Macrophage de novo NAD+ synthesis specifies immune function in aging and inflammation.

Recent advances highlight a pivotal role for cellular metabolism in programming immune responses. Here, we demonstrate that cell-autonomous generation of nicotinamide adenine dinucleotide (NAD+) via the kynurenine pathway (KP) regulates macrophage immune function in aging and inflammation. Isotope tracer studies revealed that macrophage NAD+ derives substantially from KP metabolism of tryptophan. Genetic or pharmacological blockade of de novo NAD+ synthesis depleted NAD+, suppressed mitochondrial NAD+-dependent signaling and respiration, and impaired phagocytosis and resolution of inflammation. Innate immune challenge triggered upstream KP activation but paradoxically suppressed cell-autonomous NAD+ synthesis by limiting the conversion of downstream quinolinate to NAD+, a profile recapitulated in aging macrophages. Increasing de novo NAD+ generation in immune-challenged or aged macrophages restored oxidative phosphorylation and homeostatic immune responses. Thus, KP-derived NAD+ operates as a metabolic switch to specify macrophage effector responses. Breakdown of de novo NAD+ synthesis may underlie declining NAD+ levels and rising innate immune dysfunction in aging and age-associated diseases.

Optimizing Abdominoplasty Surgical Site Morbidity Profiling Through an Effective and Nationally Validated Risk Scoring System.

BACKGROUND: Abdominoplasty complication rates are among the highest for cosmetic surgery. We sought to create a validated scoring system to predict the likelihood of wound complications after abdominoplasty using a national multi-institutional database. METHODS: Patients who underwent abdominoplasty in the American College of Surgeons National Surgical Quality Improvement Program 2007-2019 database were analyzed for surgical site complications, a composite outcome of wound disruption, and surgical site infections. The cohort was randomly divided into a 60% testing and a 40% validation sample. Multivariable logistic regression analysis was performed to identify independent predictors of complications using the testing sample (n = 11,294). The predictors were weighted according to ß coefficients to develop an integer-based clinical risk score. This system was validated using receiver operating characteristic analysis of the validation sample (n = 7528). RESULTS: A total of 18,822 abdominoplasty procedures were identified. The proportion of patients who developed a composite surgical site complication was 6.8%. Independent risk factors for composite surgical site complication included inpatient procedure (P < 0.01), smoking (P < 0.01), American Society of Anesthesiologists class ≥3 (P < 0.01), and body mass index ≥25.0 and ≤18.0 kg/m2 (P < 0.01). African American race was a protective factor against surgical site complications (P < 0.01). The factors were integrated into a scoring system, ranging from -5 to 42, and the receiver operating characteristic analysis revealed an area under the curve of 0.71. CONCLUSIONS: We present a validated scoring system for postoperative 30-day surgical site morbidity after abdominoplasty. This system will enable surgeons to optimize patient selection to decrease morbidity and unnecessary healthcare expenditure.

4HNE Impairs Myocardial Bioenergetics in Congenital Heart Disease-Induced Right Ventricular Failure.

BACKGROUND: In patients with complex congenital heart disease, such as those with tetralogy of Fallot, the right ventricle (RV) is subject to pressure overload stress, leading to RV hypertrophy and eventually RV failure. The role of lipid peroxidation, a potent form of oxidative stress, in mediating RV hypertrophy and failure in congenital heart disease is unknown. METHODS: Lipid peroxidation and mitochondrial function and structure were assessed in right ventricle (RV) myocardium collected from patients with RV hypertrophy with normal RV systolic function (RV fractional area change, 47.3±3.8%) and in patients with RV failure showing decreased RV systolic function (RV fractional area change, 26.6±3.1%). The mechanism of the effect of lipid peroxidation, mediated by 4-hydroxynonenal ([4HNE] a byproduct of lipid peroxidation) on mitochondrial function and structure was assessed in HL1 murine cardiomyocytes and human induced pluripotent stem cell-derived cardiomyocytes. RESULTS: RV failure was characterized by an increase in 4HNE adduction of metabolic and mitochondrial proteins (16 of 27 identified proteins), in particular electron transport chain proteins. Sarcomeric (myosin) and cytoskeletal proteins (desmin, tubulin) also underwent 4HNE adduction. RV failure showed lower oxidative phosphorylation (moderate RV hypertrophy, 287.6±19.75 versus RV failure, 137.8±11.57 pmol/[sec×mL]; P=0.0004), and mitochondrial structural damage. Using a cell model, we show that 4HNE decreases cell number and oxidative phosphorylation (control, 388.1±23.54 versus 4HNE, 143.7±11.64 pmol/[sec×mL]; P<0.0001). Carvedilol, a known antioxidant did not decrease 4HNE adduction of metabolic and mitochondrial proteins and did not improve oxidative phosphorylation. CONCLUSIONS: Metabolic, mitochondrial, sarcomeric, and cytoskeletal proteins are susceptible to 4HNE-adduction in patients with RV failure. 4HNE decreases mitochondrial oxygen consumption by inhibiting electron transport chain complexes. Carvedilol did not improve the 4HNE-mediated decrease in oxygen consumption. Strategies to decrease lipid peroxidation could improve mitochondrial energy generation and cardiomyocyte survival and improve RV failure in patients with congenital heart disease.

