Diannan Xu

5-Hydroxymethylcytosine signatures in circulating cell-free DNA as diagnostic biomarkers for human cancers

DNA modifications such as 5-methylcytosine (5mC) and 5-hydroxymethylcytosine (5hmC) are epigenetic marks known to affect global gene expression in mammals. Given their prevalence in the human genome, close correlation with gene expression and high chemical stability, these DNA epigenetic marks could serve as ideal biomarkers for cancer diagnosis. Taking advantage of a highly sensitive and selective chemical labeling technology, we report here the genome-wide profiling of 5hmC in circulating cell-free DNA (cfDNA) and in genomic DNA (gDNA) of paired tumor and adjacent tissues collected from a cohort of 260 patients recently diagnosed with colorectal, gastric, pancreatic, liver or thyroid cancer and normal tissues from 90 healthy individuals. 5hmC was mainly distributed in transcriptionally active regions coincident with open chromatin and permissive histone modifications. Robust cancer-associated 5hmC signatures were identified in cfDNA that were characteristic for specific cancer types. 5hmC-based biomarkers of circulating cfDNA were highly predictive of colorectal and gastric cancers and were superior to conventional biomarkers and comparable to 5hmC biomarkers from tissue biopsies. Thus, this new strategy could lead to the development of effective, minimally invasive methods for diagnosis and prognosis of cancer from the analyses of blood samples.

A single nucleotide variant in microRNA-1269a promotes the occurrence and process of hepatocellular carcinoma by targeting to oncogenes SPATS2L and LRP6.

Hepatocellular carcinoma (HCC) is one of the malignant and lethal cancers. Single nucleotide polymorphisms (SNPs) in microRNAs(miRNAs) can affect the expression and target identification of miRNAs and lead to the formation of malignant tumors. However, little is known about whether microRNA-1269a (miR-1269a) SNPs affect the susceptibility and progression of HCC or their specific mechanism. The association between microRNA-1269a rs73239138 and the susceptibility to HCC was verified by MassARRAY assay in a large case-control sample. The effect of miR-1269a and the variant on the proliferation and apoptosis of HCC cells was examined by flow cytometry (FCM), CCK8 assay and Western blot. The target of miR-1269a was identified by bioinformatics analysis and qRT-PCR and its role on cell proliferative capacity was examined by CCK8 assay. The expression level of miR-1269a was analyzed by qRT-PCR in HCC cells transfected with wild or variant type pre-miR-1269a plasmid.MiR-1269a produced a tumor suppressor effect by inhibiting cell proliferation and inducing apoptosis of human HCC cells, possibly via inhibiting the expression of its target genes SPATS2L and LRP6, which were tumor promoters. While, rs73239138 (G>A) in miR-1269a reduced the anticancer effect of miR-1269a possibly by attenuating its total amount in HCC cells or its target recognition, reduce its inhibition on target genes and promoted the susceptibility to HCC. Our findings for the first time proved that miR-1269a SNP plays a role in the occurrence and process of HCC and the relevant mechanism, in accompany with the discovery of the novel target genes of miR-1269a.

DNA 5-Hydroxymethylcytosines from Cell-free Circulating DNA as Diagnostic Biomarkers for Human Cancers

DNA modifications such as 5-methylcytosines (5mC) and 5-hydroxymethylcytosines (5hmC) are epigenetic marks known to affect global gene expression in mammals(1, 2). Given their prevalence in the human genome, close correlation with gene expression, and high chemical stability, these DNA epigenetic marks could serve as ideal biomarkers for cancer diagnosis. Taking advantage of a highly sensitive and selective chemical labeling technology(3), we report here genome-wide 5hmC profiling in circulating cell-free DNA (cfDNA) and in genomic DNA of paired tumor/adjacent tissues collected from a cohort of 90 healthy individuals and 260 patients recently diagnosed with colorectal, gastric, pancreatic, liver, or thyroid cancer. 5hmC was mainly distributed in transcriptionally active regions coincident with open chromatin and permissive histone modifications. Robust cancer-associated 5hmC signatures in cfDNA were identified with specificity for different cancers. 5hmC-based biomarkers of circulating cfDNA demonstrated highly accurate predictive value for patients with colorectal and gastric cancers versus healthy controls, superior to conventional biomarkers, and comparable to 5hmC biomarkers from tissue biopsies. This new strategy could lead to the development of effective blood-based, minimally-invasive cancer diagnosis and prognosis approaches.

