Lifelong longitudinal assessment of the contribution of multi-fractal fluctuations to heart rate and heart rate variability in aging mice: role of the sinoatrial node and autonomic nervous system.

Aging is a major risk factor for sinoatrial node (SAN) dysfunction, which can impair heart rate (HR) control and heart rate variability (HRV). HR and HRV are determined by intrinsic SAN function and its regulation by the autonomic nervous system (ANS). The purpose of this study was to use multi-scale multi-fractal detrended fluctuation analysis (MSMFDFA; a complexity-based approach to analyze multi-fractal dynamics) to longitudinally assess changes in multi-fractal HRV properties and SAN function in ECG time series recorded repeatedly across the full adult lifespan in mice. ECGs were recorded in anesthetized mice in baseline conditions and after autonomic nervous system blockade every three months beginning at 6 months of age until the end of life. MSMFDFA was used to assess HRV and SAN function every three months between 6 and 27 months of age. Intrinsic HR (i.e. HR during ANS blockade) remained relatively stable until 15 months of age, and then progressively declined until study endpoint at 27 months of age. MSMFDFA revealed sudden and rapid changes in multi-fractal properties of the ECG RR interval time series in aging mice. In particular, multi-fractal spectrum width (MFSW, a measure of multi-fractality) was relatively stable between 6 months and 15 months of age and then progressively increased at 27 months of age. These changes in MFSW were evident in baseline conditions and during ANS blockade. Thus, intrinsic SAN function declines progressively during aging and is manifested by age-associated changes in multi-fractal HRV across the lifespan in mice, which can be accurately quantified by MSMFDFA.

The Longitudinal Decline in Peak VO2 with Aging in Healthy Individuals is Associated with a Reduction in Peripheral Oxygen Utilization but not in Cardiac Output.

Aging is associated with a significant decline in exercise fitness assessed by maximal exercise oxygen consumption (VO2-max). The specific VO2-max components driving this decline, namely cardiac output (CO) and arteriovenous oxygen difference (A-V) O2, remain unclear. We examined this issue by analyzing data from 99 community-dwelling participants (baseline age 21-96 years; average follow-up 12.6 years) from the Baltimore Longitudinal Study of Aging, free of clinical cardiovascular disease. VO2-peak, a surrogate of VO2-max, was used to assess aerobic capacity during upright cycle exercise. Peak exercise left ventricular (LV) volumes, heart rate, and cardiac output were estimated using repeated gated cardiac blood pool scans. The Fick equation was used to calculate (A-V) O2-peak from CO-peak and VO2-peak. In unadjusted models, VO2-peak, (A-V) O2-peak, and CO-peakdeclined longitudinally over time at steady rates with advancing age. In multiple linear regression models adjusting for baseline values and peak workload, however, steeper declines in VO2-peak and (A-V) O2 peak were observed with advanced entry age but not in CO-peak. The association between the declines in VO2-peak and (A-V) O2-peakwas stronger among those >=50 years compared to their younger counterparts but the difference between the two age groups did not reach statistical significance. These findings suggest that age-associated impairment of peripheral oxygen utilization during maximal exercise poses a stronger limitation on peak VO2 than that of CO. Future studies examining interventions targeting the structure and function of peripheral muscles and their vasculature to mitigate age-associated declines in (A-V) O2 are warranted.

Echocardiographic characterization of age- and sex-associated differences in cardiac function and morphometry in nonhuman primates.

