Clinical Performance of Friend Leukemia Virus Integration 1 Methylation as a Biomarker for Colorectal Carcinoma

Objectives Accumulating evidence supports the reliability of molecular biomarkers and cancer, the current situation is lacking. This present study was aimed to investigate clinical performance of friend leukemia virus integration 1 (FLI1) methylation as a biomarker for colorectal cancer (CRC). Methods The datasets of UALCAN, GEPIA, cBioPortal, STRING, TIMER2.0, GEO, and KOBAS were utilized in this present study. In order to testify the methylation function of FLI1 in CRC, we studied 6 CRC cell lines and 20 pairs of tumor tissues. The transcriptional levels of FLI1 were tested by reverse transcription PCR quantitatively and western blot. Cell viability, transwell assays and plate clone assay were utilized to assess the cell function. Results FLI1 was up-regulated in the GBM, KIRC, LAML, etc. Meanwhile, it was down-regulated in BLCA, BRCA, CESC, etc (p<0.05). The CPTAC dataset showed higher total protein in the primary tissues of KIRC, lower protein in the BRCA, OV, LUAD, UCEC than normal tissues(p<0.05). Highly expressed FLI1 was linked to poor prognosis of overall survival (OS) in LGG, UVM (p<0.05). Low expression of FLI1 was associated with short OS in KIRC, LUAD, SKCM (p<0.05). Compared with normal tissues, the methylation level of FLI1 was increased in BRCA, CESC, CHOL, COAD, etc (p<0.05). In the contrary, it was decreased in KIRP, PCPG obviously (p<0.05). Moreover, it showed a reduced phosphorylation for selected probes (BRCA: NP_002008.2:S39, NP_002008.2:S3241; OV: NP_002008.2:S39; LUAD: NP_002008.2:S79, NP_002008.2:S3241; all p<0.05). Meanwhile, it showed an enhanced phosphorylation level of FLI1 in KIRC for selected probes (NP_002008.2:S241, p<0.05). Besides, a statistical negative correlation was found between FLI1 and Treg cells, neutrophils, monocytes, macrophages, NK cells, cancer-associated fibroblasts, and common immune checkpoint gene levels (p<0.05). Besides, in vivo experience showed that 5-Aza-2’-deoxycytidine diminished FLI1 methylation level and restored transcriptional levels (p>0.05) in CRC. In vitro experiments demonstrated that the proliferation, colony formation, invasion and migration of CRC cells were inhibited by FLI1 overexpression through FMNL1 (p<0.05). Conclusions This present study offers a comprehensive understanding of FLI1 in carcinomas with oncogenesis and immunotherapeutic implications. Moreover, FLI1 reduced the proliferation, colony formation, invasion and migration of CRC by FMNL1.

Iodine-125 seed represses the growth and facilitates the apoptosis of colorectal cancer cells by suppressing the methylation of miR-615 promoter

Background Colorectal cancer (CRC) represents a common malignancy in gastrointestinal tract. Iodine-125 (125I) seed implantation is an emerging treatment technology for unresectable tumors. This study investigated the mechanism of 125I seed in the function of CRC cells. Methods The CRC cells were irradiated with different doses of 125I seed (0.4, 0.6 and 0.8 mCi). miR-615 expression in CRC tissues and adjacent tissues was detected by RT-qPCR. miR-615 expression was intervened with miR-615 mimic or miR-615 inhibitor, and then the CRC cells were treated with 5-AZA (methylation inhibitor). The CRC cell growth, invasion and apoptosis were measured. The methylation level of miR-615 promoter region was detected. The xenograft tumor model irradiated by 125I seed was established in nude mice. The methylation of miR-615, Ki67 expression and CRC cell apoptosis were detected. Results 125I seed irradiation repressed the growth and facilitated apoptosis of CRC cells in a dose-dependent manner. Compared with adjacent tissues, miR-615 expression in CRC tissues was downregulated and miR-615 was poorly expressed in CRC cells. Overexpression of miR-615 suppressed the growth of CRC cells. 125I seed-irradiated CRC cells showed increased miR-615 expression, reduced growth rate and enhanced apoptosis. The methylation level of miR-615 promoter region in CRC cells was decreased after 125I seed treatment. In vivo experiments confirmed that 125I seed-irradiated xenograft tumors showed reduced methylation of the miR-615 promoter and increased miR-615 expression, as well as decreased Ki67 expression and enhanced apoptosis. The target genes of miR-615 and its regulatory downstream pathway were further predicted by bioinformatics analysis. Conclusions 125I seed repressed the growth and facilitated the apoptosis of CRC cells by suppressing the methylation of the miR-615 promoter and thus activating miR-615 expression. The possible mechanism was that miR-615-5p targeted MAPK13, thus affecting the MAPK pathway and the progression of CRC.

