Metformin has anti-inflammatory effects and induces immunometabolic reprogramming via multiple mechanisms in hidradenitis suppurativa.

BACKGROUND: Targeting immunometabolism has shown promise in treating autoimmune and inflammatory conditions. Hidradenitis suppurativa (HS) is a chronic inflammatory skin disease involving painful lesions in apocrine gland-bearing skin. Therapeutic options for HS are limited and often ineffective; thus, there is a pressing need for improved treatments. To date, metabolic dysregulation has not been investigated in HS. As HS is highly inflammatory, we hypothesized that energy metabolism is dysregulated in these patients. Metformin, an antidiabetic drug, which is known to impact on cellular metabolic and signalling pathways, has been shown to have anti-inflammatory effects in cancer and arthritis. While metformin is not licensed for use in HS, patients with HS taking metformin show improved clinical symptoms. OBJECTIVE: To assess the effect and mechanism of action of metformin in HS. METHODS: To assess the effect of metformin in vivo, we compared the immune and metabolic profiles of peripheral blood mononuclear cells (PBMCs) of patients with HS taking metformin vs. those not taking metformin. To examine the effect of metformin treatment ex vivo, we employed a skin explant model on skin biopsies from patients with HS not taking metformin, which we cultured with metformin overnight. We used enzyme-linked immunosorbent assays, multiplex cytokine assays and quantitative real-time polymerase chain reaction (RT-PCR) to measure inflammatory markers, and Seahorse flux technology and quantitative RT-PCR to assess glucose metabolism. RESULTS: We showed that metabolic pathways are dysregulated in the PBMCs of patients with HS vs. healthy individuals. In metformin-treated patients, these metabolic pathways were restored and their PBMCs had reduced inflammatory markers following long-term metformin treatment. In the skin explant model, we found that overnight culture with metformin reduced inflammatory cytokines and chemokines and glycolytic genes in lesions and tracts of patients with HS. Using in vitro assays, we found that metformin may induce these changes via the NLR family pyrin domain containing 3 (NLRP3) inflammasome and the AMP-activated protein kinase (AMPK)-mammalian target of rapamycin (mTOR) pathway, which is linked to glycolysis and protein synthesis. CONCLUSIONS: Our study provides insight into the mechanisms of action of metformin in HS. The anti-inflammatory effects of metformin support its use as a therapeutic agent in HS, while its effects on immunometabolism suggest that targeting metabolism is a promising therapeutic option in inflammatory diseases, including HS.

Alterations in immune cell phenotype and cytotoxic capacity in HER2+ breast cancer patients receiving HER2-targeted neo-adjuvant therapy.

BACKGROUND: The phase II neo-adjuvant clinical trial ICORG10-05 (NCT01485926) compared chemotherapy in combination with trastuzumab, lapatinib or both in patients with HER2+ breast cancer. We studied circulating immune cells looking for alterations in phenotype, genotype and cytotoxic capacity (direct and antibody-dependent cell-mediated cytotoxicity (ADCC)) in the context of treatment response. METHODS: Peripheral blood mononuclear cells (PBMCs) were isolated from pre- (n = 41) and post- (n = 25) neo-adjuvant treatment blood samples. Direct/trastuzumab-ADCC cytotoxicity of patient-derived PBMCs against K562/SKBR3 cell lines was determined ex vivo. Pembrolizumab was interrogated in 21 pre-treatment PBMC ADCC assays. Thirty-nine pre-treatment and 21 post-treatment PBMC samples were immunophenotyped. Fc receptor genotype, tumour infiltrating lymphocyte (TIL) levels and oestrogen receptor (ER) status were quantified. RESULTS: Treatment attenuated the cytotoxicity/ADCC of PBMCs. CD3+/CD4+/CD8+ T cells increased following therapy, while CD56+ NK cells/CD14+ monocytes/CD19+ B cells decreased with significant post-treatment immune cell changes confined to patients with residual disease. Pembrolizumab-augmented ex vivo PBMC ADCC activity was associated with residual disease, but not pathological complete response. Pembrolizumab-responsive PBMCs were associated with lower baseline TIL levels and ER+ tumours. CONCLUSIONS: PBMCs display altered phenotype and function following completion of neo-adjuvant treatment. Anti-PD-1-responsive PBMCs in ex vivo ADCC assays may be a biomarker of treatment response.

