Promoter Methylation Levels Research Articles

Promoter methylation levels of RASSF1 and ATIC genes are associated with lung cancer in Iranian patients.

Epigenetic alterations like methylation of tumor suppressor genes or oncogenes, in respiratory epithelium have been associated with lung cancer. Hypermethylation of genes promoter is an epigenetic event, and is responsible to tumor suppressor genes inactivation as well as oncogenes activation. This study aimed to assess the role of methylation status in promoter of RASSF1 and ATIC genes their potential implication in the pathogenesis of lung tumor in Iranian patients. In this study, we collected 100 tissue samples (50lung cancer tissues and 50 adjacent non-cancerous lung tissues) from Iranian lung cancer patients. The genomic DNA was extracted, and methylation status of both RASSF1 andATIC genes was investigated by methylation-sensitive high-resolution melting (MS-HRM) assay technique and Real-Time PCR. Cancer Genome Atlas (TCGA) dataset wasalso analyzed for further validation of the gene's methylation. Methylation of RASSF1 gene promoter was significantly higher in lung tumor tissues. However, promoter methylation levels of ATIC gene was significantly lower in lung tumor tissues. These results were additionally confirmed by TCGA analysis. Promoter methylation of both RASSF1 and ATIC genes was significantly associated with lymph node metastasis, and clinical stage of lung cancer. Thereceiver operating characteristic (ROC) curve analysis indicated a high accuracy of promoter methylation in these genes as a diagnostic biomarker for lung cancer. Methylation levels of both RASSF1 and ATIC genes promoters were associated with lung cancer pathogenesis in Iranian population, and may be a suitable biomarker for diagnosis and prognosis of lung cancer in early stage of tumorigenesis.

Quantitative analysis of MGMT promoter methylation in glioblastoma suggests nonlinear prognostic effect.

Epigenetic inhibition of the O6-methylguanine-DNA-methyltransferase (MGMT) gene has emerged as a clinically relevant prognostic marker in glioblastoma (GBM). Methylation of the MGMT promoter has been shown to increase chemotherapy efficacy. While traditionally reported as a binary marker, recent methodological advancements have led to quantitative methods of measuring promoter methylation, providing clearer insight into its functional relationship with survival. A CLIA assay and bisulfite sequencing was utilized to develop a quantitative, 17-point, MGMT promoter methylation index. GBMs of 240 newly diagnosed patients were sequenced and risk for mortality was assessed. Nonlinearities were captured by fitting splines to Cox proportional hazard models and plotting smoothed residuals. Covariates included age, Karnofsky performance status, IDH1 mutation, and extent of resection. Median follow-up time and progression-free survival were 16 and 9 months, respectively. A total of 176 subjects experienced death. A one-unit increase in promoter CpG methylation resulted in a 4% reduction in hazard (95% CI 0.93-0.99, P < .005). GBM patients with low levels of promoter methylation (1-6 CpG sites) fared markedly worse (HR = 1.62, 95% CI 1.03-2.54, P < .036) than individuals who were unmethylated. Subjects with medium levels of promoter methylation (7-12 sites) had the greatest reduction in hazard (HR = 0.48, 95% CI 0.29-0.80, P < .004), followed by individuals in the highest promoter methylation tertile (HR = 0.62, 95% CI 0.40-0.97, P < .035). Our findings suggest that the relationship between the extent of MGMT promoter methylation and survival in GBM may be nonlinear. These findings challenge the current understanding of MGMT and underlines the clinical importance of determining its prognostic utility. Potential limitations include censoring, sample size, and extraneous mutations.

Hypermethylated WASF2: tumor suppressive role in head and neck squamous cell carcinoma.

WASF2 regulates actin reorganization during cell migration. WASF2 has been identified as a regulator of the development of gastric cancer, breast cancer, and pancreatic cancer. But its regulatory mechanisms remain unknown. Also, its function was absent in head and neck squamous cell carcinoma (HNSCC). Consequently, we examined the effect of DNA methylation on aberrant WASF2 expression in HNSCC. TNMplot, TIMER, GSEA pathway analysis, and the Kaplan-Meier Plotter database were used to analyze the expression, function, and prognostic value of WASF2, as well as the correlation between WASF2 and infiltrating immune cells in HNSCC or pan-cancer analysis. WASF2 promoter methylation levels and the correlation between WASF2 expression and WASF2 promoter methylation in HNSCC were evaluated using the DNMIVD database. The effect of DNA methylation inhibitor on WASF2 expression was demonstrated in the GEO database. Finally, the TISIDB database determined the relationships between WASF2 methylation, immune cell infiltration, and immune inhibitors. WASF2 was significantly downregulated in HNSCC tissues where WASF2 promoter methylation was elevated. According to the GEO database, treatment with a DNA methylation inhibitor notably restored the mRNA expression of WASF2. WASF2 expression was also a favorable indicator of human papilloma virus (HPV)-positive HNSCC. Its level of promoter methylation had detrimental effects on patient survival. In addition, patients with elevated levels of WASF2 demonstrated active G2/M regulation, TGF-β signaling, Kras signaling, fatty acid metabolism, and p53 pathways. WASF2 was positively associated with tumor-killing immune cells, while WASF2 methylation was positively related to immunosuppressive cells and immune-inhibitors. Hypermethylated WASF2 acted as a tumor suppressor of HNSCC by regulating tumor formation and immune imbalance.

