Cytokine transcriptome profiling in acute experimental canine atopic dermatitis skin lesions after IL-31 inhibition with lokivetmab.

BACKGROUND: The caninised monoclonal antibody lokivetmab (LKV), directed at interleukin (IL)-31, is very effective at controlling pruritus in most dogs with atopic dermatitis (AD). However, evidence exists that IL-31 is not required for the induction of acute allergic skin inflammation, which might explain why this treatment is less efficacious in some dogs with AD. HYPOTHESIS/OBJECTIVES: To compare the comprehensive transcriptome analysis of house dust mite (HDM)-sensitised dogs with and without treatment with LKV to attest our hypothesis that LKV does not majorly affect acute cytokine/chemokine production. ANIMALS: Six HDM-sensitised atopic Maltese-beagle dogs. MATERIALS AND METHODS: In this cross-over study, the cytokine profiling of acute AD skin lesions was compared by RNA sequencing (RNA-Seq), with or without LKV-induced inhibition of IL-31. Skin biopsies were obtained from each dog at 0, 6, 12, 24, 48, and 96 h after epicutaneous HDM allergen provocation. RESULTS: Macroscopic and microscopic skin lesion scores were not significantly different between the LKV- and nontreatment groups at any time points. Likewise, the results of RNA-Seq analysis revealed no significant difference in the messenger (m)RNA expression of the major cytokines between these two groups. In LKV-treated dogs, IL6, IL9, IL13, IL33, CCL17, and CCL22 were significantly upregulated compared to their baseline expression levels, suggesting that these cytokines are unaffected by IL-31 inhibition. CONCLUSIONS AND CLINICAL RELEVANCE: IL-31 inhibition is insufficient to prevent the expression of other proinflammatory mediators in acute AD and these could be considered as other potential therapeutic targets.

Ex vivo activated CD4+ T cells from young calves exhibit Th2-biased effector function with distinct metabolic reprogramming compared to adult cows.

As maternal passive immunity wanes at 6-8 weeks, young calves must rely on their own naïve and developing immune system for protection against pathogens. Typically, an infection in the young induces a T cell-mediated response, which skews towards a Th2 phenotype and results in a reduced effector response. Our study examines the implications this transitional period of immunocompetency has on cellular metabolism in young calves, focusing on effector function of CD4+ T cells in comparison to those from adult cows. Results from sorted CD4+ T cells from young calves and adult cows activated by α-CD3:α-CD28, show that young calves exhibit a significantly greater propensity to produce the Th2 cytokine, IL-4, in comparison to IFN-γ. Concomitantly, cells from young calves and adult cows exhibit no statistical difference in cell surface marker expression induced by α-CD3:α-CD28 stimulation. Metabolically, activated CD4+ T cells from young calves show significantly greater utilization of mitochondrial respiration, measured by oxygen consumption rate (OCR), and greater glycolytic reserve, measured by extracellular acidification rate (ECAR). However, adult cows have a significantly higher change in glycolytic rate after α-CD3:α-CD28 stimulation compared to young calves. Further, CD4+ T cells from young calves have an increased mRNA expression signature associated with glycolytic metabolism (GAPDH, HK2, FBP1, HIF1A) and Th2-associated metabolic signaling (RPTOR) in comparison to adult cows. The distinct metabolic phenotype and associated gene expression in activated CD4+ T cells may be intrinsic drivers of the Th2-biased response by young calves. Additionally, CD4+ recent thymic emigrant cells (RTEs) may further contribute to altered effector function, as they are preferential precursors to Tregs, and based on the microenvironment, have the propensity to polarize toward Th2. Evaluation of T cell master transcription regulators, as well as measuring signal joint T cell receptor excision circles between young calves and adult cows, we observed a significantly increased proportion of RTEs from sorted CD4+ T cells. In this study, we show a unique metabolic profile exhibited by activated CD4+ T cells from young calves in which mitochondrial respiration and glycolytic capacity is significantly increased compared to adult cows.

Lactation stage impacts the glycolytic function of bovine CD4+ T cells during ex vivo activation.

Dairy cattle undergo dynamic physiological changes over the course of a full lactation into the dry period, which impacts their immunocompetence. During activation, T cells undergo a characteristic rewiring to increase the uptake of glucose and metabolically reprogram to favor aerobic glycolysis over oxidative phosphorylation. To date it remains to be completely elucidated how the altered energetic demands associated with lactation in dairy cows impacts T cell metabolic reprogramming. Thus, in our ex vivo studies we have examined the influence of stage of lactation (early lactation into the dry period) on cellular metabolism in activated bovine CD4+ T cells. Results showed higher rates of glycolytic function in activated CD4+ T cells from late lactation and dry cows compared to cells from early and mid-lactation cows. Similarly, protein and mRNA expression of cytokines were higher in CD4+ T cells from dry cows than CD4+ T cells from lactating cows. The data suggest CD4+ T cells from lactating cows have an altered metabolic responsiveness that could impact the immunocompetence of these animals, particularly those in early lactation, and increase their susceptibility to infection.