When do defecation function and quality of life recover for patients with non-ostomy and ostomy surgery of rectal cancer?

BACKGROUND: Rectal cancer (RC) surgery often results in permanent colostomy, seriously limiting the quality of life (QOL) in patients in terms of bowel function. This study aimed to examine defecation function and QOL in RC patients who underwent non-ostomy or ostomy surgery, at different time-points after surgery. METHODS: In total, 82 patients who underwent an ostomy and 141 who did not undergo an ostomy for the treatment of RC at our colorectal surgery department between January 2013 and January 2015 were enrolled. Surgical methods, tumor distance from the anal margin (TD), anastomosis distance from the anal margin (AD) and complications were compered between the non-ostomy and ostomy surgery groups. QOL was compared between the two groups at years 2, 3, and 4 after surgery. The Wexner score and the validated cancer-specific European Organization for Research and Treatment of Cancer (EORTC QLQ-CR30) questionnaire scores were assessed for all patients in January 2017. SPSS 21.0 was utilized for all data analyses. RESULTS: Surgical methods, TD, and AD significantly differed between the non-ostomy and ostomy surgery groups (all P < .001). However, no differences were found in the number of complications between the groups (P = .483). For the 192 patients undergoing Dixon surgery, role function (RF), global QOL (GQOL), sleep disturbance, and the incidence of constipation showed significant differences between the two groups (P = .012, P = .025, P = .036, and P = .015, respectively). In the 31 cases of permanent ostomy, we observed significant differences in GQOL scores, dyspnea incidence, and financial difficulties across the different years (P = .002, P = .036, and P < .01, respectively). Across all 223 cases, there were significant differences in social function and GQOL scores in the second year after surgery (P = .014 and P < .001, respectively). However, no differences were observed in the other indices across the three time-points. CONCLUSIONS: RC patients undergoing ostomy surgery, especially those with low and super-low RC, revealed poorer defecation function and QOL in the present study. However, 2 years after surgery, most of the defecation and QOL indicators showed recovery.

Association of CDKN2BAS polymorphism rs4977574 with coronary heart disease: a case-control study and a meta-analysis.

The goal of our study was to explore the significant association between a non-protein coding single nucleotide polymorphism (SNP) rs4977574 of CDKN2BAS gene and coronary heart disease (CHD). A total of 590 CHD cases and 482 non-CHD controls were involved in the present association study. A strong association of rs4977574 with CHD was observed in females (genotype: p=0.002; allele: p=0.002, odd ratio (OR)=1.57, 95% confidential interval (CI)=1.18-2.08). Moreover, rs4977574 was more likely to be a risk variant of CHD under the recessive model in females (χ2=10.29, p=0.003, OR=2.14, 95% CI=1.31-2.77). A breakdown analysis by age had shown that there was an 87% increased risk of CHD for females younger than 65 years (genotype: χ2=14.64, degrees of freedom (df)=2, p=0.0002; allele: χ2=11.31, df=1, p=0.0008, OR=1.87, 95% CI=1.30-2.70). Similar observation was also found in males younger than 65 years (genotype: χ2=8.63, df=2, p=0.04; allele: χ2=7.55, df=1, p=0.006, OR=1.45, 95% CI=1.11-1.90). p values were adjusted by age, sex, smoking, high density lipoprotein cholesterol (HDL-C) and low density lipoprotein cholesterol (LDL-C). Meta-analysis of 23 studies among 36,452 cases and 39,781 controls showed a strong association between rs4977574 and the risk of CHD (p<0.0001, OR=1.27, 95% CI=1.22-1.31).

Identification of Key Genes and Pathways in Oxaliplatin-Induced Neuropathic Pain Through Bioinformatic Analysis.