Hypermethylation in the promoter region inhibits AJAP1 expression and activates the JAK/STAT pathway to promote prostate cancer cell migration and stem cell sphere formation

AJAP1 is down-regulated in multiple cancer types and plays a suppressive role in cancer progression. However, its molecular regulatory mechanism in prostate cancer has not been reported. Bioinformatics methods were employed to analyze AJAP1 expression in prostate cancer tissues and its association with TNM staging. MSP and qRT-PCR were used to quantify promoter methylation and AJAP1 expression after 5-aza-20-deoxycytidine (5-AzaC) treatment. Scratch healing assay and Transwell method were adopted to analyze the effects of aberrant AJAP1 expression, 5-AzaC and AG490 on cell migration and invasion. The levels of AJAP1 protein, EMT-related and JAK/STAT pathway-related proteins were determined by Western blot. The effects of AJAP1 aberrant expression and AG490 treatment on the sphere forming ability of prostate cancer cells were analyzed by sphere formation assay. This study confirmed the significant down-regulation of AJAP1 expression in prostate cancer tissues and cells, and its negative correlation with TNM staging. 5-AzaC treatment led to a significant reduction of AJAP1 methylation level and a significant upregulation of AJAP1 expression, indicating that the methylation level of AJAP1 promoter may affect the expression of AJAP1. Cell function experiments found that overexpression or decreased methylation of AJAP1 inhibited epithelial mesenchymal transition (EMT), migration, and invasion, while silencing or increased methylation of AJAP1 had the opposite functions. JAK2/STAT3 pathway inhibiting assay found that inhibition of JAK2/STAT3 pathway significantly reduced EMT, cell migration, and stem cell sphere formation in prostate cancer. Therefore, investigating the influence of aberrant AJAP1 expression on functions of prostate cancer cells is conducive to our in-depth understanding of the mechanism of prostate cancer genesis and development.

Investigation of Methylation Levels in OPRK1 Gene Promoter among Smokers and Opium-Addicts underwent Methadone Maintenance Treatment

1.1. Background: Previous studies reported the association of the OPRK1 gene with illicit substances, nicotine, and alcohol. The present study aimed to look at the methylation levels of OPRK1 gene promoter among smokers and addicts who underwent methadone maintenance treatment (MMT). 1.2. Methods: DNAs were extracted from the whole blood of all male samples including 30 smokers, 30 opium-addicted individuals undergoing methadone treatment, and 30 healthy people, and they were treated with a sodium bisulfite kit. The studied region included 53 CpG dinucleotides investigated by sequencing technique. 1.3. Results: Results of methylation levels in addicted individuals who underwent MMT compared with healthy people showed no difference. Also, there was no change in any CpG sites of OPRK1 gene promoter in both smokers and compared healthy controls

Association of IFITM1 Promoter Methylation with Severity of SARS-CoV-2 Infection.

During viral infections such as SARS-CoV-2, epigenetic changes within the promoter region of the immune system genes would possibly occur and have an effect on the immune system response as well as disease outcome. We aimed to evaluate and compare the methylation level of the IFITM1 gene promoter in different stages of COVID-19 disease with a healthy control group. In this cross-sectional study, 75 COVID-19 patients (25 mild, 25 severe, and 25 critical in addition to 25 age- and gender-matched healthy volunteers) have been included. DNA was extracted from the peripheral white blood cells using a commercial DNA extraction kit. PCR was performed using two types of primers designed for the methylated and unmethylated forms of the IFITM1 gene promoter. The mean age of the patient and healthy volunteer groups was 52.733 ± 13.780 and 49.120 ± 12.490, respectively. Out of a hundred participants, 52 were male. The results demonstrated that severe (p = 0.03, OR 6.729) and critical (p = 0.001, OR 11.156) patients were much more likely to show methylation of the IFITM1 gene in contrast with mild patients. Moreover, IFITM1 methylation was significantly higher in COVID-19 patients in comparison with the healthy volunteer group (p = 0.004, OR 3.17). Furthermore, IFITM1 methylation in male patients with critical status, (p = 0.01) was significantly higher than in male patients with mild status. In addition, IFITM1 methylation of male (p = 0.03) and female (p = 0.01) critical patients was considerably higher compared to males and females of volunteer group. Increased methylation of the IFITM1 gene in the severe and critical stage of COVID-19 diseases may indicate the role of SARS-CoV-2 infection in increasing methylation of this antiviral gene. This might be involved in suppressing the immune system, promoting SARS-CoV-2 replication and disease outcome.

