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@#Objective To investigate the expression of centromere protein U(CENPU)in the intestinal tissues of patients with colon cancer,and to analyze the effect of CENPU expression level on the prognosis of patients with colon cancer combined with bioinformatics.Methods Firstly,the expression of CENPU in cancer tissues and normal tissues of colon cancer patients was analyzed by the expression of CENPU in tissues was further verified by real-time quantitative real time polymerase chain reaction(qRT-PCR),Western blot(WB)and immunohistochemistry(IHC).Combined with clinical data,univariate and multivariate Cox regression are used to analyze the correlation between CENPU expression and clinical case parameters of colon cancer patients.Then,the predictive effect of CENPU expression on the prognosis of colon cancer patients are explored by drawing receiver operating characteristic(ROC)curve and Kaplan-Meier survival curve.Finally,the possible molecular mechanism of the effect of CENPU expression on the progression of colon cancer are analyzed by bioinformatics.Results By qRT-PCR,WB and IHC experiments,we find that compared with normal tissues,the expression of CENPU in cancer tissues of colon cancer patients is significantly increased.Cox regression analysis show that the expression of CENPU is significantly correlated with the age and TNM stage of patients,and is a risk factor affecting the prognosis of patients.Kaplan-Meier survival curve analysis show that colon cancer patients with high CENPU expression has significantly lower survival rates.ROC curve show that the model based on CENPU expression has a high predictive power for the prognosis of colon cancer patients area under the curve(AUC=0.832).Bioinformatics analysis show that CENPI,CENPN,CENPD,CENPK,CENPP,CENPM,CENPQ,CENPH,NDC80 and ITGB3BP have significant interaction with CENPU gene.CENPU is involved in DNA repair,MYC/TARGETS/V1 and PI3K/AKT/MTOR signaling pathways.Conclusion High expression of CENPU in cancer tissues of patients with colon cancer is significantly associated with poor prognosis of patients,suggesting that CENPU is expected to be a potential target for early diagnosis and prognosis prediction of patients with colon cancer.
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Objective:To examine the expression of transmembrane emp24 domain-containing protein 4(TMED4) in liver tissue of patients with hepatocellular carcinoma, and to investigate the effects of TMED4 gene on the proliferation and migration of hepatocellular carcinoma cells and related molecular mechanisms. Methods:The expression of TMED4 at protein level in liver cancer tissue and paracancerous tissue of patients with hepatocellular carcinoma were detected by Western blotting and immunohistochemical stainning, and the correlation between its expression and clinicopathological features was analyzed. The effects of TMED4 overexpression or knockdown on proliferation, migration and healing ability of hepatocellular carcinoma cells in vitro and in vivo were determined by cell proliferation test, Transwell test, wound healing test and subcutaneous tumor formation in nude mice. The molecular mechanism of TMED4 in regulating the biological behavior of hepatocellular carcinoma cells was preliminarily explored by pathway analysis. Independent sample t test, Mann-Whitney U test and chi-square test were used for statistical analysis. Results:The results of Western blotting showed that the expression of TMED4 at protein level in hepatocellular carcinoma tissue was lower than that in paracancerous tissue(0.52±0.29 vs. 0.83±0.22), and the difference was statistically significant( t=2.54, P=0.022). The results of immunohistochemical examination indicated that the expression of TMED4 at protein level in liver cancer tissue was lower than that in paracancerous tissue(5.46±3.37 vs. 7.58±3.08), and the difference was statistically significant( t=3.49, P<0.001). The expression of TMED4 at protein level was significantly correlated with vascular invasion and Barcelona clinic liver cancer stage( χ2=6.83 and 4.20, P=0.009 and 0.040). The results of cell proliferation assay showed that the absorbance value of SMMC-7721 cells in TMED4 overexpression group was lower than that in control group(1.38±0.05 vs. 2.37±0.08), while the optical density value of HepG2 in TMED4 knockdown group was higher than that in control group(0.76±0.04 vs. 0.54±0.01), and the differences were statistically significant( t=18.23 and 8.85, both P<0.001). The results of Transwell test showed that the number of migrated SMMC-7721 cells in TMED4 overexpression group was less than that in control group(286.30±13.01 vs. 439.70±12.34), while the number of migrated HepG2 cells in TMED4 knockdown group was higher than that in control group(249.00±6.00 vs. 160.00±6.56), and the differences were statistically significant( t=14.81 and 17.34, both P<0.001). The wound healing test showed that the healing rate of SMMC-7721 cells in TMED4 overexpression group was lower than that in control group((0.21±0.01)% vs.