bluuebunny/crossref_metadata_2025 huggingface dataset:
Impaired induction of Slc26a4 promotes respiratory acidosis and severe, steroid-resistant asthma
Authors: Kim Richard Yong Hoon , Pinkerton James W , Rae Brittany E , Mayall Jemma R , Brown Alexandra C , Ali Md Khadem , Goggins Bridie J , Essilfie Ama-Tawiah , Starkey Malcolm R , To Cuong , Bosco Anthony , Horvat Jay C , Hansbro Philip M
Published: January 2023

CO2 produced by systemic cellular respiration is hydrated into carbonic acid (H2CO3) that dissociates into H+ and HCO3−. These are transported in the plasma to the lungs where HCO3− is converted back into H2CO3 and CO2 that are expelled through breathing. Evidence suggests that dysfunction of the mechanisms that govern these processes may result in the development of respiratory acidosis (RA) and the cardinal features of severe, steroid-resistant (SSR) asthma. Reduced lung function, which occurs in SSR asthma, impairs removal of volatile H2CO3 and CO2, resulting in acid accumulation and increased arterial PaCO2. Patients with severe asthma often develop complications from increased PaCO2, which skews the PaCO2/HCO3− ratio resulting in increased H+ concentration and reduced pH. We developed three mouse models of respiratory infection and ovalbumin-induced SSR allergic airways disease (SSRAAD) that are highly representative of SSR asthma in humans. We used these models to show a role for impaired homeostatic acid-base balance in SSR asthma. All three infections suppress the induction of the expression of the chloride (Cl−)/HCO− pump, Slc26a4, in the airway mucosa in AAD. Importantly, SSRAAD is associated with increased levels of free H+ ions in bronchoalveolar lavage fluid. Administration of Slc26a4-specific siRNA in steroid-sensitive AAD, which mimics the effect of decreased Slc26a4 responses in SSRAAD, induced RA and steroid-resistant airway inflammation and AHR. Importantly, treatment of RA with NaHCO3 during infection-induced SSRAAD suppressed steroid-resistant AHR. Thus, we have identified a previously unrecognised role for deficient Slc26a4 responses that result in the development of RA and the pathogenesis of SSRAAD.
Autoimmune manifestations in aged mice arise from early-life immune dysregulation
Authors: Karnell Jodi , Mahmoud Tamer I. , Wang Jingya , Wang Qiming , Wang Shu , Naiman Brian , Gross Philip , Brohawn Philip Z. , Morehouse Christopher , Aoyama Jordan , Wasserfall Clive , Carter Laura L , Atkinson Mark A. , Serreze David V , Braley-Mullen Helen , Mustelin Tomas Mikael , Kolbeck Roland , Herbst Ronald , Ettinger Rachel
Published: January 2023

Autoantibodies that arise in autoimmunity can be present years to decades prior to the onset of disease manifestations. This suggests that the initial autoimmune trigger involves a peripheral lymphoid component, which ultimately drives disease pathology in local tissues later in life. Here we show Sjögren’s Syndrome manifestations that develop in aged NOD.H-2h4 mice were driven by and dependent on peripheral dysregulation that arose in early life. Specifically, elimination of spontaneous germinal centers that developed in spleens of young NOD.H-2h4 mice by transient blockade of CD40 ligand (CD40L) or splenectomy abolished Sjögren’s pathology of aged mice. Strikingly, a single injection of anti-CD40L at 4 weeks-of-age prevented tertiary follicle neogenesis and greatly blunted the formation of key autoantibodies implicated in glandular pathology. Microarray profiling of salivary gland characterized the expression pattern of genes that increased with disease progression and showed early anti-CD40L greatly repressed B cell function, while having a broader effect on multiple biological pathways including IL-12 and interferon signaling. Importantly, a single, prophylactic treatment with anti-CD40L also inhibited the development of autoimmune thyroiditis and diabetes in NOD.H-2h4 and NOD mice, respectively, supporting a key role for CD40L in the pathophysiology of several autoimmune models. These results suggest that early peripheral immune dysregulation gives rise to autoimmune manifestations later in life and for diseases pre-dated by autoantibodies, early prophylactic intervention with biologics may prove efficacious.
The role of infiltrating macrophages and IL-6 production on seizure development after viral infection
Authors: DePaula-Silva Ana Beatriz , Hanak Tyler J. , Doty Daniel J. , Sim Jordan T. , Libbey Jane E. , Fujinami Robert S.
Published: January 2023

