Introduction: Angiopoietin like proteins (ANGPTL) 3, 4, and 8 and their complexes (ANGPTL 3/8 and 4/8) are established inhibitors of lipoprotein lipase (LPL), all of which regulate triglyceride (TG) metabolism. Regular exercise is known to decrease plasma TG levels and increase LPL activity, and we recently showed that exercise training decreases ANGPTL3/8 levels. However, the molecular underpinnings of exercise-induced changes in regulators of TG metabolism have not been fully elucidated. The purpose of this study was to identify proteins associated with TG, LPL activity, C-terminal domain-containing (CD)-ANGPTL4, ANGPTL3/8, and ANGPTL4/8 responses to exercise training. Methods: Data on cardiometabolic phenotypes and plasma proteins (Olink 5K) measured before and after 20 weeks of endurance training were available in 617 adults (55% Female, 37% Black, 17-65 yrs) of the HERITAGE Family Study. Linear mixed models were used to test the associations of change in individual proteins with changes in TG-related traits, adjusting for age, sex, race, and baseline trait and baseline protein levels. Significant (FDR<0.05) proteins were entered in LASSO regression models with 10-fold cross-validation to identify proteomic signatures of exercise-induced changes in TG-related traits. Results: At baseline, mean age was 35 yrs, BMI 26 kg/m 2 , and lipid profile values were within the normal range. Changes in the abundance of 141 unique proteins were associated with the exercise training changes of at least one of the four traits (none associated with ΔLPL; Table 1 ), with little overlap of associated proteins across traits ( Figure 1 ). The associated proteins were overrepresented in pathways related to peptide hormone metabolism, lipoprotein assembly, remodeling and clearance, assembly of LPL and LIPC complexes, G protein-coupled receptor signaling, and clotting cascade ( Table 1 ). Proteomic signatures consisting of 24, 28, 22, and 13 proteins explained 20%, 22%, 21%, and 28% of the variance in training-induced changes in TG, ANGPTL3/8, ANGPTL4/8, and CD-ANGPTL4, respectively. Top contributing proteins for each signature are shown in Figure 2 . Conclusions: We identified several plasma proteins whose changes in response to regular exercise were associated with concomitant changes in TG and ANGPTL complex and protein traits. Our findings indicate several shared but also unique biological pathways underlying the exercise responsiveness of key regulators of TG metabolism.

Background: Atherosclerotic cardiovascular disease (ASCVD) risk scores are commonly used to inform therapeutic strategies in the clinical setting. Although previous studies have shown that individual risk score components, such as HDL-C and total cholesterol, can be improved with regular exercise, few studies have investigated the effect of exercise interventions on composite measures of predicted ASCVD risk. Methods: Data from eight exercise trials (INFLAME; HART-D; CardioRACE; Queen’s; STRRIDEs 1, 2, and PD; HERITAGE) with 21 different interventions varying in exercise mode, amount, and/or intensity were analyzed. ASCVD risk scores were calculated for each participant (n=2,074) at baseline and after exercise training using the pooled cohort equations and Framingham Heart Study algorithm for 10- and 30-year risk, respectively ( Table 1 ). Meta-analysis of study group-specific mean changes in ASCVD risk with training was performed. When available, changes in exercise groups were compared against changes in the respective control group for each study. Since age is the strongest predictor of risk in the equations, additional meta-analysis was performed to examine changes in risk due to age only (phenotype levels held constant between time points), with all groups analyzed individually with no comparison to controls. Results: Exercise training resulted in an absolute decrease in 10-year ASCVD risk of -0.2% (95% CI: -0.3 to -0.1) and -1.2% (95% CI: -1.8 to -0.5) in 30-year risk, with low and high heterogeneity, respectively ( Figure 1 ). An increase in age only would have significantly increased 10- and 30-year risk across all exercise intervention groups by 0.3% (95% CI: 0.2 to 0.3), and 0.6% (95% CI: 0.5 to 0.7), respectively ( Figure 1 ). However, this age-related increase was abolished in all exercise groups except for the resistance training groups in STRRIDE 2 and CardioRACE. Conclusions: We found that a relatively short period of endurance exercise training of differing amounts and/or intensity performed alone or in combination with resistance training can prevent and even reverse the age-related increases in both short- and long-term ASCVD risk.

