CCDC12 gene methylation in peripheral blood as a potential biomarker for breast cancer detection.

BACKGROUND: Aberrant DNA methylation has been identified as biomarkers for breast cancer detection. Coiled-coil domain containing 12 gene (CCDC12) implicated in tumorigenesis. This study aims to investigate the potential of blood-based CCDC12 methylation for breast cancer detection. METHODS: DNA methylation level of CpG sites (Cytosine-phosphate Guanine dinucleotides) in CCDC12 gene was measured by mass spectrometry in 255 breast cancer patients, 155 patients with benign breast nodules and 302 healthy controls. The association between CCDC12 methylation and breast cancer risk was evaluated by logistic regression and receiver operating characteristic curve analysis. RESULTS: A total of eleven CpG sites were analyzed. The CCDC12 methylation levels were higher in breast cancer patients. Compared to the lowest tertile of methylation level in CpG_6,7, CpG_10 and CpG_11, the highest quartile was associated with 82, 91 and 95% increased breast cancer risk, respectively. The CCDC12 methylation levels were associated with estrogen receptor (ER) and human epidermal growth factor 2 (HER2) status. In ER-negative and HER2-positive (ER-/HER2+) breast cancer subtype, the combination of four sites CpG_2, CpG_5, CpG_6,7 and CpG_11 methylation levels could distinguish ER-/HER2+ breast cancer from the controls (AUC = 0.727). CONCLUSION: The hypermethylation levels of CCDC12 in peripheral blood could be used for breast cancer detection.

Breast cancer detection could be facilitated by novel blood-based DNA methylation biomarkers.The methylation levels of CpG sites in CCDC12 were higher in breast cancer than those in controls.The combination of four sites CpG_2, CpG_5, CpG_6,7 and CpG_11 methylation levels could distinguish ER-/HER2+ breast cancer subtype from the controls.

Blood FOLR3 methylation dysregulations and heterogeneity in non-small lung cancer highlight its strong associations with lung squamous carcinoma.

BACKGROUND: Non-small cell lung cancer (NSCLC) accounts for the vast majority of lung cancers. Early detection is crucial to reduce lung cancer-related mortality. Aberrant DNA methylation occurs early during carcinogenesis and can be detected in blood. It is essential to investigate the dysregulated blood methylation markers for early diagnosis of NSCLC. METHODS: NSCLC-associated methylation gene folate receptor gamma (FOLR3) was selected from an Illumina 850K array analysis of peripheral blood samples. Mass spectrometry was used for validation in two independent case-control studies (validation I: n = 2548; validation II: n = 3866). Patients with lung squamous carcinoma (LUSC) or lung adenocarcinoma (LUAD), normal controls (NCs) and benign pulmonary nodule (BPN) cases were included. FOLR3 methylations were compared among different populations. Their associations with NSCLC clinical features were investigated. Receiver operating characteristic analyses, Kruskal-Wallis test, Wilcoxon test, logistics regression analysis and nomogram analysis were performed. RESULTS: Two CpG sites (CpG_1 and CpG_2) of FOLR3 was significantly lower methylated in NSCLC patients than NCs in the discovery round. In the two validations, both LUSC and LUAD patients presented significant FOLR3 hypomethylations. LUSC patients were highlighted to have significantly lower methylation levels of CpG_1 and CpG_2 than BPN cases and LUAD patients. Both in the two validations, CpG_1 methylation and CpG_2 methylation could discriminate LUSC from NCs well, with areas under the curve (AUCs) of 0.818 and 0.832 in validation I, and 0.789 and 0.780 in validation II. They could also differentiate LUAD from NCs, but with lower efficiency. CpG_1 and CpG_2 methylations could also discriminate LUSC from BPNs well individually in the two validations. With the combined dataset of two validations, the independent associations of age, gender, and FOLR3 methylation with LUSC and LUAD risk were shown and the age-gender-CpG_1 signature could discriminate LUSC and LUAD from NCs and BPNs, with higher efficiency for LUSC. CONCLUSIONS: Blood-based FOLR3 hypomethylation was shown in LUSC and LUAD. FOLR3 methylation heterogeneity between LUSC and LUAD highlighted its stronger associations with LUSC. FOLR3 methylation and the age-gender-CpG_1 signature might be novel diagnostic markers for the early detection of NSCLC, especially for LUSC.

The association between CTSZ methylation in peripheral blood and breast cancer in Chinese women.

