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DNA Methylation Underlies the Long-Term Association Between Oral Porphyromonas gingivalis infection and Atherosclerotic Cardiovascular Disease
- Author / Creator
- Omar, Mohamed
Periodontitis, one of the most common inflammatory conditions, and the leading cause of teeth loss in adults, has been associated with cardiovascular disease (CVD) for decades, and has recently been identified, by the American Heart Association, as an independent risk factor for atherosclerotic CVD. Importantly, periodontitis is a chronic condition without a definitive treatment endpoint, and patients are managed by regular scaling and root planning. This inherent limitation renders the assessment of the effect of periodontitis treatment on the increased risk of CVD quite challenging. However, long-term studies in edentulous patients with history of periodontitis, showed persistence of the increased CVD risk for years after edentulism, suggesting that clinical elimination of the disease might not be sufficient in decreasing the periodontitis-induced CVD risk. In this project we sought to explore the validity and nature of this suggested long-term association.
We hypothesized that periodontitis induces some epigenetic changes, in the form of DNA-methylation, in hematopoietic stem cells in the bone marrow, and such changes persist after clinical elimination of the disease, and underlie the induced-CVD risk.
Using the atherosclerotic mouse model, the low-density lipoprotein receptor Knock out (LDLR-/-), all mice were fed a high fat diet (HFD) to induce atherosclerosis. Oral inoculation with Porphyromonas gingivalis (pg), a key stone periodontal pathogen, was used to induce periodontitis in one group of mice, whereas the other group was sham inoculated. We have previously shown that mice that were fed a HFD and inoculated with Pg developed more atherosclerosis than mice that were sham-inoculated. To simulate clinical elimination of periodontitis and persistence of the hypothesized epigenetic reprogramming, we used a bone marrow transplant approach. Naïve LDLR-/- mice were irradiated and transplanted with bone marrow cells form HFD fed mice that were either Pg-inoculated or sham-inoculated, creating two groups of recipient mice. Aorta morphometry showed that, like the donor mice, mice recipient of bone marrow from Pg-inoculated donors developed significantly more atherosclerosis compared to mice recipient of bone marrow from sham-inoculated donors. This increase in atherosclerosis was accompanied with a more pro-inflammatory plasma and macrophage cytokine profile. We hypothesize that changes in DNA methylation, could in part, underlie the observed increase in atherosclerosis. This was tested using whole genome bisulphite sequencing (WGBS) approach, the gold standard for a thorough and quantitative assessment for DNA methylation of the entire genome. Our data show a significant difference in the methylation profile of peritoneal macrophages between the two groups of recipient mice. There was 375 identified differentially methylated regions (DMRs) between the groups, with global hypomethylation in mice recipients of bone marrow from Pg-inoculated donors. Some of the DMRs pointed to the involvement of enzymes with major roles in the methylation and demethylation process. In mice recipients of bone marrow from Pg-inoculated donors, methionine adenosyl transferase (MAT), which catalyzes the conversion of methionine to S-adenosylmethionine (SAM), the universal methyl-donor, was hypermethylated. The protein coding gene for S-adenosylhomocysteine hydrolase (SAHH), which catalyzes the reversible conversion of the potent methylation inhibitor, S-adenosylhomocysteine (SAH), to homocysteine and adenosine was also hypermethylated. In contrast, the de-methylation enzyme, Ten-Eleven Translocase (TET 2), was hypomethylated. Accordingly, we sought to investigate the mechanism underlying the observed global hypomethylation and its potential association with the observed increase in atherosclerosis. In line with the global hypomethylation, our activity assays showed a significant increase in TET activity and a decrease in DNA methyltransferases (DNMT). In addition, plasma SAH levels were significantly higher and SAM to SAH ratio was decreased, both of which have been associated with CVD and with increased risk of myocardial infarction and stroke.
Homocysteine, is located at the intersection of the methionine cycle and the trans-sulphuration pathway, where it can either be re-methylated to methionine and go through the methionine cycle, or it can synthesize cysteine through the trans-sulphuration pathway. Glutathione (GSH), a critical antioxidant, is the end-product of the trans-sulphuration pathway. Hence, as a coping mechanism, the trans-sulphuration pathway is favoured over the methionine cycle under conditions of oxidative stress. Importantly, periodontitis has been repeatedly shown to induce oxidative stress.
In this dissertation, we propose that periodontitis-induced oxidative stress leads to global DNA hypomethylation through disruption of the methionine cycle, and that the resultant increase in SAH increases the risk of atherosclerotic CVD.
- Graduation date
- Spring 2022
- Type of Item
- Doctor of Philosophy
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