Inflammation in the pathogenesis of depression: a disorder of neuroimmune origin.

There are several hypotheses concerning the underlying pathophysiological mechanisms of major depression, which centre largely around adaptive changes in neuronal transmission and plasticity, neurogenesis, and circuit and regional connectivity. The immune and endocrine systems are commonly implicated in driving these changes. An intricate interaction of stress hormones, innate immune cells and the actions of soluble mediators of immunity within the nervous system is described as being associated with the symptoms of depression. Bridging endocrine and immune processes to neurotransmission and signalling within key cortical and limbic brain circuits are critical to understanding depression as a disorder of neuroimmune origins. Emergent areas of research include a growing recognition of the adaptive immune system, advances in neuroimaging techniques and mechanistic insights gained from transgenic animals. Elucidation of glial-neuronal interactions is providing additional avenues into promising areas of research, the development of clinically relevant disease models and the discovery of novel therapies. This narrative review focuses on molecular and cellular mechanisms that are influenced by inflammation and stress. The aim of this review is to provide an overview of our current understanding of depression as a disorder of neuroimmune origin, focusing on neuroendocrine and neuroimmune dysregulation in depression pathophysiology. Advances in current understanding lie in pursuit of relevant biomarkers, as the potential of biomarker signatures to improve clinical outcomes is yet to be fully realised. Further investigations to expand biomarker panels including integration with neuroimaging, utilising individual symptoms to stratify patients into more homogenous subpopulations and targeting the immune system for new treatment approaches will help to address current unmet clinical need.

Targeting the NLRP3 inflammasome reduces inflammation in hidradenitis suppurativa skin.

BACKGROUND: Treatment for the debilitating disease hidradenitis suppurativa (HS) is inadequate in many patients. Despite an incidence of approximately 1%, HS is often under-recognized and underdiagnosed, and is associated with a high morbidity and poor quality of life. OBJECTIVES: To gain a better understanding of the pathogenesis of HS, in order to design new therapeutic strategies. METHODS: We employed single-cell RNA sequencing to analyse gene expression in immune cells isolated from involved HS skin vs. healthy skin. Flow cytometry was used to quantify the absolute numbers of the main immune populations. The secretion of inflammatory mediators from skin explant cultures was measured using multiplex and enzyme-linked immunosorbent assays. RESULTS: Single-cell RNA sequencing analysis identified a significant enrichment in the frequency of plasma cells, T helper (Th) 17 cells and dendritic cell subsets in HS skin, and the immune transcriptome was distinct and more heterogeneous than healthy skin. Flow cytometry revealed significantly increased numbers of T cells, B cells, neutrophils, dermal macrophages and dendritic cells in HS skin. Genes and pathways associated with Th17 cells, interleukin (IL)-17, IL-1ß and the NLRP3 inflammasome were enhanced in HS skin, particularly in samples with a high inflammatory load. Inflammasome constituent genes principally mapped to Langerhans cells and a subpopulation of dendritic cells. The secretome of HS skin explants contained significantly increased concentrations of inflammatory mediators, including IL-1ß and IL-17A, and culture with an NLRP3 inflammasome inhibitor significantly reduced the secretion of these, as well as other, key mediators of inflammation. CONCLUSIONS: These data provide a rationale for targeting the NLRP3 inflammasome in HS using small-molecule inhibitors that are currently being tested for other indications.

Metformin Treatment of Hidradenitis Suppurativa: Effect on Metabolic Parameters, Inflammation, Cardiovascular Risk Biomarkers, and Immune Mediators.

