Fine tuning calcium dynamics by inhibition of Store-operated Calcium Entry as a novel therapeutic approach for the treatment of chronic pancreatitis.

Chronic pancreatitis (CP) is a complex inflammatory disorder characterized by progressive fibrosis, leading to pancreatic dysfunction, reduced quality of life and an elevated pancreatic cancer risk. Current therapeutic options for CP are restricted to symptomatic treatment. Using ex vivo and in vivo preclinical disease models, Szabó et al. now explored for the first time the involvement of Store-operated Ca2+ entry (SOCE) in the progression of CP and propose that a selective pharmacological inhibition of the SOCE signaling component Orai1 might serve as specific treatment option for CP[1,2].

Feasibility of in vivo magnetic resonance elastography of mesenteric adipose tissue in Crohn's disease.

Background: Although there is growing evidence that functional involvement and structural changes of mesenteric adipose tissue (MAT) influence the course of Crohn's disease (CD), its viscoelastic properties remain elusive. Therefore, we aimed to investigate the viscoelastic properties of MAT in CD using magnetic resonance elastography (MRE), providing reference values for CD diagnosis. Methods: In this prospective proof-of-concept study, 31 subjects (CD: n=11; healthy controls: n=20) were consecutively enrolled in a specialized care center for inflammatory bowel diseases (tertiary/quaternary care). Inclusion criteria for the CD patients were a clinically and endoscopically established diagnosis of CD based on the clinical record, absence of other concurrent bowel diseases, scheduled surgery for the following day, and age of at least 18 years. Diagnoses were confirmed by histological analysis of the resected bowel the day after MRE. Subjects were investigated using MRE at 1.5-T with frequencies of 40-70 Hz. To retrieve shear wave speed (SWS), volumes of interest (VOIs) in MAT were drawn adjacent to CD lesions (MATCD) and on the opposite side without adjacent bowel lesions in patients (MATCD_Opp) and controls (MATCTRL). The presented study is not registered in the clinical trial platform. Results: A statistically significant decrease in mean SWS of 7% was found for MATCD_Opp vs. MATCTRL (0.76±0.05 vs. 0.82±0.04 m/s, P=0.012), whereas there was a nonsignificant trend with an 8% increase for MATCD vs. MATCD_Opp (0.82±0.07 vs. 0.76±0.05 m/s, P=0.098) and no difference for MATCD vs. MATCTRL. Preliminary area under the receiver operating characteristic curve (AUC) analysis showed diagnostic accuracy in detecting CD to be excellent for SWS of MATCD_Opp [AUC =0.82; 95% confidence interval (CI): 0.64-0.96] but poor for SWS of MATCD (AUC =0.52; 95% CI: 0.34-0.73). Conclusions: This study demonstrates the feasibility of MRE of MAT and presents preliminary reference values for CD patients and healthy controls. Our results motivate further studies for the biophysical characterization of MAT in inflammatory bowel disease.

Contribution of Blood Vessel Activation, Remodeling and Barrier Function to Inflammatory Bowel Diseases.

Inflammatory bowel diseases (IBDs) consist of a group of chronic inflammatory disorders with a complex etiology, which represent a clinical challenge due to their often therapy-refractory nature. In IBD, inflammation of the intestinal mucosa is characterized by strong and sustained leukocyte infiltration, resulting in the loss of epithelial barrier function and subsequent tissue destruction. This is accompanied by the activation and the massive remodeling of mucosal micro-vessels. The role of the gut vasculature in the induction and perpetuation of mucosal inflammation is receiving increasing recognition. While the vascular barrier is considered to offer protection against bacterial translocation and sepsis after the breakdown of the epithelial barrier, endothelium activation and angiogenesis are thought to promote inflammation. The present review examines the respective pathological contributions of the different phenotypical changes observed in the microvascular endothelium during IBD, and provides an overview of potential vessel-specific targeted therapy options for the treatment of IBD.

Store-operated calcium entry controls innate and adaptive immune cell function in inflammatory bowel disease.