Background: The mechanism of Chemotherapy-induced neuropathic pain (NP) remains obscure. This study was aimed to uncover the key genes as well as protein networks that contribute to Oxaliplatin-induced NP. Material/Methods: Oxaliplatin frequently results in a type of Chemotherapy-induced NP that is marked by heightened sensitivity to mechanical and cold stimuli, which can lead to intolerance and discontinuation of medication. We investigated whether these different etiologies lead to similar pathological outcomes by targeting shared genetic targets or signaling pathways. Gene expression data were obtained from the Gene Expression Comprehensive Database (GEO) for GSE38038 (representing differential expression in the spinal nerve ligation model rats) and GSE126773 (representing differential expression among the Oxaliplatin-induced NP model rats). Differential gene expression analysis was performed using GEO2R. Results: Protein-protein interaction (PPI) analysis identified 260 co-differentially expressed genes (co-DEGs). Subsequently, Kyoto Encyclopedia of Genes and Genomes (KEGG) analysis revealed three shared pathways involved in both models: Kaposi sarcoma-associated herpesvirus (KSHV) infection, Epstein-Barr virus (EBV) infection, and AGE-RAGE signaling pathway in diabetic complications. Further bioinformatics analysis highlighted eight significantly up-regulated genes in the NP group: Mapk14, Icam1, Cd44, IL6, Cxcr4, Stat1, Casp3 and Fgf2. Our results suggest that immune dysfunction, inflammation-related factors or regulating inflammation factors may also be related to Oxaliplatin-induced NP. Additionally, we analyzed a dataset (GSE145222) involving chronic compression of DRGs (CCD) and control groups. CCD model is a classic model for studying NP. We assessed these hub genes' expression levels. In contrast with the control groups, the hub genes were up-regulated in CCD groups, the difference was statistically significant, except Stat1. Conclusion: Our research significantly contributes to elucidating the mechanisms underlying the occurrence as well as the progression of Oxaliplatin-induced NP. We have identified crucial genes and signaling pathways associated with this condition.

Correlation between mismatch repair statuses and the prognosis of stage I-IV colorectal cancer.

Background: The role of microsatellite instability (MSI) and prognosis for stage II-III colorectal cancer (CRC) has been described, but the role of MSI in stage I and IV CRC is controversial. Methods: A total of 2,540 CRC patients were collected from Huzhou Central Hospital, China, from January 2006 to 2016, and 783 cases were excluded. This retrospective study illustrates the correlation between MMR status and prognosis for 1,757 CRC patients as well as the correlation between MSI and prognosis for CRC patients. Two groups were classified as MSI-H and MSI-L&MSS. If the expression of one or more mismatch repair (MMR) proteins was negative, it was considered as microsatellite instability high expression (MSI-H), whereas positive expression was considered as microsatellite instability low expression and microsatellite stability (MSI-L&MSS), as assessed by correlation analyses. Overall and disease-free survival were analyzed using the Kaplan-Meier method. Univariable and multivariable analyses were conducted using Cox regression. Results: Preoperative serum S-CEA, positive lymph, tumor size, pathologic tumor (Pt) status, node (N) stage, differentiation, chemotherapy, and the 8th Edition of the American Joint Committee on Cancer (AJCC-8) were significantly correlated with MSI (P=0.028, 0.037, 0.019, 0.007, 0.002, <0.001, <0.001, and <0.001, respectively), whereas tumor location was not associated with MSI. Univariable and multivariable analyses showed that MSI was an independent factor for CRC. The 5-year overall survival (OS) and 5-year disease-free survival (DFS, P<0.001) rates differed significantly between the two groups in stages II, III, and IV, whereas stage I did not show a significant difference (P>0.05). Conclusion: MSI-H was associated with a good prognosis for stages II to IV, whereas stage I did not show any significant correlation. Moreover, MSI expression was an independent prognostic factor.

Pink1 promotes cell proliferation and affects glycolysis in breast cancer.