Epilepsy is a chronic neurological disorder characterized by recurrent seizures. Seizures, which are often associated with viral encephalitis, occur in response to imbalances between excitatory and inhibitory inputs within the brain. Patients with viral encephalitis are 16 times more likely to develop epilepsy. It is estimated that around 65 million people worldwide suffer from epilepsy. Although treatments to prevent seizures are available, 30% of the patients do not respond to the medication. In order to develop new immunomodulatory treatments that could lead to an eventual cure for seizures/epilepsy, a better understanding of the seizure development mechanism is required.

We have developed an experimental model of virus-induced seizures/epilepsy. In our model, C57BL/6J mice are infected intra-cranially with the Daniels strain (DAV) of Theiler’s murine encephalomyelitis virus. Between 3 and 10 days post infection (d.p.i.), around 50% of the infected mice will develop spontaneous acute seizures and approximately 50% of these mice develop epilepsy. Because seizures begin to develop at 3 d.p.i we believe it happens as a consequence of the activation of the innate immune response. We found that seizures are correlated with an increase in myeloid cells infiltrating into the brain. Also, the pro-inflammatory cytokines IL-6 and TNF alpha were elevated at 3 d.p.i. IL-6 knockout mice infected with DAV experienced significantly fewer seizures. Because infiltrating macrophages are the main producers of IL-6, we depleted macrophages from mice and we infected them with DAV. We did not observed seizures in animals depleted of macrophages, suggesting macrophages are playing a central role in the development of seizures after viral infection.
Role of SLAMF immunoregulatory molecules in MS disease processes
Authors: Mizrahi Jordan S , Park John Jaewon , Masvaker Ruturaj , Bielekova Bibi
Published: January 2023

In our proteomic analysis of cerebrospinal fluid (CSF) we found signaling lymphocyte activation molecules family (SLAMF) upregulated in multiple sclerosis (MS) patients in comparison to controls. The goal of this project is to explore the role of SLAMF molecules in MS.

To identify immune cells that express and secrete SLAMF molecules in the intrathecal (CSF) environment, we performed immunophenotyping of matched blood and CSF leukocytes and dynamic in-vitro experiments, co-culturing leukocytes with EBV and CMV viral lysates. This antigen-specific activation of a sizeable proportion of T and B cells allowed the investigating of changes in surface expression and secretion of SLAMF molecules.

SLAMF2 was broadly expressed, however CSF CD8+ T cells and monocytes had higher SLAMF2 expression in comparison to blood. Stimulation down-modulated SLAMF2 on T cells, but upregulated it on B cells. SLAMF3 was expressed on cells of adaptive, but not innate immunity and CD4+ T cells expressed higher levels of SLAMF3 in CSF than in blood. Upon activation, SLAMF3 expression did not change and the soluble form did not accumulate in supernatants. SLAMF5 was broadly expressed, with particularly higher levels on cells of innate immunity (monocytes and dendritic cells). CSF T and B cells had lower SLAMF5 levels than blood T&B cells. SLAMF6 was strongly expressed on T, B and NK cells; its expression levels did not vary between compartments or upon activation. Finally, SLAMF7 was expressed on cytotoxic T cells, NK cells and plasma cells, with significant upregulation on all antigen-presenting cells upon activation. We explored its role in cytotoxicity, but found no effect of SLAMF7 blockade or sSLAMF7 addition on NK-mediated killing of MHC-I-deficient targets.
Sex-specific regulation of CNS autoimmunity by the signal lymphocytic activation molecule (Slam) locus
Authors: Raza Abbas , Case Laure K , Osmanski Erin , Krementsov Dimitry N , Boyson Jonathan E , Teuscher Cory
Published: January 2023

