Vitamin D receptor induces oxidative stress to promote esophageal squamous cell carcinoma proliferation via the p53 signaling pathway.

Background: Esophageal squamous cell carcinoma (ESCC) is a common pathological esophageal cancer with poor prognosis. Vitamin D deficiency reportedly occurs in ESCC patients, and this is related to single nucleotide polymorphism of vitamin D receptor (VDR). Objective: We investigated the effect of VDR on ESCC proliferation, invasion, and metastasis and its potential mechanism. Methods: ESCC and normal tissues were collected from 20 ESCC patients. The ESCC tissue microarray contained 116 pairs of ESCC and normal tissues and 73 single ESCC tissues. VDR expression and its clinicopathological role were determined by real-time quantitative polymerase chain reaction, Western blot, and immunohistochemistry staining. sh-VDR and VDR overexpression were used to validate the effect of VDR on ESCC cell phenotype, and tandem mass tag-based quantitative proteomics and bioinformatics methods identified differential VDR-related proteins. The downstream pathway and regulatory effect were analyzed using ingenuity pathway analysis (IPA). Differentially expressed proteins were verified through parallel reaction monitoring and Western blot. In vivo imaging visualized subcutaneous tumor growth following tail vein injection of VDR-deficient ESCC cells. Results: High VDR expression was observed in ESCC tissues and cells. Gender, T stage, and TNM stage were related to VDR expression, which was the independent prognostic factor related to ESCC. VDR downregulation repressed ESCC cell proliferation, invasion, and migration in vitro and subcutaneous tumor growth and lung metastases in vivo. The cell phenotype changes were reversed upon VDR upregulation, and differential proteins were mainly enriched in the p53 signaling pathway. TP53 cooperated with ABCG2, APOE, FTH1, GCLM, GPX1, HMOX1, JUN, PRDX5, and SOD2 and may activate apoptosis and inhibit oxidative stress, cell metastasis, and proliferation. TP53 was upregulated after VDR knockdown, and TP53 downregulation reversed VDR knockdown-induced cell phenotype changes. Conclusions: VDR may inhibit p53 signaling pathway activation and induce ESCC proliferation, invasion, and metastasis by activating oxidative stress.

Role of a novel circRNA-CGNL1 in regulating pancreatic cancer progression via NUDT4-HDAC4-RUNX2-GAMT-mediated apoptosis.

BACKGROUND: Pancreatic cancer (PC) is an extremely malignant tumor with low survival rate. Effective biomarkers and therapeutic targets for PC are lacking. The roles of circular RNAs (circRNAs) in cancers have been explored in various studies, however more work is needed to understand the functional roles of specific circRNAs. In this study, we explore the specific role and mechanism of circ_0035435 (termed circCGNL1) in PC. METHODS: qRT-PCR analysis was performed to detect circCGNL1 expression, indicating circCGNL1 had low expression in PC cells and tissues. The function of circCGNL1 in PC progression was examined both in vitro and in vivo. circCGNL1-interacting proteins were identified by performing RNA pulldown, co-immunoprecipitation, GST-pulldown, and dual-luciferase reporter assays. RESULTS: Overexpressing circCGNL1 inhibited PC proliferation via promoting apoptosis. CircCGNL1 interacted with phosphatase nudix hydrolase 4 (NUDT4) to promote histone deacetylase 4 (HDAC4) dephosphorylation and subsequent HDAC4 nuclear translocation. Intranuclear HDAC4 mediated RUNX Family Transcription Factor 2 (RUNX2) deacetylation and thereby accelerating RUNX2 degradation. The transcription factor, RUNX2, inhibited guanidinoacetate N-methyltransferase (GAMT) expression. GAMT was further verified to induce PC cell apoptosis via AMPK-AKT-Bad signaling pathway. CONCLUSIONS: We discovered that circCGNL1 can interact with NUDT4 to enhance NUDT4-dependent HDAC4 dephosphorylation, subsequently activating HDAC4-RUNX2-GAMT-mediated apoptosis to suppress PC cell growth. These findings suggest new therapeutic targets for PC.

Molecular epidemiological characteristics of carbapenem-resistant Pseudomonas aeruginosa clinical isolates in southeast Shanxi, China.