Phosphatase and tensin homolog (PTEN)-induced kinase 1 (Pink1) is regarded as a tumor suppressor and plays an important role in cancer cell biology, while relatively few studies have examined Pink1 in breast cancer, especially in vivo. The aims of this study were to investigate Pink1 expression in different subtypes of breast cancer tissues and cell lines and explore the effect of Pink1 protein on breast cancer. In these experiments, Pink1 expression was investigated using the tissue microarray immunohistochemistry (TMA-IHC) method in 150 samples of breast cancer tissues with different subtypes, and strong staining of Pink1 was significantly correlated with the histological grade of breast cancer (p = 0.015). In addition, Pink1 messenger RNA (mRNA) displayed much higher expression levels in breast cancer cell lines than in MCF-10A breast epithelial cells. Moreover, proteomic data obtained by isobaric tags for relative and absolute quantification (iTRAQ) showed that Pink1 deletion induced a distinct proteomic profile in MDA-MB-231 cells, and enrichment analysis showed that the differential proteins were concentrated mainly in energy metabolism-related pathways. Moreover, Seahorse XF analysis showed that Pink1 knockout reduced the glycolytic ability of MDA-MB-231 cells. Our findings indicated that Pink1 may be an indicator of malignancy in breast cancer and that it presents oncogenic properties in breast cancer, which raises another perspective for understanding the regulatory role of Pink1 in breast cancer.

Screening of Key Proteins Affecting Floral Initiation of Saffron Under Cold Stress Using iTRAQ-Based Proteomics.

BACKGROUND: Saffron crocus (Crocus sativus) is an expensive and valuable species that presents preventive and curative effects. This study aimed to screen the key proteins affecting the floral initiation of saffron under cold stress and thus increasing yield by regulating the temperature. RESULTS: Protein expression profiles in flowering and non-flowering saffron buds were established using isobaric tags for relative or absolute quantitation (iTRAQ). A total of 5,624 proteins were identified, and 201 differentially abundant protein species (DAPs) were further obtained between the flowering and non-flowering groups. The most important functions of the upregulated DAPs were "sucrose metabolic process," "lipid transport," "glutathione metabolic process," and "gene silencing by RNA." Downregulated DAPs were significantly enriched in "starch biosynthetic process" and several oxidative stress response pathways. Three new flower-related proteins, CsFLK, CseIF4a, and CsHUA1, were identified in this study. The following eight key genes were validated by real-time qPCR in flowering and non-flowering top buds from five different growth phases: floral induction- and floral organ development-related genes CsFLK, CseIF4A, CsHUA1, and CsGSTU7; sucrose synthase activity-related genes CsSUS1 and CsSUS2; and starch synthase activity-related genes CsGBSS1 and CsPU1. These findings demonstrate the important roles played by sucrose/starch biosynthesis pathways in floral development at the mRNA level. During normal floral organ development, the sucrose contents in the top buds of saffron increased, and the starch contents decreased. In contrast, non-flowering buds showed significantly decreased sucrose contents under cold stress and no significant changes in starch contents compared with those in the dormancy stage. CONCLUSION: In this report, the protein profiles of saffron under cold stress and a normal environment were revealed for the first time by iTRAQ. A possible "reactive oxygen species-antioxidant system-starch/sugar interconversion flowering pathway" was established to explain the phenomenon that saffron does not bloom due to low temperature treatment.

Cutoff of 25% for Ki67 expression is a good classification tool for prognosis in colorectal cancer in the AJCC­8 stratification.

Ki­67 expression has been widely used in clinical practice as an index to evaluate the proliferative activity of tumor cells. The cutoff for Ki67 expression in order to increase the prognostic value of Ki67 expression in colorectal cancer varies. The present study assessed the relationship between the 25% cutoff for Ki67 expression and prognosis in colorectal cancer in the AJCC­8 (American Joint Committee on Cancer 8 edition) stratification. The current trial included 1,090 colorectal cancer patients enrolled from 2006 to 2012 at Huzhou Central Hospital. Ki67 expression was classified according to 25% intervals, dividing the patients into four groups. Measurement data were analyzed by ANOVA, and count data by Crosstabs. Bivariate correlation analysis was performed to assess clinicopathological indicators based on Ki67 expression. Disease­free survival (DFS) and overall survival (OS) based on Ki67 levels were analyzed by the Kaplan­Meier method. A total of 1,090 patients of the 2,080 enrolled CRC cases were evaluated (52.4%). Invasive depth, tumor differentiation, tumor size, AJCC­8, positive number of lymph nodes and chemotherapy status showed significant differences in the various Ki67 expression groups (all P<0.05), with significant correlations (Spearman rho: 0.170, 0.456, 0.22, 0.195, 0.514 and ­0.201, respectively, all P<0.001). DFS and OS for the different Ki67 level groups based on AJCC­8 stratification were analyzed, and no significance was found in stage IV (P=0.334). DFS and OS survival rates were assessed at different Ki67 expression levels, and no significant differences were found (all P>0.05). Cox regression analysis showed that invasive depth, lymph node metastasis, tumor differentiation, AJCC­8 and Ki67 were independent factors affecting colorectal cancer (P=0.030, all others P<0.001). In conclusion, a cutoff of 25% for Ki67 expression is a good classification tool. High Ki67 has a close association with poor prognosis in colorectal cancer and independently predicts prognosis in the AJCC­8 stratification.