Aging per se is a major risk factor for cardiovascular diseases and is associated with progressive changes in cardiac structure and function. Rodent models are commonly used to study cardiac aging, but do not closely mirror differences as they occur in humans. Therefore, we performed a 2D echocardiographic study in non-human primates (NHP) to establish age- and sex-associated differences in cardiac function and morphometry in this animal model. M mode and 2D echocardiography and Doppler analyses were performed cross-sectionally in 38 healthy rhesus monkeys (20 females and 18 males), both young (age 7-12 years; n = 20) and old (age 19-30 years; n = 18). The diameters of the cardiac chambers did not differ significantly by age group, but males had larger left ventricular diameters (2.43 vs 2.06 cm in diastole and 1.91 vs 1.49 cm in systole, p = 0.0004 and p = 0.0001, respectively) and left atrial diameter (1.981 vs 1.732 cm; p = 0.0101). Left ventricular mass/body surface area did not vary significantly with age and sex. Ejection fraction did not differ by age and females presented a higher ejection fraction than males (54.0 vs 50.8%, p = 0.0237). Diastolic function, defined by early to late mitral peak flow velocity ratio (E/A), was significantly lower in old rhesus monkeys (2.31 vs 1.43, p = 0.0020) and was lower in females compared to males (1.595 vs 2.230, p = 0.0406). Right ventricular function, evaluated by measuring the Tricuspid Annular Plane Systolic Excursion, did not differ by age or sex, and Right Ventricular Free Wall Longitudinal Strain, did not differ with age but was lower in males than in females (-22.21 vs -17.95%, p = 0.0059). This is the first echocardiographic study to evaluate age- and sex-associated changes of cardiac morphometry and function in young and old NHP. The findings of this work will provide a reference to examine the effect of age and sex on cardiac diseases in NHP.

RelA-mediated signaling connects adaptation to chronic cardiomyocyte stress with myocardial and systemic inflammation in the ADCY8 model of accelerated aging.

Mice with cardiac-specific overexpression of adenylyl cyclase (AC) type 8 (TGAC8) are under a constant state of severe myocardial stress. They have a remarkable ability to adapt to this stress, but they eventually develop accelerated cardiac aging and experience reduced longevity. We have previously demonstrated through bioinformatics that constitutive adenylyl cyclase activation in TGAC8 mice is associated with the activation of inflammation-related signaling pathways. However, the immune response associated with chronic myocardial stress in the TGAC8 mouse remains unexplored. Here we demonstrate that chronic activation of adenylyl cyclase in cardiomyocytes of TGAC8 mice results in activation of cell-autonomous RelA-mediated NF-κB signaling. This is associated with non-cell-autonomous activation of proinflammatory and age-associated signaling in myocardial endothelial cells and myocardial smooth muscle cells, expansion of myocardial immune cells, increase in serum levels of inflammatory cytokines, and changes in the size or composition of lymphoid organs. All these changes precede the appearance of cardiac fibrosis. We provide evidence indicating that RelA activation in cardiomyocytes with chronic activation of adenylyl cyclase is mediated by calcium-protein Kinase A (PKA) signaling. Using a model of chronic cardiomyocyte stress and accelerated aging, we highlight a novel, calcium/PKA/RelA-dependent connection between cardiomyocyte stress, myocardial inflammation, and systemic inflammation. These findings suggest that RelA-mediated signaling in cardiomyocytes might be an adaptive response to stress that, when chronically activated, ultimately contributes to both cardiac and systemic aging.

Reprogramming of cardiac phosphoproteome, proteome, and transcriptome confers resilience to chronic adenylyl cyclase-driven stress.

Our prior study (Tarasov et al., 2022) discovered that numerous adaptive mechanisms emerge in response to cardiac-specific overexpression of adenylyl cyclase type 8 (TGAC8) which included overexpression of a large number of proteins. Here, we conducted an unbiased phosphoproteomics analysis in order to determine the role of altered protein phosphorylation in the adaptive heart performance and protection profile of adult TGAC8 left ventricle (LV) at 3-4 months of age, and integrated the phosphoproteome with transcriptome and proteome. Based on differentially regulated phosphoproteins by genotype, numerous stress-response pathways within reprogrammed TGAC8 LV, including PKA, PI3K, and AMPK signaling pathways, predicted upstream regulators (e.g. PDPK1, PAK1, and PTK2B), and downstream functions (e.g. cell viability, protein quality control), and metabolism were enriched. In addition to PKA, numerous other kinases and phosphatases were hyper-phosphorylated in TGAC8 vs. WT. Hyper-phosphorylated transcriptional factors in TGAC8 were associated with increased mRNA transcription, immune responses, and metabolic pathways. Combination of the phosphoproteome with its proteome and with the previously published TGAC8 transcriptome enabled the elucidation of cardiac performance and adaptive protection profiles coordinately regulated at post-translational modification (PTM) (phosphorylation), translational, and transcriptional levels. Many stress-response signaling pathways, i.e., PI3K/AKT, ERK/MAPK, and ubiquitin labeling, were consistently enriched and activated in the TGAC8 LV at transcriptional, translational, and PTM levels. Thus, reprogramming of the cardiac phosphoproteome, proteome, and transcriptome confers resilience to chronic adenylyl cyclase-driven stress. We identified numerous pathways/function predictions via gene sets, phosphopeptides, and phosphoproteins, which may point to potential novel therapeutic targets to enhance heart adaptivity, maintaining heart performance while avoiding cardiac dysfunction.