Introduction: Plasma proteomics has been used to identify organ-specific aging signatures related to health and disease. It is unknown whether regular exercise can modulate predicted organ aging. Research Question: To examine whether proteomic aging signatures are modified by endurance exercise training (ET). Methods: We measured 4,979 plasma proteins (SomaScan assay) in the HERITAGE Family Study (N=657 adults) before and after 20 weeks of ET. Organ age was estimated for 11 major organs using different panels of proteins and age gap was calculated as the difference between predicted age and the LOWESS regression estimate of the population mean. Accelerated and decelerated aging were defined as an age gap value ≥ or < 2 SD from the mean, respectively. Paired t-tests examined changes in predicted organ age after ET. Results: The mean (SD) age of the study was 34.4 (13.5) years, and 55.1% were female. We found low-to-moderate correlations between predicted organ age and chronological age for 9 of 11 traits with the strongest correlations found in the conventional age score (r=0.81, p=2.1E-155), which included all proteins on the platform, and organismal age score, which incorporated organ-nonspecific proteins (r = 0.77, p=3.5E-132). Organismal age score was highly associated with several cardiometabolic traits including VO 2 max (r=-0.44, p=1.3E-37), body fat percentage (r=0.35, p=1.6E-21), and waist circumference (r=0.29, p=3.2E-13) after adjusting for sex and race; these associations were significant after adjustment for chronological age. We found 8 of the 11 organ age signatures changed (p<0.05) with ET ( Figure 1 ), with kidney, muscle, and pancreas the only organs to decrease age, while adipose, brain, heart, immune, and liver ages increased. Among the 23% of participants with accelerated and 15% with decelerated organ aging at baseline, 74% had fewer accelerated or decelerated organ ages after ET ( Figure 2 ). Conclusions: We found that organ age signatures replicated in the HERITAGE Family Study and were modified by ET in both positive and negative directions. These changes suggest that individuals with extreme aging tended to regress toward the mean, whereby those with accelerated aging decreased and those with decelerated aging increased organ aging.

While physical activity reduces the risk for chronic disease development, evidence suggests those experiencing early life growth-restriction do not express positive adaptations in response to physical activity. The purpose of this study was to examine the effects of low birthweight (LBW) on markers of chronic disease, adult physical activity, and the response to physical activity engagement in a longitudinal human cohort study. Data from the Framingham Offspring Cohort were organized to include participants with birthweight, physical activity, and chronic disease biomarker/treatment data available at two timepoints (exam 5 and exam 9, 19-year difference). A two-way ANCOVA was performed to determine the association of LBW and sex on physical activity engagement (63.0% female, 10.4% LBW). A multinomial logistic regression was performed to examine the associations of low birthweight and sex on chronic disease development while adjusting for physical activity. LBW was associated with elevated blood glucose and triglycerides (Exam 9). Though not statistically significant (p = 0.08), LBW females potentially spent more time in sedentary activity at exam 5 than LBW males and normal birthweight (NBW) females. LBW males spent significantly more time (p = 0.03) sedentary at exam 9 compared to NBW males and LBW females. There were no differences in the likelihood of chronic disease treatment between groups. Chronic disease biomarkers remained elevated when adjusted for total physical activity. In conclusion, LBW participants in the Framingham Offspring Cohort were not more likely to be treated for chronic diseases when controlling for physical activity engagement, though biomarkers of chronic disease remained elevated.

