Local lung epithelial IL-6TS activation in the absence of type 2 airway inflammation defines a novel subset of asthmatic patients and might drive airway inflammation and epithelial dysfunction in these patients.

Background: Group 2 innate lymphoid cells (ILC2s) are effective producers of IL-5 and IL-13 during allergic inflammation and bridge the innate and adaptive immune responses. ILC2 numbers are increased in asthmatic patients compared with healthy control subjects. Thus far, human data describing their phenotype during acute allergic inflammation in the lung are incomplete. Objectives: This study aims to characterize and compare bloodand lung-derived ILC2s before and after segmental allergen challenge in patients with mild-to-moderate asthma with high blood eosinophil counts (> _300 cells/mL). Methods: ILC2s were isolated from blood and bronchoalveolar lavage (BAL) fluid before and after segmental allergen challenge. Cells were sorted by means of flow cytometry, cultured and analyzed for cytokine release or migration, and sequenced for RNA expression. Results: ILC2s were nearly absent in the alveolar space under baseline conditions, but numbers increased significantly after allergen challenge (P < .05), whereas at the same time, ILC2 numbers in blood were reduced (P < .05). Prostaglandin D 2 and CXCL12 levels in BAL fluid correlated with decreased ILC2 numbers in blood (P 5 .004, respective P 5 .024). After allergen challenge, several genes promoting type 2 inflammation were expressed at greater levels in BAL fluid compared with blood ILC2s, whereas blood ILC2s remain unactivated. Conclusion: ILC2s accumulate at the site of allergic inflammation and are recruited from the blood. Their transcriptional and functional activation pattern promotes type 2 inflammation. (J Allergy Clin Immunol 2019;144:61-9.)

Elevated levels of glucose and lipids are characteristics of individuals with type 2 diabetes mellitus (T2DM). The enhanced nutrient levels have been connected with deterioration of b-cell function and impaired insulin secretion observed in these individuals. A strategy to improve b-cell function in individuals with T2DM has been intermittent administration of K ATP channel openers. After such treatment, both the magnitude and kinetics of insulin secretion are markedly improved. In an attempt to further delineate mechanisms of how openers of K ATP channels improve b-cell function, the effects of diazoxide on markers of endoplasmic reticulum (ER) stress was determined in b-cells exposed to the fatty acid palmitate. The eukaryotic translation factor 2-alpha kinase 3 (EIF2AK3; also known as PERK) and endoplasmic reticulum to nucleus signaling 1 (ERN1; also known as IRE1) pathways, but not the activating transcription factor (ATF6) pathway of the unfolded protein response, are activated in such lipotoxic b-cells. Inclusion of diazoxide during culture attenuated activation of the EIF2AK3 pathway but not the ERN1 pathway. This attenuation was associated with reduced levels of DNA-damage inducible transcript 3 (DDIT3; also known as CHOP) and b-cell apoptosis was decreased. It is concluded that reduction of ER stress may be a mechanism by which diazoxide improves b-cell function.

Palmitate negatively affects insulin secretion and apoptosis in the pancreatic β-cell. The detrimental effects are abolished by elongating and desaturating the fatty acid into oleate. To investigate mechanisms of how the two fatty acids differently affect β-cell function and apoptosis, lipid handling was determined in MIN6 cells cultured in the presence of the fatty acids palmitate (16:0) and oleate (18:1) and also corresponding monounsaturated fatty acid palmitoleate (16:1) and saturated fatty acid stearate (18:0). Insulin secretion was impaired and apoptosis accentuated in palmitate-, and to some extent, stearate-treated cells. Small or no changes in secretion or apoptosis were observed in cells exposed to palmitoleate or oleate. Expressions of genes associated with fatty acid esterification (SCD1, DGAT1, DGAT2, and FAS) were augmented in cells exposed to palmitate or stearate but only partially (DGAT2) in palmitoleate- or oleate-treated cells. Nevertheless, levels of triglycerides were highest in cells exposed to oleate. Similarly, fatty acid oxidation was most pronounced in oleate-treated cells despite comparable up-regulation of CPT1 after treatment of cells with the four different fatty acids. The difference in apoptosis between palmitate and stearate was paralleled by similar differences in levels of markers of endoplasmic reticulum (ER) stress in cells exposed to the two fatty acids. Palmitate-induced ER stress was not accounted for by ceramide de novo synthesis. In conclusion, although palmitate initiated stronger expression changes consistent with lipid accumulation and combustion in MIN6 cells, rise in triglyceride levels and fatty acid oxidation was favored specifically in cells exposed to oleate.

ObjectivesGenetic variations in TNFAIP3 (A20) de-ubiquitinase (DUB) domain increase the risk of systemic lupus erythematosus (SLE) and rheumatoid arthritis. A20 is a negative regulator of NF-κB but the role of its DUB domain and related genetic variants remain unclear. We aimed to study the functional effects of A20 DUB-domain alterations in immune cells and understand its link to SLE pathogenesis.MethodsCRISPR/Cas9 was used to generate human U937 monocytes with A20 DUB-inactivating C103A knock-in (KI) mutation. Whole genome RNA-sequencing was used to identify differentially expressed genes between WT and C103A KI cells. Functional studies were performed in A20 C103A U937 cells and in immune cells from A20 C103A mice and genotyped healthy individuals with A20 DUB polymorphism rs2230926. Neutrophil extracellular trap (NET) formation was addressed ex vivo in neutrophils from A20 C103A mice and SLE-patients with rs2230926.ResultsGenetic disruption of A20 DUB domain in human and murine myeloid cells did not give rise to enhanced NF-κB signalling. Instead, cells with C103A mutation or rs2230926 polymorphism presented an upregulated expression of PADI4, an enzyme regulating protein citrullination and NET formation, two key mechanisms in autoimmune pathology. A20 C103A cells exhibited enhanced protein citrullination and extracellular trap formation, which could be suppressed by selective PAD4 inhibition. Moreover, SLE-patients with rs2230926 showed increased NETs and increased frequency of autoantibodies to citrullinated epitopes.ConclusionsWe propose that genetic alterations disrupting the A20 DUB domain mediate increased susceptibility to SLE through the upregulation of PADI4 with resultant protein citrullination and extracellular trap formation.