Original article: The expression of CFL1 and N‐WASP in esophageal squamous cell carcinoma and its correlation with clinicopathological features

Cofilin1 (CFL1) is an actin‐modulating protein, which belongs to the ADF/Cofilin family. Neural Wiskott–Aldrich syndrome protein (N‐WASP) is the key regulator of the actin cytoskeleton, a member of Wiskott‐Aldrich syndrome protein family. They have been suggested to be involved in cancer cell invasion and metastasis. In this study, the expression patterns of CFL1 and N‐WASP in normal esophageal mucosa and esophageal squamous cell carcinoma (ESCC) and their correlation with clinical characteristics were investigated. Immunohistochemical staining showed that CFL1 was expressed in nuclear and cytoplasm of cancer cells. However, N‐WASP was mainly found in the cytoplasm of the cancer cells. There were significant evidences that proved that CFL1 is correlated with clinicopathological factors in ESCC, such as infiltration depth, lymph node metastasis and pathological staging (P < 0.05). It is also proved that N‐WASP is related to lymph node metastasis and pathological staging in ESCC (P < 0.05). Kaplan–Meier analysis showed that there was no correlation between CFL1 and N‐WASP protein expression and survival (P > 0.05). Moreover, the mRNA expression of CFL1 and N‐WASP was detected by quantitative real time PCR in 70 tissue specimens. The results showed that CFL1 mRNA level was over‐expressed in ESCC tissue (P < 0.05), while N‐WASP mRNA expression level was not different between cancerous tissues and adjacent normal esophageal mucosa (P > 0.05). Also, CFL1 mRNA expression was significantly associated with regional lymph node metastasis and pathological staging (P < 0.05). Kaplan–Meier analysis showed that there was no correlation between CFL1 and N‐WASP mRNA expression and survival (P > 0.05). Our findings suggested that CFL1 and N‐WASP may play an important role in the tumorigenesis of ESCC, and to be the candidate novel biomarkers for the diagnosis and prognosis of ESCC. These findings may have implications for targeted therapies in patients with ESCC.     

Pilot study special AT‐rich sequence‐binding protein 1 investigating as a potential biomarker for esophageal squamous cell carcinoma

In the present study, we aimed to evaluate the expression of special AT‐rich sequence‐binding protein 1 (SATB1) in esophageal squamous cell carcinoma (ESCC) and assess the correlation between its expression and the clinicopathological features and prognosis of the disease. SATB1 expression in ESCC tissue was determined by using immunohistochemical analysis, quantitative real‐time polymerase chain reaction, and western blot analysis. The relationship between SATB1 expression and clinicopathological features was examined by using the chi‐squared test, and the survival rate was calculated by using the Kaplan–Meier survival curve. The correlation between the indicators and patient survival was estimated by using a Cox regression analysis. High SATB1 expression in was detected in 48.3% and 7.8% of ESCC and normal esophagus tissues (P < 0.05), respectively. SATB1 expression did not significantly correlate with clinicopathological features. The Kaplan–Meier curve indicated that patients with high SATB1 expression had significantly shorter survival than those with low SATB1 expression. In a multivariate Cox regression model, high SATB1 expression was identified as an independent prognostic factor for patients with ESCC. In conclusion, these results suggest that high SATB1 expression is predictive of poor prognosis in ESCC and may be a promising new candidate for targeted therapies for ESCC.     

Prognostic relevance and therapeutic implications of centromere protein F expression in patients with esophageal squamous cell carcinoma