LINC00470 represses cell autophagy and cisplatin sensitivity of glioma via suppressing PTEN expression.

Glioma is one of primary brain tumours which has the worst clinical prognoses of patients. As an alternative chemotherapeutic drug for malignant glioma, the therapeutic effect of cisplatin (CDDP) is devastatingly affected due to resistance in patients. In this study, we investigated the effect of LINC00470/PTEN on the CDDP sensitivity of glioma cells. Differentially expressed lncRNAs and the downstream regulators in glioma tissue were obtained via bioinformatics analysis. LINC00470 and PTEN mRNA expression levels were detected using qRT-PCR. IC50 values of glioma cells were examined using Cell Counting Kit-8 (CCK-8). Cell apoptosis was revealed by flow cytometry. The expression level of autophagy-related protein was detected by western blot. Intracellular autophagosome formation was detected by immunofluorescence staining, and the methylation level of PTEN promoter was detected via methylation-specific PCR (MSP). Through the above steps, we found that LINC00470 was highly expressed in glioma cells, and the survival rate of patients was reduced in the presence of high expression of LINC00470. Silenced LINC00470 promoted LC3 II expression and autophagosome formation, and facilitated cell apoptosis to inhibit resistance to CDDP. While silenced PTEN could successfully reverse the previous effects on glioma cells. Based on the above, LINC00470 repressed cell autophagy by constraining PTEN, thereby enhancing CDDP resistance of glioma cells.

Epigenetic drugs in somatostatin type 2 receptor radionuclide theranostics and radiation transcriptomics in mouse pheochromocytoma models.

Pheochromocytomas and paragangliomas (PCCs/PGLs) are catecholamine-producing tumors. In inoperable and metastatic cases, somatostatin type 2 receptor (SSTR2) expression allows for peptide receptor radionuclide therapy with [177Lu]Lu-DOTA-TATE. Insufficient receptor levels, however, limit treatment efficacy. This study evaluates whether the epigenetic drugs valproic acid (VPA) and 5-Aza-2'-deoxycytidine (DAC) modulate SSTR2 levels and sensitivity to [177Lu]Lu-DOTA-TATE in two mouse PCC models (MPC and MTT). Methods: Drug-effects on Sstr2/SSTR2 were investigated in terms of promoter methylation, mRNA and protein levels, and radiotracer binding. Radiotracer uptake was measured in subcutaneous allografts in mice using PET and SPECT imaging. Tumor growth and gene expression (RNAseq) were characterized after drug treatments. Results: DAC alone and in combination with VPA increased SSTR2 levels along with radiotracer uptake in vitro in MPC (high-SSTR2) and MTT cells (low-SSTR2). MTT but not MPC allografts responded to DAC and VPA combination with significantly elevated radiotracer uptake, although activity concentrations remained far below those in MPC tumors. In both models, combination of DAC, VPA and [177Lu]Lu-DOTA-TATE was associated with additive effects on tumor growth delay and specific transcriptional responses in gene sets involved in cancer and treatment resistance. Effects of epigenetic drugs were unrelated to CpG island methylation of the Sstr2 promoter. Conclusion: This study demonstrates that SSTR2 induction in mouse pheochromocytoma models has some therapeutic benefit that occurs via yet unknown mechanisms. Transcriptional changes in tumor allografts associated with epigenetic treatment and [177Lu]Lu-DOTA-TATE provide first insights into genetic responses of PCCs/PGLs, potentially useful for developing additional strategies to prevent tumor recurrence.

Comprehensively analyzing the genetic alterations, and identifying key genes in ovarian cancer.