OBJECTIVES: Infection by carbapenem-resistant Pseudomonas aeruginosa (CRPA) is a serious clinical problem worldwide. However, the molecular epidemiology of the clinical isolates varies depending on the region. This study was conducted to analyse the resistance phenotype and clarify the genetic and epidemiological properties of CRPA clinical isolates from southeast Shanxi, China. METHODS: Fifty-seven isolates of CRPA were collected from a hospital in this region. These isolates were reidentified by MALDI-TOF and subjected to whole-genome sequencing by next-generation sequencing. Phylogenetic trees were constructed based on single nucleotide polymorphisms (SNPs), after which multilocus sequence typing (MLST) was performed and antimicrobial resistance genes were identified. RESULTS: All the 57 CRPA isolates carried at least one kind of gene encoding carbapenemase, such as blaIMP-1, blaIMP-10, blaOXA-10, blaOXA-395, blaOXA-396, blaOXA-485, blaOXA-486, blaOXA-488, blaOXA-494, and blaOXA-50. The isolates harboured AIM-1, CMY-51, mecD, and NmcR genes and carried one kind of Pseudomonas-derived cephalosporinase (PDC) ß-lactamase-encoding gene, such as blaPCD-1 to blaPCD-3, blaPCD-5, or blaPCD-7 to blaPCD-10. Two isolates were found to harbour the aminoglycoside-modifying enzyme genes aadA1 and aadA7; however, no isolates were found to harbour genes encoding 16S rRNA methylase or quinolone resistance-related genes. These CRPA isolates belonged to various sequence types (STs), two of which, namely, ST235 and ST277, were high-risk types. CONCLUSIONS: Our findings indicate that CRPA isolates carrying resistance genes with unique regional characteristics are spreading in this region, with a high diversity of STs, especially in high-risk clones. These findings highlight the necessity for further measures to prevent CRPA spread in Shanxi.

Raman spectroscopy and machine learning for the classification of esophageal squamous carcinoma.

Early diagnosis of esophageal squamous cell carcinoma (ESCC), a common malignant tumor with a low overall survival rate due to metastasis and recurrence, is critical for effective treatment and improved prognosis. Raman spectroscopy, an advanced detection technology for esophageal cancer, was developed to improve diagnosis sensitivity, specificity, and accuracy. This study proposed a novel, effective, and noninvasive Raman spectroscopy technique to differentiate and classify ESCC cell lines. Seven ESCC cell lines and tissues of an ESCC patient with staging of T3N1M0 and T3N2M0 at low and high differentiation levels were investigated through Raman spectroscopy. Raman spectral data analysis was performed with four machine learning algorithms, namely principal components analysis (PCA)- linear discriminant analysis (LDA), PCA-eXtreme gradient boosting (XGB), PCA- support vector machine (SVM), and PCA- (LDA, XGB, SVM)-stacked Gradient Boosting Machine (GBM). Four machine learning algorithms were able to classifiy ESCC cell subtypes from normal esophageal cells. The PCA-XGB model achieved an overall predictive accuracy of 85% for classifying ESCC and adjacent tissues. Moreover, an overall predictive accuracy of 90.3% was achieved in distinguishing low differentiation and high differentiation ESCC tissues with the same stage when PCA-LDA, XGM, and SVM models were combined. This study illustrated the Raman spectral traits of ESCC cell lines and esophageal tissues related to clinical pathological diagnosis. Future studies should investigate the role of Raman spectral features in ESCC pathogenesis.

Correlation between Vaginal Microecological Status and Prognosis of CIN Patients with High-Risk HPV Infection.

