Publications

  • Journal of the American Society of Nephrology : JASN. 07/2014;
  • Xiaonan H Wang, William E Mitch
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    ABSTRACT: In patients with chronic kidney disease (CKD), loss of cellular proteins increases the risks of morbidity and mortality. Persistence of muscle protein catabolism in CKD results in striking losses of muscle proteins as whole-body protein turnover is great; even small but persistent imbalances between protein synthesis and degradation cause substantial protein loss. No reliable methods to prevent CKD-induced muscle wasting currently exist, but mechanisms that control cellular protein turnover have been identified, suggesting that therapeutic strategies will be developed to suppress or block protein loss. Catabolic pathways that cause protein wasting include activation of the ubiquitin-proteasome system (UPS), caspase-3, lysosomes and myostatin (a negative regulator of skeletal muscle growth). These pathways can be initiated by complications associated with CKD, such as metabolic acidosis, defective insulin signalling, inflammation, increased angiotensin II levels, abnormal appetite regulation and impaired microRNA responses. Inflammation stimulates cellular signalling pathways that activate myostatin, which accelerates UPS-mediated catabolism. Blocking this pathway can prevent loss of muscle proteins. Myostatin inhibition could yield new therapeutic directions for blocking muscle protein wasting in CKD or disorders associated with its complications.
    Nature Reviews Nephrology 07/2014; · 7.94 Impact Factor
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    ABSTRACT: The mechanisms underlying the development of aging-induced muscle atrophy are unclear. By microRNA array and individual qPCR analyses, we found significant up-regulation of miR-29 in muscles of aged rodents vs. results in young. With aging, p85α, IGF-1 and B-myb muscle levels were lower while the expression of certain cell arrest proteins (p53, p16 and pRB) increased. When miR-29 was expressed in muscle progenitor cells (MPC), their proliferation was impaired while SA-βgal expression increased signifying the development of senescence. Impaired MPC proliferation resulted from interactions between miR-29 and the 3'-UTR of p85a, IGF-1 and B-myb, suppressing the translation of these mediators of myoblast proliferation. In vivo, electroporation of miR-29 into muscles of young mice suppressed the proliferation and increased levels of cellular arrest proteins, recapitulating aging-induced responses in muscle. A potential stimulus of miR-29 expression is Wnt-3a since we found that exogenous Wnt-3a stimulated miR-29 expression 2.7-fold in primary cultures of MPCs. Thus, aging-induced muscle senescence results from activation of miR-29 by Wnt-3a leading to suppressed expression of several signaling proteins (p85α, IGF-1 and B-myb) that act coordinately to impair the proliferation of MPCs contributing to muscle atrophy. The increase in miR-29 provides a potential mechanism for aging-induced sarcopenia.
    Aging 03/2014; · 4.70 Impact Factor
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    ABSTRACT: Neointima formation causes the failure of 60% of arteriovenous fistulas (AVFs) within 2 years. Neointima-forming mechanisms are controversial but possibly linked to excess proinflammatory responses and dysregulated Notch signaling. To identify how AVFs fail, we anastomosed the carotid artery to the internal jugular vein in normal and uremic mice and compared these findings with those in failed AVFs from patients with ESRD. Endothelial cells (ECs) of AVFs in uremic mice or patients expressed mesenchymal markers (FSP-1 and/or α-SMA) and exhibited increased expression and nuclear localization of Notch intracellular domain compared with ECs of AVFs in pair-fed control mice. Furthermore, expression of VE-Cadherin decreased, whereas expression of Notch1 and -4, Notch ligands, the downstream transcription factor of Notch, RBP-Jκ, and Notch target genes increased in ECs of AVFs in uremic mice. In cultured ECs, ectopic expression of Notch ligand or treatment with TGF-β1 triggered the expression of mesenchymal markers and induced endothelial cell barrier dysfunction, both of which were blocked by Notch inhibition or RBP-Jκ knockout. Furthermore, Notch-induced defects in barrier function, invasion of inflammatory cells, and neointima formation were suppressed in mice with heterozygous knockdown of endothelial-specific RBP-Jκ. These results suggest that increased TGF-β1, a complication of uremia, activates Notch in endothelial cells of AVFs, leading to accelerated neointima formation and AVF failure. Suppression of Notch activation could be a strategy for improving AFV function in uremia.
