Redox Control Of Allergic Airway Disease: Impact Of Glutaredoxin-1 On Epithelial Driven Inflammation And Allergen-Induced Airway Remodeling

Asthma is a multi-faceted chronic inflammatory disease accompanied by loss of airway epithelial integrity leading to remodeling of the airways. Perturbations to the lung redox environment, including alterations in glutathione (GSH) content, have been reported in asthma. GSH can be conjugated to protein cysteines, controlling protein function in an oxidant-dependent process known as protein S-glutathionylation (PSSG). The thioltransferase, glutaredoxin-1 (Glrx1), deglutathionylates proteins under physiological conditions, restoring sulfhydryl groups of target proteins. Glrx1 is emerging as a critical player in settings of allergic airway disease, but its function in regulating epithelial cell responses to asthma-relevant cytokines has not been examined. Furthermore, the role of Glrx1 in controlling the extent of airway remodeling in response to house dust mite (HDM) in vivo is still not well understood. Interleukin-17A (IL-17A) is a potent cytokine that stimulates epithelial cells to produce pro-inflammatory mediators, in part by activating the nuclear factor kappaB (NF-κB) pathway, a key regulator of inflammation. We demonstrate that interleukin-17A (IL-17A) induces rapid activation of both classical and alternative NF-κB, while simultaneously resulting in protein oxidation and PSSG. In particular, we show IL 17A induces S-glutathionylation of RelA (RelA-SSG) and IKKα (IKKα-SSG), which is enhanced following siRNA-mediated knockdown of Glrx1. We also demonstrate that absence of Glrx1 leads to increased nuclear content of RelA and RelB and enhanced production of NF-κB-driven pro-inflammatory genes, KC and CCL20 while decreasing IL-6 expression. Finally, we show that siRNA-mediated knockdown of IKKα attenuates nuclear RelA and RelB and dampens pro-inflammatory gene production. Together, these data indicate a crucial role for the Glrx1/PSSG axis in controlling RelA-SSG, IKKα-SSG and epithelial cell responsiveness to IL-17A. Mice lacking Glrx1 were previously shown to display enhanced resolution of allergic airway disease induced by ovalbumin (Ova) challenge. In this study, we determined the role of Glrx1 in a HDM model of allergic airway disease. Wild type (WT) mice and Glrx1 deficient (Glrx1-/-) mice demonstrated similar total lung cell counts, but Glrx1-/- mice displayed fewer neutrophils than WT mice. Conversely, mice overexpressing Glrx1 specifically in CCSP positive cells in the lung (Epi-Glrx1) showed attenuated total lung cell counts and lung eosinophils compared to control mice. Immunohistological analysis of remodeling markers revealed that Glrx1-/- mice displayed increased HDM-induced mucus metaplasia, α smooth muscle actin (αSMA) positivity and collagen staining compared to WT mice. Evaluation of total lung collagen showed that Glrx1-/- mice had significantly higher collagen content compared to WT mice. In Epi-Glrx1 mice, attenuation of mucus metaplasia, αSMA content and collagen staining was observed compared to control mice. Furthermore, Epi-Glrx1 mice also demonstrated significantly impaired collagen production compared to control mice. We also demonstrate that Glrx1 absence results in decreased expression of the epithelial cell marker, E-cadherin, and increased expression of αSMA, a mesenchymal marker. Together, these studies demonstrate a critical role for Glrx1 in controlling epithelial cell responses to IL-17A and in mediating in vivo collagen production in response to chronic allergen exposure

Extracorporeal Treatment in Phenytoin Poisoning: Systematic Review and Recommendations from the EXTRIP Workgroup