Many microorganisms live in the vagina of healthy women. They interact with and compete with the microenvironment in the female vagina to form a dynamic balance of the microenvironment in the female vagina. However, imbalanced vaginal microecology can lead to vaginal resistance to pathogenic microorganisms. Poor capacity can cause women to develop infections of the reproductive tract. This article analyzes the vaginal microecological status of women with high-risk HPV infection for more than 6 months and healthy women and explores the risk factors that cause long-term high-risk HPV infection for timely detection and regulation of possible vaginal microecological imbalance in women with high-risk HPV infection for more than 6 months to prevent further development of cervical lesions in such patients. This article covers women with a sexual life history who attended the gynecology department of a hospital from January 2020 to September 2021. There were 280 patients in the experimental group: positive high-risk HPV; and there were 140 patients in the control group: negative high-risk HPV test. The correlation between vaginal microecology of CIN patients and patient prognosis according to the subject's vaginal microecology test results and prognosis of various levels of cervical lesions was analyzed. The experiment proved that the detection rate of normal vaginal microecology in the experimental group was 12.14% (34/280) compared with the detection rate of 29.29% (41/140) in the control group, and there was a trend of decrease, and the difference was statistically significant (χ 2 = 17.23, P < 0.05). The detection rate of vaginal BV in the experimental group was 10.36% (29/280) compared with the detection rate of 5.0% (7/140) in the control group, and the difference was statistically significant (χ 2 = 5.19, P < 0.05). This indicates that women with high-risk HPV infections for 6 months or longer have a higher incidence of vaginal microecological imbalances than healthy individuals and aggressive vaginal microecological screening. It is necessary to carry out the program. Detect and treat possible abnormal conditions in time to prevent the further onset of the disease.

LncRNA SLC16A1-AS1 Suppresses Cell Proliferation in Cervical Squamous Cell Carcinoma (CSCC) Through the miR-194/SOCS2 Axis.

BACKGROUND: SLC16A1-AS1 has been characterized as an oncogenic long non-coding (lncRNA) in breast cancer and bladder cancer, while its role in cervical squamous cell carcinoma (CSCC) is unknown. METHODS: CSCC and non-tumor tissue samples were collected from 60 female patients, and qPCR was performed to detect the expression of SLC16A1-AS1, miR-194 and SOCS2. Luciferase reporter assay was performed to detect the interaction between SLC16A1-AS1 and miR-194. Colony formation assay was used to detect cell proliferation. RESULTS: SLC16A1-AS1 was down-regulated in CSCC and correlated with poor survival. Overexpression of SLC16A1-AS1 could inhibit the proliferation of cervical cancer cells. In addition, SLC16A1-AS1 could sponge miR-194 and increase the expression levels of SOCS2, ultimately inhibiting the proliferation of cervical cancer cells. CONCLUSION: SLC16A1-AS1 was downregulated in CSCC and suppressed cell proliferation in cervical squamous cell carcinoma (CSCC) through the miR-194/SOCS2 axis.

Clinical and prognostic effects of adjuvant therapy on less advanced esophageal squamous cell carcinoma patients.

BACKGROUND: This study aimed to clarify whether adjuvant therapy is suitable for less advanced esophageal squamous cell carcinoma (ESCC) patients postoperatively. METHODS: Data from 973 patients were collected. The prognosis and clinicopathological traits of these patients were calculated in both the TNM I and IIA stages. Meanwhile, 2 separate nomograms were applied in terms of the variables with a multivariate P value <0.05 in the Cox proportional hazard regression model. RESULTS: There were 471 and 502 patients in the I and IIA stage respectively; among all enrolled patients, 641, 130, 73, and 129 patients were in the no-treatment, chemotherapy, radiotherapy, and chemoradiotherapy groups, respectively. Adjuvant therapy was drawn as the independent prognostic factors for stage I (P=0.026; HR =1.081, 95% CI: 1.093, 1.308) and IIA patients (P<0.001; HR =0.788, 95% CI: 0.693, 0.896). Radiotherapy consistently obtained the best prognosis for patients when compared with the other 3 groups in the I and IIA stage. Patients in I stage had the worst prognosis after receiving chemoradiotherapy, and still, the small survival benefits of chemoradiotherapy were seen in patients of IIA stage. Two separate nomograms for the I and IIA stage were constructed. The C-index was 0.665 (95% CI: 0.569, 0.761) in the I stage and 0.645 (95% CI: 0.567, 0.723) in the IIA stage. Meanwhile, the calibration curves for predicting 3-year and 5-year survival for I and IIA patients agreed well with the actual observations. CONCLUSIONS: Among less advanced ESCC patients, adjuvant therapy was not only found to be an independent factor but also proved to be of importance in patient prognosis, and radiotherapy is recommended.

Clinical and prognostic significance of preoperative lymphocyte-monocyte ratio, neutrophil-lymphocyte ratio and neutrophil-monocyte ratio on esophageal squamous cell carcinoma patients.