Though significant improvements have been made in the treatment methods for ovarian cancer (OC), the prognosis for OC patients is still poor. Exploring hub genes associated with the development of OC and utilizing them as appropriate potential biomarkers or therapeutic targets is highly valuable. In this study, the differentially expressed genes (DEGs) were identified from an independent GSE69428 Gene Expression Omnibus (GEO) dataset between OC and control samples. The DEGs were processed to construct the protein-protein interaction (PPI) network using STRING. Later, hub genes were identified through Cytohubba analysis of the Cytoscape. Expression and survival profiling of the hub genes were validated using GEPIA, OncoDB, and GENT2. For exploring promoter methylation levels and genetic alterations in hub genes, MEXPRESS and cBioPortal were utilized, respectively. Moreover, DAVID, HPA, TIMER, CancerSEA, ENCORI, DrugBank, and GSCAlite were used for gene enrichment analysis, subcellular localization analysis, immune cell infiltration analysis, exploring correlations between hub genes and different diverse states, lncRNA-miRNA-mRNA co-regulatory network analysis, predicting hub gene-associated drugs, and conducting drug sensitivity analysis, respectively. In total, 8947 DEGs were found between OC and normal samples in GSE69428. After STRING and Cytohubba analysis, 4 hub genes including TTK (TTK Protein Kinase), (BUB1 mitotic checkpoint serine/threonine kinase B) BUB1B, (Nucleolar and spindle-associated protein 1) NUSAP1, and (ZW10 interacting kinetochore protein) ZWINT were selected as the hub genes. Further, it was validated that these 4 hub genes were significantly up-regulated in OC samples compared to normal controls, but overexpression of these genes was not associated with overall survival (OS). However, genetic alterations in those genes were found to be linked with OS and disease-free (DFS) survival. Moreover, this study also revealed some novel links between TTK, BUB1B, NUSAP1, and ZWINT overexpression and promoter methylation status, immune cell infiltration, miRNAs, gene enrichment terms, and various chemotherapeutic drugs. Four hub genes, including TTK, BUB1B, NUSAP1, and ZWINT, were revealed as tumor-promotive factors in OC, having the potential to be utilized as novel biomarkers and therapeutic targets for OC management.

TUSC3 Methylation in Peripheral Blood Cells as a Biomarker for Diagnosis of Colorectal Cancer.

Several case-control studies have suggested that global and loci-specific deoxyribonucleic acid (DNA) methylation in peripheral blood mononuclear cells (PBMCs) of DNA might be potential biomarkers of cancer diagnosis and prognosis. In this study, for the first time, we intended to assess the diagnostic power of the methylation level of tumor suppressor candidate 3 (TUSC3) gene promoter in patients with colorectal cancer (CRC). In the current study, we quantitatively assessed the promoter methylation level of TUSC3 in PBMCs of 70 CRC cases and 75 non-cancerous subjects via methylation quantification of endonuclease-resistant DNA (MethyQESD) method. The methylation level of the TUSC3 was meaningfully higher in CRC cases than in non-CRC subjects (43.55 ± 21.80% vs. 16.07 ± 13.63%, respectively; P < 0.001). The sensitivity and specificity of this gene for the detection of CRC were 88.6% and 76.0%, respectively. The receiver operating characteristic (ROC) curve examination discovered an area under the curve (AUC) of 0.880, representing a very high accuracy of the TUSC3 methylation marker in distinguishing CRC subjects from healthy individuals. However, there was no substantial diversity in methylation level between various CRC stages (P: 0.088). For CRC screening, PBMCs are a reliable source for DNA methylation analysis and TUSC3 promoter methylation can be utilized as a hopeful biomarker for early and non-invasive diagnosis of CRC.

UHRF1-mediated ferroptosis promotes pulmonary fibrosis via epigenetic repression of GPX4 and FSP1 genes

Pulmonary fibrosis (PF), as an end-stage clinical phenotype of interstitial lung diseases (ILDs), is frequently initiated after alveolar injury, in which ferroptosis has been identified as a critical event aggravating the pathophysiological progression of this disease. Here in, a comprehensive analysis of two mouse models of pulmonary fibrosis developed in our lab demonstrated that lung damage-induced ferroptosis of alveolar epithelial Type2 cells (AEC2) significantly accumulates during the development of pulmonary fibrosis while ferroptosis suppressor genes GPX4 and FSP1 are dramatically inactivated. Mechanistically, upregulation of de novo methylation regulator Uhrf1 sensitively elevates CpG site methylation levels in promoters of both GPX4 and FSP1 genes and induces the epigenetic repression of both genes, subsequently leading to ferroptosis in chemically interfered AEC2 cells. Meanwhile, specific inhibition of UHRF1 highly arrests the ferroptosis formation and blocks the progression of pulmonary fibrosis in both of our research models. This study first, to our knowledge, identified the involvement of Uhrf1 in mediating the ferroptosis of chemically injured AEC2s via de novo promoter-specific methylation of both GPX4 and FSP1 genes, which consequently accelerates the process of pulmonary fibrosis. The above findings also strongly suggested Uhrf1 as a novel potential target in the treatment of pulmonary fibrosis.

