Expression of ALDH1 and TGFβ2 in benign and malignant breast tumors and their prognostic implications

The specific mechanism underlying the role of putative stem cell marker aldehyde dehydrogenase 1 (ALDH1) playing in development and progression of breast cancer is currently unclear. Transforming growth factor β (TGFβ) signaling pathway is reported to be activated in most cancers. Thus a study was initiated to explore possible differences and correlation of ALDH1 and TGFβ2 expression in the most common malignant and benign tumors of the breast in Chinese women. Samples of 75 breast cancer tissues, 30 paracancerous normal tissues, and 39 fibroadenoma breast tissues were investigated for the expression of ALDH1 and TGFβ2 using immunohistochemistry. The positive rates of ALDH1 and TGFβ2 protein were 62.67% and 66.67%, respectively, in breast cancer tissues, which were significantly higher than that in normal fibroadenoma breast (P<0.05) and paracancerous tissues (P<0.01). ALDH1 and TGFβ2 status were significantly associated with tumor histological grade and receptor status (P<0.05). Expression of ALDH1 was found to be positively correlative to TGFβ2 in breast cancer (r = 0.33, P<0.01). Expression of both proteins remained significantly associated with reduced overall survival (OS) by univariate analysis (P<0.05). Multivariate Cox regression analysis showed that ALDH1 expression, tumor stage, and lymph node status are independent prognostic factors in invasive breast cancer patients. Thus ALDH1 and TGFβ2 play important roles in the development of breast cancer. The ALDH1 phenotype is an independent predictor of poor prognosis, and TGFβ2 signaling pathway activation might be involved in the pathological regulation of ALDH1 in breast cancer.     

Expression and clinical significances of Beclin1, LC3 and mTOR in colorectal cancer

Autophagy is related to cancer and other diseases, and compromised autophagy could promote chromosome instability associated with carcinogenesis and tumor progression. The role of autophagy in the growth and metastasis of colorectal cancer (CRC) remains poorly understood. Beclin1 mediates autophagic initiation, and LC3 is a specific marker for autophagy. Inactivation of mTOR caused by cellular hypoxia or energy deficiency induces autophagic activity. This study aims to examine the expression and clinical significance of these proteins in CRC. Immunohistochemistry results showed that the positive expression rates of Beclin1, LC3, and mTOR in cancer tissues were 90.50%, 87.19%, and 46.28%, respectively, which were higher than those in adjacent tissues (P < 0.05). Differentiation degree and lymph node metastasis were associated with LC3 overexpression (P < 0.05) but not with Beclin1 (P > 0.05). Lymph node metastasis was also related to mTOR. Spearman analysis results showed that LC3 expression was positively correlated with Beclin1 but negatively correlated with mTOR (r = 0.593 and -0.165, respectively; P < 0.01). Beclin1 expression was also not associated with mTOR (P > 0.05). Survival analysis further indicated that LC3, mTOR, and lymph node metastasis were independent prognostic factors in CRC. Real-time PCR results and Western blot indicated that Beclin1, LC3, and mTOR expression in CRC was significantly higher than that in adjacent tissues (P < 0.01). The aberrant protein expression may be associated with the development and progression of CRC. The LC3 and mTOR genes must be simultaneously detected to evaluate progression and prognosis of CRC.     

Wnt/β-catenin pathway is required for epithelial to mesenchymal transition in CXCL12 over expressed breast cancer cells

CXCL12 is positively associated with the metastasis and prognosis of various human malignancies. Cancer-associated fibroblasts (CAFs), the main cells secreting CXCL12, are capable of inducing epithelial to mesenchymal transition (EMT) of breast cancer cells. However, it has not been completely understood whether CXCL12 is involved in EMT of breast cancer cells and the underlying mechanisms. The present study aimed to investigate the effects of CXCL12 on the EMT and cancer stem cell (CSC)-like phenotypes formation by transfecting pEGFP-N1-CXCL12 plasmid into MCF-7 cells. Real time-PCR and Western blot analysis demonstrated the successful over expression of CXCL12 in MCF-7 cells. Cell counting kit-8 assay, wound healing assay and Transwell invasion analysis confirmed that over expression of CXCL12 significantly promoted the proliferation, migration and invasion in MCF-7 cells (P<0.05). In addition, ALDH activity was dramatically enhanced compared with parental (P<0.001), accompanied by the notably elevated mRNA and protein levels of OCT-4, Nanog, and SOX2 in CXCL12 overexpressed-MCF-7 cells (P<0.001). Furthermore, we observed the down regulation of E-cadherin and up regulation of vimentin, N-cadherin, and α-SMA in CXCL12 overexpressed-MCF-7 cells (P<0.01). Meanwhile, western blot and immunofluorescence assay showed that over expression of CXCL12 activated Wnt/β-catenin pathway to induce EMT of MCF-7 cells, as evidenced by the increased expression of E-cadherin after silencing β-catenin by siRNA interference (P<0.001). Collectively, our findings suggested that over expression of CXCL12 could trigger EMT by activating Wnt/β-catenin pathway and induce CSC-like phenotypes formation to promote the proliferation and metastasis in MCF-7. Hence, CXCL12 may become a promising candidate for breast cancer therapy.     