Identification of Hub Genes as Biomarkers Correlated with the Proliferation and Prognosis in Lung Cancer: A Weighted Gene Co-Expression Network Analysis.

Lung cancer is one of the most malignant tumors in the world. Early diagnosis and treatment of lung cancer are vitally important to reduce the mortality of lung cancer patients. In the present study, we attempt to identify the candidate biomarkers for lung cancer by weighted gene co-expression network analysis (WGCNA). Gene expression profile of GSE30219 was downloaded from the gene expression omnibus (GEO) database. The differentially expressed genes (DEGs) were analyzed by the limma package, and the co-expression modules of genes were built by WGCNA. UALCAN was used to analyze the relative expression of normal group and tumor subgroups based on tumor individual cancer stages. Survival analysis for the hub genes was performed by Kaplan-Meier plotter analysis with the TCGA database. A total of 2176 genes (745 upregulated and 1431 downregulated genes) were obtained from the GSE30219 database. Seven gene co-expression modules were conducted by WGCNA and the blue module might be inferred as the most crucial module in the pathogenesis of lung cancer. In the pathway enrichment analysis of KEGG, the candidate genes were enriched in the "DNA replication," "Cell cycle," and "P53 signaling pathway" pathways. Among these, the cell cycle pathway was the most significant pathway in the blue module with four hub genes CCNB1, CCNE2, MCM7, and PCNA which were selected in our study. Kaplan-Meier plotter analysis indicated that the high expressions of four hub genes were correlated with a worse overall survival (OS) and advanced tumors. qRT-PCR showed that mRNA expression levels of MCM7 (p = 0.038) and CCNE2 (0.003) were significantly higher in patients with the TNM stage. In summary, the high expression of the MCM7 and CCNE2 were significantly related with advanced tumors and worse OS in lung cancer. Thus, the MCM7 and CCNE2 genes can be good indicators for cellular proliferation and prognosis in lung cancer.

Downregulation of PTPRK Promotes Cell Proliferation and Metastasis of NSCLC by Enhancing STAT3 Activation.

OBJECTIVE: The receptor-type tyrosine-protein phosphatase κ (PTPRK) is a candidate tumor suppressor involved in the tumorigenesis of various organs. However, its expression and biological roles in non-small-cell lung cancer (NSCLC) have not yet been investigated. METHODS: PTPRK expression in NSCLC tissues and cell lines was examined using real-time PCR and western blotting. In addition, the effects of PTPRK on cell migration, invasion, and proliferation were evaluated in vitro. Furthermore, we explored whether the downregulation of PTPRK led to STAT3 activation in NSCLC cell lines by western blotting. The expression of phospho-STAT3Tyr705 in primary human NSCLC tissues was evaluated by immunohistochemistry. RESULTS: The results showed that PTPRK expression was frequently reduced in NSCLC tissues with lymph node metastasis and cell lines. The inhibition of PTPRK expression resulted in increased proliferation, invasion, and migration of NSCLC cells in vitro. Additionally, after silencing of PTPRK, phospho-STAT3Tyr705 was significantly increased in NSCLC cells. Moreover, the phospho-STAT3Tyr705 levels of NSCLC tissues were positively correlated with lymph node metastasis and significantly inversely correlated with the expression of PTPRK (p < 0.05). CONCLUSIONS: These results suggested that PTPRK functions as a novel tumor suppressor in NSCLC, and its suppressive ability may be involved in STAT3 activation.

