Description: Oxidative stress is acknowledged to play a role in kidney disease progression. Genetic variants that affect the capacity to handle oxidative stress may therefore influence the outcome of kidney disease. We examined whether genetic variants of the GSTM1 gene, a member of a superfamily of glutathione S -transferases, influence the course of kidney disease progression in participants of the African American Study of Kidney Disease (AASK) trial. Groups with and without the common GSTM1 null allele, GSTM1(0) , differed significantly in the time to a glomerular filtration rate (GFR) event or dialysis ( P = 0.04) and in the time to GFR event, dialysis, or death ( P = 0.02). The hazard ratios (HR) for the time to a GFR event or dialysis in those with two or one null allele relative to those possessing none were 1.88 [95% confidence interval (CI), 1.07 to 3.30, P = 0.03] and 1.68 (95% CI, 1.00 to 2.84, P 〈 0.05), respectively. For the time to GFR event, dialysis, or death, the HR for two null alleles was 2.06 (95% CI, 1.20 to 3.55, P = 0.01) and for one null allele 1.70 (95% CI, 1.02 to 2.81, P = 0.04). We demonstrated that GSTM1 directly regulates intracellular levels of 4-hydroxynonenal (4-HNE) in vascular smooth muscle cells. Furthermore, we showed that renal 4-HNE levels and GSTM1 are both increased after reduction of renal mass (RRM) in the mouse. We conclude that GSTM1 is normally upregulated in chronic kidney disease (CKD) in a protective response to increased oxidative stress. A genetic variant that results in loss of GSTM1 activity may be deleterious in CKD.

Description: The pathogenesis of gastroesophageal reflux disease (GERD) remains elusive, but recent evidence suggests that early secretion of inflammatory cytokines and chemokines by the mucosa leads to influx of immune cells followed by tissue damage. We previously showed that exposure of esophageal mucosa to HCl causes ATP release, resulting in activation of acetyl-CoA:1- O -alkyl- sn -glycero-3-phosphocholine acetyltransferase (lyso-PAF AT), the enzyme responsible for the production of platelet-activating factor (PAF). In addition, HCl causes release of IL-8 from the esophageal mucosa. We demonstrate that esophageal epithelial cells secrete proinflammatory mediators in response to HCl and that this response is mediated by ATP. Monolayers of the human esophageal epithelial cell line HET-1A were exposed to acidified cell culture medium (pH 5) for 12 min, a total of seven times over 48 h, to simulate the recurrent acid exposure clinically occurring in GERD. HCl upregulated mRNA and protein expression for the acid-sensing transient receptor potential cation channel, subfamily vanilloid member 1 (TRPV1), lyso-PAF AT, IL-8, eotaxin-1, -2, and -3, macrophage inflammatory protein-1α, and monocyte chemoattractant protein-1. The chemokine profile secreted by HET-1A cells in response to repeated HCl exposure parallels similar findings in erosive esophagitis patients. In HET-1A cells, the TRPV1 agonist capsaicin reproduced these findings for mRNA of the inflammatory mediators lyso-PAF AT, IL-8, and eotaxin-1. These effects were blocked by the TRPV1 antagonists iodoresiniferatoxin and JNJ-17203212. These effects were imitated by direct application of ATP and blocked by the nonselective ATP antagonist suramin. We conclude that HCl/TRPV-induced ATP release upregulated secretion of various chemoattractants by esophageal epithelial cells. These chemoattractants are selective for leukocyte subsets involved in acute inflammatory responses and allergic inflammation. The data support the validity of HET-1A cells as a model of the response of the human esophageal mucosa in GERD.

