| Item type |
文献 / Documents(1) |
| 公開日 |
2024-10-10 |
| アクセス権 |
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アクセス権 |
open access |
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アクセス権URI |
http://purl.org/coar/access_right/c_abf2 |
| 資源タイプ |
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資源タイプ識別子 |
http://purl.org/coar/resource_type/c_6501 |
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資源タイプ |
journal article |
| 出版社版DOI |
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関連識別子 |
https://doi.org/10.1371/journal.pone.0047942 |
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関連名称 |
10.1371/journal.pone.0047942 |
| 出版タイプ |
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出版タイプ |
VoR |
|
出版タイプResource |
http://purl.org/coar/version/c_970fb48d4fbd8a85 |
| タイトル |
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タイトル |
Development of a Quantitative Methylation-Specific Polymerase Chain Reaction Method for Monitoring Beta Cell Death in Type 1 Diabetes |
| タイトル別表記 |
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その他のタイトル |
qMSP Assay for Monitoring Beta Cell Death |
| 著者 |
Husseiny, Mohamed I.
黒田, 暁生
Kaye, Alexander N.
Nair, Indu
Kandeel, Fouad
Ferreri, Kevin
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| 抄録 |
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内容記述 |
DNA methylation is a mechanism by which cells control gene expression, and cell-specific genes often exhibit unique patterns of DNA methylation. We previously reported that the mouse insulin-2 gene (Ins2) promoter has three potential methylation (CpG) sites, all of which are unmethylated in insulin-producing cells but methylated in other tissues. In this study we examined Ins2 exon 2 and found a similar tissue-specific methylation pattern. These methylation patterns can differentiate between DNA from insulin-producing beta cells and other tissues. We hypothesized that damaged beta cells release their DNA into circulation at the onset of type 1 diabetes mellitus (T1DM) and sought to develop a quantitative methylation-specific polymerase chain reaction (qMSP) assay for circulating beta cell DNA to monitor the loss of beta cells. Methylation-specific primers were designed to interrogate two or more CpG in the same assay. The cloned mouse Ins2 gene was methylated in vitro and used for development of the qMSP assay. We found the qMSP method to be sensitive and specific to differentiate between insulin-producing cells and other tissues with a detection limit of 10 copies in the presence of non-specific genomic DNA background. We also compared different methods for data analysis and found that the Relative Expression Ratio method is the most robust method since it incorporates both a reference value to normalize day-to-day variability as well as PCR reaction efficiencies to normalize between the methylation-specific and bisulfite-specific components of the calculations. The assay was applied in the streptozotocin-treated diabetic mouse model and detected a significant increase in circulating beta cell DNA before the rise in blood glucose level. These results demonstrate that this qMSP assay can be used for monitoring circulating DNA from insulin-producing cells, which will provide the basis for development of assays to detect beta cell destruction in early T1DM. |
| 書誌情報 |
en : PLOS ONE
巻 7,
号 10,
p. e47942,
発行日 2012-10-29
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| 収録物ID |
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収録物識別子タイプ |
EISSN |
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収録物識別子 |
19326203 |
| 出版者 |
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出版者 |
PLOS |
| 権利情報 |
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|
権利情報 |
This is an open-access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited. |
| EID |
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|
識別子 |
260654 |
| 言語 |
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言語 |
eng |
pone_7_10_e47942.pdf (625 KB)
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