Association of Aberrant DNA Methylation Level in the CD4 and JAK-STAT-Pathway-Related Genes with Mastitis Indicator Traits in Chinese Holstein Dairy Cattle

The present study was designed to evaluate the gene expression and DNA methylation level in the promoter region of the CD4 and the JAK-STAT-pathway-related genes. A total of 24 samples were deployed in the gene expression and 118 samples were used in the DNA methylation study. Student’s t-tests were used to analyze the gene expression and DNA methylation. The evaluation of DNA methylation in promoter regions of JAK2 and STAT5A revealed hypo-methylation levels of CpG sites and higher gene expression in cows diagnosed with mastitis as compared to the healthy control, and vice versa in those with CD4. DNA methylation was negatively correlated with gene expression in JAK2, STAT5A, and CD4 genes. Six, two, and four active transcription factors were identified on the CpG sites in the promoter regions of JAK2, STAT5A, and CD4 genes, respectively. Regarding correlation analysis, the DNA methylation levels of CD4 showed significantly higher positive correlations with somatic cell counts (p < 0.05). Findings of the current study inferred that aberrant DNA methylation in the CpG sites at the 1 kb promoter region in JAK2, STAT5A, and CD4 genes due to mastitis in cows can be used as potential epigenetic markers to estimate bovine mastitis susceptibility in dairy cattle.

Long-term Changes In Aberrant DNA Methylation And Gastritis After Helicobacter Pylori Eradication Focused On Metachronous Gastric Cancer

BackgroundA persistently high methylation level in gastric mucosa after Helicobacter pylori (H. pylori) eradication is presumed to be a risk for metachronous gastric cancer (MGC); however, long-term changes in aberrant DNA methylation and histological gastritis have been unclear. Our aim was to examine changes in DNA methylation and histological gastritis according to the occurrence of MGC.MethodsSubjects were classified into 3 groups: 25 patients in whom metachronous gastric cancers occurred after the initial endoscopic resection (ER) for early gastric cancer and H. pylori eradication (MGC group), 17 patients in whom MGC did not occur for more than 5 years after initial ER and H. pylori eradication (non-MGC group) and 29 patients without a history of gastric cancer who succeeded in eradication more than 5 years ago (HP group). Aberrance of DNA methylation in 3 genes (miR-124a-3, EMX1, NKX6-1) and histological score of atrophy and intestinal metaplasia (IM) were evaluated using biopsy samples before and more than a mean of 5 years after H. pylori eradication.ResultsThe methylation level of miR-124a-3 in the HP group and non-MGC group and that of EMX1 in the HP group significantly decreased in the long term after eradication. In the MGC group, H. pylori eradication did not improve aberrant methylation, and the Z-score significantly increased. There were significant positive correlations between methylation levels in miR-124a-3 and EMX1 and histological findings after eradication.ConclusionsA persistently high methylation level after H. pylori eradication reflected precancerous mucosal conditions and led to long-term MGC.