Multiple sclerosis (MS) is a demyelinating inflammatory disease of the central nervous system (CNS). Epidemiological studies have documented a 3–6 fold increase in MS incidence and prevalence in females over the last half-century, while remaining relatively stable in males. This establishes that gene×environment×sex interactions contribute to MS susceptibility. Genome-wide association studies (GWAS) of MS patients identified multiple candidate genes, including a locus containing the signal lymphocytic activation molecule (SLAM) family of receptors, a region rich in immune-relevant genes that is highly conserved between humans and mice. There are two major Slam haplotypes segregating in laboratory mice. Slam haplotype-1 is present in C57BL/6 (B6) mice and haplotype-2 is expressed in most other common inbred laboratory strains, including 129 mice. The congenic B6.129c1 mouse possesses the 129-derived Slam haplotype-2 locus on the B6 background, which allows for assessing the potential contribution of SLAM family genes to MS susceptibility using the myelin oligodendrocyte (MOG) model of experimental autoimmune encephalomyelitis (EAE). We show that male B6.129c1 congenic mice demonstrate significant protection against MOG-EAE, but only in males. Furthermore, draining lymph nodes from immunized B6.129c1 males, but not females, had increased Foxp3+ regulatory T cells (Treg) and IL-10 levels compared with B6, suggesting that SLAM/Slam haplotypes regulate autoimmunity by modulating the generation of immunoregulatory cells in a sex-specific manner. Taken together, our results support a role for the SLAM locus is MS pathogenesis, and reveal a novel sexual dimorphism in the genetic control of this autoimmune disease
A role for the NLR family members NLRC4 and NLRP3 in sterile inflammasome activation in glial cells
Authors: Guo Haitao , Freeman Leslie , David Clément N , Brickey Willie J , Jha Sushmita , Ting Jenny P
Published: January 2023

The inflammasome is implicated in many inflammatory diseases including neurological diseases but has been primarily studied in the macrophage-myeloid lineage. Here we demonstrate a physiologic role for nucleotide-binding domain, leucine-rich repeat, CARD domain containing proteins, NLRC4 and NLRP3, in brain astrocytes and microglia under the sterile neuroinflammation condition. In vitro, lysophosphatidylcholine (LPC), a molecule associated with neurodegeneration and demyelination, elicits an inflammasome activation dependent on both NLRC4 and NLRP3 in microglia and astrocytes, which are central players in neuroinflammation. Canonical inflammasome components ASC and caspase-1 but not non-canonical inflammasome component caspase-11 is required for LPC-activated inflammasome. Mechanistically, LPC-activated inflammasome also requires cathepsin B protease, calcium mobilization and potassium efflux. In an in vivo cuprizone model of neuroinflammation and demyelination, we found only mice lacking both Nlrc4 and Nlrp3 genes show the significant reduction in astrogliosis and microglial accumulation accompanied by decreased expression of the LPC receptor, G2A, while multiple sclerosis (MS) patient samples show increased G2A. These results reveal that both NLRC4 and NLRP3 are involved in activating LPC-induced inflammasome and are important in astrogliosis and microglial accumulation in a neurological disease model.
Biological evaluation of the MILLIPLEX® map Mouse High Sensitivity T Cell Panel
Authors: Keith Robert H , Grana Paula , Schultz Christian , Bowdish Dawn M.E. , Xiao Qiang
Published: January 2023

