Introduction to Cardiovascular Aging Compendium : “Aging as the Quintessential Complex Phenotype”

Rationale Clonal hematopoiesis has been associated with increased mortality and cardiovascular disease (CVD). This condition can arise from somatic mutations in pre-leukemic driver genes within hematopoietic stem/progenitor cells (HSPC). Approximately 40 candidate driver genes have been identified, but mutations in only one of these genes, Ten-Eleven Translocation-2 (TET2), has been shown to casually contribute to CVD in murine models. Objective To develop a facile system to evaluate the disease characteristics of different clonal hematopoiesis driver genes using lentivirus vector and CRISPR/Cas9 methodology. Using this methodology, evaluate whether DNA (Cytosine-5)-methyltransferase 3a (Dnmt3a), a commonly occurring clonal hematopoiesis driver gene, causally contributes to CVD. Methods and Results Lentivirus vectors were used to deliver Cas9 and guide RNA (gRNA) to introduce inactivating mutations in Tet2 and Dnmt3a in lineage-negative bone marrow cells. Following implantation into lethally-irradiated mice, these cells were engrafted and gave rise to labeled blood cell progeny. When challenged with an infusion of angiotensin II (AngII), mice with inactivating mutations in Tet2 or Dnmt3a displayed greater cardiac hypertrophy, diminished cardiac function, and greater cardiac and renal fibrosis. In comparison to Tet2, inactivation of Dnmt3a did not lead to detectable expansion of the mutant hematopoietic cells during the time course of these experiments. Tet2 inactivation promoted the expression of IL1β, IL-6 and Ccl5 whereas Dnmt3a inactivation promoted the expression of Cxcl1, Cxcl2, IL-6 and Ccl5 in a LPS-stimulated macrophage cell line. Conclusions Experiments employing lentivirus vector/CRISPR methodology provided evidence suggesting that inactivating DNMT3A mutations in hematopoietic cells contributes to CVD. Comparative analyses showed that inactivation of Tet2 and Dnmt3 were similar in their ability to promote AngII-induced cardiac dysfunction and renal fibrosis in mice. However, gene-specific actions were indicated by differences in kinetics of HSPC expansion and different patterns of inflammatory gene expression.

The study of aging relates to changes in physical and functional dimensions that occur over time in living organisms. Yet, a model that establishes the hierarchical relationship and interlaced time courses of molecular, phenotypic, and functional hierarchical domains of aging in humans has not been established. We propose that studying the mechanisms and consequences of aging through the lens of these hierarchical domains and their connections will provide clarity in semantics and enhance a translational perspective. The study of human aging would be most informative from a life course, longitudinal perspective, given that manifestations of aging are already detectable early in life at the molecular level, yet the phenotypic responses remain masked by compensatory/resiliency mechanisms. Understanding the nature of these mechanisms is paramount for developing interventions that reduce the burden of disease and disability in older persons.