A Protocol for the Simultaneous Analysis of Gene DNA Methylation and mRNA Expression Levels in the Rodent Brain

2016 ◽  
pp. 65-85 ◽  
Author(s):  
Juzoh Umemori ◽  
Nina N. Karpova
Keyword(s):  
Dna Methylation ◽  
Mrna Expression ◽  
Simultaneous Analysis ◽  
Expression Levels ◽  
Rodent Brain ◽  
Mrna Expression Levels

scholarly journals Beta-defensins and analogs in Helicobacter pylori infections: mRNA expression levels, DNA methylation, and antibacterial activity

PLoS ONE ◽  
2019 ◽  
Vol 14 (9) ◽  
pp. e0222295 ◽  
Author(s):  
Raffaela Pero ◽  
Tiziana Angrisano ◽  
Mariarita Brancaccio ◽  
Annarita Falanga ◽  
Lucia Lombardi ◽  
...  
Keyword(s):  
Dna Methylation ◽  
Helicobacter Pylori ◽  
Antibacterial Activity ◽  
Mrna Expression ◽  
Expression Levels ◽  
Beta Defensins ◽  
Mrna Expression Levels

scholarly journals DNA Methylation and SNPs of ABCA1 Gene Promoter Affect Gene Expression and Involve in the Pathological Mechanism of Ischemic Stroke

2021 ◽  
Author(s):  
Zhihao Li ◽  
Jun Wang ◽  
Baixue Zhou ◽  
Yang Liu ◽  
Zhaojing Zhang ◽  
...  
Keyword(s):  
Gene Expression ◽  
Dna Methylation ◽  
Ischemic Stroke ◽  
Mrna Expression ◽  
Foam Cell ◽  
Gene Promoter ◽  
Control Group ◽  
Expression Levels ◽  
Pathological Mechanism ◽  
Mrna Expression Levels

Abstract ATP binding cassette subfamily A1 (ABCA1) is a key protein in the formation of mature high density lipoprotein (HDL), which plays a crucial role in atherosclerosis. Accumulating evidence has shown that the expression levels of the ABCA1 gene are upregulated in ischemic stroke (IS) patients. However, the mechanism remains elusive. We hypothesized that DNA methylation and SNPs of ABCA1 gene promoter affect the expression levels of ABCA1 gene and involve in the pathological mechanism of IS. 100 patients with IS and 100 healthy controls were enrolled in the present study. Initially, the mRNA expression levels of ABCA1 gene were examined by qPCR and the methylation levels was detected by MethyTarget sequencing. Then, rs1800976, rs1800977, rs2246298, rs2437817, rs2740483, rs539621172 in promoter region of ABCA1 gene were selected for genotyping. Finally, the relationship between the methylation of ABCA1 gene and gene expression was verified by constructing THP-1 foam cell model. The mRNA expression levels of ABCA1 gene in the IS group were significantly higher than those in controls (P<0.05). 17 CpG sites in the promoter of ABCA1 gene were analyzed and the DNA methylation levels of CpG1, CpG7 and CpG15 sites in IS group was significantly lower than control group (P<0.05). Rs2740483, rs1800977 and rs2437817 were significantly correlated with CpG1. Rs1800977 was significantly correlated with CpG3. In summary, DNA methylation and rs2740483, rs1800977, rs2437817 of ABCA1 gene promoter affect the expression levels of ABCA1 gene, change the clearance rate of intracellular lipids, and participate in the pathogenesis of IS.

Expression profile of hepatic drug transporters in patients with hepatocellular carcinoma

2009 ◽  
Vol 27 (15_suppl) ◽  
pp. e22026-e22026
Author(s):  
K. Higuchi ◽  
T. Terada ◽  
K. Ogasawara ◽  
M. Aoki ◽  
M. Fukudo ◽  
...  
Keyword(s):  
Hepatocellular Carcinoma ◽  
Dna Methylation ◽  
Mrna Expression ◽  
Expression Profile ◽  
Promoter Region ◽  
Drug Transporters ◽  
Hepatic Drug ◽  
Expression Levels ◽  
Mrna Expression Levels ◽  
Liver Tissues