Low levels of inflammation are involved in many clinical and sub-clinical disease states like autoimmune diseases. Measuring picogram levels of cytokines is critical for understanding their pathogenesis. Model organisms, such as mice, offer unique challenges of sample availability and detection of low levels of cytokines. We recently developed a Mouse High Sensitivity T Cell Panel multiplex assay for simultaneous measurement of 18 mouse cytokines using Luminex xMAP® technology. In this study, we analyzed mouse cytokine secretions both in vitro, with PMA-, PHA-, LPS-, Con-A, or calcium ionophore-challenged PBMC samples, and in vivo, using LPS-challenged mice and an aged-mouse model. Typically, the response of mouse PBMCs to in vitro stimulants is difficult to study due to low levels of cytokine secretion. Using the MILLIPLEX® map Mouse High Sensitivity T Cell Panel, distinctly different stimulant-dependent cytokine responses were observed. PMA and LPS induced the secretion of IL-1β, IL-5, IL-10, LIX, MCP-1, MIP-2 and LPS. IL-6 was secreted in response to LPS but not PMA. Th17 cell cytokine IL-17A was secreted in response to PHA and Con A but not LPS. Interestingly, low levels of secretion were seen in response to the combination of all stimulants for GM-CSF, IFNγ, IL-1α and IL-2. The in vivo challenge of LPS significantly increased mouse plasma levels of fourteen cytokines. Small increases were also seen for GM-CSF and IL-2, while IL-5 and IL-12 (p70) trended higher. In a mouse model of aging, sex-specific changes of plasma levels were observed for IL-1, IL-5, IL-10, GM-CSF, IFNγ, KC and MIP-2. Thus, the MILLIPLEX® MAP Mouse High Sensitivity T Cell Panel is a useful tool to study low levels cytokines in mouse models of disease and inflammation.
c-Maf positively regulates IL-17-producing iNKT cell-mediated airway neutrophilia
Authors: Yu Jhang-Sian , Hamada Michito , Ohtsuka Shigeo , Yoh Keigyou , Takahashi Satoru , Miaw Shi-Chuen
Published: January 2023

c-Maf belongs to the large Maf family transcription factors and plays a key role in regulation of cytokine production and differentiation in TH2, TH17, TFH and Tr1 cells. Invariant natural killer T (iNKT) cells can rapidly produce large quantity of TH-related cytokines such as IFN-γ, IL-4 and IL-17A upon stimulation by its glycolipid antigen α-galactosylceramide (α-GalCer). However, the role of c-Maf in iNKT cells and iNKT cell-mediated diseases remains poorly understood. In this study, we demonstrate that α-GalCer-stimulated iNKT cells express c-Maf transcript and protein. By using c-Maf-deficient fetal liver cell-reconstituted mice, we further show that c-Maf-deficient iNKT cells produce less IL-17A than their WT counterparts after α-GalCer stimulation, but IFN-γ and IL-4 productions were not affected. c-Maf deficiency does not affect the development and activation state of iNKT cells. c-Maf also positively regulates induction of IL-17-producing iNKT (iNKT17) cells by IL-6, TGF-β and IL-1β. c-Maf-deficient iNKT cells express lower RORγt transcripts compared with WT iNKT cells under IL-17-induction condition. Interestingly, c-Maf-deficient iNKT17 cells lose the ability to recruit neutrophils into the lungs. Taken together, c-Maf is a positive regulator for IL-17A production through transactivating RORγt expression in iNKT17 cells. It is a potential therapeutic target in iNKT17 cells-mediated inflammatory disease.
Unexpected STAT3-mediated mechanisms of regulation of effector Th17 function in autoimmunity
Authors: Raphael Itay , McGeachy Mandy J
Published: January 2023