Centromere protein F (CENP‐F), a cell cycle‐regulated centromere protein, has been shown to affect numerous tumorigenic processes. This study aimed to clarify the prognostic significance of CENP‐F expression in patients with esophageal squamous cell carcinoma (ESCC). The levels of CENP‐F messenger RNA and protein were higher in ESCC cell lines than in the normal tissues. An immunohistochemical analysis of paired tissue specimens showed that the CENP‐F expression was higher in tumorous tissues than in the adjacent non‐tumorous tissues (P < 0.001). Moreover, there was a significant correlation between CENP‐F expression and gender (P = 0.012), clinical stage (P = 0.039), and T classification (P = 0.026). Patients with higher CENP‐F expression had shorter overall survival than those with lower CENP‐F expression (P = 0.009). Multivariate Cox analysis indicated that CENP‐F expression is an independent prognostic factor for overall survival (hazard ratio = 0.582, 95% confidence interval = 0.397–0.804, P = 0.041). Importantly, it was found that zoledronic acid (ZOL) could significantly enhance the chemotherapeutic sensitivity of ESCC cell lines with high CENP‐F expression to cisplatin, although ZOL alone only exhibited a minor inhibitory effect to ESCC cells. In summary, these findings demonstrate that CENP‐F may serve as a valuable molecular marker for predicting the prognosis of ESCC patients. In addition, the data indicate a potential benefit of combining ZOL with cisplatin in ESCC, suggesting that CENP‐F expression may have therapeutic implications.     

Expression of breast cancer anti‐estrogen resistance 1 in relation to vascular endothelial growth factor,p53, and prognosis in esophageal squamous cell cancer

The purpose of this study was to clarify the role of breast cancer anti‐estrogen resistance 1 (BCAR1) expression in relation to vascular endothelial growth factor (VEGF), p53, and proliferation in esophageal squamous cell cancer (ESCC). Expression of BCAR1, VEGF, p53, and the ki‐67 proliferative index were examined by tissue microarray and immunohistochemistry in 106 specimens with ESCC and matched adjacent normal tissues. Among them, 40 cases were simultaneously examined by Western blot. Both Western blot and immunohistochemistry showed that BCAR1 expression was substantially higher in ESCC than in adjacent normal tissues (P < 0.001). BCAR1 expression was significantly connected with degree of tumor differentiation, with poorly differentiated tumors showing higher BCAR1 expression (P < 0.001). BCAR1 expression was significantly and positively correlated with VEGF and p53 expression levels (r= 0.541, P < 0.001; r= 0.374; P < 0.001) but not proliferative index (r= 0.44; P= 0.066). Additionally, a significant relationship was also observed between VEGF and p53 (r= 0.321; P= 0.001). Kaplan–Meier survival analysis revealed that patients with high BCAR1 expression had significantly shorter survival times than those with low BCAR1 expression levels (median survival 40 months vs. 27 months, P= 0.09). Multivariate analysis also revealed that levels of BCAR1 expression (hazard ratio 2.250, P= 0.015) was a significant and independent prognostic indicator. High expression of BCAR1 is associated with elevated VEGF and p53 expression levels, as well as poor prognosis in ESCC. Therefore, BCAR1 may be a potential candidate for predicting prognosis and a new therapy target for ESCC.     

Expression and prognostic relevance of p21WAF1 in stage III esophageal squamous cell carcinoma

The prognostic effect of p21^WAF1 expression on esophageal squamous cell carcinoma patients is controversial. Further clarifying the effect of this protein is beneficial for optimizing the patient outcomes. In the current study, we investigated the expression of p21^WAF1 protein in 189 specimens of stage III ESCC by immunohistochemistry. As shown by the Kaplan–Meier curve, the overall survival rate of the positive‐expression group was significantly higher than that of the negative‐expression group (P < 0.05). No significant correlation was observed between p21^WAF1 expression and clinicopathological parameters in terms of gender, age, tumor location, tumor grade, pathological stage, and number of regional lymph node metastases (P > 0.05). We concluded that p21^WAF1 played an intricate role in the tumorigenesis and development of ESCC. p21^WAF1 could serve as a positive prognostic predictor for stage III ESCC patients.     