e22026 Background: Hepatocellular carcinoma (HCC) does not respond sufficiently to chemotherapy. It has been demonstrated that several transporters can cause drug resistance in tumor cells. In human liver, various drug transporters are expressed and play pivotal roles in hepatic uptake and excretion of drugs. In HCC, the expression profile of hepatic drug transporters may be altered due to pathophysiological changes. However, limited data are available on their expression and regulation in HCC tissues. The aims of this study were to determine the expression levels of hepatic drug transporters in HCC patients and to identify transporter specifically regulated in HCC tissues. Methods: The mRNA expression levels of 17 drug transporters, including 9 solute carrier (SLC) transporters (OAT2, OAT7, OATP1B1, OATP1B3, OATP2B1, OCT1, OCTN2, MATE1 and PEPT1) and 8 ATP-binding cassette (ABC) transporters (R1, MRP1–6 and BCRP), were determined by real-time RT-PCR in cancerous and non-cancerous liver tissues from 57 patients with HCC. Protein expression level was evaluated by Western blot analysis. In addition, DNA methylation status in promoter region of transporter gene was analyzed using bisulfite sequencing. Results: The mRNA expression levels of 11 transporters (OAT7, OCTN2, MATE1, PEPT1, R1, MRP1–6) were significantly increased (1.45 to 4.74-fold, p<0.05) in HCC tissues compared with non-cancerous liver tissues, whereas only OCT1 mRNA was significantly decreased (0.61-fold, p=0.0004). Notably, MRP4 mRNA showed the highest increase, which was consistent with the marked augmentation in the protein level. The frequency of methylation in MRP4 promoter region was comparable between HCC and non-cancerous liver tissues, suggesting other mechanisms than DNA methylation for MRP4 up-regulation. Interestingly, the mRNA expression levels of MRP4 did not change with the differentiation of HCC. Conclusions: It was revealed that most of the hepatic drug transporters were elevated in HCC tissues. Furthermore, we first identified that MRP4 showed remarkable increase in protein as well as mRNA expression levels. These findings suggest that MRP4 is a novel candidate for diagnostic marker and a target molecule to reverse multidrug resistance in chemotherapy for HCC. No significant financial relationships to disclose.

scholarly journals DNA methylation profiling in recurrent miscarriage

PeerJ ◽  
2020 ◽  
Vol 8 ◽  
pp. e8196
Author(s):  
Li Pi ◽  
Zhaofeng Zhang ◽  
Yan Gu ◽  
Xinyue Wang ◽  
Jianmei Wang ◽  
...  
Keyword(s):  
Dna Methylation ◽  
Mrna Expression ◽  
Candidate Genes ◽  
Recurrent Miscarriage ◽  
Enrichment Analysis ◽  
Type I ◽  
Diagnostic Biomarkers ◽  
Expression Levels ◽  
A Genome ◽  
Mrna Expression Levels

Recurrent miscarriage (RM) is a complex clinical problem. However, specific diagnostic biomarkers and candidate regulatory targets have not yet been identified. To explore RM-related biological markers and processes, we performed a genome-wide DNA methylation analysis using the Illumina Infinium HumanMethylation450 array platform. Methylation variable positions and differentially methylated regions (DMRs) were selected using the Limma package in R language. Thereafter, gene ontology (GO) enrichment analysis and pathway enrichment analysis were performed on these DMRs. A total of 1,799 DMRs were filtered out between patients with RM and healthy pregnant women. The GO terms were mainly related to system development, plasma membrane part, and sequence-specific DNA binding, while the enriched pathways included cell adhesion molecules, type I diabetes mellitus, and ECM–receptor interactions. In addition, genes, including ABR, ALCAM, HLA-E, HLA-G, and ISG15, were obtained. These genes may be potential candidates for diagnostic biomarkers and possible regulatory targets in RM. We then detected the mRNA expression levels of the candidate genes. The mRNA expression levels of the candidate genes in the RM group were significantly higher than those in the control group. However, additional research is still required to confirm their potential roles in the occurrence of RM.

scholarly journals Lifestyle and Genetic Factors Modify Parent-of-Origin Effects on the Human Methylome