Echocardiographic heart ageing patterns predict cardiovascular and non-cardiovascular events and reflect biological age: the SardiNIA study.

AIMS: Age is a crucial risk factor for cardiovascular (CV) and non-CV diseases. As people age at different rates, the concept of biological age has been introduced as a personalized measure of functional deterioration. Associations of age with echocardiographic quantitative traits were analysed to assess different heart ageing rates and their ability to predict outcomes and reflect biological age. METHODS AND RESULTS: Associations of age with left ventricular mass, geometry, diastolic function, left atrial volume, and aortic root size were measured in 2614 healthy subjects. Based on the 95% two-sided tolerance intervals of each correlation, three discrete ageing trajectories were identified and categorized as 'slow', 'normal', and 'accelerated' heart ageing patterns. The primary endpoint included fatal and non-fatal CV events, and the secondary endpoint was a composite of CV and non-CV events and all-cause death. The phenotypic age of the heart (HeartPhAge) was estimated as a proxy of biological age. The slow ageing pattern was found in 8.7% of healthy participants, the normal pattern in 76.9%, and the accelerated pattern in 14.4%. Kaplan-Meier curves of the heart ageing patterns diverged significantly (P = 0.0001) for both primary and secondary endpoints, with the event rate being lowest in the slow, intermediate in the normal, and highest in the accelerated pattern. In the Cox proportional hazards model, heart ageing patterns predicted both primary (P = 0.01) and secondary (P = 0.03 to <0.0001) endpoints, independent of chronological age and risk factors. Compared with chronological age, HeartPhAge was 9 years younger in slow, 4 years older in accelerated (both P < 0.0001), and overlapping in normal ageing patterns. CONCLUSION: Standard Doppler echocardiography detects slow, normal, and accelerated heart ageing patterns. They predict CV and non-CV events, reflect biological age, and provide a new tool to calibrate prevention timing and intensity.

Age is the main risk factor for cardiovascular (CV) disease. Since people age and develop diseases at very different rates, biological age has been proposed as a more accurate measure of the body's functional decline. This study aimed to investigate the ageing rates of the heart and to assess their impact on CV events. The phenotypic age of the heart was also estimated as a proxy for biological age. Associations of age with Doppler echocardiographic parameters were analysed in a subgroup of 2614 clinically healthy subjects, part of a larger cohort of 3817 adults of both sexes.Three patterns of slow, normal, and accelerated ageing rates of the heart were detected. They predicted both CV and non-CV events, with different and progressively increasing event rates from the slow to the accelerated pattern. Compared with chronological age, the phenotypic (biological) age of the heart was 9 years younger in the slow pattern, 4 years older in the accelerated pattern, and comparable in the normal pattern.A standard Doppler echocardiogram is therefore able to detect three distinct heart ageing patterns, which reflect different biological susceptibilities to age-dependent diseases and provide a new tool for personalizing timeliness and intensity of prevention.

Glucose-6-phosphate dehydrogenase deficiency accelerates arterial aging in diabetes.

AIMS: High glucose levels and Glucose-6-Phosphate Dehydrogenase deficiency (G6PDd) have both tissue inflammatory effects. Here we determined whether G6PDd accelerates arterial aging (information linked stiffening) in diabetes. METHODS: Plasma glucose, interleukin 6 (IL6), and arterial stiffness (indexed as carotid-femoral Pulse Wave Velocity, PWV) and red blood cell G6PD activity were assessed in a large (4448) Sardinian population. RESULTS: Although high plasma glucose in diabetics, did not differ by G6DP status (178.2 ± 55.1 vs 169.0 ± 50.1 mg/dl) in G6DPd versus non-G6PDd subjects, respectively, IL6, and PWV (adjusted for age and glucose) were significantly increased in G6PDd vs non-G6PDd subjects (PWV, 8.0 ± 0.4 vs 7.2 ± 0.2 m/sec) and (IL6, 6.9 ± 5.0 vs 4.2 ± 3.0 pg/ml). In non-diabetics, neither fasting plasma glucose, nor IL6, nor PWV were impacted by G6PDd. CONCLUSION: G6PDd in diabetics is associated with increased inflammatory markers and accelerated arterial aging.