Th17 cells have been linked to the emergence of many autoimmune inflammatory conditions as well as defense against fungal pathogens. Early differentiation and expansion of Th17 cells relies on key cytokines that signal through STAT3, including IL-6 and IL-21. Later STAT3-dependent signals from IL-23 are critical for gain of inflammatory function by effector Th17 cells. Thus, development of drugs that target STAT3 or its upstream receptors are a major research focus to alleviate disease. However, the requirements for STAT3 signaling in effector Th17 cells remain elusive. Here, we developed a model to study the effect of STAT3 deficiency specifically in effector Th17 cells in EAE, an animal model of multiple sclerosis. Our data show that mice lacking STAT3 in effector Th17 cells are resistant to EAE development. Unexpectedly, we demonstrate that in Th17 cells, STAT3 is not required for maintenance of Th17 lineage commitment or Th17 cytokine production in vivo. To determine mechanisms by which STAT3 regulates Th17 effector cells to promote development of autoimmune inflammation, we performed bioinformatics analysis of the transcriptome of effector Th17 cells isolated from lymph nodes (LN) at day 10 (onset) of EAE. This revealed that STAT3 regulates expression of genes associated with particular cell-death pathways, and pathways associated with proliferation and T cell mediated tissue damage. Corresponding to these changes in gene expression, we found that STAT3 deficient effector Th17 cells were reduced in numbers in LN and target tissue during EAE. Our data reveal a hitherto unappreciated mechanism for STAT3 in regulating effector Th17 cells, which have major implications for use of STAT3-targeted therapies.
PD-L1 from HaCaT cells affect melanin production in B16F10 cells under poly(I:C) stimulation
Authors: Woo So-Youn , Park Minhwa , Cho Kyung-Ah , Cho Min-Sun , Lee Kyung Ho
Published: January 2023

Skin form the physical barrier protected from the stimulants outside. Keratinocytes can sense either harmless commensal organisms or harmful pathogens, and subsequently drive immune response by Toll-like receptors (TLRs). For example, activation of TLR3 has been shown to require barrier repair by stimulate essential genes, including tight junction genes and inflammatory cytokines. Melanocytes reside on the basal layer cells and connect with about 30–40 keratinocytes in the basal layer through dendritic morphology. Most observations refer to transfer of melanosome into keratinocytes but melanogenesis regulator mechanism by soluble factor(s) from keratinocytes is unclear. Upon consideration of the cellular location in the skin and ratio of residing keratinocytes to melanocytes in the epidermis, regulation of melanogenesis by controlling keratinocytes in vivo is challenging. In this study, we investigated whether HaCaT affect melanogenesis under TLR3 stimulation using B16F10 cells, and found that poly(I:C) stimulation induced PD-L1 secretion from HaCaT cells and PD-L1 inhibited expression of melanogenesis-related genes on B16F10 cells.

* This work was supported by the Basic Science Research Program through the National Research Foundation of Korea (NRF-2016R1A2B4009182).
Intestinal Regulation of C. albicans by PP Langerin+ DCs
Authors: De Jesus Magdia , Rochac-Argueta Emily , Gallagher Heather
Published: January 2023

As the largest mucosal compartment, the human gastrointestinal tract (GI) is highly colonized with a complex microbial ecosystem that includes fungal organisms. Although fewer in numbers and less extensively studied in the context of the intestinal mucosa, it is unclear whether commensal members such as Candida species play a role in intestinal mucosal immunity. Recent studies have suggested that Candida sp. in the intestinal mucosa are associated with a number of human diseases such as gastric ulcers, Crohn’s disease and Hirschsprung-associated enterocolitis. We have recently identified, a specific non-macrophage dendritic cell (DC) population within intestinal Peyer’s patches (PP) that expresses the C-type lectin receptor (CLR)-Langerin and are able to specifically uptake Candida albicans, Candida tropicalis and yeast purified β-1,3-glucan particles (GPs). We hypothesize that Langerin+ DCs are central in the sampling, presentation and regulation of C. albicans within PPs. To address this hypothesis, we are currently defining the role of Langerin+ DCs in the processing of Candida sp. after uptake using the Langerin-DTR- EGFP mouse model as well as the Langerin-DTR- EGFP-DSS model. We are also characterizing the local gene expression profiles of PP B and T- lymphocytes as well as Langerin+ DCs in response to C. albicans. Additionally, we are investigating whether Langerin+ DCs are a migratory cell subset that not only transports C. albicans near germinal centers but can also take them to tolerogenic sites such as the MLN. These studies will advance our knowledge of how commensal members that are also part of the “mycobiome” are sampled, what mucosal immune responses are elicited and how these fungal organisms are tolerated.
miR155-deficient alleviated intestinal inflammation by downregualting the development and function of CD103+CD11b+DC in the lamina propria of intestine
Authors: Xu Yingping , Zhang Binbin , Zhu Xiaoli , Li Chuanwei , Song Wengang , Tang Hua
Published: January 2023