Association of Sperm Methylation at LINE-1, Four Candidate Genes, and Nicotine/Alcohol Exposure With the Risk of Infertility.

In this study, we examined whether smoking and drinking affect sperm quality and the DNA methylation of the repetitive element LINE-1, MEST, P16, H19, and GNAS in sperm. Semen samples were obtained from 143 male residents in a minority-inhabited district of Guizhou province in southwest China. Quantitative DNA methylation analysis of the samples was performed using MassARRAY EpiTYPER assays. Sperm motility was significantly lower in both the nicotine-exposed (P = 0.0064) and the nicotine- and alcohol-exposed (P = 0.0008) groups. Follicle-stimulating hormone (FSH) levels were higher in the nicotine-exposed group (P = 0.0026). The repetitive element LINE-1 was hypermethylated in the three exposed groups, while P16 was hypomethylated in the alcohol and both the alcohol and nicotine exposure groups. Our results also show that alcohol and nicotine exposure altered sperm cell quality, which may be related to the methylation levels of MEST and GNAS. In addition, MEST, GNAS, and the repetitive element LINE1 methylation was significantly associated with the concentration of sperm as well as FSH and luteinizing hormone levels.

Elevated methylation of cyclin dependent kinase inhibitor 2B contributes to the risk of coronary heart disease in women.

Cyclin dependent kinase inhibitor 2B (CDKN2B) encodes a cyclin-dependent kinase inhibitor that may enhance the formation of atherosclerotic plaques. The aim of the present study was to investigate the contribution of CDKN2B promoter methylation on the risk of coronary heart disease (CHD). The present results indicated a significant association between increased CDKN2B methylation and the risk of CHD (adjusted P=0.043). A breakdown analysis according to sex demonstrated that CDKN2B methylation was significantly associated with the risk of CHD in women (adjusted P=0.010), but not in men. A further breakdown analysis by age indicated a significant association of CHD in the women >60 years (P=0.024). Luciferase reporter gene assay results indicated that the CDKN2B promoter fragment significantly enhanced luciferase activity (P<0.001). In addition, CDKN2B transcription was significantly enhanced following treatment with 5-aza-2'-deoxycytidine methylation inhibitor in human aortic endothelial cells (HAEC) and human primary coronary artery smooth muscle cells (HPCASMC; P<0.05 and P<0.01), but not in 293 cells. Notably, estrogen treatment reduced CDKN2B methylation of several CpGs and significantly increased CDKN2B gene expression levels in HAEC, HPCASMC and 293 cells (P<0.05 and P<0.01). Additionally, treatment of HAEC and HPCASMC with simvastatin and γ-carboxy-L-glutamic acid reduced CDKN2B promoter methylation and increased CDKN2B transcription concomitantly. The present study suggests that CDKN2B promoter methylation may be associated with sex dimorphism in the pathogenesis of CHD.

The role of tissue and serum carcinoembryonic antigen in stages I to III of colorectal cancer-A retrospective cohort study.

PURPOSE: Tissue carcinoembryonic antigen (t-CEA) and serum carcinoembryonic antigen (s-CEA) expression profiles are the most useful tumor markers for the diagnosis and evaluation of colorectal cancer (CRC) worldwide; however, their roles in CRC progression remain controversial. This study aimed to compare the prognostic values of both s-CEA and t-CEA in CRC. METHODS: A total of 517 patients from January 2006 to December 2010 with stages I-III CRC were retrospectively examined, with 5-year postoperative follow-up and death as end-points. T-CEA expression, s-CEA expression, and clinical pathological parameters were inputted into the SPSS 21.0 software. The Kaplan-Meier method was used to analyze the 5-year disease-free survival (DFS) rate of patients in different tumor node metastasis (TNM) stages based on t-CEA and s-CEA expression. RESULTS: Tumor differentiation and the number of positive lymph node harvests were significantly different among the t-CEA groups (P < 0.001, P = 0.002); however, clinicopathological features showed no significant difference. The groups with high s-CEA and t-CEA expression had a significantly poorer prognosis than those with low s-CEA (P = 0.021) and t-CEA (P < 0.01) expression, respectively. The multivariate analysis demonstrated that t-CEA was an independent prognostic factor in CRC (P < 0.001), but s-CEA was not (P = 0.339). The 5-year disease-free survival rates among the t-CEA groups were significantly different in stages I, II, and III of CRC (P = 0.001, P < 0.001, P < 0.001), whereas in the s-CEA groups, this difference was observed only in stage III (P = 0.014). CONCLUSION: This study shows that postoperative t-CEA expression is an independent factor associated with poorer CRC prognosis and has a higher prognostic value than that of preoperative s-CEA expression.