The expression of CXCR4 and its relationship with matrix metalloproteinase‐9/vascular endothelial growth factor in esophageal squamous cell cancer

Esophageal cancer (EC) is a highly aggressive neoplasm with poor prognosis. The main reason for this disappointing outcome is the strong behavior of esophageal cancer cell's invasion and metastasis. CXC chemokine receptor 4 (CXCR4) was found to be expressed in many tumors and significantly correlated with invasion, angiogenesis, metastasis, and prognosis. In the present study, we investigated the expressions of CXCR4, matrix metalloproteinase‐9 (MMP‐9), and vascular endothelial growth factor (VEGF) in esophageal squamous cell cancer (ESCC) and analyzed the relationship among the three proteins. Sections of paraffin‐embedded tissues were obtained from 127 patients with ESCC undergoing esophagectomy at Zhongshan Hospital, Fudan University in 2005. The CXCR4, MMP‐9, and VEGF expressions in EC tissues were evaluated according to the immunohistochemical staining area and intensity. The correlations between patients' prognosis and covariates were analyzed by Kaplan–Meier method (univariate analysis) and Cox regression (multivariate analysis). The overall expression rate of CXCR4, MMP‐9, and VEGF was 88.2%, 93.7%, and 79.5%, respectively. CXCR4 expression was significantly associated with tumor grade, tumor size, tumor depth, regional lymph node metastasis, and tumor, node, metastasis (TNM) stage (P < 0.05). MMP‐9 expression was significantly associated with age and tumor grade (P < 0.05). VEGF expression was significantly associated with tumor grade, tumor depth, and TNM stage (P < 0.05). CXCR4 expression was positively correlated with MMP‐9 expression (P < 0.01, r= 0.365) and VEGF expression (P < 0.01, r= 0.380). However, there was no significant correlation between MMP‐9 and VEGF expression (P > 0.05). In univariate analysis, CXCR4 expression, tumor size, tumor depth, lymph node metastasis, and TNM stage were correlated with patients' prognosis (P < 0.05); in multivariate analysis, tumor size and lymph node metastasis were the independent factors of poor prognosis. CXCR4 was highly expressed in ESCC and correlated with MMP‐9, VEGF, clinicopathological features and prognosis. We speculated CXCR4 play an important role during the progression of this disease and there might be some regulatory mechanism existing between CXCR4 and MMP‐9/VEGF in ESCC.     

Human papillomavirus (HPV) infection and the risk of esophageal squamous cell carcinoma

There is currently no consensus on the relationship between human papillomavirus (HPV) infection and the pathogenic process of esophageal squamous cell carcinoma (ESCC). Therefore, a retrospective study was performed to explore the association between HPV infection and ESCC, where 225 patients with diagnosed ESCC and 224 matched controls were enrolled in the study and stratified according to smoking and alcohol consumption. Enzyme‐linked immunosorbent assay was used to determine seropositivity to HPV by the detection of either IgG or IgM anti‐HPV antibodies. In the non‐smoking and non‐alcohol‐consuming subgroup, the incidence of ESCC of HPV seropositive subjects was similar with that of HPV seronegative subjects (P= 0.737, odds ratio [OR] 1.14, 95% confidence interval [CI] 0.54–2.40). However, in the smoking subgroup, there was a significant difference in the incidence of ESCC between HPV seropositive subjects and HPV seronegative subjects (P= 0.009, OR 2.22, 95% CI 1.22–4.04). In addition, there was a significantly higher association of the development of ESCC in HPV seropositive patients that smoke and drink than those that do not (P < 0.001, OR 10.31, 95% CI 4.04–26.29). Therefore, HPV infection is not an independent risk factor for developing ESCC in the non‐smoking and non‐alcohol‐consuming group. For smokers, however, HPV infection increases the risk of the incidence of ESCC.     