2021 ◽  
Author(s):  
Yanni Zeng ◽  
Carmen Amador ◽  
Chenhao Gao ◽  
Rosie M Walker ◽  
Stewart W Morris ◽  
...  
Keyword(s):  
Dna Methylation ◽  
Environmental Factors ◽  
Mrna Expression ◽  
Environmental Exposures ◽  
Health Study ◽  
Expression Levels ◽  
Parent Of Origin ◽  
Origin Effects ◽  
Genetic And Environmental Factors ◽  
Mrna Expression Levels

Background: parent-of-origin effects (POE) play important roles in development and complex disease and thus understanding their regulation and associated molecular and phenotypic variation are warranted. Previous studies have mainly focused on the detection of genomic regions or phenotypes regulated by POE. Understanding whether POE may be modified by environmental or genetic exposures is important for understanding of the source of POE-associated variation, but only a few case studies addressing these modifiable POE exist. Methods: in order to understand this high order of POE regulation, we screened 101 genetic and environmental factors such as "predicted mRNA expression levels" of DNA methylation/imprinting machinery genes and early/late lifestyle/environmental exposures. POE-mQTL-modifier interaction models were proposed to test the potential of these factors to modify POE at DNA methylation using data from Generation Scotland: The Scottish Family Health Study(N=2315). Results: a set of vulnerable/modifiable POE-CpGs were identified (modifiable-POE-regulated CpGs, N=3). Four factors, "lifetime smoking status" and "predicted mRNA expression levels" of TET2, SIRT1 and KDM1A, were found to significantly modify the POE on the three CpGs in both discovery and replication datasets. Importantly, the POE on one of the CpGs were modified by both genetic and environmental factors. We further identified plasma protein and health-related phenotypes associated with the methylation level of one of the identified CpGs. Conclusions: the modifiable POE identified here revealed an important yet indirect path through which genetic background and environmental exposures introduce their effect on DNA methylation, motivating future comprehensive evaluation of the role of these modifiers in complex diseases.

scholarly journals The role of CXCR3 and its ligands expression in Brucellar spondylitis

2020 ◽  
Vol 21 (1) ◽  
Author(s):  
Xin Hu ◽  
Xiaoqian Shang ◽  
Liang Wang ◽  
Jiahui Fan ◽  
Yue Wang ◽  
...  
Keyword(s):  
Protein Expression ◽  
Mrna Expression ◽  
Mononuclear Cells ◽  
Healthy Controls ◽  
Inflammatory Infiltration ◽  
Peripheral Blood Mononuclear ◽  
Expression Levels ◽  
Inflammatory Damage ◽  
Ifn Γ ◽  
Mrna Expression Levels

Abstract Aim Brucellar spondylitis (BS) is one of the most serious complications of brucellosis. CXCR3 is closely related to the severity of disease infection. This research aimed to study the degree of BS inflammatory damage through analyzing the expression levels of CXCR3 and its ligands (CXCL9 and CXCL10) in patients with BS. Methods A total of 29 BS patients and 15 healthy controls were enrolled. Real-Time PCR was used to detect the mRNA expression levels of IFN-γ, CXCR3, CXCL9 and CXCL10 in peripheral blood mononuclear cells (PBMCs) of BS patients and healthy controls. Hematoxylin-Eosin staining was used to show the pathological changes in BS lesion tissues. Immunohistochemistry staining was used to show the protein expression levels of Brucella-Ab, IFN-γ, CXCR3, CXCL9 and CXCL10 in BS lesion tissues. At the same time, ELISA was used to detect the serum levels of IFN-γ, CXCL9 CXCL10 and autoantibodies against CXCR3 in patients with BS. Results In lesion tissue of BS patients, it showed necrosis of cartilage, acute or chronic inflammatory infiltration. Brucella-Ab protein was abundantly expressed in close lesion tissue. And the protein expression levels of IFN-γ, CXCR3 and CXCL10 were highly expressed in close lesion tissue and serum of BS patients. At the same time, the mRNA expression levels of IFN-γ, CXCR3 and CXCL10 in PBMCs of BS patients were significantly higher than those in controls. Conclusion In our research, the expression levels of IFN-γ, CXCR3 and its ligands were significantly higher than those in controls. It suggested that high expression levels of IFN-γ, CXCR3 and its ligands indicated a serious inflammatory damage in patients with BS.

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