HLJ1 is a novel biomarker for colorectal carcinoma progression and overall patient survival

The implication of HLJ1, a member of the heat shock protein-40 chaperone family, in colorectal carcinoma (CRC) remains unclear. The aim of this study was to determine the dynamic changes of HLJ1 in CRC both in vitro and in vivo, and the relationship between its level and the survival rate of CRC patients. Both real-time RT-PCR and Western blot were used to detect the expression of HLJ1 in CRC cells, while the distribution of HLJ1 in CRC and its adjacent normal mucosa tissues from CRC patients was determined with immunohistochemistry. Moreover, MTT and in vitro invasive assays were performed to determine the effect of HLJ1 overexpression on cell proliferation and invasion of CRC cells. The results indicated that in highly metastatic CRC cells, the HLJ1 expression was lower than that in lowly metastatic ones, and that the overexpression of HLJ1 significantly inhibited CRC cell proliferation and invasion in vitro. Interestingly, the HLJ1 expression was significantly down-regulated in CRC or lymphatic metastatic tissues from patient, compared to that in the normal mucosa (P<0.05), and the HLJ1 expression was correlated strongly with lymph metastasis, Dukes’ stage, and remote metastasis (P<0.05). Most surprisingly, patients with a higher HLJ1 level had a better overall survival rate, compared to that in patients with lower HLJ1 level (P<0.05). Based on all these findings, we conclude that HLJ1 is a strong tumor suppressor for CRC, and thus the down-regulation of the HLJ1 expression may be used as a biomarker to predict clinical outcome of patients with CRC.     

Relationship between osteopontin and β-catenin immunohistochemical expression and prognostic parameters of colorectal carcinoma

Previous studies on the prognostic value of osteopontin (OPN) and β-catenin in colorectal carcinoma (CRC) revealed conflicting results. To date, only two immunohistochemical studies investigated their association in CRC with discrepant results. Moreover, the relevance of their co-expression to clinicopathological parameters was not previously reported. This study was designed to investigate the relationship between these markers and prognostic parameters in CRC and study further the relationship between them. Immunohistochemical expression of OPN and β-catenin was evaluated in 72 CRCs. Cytoplasmic OPN was detected in 45.83% of CRCs while normal mucosa was immunonegative. Strong continuous membranous β-catenin was present in normal mucosa. However, abnormal membranous, nuclear and cytoplasmic expressions were observed in 36.11%, 31.94% and 52.78% of CRCs, respectively. A highly significant relationship was detected between each of OPN and nuclear β-catenin expression and lymph node metastasis (P = 0.0001 and 0.004 respectively), depth of invasion (P = 0.001 and 0.004 respectively), TNM stages (P = 0.0001 and 0.001 respectively) and Dukes’ stages (P = 0.0001 and 0.004 respectively). A significant association was found between OPN and distant metastases. A strong agreement was observed between OPN and nuclear β-catenin (kappa = 0.656). A highly significant relationship was found between their co-expression and poor prognostic parameters. OPN overexpression and nuclear β-catenin expression appeared to be associated with unfavorable prognostic factors in CRC. A direct relationship was observed between them. Further understanding their role in colorectal carcinogenesis as well as targeting the interaction between them might be effective in the future development of therapeutic agents for CRC patients.     

FGFR4 and TGF-β1 Expression in Hepatocellular Carcinoma: Correlation with Clinicopathological Features and Prognosis