(0.45±0.01)%), the healing rate of HepG2 cells in TMED4 knockdown group was higher than that in control group((0.46±0.01)% vs.(0.20±0.01)%), and the differences were statistically significant( t=200.10 and 30.46, both P<0.001). The results of subcutaneous tumor formation assay in nude mice showed that the growth rate of cells in TMED4 overexpression group was slower than that in control group. After cell inoculation for 6 weeks, the subcutaneous tumor volume of mice in TMED4 overexpression group was smaller than that in control group(27.36 mm 3(138.70 mm 3) vs. 1 741.62 mm 3(1 783.39 mm 3)), the tumor weight was lower than that in control group(0.06 g(0.14 g) vs. 1.46 g(1.09 g)), and the differences were statistically significant(both Z=-2.31, both P<0.001). The results of Western blotting showed that the expression of Snail at protein level in SMMC-7721 cells of the TMED4 overexpression group was lower than that of the control group(0.32±0.01 vs. 0.90±0.03), the protein level of Snail in HepG2 cells of TMED4 knockdown group was higher than that of control group(1.03±0.01 vs. 0.97±0.01), and the differences were statistically significant( t=28.49 and 12.31, both P<0.001). The results of real time fluorescent quantitative polymerase chain reaction showed that the expression of Snail at mRNA level in SMMC-7721 cells of TMED4 overexpression group was lower than that of control group(0.13±0.05 vs. 1.00±0.15), the expression of Snail at mRNA level in HepG2 cells of TMED4 knockdown group was higher than that of control group(1.25±0.32 vs. 0.21±0.14), and the differences were statistically significant( t=9.62 and 5.10, P<0.001 and P=0.007). Conclusion:TMED4 may affect the proliferation and migration of hepatocarcinoma cells by regulating the expression of Snail, and which is expected to become a potentially therapeutic target for hepatocellular carcinoma.
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Objective:To investigate the expression and correlation of EHD2 and E-cadherin in hepatocellular carcinoma (HCC). Meth-ods:Western blot and immunohistochemistry were used to detect the expression of EHD2 and E-cadherin in HCC specimens and adja-cent noncancerous tissues. The correlations of EHD2 and E-cadherin with the clinicopathological characteristics and prognosis of pa-tients were further analyzed using Pearsonχ2 test and Kaplan-Meier method. Results:EHD2 expression, along with E-cadherin, was markedly reduced in HCC tissues than in adjacent noncancerous tissues. Moreover, EHD2 and E-cadherin expression were correlated with histological grade, microvascular invasion, and lymph node metastasis (P<0.05). Kaplan-Meier survival analysis showed that HCC patients with decreased EHD2 and E-cadherin expression had shorter overall survival time than those with higher expression. Conclu-sion:The abnormal expression of EHD2 and E-cadherin possibly promote HCC. Detection of EHD2 and E-cadherin may be valuable for diagnosing HCC and evaluating malignancy extent and prognosis.
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Objective:To explore the expression of C-terminal binding protein 2 (CtBP2) in human esophageal carcinoma and its relationship with clinicopathological parameters and survival. Methods:The expression levels of CtBP2 in eight cases of fresh frozen specimens of esophageal squamous cell carcinoma (ESCC) and the adjacent esophageal tissues were detected by Western blot. Immuno-histochemistry was used to detect CtBP2 expression in 90 samples of ESCC tissues and adjacent non-tumor tissues. Based on patient in-formation and follow-up data, the correlation of CtBP2 expression with patients' clinicopathological characteristics and overall survival was further evaluated using Fisher's exact test and Kaplan-Meier method, respectively. Results: Immunohistochemistry and Western blot analysis showed that CtBP2 expression in tumor tissues was higher than that in adjacent non-tumor tissues. CtBP2 expression was significantly correlated with histologic grade (P=0.002) and depth (P=0.032). Kaplan-Meier analysis demonstrated that high CtBP2 ex-pression was correlated with a short survival time. Conclusion:CtBP2 expression was upregulated in ESCC tissues, indicating that it may play a role in the oncogenesis and development of ESCC.