Inflammatory bowel disease (IBD) refers to inflammatory disorders of the gut with unclear pathogenesis. miR155 expression was increased in IBDs by upregulated Th1/Th17 cell response and proinflammatory factors to promote intestinal inflammation. Dendritic cell (DC) is a heterogeneous subsets in the intestine, and plays important roles in maintaining intestinal homeostasis. However, it is unclear whether miR155 regulate the differentiation and function of intestinal DC subsets to orchestrate inflammation in the intestinal microenvironment. Here, we found defiency of miR155 led to a significantly decreased number of CD103+CD11b+DC in the lamina propria (LP) of intestine at steady state, moreover, the yield of bone marrow-derived CD103+CD11b+DC induced by GM-CSF and Flt3L in vitro was also decreased compared to that of wild type (WT) mice. Further study showed that differency of miR155 had no effect on turnover rate and migration. Interestingly, we found the percentage and number of pre-cDC1 (CD103+DC precursor) in the BM were reduced significantly in the bone marrow of miR155-deficient mice compared to that of WT mice, and the transcriptor IRF4 and Notch2 expression of CD103+CD11b+DC in the LP was also decreased in miR155-deficient mice. In addition, CD103+DC and CD103+CD11b+DC in the LP of miR155-deficient mice exhibited a poor pro-inflammatory phenotype. Taken together, our results identifies a role of miR155 as a critical regular of intestinal homeostasis, which will be helpful for seeking a potential therapeutic target for the treatment of IBD.
Sensory neurons co-opt classical immune signaling pathways to mediate chronic itch
Authors: Oetjen Landon K , Mack Madison R , Whelan Timothy M , Guo Changxiong J , Yang Lihua , Hamilton Samantha L , Wang Peter L , Niu Haixia , Feng Jing , Xu Amy Z , Tripathi Shivani V , Luo Jialie , Brestoff Jonathan R , Schaffer Andras , Hsieh Chyi-Song , Gereau Robert W , Miller Mark J , Hu Hongzhen , Davidson Steve , Liu Qin , Kim Brian S
Published: January 2023

Mammals have evolved neurophysiologic reflexes such as scratching to expel invading pathogens and noxious environmental factors. It is also well established that these responses are associated with chronic inflammatory diseases such as atopic dermatitis. However, the mechanisms by which inflammatory pathways promote sensations such as itch remain poorly understood. Here, we show that the type 2 cytokines interleukin (IL)-4 and IL-13 directly stimulate sensory neurons and that chronic itch is dependent on neuronal IL-4Rα, the shared receptor subunit for IL-4 and IL-13. Based on our understanding of IL-4Rα signaling in immune cells, we hypothesized that downstream Janus kinase (JAK) signaling in sensory neurons would be a critical mediator of itch. Indeed, we find that both systemic JAK inhibition and interruption of JAK signaling in the nervous system can dramatically reduce itch. Further, sensory neuron-specific genetic deletion of JAK1 results in robust abatement of chronic itch in mice. Finally, in proof-of-concept translational studies, patients with recalcitrant chronic itch improved rapidly and dramatically in response to JAK inhibition. Thus, signaling mechanisms previously ascribed to the immune system may represent novel targets within the sensory nervous system for the treatment of pathologic sensory responses. Collectively, these studies reveal an evolutionarily conserved paradigm in which the sensory nervous system employs classical immune signaling pathways to influence mammalian behavior.
Complement protein C1q suppresses macrophage inflammasome activation during clearance of modified LDL
Authors: Manughian-Peter Ayla , Ho Minh Minh , Fraser Deborah A.
Published: January 2023