Objective: To investigate the expression and correlation of transforming growth factor-β1 (TGF-β1) and fibroblast growth factor receptor 4 (FGFR4) in human hepatocellular carcinoma (HCC) and the relationship with clinicopathological features and prognosis.Materials and methods: The expression of TGF-β1 and FGFR4 in 126 HCC samples was detected immunohistochemically. Combined with clinical postoperative follow-up data, the expression of TGF-β1 and FGFR4 in HCC and the relationship with the prognosis of patients were analyzed by statistically.Results: The positive expression rate of TGF-β1 was 84.1% (106/126) in tumors, and that in peritumoral liver tissues was 64.3% (81/126); the positive expression rate of FGFR4 in tumors was 74.6% (94/126) and that in peritumoral liver tissues was 57.1% (72/126). The expression of TGF-β1 and FGFR4 in the carcinoma tissues was significantly higher than that in peritumoral liver tissues (p < 0.05). Intratumoral TGF-β1 and FGFR4 expression was associated with TNM stage (p < 0.05). TGF-β1 and FGFR4 expression levels didn''t significantly correlate with other clinicopathological parameters, including age, sex, tumor size, serum AFP level, tumor differentiation, lymph node metastasis, etc. (p > 0.05). TGF-β1 expression was positively correlated with FGFR4 expression (r = 0.595, p < 0.05). Patients with positive FGFR4 or TGF-β1 expression had shorter overall survival compared with negative expression (p < 0.05).Conclusions: The expression of TGF-β1 and FGFR4 could make synergy on the occurrence and progression of HCC, and may be used as prognosis indicators for HCC patients.     

Effects of hydrogen sulfide on high glucose-induced glomerular podocyte injury in mice

The aim of this study was to assess the effects of hydrogen sulfide on high glucose-induced mouse podocyte (MPC) injury and the underlying mechanisms. Mouse podocytes were randomly divided into 4 groups, including high glucose (HG), normal glucose (NG), normal glucose + DL-propargylglycine (PPG), and high glucose + NaHS (HG + NaHS) groups for treatment. Then, ZO-2, nephrin, β-catenin, and cystathionine γ-lyase (CSE) protein expression levels were determined by western blot. We found that high glucose significantly reduced nephrin, ZO-2, and CSE expression levels (P<0.05), and overtly elevated β-catenin amounts (P<0.05), in a time-dependent manner. Likewise, PPG at different concentrations in normal glucose resulted in significantly lower CSE, ZO-2, and nephrin levels (P<0.05), and increased β-catenin amounts (P<0.05). Interestingly, significantly increased ZO-2 and nephrin levels, and overtly reduced β-catenin amounts were observed in the HG + NaHS group compared with HG treated cells (P<0.01). Compared with NG treated cells, decreased ZO-2 and nephrin levels and higher β-catenin amounts were obtained in the HG + NaHS group. In conclusion,CSE downregulation contributes to hyperglycemia induced podocyte injury, which is alleviated by exogenous H₂S possibly through ZO-2 upregulation and the subsequent suppression of Wnt/β-catenin pathway.     

Nuclear localizaiton of β-catenin is associated with poor survival and chemo-/radioresistance in human cervical squamous cell cancer

Nuclear expression of β-catenin has been suggested as an independent prognostic marker in a variety of cancers. The objective of this study was to investigate the clinicopathologic significance of nuclear β-catenin expression in patients with cervical squamous cell carcinoma (CSCC). In this original research article, we detected nuclear β-catenin expression in 29/171 CSCC tissues (17.0%). Patients without nuclear β-catenin expression had a significantly better outcome than patients with nuclear β-catenin expression (93.7% versus 82.7% P = 0.027). Furthermore, nuclear β-catenin expression was predictive of prognosis in CSCC patients with early stage disease (FIGO stage I or tumor size ≤ 4 cm), with well/moderately differentiated tumors, or lymph node metastasis. Interestingly, nuclear β-catenin expression correlated with poor outcome in patients who received postoperative chemotherapy or radiotherapy. Multivariate analysis suggested that nuclear β-catenin expression is an independent prognostic indicator in CSCC. Our findings suggest that nuclear β-catenin expression may be used as a prognostic biomarker in CSCC, especially for patients with early stage disease, well/moderately differentiated tumors, or lymph node metastasis. Moreover, nuclear β-catenin expression has potential as a predictive marker of chemoresistance and radioresistance in CSCC.     

Embryonic stem cell markers Sox-2 and OCT4 expression and their correlation with WNT signal pathway in cervical squamous cell carcinoma

Background: Both the expression of embryonic stem cells (ESCs) markers (Sox2, Oct4) and the Wnt signal pathway (β-catenin) are crucial for progression of various human malignancies. The purpose of this study was to investigate the clinicopathologic significance of Sox2, Oct4 and β-catenin in cervical squamous cell carcinoma (CSCC) and to study their correlation with the occurrence and prognosis. Methods: Sox2, Oct4 and β-catenin were assessed using immunohistochemistry in normal cervix tissues (n = 28) and invasive cervical squamous cell carcinoma (n = 43). Associations of Sox2, Oct4 and β-catenin levels with clinicopathological characteristics and with overall survival were studied using uni- and multivariate analysis. Results: The expression levels of Sox2, Oct4 and β-catenin were highly increased in CSCC compared with the normal cervix tissues. The ESCs markers expression (Sox2 and Oct4) correlated significantly with β-catenin expression. High expression of Sox2, but not that of Oct4 or β-catenin, was correlated with poorer differentiation (P < 0.05). Furthermore, Sox2 expression was significantly correlated with patients’ status of survival in advanced CSCC (P < 0.05), whereas there was no significant finding in Oct4 or β-catenin expression. Conclusions: These findings provide evidence that both ESCs biomarkers (Sox2, Oct4) and Wnt signal pathway (β-catenin) are activated in CSCC. Sox2 can be regarded as a novel predictor of poor prognosis for CSCC patients.     