Physiological Mitochondrial Fragmentation Is a Normal Cardiac Adaptation to Increased Energy Demand.

RATIONALE: Mitochondria play a dual role in the heart, responsible for meeting energetic demands and regulating cell death. Paradigms have held that mitochondrial fission and fragmentation are the result of pathological stresses, such as ischemia, are an indicator of poor mitochondrial health, and lead to mitophagy and cell death. However, recent studies demonstrate that inhibiting fission also results in decreased mitochondrial function and cardiac impairment, suggesting that fission is important for maintaining cardiac and mitochondrial bioenergetic homeostasis. OBJECTIVE: The purpose of this study is to determine whether mitochondrial fission and fragmentation can be an adaptive mechanism used by the heart to augment mitochondrial and cardiac function during a normal physiological stress, such as exercise. METHODS AND RESULTS: We demonstrate a novel role for cardiac mitochondrial fission as a normal adaptation to increased energetic demand. During submaximal exercise, physiological mitochondrial fragmentation results in enhanced, rather than impaired, mitochondrial function and is mediated, in part, by ß1-adrenergic receptor signaling. Similar to pathological fragmentation, physiological fragmentation is induced by activation of dynamin-related protein 1; however, unlike pathological fragmentation, membrane potential is maintained and regulators of mitophagy are downregulated. Inhibition of fission with P110, Mdivi-1 (mitochondrial division inhibitor), or in mice with cardiac-specific dynamin-related protein 1 ablation significantly decreases exercise capacity. CONCLUSIONS: These findings demonstrate the requirement for physiological mitochondrial fragmentation to meet the energetic demands of exercise, as well as providing additional support for the evolving conceptual framework, where mitochondrial fission and fragmentation play a role in the balance between mitochondrial maintenance of normal physiology and response to disease.

Time-dependent evolution of functional vs. remodeling signaling in induced pluripotent stem cell-derived cardiomyocytes and induced maturation with biomechanical stimulation.

Human induced pluripotent stem cell-derived cardiomyocytes (hiPSC-CMs) are a powerful platform for uncovering disease mechanisms and assessing drugs for efficacy/toxicity. However, the accuracy with which hiPSC-CMs recapitulate the contractile and remodeling signaling of adult cardiomyocytes is not fully known. We used ß-adrenergic receptor (ß-AR) signaling as a prototype to determine the evolution of signaling component expression and function during hiPSC-CM maturation. In "early" hiPSC-CMs (less than or equal to d 30), ß2-ARs are a primary source of cAMP/PKA signaling. With longer culture, ß1-AR signaling increases: from 0% of cAMP generation at d 30 to 56.8 ± 6.6% by d 60. PKA signaling shows a similar increase: 15.7 ± 5.2% (d 30), 49.8 ± 0.5% (d 60), and 71.0 ± 6.1% (d 90). cAMP generation increases 9-fold from d 30 to 60, with enhanced coupling to remodeling pathways (e.g., Akt and Ca(2+)/calmodulin-dependent protein kinase type II) and development of caveolin-mediated signaling compartmentalization. By contrast, cardiotoxicity induced by chronic ß-AR stimulation, a major component of heart failure, develops much later: 5% cell death at d 30vs 55% at d 90. Moreover, ß-AR maturation can be accelerated by biomechanical stimulation. The differential maturation of ß-AR functionalvs remodeling signaling in hiPSC-CMs has important implications for their use in disease modeling and drug testing. We propose that assessment of signaling be added to the indices of phenotypic maturation of hiPSC-CMs.-Jung, G., Fajardo, G., Ribeiro, A. J. S., Kooiker, K. B., Coronado, M., Zhao, M., Hu, D.-Q., Reddy, S., Kodo, K., Sriram, K., Insel, P. A., Wu, J. C., Pruitt, B. L., Bernstein, D. Time-dependent evolution of functionalvs remodeling signaling in induced pluripotent stem cell-derived cardiomyocytes and induced maturation with biomechanical stimulation.