Inflammatory bowel disease (IBD) is characterized by dysregulated intestinal immune responses. Using mass cytometry (CyTOF) to analyze the immune cell composition in the lamina propria (LP) of patients with ulcerative colitis (UC) and Crohn's disease (CD), we observed an enrichment of CD4+ effector T cells producing IL-17A and TNF, CD8+ T cells producing IFNγ, T regulatory (Treg) cells, and innate lymphoid cells (ILC). The function of these immune cells is regulated by store-operated Ca2+ entry (SOCE), which results from the opening of Ca2+ release-activated Ca2+ (CRAC) channels formed by ORAI and STIM proteins. We observed that the pharmacologic inhibition of SOCE attenuated the production of proinflammatory cytokines including IL-2, IL-4, IL-6, IL-17A, TNF, and IFNγ by human colonic T cells and ILCs, reduced the production of IL-6 by B cells and the production of IFNγ by myeloid cells, but had no effect on the viability, differentiation, and function of intestinal epithelial cells. T cell-specific deletion of CRAC channel genes in mice showed that Orai1, Stim1, and Stim2-deficient T cells have quantitatively distinct defects in SOCE, which correlate with gradually more pronounced impairment of cytokine production by Th1 and Th17 cells and the severity of IBD. Moreover, the pharmacologic inhibition of SOCE with a selective CRAC channel inhibitor attenuated IBD severity and colitogenic T cell function in mice. Our data indicate that SOCE inhibition may be a suitable new approach for the treatment of IBD.

Anti-SARS-CoV2 antibody-mediated cytokine release syndrome in a patient with acute promyelocytic leukemia.

BACKGROUND: Passive immunization against SARS-CoV-2 limits viral burden and death from COVID-19; however, it poses a theoretical risk of disease exacerbation through antibody-dependent enhancement (ADE). ADE after anti-SARS-CoV2 antibody treatment has not been reported, and therefore the potential risk and promoting factors remain unknown. CASE PRESENTATION: A 75-year-old female was admitted to the emergency room with recurrent, unexplained bruises and leukocytopenia, anemia, and thrombocytopenia. Evaluation of a bone marrow biopsy established the diagnosis of an acute promyelocytic leukemia (APL). SARS-CoV-2 RT-PCR testing of nasal and throat swabs on admission was negative. During the routine SARS-CoV-2 testing of inpatients, our patient tested positive for SARS-CoV-2 on day 14 after admission without typical COVID-19 symptoms. Due to disease- and therapy-related immunosuppression and advanced age conferring a high risk of progressing to severe COVID-19, casirivimab and imdevimab were administered as a preemptive approach. The patient developed immune activation and cytokine release syndrome (CRS) occurring within four hours of preemptive anti-SARS-CoV2 antibody (casirivimab/imdevimab) infusion. Immune activation and CRS were evidenced by a rapid increase in serum cytokines (IL-6, TNFα, IL-8, IL-10), acute respiratory insufficiency, and progressive acute respiratory distress syndrome. DISCUSSION AND CONCLUSION: The temporal relationship between therapeutic antibody administration and the rapid laboratory, radiological, and clinical deterioration suggests that CRS was an antibody-related adverse event, potentially exacerbated by APL treatment-mediated differentiation of leukemic blasts and promyelocytes. This case highlights the need for careful assessment of life-threatening adverse events after passive SARS-CoV-2 immunization, especially in the clinical context of patients with complex immune and hematological landscapes.

Activated SUMOylation restricts MHC class I antigen presentation to confer immune evasion in cancer.

Activated SUMOylation is a hallmark of cancer. Starting from a targeted screening for SUMO-regulated immune evasion mechanisms, we identified an evolutionarily conserved function of activated SUMOylation, which attenuated the immunogenicity of tumor cells. Activated SUMOylation allowed cancer cells to evade CD8+ T cell-mediated immunosurveillance by suppressing the MHC class I (MHC-I) antigen-processing and presentation machinery (APM). Loss of the MHC-I APM is a frequent cause of resistance to cancer immunotherapies, and the pharmacological inhibition of SUMOylation (SUMOi) resulted in reduced activity of the transcriptional repressor scaffold attachment factor B (SAFB) and induction of the MHC-I APM. Consequently, SUMOi enhanced the presentation of antigens and the susceptibility of tumor cells to CD8+ T cell-mediated killing. Importantly, SUMOi also triggered the activation of CD8+ T cells and thereby drove a feed-forward loop amplifying the specific antitumor immune response. In summary, we showed that activated SUMOylation allowed tumor cells to evade antitumor immunosurveillance, and we have expanded the understanding of SUMOi as a rational therapeutic strategy for enhancing the efficacy of cancer immunotherapies.

A Notch/STAT3-driven Blimp-1/c-Maf-dependent molecular switch induces IL-10 expression in human CD4+ T cells and is defective in Crohn´s disease patients.

