An increasing body of evidence points to mitochondrial dysfunction as a contributor to the molecular pathogenesis of neurodegenerative diseases such as Parkinson's disease. Recent studies of the Parkinson's disease associated genes PINK1 (ref. 2) and parkin (PARK2, ref. 3) indicate that they may act in a quality control pathway preventing the accumulation of dysfunctional mitochondria. Here we elucidate regulators that have an impact on parkin translocation to damaged mitochondria with genome-wide small interfering RNA (siRNA) screens coupled to high-content microscopy. Screening yielded gene candidates involved in diverse cellular processes that were subsequently validated in low-throughput assays. This led to characterization of TOMM7 as essential for stabilizing PINK1 on the outer mitochondrial membrane following mitochondrial damage. We also discovered that HSPA1L (HSP70 family member) and BAG4 have mutually opposing roles in the regulation of parkin translocation. The screens revealed that SIAH3, found to localize to mitochondria, inhibits PINK1 accumulation after mitochondrial insult, reducing parkin translocation. Overall, our screens provide a rich resource to understand mitochondrial quality control.

Activation of self-reactive T cells and their trafficking to target tissues leads to autoimmune organ destruction. Mice lacking the co-inhibitory receptor cytotoxic T lymphocyte antigen-4 (CTLA-4) develop fatal autoimmunity characterized by lymphocytic infiltration into nonlymphoid tissues. Here, we demonstrate that the CD28 co-stimulatory pathway regulates the trafficking of self-reactive Ctla4(-/-) T cells to tissues. Concurrent ablation of the CD28-activated Tec family kinase ITK does not block spontaneous T cell activation but instead causes self-reactive Ctla4(-/-) T cells to accumulate in secondary lymphoid organs. Despite excessive spontaneous T cell activation and proliferation in lymphoid organs, Itk(-/-); Ctla4(-/-) mice are otherwise healthy, mount antiviral immune responses and exhibit a long lifespan. We propose that ITK specifically licenses autoreactive T cells to enter tissues to mount destructive immune responses. Notably, ITK inhibitors mimic the null mutant phenotype and also prevent pancreatic islet infiltration by diabetogenic T cells in mouse models of type 1 diabetes, highlighting their potential utility for the treatment of human autoimmune disorders.

BACKGROUND & AIMS: Upper-body, i.e. visceral, obesity is associated with insulin resistance and impaired protein synthesis. It is unclear whether postprandial stimulation of protein synthesis is affected by body fat distribution. We investigated the postprandial protein anabolic response in a cohort of obese women. METHODS: Participants were studied after an overnight fast and after a mixed meal, grouped as upper-body obese (UBO, waist-to-hip ratio, WHR, >0.85, n = 6) vs. lower-body obese (LBO, WHR <0.80, n = 7). Lipid and carbohydrate metabolism were assessed by measurements of plasma free fatty acids (FFA), insulin and glucose plasma concentrations, and calculation of the Quicki index from fasting glucose and insulin values. Different labels of stable isotopes of phenylalanine were administered intravenously and orally, and leg and whole-body protein breakdown and synthesis were calculated from phenylalanine/tyrosine isotopic enrichments in femoral arterial and venous blood, using equations for steady-state kinetics. Data are denoted as mean ± SD. RESULTS: Age (38 vs. 40, p = 0.549) and body-mass index (33.7 ± 1.9 vs. 35.0 ± 1.8, p = 0.241) were similar in both groups. UBO subjects had more visceral fat (p = 0.002) and higher fat-free body mass (FFM) (p = 0.015). Plasma insulin concentrations were greater in UBO than LBO women (p = 0.013), and UBO were less insulin sensitive (Quicki = 0.32 ± 0.01 vs. 0.36 ± 0.02, p = 0.005). Protein kinetics across the leg were not different between groups. Fasting whole body protein balance was similarly negative in both groups (UBO -6.5 ± 2.4 vs. LBO -7.6 ± 0.9 μmol/kgFFM/h, p = 1.0). Postprandially, whole body protein balance became less positive in UBO than in LBO (14.8 ± 3.7 vs. 20.2 ± 3.7 μmol/kgFFM/h, p = 0.017). CONCLUSIONS: Whole-body protein balance following a meal is less positive in upper-body obese, insulin-resistant, women than in lower-body obese women.