TLR3 deficiency induces chronic inflammatory cardiomyopathy in resistant mice following coxsackievirus B3 infection: role for IL-4.

Recent findings indicate that TLR3 polymorphisms increase susceptibility to enteroviral myocarditis and inflammatory dilated cardiomyopathy (iDCM) in patients. TLR3 signaling has been found to inhibit coxsackievirus B3 (CVB3) replication and acute myocarditis in mouse models, but its role in the progression from myocarditis to iDCM has not been previously investigated. In this study we found that TLR3 deficiency increased acute (P = 5.9 × 10(-9)) and chronic (P = 6.0 × 10(-7)) myocarditis compared with WT B6.129, a mouse strain that is resistant to chronic myocarditis and iDCM. Using left ventricular in vivo hemodynamic assessment, we found that TLR3-deficient mice developed progressively worse chronic cardiomyopathy. TLR3 deficiency significantly increased viral replication in the heart during acute myocarditis from day 3 through day 12 after infection, but infectious virus was not detected in the heart during chronic disease. TLR3 deficiency increased cytokines associated with a T helper (Th)2 response, including IL-4 (P = 0.03), IL-10 (P = 0.008), IL-13 (P = 0.002), and TGF-ß(1) (P = 0.005), and induced a shift to an immunoregulatory phenotype in the heart. However, IL-4-deficient mice had improved heart function during acute CVB3 myocarditis by echocardiography and in vivo hemodynamic assessment compared with wild-type mice, indicating that IL-4 impairs cardiac function during myocarditis. IL-4 deficiency increased regulatory T-cell and macrophage populations, including FoxP3(+) T cells (P = 0.005) and Tim-3(+) macrophages (P = 0.004). Thus, TLR3 prevents the progression from myocarditis to iDCM following CVB3 infection by reducing acute viral replication and IL-4 levels in the heart.

Factors that Predict HPV Vaccination Behavior Among Young Men-Who-Have-Sex-with-Men in the Greater Philadelphia Region.

Human papillomavirus (HPV) is one of the most common sexually transmitted infections in the USA. HPV is acknowledged as one of the leading causes of anal cancer, with an increased risk in men who have sex with men (MSM), when compared to age-matched heterosexual men. This study highlights the various factors that influence and impede HPV vaccination uptake among a multiracial cohort of young-MSM (YMSM). A total of 444 participants aged 18 to 27 in the Greater Philadelphia region completed an online survey. Approximately 75.79% (n = 335) of participants did not receive at least one dose of the HPV vaccine. Having a healthcare provider recommendation (OR = 25.54, 95% CI: 25.54-85.42, p < 0.001) and a one unit increase in experiences of adverse effects of stigma and homophobia (OR = 1.06, 95% CI: 1.01-1.11, p = 0.044) were associated with a greater likelihood of receiving the HPV vaccine uptake. Having a greater number of sexual partners (OR = 0.85, 95% CI: 0.75-0.97, p = 0.014) and having had condomless anal sex in the past 6 months (OR = 0.31, 95% CI: 0.15-0.58, p < 0.001) were associated with a lower odds of HPV vaccine uptake. In conclusion, healthcare provider-focused interventions and educational programs are needed to increase awareness and uptake of the HPV vaccine to mitigate the risks associated with sexual behaviors among this population.

Sex differences in mitochondrial gene expression during viral myocarditis.