    Journal of the American Society of Nephrology 01/2014; · 8.99 Impact Factor
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    ABSTRACT: Early versus later start of dialysis is still a matter of debate. Low-protein diets have been used for many decades to delay dialysis initiation. Protein-restricted diets (0.3-0.6 g protein/kg/day) supplemented with essential amino acids and ketoanalogues (sVLPD) can be offered, in association with pharmacological treatment, to motivated stage 4-5 chronic kidney disease (CKD) patients not having severe comorbid conditions; they probably represent 30-40% of the concerned population. A satisfactory adherence to such dietary prescription is observed in approximately 50% of the patients. While the results of the studies on the effects of this diet on the rate of progression of renal failure remain inconclusive, they are highly significant when initiation of dialysis is the primary outcome. The correction of uremic symptoms allows for initiation of dialysis treatment at a level of residual renal function lower than that usually recommended. Most of the CKD-associated complications of cardiovascular and metabolic origin, which hamper both lifespan and quality of life, are positively influenced by the diet. Lastly, with regular monitoring jointly assumed by physicians and dietitians, nutritional status is well preserved as confirmed by a very low mortality rate and by the absence of detrimental effect on the long-term outcome of patients once renal replacement therapy is initiated. On account of its feasibility, efficacy and safety, sVLPD deserves a place in the management of selected patients to safely delay the time needed for dialysis.
    Seminars in Dialysis 09/2013; · 2.25 Impact Factor
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    ABSTRACT: Catabolic conditions like chronic kidney disease (CKD) cause loss of muscle mass by unclear mechanisms. In muscle biopsies from CKD patients, we found activated Stat3 (p-Stat3) and hypothesized that p-Stat3 initiates muscle wasting. We created mice with muscle-specific knockout (KO) that prevents activation of Stat3. In these mice, losses of body and muscle weights were suppressed in models with CKD or acute diabetes. A small-molecule that inhibits Stat3 activation produced similar responses, suggesting a potential for translation strategies. Using CCAAT/enhancer-binding protein δ (C/EBPδ) KO mice and C2C12 myotubes with knockdown of C/EBPδ or myostatin, we determined that p-Stat3 initiates muscle wasting via C/EBPδ, stimulating myostatin, a negative muscle growth regulator. C/EBPδ KO also improved survival of CKD mice. We verified that p-Stat3, C/EBPδ, and myostatin were increased in muscles of CKD patients. The pathway from p-Stat3 to C/EBPδ to myostatin and muscle wasting could identify therapeutic targets that prevent muscle wasting.
    Cell metabolism 09/2013; 18(3):368-79. · 17.35 Impact Factor
  • Xiaonan H Wang, William E Mitch
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    ABSTRACT: Muscle atrophy is a frequent complication of chronic kidney disease (CKD) and is associated with increased morbidity and mortality. The processes causing loss of muscle mass are also present in several catabolic conditions. Understanding the pathogenesis of CKD-induced muscle loss could lead to therapeutic interventions that prevent muscle wasting in CKD and potentially, other catabolic conditions. Major findings: Insulin or IGF-1 resistance caused by CKD, acidosis, inflammation, glucocorticoids or cancer causes defects in insulin-stimulated intracellular signaling that suppresses IRS-1 activity leading to decreased phosphorylation of Akt (p-Akt). A low p-Akt activates caspase-3 which provides muscle proteins substrates of the ubiquitin-proteasome system (UPS). A low p-Akt also leads to decreased phosphorylation of forkhead transcription factors which enter the nucleus to stimulate the expression of atrogin-1/MAFbx and MuRF1, E3 ubiquitin ligases that can be associated with proteolysis of muscle cells by the UPS. Caspase-3 also stimulates proteasome-dependent proteolysis in muscle. in CKD, diabetes, inflammatory conditions or in response to acidosis or excess glucocorticoids, insulin resistance develops, initiating reduced IRS-1/PI3K/Akt signaling. In CKD, this reduces p-Akt which stimulates muscle proteolysis by activating caspase-3 and the UPS. Second, caspase-3 cleaves actomyosin yielding substrates for the UPS and increased proteasome-mediated proteolysis. Third, p-Akt down-regulation suppresses myogenesis in CKD. Fourth, exercise in CKD stimulates insulin/IGF-1 signaling to reduce muscle atrophy. Lastly, there is evidence that microRNAs influence insulin signaling providing a potential opportunity to design therapeutic interventions.