BACKGROUND: The interaction between tumor cells and inflammatory cells has not been systematically investigated in esophageal squamous cell carcinoma (ESCC). The aim of the present study was to evaluate whether preoperative lymphocyte-monocyte ratio (LMR), neutrophil-lymphocyte ratio (NLR), and neutrophil-monocyte ratio (NMR) could predict the prognosis of ESCC patients undergoing esophagectomy. METHODS: A total of 1,883 patients with histologically diagnosed ESCC who underwent radical esophagectomy from May 2005 to May 2015 were retrospectively reviewed. Besides clinicopathological factors, "Survminer" package in R® was applied to determine the optimal cut-off point for LMR, NLR and NMR. Meanwhile, we evaluated the prognostic value of LMR, NLR, and PLR using Kaplan-Meier curves and Cox regression models. RESULTS: The median follow-up was 28.77 months (range, 1.60-247.90 months). The optimal cut-off point of LMR, NLR and NMR is 3.83, 2.06 and 7.21, respectively. Kaplan-Meier survival analysis of patients with low preoperative LMR demonstrated a significant worse prognosis for 5-year OS (P<0.001) than those with high preoperative LMR. The high NLR cohort had lower 5-year OS (P<0.001). No significant difference with 5-year OS was found in NMR (P=0.405). On multivariate analysis, preoperative LMR (P=0.018; HR =0.786, 95% CI: 0.645, 0.959) and NLR (P=0.028; HR =1.247, 95% CI: 1.024, 1.519) were the independent prognostic factors in ESCC patients. Integrating LMR and NLR, we divided the ESCC patients in four groups according to their cut-off points and we found the patients in LMR ≥3.83 and NLR <2.06 group received the best prognosis while the prognosis of patients in LMR<3.83 and NLR ≥2.06 group was the worst. The difference was statistically significant. CONCLUSIONS: Preoperative LMR and NLR better predicts cancer survival in patients with ESCC undergoing esophagectomy, especially under the circumstances of LMR ≥3.83 and NLR <2.06.

Prognostic significance and role of thoracic lymph node metastasis based on Chinese expert consensus in esophageal cancer.

BACKGROUND: The Chinese expert consensus on thoracic lymph node (LN) dissection in radical esophagectomy (Chinese Criteria, 2017 edition) was newly promoted. This study examined the prognostic significance and role of thoracic LN metastasis based on the Chinese Criteria for esophageal cancer. METHODS: Data of patients with thoracic esophageal squamous cell carcinoma (ESCC) who underwent curative esophagectomy in the West China Hospital from May 2005 to May 2015 were retrospectively analyzed. Patients' prognosis and clinicopathological features were compared to determine the role of Chinese Criteria and their relationship with Union for International Cancer Control (UICC)/American Joint Committee on Cancer (AJCC) 8th TNM staging. RESULTS: Overall, 2,285 qualified patients were divided into the no (n=1,148), skip (n=156), local (n=665), and mediastinal (n=316) metastasis groups according to the Chinese Criteria. Significant prognostic differences occurred among the four groups in all the thoracic and lower mediastinal ESCC patients (both P<0.001). The Chinese Criteria grouping was an independent prognostic factor for all thoracic [P<0.001; hazard ratio (HR) =1.261, 95% confidence interval (CI): 1.103-1.441], upper (P<0.001; HR =1.391, 95% CI: 1.264-1.530), lower mediastinal thoracic ESCC patients (P<0.001; HR =1.312, 95% CI: 1.257-1.370) and all thoracic ESCC after adjuvant therapy (P<0.001; HR =1.303, 95% CI: 1.221-1.390). Significant prognostic differences among Chinese Criteria groups occurred with N1 (P=0.014) and N2 (P=0.018) stages only. Significant differences in survival among N stages were found in local (P<0.001) and mediastinal (P=0.009) metastasis groups. CONCLUSIONS: Our study was the first to report the Chinese Criteria in measuring the degree of thoracic LN metastasis. Similar to N-stage, the Chinese Criteria were confirmed as an independent prognostic factor for thoracic ESCC. Further confirmation of our findings is warranted.