Background: Myocarditis is an inflammation of the heart muscle most often caused by an immune response to viral infections. Sex differences in the immune response during myocarditis have been well described but upstream mechanisms in the heart that might influence sex differences in disease are not completely understood. Methods: Male and female BALB/c wild type mice received an intraperitoneal injection of heart-passaged coxsackievirus B3 (CVB3) or vehicle control. Bulk-tissue RNA-sequencing was conducted to better understand sex differences in CVB3 myocarditis. We performed enrichment analysis to understand sex differences in the transcriptional landscape of myocarditis and identify candidate transcription factors that might drive sex differences in myocarditis. Results: The hearts of male and female mice with myocarditis were significantly enriched for pathways related to an innate and adaptive immune response compared to uninfected controls. When comparing females to males with myocarditis, males were enriched for inflammatory pathways and gene changes that suggested worse mitochondrial transcriptional support (e.g., mitochondrial electron transport genes). In contrast, females were enriched for pathways related to mitochondrial respiration and bioenergetics, which were confirmed by higher transcript levels of master regulators of mitochondrial function including peroxisome proliferator-activated receptor gamma coactivator 1 (PGC1α), nuclear respiratory factor 1 (NRF1) and estrogen-related receptor alpha (ERRα). TRANSFAC analysis identified ERRa as a transcription factor that may mediate sex differences in mitochondrial function during myocarditis. Conclusions: Master regulators of mitochondrial function were elevated in females with myocarditis compared to males and may promote sex differences in mitochondrial respiratory transcript expression during viral myocarditis resulting in less severe myocarditis in females following viral infection.

Testosterone and interleukin-1ß increase cardiac remodeling during coxsackievirus B3 myocarditis via serpin A 3n.

Myocarditis and dilated cardiomyopathy (DCM) are often caused by viral infections and occur more frequently in men than in women, but the reasons for the sex difference remain unclear. The aim of this study was to assess whether gene changes in the heart during coxsackievirus B3 (CVB3) myocarditis in male and female BALB/c mice predicted worse DCM in males. Although myocarditis (P = 4.2 × 10(-5)) and cardiac dilation (P = 0.008) were worse in males, there was no difference in viral replication in the heart. Fibrotic remodeling genes, such as tissue inhibitor of metalloproteinase (TIMP)-1 and serpin A 3n, were upregulated in males during myocarditis rather than during DCM. Using gonadectomy and testosterone replacement, we showed that testosterone increased cardiac TIMP-1 (P = 0.04), serpin A 3n (P = 0.007), and matrix metalloproteinase (MMP)-8 (P = 0.04) during myocarditis. Testosterone increased IL-1ß levels in the heart (P = 0.02), a cytokine known to regulate cardiovascular remodeling, and IL-1ß in turn increased cardiac serpin A 3n mRNA (P = 0.005). We found that 39 of 118 (33%) genes identified in acute DCM patients were significantly altered in the heart during CVB3 myocarditis in mice, including serpin A 3n (3.3-fold change, P = 0.0001). Recombinant serpin A 3n treatment induced cardiac fibrosis during CVB3 myocarditis (P = 0.0008) while decreasing MMP-3 (P = 0.04) and MMP-9 (P = 0.03) levels in the heart. Thus, serpin A 3n was identified as a gene associated with fibrotic cardiac remodeling and progression to DCM in male myocarditis patients and mice.

Th2 regulation of viral myocarditis in mice: different roles for TLR3 versus TRIF in progression to chronic disease.

Viral infections are able to induce autoimmune inflammation in the heart. Here, we investigated the role of virus-activated Toll-like receptor (TLR)3 and its adaptor TRIF on the development of autoimmune coxsackievirus B3 (CVB3) myocarditis in mice. Although TLR3- or TRIF-deficient mice developed similarly worse acute CVB3 myocarditis and viral replication compared to control mice, disease was significantly worse in TRIF compared to TLR3-deficient mice. Interestingly, TLR3-deficient mice developed an interleukin (IL)-4-dominant T helper (Th)2 response during acute CVB3 myocarditis with elevated markers of alternative activation, while TRIF-deficient mice elevated the Th2-associated cytokine IL-33. Treatment of TLR3-deficient mice with recombinant IL-33 improved heart function indicating that elevated IL-33 in the context of a classic Th2-driven response protects against autoimmune heart disease. We show for the first time that TLR3 versus TRIF deficiency results in different Th2 responses that uniquely influence the progression to chronic myocarditis.

Mitochondrial Quality Control in the Heart: The Balance between Physiological and Pathological Stress.

Alterations in mitochondrial function and morphology are critical adaptations to cardiovascular stress, working in concert in an attempt to restore organelle-level and cellular-level homeostasis. Processes that alter mitochondrial morphology include fission, fusion, mitophagy, and biogenesis, and these interact to maintain mitochondrial quality control. Not all cardiovascular stress is pathologic (e.g., ischemia, pressure overload, cardiotoxins), despite a wealth of studies to this effect. Physiological stress, such as that induced by aerobic exercise, can induce morphologic adaptations that share many common pathways with pathological stress, but in this case result in improved mitochondrial health. Developing a better understanding of the mechanisms underlying alterations in mitochondrial quality control under diverse cardiovascular stressors will aid in the development of pharmacologic interventions aimed at restoring cellular homeostasis.