    The international journal of biochemistry & cell biology 07/2013; · 4.89 Impact Factor
  • Biruh Workeneh, William E Mitch
    American Journal of Clinical Nutrition 07/2013; · 6.50 Impact Factor
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    ABSTRACT: Cachexia is a serious complication of many chronic diseases, such as congestive heart failure (CHF) and chronic kidney disease (CKD). Many factors are involved in the development of cachexia, and there is increasing evidence that angiotensin II (Ang II), the main effector molecule of the renin-angiotensin system (RAS), plays an important role in this process. Patients with advanced CHF or CKD often have increased Ang II levels and cachexia, and angiotensin-converting enzyme (ACE) inhibitor treatment improves weight loss. In rodent models, an increase in systemic Ang II leads to weight loss through increased protein breakdown, reduced protein synthesis in skeletal muscle and decreased appetite. Ang II activates the ubiquitin-proteasome system via generation of reactive oxygen species and via inhibition of the insulin-like growth factor-1 signaling pathway. Furthermore, Ang II inhibits 5' AMP-activated protein kinase (AMPK) activity and disrupts normal energy balance. Ang II also increases cytokines and circulating hormones such as tumor necrosis factor-α, interleukin-6, serum amyloid-A, glucocorticoids and myostatin, which regulate muscle protein synthesis and degradation. Ang II acts on hypothalamic neurons to regulate orexigenic/anorexigenic neuropeptides, such as neuropeptide-Y, orexin and corticotropin-releasing hormone, leading to reduced appetite. Also, Ang II may regulate skeletal muscle regenerative processes. Several clinical studies have indicated that blockade of Ang II signaling via ACE inhibitors or Ang II type 1 receptor blockers prevents weight loss and improves muscle strength. Thus the RAS is a promising target for the treatment of muscle atrophy in patients with CHF and CKD.
    The international journal of biochemistry & cell biology 06/2013; · 4.89 Impact Factor
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    ABSTRACT: Muscle wasting is associated with a wide range of catabolic diseases. This debilitating loss of muscle mass and functional capacity reduces the quality of life and increases the risks of morbidity and mortality. Major progress has been made in understanding the biochemical mechanisms and signaling pathways regulating muscle protein balance under normal conditions and the enhanced protein loss in atrophying muscles. It is now clear that activation of myostatin/activin signaling is critical in triggering the accelerated muscle catabolism that causes muscle loss in multiple disease states. Binding of myostatin and activin to the ActRIIB receptor complex on muscle cell membrane leads to activation of Smad2/3-mediated transcription, which in turn stimulates FoxO-dependent transcription and enhanced muscle protein breakdown via ubiquitin-proteasome system and autophagy. In addition, Smad activation inhibits muscle protein synthesis by suppressing Akt signaling. Pharmacological blockade of the myostatin/activin-ActRIIB pathway has been shown to prevent or reverse the loss of muscle mass and strength in various disease models including cancer cachexia and renal failure. Moreover, it can markedly prolong the lifespan of animals with cancer-associated muscle loss. Furthermore, inhibiting myostatin/activin actions also improves insulin sensitivity, reduces excessive adiposity, attenuates systemic inflammation, and accelerates bone fracture healing in disease models. Based on these exciting advances, the potential therapeutic benefits of myostatin/activin antagonism are now being tested in multiple clinical settings.