In the atherosclerotic environment, inflammation plays a major role. Ingestion of modified forms of low-density lipoproteins (such as oxidized LDL) by macrophages promotes macrophage secretion of inflammatory cytokines, which contributes to disease progression. We have previously shown that the complement classical pathway activator, C1q, has a novel, anti-inflammatory role independent of the complement cascade: opsonization of modified LDL with C1q polarizes macrophages towards an anti-inflammatory (M2) phenotype, and dampens the pro-inflammatory (M1) phenotype that is prevalent in the atherosclerotic environment. Differential transcriptomic analysis in human macrophages suggests that C1q may influence macrophage inflammatory phenotype by modulating the levels of components of the NLRP3 inflammasome, a protein complex responsible for activation of inflammatory cytokine IL-1β. To investigate this further, resting or M1-polarized human monocyte derived macrophages (HMDMs) were treated with 10 μg protein/mL oxLDL, with or without 75 μg/mL C1q, and inflammasome gene, protein and activity were measured. Our data suggest that when bound to oxLDL, C1q downregulates NLRP3 inflammasome related genes, and reduces inflammasome activity, leading to a reduction in IL-1β secretion from M1-polarized macrophages. This indicates that this non-complement role of C1q may be important in dampening inflammation in the early atherosclerotic lesion. Discerning the molecular pathways that regulate inflammatory states of macrophages can contribute to the development of therapeutics for chronic inflammatory illnesses such as atherosclerosis.
Elevated Prostaglandin E2 Post-Bone Marrow Transplant Mediates Interleukin-1β Related-Lung Injury
Authors: Moore Bethany B , Martinez-Colon Giovanny J. , Taylor Quincy M , Wilke Carol A , Podsiad Amy
Published: January 2023

Hematopoietic stem cell transplantation (HSCT) treats hematological and inherited disorders. Unfortunately, patients that undergo HSCT become susceptible to infections by a wide array of opportunistic pathogens. The bacterial pathogen Pseudomonas aeruginosa can have life threatening effects in HSCT patients. In the lung, P. aeruginosa triggers the secretion of Interleukin-1β (IL-1β) by alveolar macrophages (AMs) and induces features of lung injury while inhibition of IL-1β or its receptor signaling has been shown to improve outcomes. In the present study, we used a mouse model of bone marrow transplantation (BMT) and found that high levels of prostaglandin E2 (PGE2) secreted by AMs post-BMT exacerbate IL-1β-mediated pulmonary injury post-P. aeruginosa instillation. We determined that the intracellular signaling pathway by which PGE2 induces IL-1β involves EP2/EP4 signaling with subsequent cAMP increase, PKA/Epac, and CREB transcription factor activation. Immature IL-1β can then be processed via either canonical or non-canonical inflammasomes (caspase-1 and caspase 8). Furthermore, PGE2 can induce IL-1β release despite the ability of PGE2 to limit autophagy, a pathway known to regulate inflammasome accumulation and bacterial killing. Interestingly, PGE2 inhibition in BMT mice decreases bacterial load, lung injury, and IL-1β in the lung. These findings can inform new therapeutic strategies that can improve outcome of HSCT patients with pulmonary complications due to P. aeruginosa, and possibly other pathogens.
Specific role of IL-1β in instigating skin inflammation in Sharpin-deficient mice
Authors: Sharma Bhesh Raj , Gurung Prajwal , Kanneganti Thirumala-Devi
Published: January 2023

