Anatomy and Neuroscience - Theses

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    The interplay between clonal hematopoiesis and cardiometabolic diseases
    Bertuzzo Veiga, Camilla ( 2022)
    INTRODUCTION: Clonal haematopoiesis of indeterminate potential (CHIP) is a blood disorder arising from somatic mutations in haematopoietic stem and progenitor cells (HSPCs), providing these cells with a competitive advantage. This results in an increased abundance of mutant cells (>2%) in the blood. CHIP correlates with an increased risk of atherosclerotic cardiovascular disease (ACVD), proportional to the clonal expansion rate. However, the factors that control clonal outgrowth are not well understood. The most frequently mutated genes in CHIP are the epigenetic regulators DNMT3A and TET2. Dnmt3a driven-CHIP has been shown to aggravate disease progression in CVD, however the role of Dnmt3a-CHIP in ACVD remains limited. Importantly, diabetic patients present with higher incidence of CHIP. Diabetes induces TET2 loss-of-function via dysregulation of the metabolic sensor AMPK in mice and humans. In CHIP, DNMT3A and TET2 are commonly affected but their combined deficiency results in a myeloproliferative disorder. Thus, the hypothesis of this PhD thesis herein was two-fold. Firstly, that Dnmt3a-driven CHIP is accelerated in diabetes due to AMPK-TET2 dysregulation and re-activating this pathway will reverse DNMT3A clonal expansion in the setting of diabetes. Secondly, we hypothesized that Dnmt3a-driven CHIP will play a causal role in atherogenesis. METHODS AND RESULTS: In isolated BM HSPCs we observed that high glucose levels impaired the AMPK-TET2 axis, resulting in a reduction in DNA hydroxymethylation (5hmC), which was restored by activating AMPK. In a murine model of type 1 diabetes, we observed monocytosis, which was associated with TET2 dysfunction in BM HSPCs and blood myeloid cells. We next investigated how diabetes induced TET2 dysfunction would affect Dnmt3a-driven-CHIP. Using competitive bone marrow transplants (cBMT), we mimicked human mutant Dnmt3a-driven CHIP in WT mice and noted that diabetes exacerbates mutant myeloid clonal expansion due to enhanced myelopoiesis. Notably, BM HSPCs from mutant Dnmt3a CHIP diabetic mice exhibited TET2 dysfunction. Diabetes-driven TET2 dysfunction in HSPCs and DNMT3A-mutant myeloid clonal expansion were reversed by AMPK activation. Another metabolic state associated with CHIP is obesity, where AMPK activity is also reduced. We hypothesised that obesity would also induce TET2-dysfuntion. Obese models of Dnmt3a-CHIP had TET2 dysfunction in BM HSPCs along with monocyte clonal expansion and an overall decline and whole-body metabolism. This was associated with mutant macrophage infiltration in white adipose tissue and liver. Finally, we explored how Dnmt3a-CHIP affects atherosclerosis, and if clonal expansion was associated with worsened atherosclerotic outcome. Indeed, in a plaque progression model we confirmed that Dnmt3a-CHIP was associated with myeloid clonal expansion and enhanced thrombopoiesis. The increased platelet counts were associated with mutant macrophage infiltration into the livers and higher levels of TPO in the plasma. Dnmt3a-CHIP mice presented increased mutant monocytes, neutrophils and macrophages in the aortic arch which was paralleled with increased lesion size and lipid content. CONCLUSION: Herein we show that diabetes dysregulates the AMPK-TET2 axis in BM HSPCs and promotes myeloid clonal expansion in Dnmt3a-CHIP. Importantly, activating AMPK restores TET2 function and reduced clonal expansion in diabetes. CHIP associated with Dnmt3a deficiency was shown to accelerate atherosclerosis progression due to pronounced myeloid clonal expansion and thrombopoiesis. These studies suggest that targeting AMPK-TET2 pathway may provide a novel approach to limit clonal outgrowth in diabetic patients with CHIP.