Purpose: Previous studies have shown that DNA methylation in peripheral blood may be associated with breast cancer (BC). To explore the association between the methylation level of the Cathepsin Z (CTSZ) gene in peripheral blood and BC, we conducted a case-control study in the Chinese population. Methods: Peripheral blood samples were collected from 567 BC cases, 635 healthy controls, and 303 benign breast disease (BBD) cases. DNA extraction and bisulfite-specific PCR amplification were performed for all samples. The methylation levels of seven sites of the CTSZ gene were quantitatively determined by Mass spectrometry. The odds ratios (ORs) of CpG sites were evaluated for BC risk using per 10% reduction and quartiles analyses by logistic regression. Results: Our analysis showed that five out of the seven CpG sites exhibited significant associations with hypomethylation of CTSZ and BC, compared to healthy controls. The highest OR was for Q2 of CTSZ_CpG_1 (OR: 1.62, P=0.006), particularly for early-stage breast cancer in the case of per 10% reduction of CTSZ_CpG_1 (OR: 1.20, P=0.003). We also found that per 10% reduction of CTSZ_CpG_5 (OR: 1.39, P=0.004) and CTSZ_CpG_7,8 (OR: 1.35, P=0.005) were associated with increased BC risk. Our study also revealed that four out of seven CpG sites were linked to increased BC risk in women under 50 years of age, compared to healthy controls. The highest OR was for per 10% reduction of CTSZ_CpG_1 (OR: 1.47, P<0.001). Additionally, we found that BC exhibited lower methylation levels than BBD at CTSZ_CpG_4 (OR for Q1: 2.18, P<0.001) and CTSZ_CpG_7,8 (OR for Q1: 2.01, P=0.001). Furthermore, we observed a correlation between methylation levels and tumor stage, ER, and HER2 status in BC patients. Conclusion: Overall, our findings suggest that altered CTSZ methylation levels in peripheral blood may be associated with breast cancer, particularly in young women, and may serve as a potential biomarker for early-stage BC.

Amphiphilic chitosan/carboxymethyl gellan gum composite films enriched with mustard essential oil for mango preservation.

In this paper, amphiphilic chitosan and carboxymethyl modified gellan gum were synthesized to develop an active edible fresh-keeping material. The optimal weight ratio of CMCS-g-CA/CMGG was determined as 5:2 through the characterization of Fourier transform infrared (FT-IR), Thermogravimetric analysis (TGA), mechanical and barrier properties of the composite films. In addition, the water vapor permeability and oxygen permeability of CMCS-g-CA/CMGG composite films incorporated with mustard essential oil were all declined, and the antibacterial property of the composite film solutions against E. coli, S. aureus and Bacillus anthracis was distinctly improved with the increase of mustard essential oil (MEO) dosage. Furthermore, the CMCS-g-CA/CMGG + 2.0 µL/mL MEO composite film exhibited an effective preservation on mango fruits during 20 days of storage based on the characterization of surface appearance and quality indexes of fruits. Hence, the multifunctional CMCA-g-CA/CMGG/MEO composite films can be served as a prospective eco-friendly packaging material for fruit preservation.

Novel blood-based FUT7 DNA methylation is associated with lung cancer: especially for lung squamous cell carcinoma.

BACKGROUND: The death rate of lung cancer (LC) ranks first in the world. Changes of DNA methylation in peripheral blood may be related to malignant tumors. It is necessary to explore blood-based biomarkers of methylation to detect LC. METHODS: Mass spectrometry assays were conducted to measure DNA methylation levels of seven CpG sites within FUT7 gene in the peripheral blood of 428 patients with LC, 233 patients with benign pulmonary nodule (BPN) and 862 normal controls (NC). The odds ratios (ORs) of all CpG sites were evaluated for their risk to LC using per SD change and tertiles analyses by logistic regression. The predictive ability of the seven FUT7 CpG sites and risk factors were evaluated by receiver operating characteristic curve (ROC). RESULTS: The methylation levels of seven CpG sites of FUT7 in LC were significantly lower than that in NC (P < 0.05). The per SD decrement of methylation level in CpG_1-7 was significantly associated with 65%, 38%, 59%, 46%, 23%, 20% and 68% higher risk for LC versus NC, respectively, and the adjusted ORs (95% CI) were 2.92 (2.17-3.96), 1.76 (1.29-2.38), 2.83 (2.09-3.82), 3.00 (2.17-4.16), 1.81 (1.35-2.43), 1.48 (1.11-1.97) and 3.04 (2.23-4.16) for the lowest tertiles of methylation level in CpG_1-7 compared with the top tertiles, respectively. The area under the curve (AUC) of FUT7_CpG_1-7 was 0.659 (CI 0.626-0.693), 0.792 (CI 0.736-0.848) and 0.729 (CI 0.665-0.792) in distinguishing LC versus NC, LUSC versus NC and LUSC versus BPN. CONCLUSIONS: Our study revealed an association between FUT7 hypomethylation and LC, especially for LUSC, which provides novel support for the blood-based methylation signatures as potential marker for assessing lung cancer risk.