Association of mu‐opioid receptor expression with lymph node metastasis in esophageal squamous cell carcinoma

The mu‐opioid receptor (MOR), a membrane‐bound G protein‐coupled receptor, is the main target for opioids in the nervous system. MOR1 has been found in several types of cancer cells and reported to be involved in tumor progression and metastasis. However, the expression and clinical significance of MOR1 in esophageal squamous cell carcinoma (ESCC) remain unclear. In our study, the expression of MOR1 was confirmed in ESCC cell lines (KYSE180, KYSE150, and EC109) by Western blot. MOR1 was also detected on tissue microarrays of ESCC samples in 239 cases using immunohistochemical staining. We found that MOR1 was mainly located in the cytoplasm and occasionally occurred in the membrane or nucleus of ESCC cells. Moreover, results indicated that MOR1 expression in the cytoplasm was associated with lymph node metastasis (R = 0.164, P = 0.008, Kendall's tau‐b‐test). No more associations were found between MOR1 expression status and other clinical parameters. However, no statistical significant differences were found between MOR1 expression in the cytoplasm, nucleus/membrane, and the overall survival of ESCC patients (P = 0.848; P = 0.167; P = 0.428, respectively, log‐rank test). Our results suggest that the cytoplasmic MOR1 may be a high‐risk factor for lymph node metastasis of ESCC patients. We also hypothesize that MOR1 agonists used in ESCC patients should be prudent, and opioid receptor antagonists may be novel therapeutic drugs for ESCC patients.     

Diagnostic and prognostic value of serum L1-cell adhesion molecule in esophageal squamous cell carcinoma

Objective

L1 cell adhesion molecule (L1CAM) has been found to be dysregulated in several types of human cancers. Here, we aimed to determine the level of soluble L1CAM in serum of patients with esophageal squamous cell carcinoma (ESCC).

Hepatic Toll‐like receptor 3 expression in chronic hepatitis C genotype 1 correlates with treatment response to peginterferon plus ribavirin

Summary. Recent studies have shown that enhanced hepatic expression of several innate immune genes predicts non‐response to 48 weeks of peginterferon plus ribavirin in chronic hepatitis C genotype 1. This study aimed to further address how gene expression of TLR3/RIG‐I signalling correlates with the outcome of the 72‐week extended treatment regimen. Relative hepatic mRNA expression and copy numbers of positive‐ and negative‐strand hepatitis C virus (HCV) RNA were determined by real‐time PCR in 49 patients. Then, a 48‐week peginterferon‐α2b plus ribavirin treatment was commenced and extended to 72 weeks in cases of HCV RNA clearance after week 12. High rate of sustained virologic response was seen both in patients with early HCV clearance (85% [11/13]) and slow virologic responders (85% [11/13]) (per protocol analysis). The response was associated with low TLR3 expression (median, 0.9; range, 0–4.2 vs median, 1.9; range, 0.4–4.9; P = 0.004) but had no relation to the expression of TRIF (P = 0.315), RIG‐I (P = 0.953), IPS‐1 (P = 0.425), IRF3 (P = 0.329) and interferon‐β (P = 0.584). ROC curve analysis identified TLR3 expression of <1.5 as the best cut‐off for predicting response (positive and negative predictive values, 89% [16/18] and 70% [14/20], respectively). The expression was not affected by HCV replication but was higher in female patients (P = 0.043). Multivariate analysis showed TLR3 to be a single baseline predictor (odds ratio 18.5 [95% CI 3.2–111], P = 0.001). Low hepatic TLR3 expression is a novel predictor of response to peginterferon plus ribavirin in genotype 1 patients.     

Prognostic significance of preoperative absolute peripheral monocyte count in esophageal squamous cell carcinoma