Multi-omics pan-cancer study of cuproptosis core gene FDX1 and its role in kidney renal clear cell carcinoma.

The mechanism of copper-induced cellular death was newly discovered and termed cuproptosis. Inducing cuproptosis in cancer cells is well anticipated for its curative potential in treating tumor diseases. However, ferredoxin 1 (FDX1), the core regulatory gene in cuproptosis, is rarely studied, and the regulation of FDX1 in tumor biology remains obscure. A comprehensive pan-cancer analysis of FDX1 is needed. Thirty-three types of tumors were included with paired normal tissues in The Cancer Genome Atlas (TCGA) and the Genotype-Tissue Expression (GTEx) datasets. The interaction between transcription, protein, phosphorylation, and promoter methylation levels was analyzed. Survival, immune infiltration, single-cell FDX1 expression, FDX1-related tumor mutational burden (TMB), microsatellite instability (MSI), stemness, tumor immune dysfunction and exclusion (TIDE), and immunotherapy-related analyses were performed. FDX1 protein expression was assessed by kidney renal clear cell carcinoma (KIRC) tissue microarray immunohistochemistry. The function of FDX1 in KIRC was further explored by experiments in 786-O cell lines in vitro. FDX1 is highly expressed in 15 tumor types and lowly expressed in 11 tumor types. The corresponding changes in protein expression, phosphorylation, and promoter methylation level of FDX1 have been described in several tumors. Survival analysis showed that FDX1 was related to favorable or poor overall survival in eight tumors and progression-free survival in nine tumors. Immune infiltration and single-cell analysis indicated the indispensable role of FDX1 expression in macrophages and monocytes. Multiple established immunotherapy cohorts suggested that FDX1 may be a potential predictor of treatment effects for tumor patients. Tissue microarray analysis showed decreased FDX1 expression in KIRC patients' tumor tissues. Knockdown of FDX1 resulted in the downregulation of cuproptosis in kidney renal clear tumor cells. Mechanistically, the FDX1-associated gene expression signature in KIRC is related to the enrichment of genes involved in the tricarboxylic acid (TCA) cycle, NOTCH pathway, etc. Several NOTCH pathway genes were differentially expressed in the high- and low-FDX1 groups in KIRC. Our analysis showed that the central regulatory gene of cuproptosis, FDX1, has differential expression and modification levels in various tumors, which is associated with cellular function, immune modulation, and disease prognosis. Thus, FDX1-dependent cuproptosis may serve as a brand-new target in future therapeutic approaches against tumors.

USPs in Pancreatic Ductal Adenocarcinoma: A Comprehensive Bioinformatic Analysis of Expression, Prognostic Significance, and Immune Infiltration.

Pancreatic ductal adenocarcinoma (PDAC), as an intractable malignancy, still causes an extremely high mortality worldwide. The ubiquitin-specific protease (USP) family constitutes the major part of deubiquitinating enzymes (DUBs) which has been reported to be involved in initiation and progression of various malignancies via the function of deubiquitination. However, the biological function and clinical values of USPs in PDAC have not been comprehensively elucidated. In this study, Gene Expression Profiling Interactive Analysis (GEPIA), Gene Expression Omnibus (GEO) datasets, UALCAN database, and the Human Protein Atlas (HPA) online tool were used to analyze the expression level and the relationship between USP expression and clinicopathological features in PDAC. Survival module of HPA and Kaplan-Meier plotter (KMP) databases was recruited to explore the prognostic value of USPs. Tumor Immune Estimation Resource (TIMER) online tool and KMP databases were utilized to elucidate tumor immune infiltration and immune-related survival of USPs. CBioPortal online tool was used to identify the gene mutation level of USPs in PDAC. Both cBioPortal and LinkedOmics were used to confirm the potential biological functions of USPs in PDAC. Our study showed that USP10, USP14, USP18, USP32, USP33, and USP39 (termed as six-USPs) expressions were significantly elevated in tumor tissues. The high expression of the four USPs (USP10, USP14, USP18, and USP39) indicated a poor prognosis. A significant relationship was indicated between the expression of six-USPs and clinicopathological features. Also, the expression of six-USPs was related to promoter methylation level. Moreover, more than 40% genetic alterations and mutations were discovered in six-USPs. Furthermore, the six-USP expression was correlated with immune infiltration and immune-related prognosis. The functional analysis found that the six-USPs were involved in various biological processes and signaling pathways, such as nucleocytoplasmic transport, choline metabolism in cancer, cell cycle, ErbB signaling pathway, RIG-I-like receptor signaling pathway, TGF-β signaling pathway, and TNF signaling pathway. In conclusion, the results showed that six-USPs are potential prognostic biomarkers and can be recruited as possible therapeutic targets of PDAC.

