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Methylation status of DJ-1 in leukocyte DNA of Parkinson's disease patients.

Tan Y, Wu L, Li D, Liu X, Ding J, Chen S - Transl Neurodegener (2016)

Bottom Line: It was found reduced in PD brains, CSF and saliva, although there were conflicting results.In SH-SY5Y cell model treated by methylation inhibitor, there was no significant change of DJ-1 expression in either mRNA level or protein level.Our results indicated that DNA methylation inhibitor didn't alter DJ-1 gene expression in SH-SY5Y cell model, and DNA methylation of DJ-1 promoter region in PBLs level might not be an efficient biomarker for PD patients.

View Article: PubMed Central - PubMed

Affiliation: Department of Neurology, and Institute of Neurology, Ruijin Hospital Affiliated to Shanghai Jiao Tong University School of Medicine, Shanghai, 200025 China.

ABSTRACT

Background: DJ-1 has been thought as a candidate biomarker for Parkinson's disease (PD). It was found reduced in PD brains, CSF and saliva, although there were conflicting results. How DJ-1 expression may be regulated is not clear. Recently, blood-based DNA methylation represents a highly promising biomarker for PD by regulating the causative gene expression. Thus, in this study, we try to explore whether blood-based DNA methylation of DJ-1 could be used as a biomarker to differentiate PD patients from normal control (NC), and whether DNA methylation could regulate DJ-1 expression in a SH-SY5Y cell model.

Methods: Forty PD patients and 40 NC were recruited in this study. DNA was extracted from peripheral blood leukocytes (PBLs). Methylation status of two CpG islands (CpG1 and CpG2) in promoter region of DJ-1 was explored by bisulfite specific PCR-based sequencing method. Methylation inhibitor 5-Aza-dC was used to treat SH-SY5Y cell line, DJ-1 level was detected in both mRNA and protein level.

Results: CpG sites in these two CpG islands (CpG1 and CpG2) of DJ-1 were unmethylated in both PD and NC group. In SH-SY5Y cell model treated by methylation inhibitor, there was no significant change of DJ-1 expression in either mRNA level or protein level.

Conclusions: Our results indicated that DNA methylation inhibitor didn't alter DJ-1 gene expression in SH-SY5Y cell model, and DNA methylation of DJ-1 promoter region in PBLs level might not be an efficient biomarker for PD patients.

No MeSH data available.


Related in: MedlinePlus

Bisulfite specific PCR-based sequencing analysis of two CpG islands methylation in 40 PD patients and 40 normal controls. a and b Unmethylated cytosines (c) were converted to uracil (U) after bisulfite treatment and amplified as thymines (T) in PCR amplication. Unmethylated CpG sites 1–5 in CpG1 island and CpG sites 9–16 in CpG2 island were shown here as an example. c and d All the CpG sites of these two CpG islands were unmethylated. Here are the methylation of CpG1 island and CpG2 island from 5 PD and 5 NC cases, and the rest cases showed the exact same methylation pattern
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Fig2: Bisulfite specific PCR-based sequencing analysis of two CpG islands methylation in 40 PD patients and 40 normal controls. a and b Unmethylated cytosines (c) were converted to uracil (U) after bisulfite treatment and amplified as thymines (T) in PCR amplication. Unmethylated CpG sites 1–5 in CpG1 island and CpG sites 9–16 in CpG2 island were shown here as an example. c and d All the CpG sites of these two CpG islands were unmethylated. Here are the methylation of CpG1 island and CpG2 island from 5 PD and 5 NC cases, and the rest cases showed the exact same methylation pattern

Mentions: Based on the NCBI database, the promoter region of human DJ-1 gene (NM_001123377) has two CpG islands (CGIs), locating at−1545 ~−1244 bp(CpG1),−1178 ~−732 bp(CpG2) upstream of the translation initiation site (Fig. 1a). CpG1 has 15 CpG sequences (Fig. 1b, a). CpG2 contains 21 CpG sequences (Fig. 1b, b). Bisulfite specific PCR-based sequencing method was used to examine the average methylation level at each CpG site (See methods). Our results showed that all the CpG sites probed in the two CGIs were unmethylated in both PD and NC (Fig. 2). The stable unmethylated status of CpG sites in both PD and NC group indicated that CpG methylation in the promoter region of DJ-1 in PBLs might have very limited or no regulatory effects on DJ-1 expression.Fig 1


Methylation status of DJ-1 in leukocyte DNA of Parkinson's disease patients.