Clinical value of MRI, serum SCCA, and CA125 levels in the diagnosis of lymph node metastasis and para-uterine infiltration in cervical cancer.

BACKGROUND: Cervical cancer shows great differences in depth of invasion, metastasis, and other biological behaviors. The location of the lesion is special, so it is usually difficult to determine the clinical stage. This study aimed to explore the clinical value of magnetic resonance imaging (MRI) and tumor serum markers for the preoperative diagnosis of cervical cancer lymph node metastasis and para-uterine invasion. METHODS: A total of 200 patients with cervical cancer admitted to our hospital from January 2019 to January 2020 were collected as the research subjects. Comparing the diagnosis results of preoperative MRI scan, serum tumor markers, and postoperative pathological examination using single factor comparison, we determined the MRI scan results, the comprehensive matching rate between serum tumor markers (squamous cell carcinoma antigen (SCCA), carbohydrate antigen 125 (CA125)) and postoperative pathological results, and the differences of sensitivity, specificity, and accuracy in the prediction of lymph node metastasis and para-uterine infiltration of cervical cancer. RESULTS: The levels of SCCA and CA125 in patients with para-uterine invasion and lymph node metastasis were higher than those of patients without invasion and metastasis. Among them, the level of SCCA was significantly different (P<0.05). The level of CA125 was not statistically significant (P>0.05), so MRI combined with serum SCCA was selected for combined diagnosis in the later period. The sensitivity, specificity, and accuracy of MRI diagnosis of cervical cancer and para-uterine infiltrating lymph node metastasis and metastasis were 55.2, 91.6, and 89.5% and 55.2, 91.6, and 89.5%, respectively. These data in MRI combined with serum SCCA were 76.3, 95.3, and 94.3% and 63.2, 96.0, and 95.1%, respectively. The accuracy of tumor markers combined with MRI in the diagnosis of cervical cancer lymph node metastasis and para-uterine invasion was higher than that of MRI. CONCLUSIONS: MRI combined with serum SCCA can more accurately identify cervical cancer lymph node metastasis and para-uterine invasion compared with MRI alone. Tumor marker combined with MRI diagnosis is an important auxiliary method for cervical cancer treatment and can provide comprehensive and reliable clinical evidence for evaluation before cervical cancer surgery.

Diagnostic value of multi-slice spiral CT (MSCT) combined with digestive tract angiography in gastric fundus cardia carcinoma.

PURPOSE: To evaluate the diagnostic value of multi-slice spiral CT (MSCT) combined with digestive tract angiography in patients with gastric fundus cardia carcinoma. METHODS: A total of 185 patients with suspected gastric fundus cardia carcinoma admitted in our hospital were collected. Among them, 93 patients were examined with MSCT combined with digestive tract angiography and were enrolled in the research group. Another 92 patients examined by MSCT alone comprised the control group. The diagnostic value of MSCT combined with digestive tract angiography in patients with gastric fundus cardia carcinoma was investigated. 185 patients were diagnosed by pathological examination and 166 had gastric fundus cardia carcinoma, with 84 patients in the research group, and 84 patients in the control group. Fifty nine patients with gastric fundus cardia carcinoma in the control group were diagnosed by MSCT. RESULTS: There were significant differences compared with pathological diagnosis (p<0.05). Eighty-two patients with gastric fundus cardia carcinoma in the research group were diagnosed by MSCT combined with digestive tract angiography. There were no significant differences compared with pathological diagnosis (p>0.05). Sensitivity, specificity and accuracy of the research group were significantly higher than those of the control group (p<0.05). The detectable rate in imaging results of the research group was higher than that of the control group (p<0.05). CONCLUSION: MSCT combined with digestive tract angiography is more accurate than single MSCT in the diagnosis of gastric fundus cardia carcinoma, which can effectively reduce the misdiagnosis and missed diagnosis and is worthy of clinical promotion.

lncRNA MAFG-AS1 Contributes to Esophageal Squamous-Cell Carcinoma Progression via Regulating miR143/LASP1.