Immunosuppressive Interleukin (IL)-10 production by pro-inflammatory CD4+ T cells is a central self-regulatory function to limit aberrant inflammation. Still, the molecular mediators controlling IL-10 expression in human CD4+ T cells are largely undefined. Here, we identify a Notch/STAT3 signaling-module as a universal molecular switch to induce IL-10 expression across human naïve and major effector CD4+ T cell subsets. IL-10 induction was transient, jointly controlled by the transcription factors Blimp-1/c-Maf and accompanied by upregulation of several co-inhibitory receptors, including LAG-3, CD49b, PD-1, TIM-3 and TIGIT. Consistent with a protective role of IL-10 in inflammatory bowel diseases (IBD), effector CD4+ T cells from Crohn's disease patients were defective in Notch/STAT3-induced IL-10 production and skewed towards an inflammatory Th1/17 cell phenotype. Collectively, our data identify a Notch/STAT3-Blimp-1/c-Maf axis as a common anti-inflammatory pathway in human CD4+ T cells, which is defective in IBD and thus may represent an attractive therapeutic target.

An Integrated Taxonomy for Monogenic Inflammatory Bowel Disease.

BACKGROUND & AIMS: Monogenic forms of inflammatory bowel disease (IBD) illustrate the essential roles of individual genes in pathways and networks safeguarding immune tolerance and gut homeostasis. METHODS: To build a taxonomy model, we assessed 165 disorders. Genes were prioritized based on penetrance of IBD and disease phenotypes were integrated with multi-omics datasets. Monogenic IBD genes were classified by (1) overlapping syndromic features, (2) response to hematopoietic stem cell transplantation, (3) bulk RNA-sequencing of 32 tissues, (4) single-cell RNA-sequencing of >50 cell subsets from the intestine of healthy individuals and patients with IBD (pediatric and adult), and (5) proteomes of 43 immune subsets. The model was validated by addition of newly identified monogenic IBD defects. As a proof-of-concept, we explore the intersection between immunometabolism and antimicrobial activity for a group of disorders (G6PC3/SLC37A4). RESULTS: Our quantitative integrated taxonomy defines the cellular landscape of monogenic IBD gene expression across 102 genes with high and moderate penetrance (81 in the model set and 21 genes in the validation set). We illustrate distinct cellular networks, highlight expression profiles across understudied cell types (e.g., CD8+ T cells, neutrophils, epithelial subsets, and endothelial cells) and define genotype-phenotype associations (perianal disease and defective antimicrobial activity). We illustrate processes and pathways shared across cellular compartments and phenotypic groups and highlight cellular immunometabolism with mammalian target of rapamycin activation as one of the converging pathways. There is an overlap of genes and enriched cell-specific expression between monogenic and polygenic IBD. CONCLUSION: Our taxonomy integrates genetic, clinical and multi-omic data; providing a basis for genomic diagnostics and testable hypotheses for disease functions and treatment responses.

Environmental Microbial Factors Determine the Pattern of Inflammatory Lesions in a Murine Model of Crohn's Disease-Like Inflammation.

Crohn's disease (CD) patients can be grouped into patients suffering from ileitis, ileocolitis, jejunoileitis, and colitis. The pathophysiological mechanism underlying this regional inflammation is still unknown. Although most murine models of inflammatory bowel disease (IBD) develop inflammation in the colon, there is an unmet need for novel models that recapitulate the spontaneous and fluctuating nature of inflammation as seen in CD. Recently, mice with an intestinal epithelial cell-specific deletion for Caspase-8 (Casp8ΔIEC mice), which are characterized by cell death-driven ileitis and disrupted Paneth cell homeostasis, have been identified as a novel model of CD-like ileitis. Here we uncovered that genetic susceptibility alone is sufficient to drive ileitis in Casp8ΔIEC mice. In sharp contrast, environmental factors, such as a disease-relevant microbial flora, determine colonic inflammation. Accordingly, depending on the microbial environment, isogenic Casp8ΔIEC mice either exclusively developed ileitis or suffered from pathologies in several parts of the gastrointestinal tract. Colitis in these mice was characterized by massive epithelial cell death, leading to spread of commensal gut microbes to the extra-intestinal space and hence an aberrant activation of the systemic immunity. We further uncovered that Casp8ΔIEC mice show qualitative and quantitative changes in the intestinal microbiome associated with an altered mucosal and systemic immune response. In summary, we identified that inflammation in this murine model of CD-like inflammation is characterized by an immune reaction, presumably directed against a disease-relevant microbiota in a genetically susceptible host, with impaired mucosal barrier function and bacterial clearance at the epithelial interface.

Leptin induces TNFα-dependent inflammation in acquired generalized lipodystrophy and combined Crohn's disease.