A novel conceptual model of heart rate autonomic modulation based on a small-world modular structure of the sinoatrial node.

The present view on heartbeat initiation is that a primary pacemaker cell or a group of cells in the sinoatrial node (SAN) center paces the rest of the SAN and the atria. However, recent high-resolution imaging studies show a more complex paradigm of SAN function that emerges from heterogeneous signaling, mimicking brain cytoarchitecture and function. Here, we developed and tested a new conceptual numerical model of SAN organized similarly to brain networks featuring a modular structure with small-world topology. In our model, a lower rate module leads action potential (AP) firing in the basal state and during parasympathetic stimulation, whereas a higher rate module leads during ß-adrenergic stimulation. Such a system reproduces the respective shift of the leading pacemaker site observed experimentally and a wide range of rate modulation and robust function while conserving energy. Since experimental studies found functional modules at different scales, from a few cells up to the highest scale of the superior and inferior SAN, the SAN appears to feature hierarchical modularity, i.e., within each module, there is a set of sub-modules, like in the brain, exhibiting greater robustness, adaptivity, and evolvability of network function. In this perspective, our model offers a new mainframe for interpreting new data on heterogeneous signaling in the SAN at different scales, providing new insights into cardiac pacemaker function and SAN-related cardiac arrhythmias in aging and disease.

Transcriptome of Left Ventricle and Sinoatrial Node in Young and Old C57 Mice.

Advancing age is the most important risk factor for cardiovascular diseases (CVDs). Two types of cells, within the heart pacemaker, sinoatrial node (SAN), and within the left ventricle (LV), control two crucial characteristics of heart function, heart beat rate and contraction strength. As age advances, the heart's structure becomes remodeled, and SAN and LV cell functions deteriorate, thus increasing the risk for CVDs. However, the different molecular features of age-associated changes in SAN and LV cells have never been compared in omics scale in the context of aging. We applied deep RNA sequencing to four groups of samples, young LV, old LV, young SAN and old SAN, followed by numerous bioinformatic analyses. In addition to profiling the differences in gene expression patterns between the two heart chambers (LV vs. SAN), we also identified the chamber-specific concordant or discordant age-associated changes in: (1) genes linked to energy production related to cardiomyocyte contraction, (2) genes related to post-transcriptional processing, (3) genes involved in KEGG longevity regulating pathway, (4) prolongevity and antilongevity genes recorded and curated in the GenAge database, and (5) CVD marker genes. Our bioinformatic analysis also predicted the regulation activities and mapped the expression of upstream regulators including transcription regulators and post-transcriptional regulator miRNAs. This comprehensive analysis promotes our understanding of regulation of heart functions and will enable discovery of gene-specific therapeutic targets of CVDs in advanced age.

Cardiomyocyte-specific adenylyl cyclase type-8 overexpression induces activation of RelA together with myocardial and systemic inflammation.

Background: Mice with cardiac-specific overexpression of adenylyl cyclase (AC) type 8 (TG AC8 ) are under a constant state of severe myocardial stress. They have a remarkable ability to adapt to this stress, but they eventually develop accelerated cardiac aging and experience reduced longevity. Results: Here we demonstrate that activation of ACVIII in cardiomyocytes results in cell-autonomous RelA-mediated NF-κB signaling. This is associated with non-cell-autonomous activation of proinflammatory and age-associated signaling in myocardial endothelial cells and myocardial smooth muscle cells, expansion of myocardial immune cells, increase in serum levels of inflammatory cytokines, and changes in the size or composition of lymphoid organs. These changes precede the appearance of cardiac fibrosis. We provide evidence indicating that ACVIII-driven RelA activation in cardiomyocytes is mediated by calcium-Protein Kinase A (PKA) signaling. Conclusions: Using a model of chronic cardiomyocyte stress and accelerated aging we highlight a novel, PKA/RelA-dependent connection between cardiomyocyte stress, myocardial para-inflammation and systemic inflammation. These findings point to RelA-mediated signaling in cardiomyocytes and inter-organ communication between the heart and lymphoid organs as novel potential therapeutic targets to reduce age-associated myocardial deterioration.

