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dc.contributor.advisorSapienza, Carmen
dc.creatorVaidya, Himani
dc.date.accessioned2022-05-26T18:04:09Z
dc.date.available2022-05-26T18:04:09Z
dc.date.issued2022
dc.identifier.urihttp://hdl.handle.net/20.500.12613/7654
dc.description.abstractEpigenetic marks encoded by DNA methylation, drifts with age, this results in gain of methylation in CpG islands and loss of methylation in repeat regions. This drift correlates with lifespan and is conserved across species mice, rhesus monkeys, and humans. However, the biology of how this drift occurs is still unexplained. Our hypothesis is that epigenetic drift occurs in aging stem cells. i.e. young stem cells have a stable, uniform DNA methylation pattern. However, as stem cells age, due to cell division and environmental effects, old stem cells have an unstable mosaic methylation pattern which leads to stem cell depletion and exhaustion, focal proliferation, etc., causing cancer and various age-related diseases. To evaluate this hypothesis, we analyzed DNA methylation pattern in mouse colon intestinal stem cells (Lgr5-gfp+) mice from four different age groups (4,12,18, and 24-months) against nonstem cells (Lgr5-GFP-). We found that there are not many methylation changes between stem cells and nonstem cells of the same age (0% change in 4 month and +- 0.2% in 24-month-old samples). This demonstrates that DNA methylation changes primarily occur in stem cells. Permutation analysis of all four aging time points (stem and nonstem) revealed 8102 CpG sites that changed methylation with age, with hypermethylated CpG sites in the promoter-nonCpG islands having the highest odds of gaining methylation with age. CpG sites that changed in DNA methylation levels between tissues (colon and small intestine) were also analyzed through permutation testing. CpG sites that changed between tissues did not overlap with age, showing that CpG sites that change with age and differentiation are different. Gene expression changes with aging was also analyzed, however, compared to DNA methylation not many genes changed in expression with age. We compared the correlation between levels of DNA methylation and gene expression. Low levels of methylation (0-10%) had higher levels of gene expression and increase in DNA methylation led to a decrease in RNA expression. We compared gene expression with CpG sites that change significantly with age in the promoter region from the permutation analysis and found that gene expression is correlated with DNA methylation in CpG sites in promoter-CpG island (r= -0.18). We used principles of entropy and information theory to better analyze the pattern of methylation drift with age in the intestine and found that entropy, measured through Jensen-Shannon Distance (JSD), increased with age but not with differentiation from stem to nonstem cells and the entropy increased 2.5-3 folds in the old when compared to the young. We extended this analysis to other organs with different rates of cell division and found that entropy does not change with age in organs with low rate of cell division such as the heart and kidney. Thus, the rate of cell division and change in entropy with age are highly correlated (r=0.89, p =<0.001). This demonstrates that we can use DNA methylation entropy to measure stem cell replication and aging. We also did not find many loci that changed with age overlapping across tissues, showing that change in methylation entropy is tissue specific. Thus, our data suggests that tissue specific clocks that measure entropy may provide a more accurate measurement of methylation age when compared to clocks based solely on percent methylation. Finally, we looked at DNA methylation changes in intestinal organoids and compared them to primary stem and nonstem cells and found a very weak correlation between them (r=0.23, p=<0.001) and no correlation as the organoids are passaged. We assessed DNA methylation changes caused by long-term passaging (28 weeks). DNA methylation changes with passage, in 4-month and 24-month upper small intestine (USI) organoids, was compared to aging in USI primary cells and we found a weak correlation with 4-month organoids (r=0.28, p=<0.001) and no correlation in 24-month organoids passaged long-term. This demonstrates there are changes in DNA methylation when transitioning from in vivo to in vitro and could result in gene expression and function of the organoid. More studies need to be done to assess the impact of these changes.
dc.format.extent128 pages
dc.language.isoeng
dc.publisherTemple University. Libraries
dc.relation.ispartofTheses and Dissertations
dc.rightsIN COPYRIGHT- This Rights Statement can be used for an Item that is in copyright. Using this statement implies that the organization making this Item available has determined that the Item is in copyright and either is the rights-holder, has obtained permission from the rights-holder(s) to make their Work(s) available, or makes the Item available under an exception or limitation to copyright (including Fair Use) that entitles it to make the Item available.
dc.rights.urihttp://rightsstatements.org/vocab/InC/1.0/
dc.subjectCellular biology
dc.subjectAging
dc.subjectGenetics
dc.subjectAging
dc.subjectDNA methylation
dc.subjectStem cell
dc.titleDNA METHYLATION ENTROPY AS A MEASURE OF STEM CELL REPLICATION AND AGING
dc.typeText
dc.type.genreThesis/Dissertation
dc.contributor.committeememberIssa, Jean-Pierre
dc.contributor.committeememberGra�a-Amat, Xavier
dc.contributor.committeememberTempera, Italo
dc.contributor.committeememberWhelan, Kelly A.
dc.contributor.committeememberSawaya, Bassel E.
dc.description.departmentBiomedical Sciences
dc.relation.doihttp://dx.doi.org/10.34944/dspace/7626
dc.ada.noteFor Americans with Disabilities Act (ADA) accommodation, including help with reading this content, please contact scholarshare@temple.edu
dc.description.degreePh.D.
dc.identifier.proqst14757
dc.creator.orcid0000-0002-2779-3069
dc.date.updated2022-05-11T16:08:23Z
refterms.dateFOA2022-05-26T18:04:10Z
dc.identifier.filenameVaidya_temple_0225E_14757.pdf


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