BACKGROUND: Increasing investigations indicate that long noncoding RNA (lncRNA) is responsible for diverse biological functions during the progression of cancer. However, its functions and underlying mechanisms remain elusive. Here, we investigated the MAFG-AS1- expression profile in esophageal squamous-cell carcinoma (ESCC) patients and explored its biological function and potential molecular mechanisms. METHODS: qRT-PCR and the GEPIA data base were used to evaluate expression levels of MAFG-AS1 in ESCC tissue and cells. WST1-proliferation, -migration, and -invasion assays were performed to define the role of MAFG-AS1 in ESCC. Potential molecular mechanisms of MAFG-AS1 were investigated with online bioinformatic analysis, qRT-PCR, and rescue assays. RESULTS: MAFG-AS1 was upregulated in 45 ESCC-tissue samples and cell lines compared to that of adjacent nontumor tissue and normal esophageal cells. Higher MAFG-AS1 expressionindicated poor survival. Gain- and loss-of-function experiments suggested that MAFG-AS1 promoted ESCC-cell proliferation, migration, and invasion. Molecular mechanism analysis and rescue assay showed that miR143 inhibitors partly abolished the suppression of MAFG-AS1 knockdown on EC109-cells proliferation. Moreover, we found that LASP1 specifically targeted miR143. Collectively, these data indicated that MAFG-AS1 served as a ceRNA to elevate LASP1 levels by sponging miR143, and played an oncogenic role in ESCC. CONCLUSION: Our research findings demonstrate that MAFG-AS1 is a key regulator through a novel MAFG-AS1-miR143-LASP1 axis in ESCC development and progression, which may offer a potential therapeutic target for ESCC.

LncRNA MIR31HG functions as a ceRNA to regulate c-Met function by sponging miR-34a in esophageal squamous cell carcinoma.

Accumulating evidence has demonstrated that long non-coding RNAs (lncRNAs) function as essential regulators in the development and progression of multiple tumors. However, the molecular mechanisms of MIR31HG in regulating ESCC progression remain unknown. Here, we confirmed that MIR31HG facilitated ESCC cells proliferation in vivo. Besides, MIR31HG knockdown increases the percentage of cells at the G1 phase, along with reduced arrest in S phase and MIR31HG overexpression exhibits the opposite effects. Overexpressed MIR31HG decreases the percentage of apoptotic ESCC cells. Interestingly, MIR31HG can function as a competing endogenous RNA by sponging miR-34a. The rescue experiments demonstrated that MIR31HG function is partially reversed by inhibiting miR-34a. In addition, we found c-Met is a target gene of miR-34a and is indirectly regulated by MIR31HG. Taken together, our findings revealed that MIR31HG promotes ESCC progression by regulating miR-34a/ c-Met axis and may provide a new prospective for exploration and understanding of the biological effects of esophageal squamous cell carcinoma.

Exosomes Derived From MicroRNA-148b-3p-Overexpressing Human Umbilical Cord Mesenchymal Stem Cells Restrain Breast Cancer Progression.

Exosomes derived from human umbilical cord mesenchymal stem cells (HUCMSCs) expressing microRNAs (miRs) have been highlighted as important carriers for gene or drug therapy. Hence, this study aimed to explore the role of exosomal miR-148b-3p from HUCMSCs in breast cancer. Clinical samples subjected to RT-qPCR detection revealed that miR-148b-3p was poorly expressed, while tripartite motif 59 (TRIM59) was highly expressed in breast cancer tissues. Online analyses available at miRanda, TargetScan, and miRbase databases revealed that miR-148b-3p could bind to TRIM59, while dual-luciferase reporter gene assay further verified that TRIM59 was a target gene of miR-148b-3p. Next, miR-148b-3p mimic or inhibitor and siRNA against TRIM59 were delivered into the breast cancer cells (MDA-MB-231) to alter the expression of miR-148b-3p and TRIM59 so as to evaluate their respective effects on breast cancer cellular processes. Evidence was obtained demonstrating that miR-148b-3p inhibited cell proliferation, invasion, and migration, but promoted cell apoptosis in breast cancer by down-regulating TRIM59. Next, MDA-MB-231 cells were co-cultured with the exosomes derived from HUCMSCs expressing miR-148b-3p. The results of co-culture experiments demonstrated that HUCMSCs-derived exosomes carrying miR-148b-3p exerted inhibitory effects on MDA-MB-231 progression in vitro. In vivo experimentation further confirmed the anti-tumor effects of HUCMSCs-derived exosomes carrying miR-148b-3p. Taken together, HUCMSC-derived exosomes carrying miR-148b-3p might suppress breast cancer progression, which highlights the potential of exosomes containing miR-148b-3p as a promising therapeutic approach for breast cancer treatment.