    The international journal of biochemistry & cell biology 05/2013; · 4.89 Impact Factor
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    ABSTRACT: Insulin resistance from chronic kidney disease (CKD) stimulates muscle protein wasting but mechanisms causing this resistance are controversial. To help resolve this, we used microarray analyses to identify initiators of insulin resistance in the muscles of mice with CKD, glucose intolerance, and insulin resistance. CKD raised mRNAs of inflammatory cytokines in muscles and there was a 5.2-fold increase in signal regulatory protein-α (SIRP-α), a transmembrane glycoprotein principally present in muscle membranes. By immunoprecipitation we found it interacts with the insulin receptor and insulin receptor substrate-1 (IRS-1). Treatment of myotubes with a mixture of inflammatory cytokines showed that SIRP-α expression was increased by a NF-κB-dependent pathway. Blockade of NF-κB using a small-molecule chemical inhibitor or a dominant-negative IKKβ reduced cytokine-induced SIRP-α expression. The overexpression of SIRP-α in myotubes impaired insulin signaling and raised proteolysis while SIRP-α knockdown with siRNAs in skeletal muscle cells increased tyrosine phosphorylation of the insulin receptor and IRS-1 despite inclusion of cytokines. This led to increased p-Akt and suppression of protein degradation. Thus, SIRP-α is part of a novel mechanism for inflammation-mediated insulin resistance in muscle. In catabolic conditions with impaired insulin signaling, targeting SIRP-α may improve insulin sensitivity and prevent muscle atrophy.Kidney International advance online publication, 20 March 2013; doi:10.1038/ki.2013.97.
    Kidney International 03/2013; · 8.52 Impact Factor
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    ABSTRACT: Glutathione transferase isozyme A4 (GSTA4) exhibits high catalytic efficiency to metabolize 4-hydroxynonenal (4-HNE), a highly reactive lipid peroxidation product that has been implicated in the pathogenesis of various chronic diseases. We investigated the role of 4-HNE in the mechanisms of unilateral ureteral obstruction (UUO)-induced fibrosis and its modulation by GSTA4-4 in a mouse model. Our data indicate that after UUO, accumulation of 4-HNE and its adducts were increased in renal tissues, with a concomitant decrease in the expression of GSTA4-4 in mice. As compared to wild-type (WT) mice, UUO caused an increased expression of fibroblast markers in the interstitium of GSTA4 KO mice. Additionally, increased autophagy and tubular cell damage were more severe in UUO-treated GSTA4 KO mice than in WT mice. Furthermore, GSK-3β phosphorylation and expression of Snail, a regulator of E-cadherin and Occludin, was found to be significantly higher in UUO-inflicted GSTA4 KO mice. GSTA4 over-expression prevented 4-HNE-induced autophagy activation, tubular cell damage and Snail nuclear translocation in vitro. The effects of long-term expression of GSTA4 in restoration of UUO-induced damage in mice with the GSTA4 inducible transposon system indicated that release of obstruction after 3 days of UUO resulted in the attenuation of interstitial SMAα and collagen I expression. This transposon-delivered GSTA4 expression also suppressed UUO-induced loss of tubular cell junction markers and autophagy activation. Together, these results indicate that 4-HNE significantly contributes to the mechanisms of tubule injury and fibrosis and that these effects can be inhibited by the enhanced expression of GSTA4-4. Copyright © 2012 Pathological Society of Great Britain and Ireland. Published by John Wiley & Sons, Ltd.
