Clonal Hematopoiesis Driven By Mutations in DNMT3A Promotes Metabolic Disease

Background: Clonal hematopoiesis (CH) is characterized by the clonal expansion of hematopoietic stem cells (HSCs) marked with somatic mutations and of their progeny in the absence of quantitative blood abnormalities. In addition to an elevated risk of developing blood malignancies, CH is notably associated with various chronic diseases beyond the hematopoietic system, such as inflammatory disorders and cardiovascular disease. DNA methyltransferase 3A (DNMT3A) is the most commonly mutated gene in CH (up to 40% of all cases). While the DNMT3A Arg882His (RH) hotspot variant is enriched in acute myeloid leukemia (AML), most DNMT3A alterations observed in CH are consistent with the loss-of-function (LOF), indicating these types of mutations may not be fully mechanistically and functionally equivalent. However, the specific impact of CH driven by different types of DNMT3A mutations on chronic disease development has not been investigated.

Methods: To elucidate the functional impact of CH with DNMT3A LOF and RH mutations in the hematopoietic system on metabolic disease development, we created a murine bone marrow transplantation (BMT)-based chimeric model with 20% of either Dnmt3a^+/-, Dnmt3a^+/RH or wild-type (WT) control cells (CD45.2) mixed with 80% WT support (CD45.1). To induce metabolic disease, mice were maintained on high-fat high-carbohydrate “western” diet (HFD) over 5 months or remained on normal chow as control.

Results: We found that animals harboring a proportion of Dnmt3a^+/- or Dnmt3a^+/RH cells in the bone marrow gained more body weight even on normal chow. This was accompanied by increased food intake and elevated serum leptin levels compared to WT controls; the size of fat cells within white adipose tissue was consistently enlarged. Notably, these effects were more pronounced in the Dnmt3a^+/- group compared to the Dnmt3a^+/RH CH animals. These effects were dramatically exacerbated on HFD. Importantly, these mice exhibited higher fasting glucose levels and impaired glucose tolerance indicative of an increased tendency to developing diet-induced diabetes.

Lineage composition analysis of the blood system revealed Dnmt3a^+/- cells showed increased abundance of inflammatory monocytes in peripheral circulation and spleen compared to WT cells. This was further amplified by HFD in both Dnmt3a^+/- and Dnmt3a^+/RH CH mice, with Dnmt3a^+/- cells displaying a more robust pro-inflammatory profile. Notably, this phenotype was present in both CH (CD45.2) and WT (CD45.1) cells within the same animal reflecting a generalized pro-inflammatory state. Consistently, Dnmt3a^+/- CH mice had elevated levels of serum pro-inflammatory cytokines such as IL-1α, IL-15 and MIP-1α, compared to WT controls, especially under HFD conditions.

The generalized pro-inflammatory state was further evident in the livers of both Dnmt3a^+/- and Dnmt3a^+/RH CH animals that exhibited signs of lobular hepatitis with prominent leukocyte infiltration even on normal chow compared to WT controls. Dnmt3a^+/- CH mice further developed hepatic steatosis characterized by an increased number and size of macrovesicular fat droplets, which progressed to severe chronic liver disease under HFD evidenced by more pronounced liver inflammation, extensive fibrosis, and a high non-alcoholic fatty liver disease activity score (NAS).

Conclusions: These findings indicate that in a mouse model, the presence of Dnmt3a LOF or RH CH promotes the development of obesity, impaired glucose metabolism, and chronic inflammatory liver disease. These metabolic diseases are further exacerbated by a high-fat high-carbohydrate diet, most strongly in the Dnmt3a^+/- context. Our findings highlight the differential impact of different CH mutations and urge detailed mutation- and gene-specific investigation of CH in human chronic disease pathogenesis, risk stratification, and mitigation through dietary interventions.