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Objective To study the value of serum galectin-3 detected by nanomagnetic beads sorting time resolved fluoroimmunoassay (TRFIA) tandem analysis in the diagnosis of pancreatic cancer.Methods The serum of 88 cases of pancreatic cancer,50 cases of acute pancreatitis,10 cases of pancreatic cysts,and 20 cases of healthy volunteers as control were collected.First,galectin-3 antibody coupled nanomagnetic-beads was used to sort the semm,then TRFIA was applied to detect the level of galectin 3,and the result was compared with that of routine TRFIA.ROC curve was constructed to determine the cutoff value and sensitivity for pancreatic cancer diagnosis.Results The method of nanomagnetic beads sorting TRFIA detected the level of galectin 3 between O.78 and 100 ng/ml in a linear manner,the intra-CV was ≤6.38%,and inter-CV was ≤7.22%,and average recovery rate was 105.3%.The serum level of galectin-3 in control group by nanomagnetic beads sorting TRFIA was 0.38 (0.02 ~ 3.06) ng/ml,which was significantly higher than that detected by routine TRFIA [0.18 (0.00 ~ 2.64) ng/ml,P =0.000).The serum levels of galectin-3 by nanomagnetic beads sorting TRFIA in pancreatic cancer,acute pancreatitis,pancreatic cysts and healthy volunteers were 4.85(0.00 ~42.67),0.69(0.00~ 13.62),0.70(0.00 ~ 14.54),0.38(0.02 ~ 3.06) ng/ml,and the level of galectin-3 in pancreatic cancer was significantly higher than that in acute pancreatitis,pancreatic cysts and healthy volunteers group (P <0.01),while the difference among the other three group was not significantly different.The AUC of galectin-3 for pancreatic cancer was 0.851 ± 0.040,95% CI:0.772 ~ 0.929.While using 1.07 ng/ml as the cut-off value,the positive rates for the diagnosis of pancreatic cancer,acute pancreatitis,pancreatic cysts and healthy volunteers were 84.1% (74/88),34.0% (17/50),20.0% (2/10),10.0% (2/20),and the sensitivity for diagnosis of pancreatic cancer was significantly higher than those in other 3 groups (P < 0.01).Conclusions Nanomagnetic beads sorting TRFIA tandem analysis method can enrich serum galectin-3,and increase the sensitivity of detection for pancreatic cancer and enhance the diagnostic value of galectin-3 for pancreatic cancer.
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ObjectiveTo establish the time-resolved fluoroimmunoassay (TRFIA) method for the detection of serum galectin-3 and investigate the clinical value of serum galectin-3 for the diagnosis of pancreas cancer.MethodsMonoclonal anti-human galectin - 3 antibody and biotinylated polyclonal antibody were used to establish the sandwich TRFIA for detection of serum galectin-3.The optimal experimental condition was studied.Serum levels of galectin-3,CEA and CA19-9 in the patients with pancreatic cancer,benign pancreatic mass,pancreatitis,and healthy controls were measured.The diagnostic value of serum galectin-3,CEA and CA19-9 for pancreas cancer was studied.ResultsThe linearity of the TRFIA for detection of serum galectin3 tanged between 0 to 100 μg/L.The within-run CV and between-run CV were ≤6.45% and ≤8.68%,respectively,and the average recovery was 106.6%.The level of serum galectin-3 was 4.93 ( 0.85 ~ 23.80) μg/L in pancreatic cancer group,which were significantly higher than those in benign pancreatic mass [2.83 ( 2.17 ~ 4.06) μg/L ],pancreatitis [ 2.62 (0.55 ~ 9.76 ) μg/L ],and healthy controls group [ 1.88 ( 0.59 ~ 3.94) μg/L] (P <0.05).By using 3.77 μg/L as the cut-off point,the smsitivity,specificity for the diagnosis of pancreatic cancer was 75.5% and 90.9%.The levels of Gal 3 and CEA,CA19-9 was not correlated ( r =0.1321,P =0.3761 ; r =0.0920,P =0.5384).Combined determination of galactin-3 and CEA,CA19-9 levels could increase the diagnostic sensitivity to 92%.ConclusionsTRFIA method for the detection of galactin-3 is sensitive and stable.Galectin-3 could be a potentially novel serum tumor marker of pancreatic cancer.
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Objective To explore the expression of calcium binding protein (S100A11) and its clinical significances in human hepatocellular carcinoma (HCC) tissue and blood plasma. Methods The expressions of S100A11 in 46 cases of HCC tissues and their paracancerous tissues were detected by immunohistochemistry. The relationship between S100A11 expression level in HCC tissues and clinical parameters was analyzed. The S100A11 expression levels in blood plasma of HCC patients (62 cases), liver cirrhosis patients (32 cases), chronic hepatitis patients (30 cases) and healthy subjects (30 cases) were detected. The sensitivity and specificity of S100A11, alpha fetoprotein (AFP) and γ-glutamyl transpeptidase Ⅱ (GGT-Ⅱ ) in HCC diagnosis were compared. Results The positive rate of S100A11 in HCC tissue (78. 3%) was significantly higher than that in paracancerous tissues (19. 6%) (P<0. 05). The expression level was correlated with the degree of differentiation, the lower differentiation degree with the higher expression level. According to ROC curve, if the cutoff points for diagnosis was set at 7. 3 μg/L, the positive rate of S100A11 in HCC patients blood plasma was 30. 6% , which was significantly higher than that in the blood plasma of patients with liver cirrhosis, patients with chronic hepatitis and healthy persons (P<0. 05). There was no correlation between S100A11 and AFP or GGT-Ⅱ in the blood plasma of HCC patients. These three indicators were complementary in HCC diagnosis, and the diagnostic sensitivity increased to 84.5% with combined detection. Conclusions S100All may be related to HCC genesis and development. The HCC diagnostic sensitivity may be increased with combined detection of S100All ,AFP and GOT- Ⅱ.