    The Journal of Pathology 06/2012; · 7.59 Impact Factor
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    ABSTRACT: Chronic kidney disease (CKD) is increasingly common, and there is an increasing awareness that every strategy should be used to avoid complications of CKD. Restriction of dietary protein intake has been a relevant part of the management of CKD for more than 100 years, but even today, the principal goal of protein-restricted regimens is to decrease the accumulation of nitrogen waste products, hydrogen ions, phosphates, and inorganic ions while maintaining an adequate nutritional status to avoid secondary problems such as metabolic acidosis, bone disease, and insulin resistance, as well as proteinuria and deterioration of renal function. This supplement focuses on recent experimental and clinical findings related to an optimized dietary management of predialysis, dialysis, and transplanted patients as an important aspect of patient care. Nutritional treatment strategies are linked toward ameliorating metabolic and endocrine disturbances, improving/maintaining nutritional status, as well as delaying the renal replacement initiation and improving outcomes in CKD patients. A final consensus states that dietary manipulations should be considered as one of the main approaches in the management program of CKD patients and that a reasonable number of patients with moderate or severe CKD benefit from dietary protein/phosphorus restriction.
    Journal of Renal Nutrition 03/2012; 22(2 Suppl):S1-21. · 1.75 Impact Factor
  • Journal of Renal Nutrition 03/2012; 22(2 Suppl):S22-4. · 1.75 Impact Factor
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    H Q Han, William E Mitch
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    ABSTRACT: To understand the mechanisms of muscle wasting and how inhibiting myostatin signaling affects them. Myostatin signaling is critical for the understanding of the pathogenesis of muscle wasting as blocking signaling mitigates muscle losses in rodent models of catabolic diseases including cancer, chronic kidney, or heart failure. Muscle wasting increases the risks of morbidity and mortality. But, the reliability of estimates of the degree of muscle wasting is controversial as are definitions of terms like cachexia. Much information has been learnt about the pathophysiology of muscle wasting, including the major role of the ubiquitin-proteasome system (UPS) which along with other proteases degrades protein and limits protein synthesis. In contrast, few successful strategies for reversing muscle loss have been tested. Several catabolic conditions are characterized by inflammation, increased glucocorticoid production, and impaired intracellular signaling in response to insulin and IGF-1. These characteristics lead to activation of the UPS and other proteases producing muscle wasting. Another potential initiator of muscle wasting is myostatin and its expression is increased in muscles of animal models and patients with certain catabolic conditions. Myostatin is a member of the TGF-β family; it suppresses muscle growth and its absence stimulates muscle growth substantially. Recently, pharmacologic suppression of myostatin was found to counteract inflammation, increased glucocorticoids and impaired insulin/IGF-1 signaling and most importantly, prevents muscle wasting in rodent models of cancer and kidney failure. Myostatin antagonism as a therapy for patients with muscle wasting should become a topic of clinical investigation.
    Current opinion in supportive and palliative care 12/2011; 5(4):334-41.
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    ABSTRACT: Nitric oxide (NO) is crucial in diverse physiological and pathological processes. We show that a hypomorphic mouse model of argininosuccinate lyase (encoded by Asl) deficiency has a distinct phenotype of multiorgan dysfunction and NO deficiency. Loss of Asl in both humans and mice leads to reduced NO synthesis, owing to both decreased endogenous arginine synthesis and an impaired ability to use extracellular arginine for NO production. Administration of nitrite, which can be converted into NO in vivo, rescued the manifestations of NO deficiency in hypomorphic Asl mice, and a nitric oxide synthase (NOS)-independent NO donor restored NO-dependent vascular reactivity in humans with ASL deficiency. Mechanistic studies showed that ASL has a structural function in addition to its catalytic activity, by which it contributes to the formation of a multiprotein complex required for NO production. Our data demonstrate a previously unappreciated role for ASL in NOS function and NO homeostasis. Hence, ASL may serve as a target for manipulating NO production in experimental models, as well as for the treatment of NO-related diseases.