A national multi-institutional analysis of predictors of surgical site complications and unplanned reoperation after paramedian forehead flap reconstruction.

Background: Although postoperative complications of paramedian forehead flap (PMFF) are generally low, surgical site complications and unplanned reoperation can still occur. Recent literature suggests infection to be the most common complication following PMFF reconstruction. This study sought to determine the patient and preoperative factors associated with surgical site complications and unplanned reoperations at a national level. Methods: Patients who underwent PMFF reconstruction from the ACS-NSQIP 2007 - 2019 database were analyzed to determine composite surgical site morbidity and unplanned return to the operating room. Patient and operative factors were also analyzed to assess independent risk factors for surgical site morbidity and unplanned reoperation in the first 30 postoperative days. Results: A total of 1,592 PMFF were analyzed between 2007 and 2019. Of these, 2.7% (43/1592) developed a composite surgical site complication in the first 30 postoperative days. Risk factors for composite surgical site complication included >10% weight loss in the previous 6 months (p<0.05), disseminated cancer (p<0.01), class 4 wounds (dirty/infected) (p<0.01), and operative time greater than 123 min (p<0.01). Based on the univariate analysis, low preoperative albumin and hematocrit were also associated with increased odds of composite surgical site complication. 2.5% (40/1592) of patients underwent unplanned reoperation. Higher ASA class (p<0.05) and class 4 wounds (p<0.05) were associated with unplanned return to the operating room. Conclusion: Significant weight loss, disseminated cancer, prolonged operation time, low preoperative albumin, and hematocrit are associated with higher PMFF composite surgical site complications. Higher ASA and class 4 wound status are associated with an increased risk of unplanned reoperation.

ß2-Adrenergic Signaling Modulates Mitochondrial Function and Morphology in Skeletal Muscle in Response to Aerobic Exercise.

The molecular mechanisms underlying skeletal muscle mitochondrial adaptations induced by aerobic exercise (AE) are not fully understood. We have previously shown that AE induces mitochondrial adaptations in cardiac muscle, mediated by sympathetic stimulation. Since direct sympathetic innervation of neuromuscular junctions influences skeletal muscle homeostasis, we tested the hypothesis that ß2-adrenergic receptor (ß2-AR)-mediated sympathetic activation induces mitochondrial adaptations to AE in skeletal muscle. Male FVB mice were subjected to a single bout of AE on a treadmill (80% Vmax, 60 min) under ß2-AR blockade with ICI 118,551 (ICI) or vehicle, and parameters of mitochondrial function and morphology/dynamics were evaluated. An acute bout of AE significantly increased maximal mitochondrial respiration in tibialis anterior (TA) isolated fiber bundles, which was prevented by ß2-AR blockade. This increased mitochondrial function after AE was accompanied by a change in mitochondrial morphology towards fusion, associated with increased Mfn1 protein expression and activity. ß2-AR blockade fully prevented the increase in Mfn1 activity and reduced mitochondrial elongation. To determine the mechanisms involved in mitochondrial modulation by ß2-AR activation in skeletal muscle during AE, we used C2C12 myotubes, treated with the non-selective ß-AR agonist isoproterenol (ISO) in the presence of the specific ß2-AR antagonist ICI or during protein kinase A (PKA) and Gαi protein blockade. Our in vitro data show that ß-AR activation significantly increases mitochondrial respiration in myotubes, and this response was dependent on ß2-AR activation through a Gαs-PKA signaling cascade. In conclusion, we provide evidence for AE-induced ß2-AR activation as a major mechanism leading to alterations in mitochondria function and morphology/dynamics. ß2-AR signaling is thus a key-signaling pathway that contributes to skeletal muscle plasticity in response to exercise.

Sex differences in inflammation, redox biology, mitochondria and autoimmunity.