Systematic analysis of virus nucleic acid sensor DDX58 in malignant tumor.

In December 2019, a novel epidemic of coronavirus pneumonia (COVID-19) was reported，and population-based studies had shown that cancer was a risk factor for death from COVID-19 infection. However, the molecular mechanism between COVID-19 and cancer remains indistinct. In this paper, we analyzed the nucleic acid sensor (DDX58) of SARS-CoV-2 virus, which is a significant gene related to virus infection. For purpose of clarifying the characteristics of DDX58 expression in malignant tumors, this study began to systematically analyze the DDX58 expression profile in the entire cancer type spectrum. Using TCGA pan-cancer database and related data resources, we analyzed the expression, survival analysis, methylation expression, mutation status, microsatellite instability (MSI), immune related microenvironment, gene related network, function and drug sensitivity of DDX58. The expression level of DDX58 mRNA in most cancers was higher than the expression level in normal tissues. Through TIMER algorithm mining, we found that DDX58 expression was closely related to various levels of immune infiltration in pan-cancer. The promoter methylation level of DDX58 was significantly increased in multiple cancers. In addition, abnormal expression of DDX58 was related to MSI and TMB in multiple cancers, and the most common type of genomic mutation was "mutation." In the protein-protein interaction (PPI) network, we found that type I interferon, phagocytosis, ubiquitinase, and tumor pathways were significantly enriched. Finally, according to the expression of DDX58 indicated potential sensitive drugs such as Cediranib, VE-821, Itraconazole, JNJ-42756493, IWR-1, and Linsitinib. In conclusion, we had gained new insights into how DDX58 might contribute to tumor development, and DDX58 could be used as an immune-related biomarker and as a potential immunotherapeutic target for COVID-19 infected cancer patients.

Involvement of brain-derived neurotrophic factor methylation in the prefrontal cortex and hippocampus induced by chronic unpredictable mild stress in male mice.

Early life stress alters brain-derived neurotrophic factor (BDNF) promoter IV methylation and BDNF expression, which is closely related to the pathophysiological process of depression. However, the role of abnormal methylation of BDNF induced by stress during adolescence due to depression has not yet been clarified. In this study, adolescent mice were exposed to chronic unpredictable mild stress (CUMS). Depression-like behaviors, BDNF promoter IV methylation, expression of DNA methyltransferases (DNMTs), demethylation machinery enzymes, BDNF protein levels, and neuronal development in the prefrontal cortex (PFC) and hippocampus (HIP) were assessed in adolescent and adult mice. The DNMT inhibitor, 5-Aza-2-deoxycytidine (5-AzaD), was used as an intervention. Stress in adolescence induces behavioral dysfunction, elevated methylation levels of BDNF promoter IV, changes in the expression of DNMT, and demethylation machinery enzymes in adolescent and adult mice. Additionally, the stress in adolescence induced lower levels of BDNF and abnormal hippocampal doublecortin (DCX) expression in adolescent and adult mice. However, DNMT inhibitor treatment in adolescent-stressed mice relieved the abnormal behaviors, normalized the methylation level of BDNF promoter IV, BDNF protein expression, expression of DNMTs, and demethylation machinery enzymes, and improved the neuronal development of adult mice. These results suggest that stress in adolescence induces short- and long-term hypermethylation of BDNF promoter IV, which is regulated by DNMTs, and leads to the development of depression.

Pan-cancer analysis reveals NAA50 as a cancer prognosis and immune infiltration-related biomarker.