Enhanced Myocardial Adenylyl Cyclase Activity Alters Heart-Brain Communication.

BACKGROUND: The central nervous system's influence on cardiac function is well described; however, direct evidence for signaling from heart to brain remains sparse. Mice with cardiac-selective overexpression of adenylyl cyclase type 8 (TGAC8) display elevated heart rate/contractility and altered neuroautonomic surveillance. OBJECTIVES: In this study the authors tested whether elevated adenylyl cyclase type 8-dependent signaling at the cardiac cell level affects brain activity and behavior. METHODS: A telemetry system was used to record electrocardiogram (ECG) and electroencephalogram (EEG) in TGAC8 and wild-type mice simultaneously. The Granger causality statistical approach evaluated variations in the ECG/EEG relationship. Mouse behavior was assessed via elevated plus maze, open field, light-dark box, and fear conditioning tests. Transcriptomic and proteomic analyses were performed on brain tissue lysates. RESULTS: Behavioral testing revealed increased locomotor activity in TGAC8 that included a greater total distance traveled (+43%; P < 0.01), a higher average speed (+38%; P < 0.01), and a reduced freezing time (-45%; P < 0.01). Dual-lead telemetry recording confirmed a persistent heart rate elevation with a corresponding reduction in ECG-R-waves interval variability and revealed increased EEG-gamma activity in TGAC8 vs wild-type. Bioinformatic assessment of hippocampal tissue indicated upregulation of dopamine 5, gamma-aminobutyric acid A, and metabotropic glutamate 1/5 receptors, major players in gamma activity generation. Granger causality analyses of ECG and EEG recordings showed a marked increase in informational flow between the TGAC8 heart and brain. CONCLUSIONS: Perturbed signals arising from the heart cause changes in brain activity, altering mouse behavior. More specifically, the brain interprets augmented myocardial humoral/functional output as a "sustained exercise-like" situation and responds by activating central nervous system output controlling locomotion.

Silencing of PKG1 Gene Mimics Effect of Aging and Sensitizes Rat Vascular Smooth Muscle Cells to Cardiotonic Steroids: Impact on Fibrosis and Salt Sensitivity.

Background Marinobufagenin, NKA (Na/K-ATPase) inhibitor, causes vasoconstriction and induces fibrosis via inhibition of Fli1 (Friend leukemia integration-1), a negative regulator of collagen synthesis. In vascular smooth muscle cells (VSMC), ANP (atrial natriuretic peptide), via a cGMP/PKG1 (protein kinase G1)-dependent mechanism, reduces NKA sensitivity to marinobufagenin. We hypothesized that VSMC from old rats, due to downregulation of ANP/cGMP/PKG-dependent signaling, would exhibit heightened sensitivity to the profibrotic effect of marinobufagenin. Methods and Results Cultured VSMC from the young (3-month-old) and old (24-month-old) male Sprague-Dawley rats and young VSMC with silenced PKG1 gene were treated with 1 nmol/L ANP, or with 1 nmol/L marinobufagenin, or with a combination of ANP and marinobufagenin. Collagen-1, Fli1, and PKG1 levels were assessed by Western blotting analyses. Vascular PKG1 and Fli1 levels in the old rats were reduced compared with their young counterparts. ANP prevented inhibition of vascular NKA by marinobufagenin in young VSMC but not in old VSMC. In VSMC from the young rats, marinobufagenin induced downregulation of Fli1 and an increase in collagen-1 level, whereas ANP blocked this effect. Silencing of the PKG1 gene in young VSMC resulted in a reduction in levels of PKG1 and Fli1; marinobufagenin additionally reduced Fli1 and increased collagen-1 level, and ANP failed to oppose these marinobufagenin effects, similar to VSMC from the old rats with the age-associated reduction in PKG1. Conclusions Age-associated reduction in vascular PKG1 and the resultant decline in cGMP signaling lead to the loss of the ability of ANP to oppose marinobufagenin-induced inhibition of NKA and fibrosis development. Silencing of the PKG1 gene mimicked these effects of aging.