Background: The lipoprotein insulin resistance (LP-IR) score has been shown to assess insulin resistance, predict future type 2 diabetes, and improve with regular exercise. The lipidomic profile is known to differ with insulin resistance and type 2 diabetes, but few studies have examined lipidome associations with LP-IR, particularly in response to an exercise intervention. Methods: Plasma lipids were measured using the C8-positive LC-MS method in 671 participants from the HERITAGE Family Study (56% Female, 35% Black, 35.2 yrs) before and after a 20-week exercise intervention. LP-IR, a weighted index of six lipoprotein parameters, was measured before and after training through nuclear magnetic resonance spectroscopy (Labcorp, NC). Linear mixed models were used to test the associations of 193 known plasma lipids with LP-IR before and after exercise training. All models were adjusted for age, sex, and race, while post-training models also adjusted for baseline lipid and baseline LP-IR. Results: A total of 162 lipids (84%) were associated (FDR<0.05) with LP-IR at baseline ( Figure 1 ). The top positive associations were found for TG species, while cholesterol ester species showed the top inverse associations. Following training, the change in 143 (74%) lipids were associated (FDR<0.05) with change in LP-IR. There were 129 lipid species associated with both baseline and changes in LP-IR, while 33 were only associated with baseline measures and 14 only associated with changes in LP-IR following exercise ( Figure 1 ). Conclusions: We found that most lipids were associated with LP-IR regardless of exercise training status. Importantly, we identified a subset of lipids that were only associated with changes in LP-IR, which may represent exercise responsive biomarkers of exercise induced changes in LP-IR. However, further research is needed to identify the biological mechanisms connecting the changes in these lipid species and metabolic changes following exercise training.

Postnatal growth restriction (PGR) can increase the risk of cardiovascular disease (CVD) potentially due to impairments in oxidative phosphorylation (OxPhos) within cardiomyocyte mitochondria. The purpose of this investigation was to determine if PGR impairs cardiac metabolism, specifically OxPhos. FVB (Friend Virus B-type) mice were fed a normal-protein (NP: 20% protein), or low-protein (LP: 8% protein) isocaloric diet 2 weeks before mating. LP dams produce ∼20% less milk, and pups nursed by LP dams experience reduced growth into adulthood as compared to pups nursed by NP dams. At birth (PN1), pups born to dams fed the NP diet were transferred to LP dams (PGR group) or a different NP dam (control group: CON). At weaning (PN21), all mice were fed the NP diet. At PN22 and PN80, mitochondria were isolated for respirometry (oxygen consumption rate, J O 2 ${J_{{{\mathrm{O}}_{\mathrm{2}}}}}$ ) and fluorimetry (reactive oxygen species emission, J H 2 O 2 ${J_{{{\mathrm{H}}_{\mathrm{2}}}{{\mathrm{O}}_{\mathrm{2}}}}}$ ) analysis measured as baseline respiration (LEAK) and with saturating ADP (OxPhos). Western blotting at PN22 and PN80 determined protein abundance of uncoupling protein 3, peroxiredoxin-6, voltage-dependent anion channel and adenine nucleotide translocator 1 to provide further insight into mitochondrial function. ANOVAs with the main effects of diet, sex and age with α-level of 0.05 was set a priori. Overall, PGR (7.8 ± 1.1) had significant (P = 0.01) reductions in respiratory control in complex I when compared to CON (8.9 ± 1.0). In general, our results show that PGR led to higher electron leakage in the form of free radical production and reactive oxygen species emission. No significant diet effects were found in protein abundance. The observed reduced respiratory control and increased ROS emission in PGR mice may increase risk for CVD in mice.