Background: N-Alpha-Acetyltransferase 50 (NAA50) has acetyltransferase activity and is important for chromosome segregation. However, the function and mechanism of NAA50 expression in cancer development was still unclear. Here, we systematically researched the function and mechanism of NAA50 in pan-cancer, and further verified the results of NAA50 in lung adenocarcinoma (LUAD). Methods: In this study, using the online databases TIMER2.0, SangerBox3.0, HPA, UCSC, GEPIA, cBioPortal, UALCAN, TISIDB, CancerSEA and LinkedOmics, we focused on the relevance between NAA50 and oncogenesis, progression, methylation, immune infiltration, function and prognosis. In addition, the proliferation of cells was detected by CCK-8 and Edu assay. Finally, we analyzed the relationship between the expression of NAA50 and cell cycle related proteins. Results: Pan-cancer analysis indicated that NAA50 was overexpressed in most cancers. And there was a significant correlation between NAA50 expression and the prognosis of cancer patients. In the meantime, NAA50 gene changes occur in a variety of tumors. Compared with normal tissues, the methylation level of NAA50 promoter increased in most cancer tissues. In addition, the results exhibited that in most cancers, NAA50 was significantly positively correlated with bone myeloid-derived suppressor cell (MDSC) infiltration and negatively correlated with T cell NK infiltration. Moreover, functional enrichment indicated that NAA50 regulates cell cycle and proliferation in LUAD. In vitro experiments testified that knockout of NAA50 could significantly inhibit the proliferation of LUAD. Conclusion: NAA50 may be a potential biomarker and oncogene of pan-cancer, especially LUAD, which may promote the occurrence and development of tumors through different mechanisms. Furthermore, NAA50 was bound up with to immune cell infiltration in pan-cancer, meaning NAA50 may be an important therapeutic target for human cancers.

Hsa_circ_0000119 promoted ovarian cancer development via enhancing the methylation of CDH13 by sponging miR-142-5p.

Ovarian cancer is the leading cause of gynecological cancer-related death in women, and is difficult to treat. The aim of our study is to explore the role and action mechanism of hsa_circ_0000119 in ovarian cancer, thus to analyze whether the circular RNA is a potential target for the treatment of the disease. In this present study, our data shows that hsa_circ_0000119 and DNA methyltransferase 1 (DNMT1) was increased, while miR-142-5p was decreased in ovarian cancer. Overexpression of hsa_circ_0000119 promoted tumor growth, while silencing of hsa_circ_0000119 resulted in an opposite effects. Decreasing of hsa_circ_0000119 also notably inhibited the proliferation, migration, and invasion of the ovarian cancer cells. Moreover, the data proves that hsa_circ_0000119 negatively regulated miR-142-5p and cadherin 13 (CDH13) expression, but positively regulated DNMT1 expression. miR-142-5p could interact with hsa_circ_0000119 and DNMT1 3'-UTR. Silencing of DNMT1 could reverse the inhibition of hsa_circ_0000119 to miR-142-5p and CDH13 expression. Importantly, higher level of CDH13 promoter methylation existed in the ovarian tumors than that in matched normal tissues. DNA methyltransferase inhibitor could increase the expression of CDH13 in ovarian cancer cells. In addition, our results also prove that increasing of CDH13 or miR-142-5p effectively reversed the inhibition of hsa _circ_0000119 to the cell malignant phenotypes. Overall, our data demonstrate that hsa_circ_0000119 facilitated ovarian cancer development through increasing CDH13 expression via promoting DNMT1 expression by sponging miR-142-5p. Our data demonstrate the potential role of hsa_circ_0000119 in the treatment of ovarian cancer.

DNMT3B attenuated the inhibition of TET3 on epithelial-mesenchymal transition in TGF-β1-induced ovarian cancer by methylating the TET3 promoter.

This study intends to investigate the effects of DNA methyltransferase 3B (DNMT3B) and ten-eleven translocation 3 (TET3) on transforming growth factor-β1 (TGF-β1)-induced epithelial-mesenchymal transition (EMT) in ovarian cancer (OV) cells. According to the specific experiments, the cells were treated with TGF-β1 for 48h, and then the expressions of EMT-related proteins (E-cadherin, Vimentin and Snail), TET3 and DNMT3B were quantified by Western blot and quantitative reverse transcription polymerase chain reaction (qRT-PCR). Methprimer, methylation-specific PCR (MSP), bisulfite sequencing PCR (BSP) and chromatin immunoprecipitation (ChIP) were used to determine the regulation of DNMT3B on TET3 promoter. The impacts of DNMT3B and TET3 upon the EMT-related proteins and OV cell migration and invasion abilities were evaluated through the rescue experiments and the loss- and gain-of-function experiments. In line with the results, TGF-β1 down-regulated TET3 and E-cadherin levels, up-regulated Vimentin and Snail levels, promoted migratory and invasive abilities, and increased methylation level of TET3 promoter in OV cells, which however were reversed by shDNMT3B. The binding of DNMT3B to TET3 promoter facilitated the methylation of TET3 promoter. Overexpressed TET3 inhibited the migratory and invasive abilities and EMT of OV cells, whereas shTET3 did the opposite. ShTET3 also offset the regulatory effects of shDNMT3B on EMT, migration and invasion of OV cells. To conclude, DNMT3B mitigates the suppression of TET3 on TGF-β1-induced EMT in OV cells by methylating the promoter region of TET3.