A human in vitro cardiac tissue model would be a significant advancement for understanding, studying, and developing new strategies for treating cardiac arrhythmias and related cardiovascular diseases. We developed an in vitro model of three-dimensional (3D) human cardiac tissue by populating synthetic filamentous matrices with cardiomyocytes derived from healthy wild-type volunteer (WT) and patient-specific long QT syndrome type 3 (LQT3) induced pluripotent stem cells (iPS-CMs) to mimic the condensed and aligned human ventricular myocardium. Using such a highly controllable cardiac model, we studied the contractility malfunctions associated with the electrophysiological consequences of LQT3 and their response to a panel of drugs. By varying the stiffness of filamentous matrices, LQT3 iPS-CMs exhibited different level of contractility abnormality and susceptibility to drug-induced cardiotoxicity.

Over the last two decades, essential hypertension has become common in adolescents, yet remains under-diagnosed in absence of symptoms. Diagnosis is based on normative percentiles that factor in age, sex and height. Evaluation is more similar to adult essential hypertension than childhood secondary hypertension. Modifiable risk factors such as obesity, sodium consumption and low exercise should be addressed first. Many anti-hypertensive medications now have specific regulatory approval for children. Sports participation need not be limited in mild or well-controlled cases. Primary care physicians play an important role in reduction of cardiovascular mortality by early detection and referral when needed.

OBJECTIVE: Dual specificity phosphatase 1 (DUSP1) inhibits mitogen activated protein kinase activity, and is activated by several stimuli such as sustained hypoxia, oxidative stress, and hormones. However, the effect of intermittent hypoxia is not known. The aim of this study was to evaluate the role of intermittent hypoxia on DUSP1 expression, and to validate its role in a human model of intermittent hypoxia, as seen in obstructive sleep apnea (OSA). OSA is characterized by recurrent episodes of hypoxemia/reoxygenation and is a known risk factor for cardiovascular morbidity. METHODS: In-vitro studies using human coronary artery endothelial cells (HCAEC) and ex-vivo studies using white blood cells isolated from healthy and OSA subjects. RESULTS: Intermittent hypoxia induced DUSP1 expression in human coronary artery endothelial cells (HCAEC), and in granulocytes isolated from healthy human subjects. Functionally, DUSP1 increased the expression and activity of manganese superoxide dismutase (MnSOD) in HCAEC. Further, significant increases in DUSP1 mRNA from total blood, and in DUSP1 protein in mononuclear cells and granulocytes isolated from OSA subjects, were observed in the early morning hours after one night of intermittent hypoxemia due to untreated OSA. This early-morning OSA-induced augmentation of DUSP1 gene expression was attenuated by continuous positive airway pressure (CPAP) treatment of OSA. CONCLUSION: Intermittent hypoxia increases MnSOD activity via increased DUSP1 expression in HCAEC. Similarly, overnight intermittent hypoxemia in patients with OSA induces expression of DUSP1, which may mediate increases of MnSOD expression and activity. This may contribute significantly to neutralizing the effects of reactive oxygen species, a consequence of the intermittent hypoxemia/reperfusion elicited by OSA.

Women in prison have a higher prevalence of HIV than men. After release from prison, former inmates have the opportunity to engage in risk behaviors for HIV and other sexually transmitted infections (STIs). We sought to assess change in risk behaviors over time and the association of gender with risk behavior in the postrelease period. In this prospective cohort study, we interviewed 200 former inmates (51 women) approximately two weeks (baseline) and three months (follow-up) after release and tested them for HIV infection at follow-up. We examined the association of gender with unprotected vaginal or anal sex in the last seven days using chi-square and Fisher's exact tests and multivariable logistic regression. At baseline, 22% of men and 41% of women reported unprotected vaginal sex (p < 0.01) and 5% of men and 8% of women reported unprotected anal sex (p = 0.51). Being younger (OR for each decade increase 0.48, 95% CI = 0.29-0.80), being gay/lesbian or being bisexual (compared with being heterosexual, OR = 4.74, 95% CI = 1.01-22.17 and OR = 3.98, 95% CI = 1.41-11.26, respectively), or reporting a drug of choice of heroin/speedballs or cocaine/crack (compared with marijuana/no drug of choice, OR = 24.00, 95% CI = 5.15-111.81 and OR = 3.49, 95% CI = 1.20-10.18, respectively) was associated with unprotected vaginal or anal sex after adjusting for race, homelessness, and hazardous drinking. At follow-up, 21% of men and 44% of women reported unprotected sex (p = 0.005), and female gender (OR = 4.42, 95% CI = 1.79-10.94) and hazardous drinking (compared with not meeting criteria for hazardous drinking, OR = 3.64, 95% CI = 1.34-9.86) were associated with unprotected sex, adjusting for race and homelessness. In this population with a high prevalence of HIV, we demonstrated persistent engagement in sexual risk behavior during the postrelease period. Enhanced efforts to promote sexual health and reduced risk behavior among both male and female current and former prison inmates are needed, including improved access to preventive care and HIV and STI screening, testing, and treatment.