Description: Background— The cardiac cytoskeleton plays key roles in maintaining myocyte structural integrity in health and disease. In fact, human mutations in cardiac cytoskeletal elements are tightly linked to cardiac pathologies, including myopathies, aortopathies, and dystrophies. Conversely, the link between cytoskeletal protein dysfunction and cardiac electric activity is not well understood and often overlooked in the cardiac arrhythmia field. Methods and Results— Here, we uncover a new mechanism for the regulation of cardiac membrane excitability. We report that βII spectrin, an actin-associated molecule, is essential for the posttranslational targeting and localization of critical membrane proteins in heart. βII spectrin recruits ankyrin-B to the cardiac dyad, and a novel human mutation in the ankyrin-B gene disrupts the ankyrin-B/βII spectrin interaction, leading to severe human arrhythmia phenotypes. Mice lacking cardiac βII spectrin display lethal arrhythmias, aberrant electric and calcium handling phenotypes, and abnormal expression/localization of cardiac membrane proteins. Mechanistically, βII spectrin regulates the localization of cytoskeletal and plasma membrane/sarcoplasmic reticulum protein complexes, including the Na/Ca exchanger, ryanodine receptor 2, ankyrin-B, actin, and αII spectrin. Finally, we observe accelerated heart failure phenotypes in βII spectrin–deficient mice. Conclusions— Our findings identify βII spectrin as critical for normal myocyte electric activity, link this molecule to human disease, and provide new insight into the mechanisms underlying cardiac myocyte biology.

Description: Background— In the general population, blacks experience higher mortality than their white peers, attributed in part to their lower socioeconomic status, reduced access to care, and possibly intrinsic biological factors. Patients with kidney disease are a notable exception, among whom blacks experience lower mortality. It is unclear if similar differences affecting outcomes exist in patients with no kidney disease but with equal or similar access to health care. Methods and Results— We compared all-cause mortality, incident coronary heart disease, and incident ischemic stroke using multivariable-adjusted Cox models in a nationwide cohort of 547 441 black and 2 525 525 white patients with baseline estimated glomerular filtration rate ≥60 mL·min –1 ·1.73 m –2 receiving care from the US Veterans Health Administration. In parallel analyses, we compared outcomes in black versus white individuals in the National Health and Nutrition Examination Survey (NHANES) 1999 to 2004. After multivariable adjustments in veterans, black race was associated with 24% lower all-cause mortality (adjusted hazard ratio, 0.76; 95% confidence interval, 0.75–0.77; P 〈0.001) and 37% lower incidence of coronary heart disease (adjusted hazard ratio, 0.63; 95% confidence interval, 0.62–0.65; P 〈0.001) but a similar incidence of ischemic stroke (adjusted hazard ratio, 0.99; 95% confidence interval, 0.97–1.01; P =0.3). Black race was associated with a 42% higher adjusted mortality among individuals with estimated glomerular filtration rate ≥60 mL·min –1 ·1.73 m –2 in NHANES (adjusted hazard ratio, 1.42; 95% confidence interval, 1.09–1.87). Conclusions— Black veterans with normal estimated glomerular filtration rate and equal access to healthcare have lower all-cause mortality and incidence of coronary heart disease and a similar incidence of ischemic stroke. These associations are in contrast to the higher mortality experienced by black individuals in the general US population.