H19/miR-148a/USP4 axis facilitates liver fibrosis by enhancing TGF-β signaling in both hepatic stellate cells and hepatocytes: ZHU et al.

Liver fibrosis is a wound‐healing response represented by excessive extracellular matrix deposition. Activation of hepatic stellate cell (HSC) is the critical cellular basis for hepatic fibrogenesis, whereas hepatocyte undergoes epithelial‐mesenchymal transition (EMT) which is also involved in chronic liver injury. Long noncoding RNA H19 has been found to be associated with cholestatic liver fibrosis lately. However, the role of H19 in liver fibrosis remains largely to be elucidated. In this study, we found that the expression of H19 was significantly upregulated in the liver tissue of CCl4‐induced mice, a toxicant‐induced liver fibrogenesis model. Overexpression of H19 significantly aggravated activation of HSC and EMT of hepatocyte both by stimulating transforming growth factor‐β (TGF‐β) pathway. In terms of mechanism, H19 functioned as a competing endogenous RNA to sponge miR‐148a and subsequently sustained the level of ubiquitin‐specific protease 4 (USP4), which was an identified target of miR‐148a and was able to stabilize TGF‐β receptor I. In conclusion, our findings revealed a novel H19/miR‐148a/USP4 axis which promoted liver fibrosis via TGF‐β pathway in both HSC and hepatocyte, indicating that H19 could become a promising target for the treatment of liver fibrosis.

Downregulated miRNA‑1269a variant (rs73239138) decreases the susceptibility to gastric cancer via targeting ZNF70

Although emerging evidence has indicated that single nucleotide polymorphisms (SNPs) in microRNAs (miRNAs) are associated with susceptibility to gastric cancer, a limited number of studies have revealed the underlying molecular mechanisms. In the present study, the results suggested that miR-1269a rs73239138 has a role in decreasing the risk of gastric cancer. The level of miR-1269a variant expression was significantly downregulated compared with the wild-type miR-1269a in the gastric cells (Fig. 1). Furthermore, overexpression of miR-1269a inhibited apoptosis of gastric cancer cells. Expression of the miR-1269a variant inhibited the function of miR-1269a by increasing the apoptotic rate and the expression of Bik, Bim and Bak was upregulated consistently. In addition, zinc-finger protein 70 (ZNF70) was identified to be a target gene of miR-1269a, which was downregulated by miR-1269a and upregulated by miR-1269a variant. ZNF70 was indicated to exert a role as a tumor suppressor in gastric cancer. To the best our knowledge, the present study for the first time highlights a critical role of miR-1269a variant rs73239138 in decreasing the susceptibility to gastric cancer by downregulating its expression and targeting ZNF70, which promotes apoptosis of gastric cancer cells. This SNP is indicated to serve as a potential biomarker and therapeutic target for gastric cancer.

Global transcriptome‑wide analysis of the function of GDDR in acute gastric lesions

Acute gastric lesions induced by stress are frequent occurrences in medical establishments. The gastric dramatic downrelated gene (GDDR) is a secreted protein, which is abundantly expressed in normal gastric epithelia and is significantly decreased in gastric cancer. In our previous study, it was found that GDDR aggravated stress-induced acute gastric lesions. However, the role of GDDR in acute gastric lesions remains to be fully elucidated. In the present study, RNA sequencing was performed in order to examine the gene expression profile regulated by GDDR in acute gastric lesions. The dataset comprised four stomach samples from wild-type (WT) mice and four stomach samples from GDDR-knockout mice. Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway enrichment analyses were performed to analyze the differentially-expressed genes (DEGs). Weighted correlation network analysis was used to identify clusters of highly correlated genes. Cytoscape was used to construct a protein-protein interaction network (PPI) of the DEGs. Based on the GO analysis, the upregulated DEGs were distinctly enriched in muscle contraction and response to wounding; and the downregulated DEGs were significantly enriched in the regulation of nitrogen compound metabolic process and regulation of RNA metabolic process. The results of the KEGG pathway analysis showed that the upregulated DEGs were enriched in ECM-receptor interaction and the signaling pathway of cGMP-PKG, and the downregulated DEGs were enriched in the renin-angiotensin system and glycerolipid metabolism. The co-expression network revealed a group of genes, which were associated with increased wound healing in the WT mice. Significant pathways were identified through the PPI network, including negative regulation of the signaling pathway of glucocorticoid receptor, regulation of cellular stress response, and regulation of hormone secretion. In conclusion, the present study improves current understanding of the molecular mechanism underlying acute gastric lesions and may assist in the treatment of gastric lesions.