Comparison of the eighth version of the American Joint Committee on Cancer manual to the seventh version for colorectal cancer: A retrospective review of our data.

AIM: To analyze the survival trends in colorectal cancer (CRC) based on the different classifications recommended by the seventh and eighth editions of the American Joint Committee on Cancer staging system (AJCC-7th and AJCC-8th). METHODS: The database from our institution was queried to identify patients with pathologically confirmed stage 0-IV CRC diagnosed between 2006 and 2012. Data from 2080 cases were collected and 1090 cases were evaluated through standardized inclusion and exclusion criteria. CRC was staged by AJCC-7th and then restaged by AJCC-8th. Five-year disease-free survival (DFS) and overall survival (OS) were compared. SPSS 21.0 software was used for all data. DFS and OS were compared and analyzed by Kaplan-Meier and Log-rank test. RESULTS: Linear regression and automatic linear regression showed lymph node positive functional equations by tumor-node-metastasis staging from AJCC-7th and tumor-node-metastasis staging from AJCC-8th. Neurological invasion, venous infiltration, lymphatic infiltration, and tumor deposition put forward stricter requirements for pathological examination in AJCC-8th compared to AJCC-7th. After re-analyzing our cohort with AJCC-8th, the percentage of stage IVB cases decreased from 2.8% to 0.8%. As a result 2% of the cases were classified under the new IVC staging. DFS and OS was significantly shorter (P = 0.012) in stage IVC patients compared to stage IVB patients. CONCLUSION: The addition of stage IVC in AJCC-8th has shown that peritoneal metastasis has a worse prognosis than distant organ metastasis in our institution's CRC cohort. Additional datasets should be analyzed to confirm these findings.

Astrocyte elevated gene-1 is overexpressed in non-small-cell lung cancer and associated with increased tumour angiogenesis.

OBJECTIVES: Astrocyte elevated gene-1 (AEG-1) functions to mediate angiogenesis, and its upregulation is responsible for tumour angiogenesis during cancer development. This study analysed AEG-1 expression in non-small-cell lung cancer (NSCLC) for association with NSCLC clinicopathological features and tumour angiogenesis. METHODS: The expression of AEG-1, vascular endothelial growth factor and intratumoural microvessel density (assessed using the expression of CD105) was detected by immunohistochemistry in 88 paired tumour tissue and adjacent normal tissue specimens obtained from NSCLC patients. The Kaplan-Meier curves were used for survival analysis through an online tool (http://kmplot.com/analysis/). RESULTS: AEG-1 was overexpressed in 61.3% of NSCLC tissues vs 6.8% (6/88) of normal tissues (P < 0.001). AEG-1 expression in NSCLC was significantly associated with advanced pTNM stage (P = 0.021), tumour dedifferentiation (P = 0.034), vascular invasion (P = 0.035), lymph node metastasis (P < 0.001) and poor overall survival (P = 0.024). Moreover, the expression of AEG-1 in NSCLC was associated with tumour angiogenesis; that is, vascular endothelial growth factor overexpression (P < 0.001) and intratumoural microvessel density (P < 0.001). CONCLUSIONS: This study demonstrates that AEG-1 expression is associated with NSCLC development, angiogenesis, progression and poor prognosis, indicating that the adjuvant therapy with antiangiogenic agent be adopted for the early postoperative period before the start of conventional chemotherapy in patients with AEG-1 overexpressed NSCLC.

