Journal: Scientific Reports

Article Title: Alcohol and nicotine codependence-associated DNA methylation changes in promoter regions of addiction-related genes

doi: 10.1038/srep41816

Figure Lengend Snippet: The top panel contains the -log P values for the association between DNA methylation of three HTR2B promoter CpGs and alcohol-nicotine codependence. The other panels indicate the presence of coding exons (boxes in dark red) and noncoding introns (grey lines) of HTR2B (the second panel), DNase hypersensitivity cluster sites (the third panel), the location of a H3K27ac histone modification (the fourth panel), the conservative area (the fifth panel), the percentage of G (guanine) and C (cytosine) bases (the 6th panel), and the location of CpGs in transcription factor binding sites (TFBSs) (the seventh panel). The function of the most significant CpG (i.e., cg27531267 in the promoter region of HTR2B ) is annotated by a brown line across all panels.

Article Snippet: For a more complete understanding of the epigenetic mechanism of AD-ND codependence, it will be necessary to use a high-resolution DNA methylation array (such as the Illumina MethylationEPIC BeadChip) assays or whole genome bisulfite sequencing (WGBS) to identify DNA methylomic changes.

Techniques: DNA Methylation Assay, Modification, Binding Assay

Journal: Carcinogenesis

Article Title: Aberrant DNA methylation in non-small cell lung cancer-associated fibroblasts

doi: 10.1093/carcin/bgv146

Figure Lengend Snippet: Primary lung TAFs exhibit global DNA hypomethylation and focal gain of DNA methylation. ( A ) Representative fluorescence images illustrating α-SMA overexpression in lung TAFs compared to paired CFs obtained with a ×20 objective (top). Scale bars here and thereafter, 30 μm. The bottom plot shows the quantification of fold α-SMA intensity per cell of fibroblasts from four randomized patients. ( B ) Unsupervised clustering of 1452 CpG sites with marked differential methylation in TAFs and CFs from 12 randomized patients of our cohort and ( C ) normalized distribution (relative density) of the corresponding β-values. Dashed vertical lines indicate median β-values. * P < 0.05; ** P < 0.01; *** P < 0.001 (here and thereafter).

Article Snippet: In the present study, we interrogated for the first time the DNA methylation landscape changes between CFs and paired TAFs from surgical patients with early stage NSCLC using the high-resolution Infinium 450K DNA Methylation Array.

Techniques: DNA Methylation Assay, Fluorescence, Over Expression, Methylation

Journal: Carcinogenesis

Article Title: Aberrant DNA methylation in non-small cell lung cancer-associated fibroblasts

doi: 10.1093/carcin/bgv146

Figure Lengend Snippet: Activating CFs with TGF-β1 partially mimics the genomic methylation changes in TAFs. ( A ) Effect of TGF-β1 on α-SMA fluorescence staining in CFs. ( B ) Effect of TGF-β1 on the DNA methylation distribution of the list of 1452 differential CpG sites in CFs from two randomly selected patients (P5 and P28). Dashed horizontal lines indicate the median of each distribution. ( C ) DNA methylation distribution of the list of 1452 differential CpG sites in CFs and paired TAFs from the same patients. ( D ) Outline of the culture assay based on polyacrylamide gels with normal- or tumor-like rigidities in the presence of TGF-β1. ( E ) Effect of culture conditions shown in ( D ) on the DNA methylation distribution of the list of 1452 differential CpG sites in CFs from the same patients.

Article Snippet: In the present study, we interrogated for the first time the DNA methylation landscape changes between CFs and paired TAFs from surgical patients with early stage NSCLC using the high-resolution Infinium 450K DNA Methylation Array.

Techniques: Methylation, Fluorescence, Staining, DNA Methylation Assay

Journal: Carcinogenesis

Article Title: Aberrant DNA methylation in non-small cell lung cancer-associated fibroblasts

doi: 10.1093/carcin/bgv146

Figure Lengend Snippet: Pathway-enrichment analysis reveals that a fraction of TAFs are fibrocyte or fibrocyte-like cells in origin. ( A ) Statistically significant overrepresented KEGG pathways within the 750 distinct genes corresponding to the list of 1452 CpG sites with marked differential methylation between TAFs and CFs. Pathway’s circle size is proportional to the number of annotated genes. Genes annotated to each pathway are color-coded according to their fold DNA methylation. None, one or two circles around each gene indicate that differential methylation was found within non-promoter, promoter, or both non-promoter and promoter regions, respectively. ( B ) Representative images of tumor histologic sections of a randomly selected patient from our cohort stained for fibrocyte markers CD34 (left) and CD45 (right). Black arrows point to CD34+ and CD45+ (non-endothelial) spindle-shaped stromal mesenchymal cells. ( C ) Histograms of CD34+ (left) and CD45+ (right) cultured CFs and TAFs from the same patient as in (B) assessed by flow cytometry. ( D ) Fold (TAFs/CFs) percentages of CD34+ and CD45+ fibroblasts from three randomly selected patients assessed as in (C).

Article Snippet: In the present study, we interrogated for the first time the DNA methylation landscape changes between CFs and paired TAFs from surgical patients with early stage NSCLC using the high-resolution Infinium 450K DNA Methylation Array.

Techniques: Methylation, DNA Methylation Assay, Staining, Cell Culture, Flow Cytometry

Journal: Carcinogenesis

Article Title: Aberrant DNA methylation in non-small cell lung cancer-associated fibroblasts

doi: 10.1093/carcin/bgv146

Figure Lengend Snippet: Validation of differential methylated promoters and the prognostic value of EDARADD in NSCLC with high stromal mesenchymal expression in-vivo . ( A ) DNA methylation of selected genes measured by pyrosequencing in TAFs and paired CFs from 12 patients. ( B ) Fold (TAFs/CFs) relative mRNA expression of selected genes in five randomized patients by qRT-PCR. Horizontal dashed lines here and thereafter are added as a reference. ( C ) Kaplan–Meier survival curve using publicly available data of a cohort of 204 NSCLC patients (161 ADC, 43 SCC) sorted by their EDARADD DNA methylation status assessed in primary tumoral DNA . ( D ) Kaplan–Meier survival curve using publicly available data of a cohort of 226 NSCLC patients (226 ADC) sorted by their EDARADD expression assessed in primary tumors . ( E ) Representative images of histologic sections from our cohort stained for EDARADD obtained from unaffected lung parenchyma (control, left) or tumor (right). Black arrows within inserts point to EDARADD+ spindle-shaped mesenchymal cells. ( F ) Scoring of EDARADD protein expression in (non-endothelial) spindle-shaped stromal cells in our cohort of 20 NSCLC patients (10 ADC, 10 SCC).

Article Snippet: In the present study, we interrogated for the first time the DNA methylation landscape changes between CFs and paired TAFs from surgical patients with early stage NSCLC using the high-resolution Infinium 450K DNA Methylation Array.

Techniques: Biomarker Discovery, Methylation, Expressing, In Vivo, DNA Methylation Assay, Quantitative RT-PCR, Staining, Control