Background and aims: Previous studies have derived and validated an HDL apolipoproteomic score (pCAD) that predicts coronary artery disease (CAD) risk. However, the associations between pCAD and markers of cardiometabolic health in healthy adults are not known, nor are the effects of regular exercise on pCAD. Methods: A total of 641 physically inactive adults free of cardiovascular disease from the HERITAGE Family Study completed 20 weeks of exercise training. The pCAD index (range 0-100) was calculated using measurements of apolipoproteins A-I, C-I, C-II, C-III, and C-IV from ApoA-I-tagged serum (higher index = higher CAD risk). The associations between pCAD index and cardiometabolic traits at baseline and their training responses were assessed with Spearman correlation and general linear models. A Bonferroni correction of p < 8.9 x 10(-04) was used to determine statistical significance. Results: The mean +/- SD baseline pCAD index was 29 +/- 32, with 106 (16.5 %) participants classified as high CAD risk. At baseline, pCAD index was positively associated with blood pressure, systemic inflammation, and body composition. HDL size, VO2max, and HDL-C were negatively associated with pCAD index at baseline. Of those classified as high CAD risk at baseline, 52 (49 %) were reclassified as normal risk after training. Following training, pCAD index changes were inversely correlated (p < 1.4 x 10(-04)) with changes in HDL-C, HDL size, and LDL size. Conclusions: A higher pCAD index was associated with a worse cardiometabolic profile at baseline but improved with regular exercise. The results from this study highlight the potential role of HDL apolipoproteins as therapeutic targets for lifestyle interventions, particularly in high-risk individuals.

Background: Prospective cohort studies have shown plasma cholesterol ester (CE) 18:2 is inversely associated with all-cause and cardiovascular mortality. However, less is known about the association of CE18:2 with cardiometabolic risk factors and whether it is responsive to regular exercise. Methods: A total of 193 known plasma lipids, including 11 CEs, were measured using the C8-positive LC-MS method in 671 participants from the HERITAGE Family Study (56% Female, 35% Black, 35 yrs) before and after a 20-week exercise intervention. Linear mixed models were used to test the associations of all lipids with 6 cardiometabolic traits at baseline and post-training adjusting for age, sex, and race, with post-training models also adjusted for baseline lipid and trait values. Paired t-tests were used to test the difference in CE18:2 abundance before and after exercise training. Results: Out of the 193 lipids, CE18:2 was consistently among the top 2 associations with each cardiometabolic trait at baseline ( Table 1 ). Levels of CE18:2 were inversely associated with triglycerides, visceral fat, glycA, and small LDL, while positively associated with HDL-C and insulin sensitivity. Importantly, CE18:2 abundance was increased following the exercise intervention (p=0.0003). Exercise-induced change in CE18:2 was significantly (FDR<0.05) associated with concomitant changes in all traits except visceral fat in the same directions as baseline, with CE18:2 among the top 3 associations with each training response trait ( Table 1 ). Conclusions: Plasma CE18:2 levels were associated with cardiometabolic traits in a favorable direction both before and after an exercise intervention. Thus, this plasma lipid may be an exercise inducible metabolite that is indicative of improvements in cardiometabolic health. However, additional studies are needed to confirm the observed findings and determine the mechanisms underlying these beneficial associations.

INTRODUCTION:Early life growth restriction significantly increases the risk of adulthood physical inactivity and thereby chronic disease incidence. Improvements in motor skill acquisition could result in greater physical activity engagement in the growth-restricted population, thus reducing chronic disease risk. The purpose of this study was to implement an early life motor training intervention to improve physical activity engagement in control and growth-restricted mice. METHODS:Mice were growth restricted in early life utilizing a validated nutritive model or remained fully nourished in early life as a control. All mice were tested throughout early life for various components of motor skill acquisition. On postnatal day 10, mice were randomly assigned to engage in an early life motor skill intervention daily until postnatal day 21 or remained as a sedentary control. All mice were given access to an in-cage running wheel from postnatal days 45-70. RESULTS:Growth-restricted group (PGR) mice had impaired trunk and postural control, coordination/vestibular development, and hindlimb strength in early life compared with control mice. There were no differences in wheel running behavior between the trained and sedentary mice, although control mice ran at a faster average speed compared with PGR mice. Control female mice ran more than PGR female mice during the week 2 dark cycle. CONCLUSIONS:Early life growth restriction reduced motor skill attainment throughout early life, which may be associated with reduced ability to engage in physical activity in adulthood. The early life motor skill intervention did not elicit changes in body weight or physical activity engagement in control or PGR mice, indicating that a more intense/different intervention specifically targeting skeletal muscle may be necessary to counteract the detrimental effects of early life growth restriction.