Hypermethylation of the κ1 opioid receptor promoter in Chinese heroin and methamphetamine addicts.

Heroin and methamphetamine (METH) addiction continues to be a major social, economic and therapeutic problem worldwide. The opioid pathway may mediate the effects of addictive drugs. However, the potential correlation between the κ1 opioid receptor (OPRK1) and drug addiction has not yet been characterized. The aim of the present study was to investigate the potential association between methylation of the OPRK1 promoter and substance abuse. Bisulfite pyrosequencing technology was used to determine the levels of OPRK1 promoter methylation in 60 drug abusers (30 heroin and 30 METH addicts) and 52 controls, observed to exhibit no significant differences in age or gender. The results indicated that levels of OPRK1 promoter methylation were significantly higher in drug addicts when compared with controls (P=2.43×10-4). Significant correlations between OPRK1 promoter methylation and the length and frequency of drug use were also observed in male heroin addicts (length: r=0.661, P=0.007; frequency: r=-0.684, P=0.005). In addition, a luciferase reporter gene assay indicated that the OPRK1 promoter fragment was able to regulate gene expression (fold change between two groups >32.12, P≤0.0001). In conclusion, results of the present study indicate that methylation of the OPRK1 promoter contributes to the pathophysiology of drug addiction.

Dopamine receptor D4 promoter hypermethylation increases the risk of drug addiction.

Heroin and methylamphetamine (METH) are two addictive drugs that cause serious problems for society. Dopamine receptor D4 (DRD4), a key receptor in the dopaminergic system, may facilitate the development of drug addiction. The aim of the present study was to investigate the association between the promoter methylation level of DRD4 gene and drug addiction. Bisulfite pyrosequencing technology was used to measure the methylation levels of DRD4 promoter in 60 drug addicts and 52 matched controls. Significantly higher levels of DRD4 CpG1 and CpG4 methylation were detected in METH and heroin drug addicts compared with controls (P<0.05). Male METH addicts exhibited significantly higher DRD4 CpG1, CpG2 and CpG4 methylation levels compared with sex-matched controls (P<0.05). In heroin addicts, a positive correlation was observed between depression-dejection and DRD4 CpG5 methylation (r=0.537, P=0.039) whereas there was a negative correlation between drug usage frequency and CpG1 methylation (r=-0.632, P=0.011). In METH addicts, methylation levels were not significantly associated with depression-dejection and drug usage frequency. In addition, luciferase assays demonstrated that the target sequence of the DRD4 promoter upregulates gene expression. The results of the present study suggest that DNA methylation of DRD4 may be responsible for the pathophysiology of drug addiction.

MiR-1260b promotes the migration and invasion in non-small cell lung cancer via targeting PTPRK.

OBJECTIVE: Non-small cell lung cancer (NSCLC) accounts for 80-85% of lung cancer cases which cause most of cancer-related deaths globally. As our previous study discovered miR-1260b can be regarded as a specific signature for metastasis in NSCLC patients. However, the molecular mechanisms of miR-1260b underlying NSCLC progression and metastasis remain dismal. METHODS: The expression of miR-1260b in NSCLC tissues and cell lines were examined by real-time PCR, the effects of miR-1260b on cell migration, invasion and proliferation were evaluated in vitro. Furthermore, luciferase reporter assay was performed to identify the targets of miR-1260b, and the association between miR-1260b and its target gene was determined by real-time PCR and western blot assay. RESULTS: The results showed that miR-1260b was significantly upregulated in NSCLC cell lines. The inhibition of miR-1260b expression decreased the migratory and invasive rates in A549 cells while miR-1260b overexpression had the opposite effect. Furthermore, PTPRK was identified as a direct target of miR-1260b, and PTPRK expression was inversely correlated with miR-1260b in NSCLC cell lines and clinical tissues. CONCLUSIONS: These results suggested that miR-1260b may play an important role in NSCLC metastasis progression and could serve as a putative target for diagnosis and treatment of NSCLC.

AEG-1 mRNA expression in non-small cell lung cancer is associated with increased tumor angiogenesis.