Long Non-coding RNA ITIH4-AS1 Accelerates the Proliferation and Metastasis of Colorectal Cancer by Activating JAK/STAT3 Signaling.

Accumulating evidence has uncovered long non-coding RNAs (lncRNAs) as central regulators in the pathogenesis of diverse human cancers including colorectal cancer (CRC). The present study discovered that a novel lncRNA ITIH4 antisense RNA 1 (ITHI4-AS1) was frequently under-expressed in most normal human tissues, including colon tissues. Therefore, we aimed to investigate the role of ITHI4-AS1 in CRC. Interestingly, a significant overexpression of ITIH4-AS1 was observed in CRC cell lines relative to normal NCM460 cells. Also, we investigated the facilitating role of ITIH4-AS1 in CRC cell growth and metastasis both in vitro and in vivo. Additionally, we explained that ITIH4-AS1 upregulation in CRC was attributed to downregulation or even depletion of RE1 silencing transcription factor (REST), a presently identified transcriptional repressor for ITIH4-AS1. Meanwhile, the contribution of ITIH4-AS1 to CRC development was validated to rely on the activation of the JAK/STAT3 pathway. More importantly, we verified that FUS interacted with both ITIH4-AS1 and STAT3, and that ITIH4-AS1 evoked nuclear translocation of phosphorylated (p)-STAT3 in CRC through recruiting FUS. In summary, our findings unveiled for the first time that REST downregulation-enhanced ITIH4-AS1 exerts pro-tumor functions in CRC through FUS-dependent activation of the JAK/STAT3 pathway, implying that targeting ITIH4-AS1 may be a novel effective strategy for CRC therapy.

Long noncoding RNA FEZF1-AS1 promotes the motility of esophageal squamous cell carcinoma through Wnt/ß-catenin pathway.

Background: Long noncoding RNAs (lncRNAs), a class of noncoding RNA nucleotides >200 bp, has been demonstrated to play vital role in the development of cancer. FEZ family zinc finger 1 antisense RNA 1 (FEZF1-AS1) has been reported as an lncRNA which acts as a tumor-promoting effect in some cancers. However, the role of it in esophageal squamous cell carcinoma (ESCC) and its potential regulatory mechanism was unclear now. Methods: qRT-PCR was used to detect the levels of FEZF1-AS1 and mRNA CTNNB1 (ß-catenin) in ESCC tissues and cells. Cell transfection experiments were used to knock down or overexpress the level of FEZF1-AS1 in EC1 and EC9706 cell lines. WST-1 assays, cell cycle assays, scratch wound assays, migration, and invasion assays were used to evaluate the function of FEZF1-AS1 in ESCC progression. Results: FEZF1-AS1 was remarkably upregulated in ESCC tissues and cell lines. Silencing of FEZF1-AS1 significantly inhibited the migration and invasion of ESCC cells, while overexpression of FEZF1-AS1 notably accelerated ESCC migration and invasion. Meanwhile, the levels of FEZF1-AS1 had no effect on ESCC cell proliferation and cell cycle. We also found that ß-catenin was upregulated in ESCC tissues, and the level of it was positively correlated with the expression of FEZF1-AS1. Silencing of FEZF1-AS1 could decrease the mRNA and protein level of ß-catenin, while overexpression FEZF1-AS1 could lead to the contrary. Conclusion: Our results suggested that the expression of lncRNA FEZF1-AS1 played an important role in ESCC progression, especially the motility of the tumor. FEZF1-AS1 may provide us with a new sight for ESCC treatment.

LncRNA MNX1-AS1 promotes progression of esophageal squamous cell carcinoma by regulating miR-34a/SIRT1 axis.