    Nature medicine 11/2011; 17(12):1619-26. · 27.14 Impact Factor
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    ABSTRACT: The mechanisms underlying the muscle wasting that accompanies CKD are not well understood. Animal models suggest that impaired differentiation of muscle progenitor cells may contribute. Expression of the myogenesis-suppressing transcription factor Ying Yang-1 increases in muscle of animals with CKD, but the mechanism underlying this increased expression is unknown. Here, we examined a profile of microRNAs in muscles from mice with CKD and observed downregulation of both microRNA-29a (miR-29a) and miR-29b. Because miR-29 has a complementary sequence to the 3'-untranslated region of Ying Yang-1 mRNA, a decrease in miR-29 could increase Ying Yang-1. We used adenovirus-mediated gene transfer to express miR-29 in C2C12 myoblasts and measured its effect on both Ying Yang-1 and myoblast differentiation. An increase in miR-29 decreased the abundance of Ying Yang-1 and improved the differentiation of myoblasts into myotubes. Similarly, using myoblasts isolated from muscles of mice with CKD, an increase in miR-29 improved differentiation of muscle progenitor cells into myotubes. In conclusion, CKD suppresses miR-29 in muscle, which leads to higher expression of the transcription factor Ying Yang-1, thereby suppressing myogenesis. These data suggest a potential mechanism for the impaired muscle cell differentiation associated with CKD.
    Journal of the American Society of Nephrology 09/2011; 22(11):2068-76. · 8.99 Impact Factor
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    ABSTRACT: Congestive heart failure and chronic kidney disease are characterized by chronically elevated angiotensin II (Ang II) and muscle wasting. Ang II causes skeletal muscle wasting by reducing appetite and by enhancing catabolism. The serine/threonine kinase AMP-activated protein kinase (AMPK) functions mainly as a sensor of cellular energy status. It is energy sparing and favors ATP generation. We hypothesized that Ang II induces muscle wasting in part by inhibiting AMPK signaling and altering cellular energy balance. Our results show that Ang II infusion in mice reduced gastrocnemius muscle weight by 26% and depleted ATP by 74%. In addition, Ang II upregulated protein phosphatase 2Cα by 2.6-fold and reduced AMPK phosphorylation and signaling in muscle. Importantly, the pharmacological AMPK activator 5-aminoimidazole-4-carboxamide ribonucleoside restored AMPK activity to levels of pair-fed controls and reversed Ang II-mediated ATP depletion and muscle wasting. Moreover, 5-aminoimidazole-4-carboxamide ribonucleoside activated Akt and inhibited Ang II-induced increases in E3 ubiquitin ligase expression. These novel results demonstrate critical roles for energy depletion and AMPK inhibition in Ang II-induced skeletal muscle wasting and suggest a therapeutic potential for AMPK activators in diseases characterized by muscle wasting.
    Hypertension 08/2011; 58(4):643-9. · 6.87 Impact Factor
  • Stewart H Lecker, William E Mitch
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    ABSTRACT: Proteins in all cells turnover continuously such that rigorous control of proteolysis is required to govern levels of proteins with vastly different half-lives and actions including those regulating transcription, metabolic pathways, or the breakdown of muscle proteins to amino acids used in gluconeogenesis or the synthesis of new proteins. Critical cellular functions would be disrupted without precise regulation of protein degradation. Thus, it is surprising that the bulk of protein in all cells is degraded by the ATP-dependent, ubiquitin-proteasome system. The system achieves remarkable specificity by selective conjugation of ubiquitin (Ub) to a doomed protein in a process catalyzed by >1000 ubiquitin ligases that recognize individual substrate proteins. Because the pathogenesis of certain kidney diseases and their complications are linked to the function of the ubiquitin-proteasome system, understanding its mechanisms could lead to novel therapies.
    Journal of the American Society of Nephrology 05/2011; 22(5):821-4. · 8.99 Impact Factor
  • Yanlin Wang, William E Mitch
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    ABSTRACT: The mechanisms underlying beneficial influences of low-protein diets on the progressive nature of chronic kidney disease are incompletely understood. Gao et al. identify a downregulation of Kruppel-like factor-15 (KLF15) associated with increased renal fibrosis. The KLF15 downregulation and renal fibrosis are reversed with dietary protein restriction, especially when ketoacids are added. The results suggest that KLF15 may play a role in suppressing renal fibrosis and could contribute to the benefits of dietary protein restriction.
    Kidney International 05/2011; 79(9):933-4. · 8.52 Impact Factor

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