Quantitative analysis of chromatin accessibility in mouse embryonic fibroblasts.

Genomic DNA of eukaryotic cells is hierarchically packaged into chromatin by histones. The dynamic organization of chromatin fibers plays a critical role in the regulation of gene transcription and other DNA-associated biological processes. Recently, numerous approaches have been developed to map the chromatin organization by characterizing chromatin accessibilities in genome-wide. However, reliable methods to quantitatively map chromatin accessibility are not well-established, especially not on a genome-wide scale. Here, we developed a modified MNase-seq for mouse embryonic fibroblasts, wherein chromatin was partially digested at multiple digestion times using micrococcal nuclease (MNase), allowing quantitative analysis of local yet genome-wide chromatin compaction. Our results provide strong evidence that the chromatin accessibility at promoter regions are positively correlated with gene activity. In conclusion, our assay is an ideal tool for the quantitative study of gene regulation in the perspective of chromatin accessibility.

Generation of a human induced pluripotent stem cell line (SMUSHi002-A) from an ALS patient carrying a heterozygous mutation c.1562G > A in the FUS gene.

Amyotrophic lateral sclerosis (ALS) is a fatal neurodegenerative disease. Affected patients experience gradual loss of their spinal cord and cortical motor neurons with consequent muscle weakness and emaciation, and eventual respiratory failure. The pathogenesis of ALS remains largely unknown although the FUS (sarcoma fusion gene) gene is known to be one of the major pathogenic genes. We have generated an induced pluripotent stem cell line SMUSHi002-A from an ALS patient who carries a heterozygous mutation c.1562G > A in FUS. This cell line will serve as a useful model to investigate disease pathogenesis and develop potential therapeutic approaches for ALS.

Periconceptional folate and gestational diabetes mellitus: a systematic review and meta-analysis of cohort studies.

OBJECTIVE: To examine the relationship between periconceptional folate exposure and risk of gestational diabetes mellitus (GDM). METHODS: Several electronic databases, including PubMed, Embase, China National Knowledge Infrastructure (CNKI), China Biology Medicine (CBM), and Cochrane Library, were searched for all relevant cohort studies by January 2021. Studies on relationship between folate exposure (intake or status) and GDM risk were included. Quality of included studies was assessed using Newcastle-Ottawa Scale. Random effects meta-analysis was performed to estimate overall odds ratio (OR) and 95% confidence intervals (CIs) by Stata software (Stata Corp., College Station, TX). RESULTS: Ten cohort studies with 40,244 pregnancies were eligible for quantitative meta-analysis. Significant association was observed between folate exposure and risk of GDM (OR = 1.24, p=.036, 95% CI: 1.01-1.52). Subgroup analysis revealed that periconceptional folate exposure of population in China (OR = 1.35, 95% CI: 1.09-1.67) but not in western countries, folate exposure during pregnancy (OR = 1.49, 95% CI: 1.22-1.81) but not before pregnancy, and internal folate exposure (OR = 1.36, 95% CI: 1.10-1.67), were significantly associated with increased GDM risk. CONCLUSIONS: Overall, periconceptional folate exposure is positively associated with GDM risk, especially the exposure during pregnancy and exposure in Chinese populations.

Generation of a human induced pluripotent stem cell line (SBWCHi001-A) from a patient with NEDSDV carrying a pathogenic mutation in CTNNB1 gene.

Neurodevelopmental disorder with spastic diplegia and visual defects (NEDSDV) is a rare disease. Patients with NEDSDV are usually accompanied by microcephaly, severe mental retardation, spasticity, and global developmental delay. Recent studies showed that mutations in CTNNB1 are responsible for the phenotype. Here, we generated an induced pluripotent stem cell (iPSC) line (SBWCHi001-A) from an 18-month-old patient with NEDSDV, who harbored a de novo heterozygous mutation in CTNNB1. The transduced iPSCs expressed pluripotency markers, and could differentiate into three germ layers in vitro. This cell line will be a cell model to explore the pathogenesis of NEDSDV and discover potential therapies.

Identification of novel FUS and TARDBP gene mutations in Chinese amyotrophic lateral sclerosis patients with HRM analysis.

Amyotrophic lateral sclerosis (ALS) is a neurodegenerative disease characterized by progressive loss of motor neurons. More than 30 genes have been linked to ALS to date, including FUS and TARDBP, which exhibit similar roles in RNA metabolism. This study explored the use of high-resolution melting (HRM) analysis to screen for FUS and TARDBP mutation hotspot regions in 146 Chinese ALS patients, which achieved 100% detection. Two FUS mutations were observed in two different familial ALS probands, a missense mutation (p.R521H) and a novel splicing mutation (c.1541+1G>A). Five TARDBP mutations were identified in six ALS patients, including a novel 3'UTR mutation (c.*731A>G) and four missense mutations (p.G294V, p.M337V, p.G348V, and p.I383V). We found that FUS mutations were present in 1.4% of Chinese ALS patients, whereas TARDBP mutations were responsible for 4.1% of Chinese ALS cases. Here, we describe the accuracy of using highly sensitive HRM analysis to identify two novel FUS and TARDBP mutations in Chinese sporadic and familial ALS cases. Our study contributes to the further understanding of the genetic and phenotypic diversity of ALS.

Effects of long term low- and high-dose sodium arsenite exposure in human transitional cells.

Epidemiological studies have revealed the association between increased risk of bladder cancer and chronic arsenic exposure. Here, we explored biological effects of arsenic in T24. Microarray analysis was applied to analyze mRNA in T24 following 0, 2 or 5 µM sodium arsenite (As) exposure for 72 hours. Long term (up to 140 days) low-dose (200 nM) and high-dose (1,000 nM) As decreased E-cadherin protein level through different mechanisms because the mRNA levels of E-cadherin increased following low-dose As exposure but decreased following high-dose As exposure. Long term As increased the protein levels of N-cadherin, vimentin, ß-catenin, and slug. Low-dose As exposure resulted in a change in the morphology of T24 cells from an epithelial to a mesenchymal-like appearance. Knockdown of E-cadherin increased the protein levels of N-cadherin, vimentin, ß-catenin, and slug. Cell proliferation and growth of T24 with or without As exposure for 100 days were assayed using EdU and WST, respectively. Low-dose As exposure increased cell proliferation and growth while high-dose As exposure decreased both. Long term As activated p53 on account of increasing protein levels of p53, p-p53 (Ser15), and mRNA levels of p21. These demonstrate that arsenic exposure exerts multiple effects. Long term low- or high-dose arsenic induces epithelial-mesenchymal transition, likely via downregulation of E-cadherin, activates p53, and differently affects cell proliferation/growth.