Hidradenitis suppurativa (HS) is a common cutaneous and systemic inflammatory disease with a significant impact on mental health and quality of life. It is associated with obesity, insulin resistance, metabolic syndrome, cardiovascular (CV) disease, and increased all-cause mortality. Metformin is used frequently in HS treatment and is effective for some patients. The mechanism of action of metformin in HS is unknown. A case-control study of 40 patients with HS (20 on metformin and 20 controls) was conducted to assess differences in metabolic markers, inflammation (C-reactive protein [CRP], serum adipokines, and CV risk biomarkers), and serum immune mediators. Body mass index (BMI), insulin resistance (77%), and metabolic syndrome (44%) were high overall, but not significantly different between the groups. This highlights the need for co-morbidity screening and management. A significant reduction in fasting insulin and a trend towards a reduction in insulin resistance were identified in the metformin group compared with pre-treatment levels. CV risk biomarkers were significantly favourable in the metformin group (lymphocytes, monocyte-lymphocyte ratio, neutrophil-lymphocyte ratio, and platelet-lymphocyte ratio). CRP was lower in the metformin group but was not statistically significant. Adipokines were dysregulated overall but were not different between the two groups. Serum IFN-γ, IL-8, TNF-α, and CXCL1 trended lower in the metformin group but did not reach significance. These results suggest that metformin improves CV risk biomarkers and insulin resistance in patients with HS. When the results of this study are considered alongside other studies in HS and related conditions, it is likely that metformin also has beneficial effects on metabolic markers and systemic inflammation in HS (CRP, serum adipokines, and immune mediators), warranting further research.

The mammalian target of rapamycin contributes to synovial fibroblast pathogenicity in rheumatoid arthritis.

Objectives: The mammalian target of Rapamycin (mTOR) is a metabolic master regulator of both innate and adaptive immunity; however, its exact role in stromal cell biology is unknown. In this study we explored the role of the mTOR pathway on Rheumatoid Arthritis synovial fibroblast (RASF) metabolism and activation and determined if crosstalk with the Hippo-YAP pathway mediates their effects. Methods: Primary RA synovial fibroblasts (RASF) were cultured with TNFα alone or in combination with the mTOR inhibitor Rapamycin or YAP inhibitor Verteporfin. Chemokine production, matrix metalloproteinase (MMP) production, and adhesion marker expression were quantified by real-time PCR, ELISA, and/or Flow Cytometry. Invasion assays were performed using Transwell invasion chambers, while wound repair assays were used to assess RASF migration. Cellular bioenergetics was assessed using the Seahorse XFe96 Analyzer. Key metabolic genes (GLUT-1, HK2, G6PD) were measured using real-time PCR. Reanalysis of RNA-Seq analysis was performed on RA (n = 151) and healthy control (HC) (n = 28) synovial tissue biopsies to detect differential gene and pathway expression. The expression of YAP was measured by Western Blot. Results: Transcriptomic analysis of healthy donor and RA synovial tissue revealed dysregulated expression of several key components of the mTOR pathway in RA. Moreover, the expression of phospho-ribosomal protein S6 (pS6), the major downstream target of mTOR is specifically increased in RA synovial fibroblasts compared to healthy tissue. In the presence of TNFα, RASF display heightened phosphorylation of S6 and are responsive to mTOR inhibition via Rapamycin. Rapamycin effectively alters RASF cellular bioenergetics by inhibiting glycolysis and the expression of rate limiting glycolytic enzymes. Furthermore, we demonstrate a key role for mTOR signaling in uniquely mediating RASF migratory and invasive mechanisms, which are significantly abrogated in the presence of Rapamycin. Finally, we report a significant upregulation in several genes involved in the Hippo-YAP pathway in RA synovial tissue, which are predicted to converge with the mTOR pathway. We demonstrate crosstalk between the mTOR and YAP pathways in mediating RASF invasive mechanism whereby Rapamycin significantly abrogates YAP expression and YAP inhibition significantly inhibits RASF invasiveness. Conclusion: mTOR drives pathogenic mechanisms in RASF an effect which is in part mediated via crosstalk with the Hippo-YAP pathway.