What makes the sinoatrial node tick? A question not for the faint of heart.

Even before the sinoatrial node (SAN) was discovered, cardiovascular science was engaged in an active investigation of when and why the heart would beat. After the electrochemical theory of bioelectric membrane potentials was formulated and the first action potentials were measured in contracting muscle cells, the field became divided: some investigators studied electrophysiology and ion channels, others studied muscle contraction. It later became known that changes in intracellular Ca2+ cause contraction. The pacemaking field was reunited by the coupled-clock theory of pacemaker cell function, which integrated intracellular Ca2+ cycling and transmembrane voltage into one rhythmogenic system. In this review, we will discuss recent discoveries that contextualize the coupled-clock system, first described in isolated SAN cells, into the complex world of SAN tissue: heterogeneous local Ca2+ releases, generated within SAN pacemaker cells and regulated by the other cell types within the SAN cytoarchitecture, variably co-localize and synchronize to give rise to relatively rhythmic impulses that emanate from the SAN to excite the heart. We will ultimately conceptualize the SAN as a brain-like structure, composed of intercommunicating meshworks of multiple types of pacemaker cells and interstitial cells, intertwined networks of nerves and glial cells and more. This article is part of the theme issue 'The heartbeat: its molecular basis and physiological mechanisms'.

DPP4 inhibition impairs senohemostasis to improve plaque stability in atherosclerotic mice.

Senescent vascular smooth muscle cells (VSMCs) accumulate in the vasculature with age and tissue damage and secrete factors that promote atherosclerotic plaque vulnerability and disease. Here, we report increased levels and activity of dipeptidyl peptidase 4 (DPP4), a serine protease, in senescent VSMCs. Analysis of the conditioned media from senescent VSMCs revealed a unique senescence-associated secretory phenotype (SASP) signature comprising many complement and coagulation factors; silencing or inhibiting DPP4 reduced these factors and increased cell death. Serum samples from persons with high risk for cardiovascular disease contained high levels of DPP4-regulated complement and coagulation factors. Importantly, DPP4 inhibition reduced senescent cell burden and coagulation and improved plaque stability, while single-cell resolution of senescent VSMCs reflected the senomorphic and senolytic effects of DPP4 inhibition in murine atherosclerosis. We propose that DPP4-regulated factors could be exploited therapeutically to reduce senescent cell function, reverse senohemostasis, and improve vascular disease.

cAMP signaling affects age-associated deterioration of pacemaker beating interval dynamics.

Sinoatrial node (SAN) beating interval variability (BIV) and the average beating interval (BI) are regulated by a coupled-clock system, driven by Ca2+-calmodulin activated adenylyl cyclase, cAMP, and downstream PKA signaling. Reduced responsiveness of the BI and BIV to submaximal, [X]50, ß-adrenergic receptor (ß-AR) stimulation, and phosphodiesterase inhibition (PDEI) have been documented in aged SAN tissue, whereas the maximal responses, [X]max, do not differ by age. To determine whether age-associated dysfunction in cAMP signaling leads to altered responsiveness of BI and BIV, we measured cAMP levels and BI in adult (2-4 months n = 27) and aged (22-26 months n = 25) C57/BL6 mouse SAN tissue in control and in response to ß-AR or PDEI at X50 and [X]max. Both cAMP and average BI in adult SAN were reduced at X50, whereas cAMP and BI at Xmax did not differ by age. cAMP levels and average BI were correlated both within and between adult and aged SAN. BIV parameters in long- and short-range terms were correlated with cAMP levels for adult SAN. However, due to reduced cAMP within aged tissues at [X]50, these correlations were diminished in advanced age. Thus, cAMP level generated by the coupled clock mechanisms is tightly linked to average BI. Reduced cAMP level at X50 in aged SAN explains the reduced responsiveness of the BI and BIV to ß-AR stimulation and PDEI.

Age-dependent Lamin changes induce cardiac dysfunction via dysregulation of cardiac transcriptional programs.