The objective of this study was to investigate the prognostic value of peripheral blood monocytes in esophageal squamous cell carcinoma (ESCC) patients who underwent esophagectomy. Records from 218 consecutive patients with histologically diagnosed ESCC who underwent esophagectomy at Qilu Hospital of Shandong University from January 2007 to December 2008 were retrospectively reviewed. The median disease‐free survival (DFS) of this cohort was 29.0 months, and the 5‐year DFS rate was 34.4%. The median overall survival (OS) was 35.0 months, and the 5‐year OS rate was 37.6%. The cut‐off value of 0.42 × 10⁹/L for the absolute monocyte count (AMC) was chosen as optimal to discriminate between survival and death by applying receiver operating curve analysis. There were 131 patients (60.1%) who had high AMC (≥0.42 × 10⁹/L) preoperatively. We found that AMC was significantly associated with gender, tumor location, and platelet count. Kaplan–Meier survival analysis of patients with high preoperative AMC had a significant worse prognosis for DFS (high vs. low: 27.5% vs. 39.0%, P = 0.015) and OS (high vs. low: 31.1% vs. 44.8%, P = 0.009) than those with low preoperative AMC. In a multivariate analysis, preoperative AMC was an independent prognostic factor for DFS (P = 0.025, hazard ratio [HR]: 1.469, 95% confidence interval [CI]: 1.050–2.054) and OS (P = 0.015, HR: 1.547, 95% CI: 1.088–2.200). In addition, among 140 patients without both preoperative and postoperative therapy, significantly worse OS (P = 0.012) and marginally reduced DFS (P = 0.079) were found in the high AMC cohort versus the low AMC cohort. A higher preoperative absolute peripheral monocyte count can be considered as a useful prognostic marker of ESCC patients who underwent esophagectomy.     

Cognitive test scores in male adolescent cigarette smokers compared to non‐smokers: a population‐based study

Background Although previous studies indicate that people with lower intelligence quotient (IQ) scores are more likely to become cigarette smokers, IQ scores of siblings discordant for smoking and of adolescents who began smoking between ages 18–21 years have not been studied systematically. Methods Each year a random sample of Israeli military recruits complete a smoking questionnaire. Cognitive functioning is assessed by the military using standardized tests equivalent to IQ. Results Of 20 221 18‐year‐old males, 28.5% reported smoking at least one cigarette a day (smokers). An unadjusted comparison found that smokers scored 0.41 effect sizes (ES, P < 0.001) lower than non‐smokers; adjusted analyses remained significant (adjusted ES = 0.27, P < 0.001). Adolescents smoking one to five, six to 10, 11–20 and 21+ cigarettes/day had cognitive test scores 0.14, 0.22, 0.33 and 0.5 adjusted ES poorer than those of non‐smokers (P < 0.001). Adolescents who did not smoke by age 18, and then began to smoke between ages 18–21 had lower cognitive test scores compared to never‐smokers (adjusted ES = 0.14, P < 0.001). An analysis of brothers discordant for smoking found that smoking brothers had lower cognitive scores than non‐smoking brothers (adjusted ES = 0.27; P = 0.014). Conclusion Controlled analyses from this large population‐based cohort of male adolescents indicate that IQ scores are lower in male adolescents who smoke compared to non‐smokers and in brothers who smoke compared to their non‐smoking brothers. The IQs of adolescents who began smoking between ages 18–21 are lower than those of non‐smokers. Adolescents with poorer IQ scores might be targeted for programmes designed to prevent smoking.     

Original article: Upregulation of caspase‐3 expression in esophageal cancer correlates with favorable prognosis: an immunohistochemical study from a high incidence area in northern China

Caspase‐3 plays an important role as the key effector during apoptosis, but there are very few studies of caspase‐3 in esophageal squamous cell carcinoma (ESCC). The purpose of this study was to investigate the expression and prognostic significance of caspase‐3 in ESCC from Linzhou City, a high incidence area in northern China. All 64 patients underwent esophagectomy for ESCC between January 2002 and December were enrolled in this study. Caspase‐3 expression was assessed by immunohistochemistry (IHC) in primary ESCC and paired normal esophageal epithelium. The positive rate of caspase‐3 expression was higher in ESCC than in normal esophageal epithelium (79.7% vs. 50.0%, Chi‐square = 12.372, P= 0.001). Caspase‐3 expression was correlated with tumor cell differentiation (Phi = 0.717, P < 0.001), tumor infiltration depth (Phi =?0.334, P= 0.008), and pathologic TNM (pTNM) staging (rs =?0.268, P= 0.032). Patients in caspase‐3 positive group had a significantly better 5‐year overall survival than those in the negative group (77.4% vs. 35.9%, χ²= 7.344, P= 0.007). Our results showed that caspase‐3 expression was upregulated in ESCC compared with normal esophageal epithelium in population of Chinese high incidence area, and patients with caspase‐3 positive expression had better prognosis. Therefore, caspase‐3 immunostaining could be a simple and useful tool for predicting survival in ESCC patients.     