Innate lymphoid cell (ILC) subsets are enriched in the skin of patients with hidradenitis suppurativa.

Hidradenitis suppurativa (HS) is a chronic relapsing inflammatory skin disease manifested as painful inflamed lesions including deep-seated nodules, abscesses and sinus tracts. The exact aetiology of HS is unclear. Recent evidence suggests that immune dysregulation plays a crucial role in pathogenesis and disease progression. Innate lymphoid cells (ILC) are a recently identified immune cell subset involved in mediating immunity, however their role in HS has not yet been investigated. Three distinct subsets of ILC- ILC1, ILC2 and ILC3 have been described, and these are involved in skin tissue homeostasis and pathologic inflammation associated with autoimmunity and allergic diseases. In this study, we analysed by multiparameter flow cytometry the frequencies of ILC subsets in skin and peripheral blood mononuclear cells (PBMC) of HS patients and compared these to healthy control subjects and psoriasis patients. The absolute numbers of total ILC and subsets thereof were significantly reduced in the blood of HS patients relative to healthy controls. However, when patients were stratified according to treatment, this reduction was no longer observed in patients undergoing anti-TNF treatment. In HS lesional skin the absolute numbers of ILC were significantly increased relative to control skin. Furthermore, the frequencies of total ILC as well as ILC2 and ILC3 were significantly higher in non-lesional than lesional HS skin. This study analysed for the first time the presence of ILC subsets in the blood and skin of HS patients. Our findings suggest that ILC may participate in HS pathogenesis.

B-cell and complement signature in severe hidradenitis suppurativa that does not respond to adalimumab.

BACKGROUND: Hidradenitis suppurativa (HS) is a chronic inflammatory skin disorder with significant morbidity. The pathogenesis remains incompletely understood although immune dysregulation plays an important role. It is challenging to treat and approximately 50% of patients respond clinically to adalimumab, the only licensed treatment. OBJECTIVES: To examine differences between lesional and nonlesional HS skin at baseline using bulk RNA sequencing, and to compare the transcriptome in the skin before and after 12 weeks of treatment with adalimumab. To examine transcriptomic differences between adalimumab responders and nonresponders using Hidradenitis Suppurativa Clinical Response and the International Hidradenitis Suppurativa Severity Score System (IHS4); and to compare transcriptomic differences based on disease severity (Hurley stage and IHS4). METHODS: We completed bulk RNA sequencing on lesional and nonlesional skin samples of patients before and after 12 weeks of treatment with adalimumab. RESULTS: Baseline differentially expressed genes and pathways between lesional and nonlesional skin highlighted chemokines and antimicrobial peptides produced by keratinocytes; B-cell function; T-cell-receptor, interleukin-17 and nuclear factor-κB signalling; and T-helper-cell differentiation. Transcriptomic differences were identified in lesional skin at baseline, between subsequent responders and nonresponders. Patients with severe HS who did not respond to adalimumab had enriched complement and B-cell activation pathways at baseline. In addition, logistic regression identified CCL28 in baseline lesional HS skin as a potential biomarker of treatment response. CONCLUSIONS: This highlights the potential for targeting B-cell and complement pathways in HS treatment and the potential of stratifying patients at baseline to the most suitable treatment based on the skin transcriptome. CCL28 has not previously been identified in HS skin and has potential clinical relevance due to its antimicrobial function and homing of B and T cells at epithelial surfaces. Our results provide data to inform future translational and clinical studies on therapeutics in HS.

Animated teaching improves student learning of human gastrulation and neurulation.