Leptin has been shown to modulate intestinal inflammation in mice. However, clinical evidence regarding its immune-stimulatory potential in human Crohn's disease remains sparse. We here describe a patient with the unique combination of acquired generalized lipodystrophy and Crohn's disease (AGLCD) featuring a lack of adipose tissue, leptin deficiency and intestinal inflammation. Using mass and flow cytometry, immunohistochemistry and functional metabolic analyses, the AGLCD patient was compared to healthy individuals and Crohn's disease patients regarding immune cell composition, function and metabolism and the effects of recombinant N-methionylleptin (rLeptin) were evaluated. We provide evidence that rLeptin exerts diverse pro-inflammatory effects on immune cell differentiation and function, including the metabolic reprogramming of immune cells and the induction of TNFα, ultimately aggravating Crohn's disease in the AGLCD patient, which can be reversed by anti-TNFα therapy. Our results indicate that leptin is required for human immune homeostasis and contributes to autoimmunity in a TNFα-dependent manner.

Metabolic Control of Epigenetics and Its Role in CD8+ T Cell Differentiation and Function.

Epigenetic programs that control posttranslational modifications of histone proteins and DNA itself tightly regulate transcriptional networks determining the identity and function of CD8+ T cells. Chromatin-modifying enzymes such as histone acetyltransferases and deacetylases, represent key molecular determinants of the epigenetic imprinting of CD8+ T cells. The functions of these enzymes highly depend on the availability of key products of cellular metabolism pathways such as acetyl-CoA, NAD (Nicotinamide adenine dinucleotide) and SEM (S-adenosylmethionine), suggesting that there is a close crosstalk between the metabolic and the epigenetic regulation of CD8+ T cells. In this review, we will discuss the metabolic regulation of CD8+ T cell epigenetics during activation and differentiation. We will furthermore summarize how metabolic signals from the tumor microenvironment (TME) shape the epigenetic landscape of CD8+ T cells to better understand the mechanism underlying CD8+ T cell exhaustion in anti-tumor and anti-viral immunity, which might help to overcome limitations of current CD8+ T cell-based therapies.

HDAC inhibitors promote intestinal epithelial regeneration via autocrine TGFß1 signalling in inflammation.

Intact epithelial barrier function is pivotal for maintaining intestinal homeostasis. Current therapeutic developments aim at restoring the epithelial barrier in inflammatory bowel disease. Histone deacetylase (HDAC) inhibitors are known to modulate immune responses and to ameliorate experimental colitis. However, their direct impact on epithelial barrier function and intestinal wound healing is unknown. In human and murine colonic epithelial cell lines, the presence of the HDAC inhibitors Givinostat and Vorinostat not only improved transepithelial electrical resistance under inflammatory conditions but also attenuated the passage of macromolecules across the epithelial monolayer. Givinostat treatment mediated an accelerated wound closure in scratch assays. In vivo, Givinostat treatment resulted in improved barrier recovery and epithelial wound healing in dextran sodium sulphate-stressed mice. Mechanistically, these regenerative effects could be linked to an increased secretion of transforming growth factor beta1 and interleukin 8, paralleled by differential expression of the tight junction proteins claudin-1, claudin-2 and occludin. Our data reveal a novel tissue regenerative property of the pan-HDAC inhibitors Givinostat and Vorinostat in intestinal inflammation, which may have beneficial implications by repurposing HDAC inhibitors for therapeutic strategies for inflammatory bowel disease.

The role of adipose tissue in inflammatory bowel diseases.

PURPOSE OF REVIEW: The occurrence of creeping fat wrapping segments of inflamed gut represents a characteristic yet incompletely understood hallmark of Crohn's disease. Over the last decade, numerous studies have provided a limited understanding of this feature. Still, deciphering the detailed mechanisms and the pathophysiologic relevance of the interplay between creeping fat, barrier function and intestinal inflammation will be the aim of future studies. RECENT FINDINGS: The last 18 months have substantially contributed to this field, starting with an elegant three-dimensional study revealing B cell aggregates around lymphatic vessels embedded in the mesenteric fat, thus bringing back the idea that Crohn's disease might represent a 'lymphatic disease'. Furthermore, studies on a cellular level elucidated the interplay of mesenteric adipocytes, immune cells and intestinal epithelial cells. Last, imaging studies provide evidence indicating that changes depicted by computed tomography within the mesenteric fat compartment rather than of the bowel wall are predictive for the presence of endoscopic lesions. This underlines the impact of mesenteric changes on Crohn's disease activity. SUMMARY: The findings of the last 18 months further contribute to solving the puzzle that will ultimately reveal the role of the mesenteric fat tissue in the control of intestinal immunity and inflammation.