Elevated expression of UHRF1 predicts unfavorable prognosis for patients with hepatocellular carcinoma

Aims: The present study was designed to evaluate the different expression of ubiquitin-like with PHD and ring finger domains 1 (UHRF1) in hepatocellular carcinoma (HCC) tissues and the adjacent normal tissues, further explore the correlation between UHRF1 expression and the prognosis of HCC patients. Methods: The UHRF1 expression at protein level in HCC tissues and the adjacent normal tissues were measured by high performance liquid chromatography (HPLC). Chi-square test was used to estimate the relationship between UHRF1 expression and clinicopathologic characteristics of HCC patients. The overall survival of HCC patients with diverse expression of UHRF1 was measured by Kaplan-Meier analysis. Cox regression analysis was conducted to judge the prognostic value of UHRF1 in HCC patients. Results: The UHRF1 was over-expressed in HCC tissues compared with the adjacent normal tissues according to the outcome of HPLC (P<0.001). Besides, the UHRF1 expression was tightly related to distant metastasis, cancer area, and HBV (P<0.05), but shared no correlation with gender, cirrhosis, and bilirubin (P>0.05). Patients with high UHRF1 expression had a shorter overall survival time than those with low UHRF1 expression (P<0.001). Cox regression analysis showed that UHRF1 was significantly linked with the prognosis of HCC patients (P=0.002, HR=5.807, 95% CI=1.901-17.742). Conclusion: UHRF1 was over-expressed in HCC tissues compared to the adjacent normal tissues and UHRF1 expression shared significant relevance with distant metastasis, cancer area and HBV. It could be an important and independent prognostic biomarker for HCC patients.     

Expression of chemokine receptor CXCR7 in colorectal carcinoma and its prognostic significance

Previous studies have shown that chemokine receptor CXCR7 plays critical roles in tumor development. However, the clinicopathological and prognositic significance of CXCR7 in colorectal carcinoma (CRC) has not been fully understood. The aim of our study is to investigate the expression of CXCR7 and its clinical significance in CRC. First, quantitative RT-PCR and Western blot assays were performed to determine the expression of CXCR7 mRNA and protein in 20 paired of CRC tissues and corresponding adjacent non-tumor tissues. Next, immunohistochemistry was performed to detect the expression of CXCR7 protein in another 96 cases of CRC tissues, and analyze its correlation with clinicopathological factors of patients. Finally, the correlation of CXCR7 with 5-year overall survival (OS) and progression free survival (PFS) was statistically analyzed by the Kaplan-Meier method and Cox proportional hazards model. Results showed that the expression levels of CXCR7 mRNA and protein were significantly higher in CRC tissues than in normal tissues. Positive CXCR7 expression was observed to be significantly correlated with lymph nodal metastasis (P < 0.001), distant metastasis (P = 0.017), and advanced TNM stage (P < 0.001). Patients with positive expression of CXCR7 were demonstrated to be associated with worse OS and PFS (P < 0.001, P < 0.001, respectively). Moreover, multivariate survival analysis revealed that CXCR7 expression level might be an independent predictive factor for OS and PFS of CRC patients. Collectively, positive CXCR7 expression in CRC was correlated with tumor development and poor prognosis of patients.     

Prognostic correlation between MTA2 expression level and colorectal cancer

Objectives: Association of MTA2 expression with presence, development, metastasis and prognosis of colorectal cancer (CRC) was investigated. Methods: 90 CRC-related cases with follow-up information were made into tissue microarrays according to the paired principle of cancer tissues and the adjacent tissues. Subsequently, the expression of MAT2 was detected with immunohistochemical analysis and SPSS software was finally utilized to analyze the relationships between experimental data and clinical indicatives. Results: Expression of MTA2 in CRC tissues were notably higher than their adjacent tissues (P < 0.001) and showed significant positive correlation with tumor grade (r² > 0, P < 0.01). Moreover, survival analysis indicated that MTA2 expression in cancer tissues, serving as an independent correlation factor, was significantly correlated with poor prognosis (P = 0.004). Conclusions: MTA2 is a crucial biomarker that is closely related with prognosis of CRC and also a potential molecular target for evaluating the prognosis and treatment of CRC.     