Human embryology is a complex topic that brings together core components of anatomy and physiology to describe the developmental process from fertilisation to birth. Embryonic development is a challenging topic of study that is core to the curricula for health science students. There are challenges ingrained in teaching and learning embryology, due to the three-dimensional dynamic processes that occur as the embryo develops. This study aimed to develop and assess two newly developed animations depicting key processes in embryology, namely gastrulation and neurulation, as supplemental learning aids for students. Indeed, animated teaching tools to enhance the learning of gastrulation and neurulation are not widely available. A multi-disciplinary team of physiologists, biochemists, anatomists, and a computer scientist developed the animation sets. A student cohort of 81 first-year health science students were enrolled in this study over a period of three academic years. Both animations are in line with the course content of the first-year health science students undertaking the Human Health and Disease BSc at Trinity College Dublin, who were the study participants. Participants were randomly assigned to a non-animation control group and an animation group. Each set of animated teaching aids was broken down into individual clips which were given identifiable headings to allow the user to interchange between clips to facilitate a more personal learning experience. The animation group had open access to the animations for a three-week period. Questionnaires were designed to assess participants' attitude to the animations and their knowledge of embryology, both at the start of the study and three weeks later following access to the animations. Data presented herein indicate that students incorporated the animated teaching aids into digital home study and that the use of the animations acted as a supplemental tool that increased student knowledge in key areas of human embryology. From a qualitative point of view, students described the animations as enjoyable and helpful in visualising complex processes. This study indicates that the development of gastrulation and neurulation animated learning tools allow for a more engaging learning experience, facilitating student's engagement with academically challenging concepts in human embryology.

Distinct stromal and immune cell interactions shape the pathogenesis of rheumatoid and psoriatic arthritis.

OBJECTIVES: Immune and stromal cell communication is central in the pathogenesis of rheumatoid arthritis (RA) and psoriatic arthritis (PsA), however, the nature of these interactions in the synovial pathology of the two pathotypes can differ. Identifying immune-stromal cell crosstalk at the site of inflammation in RA and PsA is challenging. This study creates the first global transcriptomic analysis of the RA and PsA inflamed joint and investigates immune-stromal cell interactions in the pathogenesis of synovial inflammation. METHODS: Single cell transcriptomic profiling of 178 000 synovial tissue cells from five patients with PsA and four patients with RA, importantly, without prior sorting of immune and stromal cells. This approach enabled the transcriptomic analysis of the intact synovial tissue and identification of immune and stromal cell interactions. State of the art data integration and annotation techniques identified and characterised 18 stromal and 14 immune cell clusters. RESULTS: Global transcriptomic analysis of synovial cell subsets identifies actively proliferating synovial T cells and indicates that due to differential λ and κ immunoglobulin light chain usage, synovial plasma cells are potentially not derived from the local memory B cell pool. Importantly, we report distinct fibroblast and endothelial cell transcriptomes indicating abundant subpopulations in RA and PsA characterised by differential transcription factor usage. Using receptor-ligand interactions and downstream target characterisation, we identify RA-specific synovial T cell-derived transforming growth factor (TGF)-ß and macrophage interleukin (IL)-1ß synergy in driving the transcriptional profile of FAPα+THY1+ invasive synovial fibroblasts, expanded in RA compared with PsA. In vitro characterisation of patient with RA synovial fibroblasts showed metabolic switch to glycolysis, increased adhesion intercellular adhesion molecules 1 expression and IL-6 secretion in response to combined TGF-ß and IL-1ß treatment. Disrupting specific immune and stromal cell interactions offers novel opportunities for targeted therapeutic intervention in RA and PsA.

The Trypanosoma brucei-Derived Ketoacids, Indole Pyruvate and Hydroxyphenylpyruvate, Induce HO-1 Expression and Suppress Inflammatory Responses in Human Dendritic Cells.