Oleate but not stearate induces the regulatory phenotype of myeloid suppressor cells.

Tumor infiltrating myeloid cells play contradictory roles in the tumor development. Dendritic cells and classical activated macrophages support anti-tumor immune activity via antigen presentation and induction of pro-inflammatory immune responses. Myeloid suppressor cells (MSCs), for instance myeloid derived suppressor cells (MDSCs) or tumor associated macrophages play a critical role in tumor growth. Here, treatment with sodium oleate, an unsaturated fatty acid, induced a regulatory phenotype in the myeloid suppressor cell line MSC-2 and resulted in an increased suppression of activated T cells, paralleled by increased intracellular lipid droplets formation. Furthermore, sodium oleate potentiated nitric oxide (NO) production in MSC-2, thereby increasing their suppressive capacity. In primary polarized bone marrow cells, sodium oleate (C18:1) and linoleate (C18:2), but not stearate (C18:0) were identified as potent FFA to induce a regulatory phenotype. This effect was abrogated in MSC-2 as well as primary cells by specific inhibition of droplets formation while the inhibition of de novo FFA synthesis proved ineffective, suggesting a critical role for exogenous FFA in the functional induction of MSCs. Taken together our data introduce a new unsaturated fatty acid-dependent pathway shaping the functional phenotype of MSCs, facilitating the tumor escape from the immune system.

Store-operated Ca2+ entry regulates Ca2+-activated chloride channels and eccrine sweat gland function.

Eccrine sweat glands are essential for sweating and thermoregulation in humans. Loss-of-function mutations in the Ca2+ release-activated Ca2+ (CRAC) channel genes ORAI1 and STIM1 abolish store-operated Ca2+ entry (SOCE), and patients with these CRAC channel mutations suffer from anhidrosis and hyperthermia at high ambient temperatures. Here we have shown that CRAC channel-deficient patients and mice with ectodermal tissue-specific deletion of Orai1 (Orai1K14Cre) or Stim1 and Stim2 (Stim1/2K14Cre) failed to sweat despite normal sweat gland development. SOCE was absent in agonist-stimulated sweat glands from Orai1K14Cre and Stim1/2K14Cre mice and human sweat gland cells lacking ORAI1 or STIM1 expression. In Orai1K14Cre mice, abolishment of SOCE was associated with impaired chloride secretion by primary murine sweat glands. In human sweat gland cells, SOCE mediated by ORAI1 was necessary for agonist-induced chloride secretion and activation of the Ca2+-activated chloride channel (CaCC) anoctamin 1 (ANO1, also known as TMEM16A). By contrast, expression of TMEM16A, the water channel aquaporin 5 (AQP5), and other regulators of sweat gland function was normal in the absence of SOCE. Our findings demonstrate that Ca2+ influx via store-operated CRAC channels is essential for CaCC activation, chloride secretion, and sweat production in humans and mice.

Ca2+ Signaling but Not Store-Operated Ca2+ Entry Is Required for the Function of Macrophages and Dendritic Cells.

Store-operated Ca(2+) entry (SOCE) through Ca(2+) release-activated Ca(2+) (CRAC) channels is essential for immunity to infection. CRAC channels are formed by ORAI1 proteins in the plasma membrane and activated by stromal interaction molecule (STIM)1 and STIM2 in the endoplasmic reticulum. Mutations in ORAI1 and STIM1 genes that abolish SOCE cause severe immunodeficiency with recurrent infections due to impaired T cell function. SOCE has also been observed in cells of the innate immune system such as macrophages and dendritic cells (DCs) and may provide Ca(2+) signals required for their function. The specific role of SOCE in macrophage and DC function, as well as its contribution to innate immunity, however, is not well defined. We found that nonselective inhibition of Ca(2+) signaling strongly impairs many effector functions of bone marrow-derived macrophages and bone marrow-derived DCs, including phagocytosis, inflammasome activation, and priming of T cells. Surprisingly, however, macrophages and DCs from mice with conditional deletion of Stim1 and Stim2 genes, and therefore complete inhibition of SOCE, showed no major functional defects. Their differentiation, FcR-dependent and -independent phagocytosis, phagolysosome fusion, cytokine production, NLRP3 inflammasome activation, and their ability to present Ags to activate T cells were preserved. Our findings demonstrate that STIM1, STIM2, and SOCE are dispensable for many critical effector functions of macrophages and DCs, which has important implications for CRAC channel inhibition as a therapeutic strategy to suppress pathogenic T cells while not interfering with myeloid cell functions required for innate immunity.