BACKGROUND: Long non-coding RNAs (lncRNAs) are powerful factors influencing the tumorigenesis and metastasis of multiple carcinomas. LncRNA MNX1-AS1 plays critical roles in the progression of tumor formation according to recent research, while its roles in esophageal squamous cell carcinoma (ESCC) remains unknown. METHODS: The expression levels of lncRNA MNX1-AS1 were examined in ESCC tissues by quantitative reverse transcriptase polymerase chain reaction (qRT-PCR). The role of lncRNA MNX1-AS1 was performed by WST-1 proliferation assays, migration and invasion assays. Besides, the molecular mechanism of lncRNA MNX1-AS1 was verified by online bioinformatics, qRT-PCR and rescue assays. RESULTS: MNX1-AS1 was signifcantly upregulated in ESCC tissues. It was conformed that high MNX1-AS1 expression was associated with ESCC lymph node metastasis. Moreover, we found that knockdown of MNX1-AS1 apparently suppressed the cell proliferation, migration, and invasion capacity. Flow cytometry analysis showed MNX1-AS1 regulated ESCC cell cycle and apoptosis progression. Mechanism analysis revealed that miR-34a inhibitor could rescue the influence of inhibiting MNX1-AS1 on ESCC cells migration by serving as competing endogenous RNA (ceRNAs). Furthermore, we found that miR-34a specifically targeted SIRTI. CONCLUSIONS: Taken together, we demonstrated that lncRNA MNX1-AS1/miR-34a/SIRT1 regulatory axis could play an important role in ESCC progression, and MNX1-AS1 may act as a novel potential biomarker for esophageal squamous cell carcinoma.

Long non-coding RNA FTH1P3 regulated metastasis and invasion of esophageal squamous cell carcinoma through SP1/NF-kB pathway.

AIMS：: Recent research showed that Long non-protein coding RNA ferritin heavy chain 1 pseudogene 3 (FTH1P3) plays a crucial role in the course of tumor formation. The present study was aimed to explore its role in esophageal squamous cell carcinoma (ESCC). MAIN METHODS: Quantitative reverse transcriptase polymerase chain reaction (qRT-PCR) was used to examine the expression levels of FTH1P3, mRNA SP1 and NF-kB in ESCC samples and cell lines. The impact of FTH1P3 knockdown was evaluated by WST-1 assays, colony formation assays, scratch wound assays, migration and invasion assays. KEY FINDINGS: FTH1P3 was significantly upregulated in ESCC tissues and cells (P < 0.001). Knockdown of FTH1P3 notably decreased the proliferation, migration, and invasion capacity of ESCC cells. Silencing of FTH1P3 decreased the expression of specificity protein 1 (Sp1) and NF-kB (p65) in EC9706 and EC1. SIGNIFICANCE: FTH1P3 plays a crucial role in ESCC tumorigenesis, and can be used as a potential therapeutic target for ESCC.

YAP Inhibits the Apoptosis and Migration of Human Rectal Cancer Cells via Suppression of JNK-Drp1-Mitochondrial Fission-HtrA2/Omi Pathways.

BACKGROUND/AIMS: The Hippo-Yap pathway is associated with tumor development and progression. However, little evidence is available concerning its role in cancer cell apoptosis and migration via mitochondrial homeostasis. Here, we identify mitochondrial fission as a regulator of the Hippo-Yap pathway in human rectal cancer tumorigenesis and metastasis. METHODS: In this study, we performed loss-of function assays concerning Yap in RCC via shRNA. Cellular viability and apoptosis were measured via MTT, the TUNEL assay and trypan blue staining. Mitochondrial function was assessed via JC1 staining, the mPTP opening assay, mitochondrial respiratory function analysis, electron microscopy and immunofluorescence analysis of HtrA2/Omi. Mitophagy and mitochondrial fission were assessed via western blots and immunofluorescence. Cell migration was evaluated via the Transwell assay, wound-healing assay and immunofluorescence analysis of F-actin. The interaction between JNK and Yap was detected via co-immunoprecipitation and Yap recombinant mutagenic plasmid transfection. Western blots were used to analyze signaling pathways in conjunction with JNK inhibitors or HtrA2/Omi siRNA. RESULTS: Yap is upregulated in human rectal cancer cells, where its expression correlates positively with cell survival and migration. Functional studies established that silencing of Yap drove JNK phosphorylation, which induced Drp1 activation and translocation to the surface of mitochondria, initiating mitochondrial fission. Excessive mitochondrial fission mediated HtrA2/Omi leakage from the mitochondria into the cytoplasm, where HtrA2/Omi triggered cellular apoptosis via the mitochondrial apoptosis pathway. Moreover, released HtrA2/Omi also phosphorylated cofilin and inhibited cofilin-mediated F-actin polymerization. F-actin collapse perturbed lamellipodia formation and therefore impaired cellular migration and invasion. CONCLUSION: Collectively, our results demonstrate that Hippo-Yap can serve as a tumor promoter in human rectal cancer and acts by restricting JNK/Drp1/mitochondrial fission/ HtrA2/Omi, with potential implications for new approaches to human rectal cancer therapy.