Insulin resistance raises the risk for recurrence of stage I hepatocellular carcinoma after curative radiofrequency ablation in hepatitis C virus‐positive patients: A prospective,case series study

Aim: Several studies have reported that insulin resistance raises the risk of primary hepatocellular carcinoma (HCC). We conducted a prospective, case series study to test the impact of insulin resistance on the recurrence after curative radiofrequency ablation (RFA) of stage I HCC in HCV‐positive patients. Methods: From January 2006 to December 2007, 226 consecutive patients underwent treatment for primary HCC at our institutions, including 37 stage I cases. Among them, 33 were HCV‐positive, and three, six and 24 received curative surgery, transarterial chemoembolization or RFA, respectively. In the 24 patients treated with RFA, recurrence‐free survival was analyzed using the Kaplan–Meier method. The factors contributing to recurrence of HCC were subjected to univariate and multivariate analyses using the Cox proportional hazards model. Insulin resistance was estimated by the Homeostatic Model Assessment of Insulin Resistance (HOMA‐IR). Results: Kaplan–Meier analysis showed that the recurrence‐free survival was lower in patients with higher HOMA‐IR (>2.3, P = 0.0252) or with lower serum albumin level (<3.3 g/dL, P = 0.0004). In the univariate analysis, HOMA‐IR (P = 0.0420) and albumin (P = 0.0036) were significantly associated with recurrence of HCC. Multivariate analysis revealed albumin (odds ratio = 0.01, 95% confidence interval = 0.0002–0.015, P = 0.0001) and HOMA‐IR (odds ratio = 3.85, 95% confidence interval = 1.57–14.2, P = 0.0015) to be independent predictors for recurrence of HCC. Conclusion: Serum albumin level and HOMA‐IR were independent risk factors for recurrence of stage I HCC after curative RFA in HCV‐positive patients. Patients with these factors require closer surveillance.     

Association between positive murine double minute 2 expression and clinicopathological characteristics of esophageal squamous cell carcinoma: a meta‐analysis

The correlations of murine double minute 2 (MDM2) T309G and esophageal cancer were elucidated because the association between MDM2 expression states and clinicopathological parameters of esophageal squamous cell carcinoma (ESCC) is controversial. We conducted a meta‐analysis on studies screened from PubMed, Web of Science, Embase, the Cochrane Library, and the Chinese Biomedical Literature Databases that were published before October 2014. All studies describing the association between MDM2 and ESCC were traced. Meta‐analysis was performed using the STATA software (Stata Corp., College Station, TX, USA). A total of 9 studies with 707 cases and 324 controls were included. MDM2 expression was higher in ESCC than in normal esophageal epithelium (odds ratio [OR] 10.38, 95% confidence interval [CI] 6.42–16.78, P < 0.001). High MDM2 expression was associated with early primary tumor stage (T1/T2 vs. T3/T4, OR 0.59, 95% CI 0.38–0.92, P = 0.018) and increased risk of regional lymph node metastasis (N0 vs. N1, OR 1.66, 95% CI 1.03–2.67, P = 0.039). However, no relationship was observed between MDM2 expression and the risk of distant metastasis (OR = 2.09, 95% CI 1.00–4.36, P = 0.050), and MDM2 was not significantly correlated with TP53 expression (OR 1.22, 95% CI 0.53–2.77, P = 0.643). Our analysis suggests that MDM2 acts as a potent marker of early primary tumor stage but higher risk of regional lymph node metastasis in ESCC. However, because of the limited number of studies included, the result should be further clarified by well‐designed prospective studies.