Docosahexaenoic acid enhances amphiregulin-mediated bronchial epithelial cell repair processes following organic dust exposure.

Injurious dust exposures in the agricultural workplace involve the release of inflammatory mediators and activation of epidermal growth factor receptor (EGFR) in the respiratory epithelium. Amphiregulin (AREG), an EGFR ligand, mediates tissue repair and wound healing in the lung epithelium. Omega-3 fatty acids such as docosahexaenoic acid (DHA) are also known modulators of repair and resolution of inflammatory injury. This study investigated how AREG, DHA, and EGFR modulate lung repair processes following dust-induced injury. Primary human bronchial epithelial (BEC) and BEAS-2B cells were treated with an aqueous extract of swine confinement facility dust (DE) in the presence of DHA and AREG or EGFR inhibitors. Mice were exposed to DE intranasally with or without EGFR inhibition and DHA. Using a decellularized lung scaffolding tissue repair model, BEC recolonization of human lung scaffolds was analyzed in the context of DE, DHA, and AREG treatments. Through these investigations, we identified an important role for AREG in mediating BEC repair processes. DE-induced AREG release from BEC, and DHA treatment following DE exposure, enhanced this release. Both DHA and AREG also enhanced BEC repair capacities and rescued DE-induced recellularization deficits. In vivo, DHA treatment enhanced AREG production following DE exposure, whereas EGFR inhibitor-treated mice exhibited reduced AREG in their lung homogenates. These data indicate a role for AREG in the process of tissue repair after inflammatory lung injury caused by environmental dust exposure and implicate a role for DHA in regulating AREG-mediated repair signaling in BEC.

ß2-Adrenergic agonists attenuate organic dust-induced lung inflammation.

Agricultural dust exposure results in significant lung inflammation, and individuals working in concentrated animal feeding operations (CAFOs) are at risk for chronic airway inflammatory diseases. Exposure of bronchial epithelial cells to aqueous extracts of hog CAFO dusts (HDE) leads to inflammatory cytokine production that is driven by protein kinase C (PKC) activation. cAMP-dependent protein kinase (PKA)-activating agents can inhibit PKC activation in epithelial cells, leading to reduced inflammatory cytokine production following HDE exposure. ß2-Adrenergic receptor agonists (ß2-agonists) activate PKA, and we hypothesized that ß2-agonists would beneficially impact HDE-induced adverse airway inflammatory consequences. Bronchial epithelial cells were cultured with the short-acting ß2-agonist salbutamol or the long-acting ß2-agonist salmeterol prior to stimulation with HDE. ß2-Agonist treatment significantly increased PKA activation and significantly decreased HDE-stimulated IL-6 and IL-8 production in a concentration- and time-dependent manner. Salbutamol treatment significantly reduced HDE-induced intracellular adhesion molecule-1 expression and neutrophil adhesion to epithelial cells. Using an established intranasal inhalation exposure model, we found that salbutamol pretreatment reduced airway neutrophil influx and IL-6, TNF-α, CXCL1, and CXCL2 release in bronchoalveolar lavage fluid following a one-time exposure to HDE. Likewise, when mice were pretreated daily with salbutamol prior to HDE exposure for 3 wk, HDE-induced neutrophil influx and inflammatory mediator production were also reduced. The severity of HDE-induced lung pathology in mice repetitively exposed to HDE for 3 wk was also decreased with daily salbutamol pretreatment. Together, these results support the need for future clinical investigations to evaluate the utility of ß2-agonist therapies in the treatment of airway inflammation associated with CAFO dust exposure.

Pharmacological inhibition of soluble epoxide hydrolase prevents renal interstitial fibrogenesis in obstructive nephropathy.

Treating chronic kidney disease (CKD) has been challenging because of its pathogenic complexity. Epoxyeicosatrienoic acids (EETs) are cytochrome P-450-dependent derivatives of arachidonic acid with antihypertensive, anti-inflammatory, and profibrinolytic functions. We recently reported that genetic ablation of soluble epoxide hydrolase (sEH), an enzyme that converts EETs to less active dihydroxyeicosatrienoic acids, prevents renal tubulointerstitial fibrosis and inflammation in experimental mouse models of CKD. Here, we tested the hypothesis that pharmacological inhibition of sEH after unilateral ureteral obstruction (UUO) would attenuate tubulointerstitial fibrosis and inflammation in mouse kidneys and may provide a novel approach to manage the progression of CKD. Inhibition of sEH enhanced levels of EET regioisomers and abolished tubulointerstitial fibrosis, as demonstrated by reduced collagen deposition and myofibroblast formation after UUO. The inflammatory response was also attenuated, as demonstrated by decreased influx of neutrophils and macrophages and decreased expression of inflammatory cytokines keratinocyte chemoattractant, macrophage inflammatory protein-2, monocyte chemotactic protein-1, TNF-α, and ICAM-1 in kidneys after UUO. UUO upregulated transforming growth factor-ß1/Smad3 signaling and induced NF-κB activation, oxidative stress, tubular injury, and apoptosis; in contrast, it downregulated antifibrotic factors, including peroxisome proliferator-activated receptor (PPAR) isoforms, especially PPAR-γ. sEH inhibition mitigated the aforementioned malevolent effects in UUO kidneys. These data demonstrate that pharmacological inhibition of sEH promotes anti-inflammatory and fibroprotective effects in UUO kidneys by preventing tubular injury, downregulation of NF-κB, transforming growth factor-ß1/Smad3, and inflammatory signaling pathways, and activation of PPAR isoforms. Our data suggest the potential use of sEH inhibitors in treating fibrogenesis in the UUO model of CKD.

Replicated, replicable and relevant-target engagement and pharmacological experimentation in the 21st century.

A pharmacological experiment is typically conducted to: i) test or expand a hypothesis regarding the potential role of a target in the mechanism(s) underlying a disease state using an existing drug or tool compound in normal and/or diseased tissue or animals; or ii) characterize and optimize a new chemical entity (NCE) targeted to modulate a specific disease-associated target to restore homeostasis as a potential drug candidate. Hypothesis testing necessitates an intellectually rigorous, null hypothesis approach that is distinct from a high throughput fishing expedition in search of a hypothesis. In conducting an experiment, the protocol should be transparently defined along with its powering, design, appropriate statistical analysis and consideration of the anticipated outcome (s) before it is initiated. Compound-target interactions often involve the direct study of phenotype(s) unique to the target at the cell, tissue or animal/human level. However, in vivo studies are often compromised by a lack of sufficient information on the compound pharmacokinetics necessary to ensure target engagement and also by the context-free analysis of ubiquitous cellular signaling pathways downstream from the target. The use of single tool compounds/drugs at one concentration in engineered cell lines frequently results in reductionistic data that have no physiologically relevance. This overview, focused on trends in the peer-reviewed literature, discusses the execution and reporting of experiments and the criteria recommended for the physiologically-relevant assessment of target engagement to identify viable new drug targets and facilitate the advancement of translational studies.