The Extracorporeal Treatments in Poisoning (EXTRIP) Workgroup conducted a systematic literature review using a standardized process to develop evidence-based recommendations on the use of extracorporeal treatment (ECTR) in patients with phenytoin poisoning. The authors reviewed all articles, extracted data, summarized findings, and proposed structured voting statements following a predetermined format. A 2-round modified Delphi method was used to reach a consensus on voting statements, and the RAND/UCLA Appropriateness Method was used to quantify disagreement. 51 articles met the inclusion criteria. Only case reports, case series, and pharmacokinetic studies were identified, yielding a very low quality of evidence. Clinical data from 31 patients and toxicokinetic grading from 46 patients were abstracted. The workgroup concluded that phenytoin is moderately dialyzable (level of evidence = C) despite its high protein binding and made the following recommendations. ECTR would be reasonable in select cases of severe phenytoin poisoning (neutral recommendation, 3D). ECTR is suggested if prolonged coma is present or expected (graded 2D) and it would be reasonable if prolonged incapacitating ataxia is present or expected (graded 3D). If ECTR is used, it should be discontinued when clinical improvement is apparent (graded 1D). The preferred ECTR modality in phenytoin poisoning is intermittent hemodialysis (graded 1D), but hemoperfusion is an acceptable alternative if hemodialysis is not available (graded 1D). In summary, phenytoin appears to be amenable to extracorporeal removal. However, because of the low incidence of irreversible tissue injury or death related to phenytoin poisoning and the relatively limited effect of ECTR on phenytoin removal, the workgroup proposed the use of ECTR only in very select patients with severe phenytoin poisoning

Exercise-induced alterations in phospholipid hydrolysis, airway surfactant, and eicosanoids and their role in airway hyperresponsiveness in asthma

The mechanisms responsible for driving endogenous airway hyperresponsiveness (AHR) in the form of exercise-induced bronchoconstriction (EIB) are not fully understood. We examined alterations in airway phospholipid hydrolysis, surfactant degradation, and lipid mediator release in relation to AHR severity and changes induced by exercise challenge. Paired induced sputum

Attenuation of lung fibrosis in mice with a clinically relevant inhibitor of glutathione-S-transferase π

Idiopathic pulmonary fibrosis (IPF) is a debilitating lung disease characterized by excessive collagen production and fibrogenesis. Apoptosis in lung epithelial cells is critical in IPF pathogenesis, as heightened loss of these cells promotes fibroblast activation and remodeling. Changes in glutathione redox status have been reported in IPF patients. S-glutathionylation, the conjugation of glutathione to reactive cysteines, is catalyzed in part by glutathione-S-transferase π (GSTP). To date, no published information exists linking GSTP and IPF to our knowledge. We hypothesized that GSTP mediates lung fibrogenesis in part through FAS S-glutathionylation, a critical event in epithelial cell apoptosis. Our results demonstrate that GSTP immunoreactivity is increased in the lungs of IPF patients, notably within type II epithelial cells. The FAS-GSTP interaction was also increased in IPF lungs. Bleomycin- and AdTGFβ-induced increases in collagen content, α-SMA, FAS S-glutathionylation, and total protein S-glutathionylation were strongly attenuated in Gstp(–/–) mice. Oropharyngeal administration of the GSTP inhibitor, TLK117, at a time when fibrosis was already apparent, attenuated bleomycin- and AdTGFβ-induced remodeling, α-SMA, caspase activation, FAS S-glutathionylation, and total protein S-glutathionylation. GSTP is an important driver of protein S-glutathionylation and lung fibrosis, and GSTP inhibition via the airways may be a novel therapeutic strategy for the treatment of IPF

Chronic kidney disease and arrhythmias: conclusions from a Kidney Disease: Improving Global Outcomes (KDIGO) Controversies Conference.

Patients with chronic kidney disease (CKD) are predisposed to heart rhythm disorders, including atrial fibrillation (AF)/atrial flutter, supraventricular tachycardias, ventricular arrhythmias, and sudden cardiac death (SCD). While treatment options, including drug, device, and procedural therapies, are available, their use in the setting of CKD is complex and limited. Patients with CKD and end-stage kidney disease (ESKD) have historically been under-represented or excluded from randomized trials of arrhythmia treatment strategies,1 although this situation is changing.2 Cardiovascular society consensus documents have recently identified evidence gaps for treating patients with CKD and heart rhythm disorders [...