Merkel cell carcinoma (MCC) is an aggressive skin cancer with rising incidence. In this study, we demonstrate that mTOR activation and suppressed autophagy is common in MCCs. mTOR inhibition in two primary human MCC cell lines induces autophagy and cell death that is independent of caspase activation but can be attenuated by autophagy inhibition. This is the first study to evaluate mTOR and autophagy in MCC. Our data suggests a potential role of autophagic cell death upon mTOR inhibition and thus uncovers a previously underappreciated role of mTOR signaling and cell survival, and merits further studies for potential therapeutic targets.

We examined factors that affect decision-making for families presented with a phase I clinical trial of hepatocyte transplant as a potential alternative to liver transplant for their children among two groups: (i) families who were actually offered enrollment in the hepatocyte trial and; (ii) families whose children had liver transplants before the trial was available. We conducted semi-structured interviews about actual and hypothetical decision-making regarding trial participation and used grounded theory analysis to identify common themes. The most common motivator for participation was decline in the child's health. The most common deterrent was lack of data from prior hepatocyte transplants, particularly when compared with data available about liver transplant. Interviewees' point of comparison for evaluating relative benefits and risks of hepatocyte transplant oscillated between the alternative of doing nothing while waiting for a liver (the relevant alternative) vs. the alternative of getting a liver. These results suggest that families' reluctance to participate may result from misconceptions about severity of the child's disease, underestimating risks of liver transplant, or confusion about the role of hepatocyte transplant in the treatment pathway. Clarification of available treatment alternatives and associated risks as part of informed consent may improve the quality of decision-making regarding trial enrollment.

IMPORTANCE: Small studies suggest that low-dose dopamine or low-dose nesiritide may enhance decongestion and preserve renal function in patients with acute heart failure and renal dysfunction; however, neither strategy has been rigorously tested. OBJECTIVE: To test the 2 independent hypotheses that, compared with placebo, addition of low-dose dopamine (2 μg/kg/min) or low-dose nesiritide (0.005 μg/kg/min without bolus) to diuretic therapy will enhance decongestion and preserve renal function in patients with acute heart failure and renal dysfunction. DESIGN, SETTING, AND PARTICIPANTS: Multicenter, double-blind, placebo-controlled clinical trial (Renal Optimization Strategies Evaluation [ROSE]) of 360 hospitalized patients with acute heart failure and renal dysfunction (estimated glomerular filtration rate of 15-60 mL/min/1.73 m2), randomized within 24 hours of admission. Enrollment occurred from September 2010 to March 2013 across 26 sites in North America. INTERVENTIONS: Participants were randomized in an open, 1:1 allocation ratio to the dopamine or nesiritide strategy. Within each strategy, participants were randomized in a double-blind, 2:1 ratio to active treatment or placebo. The dopamine (n = 122) and nesiritide (n = 119) groups were independently compared with the pooled placebo group (n = 119). MAIN OUTCOMES AND MEASURES: Coprimary end points included 72-hour cumulative urine volume (decongestion end point) and the change in serum cystatin C from enrollment to 72 hours (renal function end point). RESULTS: Compared with placebo, low-dose dopamine had no significant effect on 72-hour cumulative urine volume (dopamine, 8524 mL; 95% CI, 7917-9131 vs placebo, 8296 mL; 95% CI, 7762-8830 ; difference, 229 mL; 95% CI, -714 to 1171 mL; P = .59) or on the change in cystatin C level (dopamine, 0.12 mg/L; 95% CI, 0.06-0.18 vs placebo, 0.11 mg/L; 95% CI, 0.06-0.16; difference, 0.01; 95% CI, -0.08 to 0.10; P = .72). Similarly, low-dose nesiritide had no significant effect on 72-hour cumulative urine volume (nesiritide, 8574 mL; 95% CI, 8014-9134 vs placebo, 8296 mL; 95% CI, 7762-8830; difference, 279 mL; 95% CI, -618 to 1176 mL; P = .49) or on the change in cystatin C level (nesiritide, 0.07 mg/L; 95% CI, 0.01-0.13 vs placebo, 0.11 mg/L; 95% CI, 0.06-0.16; difference, -0.04; 95% CI, -0.13 to 0.05; P = .36). Compared with placebo, there was no effect of low-dose dopamine or nesiritide on secondary end points reflective of decongestion, renal function, or clinical outcomes. CONCLUSION AND RELEVANCE: In participants with acute heart failure and renal dysfunction, neither low-dose dopamine nor low-dose nesiritide enhanced decongestion or improved renal function when added to diuretic therapy. TRIAL REGISTRATION: clinicaltrials.gov Identifier: NCT01132846.