Autoimmune diseases are characterized by circulating antibodies and immune complexes directed against self-tissues that result in both systemic and organ-specific inflammation and pathology. Most autoimmune diseases occur more often in women than men. One exception is myocarditis, which is an inflammation of the myocardium that is typically caused by viral infections. Sex differences in the immune response and the role of the sex hormones estrogen and testosterone are well established based on animal models of autoimmune viral myocarditis as well as in mitochondrial function leading to reactive oxygen species production. RNA viruses like coxsackievirus B3, the primary cause of myocarditis in the US, activate the inflammasome through mitochondrial antiviral signaling protein located on the mitochondrial outer membrane. Toll-like receptor 4 and the inflammasome are the primary signaling pathways that increase inflammation during myocarditis, which is increased by testosterone. This review describes what is known about sex differences in inflammation, redox biology and mitochondrial function in the male-dominant autoimmune disease myocarditis and highlights gaps in the literature and future directions.

Gonadectomy of male BALB/c mice increases Tim-3(+) alternatively activated M2 macrophages, Tim-3(+) T cells, Th2 cells and Treg in the heart during acute coxsackievirus-induced myocarditis.

The incidence of cardiovascular disease, including inflammatory heart diseases like myocarditis, is increased in men. Similarly, male BALB/c mice infected with coxsackievirus B3 (CVB3) develop more severe acute inflammation in the heart compared to females. To better understand the effect of male sex hormones on cardiac inflammation, we gonadectomized (Gdx) male BALB/c mice and examined acute CVB3-induced myocarditis compared to sham controls. Viral replication in the heart was not significantly altered between Gdx and sham mice. However, gonadectomy significantly reduced testosterone levels and inflammation in the heart. FACS analysis of cell populations isolated from the heart revealed that CD11b(+) cells were significantly reduced in Gdx males. However, a GR1(+)F4/80(+) subset of CD11b(+) cells was significantly increased. Because this subset also expressed the interleukin (IL)-4R and IL-10, we refer to these cells as "alternatively activated" or M2 macrophages. A greater percentage of M2 macrophages in Gdx males expressed the inhibitory receptor Tim-3, while fewer expressed IL-1beta and IL-10. Only M2 macrophages upregulated TLR4 and Tim-3, whereas GR1(-)IL-4R(lo) macrophages did not. Additionally, IL-4(+)CD4(+) Th2 cells, Foxp3(+) regulatory T (Treg) cells and Tim-3(+)CD4(+) T cells were significantly increased in the heart following Gdx. Thus, we report for the first time that the inhibitory receptor Tim-3 is expressed on M2 macrophages. Our findings show that sex hormones and/or other mediators released from the testes inhibit anti-inflammatory populations in the heart including Tim-3(+) M2, Tim-3(+)CD4(+) T cells, Th2 and Treg resulting in more severe acute cardiac inflammation in males following CVB3 infection.

Elevated Sera sST2 Is Associated With Heart Failure in Men ≤50 Years Old With Myocarditis.

Background Myocarditis is an important cause of acute and chronic heart failure. Men with myocarditis have worse recovery and an increased need for transplantation compared with women, but the reason for the sex difference remains unclear. Elevated sera soluble (s) ST2 predicts mortality from acute and chronic heart failure, but has not been studied in myocarditis patients. Methods and Results Adults with a diagnosis of clinically suspected myocarditis (n=303, 78% male) were identified according to the 2013 European Society of Cardiology position statement. Sera sST2 levels were examined by ELISA in humans and mice and correlated with heart function according to sex and age. Sera sST2 levels were higher in healthy men ( P=8×10-6) and men with myocarditis ( P=0.004) compared with women. sST2 levels were elevated in patients with myocarditis and New York Heart Association class III - IV heart failure ( P=0.002), predominantly in men ( P=0.0003). Sera sST2 levels were associated with New York Heart Association class in men with myocarditis who were ≤50 years old ( r=0.231, P=0.0006), but not in women ( r=0.172, P=0.57). Sera sST2 levels were also significantly higher in male mice with myocarditis ( P=0.005) where levels were associated with cardiac inflammation. Gonadectomy with hormone replacement showed that testosterone ( P<0.001), but not estradiol ( P=0.32), increased sera sST2 levels in male mice with myocarditis. Conclusions We show in a well-characterized subset of heart failure patients with clinically suspected and biopsy-confirmed myocarditis that elevated sera sST2 is associated with an increased risk of heart failure based on New York Heart Association class in men ≤50 years old.

miR-21 is associated with fibrosis and right ventricular failure.