MicroRNA-16-5p overexpression suppresses proliferation and invasion as well as triggers apoptosis by targeting VEGFA expression in breast carcinoma.

MicroRNAs (miRNAs), a class of small noncoding RNA molecules, can manipulate the expressions of endogenous tumor-related genes, and are implicated in the development and progression of a wide type of tumors. In this study, the investigation from real-time quantitative PCR revealed that miRNA-16-5p was downregulated in breast carcinoma tissues and cells, coupled with the elevations of HIF-α and VEGFA protein expressions, compared with normal tissues. Lentiviral armed with miR-16-5p markedly increased the miR-16-5p levels in MCF-7 and MDA-MB-231 cells, compared to blank and NC groups, and miR-16-5p overexpression significantly inhibited the proliferation and colony formation in MCF-7 and MDA-MB-231 cells. Besides, miR-16-5p upregulation markedly induced apoptosis and reduced invasion ability in MCF-7 and MDA-MB-231 cells. Notably, VEGFA was direct target of miR-16-5p. Stepwise investigation from in vitro and in vivo experiments demonstrated that miR-16-5p overexpression suppressed tumor growth and reduced HIF-α and VEGFA expressions in breast carcinoma cells and nude mice tumor tissues. These findings provide novel insights into molecular mechanism involved in the roles of miR-16-5p in tumor development and progression of breast carcinoma, and thus manipulation of miR-16-5p may be a novel potential therapeutic target for future therapies of the patients with breast carcinoma.

SIRT1 rs10823108 and FOXO1 rs17446614 responsible for genetic susceptibility to diabetic nephropathy.

SIRT1 and FOXO1 play an important role in the pathogenesis of diabetic nephropathy (DN). However, the association between genetic polymorphisms and susceptibility to type 2 DN (T2DN) has not been explored. In this study, a total of 1066 patients with type 2 diabetes mellitus (T2DM) (413 without and 653 with DN) were enrolled. The genotypes of three htSNPs (rs3818292, rs4746720, rs10823108) within SIRT1 and two htSNPs (rs2721068, rs17446614) in FOXO1 were determined by PCR-RFLP. HbA1C, LDL, HDL, TC, and TG levels were also examined. SIRT1 rs10823108 AA genotype was significantly associated with a decreased risk of DN (OR = 0.60, 95%CI: 0.38-0.97), while GA genotype (OR = 1.77, 95%CI: 1.33-2.35) and AA genotype (OR = 2.32, 95%CI: 1.25-4.34) of FOXO1 rs17446614 was associated with an increased T2DN risk. The interactions among rs1744 6614, BMI and duration of diabetes (OR: 2.63, 95%CI: 1.23-4.31) were also observed. Subsequent haplotype analysis revealed that two haplotype defined by AC (OR: 1.50, 95%CI: 1.15-1.94) and AT (OR: 1.79, 95%CI: 1.06-2.80) within FOXO1 gene may increase the risk of T2DN. In conclusion, genetic variant rs10823108 in SIRT1 and variant rs17446614 in FoxO1 may contribute to the risk of DN in T2DM patients.

HDAC2 regulates cell proliferation, cell cycle progression and cell apoptosis in esophageal squamous cell carcinoma EC9706 cells.