Description: We have recently shown that in vivo inhibition of histone deacetylase (HDAC) stimulates endogenous myocardial regeneration in infarcted hearts (Zhang L et al. J Pharmacol Exp Ther 341: 285–293, 2012). Furthermore, our observation demonstrates that HDAC inhibition promotes cardiogenesis, which is associated with HDAC4 reduction. However, it remains unknown as to whether specific inhibition of HDAC4 modulates cardiac stem cells (CSCs) to facilitate myocardial repair and to preserve cardiac performance. c-kit + CSCs were isolated from adult mouse hearts and were transfected with HDAC4 siRNA to knockdown HDAC4 of c-kit + CSCs. The transfection of HDAC4 siRNA caused a marked reduction of HDAC4 mRNA and proteins in c-kit + CSCs. Mouse myocardial infarction (MI) was created to assess the effect of HDAC4 inhibition in c-kit + CSCs on myocardial regeneration in vivo when cells were introduced into MI hearts. Transplantation of HDAC4 siRNA-treated c-kit + CSCs into MI hearts improved ventricular function, attenuated ventricular remodeling, and promoted CSC-derived regeneration and neovascularization. Furthermore, Ki67 and BrdU positively proliferative myocytes increased in MI hearts receiving HDAC4 siRNA-treated c-kit + CSCs compared with MI hearts engrafted with control siRNA-treated c-kit + CSCs. In addition, compared with MI hearts engrafted with control adenoviral GFP-infected c-kit + CSCs, MI hearts receiving adenoviral HDAC4-infected c-kit + CSCs exhibited attenuated cardiac functional recovery, CSC-derived regeneration, and neovascularization, which was accompanied with adverse ventricular remodeling and decrease in Ki67 and BrdU positively proliferative myocytes. HDAC4 inhibition facilitated c-kit + CSCs into the differentiation into cardiac lineage commitments in vitro, while HDAC4 overexpression attenuated c-kit + CSC-derived cardiogenesis. Our results indicate that HDAC4 inhibition promotes CSC-derived cardiac regeneration and improves the restoration of cardiac function.

Description: Background— Sugar-sweetened beverage (SSB) intake has been linked to abnormal abdominal adipose tissue. We examined the prospective association of habitual SSB intake and change in visceral adipose tissue (VAT) and subcutaneous adipose tissue. Methods and Results— The quantity (volume, cm 3 ) and quality (attenuation, Hounsfield Unit) of abdominal adipose tissue were measured using computed tomography in 1003 participants (mean age 45.3 years, 45.0% women) at examination 1 and 2 in the Framingham’s Third Generation cohort. The 2 exams were 6 years apart. At baseline, SSB and diet soda intake were assessed using a valid food frequency questionnaire. Participants were categorized into 4 groups: none to 〈1 serving/mo (nonconsumers), 1 serving/mo to 〈1 serving/week, 1 serving/week to 1 serving/d, and ≥1 serving/d (daily consumers) of either SSB or diet soda. After adjustment for multiple confounders including change in body weight, higher SSB intake was associated with greater change in VAT volume ( P trend〈0.001). VAT volume increased by 658 cm 3 (95% confidence interval [CI], 602 to 713), 649 cm 3 (95% CI, 582 to 716), 707 cm 3 (95% CI, 657 to 757), and 852 cm 3 (95% CI, 760 to 943) from nonconsumers to daily consumers. Higher SSB intake was also associated with greater decline of VAT attenuation ( P trend=0.007); however, the association became nonsignificant after additional adjustment for VAT volume change. In contrast, diet soda consumption was not associated with change in abdominal adipose tissue. Conclusions— Regular SSB intake was associated with adverse change in both VAT quality and quantity, whereas we observed no such association for diet soda.

Description: Silicon (Si) has long been known to play a major physiological and structural role in certain organisms, including diatoms, sponges, and many higher plants, leading to the recent identification of multiple proteins responsible for Si transport in a range of algal and plant species. In mammals, despite several convincing studies suggesting that silicon is an important factor in bone development and connective tissue health, there is a critical lack of understanding about the biochemical pathways that enable Si homeostasis. Here we report the identification of a mammalian efflux Si transporter, namely Slc34a2 (also termed NaPiIIb), a known sodium-phosphate cotransporter, which was upregulated in rat kidney following chronic dietary Si deprivation. Normal rat renal epithelium demonstrated punctate expression of Slc34a2, and when the protein was heterologously expressed in Xenopus laevis oocytes, Si efflux activity (i.e., movement of Si out of cells) was induced and was quantitatively similar to that induced by the known plant Si transporter Os Lsi2 in the same expression system. Interestingly, Si efflux appeared saturable over time, but it did not vary as a function of extracellular HPO42– or Na + concentration, suggesting that Slc34a2 harbors a functionally independent transport site for Si operating in the reverse direction to the site for phosphate. Indeed, in rats with dietary Si depletion-induced upregulation of transporter expression, there was increased urinary phosphate excretion. This is the first evidence of an active Si transport protein in mammals and points towards an important role for Si in vertebrates and explains interactions between dietary phosphate and silicon.