Combined pulmonary insufficiency (PI) and stenosis (PS) is a common long-term sequela after repair of many forms of congenital heart disease, causing progressive right ventricular (RV) dilation and failure. Little is known of the mechanisms underlying this combination of preload and afterload stressors. We developed a murine model of PI and PS (PI+PS) to identify clinically relevant pathways and biomarkers of disease progression. Diastolic dysfunction was induced (restrictive RV filling, elevated RV end-diastolic pressures) at 1 month after generation of PI+PS and progressed to systolic dysfunction (decreased RV shortening) by 3 months. RV fibrosis progressed from 1 month (4.4% ± 0.4%) to 3 months (9.2% ± 1%), along with TGF-ß signaling and tissue expression of profibrotic miR-21. Although plasma miR-21 was upregulated with diastolic dysfunction, it was downregulated with the onset of systolic dysfunction), correlating with RV fibrosis. Plasma miR-21 in children with PI+PS followed a similar pattern. A model of combined RV volume and pressure overload recapitulates the evolution of RV failure unique to patients with prior RV outflow tract surgery. This progression was characterized by enhanced TGF-ß and miR-21 signaling. miR-21 may serve as a plasma biomarker of RV failure, with decreased expression heralding the need for valve replacement.

Nat1 Deficiency Is Associated with Mitochondrial Dysfunction and Exercise Intolerance in Mice.

We recently identified human N-acetyltransferase 2 (NAT2) as an insulin resistance (IR) gene. Here, we examine the cellular mechanism linking NAT2 to IR and find that Nat1 (mouse ortholog of NAT2) is co-regulated with key mitochondrial genes. RNAi-mediated silencing of Nat1 led to mitochondrial dysfunction characterized by increased intracellular reactive oxygen species and mitochondrial fragmentation as well as decreased mitochondrial membrane potential, biogenesis, mass, cellular respiration, and ATP generation. These effects were consistent in 3T3-L1 adipocytes, C2C12 myoblasts, and in tissues from Nat1-deficient mice, including white adipose tissue, heart, and skeletal muscle. Nat1-deficient mice had changes in plasma metabolites and lipids consistent with a decreased ability to utilize fats for energy and a decrease in basal metabolic rate and exercise capacity without altered thermogenesis. Collectively, our results suggest that Nat1 deficiency results in mitochondrial dysfunction, which may constitute a mechanistic link between this gene and IR.

Human induced pluripotent stem cell-derived cardiomyocytes recapitulate the predilection of breast cancer patients to doxorubicin-induced cardiotoxicity.

Doxorubicin is an anthracycline chemotherapy agent effective in treating a wide range of malignancies, but it causes a dose-related cardiotoxicity that can lead to heart failure in a subset of patients. At present, it is not possible to predict which patients will be affected by doxorubicin-induced cardiotoxicity (DIC). Here we demonstrate that patient-specific human induced pluripotent stem cell-derived cardiomyocytes (hiPSC-CMs) can recapitulate the predilection to DIC of individual patients at the cellular level. hiPSC-CMs derived from individuals with breast cancer who experienced DIC were consistently more sensitive to doxorubicin toxicity than hiPSC-CMs from patients who did not experience DIC, with decreased cell viability, impaired mitochondrial and metabolic function, impaired calcium handling, decreased antioxidant pathway activity, and increased reactive oxygen species production. Taken together, our data indicate that hiPSC-CMs are a suitable platform to identify and characterize the genetic basis and molecular mechanisms of DIC.

Sex differences in translocator protein 18 kDa (TSPO) in the heart: implications for imaging myocardial inflammation.

Myocarditis is more severe in men than in women and difficult to diagnose due to a lack of imaging modalities that directly detect myocardial inflammation. Translocator protein 18 kDa (TSPO) is used extensively to image brain inflammation due to its presence in CD11b(+) brain microglia. In this study, we examined expression of TSPO and CD11b in mice with coxsackievirus B3 (CVB3) myocarditis and biopsy sections from myocarditis patients in order to determine if it could be used to image myocarditis. We found that male mice with CVB3 myocarditis upregulated more genes associated with TSPO activation than female mice. TSPO expression was increased in the heart of male mice and men with myocarditis compared with female subjects due to testosterone, where it was expressed predominantly in CD11b(+) immune cells. We show that TSPO ligands detect myocardial inflammation using microSPECT, with increased uptake of [(125)I]-IodoDPA-713 in male mice with CVB3 myocarditis compared with undiseased controls.