Maternal Glucose and LDL-Cholesterol Levels Are Related to Placental Leptin Gene Methylation, and, Together With Nutritional Factors, Largely Explain a Higher Methylation Level Among Ethnic South Asians

BackgroundLeptin, mainly secreted by fat cells, plays a core role in the regulation of appetite and body weight, and has been proposed as a mediator of metabolic programming. During pregnancy leptin is also secreted by the placenta, as well as being a key regulatory cytokine for the development, homeostatic regulation and nutrient transport within the placenta. South Asians have a high burden of type 2 diabetes, partly attributed to a “thin-fat-phenotype”.ObjectiveOur aim was to investigate how maternal ethnicity, adiposity and glucose- and lipid/cholesterol levels in pregnancy are related to placental leptin gene (LEP) DNA methylation.MethodsWe performed DNA methylation analyses of 13 placental LEP CpG sites in 40 ethnic Europeans and 40 ethnic South Asians participating in the STORK-Groruddalen cohort.ResultsSouth Asian ethnicity and gestational diabetes (GDM) were associated with higher placental LEP methylation. The largest ethnic difference was found for CpG11 [5.8% (95% CI: 2.4, 9.2), p&lt;0.001], and the strongest associations with GDM was seen for CpG5 [5.2% (1.4, 9.0), p=0.008]. Higher maternal LDL-cholesterol was associated with lower placental LEP methylation, in particular for CpG11 [-3.6% (-5.5, -1.4) per one mmol/L increase in LDL, p&lt;0.001]. After adjustments, including for nutritional factors involved in the one-carbon-metabolism cycle (vitamin D, B12 and folate levels), ethnic differences in placental LEP methylation were strongly attenuated, while associations with glucose and LDL-cholesterol persisted.ConclusionsMaternal glucose and lipid metabolism is related to placental LEP methylation, whilst metabolic and nutritional factors largely explain a higher methylation level among ethnic South Asians.

Methylation of SDC2/TFPI2 and Its Diagnostic Value in Colorectal Tumorous Lesions

Background:SDC2 methylation is a feasible biomarker for colorectal cancer detection. Its specificity for colorectal cancer is higher than 90%, but the sensitivity is normally lower than 90%. This study aims to improve the sensitivity of SDC2 detection through finding a high positive target from the false-negative samples of SDC2 detection based on analysis of the bowel subsite difference in methylation.Methods: Hypermethylated TFPI2 was identified in SDC2 hypomethylated colorectal cancer samples retrieved from TCGA database with the methylation level lower than 0.2. The methylation-specific PCR assay was developed and then evaluated using tissue samples (184 cancer and 54 healthy control samples) and stool samples (289 cancer, 190 adenoma, and 217 healthy control samples).Results:TFPI2 was hypermethylated in most SDC2 hypomethylated colorectal cancer samples. When the SDC2/TFPI2-combined PCR assay was performed in stool specimens, the AUC value of cancer vs. control was 0.98, with the specificity of 96.40% and sensitivity of 96.60%, and the AUC value of adenoma vs. control was 0.87, with the specificity of 95.70% and the sensitivity of 80.00%. The improvement in sensitivity was the most momentous in the left colon. As the detection index, the Ct value was better in improving the sensitivity of detection than the methylation level based on the 2−ΔΔCt value.Conclusion:TFPI2 can improve the sensitivity of SDC2 methylation–specific detection of colorectal tumorous lesions while maintaining high specificity, in particular reducing the missed detection of left colon cancer and adenoma.

Interactive Regulations of Dynamic Methylation and Transcriptional Responses to Recurring Environmental Stresses During Biological Invasions

Deoxyribonucleic acid methylation and gene transcription have been proved as two underlying mechanisms involved in rapid plastic response to environmental stresses. However, it remains elusive on how DNA methylation regulates gene transcription under acute and recurring environmental challenges to form the stress memory, further contributing to invasion success during range expansions. Using a model invasive species Ciona robusta, we investigated the regulatory roles of DNA methylation on gene transcription and their contribution to the formation of stress memory at 30 genes under acute and recurring osmotic challenges simulated during the invasion process. We found the bimodal distribution of methylation level for the 68 mCpGs identified across all the genes after challenges, but only five sites were significantly correlated with the expression of their corresponding genes. These genes participated in the biological processes of Ca2+ transport and metabolism of lipid and proline. At the DNA methylation level, we found two early-responding and four tardy-responding sites of stress memory and these sites were functionally related to genes involved in the biosynthesis of proline, metabolism of lipid, and transport of taurine and Ca2+. At the transcriptional level, three tardy-responding and five early-responding memory genes were involved in the transport of ions, regulation of water channels, biosynthesis of taurine, and metabolism of lipid. Altogether, the findings here suggest that DNA methylation and gene transcription should work in concert to facilitate the formation of stress memory, thus further improving the performance of invaders under recurring environmental challenges during biological invasions.