The extracellular parasite and causative agent of African sleeping sickness Trypanosoma brucei (T. brucei) has evolved a number of strategies to avoid immune detection in the host. One recently described mechanism involves the conversion of host-derived amino acids to aromatic ketoacids, which are detected at relatively high concentrations in the bloodstream of infected individuals. These ketoacids have been shown to directly suppress inflammatory responses in murine immune cells, as well as acting as potent inducers of the stress response enzyme, heme oxygenase 1 (HO-1), which has proven anti-inflammatory properties. The aim of this study was to investigate the immunomodulatory properties of the T. brucei-derived ketoacids in primary human immune cells and further examine their potential as a therapy for inflammatory diseases. We report that the T. brucei-derived ketoacids, indole pyruvate (IP) and hydroxyphenylpyruvate (HPP), induce HO-1 expression through Nrf2 activation in human dendritic cells (DC). They also limit DC maturation and suppress the production of pro-inflammatory cytokines, which, in turn, leads to a reduced capacity to differentiate adaptive CD4+ T cells. Furthermore, the ketoacids are capable of modulating DC cellular metabolism and suppressing the inflammatory profile of cells isolated from patients with inflammatory bowel disease. This study therefore not only provides further evidence of the immune-evasion mechanisms employed by T. brucei, but also supports further exploration of this new class of HO-1 inducers as potential therapeutics for the treatment of inflammatory conditions.

The Pathogenesis of Parkinson's Disease: A Complex Interplay Between Astrocytes, Microglia, and T Lymphocytes?

Parkinson's disease (PD), the second most common neurodegenerative disease, is characterised by the motor symptoms of bradykinesia, rigidity and resting tremor and non-motor symptoms of sleep disturbances, constipation, and depression. Pathological hallmarks include neuroinflammation, degeneration of dopaminergic neurons in the substantia nigra pars compacta, and accumulation of misfolded α-synuclein proteins as intra-cytoplasmic Lewy bodies and neurites. Microglia and astrocytes are essential to maintaining homeostasis within the central nervous system (CNS), including providing protection through the process of gliosis. However, dysregulation of glial cells results in disruption of homeostasis leading to a chronic pro-inflammatory, deleterious environment, implicated in numerous CNS diseases. Recent evidence has demonstrated a role for peripheral immune cells, in particular T lymphocytes in the pathogenesis of PD. These cells infiltrate the CNS, and accumulate in the substantia nigra, where they secrete pro-inflammatory cytokines, stimulate surrounding immune cells, and induce dopaminergic neuronal cell death. Indeed, a greater understanding of the integrated network of communication that exists between glial cells and peripheral immune cells may increase our understanding of disease pathogenesis and hence provide novel therapeutic approaches.

Targeting bioenergetics prevents CD4 T cell-mediated activation of synovial fibroblasts in rheumatoid arthritis.

OBJECTIVES: We investigated the reciprocal relationship linking fibroblast-like synoviocytes (FLS) and T lymphocytes in the inflamed RA synovium and subsequently targeted cellular metabolic pathways in FLS to identify key molecular players in joint inflammation. METHODS: RA FLS were cultured with CD4 T cells or T cell conditioned medium (CD4CM); proliferation, expression of adhesion molecules and intracellular cytokines were examined by flow cytometry. FLS invasiveness and secreted cytokines were measured by transwell matrigel invasion chambers and ELISA, while metabolic profiles were determined by extracellular Seahorse flux analysis. Gene expression was quantified by real-time quantitative RT-PCR. RESULTS: Our results showed mutual activation between CD4 T cells and FLS, which resulted in increased proliferation and expression of intercellular adhesion molecule 1 and vascular cell adhesion molecule 1 by both CD4 T cells and FLS. Furthermore, interaction between CD4 T cells and FLS resulted in an increased frequency of TNF-α+, IFN-γ+ and IL-17A+ CD4 T cells and augmented TNF-α, IFN-γ, IL-17A, IL-6, IL-8 and VEGF secretion. Moreover, CD4CM promoted invasiveness and boosted glycolysis in FLS while downregulating oxidative phosphorylation, effects paralleled by increased glucose transporters GLUT1 and GLUT3; key glycolytic enzymes GSK3A, HK2, LDHA and PFKFB3; angiogenic factor VEGF and MMP-3 and MMP-9. Importantly, these effects were reversed by the glycolytic inhibitor 2-DG and AMP analogue 5-aminoimidazole-4-carboxamide ribonucleotide (AICAR). CONCLUSION: This study demonstrates that CD4 T cells elicit an aggressive phenotype in FLS, which subsequently upregulate glycolysis to meet the increased metabolic demand. Accordingly, 2-DG and AICAR prevent this activation, suggesting that glycolytic manipulation could have clinical implications for RA treatment.