Follistatin-like 1 regulates renal IL-1β expression in cisplatin nephrotoxicity

Follistatin-like 1 (FSTL1) is a secreted protein with homology to both Follistatin and the SPARC/BM40 family of matricellular proteins. In this study, we sought to determine the expression patterns of Fstl1 and its cognate receptor Dip2a in the adult, and to assess the consequences of Fstl1 inactivation on development and homeostasis of the kidney. We find that FSTL1 circulates at high levels in both the human and the mouse and that it is also locally expressed in the loop of Henle in the kidney. To begin to understand the in vivo functions of Fstl1, we generated a mouse mutant using a genetrap approach. The hypomorphic Fstl1 genetrap strain displays a strong reduction in FSTL1 expression at the protein level, but it does not show overt developmental defects. FSTL1 has previously been implicated in diverse disease processes as a regulator of inflammatory cytokine expression, and we therefore evaluated the response of the genetrap strain to cisplatin-mediated acute kidney injury, a disease model with highly cytokine-dependent pathology. We find that although TNF-α and Il6 levels are unchanged relative to wild-type, renal Il-1β expression is increased in genetrap mice following cisplatin treatment. Furthermore, histopatological analysis, expression of the tissue injury marker Havcr1, and measurement of serum creatinine demonstrate that reduction of Fstl1 expression sensitizes the kidney to acute cisplatin nephrotoxicity, suggesting a role for FSTL1-mediated Il-1β suppression in protection of the kidney from acute nephrotoxic injury

Recommendations for the role of extracorporeal treatments in the management of acute methanol poisoning: a systematic review and consensus statement

Objective: Methanol poisoning can induce death and disability. Treatment includes the administration of antidotes (ethanol or fomepizole and folic/folinic acid) and consideration of extracorporeal treatment for correction of acidemia and/or enhanced elimination. The Extracorporeal Treatments in Poisoning workgroup aimed to develop evidence-based consensus recommendations for extracorporeal treatment in methanol poisoning. Design and Methods: Utilizing predetermined methods, we conducted a systematic review of the literature. Two hundred seventy- Two relevant publications were identified but publication and selection biases were noted. Data on clinical outcomes and dialyzability were collated and a two-round modified Delphi process was used to reach a consensus. Results: Recommended indications for extracorporeal treatment: Severe methanol poisoning including any of the following being attributed to methanol: coma, seizures, new vision deficits, metabolic acidosis with blood pH ≤7.15, persistent metabolic acidosis despite adequate supportive measures and antidotes, serum anion gap higher than 24 mmol/L; or, serum methanol concentration 1) greater than 700 mg/L (21.8 mmol/L) in the context of fomepizole therapy, 2) greater than 600 mg/L or 18.7 mmol/L in the context of ethanol treatment, 3) greater than 500 mg/L or 15.6 mmol/L in the absence of an alcohol dehydrogenase blocker; in the absence of a methanol concentration, the osmolal/ osmolar gap may be informative; or, in the context of impaired kidney function. Intermittent hemodialysis is the modality of choice and continuous modalities are acceptable alternatives. Extracorporeal treatment can be terminated when the methanol concentration i

Glutathione S-transferase pi modulates NF-κB activation and pro-inflammatory responses in lung epithelial cells

Nuclear Factor kappa B (NF-κB) is a transcription factor family critical in the activation of pro- inflammatory responses. The NF-κB pathway is regulated by oxidant-induced post-translational modifications. Protein S-glutathionylation, or the conjugation of the antioxidant molecule, glutathione to reactive cysteines inhibits the activity of inhibitory kappa B kinase beta (IKKβ), among other NF-κB proteins. Glutathione S-transferase Pi (GSTP) is an enzyme that has been shown to catalyze protein S-glutathionylation (PSSG) under conditions of oxidative stress. The objective of the present study was to determine whether GSTP regulates NF-κB signaling, S-glutathionylation of IKK, and subsequent pro-inflammatory signaling. We demonstrated that, in unstimulated cells, GSTP associated with the inhibitor of NF-κB, IκBα. However, exposure to LPS resulted in a rapid loss of association between IκBα and GSTP, and instead led to a protracted association between IKKβ and GSTP. LPS exposure also led to increases in the S-glutathionylation of IKKβ. SiRNA-mediated knockdown of GSTP decreased IKKβ-SSG, and enhanced NF-κB nuclear translocation, transcriptional activity, and pro-inflammatory cytokine production in response to lipopolysaccharide (LPS). TLK117, an isotype-selective inhibitor of GSTP, also enhanced LPS-induced NF-κB transcriptional activity and pro-inflammatory cytokine production, suggesting that the catalytic activity of GSTP is important in repressing NF-κB activation. Expression of both wild-type and catalytically-inactive Y7F mutant GSTP significantly attenuated LPS- or IKKβ-induced production of GM-CSF. These studies indicate a complex role for GSTP in modulating NF-κB, which may involve S-glutathionylation of IKK proteins, and interaction with NF-κB family members. Our findings suggest that targeting GSTP is a potential avenue for regulating the activity of this prominent pro-inflammatory and immunomodulatory transcription factor