Description: Pulmonary hypertension (PH) is characterized by increased pulmonary vascular resistance, pulmonary vascular remodeling, and increased pulmonary vascular pressures that often result in right ventricular dysfunction, leading to right heart failure. Evidence suggests that reactive oxygen species (ROS) contribute to PH pathogenesis by altering pulmonary vascular cell proliferation and intracellular signaling pathways. However, the role of mitochondrial antioxidants and oxidant-derived stress signaling in the development of hypoxia-induced PH is largely unknown. Therefore, we examined the role of the major mitochondrial redox regulator thioredoxin 2 (Trx2). Levels of Trx2 mRNA and protein were examined in human pulmonary arterial endothelial cells (HPAECs) and smooth muscle cells (HPASMCs) exposed to hypoxia, a common stimulus for PH, for 72 h. Hypoxia decreased Trx2 mRNA and protein levels. In vitro overexpression of Trx2 reduced hypoxia-induced H 2 O 2 production. The effects of increased Trx2 protein level were examined in transgenic mice expressing human Trx2 (Tg hTrx2 ) that were exposed to hypoxia (10% O 2 ) for 3 wk. Tg hTrx2 mice exposed to hypoxia had exacerbated increases in right ventricular systolic pressures, right ventricular hypertrophy, and increased ROS in the lung tissue. Trx2 overexpression did not attenuate hypoxia-induced increases in Trx2 oxidation or Nox4 expression. Expression of a dominant negative C93S Trx2 mutant that mimics Trx2 oxidation exacerbated hypoxia-induced increases in HPASMC H 2 O 2 levels and cell proliferation. In conclusion, Trx2 overexpression failed to attenuate hypoxia-induced HPASMC proliferation in vitro or hypoxia-induced PH in vivo. These findings indicate that strategies to enhance Trx2 expression are unlikely to exert therapeutic effects in PH pathogenesis.

Description: An increase in intracellular Ca 2+ concentration ([Ca 2+ ] i ) augments late sodium current ( I Na.L ) in cardiomyocytes. This study tests the hypothesis that both Ca 2+ -calmodulin-dependent protein kinase II (CaMKII) and protein kinase C (PKC) mediate the effect of increased [Ca 2+ ] i to increase I Na.L . Whole cell and open cell-attached patch clamp techniques were used to record I Na.L in rabbit ventricular myocytes dialyzed with solutions containing various concentrations of [Ca 2+ ] i . Dialysis of cells with [Ca 2+ ] i from 0.1 to 0.3, 0.6, and 1.0 μM increased I Na.L in a concentration-dependent manner from 0.221 ± 0.038 to 0.554 ± 0.045 pA/pF ( n = 10, P 〈 0.01) and was associated with an increase in mean Na + channel open probability and prolongation of channel mean open-time ( n = 7, P 〈 0.01). In the presence of 0.6 μM [Ca 2+ ] i , KN-93 (10 μM) and bisindolylmaleimide (BIM, 2 μM) decreased I Na.L by 45.2 and 54.8%, respectively. The effects of KN-93 and autocamtide-2-related inhibitory peptide II (2 μM) were not different. A combination of KN-93 and BIM completely reversed the increase in I Na.L as well as the Ca 2+ -induced changes in Na + channel mean open probability and mean open-time induced by 0.6 μM [Ca 2+ ] i . Phorbol myristoyl acetate increased I Na.L in myocytes dialyzed with 0.1 μM [Ca 2+ ] i ; the effect was abolished by Gö-6976. In summary, both CaMKII and PKC are involved in [Ca 2+ ] i -mediated augmentation of I Na.L in ventricular myocytes. Inhibition of CaMKII and/or PKC pathways may be a therapeutic target to reduce myocardial dysfunction and cardiac arrhythmias caused by calcium overload.