Plant-Derived Polyphenols Modulate Human Dendritic Cell Metabolism and Immune Function via AMPK-Dependent Induction of Heme Oxygenase-1.

Polyphenols are important immunonutrients which have been investigated in the context of inflammatory and autoimmune disease due to their significant immunosuppressive properties. However, the mechanism of action of many polyphenols is unclear, particularly in human immune cells. The emerging field of immunometabolism has highlighted the significance of metabolic function in the regulation of immune cell activity, yet the effects of polyphenols on immune cell metabolic signaling and function has not been explored. We have investigated the effects of two plant-derived polyphenols, carnosol and curcumin, on the metabolism of primary human dendritic cells (DC). We report that human DC display an increase in glycolysis and spare respiratory capacity in response to LPS stimulation, which was attenuated by both carnosol and curcumin treatment. The regulation of DC metabolism by these polyphenols appeared to be mediated by their activation of the cellular energy sensor, AMP-activated Protein Kinase (AMPK), which resulted in the inhibition of mTOR signaling in LPS-stimulated DC. Previously we have reported that both carnosol and curcumin can regulate the maturation and function of human DC through upregulation of the immunomodulatory enzyme, Heme Oxygenase-1 (HO-1). Here we also demonstrate that the induction of HO-1 by polyphenols in human DC is dependent on their activation of AMPK. Moreover, pharmacological inhibition of AMPK was found to reverse the observed reduction of DC maturation by carnosol and curcumin. This study therefore describes a novel relationship between metabolic signaling via AMPK and HO-1 induction by carnosol and curcumin in human DC, and characterizes the effects of these polyphenols on DC immunometabolism for the first time. These results expand our understanding of the mechanism of action of carnosol and curcumin in human immune cells, and suggest that polyphenol supplementation may be useful to regulate the metabolism and function of immune cells in inflammatory and metabolic disease.

Differential Regulation of Human Treg and Th17 Cells by Fatty Acid Synthesis and Glycolysis.

In this study we examined the metabolic requirements of human T helper cells and the effect of manipulating metabolic pathways in Th17 and Treg cells. The Th17:Treg cell axis is dysregulated in a number of autoimmune or inflammatory diseases and therefore it is of key importance to identify novel strategies to modulate this axis in favor of Treg cells. We investigated the role of carbohydrate and fatty acid metabolism in the regulation of human memory T helper cell subsets, in order to understand how T cells are regulated at the site of inflammation where essential nutrients including oxygen may be limiting. We found that Th17 lineage cells primarily utilize glycolysis, as glucose-deprivation and treatment with rapamycin resulted in a reduction in these cells. On the other hand, Treg cells exhibited increased glycolysis, mitochondrial respiration, and fatty acid oxidation, whereas Th17 cells demonstrated a reliance upon fatty acid synthesis. Treg cells were somewhat reliant on glycolysis, but to a lesser extent than Th17 cells. Here we expose a fundamental difference in the metabolic requirements of human Treg and Th17 cells and a possible mechanism for manipulating the Th17:Treg cell axis.

Enriched Cd141+ DCs in the joint are transcriptionally distinct, activated, and contribute to joint pathogenesis.