Positive ALDH1A3 and Negative GPX3 Expressions Are Biomarkers for Poor Prognosis of Gallbladder Cancer

Background. Gallbladder cancers (GBCs) are highly aggressive cancers with high mortality. However, biological markers for the progression and prognosis of GBC are currently unavailable in the clinic. Objective. To identify biomarkers for predicting GBC metastasis and prognosis. Methods. We examined ALDH1A3 and GPX3 expressions in 46 squamous cell/adenosquamous carcinomas (SC/ASC) and 80 adenocarcinomas (AC) by using immunohistochemistry. Results. Positive ALDH1A3 and negative GPX3 expressions were significantly associated with lymph node metastasis and invasion of SC/ASCs and ACs. Univariate Kaplan-Meier analysis showed that either positive ALDH1A3 (P < 0.001) or negative GPX3 (P < 0.001) expression significantly correlated with decreased overall survival in both SC/ASC and AC patients. Multivariate Cox regression analysis showed that positive ALDH1A3 expression or negative GPX3 expression was an independent poor-prognostic predictor in both SC/ASC and AC patients. Conclusions. Our study suggested that positive ALDH1A3 and negative GPX3 expressions are closely associated with clinical pathological behaviors and poor prognosis of gallbladder cancer.     

Estrogen receptor β promoter methylation: a potential indicator of malignant changes in breast cancer

Introduction

Estrogen receptor β (ERβ) always lacks expression in estrogen-dependent tumors, which may result from gene inactivation by methylation. In this study, we aimed to determine whether aberrant methylation of the ERβ promoter is associated with decreased ERβ gene expression in breast cancer.

Material and methods

ERβ methylation status was determined for 132 pairs of breast cancer and adjacent normal tissues via the MethyLight method. Additionally, mRNA relative expression was quantified by real-time polymerase chain reaction (RT-PCR) to determine whether aberrant methylation had a negative correlation with expression. The correlation of ERβ promoter methylation and clinical parameters is also discussed.

Results

Methylation was observed in 96 (72.7%) breast cancer samples, and the median percentage of fully methylated reference (PMR) among methylated tissues was 0.83. Meanwhile, 94 (71.2%) adjacent normal tissues were methylated and the median PMR was 0.48. Compared to adjacent normal tissues, the methylation level of breast cancer was significantly higher (p < 0.001) and mRNA expression was much lower (p < 0.001). There was a significant correlation between ERβ methylation and mRNA expression in adjacent normal breast tissues (p = 0.004). In addition, the methylation rate of cancer tissues whose maximum diameter < 3 cm was significantly higher than those > 3 cm (p = 0.025).

Conclusions

ERβ promoter methylation level varies between cancerous and adjacent normal breast tissues. There was significant downregulation of ERβ methylation expression in pre-cancerous stages of breast cancer. Therefore, demethylation drugs may offer a potential strategy for preventing the development of pre-cancerous cells.     

Overexpression of β-catenin and cyclinD1 predicts a poor prognosis in ovarian serous carcinomas

Ovarian serous cancer is the most common subtype of epithelial ovarian cancer, and is the leading cause of death from gynecologic cancer. There is an important need for exploration of diagnostic and prognostic markers for this disease. β-catenin and cyclinD1 play central roles in the tumorigenesis for certain cancers. The role of β-catenin and cyclinD1 in diagnosis and prognosis of ovarian serous carcinoma is uncertain. In the present study, the expression of β-catenin and cyclinD1 was examined in 60 ovarian serous carcinomas patients with immunohistochemical staining. The relationship between expression of β-catenin and cyclinD1 and FIGO stage, pathological grade was analyzed. Kaplan-Meier survival function was used to analyze the prognosis. Overexpression of β-catenin is more often detected in patients with FIGO stage III and IV than in those with stage I, and II (P=0.003). No significant relationship was found between expression of β-catenin and pathological grade (P=0.817). Positive expression of β-catenin related to lower survival rate (P=0.034). The expression of cyclinD1 had no relationship with FIGO stage (P=0.829). Overexpression of cyclinD1 was positively to pathological grade (P=0.017) and survival rate (P=0.009). There is a significantly positive relationship between expression of β-catenin and cyclinD1 (P=0.014). No statistical significance was found between expression of β-catenin and cyclinD1 and other pathological parameters. Conclusions: Expression of β-catenin and cyclinD1 may be used as predict markers for poor prognosis.