Vitamin D Signaling Pathways Confer the Susceptibility of Esophageal Squamous Cell Carcinoma in a Northern Chinese Population.

Experimental studies have determined the chemopreventive effects of vitamin D against the esophageal squamous cell carcinoma (ESCC); however, results from the epidemiological studies are not yet well established. The current study aimed to evaluate the associations between plasma vitamin D levels and variants on vitamin D metabolic-related genes with the risks for ESCC. A hospital-based case-control study was performed. Five hundred eighty-two ESCC patients and 569 controls were recruited in a Northern Chinese population. Common variants on vitamin D metabolism-related genes CYP24A1, DHCR7, GC, CYP27B1, and vitamin D receptor (VDR) and the plasma 25(OH)D level were determined. The unconditional logistic regression method was applied to determine the associations between the variants and vitamin D level and ESCC. Higher plasma 25(OH)D was associated with a reduced risk for ESCC, especially for rs2296241, rs11568820, and rs4646536. The variants rs2296241 on CYP24A1 and rs11568820 on VDR are significantly associated with ESCC cancer. Vitamin D signaling pathways may participate in the ESCC development. Further studies with larger sample size are warranted to confirm the results. Intervention studies are needed to determine whether vitamin D supplementation may reduce the ESCC risk in the Chinese population.

miR-502 inhibits cell proliferation and tumor growth in hepatocellular carcinoma through suppressing phosphoinositide 3-kinase catalytic subunit gamma.

MicroRNAs (miRNAs) play a key role in carcinogenesis and tumor progression in hepatocellular carcinoma (HCC). In the present study, we demonstrated that miR-502 significantly inhibits HCC cell proliferation in vitro and tumor growth in vivo. G1/S cell cycle arrest and apoptosis of HCC cells were induced by miR-502. Phosphoinositide 3-kinase catalytic subunit gamma (PIK3CG) was identified as a direct downstream target of miR-502 in HCC cells. Notably, overexpression of PIK3CG reversed the inhibitory effects of miR-502 in HCC cells. Our findings suggest that miR-502 functions as a tumor suppressor in HCC via inhibition of PI3KCG, supporting its utility as a promising therapeutic gene target for this tumor type.

[Identification of a novel mutation of COL2A1 gene in a Chinese family affected with spondyloepiphyseal dysplasia congenita].

OBJECTIVE: To identify potential disease-causing mutation in the COL2A1 gene in a Chinese family affected with autosomal dominant spondyloepiphyseal dysplasia congenita (SEDC; OMIM 183900) and to analyze the phenotype-genotype correlation. METHODS: Complete physical, and radiographic examinations of 4 affected individuals from the family were conducted. Genomic DNA was isolated from peripheral blood leukocytes. Whole-exome sequencing was performed using a HiSeq2000 sequencer. All 54 exons and exon-intron boundaries of the COL2A1 gene were amplified by polymerase chain reaction (PCR) and bidirectionally sequenced. RESULTS: All of the 4 individuals were found to carry a novel missense mutation of c.2224G>A (p.Gly687Ser) in the COL2A1 gene, while the same mutation was not found in the normal members of the family and 50 healthy controls. Protein prediction of missense mutation by Polyphen-2 and SIFT software indicated severe damage to the function. CONCLUSION: The mutation c.2224G>A (p.Gly687Ser) of the COL2A1 gene is responsible for this family. There are heterozygous of phenotype for the mutation.

Effects of bone marrow-derived mesenchymal stem cells transplanted via the portal vein or tail vein on liver injury in rats with liver cirrhosis.

