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DNA methylation plays key roles in diverse biological processes such as X chromosome inactivation, transposable element repression, genomic imprinting, and tissue-specific gene expression. Sequencing-based DNA methylation profiling provides an unprecedented opportunity to map and compare complete DNA methylomes. This includes one of the most widely applied technologies for measuring DNA methylation: methylated DNA immunoprecipitation followed by sequencing (MeDIP-seq), coupled with a complementary method, methylation-sensitive restriction enzyme sequencing (MRE-seq). A computational approach that integrates data from these two different but complementary assays and predicts methylation differences between samples has been unavailable. Here, we present a novel integrative statistical framework M&M (for integration of MeDIP-seq and MRE-seq) that dynamically scales, normalizes, and combines MeDIP-seq and MRE-seq data to detect differentially methylated regions. Using sample-matched whole-genome bisulfite sequencing (WGBS) as a gold standard, we demonstrate superior accuracy and reproducibility of M&M compared to existing analytical methods for MeDIP-seq data alone. M&M leverages the complementary nature of MeDIP-seq and MRE-seq data to allow rapid comparative analysis between whole methylomes at a fraction of the cost of WGBS. Comprehensive analysis of nineteen human DNA methylomes with M&M reveals distinct DNA methylation patterns among different tissue types, cell types, and individuals, potentially underscoring divergent epigenetic regulation at different scales of phenotypic diversity. We find that differential DNA methylation at enhancer elements, with concurrent changes in histone modifications and transcription factor binding, is common at the cell, tissue, and individual levels, whereas promoter methylation is more prominent in reinforcing fundamental tissue identities.

Aim: Zinc finger and BTB domain‐containing protein 28 (ZBTB28) is a potential tumor suppressor for some cancers. However, its epigenetic regulation and functions in renal cell carcinoma (RCC) remain to be elucidated. Methods: The expression of ZBTB28 mRNA was analyzed by semi‐quantitative reverse transcription polymerase chain reaction (PCR) in nine RCC cell lines and normal kidney tissues. Methylation status of ZBTB28 promoter was assessed by methylation‐specific PCR in RCC cell lines, primary RCC, tumors and adjacent tissues. The involvement of ZBTB28 in cell proliferation and migration was investigated. Results: ZBTB28 promoter was hypermethylated in 88.9% (8/9) of RCC cell lines with reduced ZBTB28 mRNA expression, and could be reversed by DNA methyltransferase inhibitors. The methylation of ZBTB28 promoter was detected in 73.5% (36/49) of primary RCC tissues, compared with 7.1% (1/14) in normal tissues. Overexpression of ZBTB28 significantly inhibited RCC cell proliferation and migration, and induced apoptosis. Further analyses revealed that ZBTB28 upregulation could inhibit multiple oncogenic signaling transduction pathways. Conclusion: ZBTB28 is frequently silenced by promoter methylation in RCC pathogenesis and functions as a novel tumor suppressive gene. ZBTB28 may be a potential target for the development of RCC therapeutic strategies. [ABSTRACT FROM AUTHOR]

Objective To explore the relation between micro RNA-30a (miR-30a) and Notch1, and to evaluate the potential prognostic role of miR-30a in invasive urothelial carcinoma of the bladder ( UCB). Patients and methods In all, 50 invasive UCB tissue specimens, along with the adjacent bladder tissue specimens were obtained, and the clinical parameters of the 50 patients were analysed. Bioinformatics analysis was performed and miR-30a was selected as a potential mi RNA targeting Notch1, with a luciferase assay performed to verify the binding site between miR-30a and Notch1. Quantitative real-time reverse transcriptase-polymerase chain reaction was used to assess the RNA expressions of miR-30a and Notch1, while Western Blotting and immunohistochemical staining were used to assess the protein expression of Notch1. Finally, cell proliferation, cell cycle, cell migration and invasion assays were used to evaluate the cellular effects of miR-30a and Notch1 on the UCB cell lines T24 and 5637. Results MiR-30a was downregulated in tumour tissues when compared with adjacent bladder tissues ( P < 0.001), negatively correlating with Notch1 messenger RNA ( R 2 0.106, P = 0.021) in invasive UCB, and miR-30a expression further decreased in patients with shorter overall survival and disease-free survival ( P = 0.039 and P = 0.037, respectively). The luciferase assay showed that miR-30a inhibited the Notch1 3′-untranslated region reporter activities in the T24 and 5637 cell lines (both P < 0.001). Both miR-30a and small interfering RNA Notch1 negatively regulated cell proliferation ( P = 0.002 and P = 0.035 in T24; P = 0.029 and P = 0.037 in 5637 cell lines), activated cell cycle arrest (both P < 0.001 in T24; both P < 0.001 in 5637 cell lines), and prevented cellular migration (both P < 0.001 in T24; P = 0.003 and P < 0.001 in 5637 cell lines) and invasion ( P = 0.009 and P = 0.006 in T24; P = 0.006 and P = 0.002 in 5637 cell lines) abilities. Ectopic Notch1 without the 3′untranslated region partially rescued the above-mentioned cellular effects of over-expressed miR-30a on T24 and 5637 cells. Conclusions MiR-30a lessens cellular malignancy by antagonising oncogene Notch1 and plays an effective prognostic role in invasive UCB. [ABSTRACT FROM AUTHOR]

Three-dimensional (3D) cell culture has been reported to increase the therapeutic potentials of mesenchymal stem cells ( MSCs). In this study, we aimed to investigate the therapeutic effects of 3D spheroids of human adipose-derived MSCs for acute kidney injury ( AKI). In vitro studies indicated that 3D spheroids of MSCs produced higher levels of extracellular matrix proteins (including collagen I, fibronectin and laminin), and exhibited stronger anti-apoptotic and anti-oxidative capacities than two-dimensional (2D) cultured cells. Furthermore, 3D culture increased the paracrine secretion of cytokines by MSCs, including angiogenic factors ( VEGF and basic fibroblast growth factor), anti-apoptotic factors (epidermal growth factor and hepatocyte growth factor), the anti-oxidative factor insulin-like growth factor and the anti-inflammatory protein tumour necrosis factor-alpha stimulated gene/protein 6. Consistent with in vitro experiments, 3D spheroids of MSCs showed enhanced survival and paracrine effects in vivo. More importantly, when injected into the kidney of model rats with ischemia-reperfusion (I/R)-induced AKI, 3D spheroids were more beneficial in protecting the I/R kidney against apoptosis, reducing tissue damage, promoting vascularization and ameliorating renal function compared with 2D cultured cells. Therefore, the 3D culture strategy improved the therapeutic effects of MSCs, and might be promising for AKI treatment. [ABSTRACT FROM AUTHOR]