Tan Y, Wu L, Li D, Liu X, Ding J, Chen S - Transl Neurodegener (2016)

Bisulfite specific PCR-based sequencing analysis of two CpG islands methylation in 40 PD patients and 40 normal controls. a and b Unmethylated cytosines (c) were converted to uracil (U) after bisulfite treatment and amplified as thymines (T) in PCR amplication. Unmethylated CpG sites 1–5 in CpG1 island and CpG sites 9–16 in CpG2 island were shown here as an example. c and d All the CpG sites of these two CpG islands were unmethylated. Here are the methylation of CpG1 island and CpG2 island from 5 PD and 5 NC cases, and the rest cases showed the exact same methylation pattern
© Copyright Policy - OpenAccess
Related In: Results  -  Collection

License 1 - License 2
Show All Figures
getmorefigures.php?uid=PMC4815061&req=5

Fig2: Bisulfite specific PCR-based sequencing analysis of two CpG islands methylation in 40 PD patients and 40 normal controls. a and b Unmethylated cytosines (c) were converted to uracil (U) after bisulfite treatment and amplified as thymines (T) in PCR amplication. Unmethylated CpG sites 1–5 in CpG1 island and CpG sites 9–16 in CpG2 island were shown here as an example. c and d All the CpG sites of these two CpG islands were unmethylated. Here are the methylation of CpG1 island and CpG2 island from 5 PD and 5 NC cases, and the rest cases showed the exact same methylation pattern
Mentions: Based on the NCBI database, the promoter region of human DJ-1 gene (NM_001123377) has two CpG islands (CGIs), locating at−1545 ~−1244 bp(CpG1),−1178 ~−732 bp(CpG2) upstream of the translation initiation site (Fig. 1a). CpG1 has 15 CpG sequences (Fig. 1b, a). CpG2 contains 21 CpG sequences (Fig. 1b, b). Bisulfite specific PCR-based sequencing method was used to examine the average methylation level at each CpG site (See methods). Our results showed that all the CpG sites probed in the two CGIs were unmethylated in both PD and NC (Fig. 2). The stable unmethylated status of CpG sites in both PD and NC group indicated that CpG methylation in the promoter region of DJ-1 in PBLs might have very limited or no regulatory effects on DJ-1 expression.Fig 1

Bottom Line: It was found reduced in PD brains, CSF and saliva, although there were conflicting results.In SH-SY5Y cell model treated by methylation inhibitor, there was no significant change of DJ-1 expression in either mRNA level or protein level.Our results indicated that DNA methylation inhibitor didn't alter DJ-1 gene expression in SH-SY5Y cell model, and DNA methylation of DJ-1 promoter region in PBLs level might not be an efficient biomarker for PD patients.

View Article: PubMed Central - PubMed

Affiliation: Department of Neurology, and Institute of Neurology, Ruijin Hospital Affiliated to Shanghai Jiao Tong University School of Medicine, Shanghai, 200025 China.

ABSTRACT

Background: DJ-1 has been thought as a candidate biomarker for Parkinson's disease (PD). It was found reduced in PD brains, CSF and saliva, although there were conflicting results. How DJ-1 expression may be regulated is not clear. Recently, blood-based DNA methylation represents a highly promising biomarker for PD by regulating the causative gene expression. Thus, in this study, we try to explore whether blood-based DNA methylation of DJ-1 could be used as a biomarker to differentiate PD patients from normal control (NC), and whether DNA methylation could regulate DJ-1 expression in a SH-SY5Y cell model.

Methods: Forty PD patients and 40 NC were recruited in this study. DNA was extracted from peripheral blood leukocytes (PBLs). Methylation status of two CpG islands (CpG1 and CpG2) in promoter region of DJ-1 was explored by bisulfite specific PCR-based sequencing method. Methylation inhibitor 5-Aza-dC was used to treat SH-SY5Y cell line, DJ-1 level was detected in both mRNA and protein level.

Results: CpG sites in these two CpG islands (CpG1 and CpG2) of DJ-1 were unmethylated in both PD and NC group. In SH-SY5Y cell model treated by methylation inhibitor, there was no significant change of DJ-1 expression in either mRNA level or protein level.

Conclusions: Our results indicated that DNA methylation inhibitor didn't alter DJ-1 gene expression in SH-SY5Y cell model, and DNA methylation of DJ-1 promoter region in PBLs level might not be an efficient biomarker for PD patients.

No MeSH data available.


Related in: MedlinePlus