Association Between Promoter Methylation of Vitamin D Metabolic Pathway Genes and Tuberculosis and Diabetes Comorbidity in a Chinese Han Population: A Case-Control Study.

Deficiency vitamin D and hyperglycemia could be related to weakened innate immune response and aggravate the progression of tuberculosis (TB). This study hypothesized that DNA promoter methylation of the pivotal genes in the vitamin D metabolic pathway might be related to diabetes and tuberculosis co-morbidity (TB-DM) susceptibility. A total of 50 TB-DM and 50 healthy subjects (HS) were included in the present study. Targeted bisulfite sequencing was applied to detect the methylation of the promoter regions of candidate genes in the vitamin D metabolic pathway (CYP24A1, CYP27B1, CYP2R1, DHCR7, and VDR) in whole blood. The overall methylation level of candidate genes in this study was lower in patients with TB-DM than HS, except for CYP2R1. The results of the ROC demonstrated the potential of CYP24A1, CYP27B1, DHCR7, and VDR promoter methylation as a biomarker for diagnosing TB-DM, with all the AUC above 0.7. In subgroup analysis, we found that lower circulating vitamin D is related to a low level of CYP24A1, CYP27B1, and DHCR7 promoter methylation in patients with TB-DM. With decreasing methylation level, risk of TB-DM was significantly increased (odds ratio, 95% CI 0.343, 0.144-0.821 for CYP24A1; 0.461, 0.275-0.773 for CYP27B1; 0.09, 0.015-0.530 for DHCR7; 0.006, 0.0003-0.115 for VDR). Besides, our results revealed that there was a significant correlation between DNA promoter methylation of selected genes in the vitamin D metabolic pathway and platelet indices in TB-DM. However, there was no correlation between DNA methylation of the four genes and fasting glucose and HbA1c. Our results could suggest that the selected genes in the vitamin D metabolic pathway may be involved in the pathological process of TB-DM, but independent of the process of hyperglycemia to impaired immune responses to Mtb.

Identification and validation of DNA methylation markers to predict axillary lymph node metastasis of breast cancer.

Axillary lymph node metastasis (ALNM) is one of the most important prognostic factors for breast cancer patients, and DNA methylation is involved in ALNM of breast cancer. However, the methylation profile of breast cancer ALNM remains unknown. Breast cancer tissues were collected from patients with and without ALNM. We investigated the genome-wide DNA methylation profile in breast cancer with and without ALNM using reduced representation bisulfite sequencing (RRBS). Then, differentially methylated regions (DMRs) were verified by targeted bisulfite sequencing. A total of 21491 DMRs were identified between the lymph node positive group and negative group. Compared to the LN-negative breast cancer, LN-positive breast cancer had 10,920 hypermethylated DMRs and 10,571 hypomethylated DMRs. Then, 10 DMRs in the gene promoter region were detected by targeted bisulfite sequencing, these gene included HOXA5, PTOV1-AS1, RHOF, PAX6, GSTP1, RASGRF2, AKR1B1, BNIP3, CRMP1, ING5. Compared with negative lymph node, the promoter methylation levels of RASGRF2, AKR1B1 and CRMP1 increased in positive lymph node, while the promoter methylation level of RHOF decreased in positive lymph node. In addition, Cancer Genome Atlas (TCGA) data showed that RASGRF2, AKR1B1 and CRMP1 were low expressed in breast Cancer tissues, while RHOF was high expressed in breast Cancer tissues. Furthermore, in addition to highly methylated AKR1B1, RASGRF2 and CRMP1 gene promoters, BNIP3, GSTP1, HOXA5 and PAX6 gene promoters were also methylated in ER-positive and HER2-negative breast cancer with ALNM. When compared to negative lymph node breast cancer, the positive lymph node breast cancer has a differential methylation status. Promoter methylation of RASGRF2, AKR1B1, CRMP1 and RHOF in lymph node positive breast cancer tissues was significantly different from that in lymph node negative breast cancer tissues. AKR1B1, RASGRF2, CRMP1, BNIP3, GSTP1, HOXA5 and PAX6 genes were methylated in ER-positive and HER2-negative breast cancer with ALNM. The study provides an important biological base for understanding breast cancer with ALNM and developing therapeutic targets for breast cancer with ALNM.