Astrocyte-elevated gene-1 (AEG-1) is implicated in the oncogenesis and angiogenesis of various types of human malignant disease. However, the angiogenesis roles of AEG-1 in non-small cell lung cancer (NSCLC) remain to be further elucidated. In the present study, the expression level of AEG-1 mRNA in seven human lung cell lines and 89 paired tissue samples (tumor tissues (TTs) and pair-matched normal adjacent tissues (PMNATs)) from NSCLC patients was detected by real-time PCR. Staining of vascular endothelial growth factor (VEGF) and intratumoral microvessel density (iMVD, labeled by CD105) were assessed by immunohistochemistry. Furthermore, cell migration and invasion were evaluated by wound healing assay and transwell assays. AEG-1 mRNA level was significantly higher in human lung cancer cells and TTs than that in human normal bronchial epithelial cell line 16HBE and PMNATs, respectively (P<0.001). Higher AEG-1 mRNA level in patients with NSCLC was correlated with clinical stages (P=0.028), differentiation (P=0.042), and lymph node metastasis (P=0.004). Moreover, Upregulated AEG-1 mRNA expression level was associated with higher tumor angiogenesis, reflected by the increase of VEGF expression and iMVD counting (P=0.021, P<0.001). However, 95D cell line transfected with AEG-1 siRNA oligos (siAEG-1) exhibited no significant decrease of cell invasion or migration capacities when compared with the control cells (P>0.05).These results suggested that AEG-1 may play important roles at the transcription level in malignant transformation and tumor angiogenesis in NSCLC, and anti-AEG-1 mRNA expression may be a novel potential strategy for anti-angiogenic therapy of NSCLC.

Elevated OPRD1 promoter methylation in Alzheimer's disease patients.

Aberrant DNA methylation has been observed in the patients with Alzheimer's disease (AD), a common neurodegenerative disorder in the elderly. OPRD1 encodes the delta opioid receptor, a member of the opioid family of G-protein-coupled receptors. In the current study, we compare the DNA methylation levels of OPRD1 promoter CpG sites (CpG1, CpG2, and CpG3) between 51 AD cases and 63 controls using the bisulfite pyrosequencing technology. Our results show that significantly higher CpG3 methylation is found in AD cases than controls. Significant associations are found between several biochemical parameters (including HDL-C and ALP) and CpG3 methylation. Subsequent luciferase reporter gene assay shows that DNA fragment containing the three OPRD1 promoter CpGs is able to regulate gene expression. In summary, our results suggest that OPRD1 promoter hypermethylation is associated with the risk of AD.

H4K5 histone acetylation of BRG1 is associated with heroin administration rather than addiction.

Diacetylmorphine hydrochloride (heroin) addiction is a chronic relapsing brain disorder that is a heavy public health burden worldwide. Brm/SWI2-related gene-1 (BRG1) is a tumor suppressor gene that can influence embryogenesis and the development of the cerebellum. The current study aimed to investigate the effect of histone H4 lysine 5 (H4K5) modifications on the BRG1 gene in brain tissue of the ventral tegmental area (VTA) of heroin-addicted rats. A total of 21 male Sprague Dawley rats were raised in a standard manner and underwent heroin self-administration training. Rats were randomly divided into three equal groups: Group A, self-administered delivery of heroin; group B, yoked delivery of heroin; and group C, yoked delivery of saline. The VTA was harvested and subjected to chromatin immunoprecipitation (ChIP) analysis. Gene expression was evaluated by quantitative polymerase chain reaction. We calculated the recovery rate, which indicated the percentage of the total input BRG1 recovered by ChIP. Our results showed that BRG1 was less associated with H4K5 histone modification in the group of rats that underwent heroin self-administration than in the other two groups (A vs. B, P=0.031; A vs. C, P=0.067). The recovery fold changes of the self-administration group and the passive-administration group were significantly different from those of the group with yoked saline (A vs. C, P=0.013; B vs. C, P=0.009; A vs. B, P=0.731). The results of the current study demonstrated that H4K5 histone acetylation of BRG1 in the VTA may be associated with heroin administration, but not addiction.