As we age, structural changes contribute to progressive decline in organ function, which in the heart act through poorly characterized mechanisms. Taking advantage of the short lifespan and conserved cardiac proteome of the fruit fly, we found that cardiomyocytes exhibit progressive loss of Lamin C (mammalian Lamin A/C homologue) with age, coincident with decreasing nuclear size and increasing nuclear stiffness. Premature genetic reduction of Lamin C phenocopies aging's effects on the nucleus, and subsequently decreases heart contractility and sarcomere organization. Surprisingly, Lamin C reduction downregulates myogenic transcription factors and cytoskeletal regulators, possibly via reduced chromatin accessibility. Subsequently, we find a role for cardiac transcription factors in regulating adult heart contractility and show that maintenance of Lamin C, and cardiac transcription factor expression, prevents age-dependent cardiac decline. Our findings are conserved in aged non-human primates and mice, demonstrating that age-dependent nuclear remodeling is a major mechanism contributing to cardiac dysfunction.

Photobiomodulation therapy mitigates cardiovascular aging and improves survival.

BACKGROUND: Photobiomodulation (PBM) therapy, a form of low-dose light therapy, has been noted to be effective in several age-associated chronic diseases such as hypertension and atherosclerosis. Here, we examined the effects of PBM therapy on age-associated cardiovascular changes in a mouse model of accelerated cardiac aging. METHODS: Fourteen months old Adenylyl cyclase type VIII (AC8) overexpressing transgenic mice (n = 8) and their wild-type (WT) littermates (n = 8) were treated with daily exposure to Near-Infrared Light (850 nm) at 25 mW/cm2 for 2 min each weekday for a total dose of 1 Einstein (4.5 p.J/cm2 or fluence 3 J/cm2 ) and compared to untreated controls over an 8-month period. PBM therapy was administered for 3.5 months (Early Treatment period), paused, due to Covid-19 restrictions for the following 3 months, and restarted again for 1.5 months. Serial echocardiography and gait analyses were performed at monthly intervals, and serum TGF-ß1 levels were assessed following sacrifice. RESULTS: During the Early Treatment period PBM treatments: reduced the age-associated increases in left ventricular (LV) mass in both genotypes (p = 0.0003), reduced the LV end-diastolic volume (EDV) in AC8 (p = 0.04); and reduced the left atrial dimension in both genotypes (p = 0.02). PBM treatments substantially increased the LV ejection fraction (p = 0.03), reduced the aortic wall stiffness (p = 0.001), and improved gait symmetry, an index of neuro-muscular coordination (p = 0.005). The effects of PBM treatments, measured following the pause, persisted. Total TGF-ß1 levels were significantly increased in circulation (serum) in AC8 following PBM treatments (p = 0.01). We observed a striking increase in cumulative survival in PBM-treated AC8 mice (100%; p = 0.01) compared to untreated AC8 mice (43%). CONCLUSION: PBM treatment mitigated age-associated cardiovascular remodeling and reduced cardiac function, improved neuromuscular coordination, and increased longevity in an experimental animal model. These responses correlate with increased TGF-ß1 in circulation. Future mechanistic and dose optimization studies are necessary to assess these anti-aging effects of PBM, and validation in future controlled human studies is required for effective clinical translation.

Characterization of diverse populations of sinoatrial node cells and their proliferation potential at single nucleus resolution.

Background: Each heartbeat is initiated in the sinoatrial node (SAN), and although a recent study (GSE130710) using single nucleus RNA-seq had discovered different populations of cell types within SAN tissue, the distinct potential functions of these cell types have not been delineated. Methods: To infer some special potential functions of different SAN cell clusters, we applied principal component analysis (PCA), t-distributed stochastic neighbor embedding (t-SNE) and uniform manifold approximation and projection (UMAP) to the GSE130710 dataset to reduce dimensions, followed by Pseudotime trajectory and AUCell analyses, ANOVA and Hurdle statistical models, and Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) enrichments to determine functional potential of cell types. Nuclear EdU immuno-labeling of SAN tissue confirmed cell type proliferation. Findings: We identified elements of a coupled clock system known to drive SAN cell pacemaking within the GSE130710 sinus node myocyte cluster, which, surprisingly, manifested signals of suppressed fatty acid and nitrogen metabolism and reduced immune gene expression. Proliferation signaling was enriched in endocardial, epicardial, epithelial cells, and macrophages, in which, fatty acid and nitrogen metabolic signals were also suppressed, but immune signaling was enhanced. EdU labeling was rare in pacemaker cells but was robust in interstitial cells. Interpretation: Pacemaker cells that initiate each heartbeat manifest suppressed fatty acid and nitrogen metabolism and limited immune signaling and proliferation potential. In contrast, other populations of SAN cells not directly involved in the initiation of heartbeats, manifest robust proliferation and immune potential, likely to ensure an environment required to sustain healthy SAN tissue pacemaker function.