The aim of the present study was to compare the effects of bone marrow-derived mesenchymal stem cells (BMSCs) transplanted via the portal vein or tail vein on liver injury in rats with liver cirrhosis. BMSCs were isolated from rat bone marrow and labeled with green fluorescent protein (GFP). Then, the labeled BMSCs were injected into rats with liver injury via the portal vein or tail vein. Two weeks after transplantation, three rats in each group were sacrificed to test the distribution of GFP in the liver and the serum levels of alanine aminotransferase (ALT), aspartate aminotransferase (AST) and albumin. Six weeks later, the remaining rats were sacrificed, and serum ALT, AST, albumin, hyaluronic acid (HA), laminin (LN) and procollagen type III (PC-III) levels were measured. The expression of albumin in the liver was analyzed by immunohistochemistry. Two weeks after BMSC transplantation, GFP-positive cells were detected in the livers of rats with BMSCs transplanted via the portal vein and tail vein. Compared with pre-transplantation levels, the ALT levels of the groups with BMSC transplantation via the portal vein and tail vein were significantly decreased after two and six weeks of BMSC transplantation (P<0.05), whereas the AST and albumin levels were not significantly different at two weeks after BMSC transplantation in the two groups (all P>0.05). However, the AST and albumin levels were significantly reduced at six weeks after BMSC transplantation (all P<0.05). At six weeks after BMSC transplantation, the serum HA, LN and PC-III levels in rats transplanted with BMSCs via the portal vein or tail vein had decreased significantly (all P<0.05), as compared with the levels prior to BMSC transplantation. BMSCs transplanted via the portal vein and tail vein achieved similar improvements in liver function in rats with liver cirrhosis, which suggests that peripheral venous administration is a convenient and effective route for BMSC transplantation.

The functional SNP rs3746804 in C20orf54 modifies susceptibility to esophageal squamous cell carcinoma.

OBJECTIVE: The aim of the present study was to explore the association of the functional single-nucleotide polymorphism (SNP) rs3746804 in C20orf54 with the susceptibility to esophageal squamous cell carcinoma (ESCC). METHODS: SNP rs3746804 in C20orf54 was detected by direct sequencing in 434 ESCC patients and 554 healthy controls from Shanxi and Henan Provinces in China, geographically a high incidence area for ESCC. The frequencies and distribution of TT, CT, and CC genotypes of SNP rs3746804 between those 2 cohorts were compared and its association with ESCC was assessed. RESULTS: For SNP rs3746804 the genotype distribution of CT and CC in ESCC patients both significantly differed from those in healthy controls (p=0.002, 0.001, respectively), while the distribution of TT did not differ significantly between the groups (p=0.757). The ratio of T:C was high in healthy individuals and low in ESCC patients. Compared to genotype CC, both genotypes CT (odds ratio (OR)=0.63, 95% confidence interval (CI) 0.48l-0.826), and CT+TT (OR=0.654, 95% CI 0.507-0.844) were associated with significantly decreased risk for ESCC. CONCLUSION: A close association exists between functional SNP rs3746804 in C20orf54 and susceptibility to ESCC.

Effects of PLCE1 gene silencing by RNA interference on cell cycling and apoptosis in esophageal carcinoma cells.

Esophageal squamous cell carcinoma (ESCC) is one of the most malignancies with a poor prognosis. The phospholipase C? gene (PLCE1) encodes a novel ras-related protein effector mediating the effects of R-Ras on the actin cytoskeleton and membrane protrusion. However, molecular mechanisms pertinent to ESCC are unclear. We therefore designed PLCE1-special small interfering RNA and transfected to esophageal squamous cell (EC) 9706 cells to investigate the effects of PLCE1 gene silencing on the cell cycle and apoptosis of ESCC and indicate its important role in the development of ESCC. Esophageal cancer tissue specimens and normal esophageal mucosa were obtained and assayed by immunohistochemical staining to confirm overexpression of PLCE1 in neoplasias. Fluorescence microscopy was used to examine transfection efficiency, while the result of PLCE1 silencing was examined by reverse transcription (RT-PCR). Flow cytometry and annexin V apoptosis assays were used to assess the cell cycle and apoptosis, respectively. Expression of cyclin D1 and caspase-3 was detected by Western-blotting. The level of PLCE1 protein in esophageal cancer tissue was significantly higher than that in normal tissue. After transfection, the expression of PLCE1 mRNA in EC 9706 was significantly reduced, compared with the control group. Furthermore, flow cytometry results suggested that the PLCE1 gene silencing arrested the cell cycle in the G0/G1 phase; apoptosis was significantly higher than in the negative control group and mock group. PLCE1 gene silencing by RNAi resulted in decreased expression of cyclin D1 and increased expression of caspase-3. Our study suggests that PLCE1 may be an oncogene and play an important role in esophageal carcinogenesis through regulating proteins which control cell cycling and apoptosis.

[Inducing-apoptosis effect of brucine on human monocytic leukemia cell line THP-1 and its mechanism].

