratio=1.846, 2.206, 2.040, 2.177, 95% confidence interval as 1.231-2.768, 1.419-3.429, 1.322-3.148, 1.377-3.254, P<0.05). (3) Effects of NUP43 overexpression on the proliferation and migration of HepG2 and SK-HEP-1 hepatocellular carcinoma cells. Western blot analysis showed that transfection of Flag-NUP43 overexpression plasmid significantly increased the expression of NUP43 in HepG2 and SK-HEP-1 cells. Results of CCK-8 experiment showed that after transfection with the control plasmid and Flag-NUP43 overexpression plasmid, the cell proliferation indices of HepG2 were 0.79±0.07 and 1.47±0.05, respectively, showing a significant difference between them ( t=19.402, P<0.05). After transfection with the control plasmid and Flag-NUP43 overexpression plasmid, the cell proliferation indices of SK-HEP-1 cells were 0.59±0.05 and 0.95±0.05, respectively, showing a significant difference between them ( t=15.753, P<0.05). Results of cell migration experiments showed that after transfection with the control plasmid and Flag-NUP43 overexpression plasmid, the number of cell migrations in HepG2 was 188±8 and 595±13, respectively, showing a significant difference between them ( t=46.192, P<0.05). After transfection with the control plasmid and Flag-NUP43 overexpre-ssion plasmid, the number of cell migrations in SK-HEP-1 cells were 136±10 and 447±20, respectively, showing a significant difference between them ( t=24.721, P<0.05). Conclusions:The expression of NUP43 in hepatocellular carcinoma tissues is significantly higher than that in adjacent tissues. Gender, NUP43 expression, TNM staging, and pathological grading are independent influen-cing factors for postoperative survival of hepatocellular carcinoma patients. Overexpression of NUP43 can significantly promote the proliferation and migration of HepG2 and SK-HEP-1 hepatocellular carcinoma cells.

Objective:To investigate the expression of nucleoporin 43 (NUP43) in hepato-cellular carcinoma tissues and its impact on prognosis of patients and proliferation and migration of hepatocellular carcinoma cells.Methods:The retrospective cohort study and experi-mental study were conducted. The clinicopathological data of 102 hepatocellular carcinoma patients who were admitted to The First Affiliated Hospital of Army Medical University from January 2008 to December 2012 were collected. There were 83 males and 19 females, aged 56(range, 19-87)years. The expression of NUP43 in hepatocellular carcinoma tissues was analyzed by immunohistochemical staining. HepG2 and SK-HEP-1 hepatocellular carcinoma cells were cultured in vitro. The Western blot was used to verify the effects of Flag-NUP43 overexpression plasmid in transfected cells. The CCK-8 and cell migration experiments were used to analyze the effect of NUP43 overexpression on HepG2 and SK-HEP-1 hepa-tocellular carcinoma cells. Measurement data of normal distribution were expressed as Mean± SD, and comparison between groups was conducted using the independent sample t test. Measurement data of skewed distribution were represented as M(range). Count data were expressed as absolute numbers, and comparisons between groups was conducted using the paired chi-square test. The Kaplan-Meier method was used to calculate survival time, and Log-rank test was used for survival analysis. The R 4.2.1 software was used to draw survival curves. The COX proportional hazards regre-ssion model was used for univariate and multivariate analyses. Results:(1) Expression of NUP43 in hepatocellular carcinoma and adjacent tissues, and analysis of clinicopathological characteristics of patients with high and low expression of NUP43. Results of immunohistochemical staining showed that NUP43 was mainly expressed in the cytoplasm and nuclear membrane of cells. Of 102 hepatocellular carcinoma tissue samples, there were 49 samples with low expression of NUP43 and 53 samples with high expression of NUP43. Of 102 hepatocellular carcinoma adjacent tissue samples, there were 80 samples with low expression of NUP43 and 22 samples with high expression of NUP43. There was a significant difference in the expression of NUP43 between hepatocellular carcinoma and adjacent tissues ( χ2=16.505, P<0.05). Of 102 hepatocellular carcinoma tissue samples, there were significant differences in tumor diameter, pathological grading, and intrahepatic metastasis between the patients with low expression of NUP43 and the patients with high expression of NUP43 ( χ2=5.104, 23.217, 4.169, P<0.05). (2) Survival of hepatocellular carcinoma patients and prognostic factors analysis. The follow-up time of 102 hepatocellular carcinoma patients was 17.9(range, 0.1-107.9)months. The postoperative 1-, 3-, and 5-year overall survival rates were 79.59%, 53.06% and 34.69% for the patients with low expression of NUP43, versus 52.83%, 18.87%, and 9.43% for the patients with high expre-ssion of NUP43, showing a significant difference between them ( χ2=27.071, P<0.05). Results of multi-variate analysis showed that gender, NUP43 expression, TNM staging, and pathological grading were independent influencing factors for postoperative survival in patients with hepatocellular carcinoma ( hazard ratio=1.846, 2.206, 2.040, 2.177, 95% confidence interval as 1.231-2.768, 1.419-3.429, 1.322-3.148, 1.377-3.254, P<0.05). (3) Effects of NUP43 overexpression on the proliferation and migration of HepG2 and SK-HEP-1 hepatocellular carcinoma cells. Western blot analysis showed that transfection of Flag-NUP43 overexpression plasmid significantly increased the expression of NUP43 in HepG2 and SK-HEP-1 cells. Results of CCK-8 experiment showed that after transfection with the control plasmid and Flag-NUP43 overexpression plasmid, the cell proliferation indices of HepG2 were 0.79±0.07 and 1.47±0.05, respectively, showing a significant difference between them ( t=19.402, P<0.05). After transfection with the control plasmid and Flag-NUP43 overexpression plasmid, the cell proliferation indices of SK-HEP-1 cells were 0.59±0.05 and 0.95±0.05, respectively, showing a significant difference between them ( t=15.753, P<0.05). Results of cell migration experiments showed that after transfection with the control plasmid and Flag-NUP43 overexpression plasmid, the number of cell migrations in HepG2 was 188±8 and 595±13, respectively, showing a significant difference between them ( t=46.192, P<0.05). After transfection with the control plasmid and Flag-NUP43 overexpre-ssion plasmid, the number of cell migrations in SK-HEP-1 cells were 136±10 and 447±20, respectively, showing a significant difference between them ( t=24.721, P<0.05). Conclusions:The expression of NUP43 in hepatocellular carcinoma tissues is significantly higher than that in adjacent tissues. Gender, NUP43 expression, TNM staging, and pathological grading are independent influen-cing factors for postoperative survival of hepatocellular carcinoma patients. Overexpression of NUP43 can significantly promote the proliferation and migration of HepG2 and SK-HEP-1 hepatocellular carcinoma cells.