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" זיהוי לא פולשני של תמותת תאים על ידי שימוש בתבנית המתילציה של דנ'א חופשי בדם "
Werman, Roni
Dor, Yuval
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Latin Dissertation
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English
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| Record Number
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1059105
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TL58222
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| Main Entry
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Werman, Roni
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| Title & Author
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זיהוי לא פולשני של תמותת תאים על ידי שימוש בתבנית המתילציה של דנ'א חופשי בדם\ Werman, RoniDor, Yuval
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| College
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The Hebrew University of Jerusalem (Israel)
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| Date
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2019
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Ph.D.
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2019
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| Note
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65 p.
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| Abstract
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Cell-free circulating DNA (cfDNA) released from dying cells is emerging as an important tool for the diagnosis of disease. However, the utility of cfDNA is limited in conditions in which involved tissues have a normal genome or unknown mutations. I have developed an approach to identify the tissue origins of human cfDNA, based on differential tissue-specific methylation patterns retained in circulating DNA fragments. We identified multiple CpG clusters whose methylation pattern is unique for specific cell types, and utilized this information to detect cfDNA derived from those tissues. The principles of the method are as follows. Differentially methylated CpGs are found using downloaded data from publicly available databases, or experiments done in our lab. Since methylation is a regional characteristic, we then expand the area we examine to CpGs surrounding it. We design primers that complement bisulfite-converted DNA in the area surrounding the located CpG. We then extract DNA from various tissues of the body, treat it with bisulfite, amplify the designated area with a PCR reaction and sequence it by next generation sequencing. We then analyze the data and after the area is ascertained as having methylation pattern specific to the tissue of interest, we extract cell-free DNA from serum/plasma samples of healthy controls and of patients with diseases involving cell death from that tissue and examine the methylation pattern in their DNA. We used the assay to examine several types of tissues and pathologies. We demonstrated the presence of brain derived DNA in the circulation of patients with multiple sclerosis during relapse, but not in healthy individuals. On the other hand, when examining liver derived cfDNA we found a basal level in healthy individuals, reflecting hepatic turnover. We also examined cfDNA from patients with pancreatic or colorectal cancer and were able not only to detect elevation of pancreatic and intestinal derived cfDNA, but also to distinguish between the cancers. This approach can be adapted to identify cfDNA derived from any cell type in the body, thus offering a minimally invasive window for monitoring and diagnosis of a broad spectrum of human pathologies, as well as a better understanding of normal tissue dynamics.
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| Descriptor
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Antibodies
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Apoptosis
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Biochemistry
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Biomarkers
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Biopsy
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Cellular biology
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Colorectal cancer
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DNA methylation
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Genes
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Genomes
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Haplotypes
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Lymphocytes
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Medical research
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Metastasis
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Mutation
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Neurons
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Pancreatic cancer
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Pancreatitis
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Pathology
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Plasma
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Tumors
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| Added Entry
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Dor, Yuval
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The Hebrew University of Jerusalem (Israel)
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