Background: Excessive visceral fat is associated with metabolic alterations and is a causal risk factor for CVD. The plasma lipidome is altered in obesity and lipidome signatures of BMI and obesity have been identified. However, few studies have examined the plasma lipidome in relation to visceral fat. Methods: Plasma lipids were measured using the C8-positive LC-MS method in 671 participants from the HERITAGE Family Study (56% Female, 35% Black, 35 yrs). Visceral fat was measured using CT scans. Linear mixed models were used to test the associations of 193 known plasma lipids with visceral fat adjusting for age, sex, race, and BMI. A FDR<5% was used to determine significance. Results: In individual models, 156 lipid species were significantly associated with visceral fat, with 109 species associated after additional adjustment for BMI ( Fig 1 ). The top positively associated lipids were triglycerides, while cholesterol esters showed the strongest inverse associations with visceral fat. A LASSO regression model retained 73 lipids and explained 78.5% of the variance in visceral fat. Delta visceral fat was calculated as the difference between predicted (from LASSO) and measured visceral fat. Examining quartiles of delta visceral fat showed that discordance between predicted and actual visceral fat was associated with differing cardiometabolic profiles independent of age, sex, race, visceral fat, and BMI. Individuals with higher delta visceral fat (Q4, overpredicted) had significantly (p<1.0x10 -04 ) higher levels of TG, apoB, total cholesterol, LDL-C, and fasting insulin and lower levels of large HDL particles and LPL activity compared to those with lower delta visceral fat (Q1, underpredicted). Conclusions: The plasma lipidome is widely associated with visceral fat levels. A lipidome-based visceral fat score may provide additional information over measured visceral fat for assessment of cardiometabolic health. Further studies are needed to test and validate the clinical utility of such lipidome-based scores.

ABSTRACT Introduction Growth restriction (GR) reduces ribosome abundance and skeletal muscle mass in mice. A reduction in skeletal muscle mass increases the risk of frailty and is associated with high morbidity and mortality rates. As eccentric type exercise increases muscle mass, this investigation aimed to determine if eccentric loading of skeletal muscle via downhill running (DHR) increased muscle mass in GR mice. Methods Mice were growth-restricted either gestational undernutrition (GUN, n = 8 litters), postnatal undernutrition (PUN, n = 8 litters), or were not restricted (CON, n = 8 litters) via a validated cross-fostering nutritive model. On postnatal day (PN) 21, all mice were weaned to a healthy diet, isolating the period of GR to early life as seen in humans. At PN45, mice were assigned to either a DHR (CON, n = 4 litters; GUN, n = 4 litters; PUN, n = 4 litters) or sedentary (SED: CON, n = 4 litters; GUN, n = 4 litters; PUN, n = 4 litters) group. Downhill running (16% decline: 18 m·min−1) was performed in 30-min bouts, three times per week, for 12 wk on a rodent treadmill. At PN129, the quadriceps femoris was dissected and evaluated for mass, myofiber size and type, and molecular markers of growth. Results Following training, CON-DHR mice having larger cells than CON-SED, GUN-SED, PUN-SED, and PUN-DHR mice (P < 0.05). The PUN group (as compared with CON) had reduced body mass (P < 0.001), upstream binding factor abundance (P = 0.012), phosphor-mTOR (P < 0.001), and quadriceps mass (P = 0.02). The GUN and PUN groups had increased MuRF1 abundance (P < 0.001) compared with CON (P < 0.001). Conclusions The blunted response to training suggests GR mice may have anabolic resistance when exposed to eccentric type exercise.