TMEM130 regulates cell migration through DNA methylation in nasopharyngeal carcinoma

BACKGROUND: Nasopharyngeal carcinoma (NPC), the common malignant head and neck cancer, is highly prevalent in southern China. The molecular mechanism underlying NPC tumorigenesis is unclear. We used 5-Aza-CdR, a DNA methyltransferase inhibitor, to treat NPC cell lines and discovered that the expression of TMEM130 changed significantly compared with the untreatment cells. This study aimed to identify the relationship between the DNA methylation status of TMEM130 and NPC, and to explore the function of TMEM130 in NPC cell migration. METHODS: qRT-PCR was performed to investigate the transcriptional expression of TMEM130 in NPC. Bisulfite sequencing PCR and 5-Aza-CdR treatment were used to detect the methylation level of the TMEM130 promoter. Gene Expression Omnibus (GEO) datasets were obtained to identifiy the methylation status and mRNA expression of TMEM130 in NPC and normal control tissues. Transwell and western blot analyses were used to detect cell migration ability after transfection of TMEM130/NC plasmids in NPC cells. RESULTS: The transcriptional expression of TMEM130 was decreased in NPC cell lines compared with in the NP69 cell line. TMEM130 promoter was significantly hyper methylated in three NPC cell lines (C666, CNE, and HONE) but hypo methylated in NP69 cells. The methylation level was higher in NPC than normal control tissues. Additionally, treatment of NPC cells with 5-Aza-CdR increased the TMEM130 mRNA expression level. Overexpression of TMEM130 in NPC cell lines suppressed cell migration ability and affected some epithelial-mesenchymal transition-associated gene expression. CONCLUSIONS: This study is the first to investigate the expression and function of TMEM130 in NPC. It was found that TMEM130 hyper methylation might contribute to NPC migration and this gene might act as a tumor suppressor gene. TMEM130 is a promising biomarker for NPC diagnosis.

The Impact of the Serotonin Transporter Gene Promoter DNA Methylation on Anorexia Nervosa：A Preliminary Longitudinal Study

BackgroudThe serotonin system has been reported to be involved in the pathogenesis of anorexia nervosa (AN). The promoter region of serotonin transporter gene (SLC6A4), also called 5-HTTLPR, responsible for serotonin reuptake, has received much attention, especially 5-HTTLPR methylation, which was associated with abnormal eating behaviors. The study was aimed to explore the association between 5-HTTLPR methylation and AN, as well as its predictive effect on therapeutic response.Methods91 AN patients and 87 healthy controls (HCs) were recruited. Only 30 patients completed the 12-week follow-up. 5-HTTLPR methylation levels and clinical symptoms were assessed at baseline for all participants, at the fourth and the twelfth week for follow-up patients. 5-HTTLPR methylation was measured by MassArray methods and clinical symptoms were mainly evaluated by the EDE-Q6.0 scale. ResultsAN patients had higher methylation at CpG 11, CpG 24.25, CpG 31.32 and CpG_mean than healthy controls (P=0.039, 0.042, 0.018, 0.001). Furthermore, it was the binge/purging subtype that revealed significant hypermethylation at CpG4 and CpG_mean (P=0.027, 0.031). However, it failed to discover significant differences between AN-R and AN-BP groups at all units. Besides, CpG 11 methylation level was positively correlated with EDEQ-6.0 total score in patients (r=0.226, P=0.047). The methylation level of CpG11, CpG26.27.28, CpG31.32, CpG33.34.35.36 and CpG_mean decreased after treatment, but not significantly. 5-HTTLPR methylation was not significantly different between the two groups after treatment.ConclusionsOur study provided preliminary evidence that 5-HTTLPR methylation played vital roles in AN pathogenesis. The characteristic of 5-HTTLPR among untreated AN patients was hypermethylation. However, whether it is the biomarker to distinguish the subtypes and therapeutic response of AN needs further investigation.