Increasing evidence has demonstrated that histone deacetylase 2 (HDAC2) participates in the regulation of a variety of biological processes in numerous tumors. However, the potential role of HDAC2 in the development and progression of esophageal squamous cell carcinoma (ESCC) remains elusive. Immunohistochemistry was utilized to detect the expression of HDAC2, Cell Counting Kit-8 was used to determine the cell proliferation, and flow cytometry was employed to investigate cell cycle and cell apoptosis. Finally, western blotting was employed to detect the protein expression of cyclin D1, p21, B cell lymphoma-2 (Bcl-2) and Bcl-2-associated X protein (Bax). The present study found that expression of HDAC2 protein in ESCC tissues was significantly increased compared with atypical hyperplasia tissues and normal esophageal mucosa (P<0.001). The expression of HDAC2 was not associated with the age or gender of patients (P>0.05), but was closely associated with the histological grade, invasion depth, tumor-node-metastasis stage and lymph node metastasis, respectively (all P<0.001). HDAC2 small interfering RNA effectively downregulated the expression of HDAC2 protein in ESCC EC9706 cells. Downregulation of HDAC2 expression evidently inhibited cell proliferation, arrested cell cycle at the G0/G1 phase and induced cell apoptosis in ESCC EC9706 cells, coupled with increased expression of p21 and Bax proteins and decreased expression of cyclin D1 and Bcl-2 proteins. Overall, the present findings suggest that HDAC2 may play an important role in the development and progression of ESCC and be considered as a novel molecular target for the treatment of ESCC.

Expression of circulating microRNA-20a and let-7a in esophageal squamous cell carcinoma.

AIM: To investigate the expressions of microRNA-20a (miR-20a) and let-7a in esophageal squamous cell carcinoma (ESCC) and their diagnostic value. METHODS: Seventy patients with ESCC and 40 healthy subjects were enrolled to investigate the expression of miR-20a and let-7a using quantitative real-time PCR. The expression of miR-20a and let-7a was compared between ESCC patients and healthy subjects. The plasma levels of miR-20a and let-7a in relation to patient clinicopathologic parameters, the receiver operating characteristic (ROC) curve, and the sensitivity and specificity of miR-20a and let-7a in ESCC diagnosis were analyzed. RESULTS: Plasma levels of miR-20a were significantly higher in ESCC patients than in healthy controls, and plasma levels of let-7 were lower in ESCC patients than in healthy controls (both P < 0.05). The area under the ROC curve of miR-20a was 0.767 (95%CI: 0.677-0.857; P < 0.001), when the cut-off value was set at 4.77, the sensitivity and specificity were 64.3% and 75.0%, respectively. The area under the ROC curve of let-7a was 0.829 (95%CI: 0.754-0.904; P < 0.001), when the cut-off value was set at 6.22, the sensitivity and specificity were 74.3% and 85.0%, respectively. Thus, the sensitivity and specificity of let-7a were higher than those of miR-20a. The median relative plasma expression of let-7a in clinical stage III/IV (0.24) was lower than that in stage I/II (0.42), while the expression of miR-20a according to stage was not statistically different. The expressions of miR-20a and let-7a were not related to gender, age, tumor diameter, tumor grade, or pathologic stage. CONCLUSION: Plasma miR-20a and let-7a levels are significantly altered in patients with ESCC and can be used as potential biomarkers in the diagnosis of ESCC.

[Expressions of Dab2 and ERK1 in esophageal carcinoma tissues and their clinical significance].

OBJECTIVE: To explore the expressions of protein and mRNA of disabled-2 (Dab2) and extracellular signal-regulated kinase1 (ERK1) in esophageal squamous cell carcinoma tissue (ESCC) and their clinical significance. METHODS: The expressions of protein and mRNA of Dab2 and ERK1 were detected in ESCC (n = 59), atypical hyperplasia (n = 27) and normal esophageal mucosa (n = 36) tissues by immunohistochemistry, in situ hybridization and reverse transcription polymerase chain reaction (RT-PCR). RESULTS: The expressions of protein and mRNA of Dab2 were lesser in ESCC than those in atypical hyperplasia and normal esophageal mucosa tissues (23.7% vs 94.4%, 44.4% and 18.6% vs 97.2%, 33.3%). And the expressions of protein and mRNA ERK1 were higher in ESCC than those in atypical hyperplasia and normal esophageal mucosa tissues (81.4% vs 44.4%, 63.0% and 78.0% vs 30.6%, 51.9%) (all P < 0.05). Additionally, the expressions of protein and mRNA of Dab2 and ERK1 were closely related with lymph node metastasis in ESCC tissue and the expressions of protein and mRNA of ERK1 closely related with the depth of tumor invasion in ESCC tissue (all P < 0.05). Furthermore, the results of correlation analysis revealed that expressions of protein and mRNA of Dab2 and ERK1 in ESCC had a negative correlation (all P < 0.05). CONCLUSION: The expressions of protein and mRNA of Dab2 and ERK1 play important roles in the infiltration, metastasis and carcinogenesis of ESCC and their effects may be antagonistic to each other.