Changes in cAMP signaling are associated with age-related downregulation of spontaneously beating atrial tissue energetic indices.

The prevalence of atria-related diseases increases exponentially with age and is associated with ATP supply-to-demand imbalances. Because evidence suggests that cAMP regulates ATP supply-to-demand, we explored aged-associated alterations in atrial ATP supply-to-demand balance and its correlation with cAMP levels. Right atrial tissues driven by spontaneous sinoatrial node impulses were isolated from aged (22-26 months) and adult (3-4 months) C57/BL6 mice. ATP demand increased by addition of isoproterenol or 3-Isobutyl-1-methylxanthine (IBMX) and decreased by application of carbachol. Each drug was administrated at the dose that led to a maximal change in beating rate (Xmax) and to 50% of that maximal change in adult tissue (X50). cAMP, NADH, NAD + NADH, and ATP levels were measured in the same tissue. The tight correlation between cAMP levels and the beating rate (i.e., the ATP demand) demonstrated in adult atria was altered in aged atria. cAMP levels were lower in aged compared to adult atrial tissue exposed to X50 of ISO or IBMX, but this difference narrowed at Xmax. Neither ATP nor NADH levels correlated with ATP demand in either adult or aged atria. Baseline NADH levels were lower in aged as compared to adult atria, but were restored by drug perturbations that increased cAMP levels. Reduction in Ca2+-activated adenylyl cyclase-induced decreased cAMP and prolongation of the spontaneous beat interval of adult atrial tissue to their baseline levels in aged tissue, brought energetics indices to baseline levels in aged tissue. Thus, cAMP regulates right atrial ATP supply-to-demand matching and can restore age-associated ATP supply-to-demand imbalance.

Dilated hypertrophic phenotype of the carotid artery is associated with accelerated age-associated central arterial stiffening.

Hypertrophic carotid geometric phenotypes (h-CGP) are predictors of incident cardiovascular disease (CVD). While arterial aging is hypothesized as a contributor to this associated risk, the association of CGPs with chronological age is not clear. In this manuscript we examine whether hypertrophic CGPs represent accelerated biological, rather than chronological, aging by examining their association with carotid-femoral pulse wave velocity (PWV), the hallmark of arterial aging. We analyzed data from 5516 participants of the SardiNIA study with a wide range of age at baseline (20-101 years), and a median follow-up time of 13 years (mean 11.5 years; maximum 17.9 years). Baseline CGPs were defined based on the common carotid lumen diameter, wall thickness, and their ratio. Subject-specific rates of change of PWV, blood pressure parameters, body mass index, glucose, and lipids were estimated using linear mixed effects models. Compared to those with typical(t-) CGP, those with dilated hypertrophy (dh-) CGP had a greater longitudinal increase in PWV; this increase was significantly greater among older individuals and men. The greater PWV longitudinal increase in dh-CGP remained significant after adjusting for baseline values and rates of change of covariates. Dilated hypertrophic CGP is independently associated with accelerated increase in age-associated arterial stiffening over time, with a strong association in men than in women. Future studies are needed to examine if this association mediates the increased risk for CVD observed in individuals with hypertrophic cardiac remodelling and the role of retarding it to reduce this risk. HIGHLIGHTS: • Individuals with dilated hypertrophic geometric phenotypes of the common carotid artery (increased age- and sex-specific wall thickness and lumen diameter) have greater future central arterial stiffening, independently of other determinants of arterial stiffening. • The dilated hypertrophic phenotype group has a greater age-specific arterial dilation, wall thickening, and stiffness (the arterial aging triad). This suggests that this phenotype is a form of accelerated aging that might explain the worse clinic outcomes observed in this group. • Understanding the natural history of the carotid geometric phenotype across the lifespan and the determinants of the deleterious progression towards the dilated hypertrophic phenotype are needed to develop interventions that reduce the adverse clinical outcomes associated with it.