Physical activity engagement results in a variety of positive health outcomes, including a reduction in cardiovascular disease risk partially due to eccentric remodeling of the heart. The purpose of this investigation was to determine if four replicate lines of High Runner mice that have been selectively bred for voluntary exercise on wheels have a cardiac phenotype that resembles the outcome of eccentric remodeling. Adult females (average age 55 days) from the 4 High Runner and 4 non-selected control lines were anaesthetized via vaporized isoflurane, then echocardiographic images were collected and analyzed for structural and functional differences. High Runner mice in general had lower ejection fractions compared to control mice lines (2-tailed p ​= ​0.023 6) and tended to have thicker walls of the anterior portion of the left ventricle (p ​= ​0.065). However, a subset of the High Runner individuals, termed mini-muscle mice, had greater ejection fraction (p ​= ​0.000 6), fractional shortening percentage (p ​< ​0.000 1), and ventricular mass at dissection (p ​< ​0.002 7 with body mass as a covariate) compared to non-mini muscle mice. Mice from replicate lines bred for high voluntary exercise did not all have inherent positive cardiac functional or structural characteristics, although a genetically unique subset of mini-muscle individuals did have greater functional cardiac characteristics, which in conjunction with their previously described peripheral aerobic enhancements (e.g., increased capillarity) would partially account for their increased V˙ O2max.

Purpose: Growth restriction during postnatal development is linked to increased cardiovascular disease risk in adulthood, with greater impairments seen in females. Postnatal growth restricted (PNGR) female mice display diastolic dysfunction and impaired calcium (Ca2+) flux. Regular aerobic exercise promotes cardiovascular health through increases in Ca2+ handling proteins, specifically SERCA2. Thus, the purpose of this investigation was to determine if moderate-intensity aerobic exercise improves Ca2+ handling protein abundance in PNGR mice. Methods: To induce postnatal growth-restriction, FVB mouse dams were fed either a 20% protein control diet or 8% low-protein (LP) diet 2 weeks before mating. Only pups born to control-fed dams were cross-fostered on postnatal day (PN) 1 to either a LP fed dam (PNGR) or a different control fed dam (CON). LP fed dams produce less milk leading to permanent growth restriction in pups. On PN 21, all pups were weaned onto the CON diet. At PN 45, mice were assigned to down-hill running (PNGRDHR; n = 4, CONDHR; n=3) or sedentary (PNGRSED; n = 4, CONSED; n = 4). DHR elicits moderate intensity aerobic exercise as it does not allow mice to stop and re-start running during sessions. The exercise protocol consisted of treadmill running 3 days/week for 12 weeks at 18 m/min and -16% decline for 28 mins/session. On PN 129, hearts were dissected and analyzed via western blot for Ca2+ proteins: SERCA1/2, Phospholamban (PLN), Phosphorylated-PLN (P-PLN), Protein Kinase A (PKA) and Phosphorylated-PKA (P-PKA). The effect of exercise on Ca2+ protein abundances was calculated as percent change from respective SED groups and then analyzed with a t-test (alpha p ≤ 0.05). Results: PNGR-EX had lower abundance of SERCA2 (-45 ± 13 vs. 70 ± 31, p = 0.01), PLN (-13 ± 7 vs. 45 ± 7, p = 0.0022), P-PLN (-20 ± 14 vs. 48 ± 16, p = 0.01), and P-PKA (-3 ± 3 vs. 11 ± 4, p = 0.02) compared to CON-EX. Neither PNGR nor CON mice showed a change in SERCA1(-21 ± 13 vs. -19 ± 9, p = 0.45) or PKA post exercise (32 + 13 vs. -.66 ± 4, p = 0.90). Conclusion: Aerobic exercise decreased the abundance of SERCA2, PLN, P-PLN and P-PKA in PNGR mice. The reduced SERCA2 abundance is associated with impaired contraction dynamics (diastolic dysfunction myocardial stiffness, or fibrosis) which can increase mortality from cardiovascular disease. Supported by MSU startup funds