This study was aimed to investigate the inducing-apoptosis effect of brucine on human monocytic leukemia cell line THP-1 cells and its possible mechanism. The inhibition effect of brucine on growth of THP-1 cells was measured by CCK-8 method. Morphological changes of THP-1 cells treated with brucine was detected by acridine orange/ethidium bromide (AO/EB)double staining. Annexin-V/PI double labeling method was used to assay the apoptosis rate of THP-1 cells. The effect of brucine on THP-1 cell cycle distribution was detected by PI single staining. RT-PCR was used to detect the expression of BCL-2 and BAX. The results showed that the brucine could inhibit the THP-1 cell growth in concentration and time-dependent manners at the range of 50 to 400 µg/ml. The cells stained with AO/EB revealed that the brucine induced the nuclear chromatin condensation. After the THP-1 cells were treated with brucine of 400µg/ml for 48 hours, most nucleic were stained as orange-red, and condensed, displaying the late apoptotic cell morphology. Annexin-V/PI detection showed that brucine could induce apoptosis of THP-1 cells in a concentration-dependent manner. Compared with the control group, more cells in brucine-treated group were arrested at G0/G1 phase in a concentration-dependent manner. RT-PCR detection revealed that the expression of BCL-2 was down-regulated strikingly and BAX was up-regulated. It is concluded that brucine can efficiently inhibit cell growth and block THP-1 cells in G0/G1 phase. The mechanism of THP-1 cell apoptosis induced by brucine may be related to the inhibition of BCL-2 and activation of BAX.

Association between Ras association domain family 1A promoter methylation and esophageal squamous cell carcinoma: a meta-analysis.

RASSF1A has been reported to be a candidate tumor suppressor in esophageal squamous cell carcinoma (ESCC). However, the association between RASSF1A promoter methylation and ESCC remains unclear. Eligible studies were identified through searching PubMed, Medline, Web of Science, and the China National Knowledge Infrastucture database. Studies were pooled and odds ratios (ORs) with corresponding confidence intervals (CIs) were calculated. Funnel plots were also performed to evaluate publication bias. Twelve studies involving 859 cases and 675 controls were included in this meta-analysis. A significant association was observed between RASSF1A methylation and ESCC overall (OR = 11.7, 95% CI: 6.59-20.9, z=8.36, P<0.00001). Subgroup analysis showed that the OR for heterogeneous tissues was 5.35 (95% CI = 2.95-9.71) while for autologous tissues it was 16.0 (8.31-30.96). For patient sample size, the OR for the <50 subgroup was 9.92 (95% CI = 2.88-34.2) and for the 50 case group was 13.1 (95% CI = 6.59-25.91). The OR for a relationship between RASSF1A methylation and TNM stages was 0.27 (95% CI=0.10-0.77), whereas there were no significant differences in RASSF1A methylation in relation to gender and differentiation among ESCC cases. This meta-analysis suggests a significant association between RASSF1A methylation and ESCC.

[A new photodynamic therapy drug toward gastric cancer MGC803 cell].

OBJECTIVE: To investigate the treatment efficiency of a new photodynamic therapeutic(PDT) drug synthesized by our laboratory toward MGC803 cells and related mechanisms. METHODS: Bleaching method was used to evaluate the photostability of drug upon repetitive illumination. MTT assay was used to determine the ability of new drug killing MGC803 cells after PDT. Laser scanning confocal microscopy (LSCM) was applied to investigate the subcellular localization of drug in MGC803 cells (mitochondria and/or lysosomes). Hoechst staining and flow cytometry(Annexin V/PI double-staining) were performed to detect the death mode of MGC803 cells after PDT. RESULTS: This new PDT drug had good stability to light irradiation after repetitive illumination. MTT assay showed no cytotoxicity towards MGC803 cells only by drug or only by irradiation(P>0.05), but intense lethal effect was observed with drug and light combination(P<0.05). The phototoxicity of medicine increased with the elevation of concentration, the LD50 was 1.74 µmol/L, and reaching plateau at the concentration of 3.12 µmol/L, even increasing the concentration. LSCM found that drug localized in lysosomes of MGC803 cells. Hoechst staining showed that the death mode of cells was mainly necrosis and Annexin V/PI double-staining proved this result further. CONCLUSION: This new PDT drug is an effective PDT sensitizer for MGC803 cells and the death mode of cells is mainly necrosis.