CD141+ DC are implicated in antiviral and antitumor immunity. However, mechanistic studies in autoimmune disease are limited. This is the first study to our knowledge examining CD141+ DC in autoimmune disease, specifically inflammatory arthritis (IA). We identified significant enrichment of CD141+ DC in the inflamed synovial joint, which were transcriptionally distinct from IA and healthy control (HC) blood CD141+ DC and significantly more activated, and they exhibited increased responsiveness to TLR3. Synovial CD141+ DC represent a bone fide CD141+ DC population that is distinct from CD1c+ DC. Synovial CD141+ DC induced higher levels of CD4+ and CD8+ T cell activation compared with their peripheral blood counterparts, as made evident by expression of IFN-γ, TNF-α, and granulocyte-macrophage CSF (GMCSF). Autologous synovial CD141+ DC cocultures also induce higher levels of these cytokines, further highlighting their contribution to synovial inflammation. Synovial CD141+ DC-T cell interactions had the ability to further activate synovial fibroblasts, inducing adhesive and invasive pathogenic mechanisms. Furthermore, we identify a mechanism in which synovial CD141+ DC are activated, via ligation of the hypoxia-inducible immune-amplification receptor TREM-1, which increased synovial CD141+ DC activation, migratory capacity, and proinflammatory cytokines. Thus, synovial CD141+ DC display unique mechanistic and transcriptomic signatures, which are distinguishable from blood CD141+ DC and can contribute to synovial joint inflammation.

Naturally derived Heme-Oxygenase 1 inducers attenuate inflammatory responses in human dendritic cells and T cells: relevance for psoriasis treatment.

Psoriasis is a chronic autoimmune disease mediated by dysregulated immune responses in dendritic cells (DC) and T cells. The stress-response enzyme heme oxygenase-1 (HO-1) has been described as protective in animal models of psoriasis, however, implementation of HO-1-based therapies is hindered by the lack of clinically-suitable HO-1 inducers. The plant-derived polyphenols, carnosol and curcumin, have been identified as candidate HO-1 inducers however there has been little investigation into their effects on human immune cells. We demonstrate that treatment of human DC with these polyphenols limits DC maturation, reduces pro-inflammatory cytokine production, and prevents induction of allospecific T cell responses, in a manner partially dependent on carbon monoxide (CO). We also characterised their effects in ex-vivo psoriasis PBMC and report that curcumin, but not carnosol, strongly reduces T cell proliferation and cytokine poly-functionality, with reduced expression of psoriatic cytokines IFNγ, IL-17, GM-CSF and IL-22. This study therefore supports reports highlighting the therapeutic potential of curcumin in psoriasis by providing insight into its immunological effects on healthy human DC and psoriasis PBMC. We also demonstrate, for the first time, the anti-inflammatory effects of carnosol in human immune cells.

Effects of vitamin D3 in clinically isolated syndrome and healthy control participants: A double-blind randomised controlled trial.

BACKGROUND: Lowserum vitamin D levels are associated with susceptibility to, and severity of, multiple sclerosis. High dose vitamin D has been proposed as a potential immunomodulator in multiple sclerosis. OBJECTIVES: We performed a single centre, investigator-led, exploratory, double-blind, randomised, placebo controlled, trial of vitamin D3 in clinically isolated syndrome and healthy control participants to assess its immunological effects. Secondary end-points included clinical and magnetic resonance imaging outcomes and safety. METHODS: Clinically isolated syndrome patients and healthy control participants were randomised to: placebo, 5000 IU or 10,000 IU vitamin D3/day (Vigantol oil). Study duration was 24 weeks. RESULTS: The trial did not meet its primary end point, with no difference in the frequency of pro-inflammatory CD4+ T cells (interleukin (IL)-17+/interferon (IFN)-γ+) seen. A higher level of disease freedom (67% versus 50%) was seen in those with serum 1,25 (OH) vitamin D levels>100 nmol/l but this did not reach significance. High dose vitamin D3 was well tolerated with no safety signal. CONCLUSIONS: High dose vitamin D3 over 24 weeks was well tolerated but without immunological, magnetic resonance imaging or clinical evidence of benefit. The hypothesised therapeutic effects in clinically isolated syndrome or multiple sclerosis patients may require longer periods of administration or may only be seen in patients treated with vitamin D3 as an adjunct to established disease modifying therapies.