Gene silencing of ß-catenin by RNAi inhibits proliferation of human esophageal cancer cells by inducing G0/G1 cell cycle arrest.

OBJECTIVES: The aim of the present study was to explore mechanisms underlying the effects of down-regulating ß-catenin expression on esophageal carcinoma (EC) cells. METHODS: Cell cycle distribution and apoptosis were determined using flow cytometry and annexin V apoptosis assay, respectively. Transmission electron microscopy (TEM) was used to examine changes in ultrastructure, while expression of cyclin D1 protein and mRNA was detected by western blot and real-time PCR. Proliferating cell nuclear antigen (PCNA) and extracellular signal-regulated kinase (ERK) 1/2 were evaluated by Western blot analysis. PCNA labeling index (LI) was determined by immunocytochemistry. RESULTS: Compared with pGen-3-con transfected and Eca-109 cells, the percentage of G0/G1-phase pGen-3-CTNNB1 transfected cells was obviously increased (P<0.05), with no significant difference among the three groups with regard to apoptosis (P>0.05). pGen-3-CTNNB1 transfected cells exhibited obvious decrease in cyclin D1 mRNA and protein expression (P<0.05) and the ultrastructure of Eca-109 cells underwent a significant change after being transfected with pGen-3-CTNNB1, suggesting that down-regulating ß-catenin expression can promote the differentiation and maturation. The expression of PCNA and the ERKI/2 phosphorylation state were also down-regulated in pGen-3-CTNNB1 transfected cells (P<0.05). At the same time, the PCNA labeling index was decreased accordingly (P<0.05). CONCLUSION: Inhibition of EC Eca-109 cellproliferation by down-regulating ß-catenin expression could improve cell ultrastructure by mediating blockade in G0/G1 through inhibiting cyclin D1, PCNA and the MAPK pathway (p-ERK1/2).

[The relationship between C20orf54 gene rs3746804 position single nucleotide polymorphism and susceptibility to esophageal squamous cell carcinoma].

OBJECTIVE: To explore the association of C20orf54 gene rs3746804 position single nucleotide polymorphism and susceptibility to esophageal squamous cell carcinoma (ESCC). METHODS: Purification of genomic DNA from whole blood was used the Maxwell(16) System. rs3746804 in C20orf54 was detected by direct sequencing in 434 ESCC patients from Changzhi (Shanxi province) and Linzhou (Henan province) and 554 healthy controls from Changzhi, Linzhou and including immigrators from Linzhou to Changzhi. RESULTS: For rs3746804, the genotypic frequencies of CT (37.5% vs 51.0%, 37.5% vs 52.0%), CC (44.2% vs 34.8%, 44.2% vs 33.0%) in Changzhi ESCC patients showed significant differences with healthy Changzhi controls and the healthy immigrator controls (all P < 0.05), and the frequencies of TT (18.3% vs 4.1%) and CC (44.2% vs 54.6%) in Changzhi ESCC patients showed significant differences with Linzhou ESCC patients (all P < 0.05). The genotypic frequencies of TT (4.1% vs 15.0%), CT (41.2% vs 52.0%) and CC(54.6% vs 33.0%) showed significant differences between Linzhou ESCC patients and the healthy immigrator controls (all P < 0.05), and the frequencies of TT (4.1% vs 14.1%) and CC(54.6% vs 34.8%) showed significant differences between Linzhou ESCC patients and Changzhi healthy controls (all P < 0.01). Meanwhile, there were significant differences between ESCC patients (including Changzhi and Linzhou ESCC patients) and healthy controls (including the healthy Changzhi, Linzhou and immigrator controls) in genotypic frequencies of CT (39.2% vs 48.7%) and CC (48.8% vs 38.2%) (all P < 0.01). CT and CT + TT genotype could decrease the risk of ESCC compared with the CC genotype (OR = 0.630, 95%CI 0.481 - 0.826; OR = 0.654, 95%CI 0.507 - 0.844). CONCLUSION: There is a closed relationship between SNP rs3746804 in C20orf54 and susceptibility to ESCC.