Genetically inherited autoinflammatory skin disorders are characterized by spontaneous skin inflammation. Mutations in the members of linear ubiquitin chain assembly complex (LUBAC) are associated with spontaneous skin inflammation. Mice, deficient in SHARPIN (Sharpincpdm mice), a member of LUBAC, develop severe dermatitis associated with systemic autoinflammation. Previous studies have demonstrated that NLRP3 inflammasome provoke skin inflammation in Sharpin-deficient mice. However, whether IL-1α or IL-1β instigates disease and, if skin inflammation precedes systemic autoinflammation is not known. Here, we have performed thorough cellular analysis of pre-diseased and diseased Sharpin-deficient mice and demonstrated that systemic inflammation precedes skin inflammation. Furthermore, we demonstrate a specific role for IL-1β, but not IL-1α in instigating disease in Sharpincpdm mice. Our results altogether demonstrate distinct roles of SHARPIN in initiating systemic autoinflammation and skin dermatitis. Furthermore, skin inflammation in Sharpincpdm mice is specifically mediated by IL-1β, highlighting the importance of specific targeted therapies in IL-1 signaling blockade.
Characterizations of mucosal associated invariant T cells in alcoholic liver disease
Authors: Sa Moa , Kim Sojeong , Kim Won , Park Su-Hyung
Published: January 2023

The liver is a critical immunological site for chronic alcohol consumption as it filters various antigens activating both innate and adaptive responses. The pathogenesis of alcoholic liver disease (ALD) in human is not clear in terms of the specific role of immune cell population. Mucosal associated invariant T (MAIT) cells are found to be abundant in the mucosal organs, especially within the liver organ in which the cells comprise up to 50 % of T cells. Here, we show that the percentage of circulating MAIT cells is severely reduced in ALD patients (p=0.0028), including patients with steatohepatitis (ASH, n=20), steatohepatitis with cirrhosis (ASHAC, n=15) and cirrhosis (AC, n=12), compared to age-matched healthy donors (HD, n=8). Moreover, compared to non-liver disease group, MAIT cells are depleted among liver infiltrating lymphocytes in ASH (p=0.0276) and ASHAC (p=0.0049) patients, whereas that of AC group is recovered. Although the percentage of MAIT cells is decreased, the cells display increased expression level of activation markers. Pro-inflammatory cytokines such as IL-17A and MIP-1 beta are up-regulated in ASH and ASHAC patients by PMA/Ionomycin stimulation. In addition, Perforin and Granzyme B, which are cytotoxic markers in granule, are significantly increased in both peripheral blood and liver of ASHAC group. Taken together, our results suggest that activated MAIT cell population in the liver may contribute to the pathogenesis of ALD.
The xenobiotic transporter Mdr1 permits T cell adaptation to mucosa-associated bile acids in the ileum
Authors: Sundrud Mark S , Cao Wei , Kayama Hisako , Chen Mei Lan , Delmas Amber , Sun Amy , Kim Sang Yong , McKevitt Kelly , Rangarajan Erumbi S. , Beck Amanda P. , Oikonomopoulos Angelos , Lacey Precious N. , Martinez Gustavo J , Izard Tina , Lorenz Robin G , Rodriguez-Palacios Alex , Cominelli Fabio , Abreu Maria T , Hommes Daniel W. , Koralov Sergei B , Takeda Kiyoshi
Published: January 2023

Intestinal CD4+ T helper (TH) cells are subject to extensive regulation by microbiota. By contrast, it is not known whether or how TH cells interface with other, host-derived intestinal metabolites. Here we show that bile acids directly regulate mucosal TH cell function in the distal small intestine (i.e., ileum) via the xenobiotic transporter, Mdr1. Using both Mdr1-dependent dye efflux and a novel CRISPR-generated Mdr1 reporter mouse, we show that wild type RORγt+IL-17A+ (Th17) and RORγt-IFNγ+ (Th1) cells upregulate Mdr1 expression upon migration into the ileum. By contrast, germline ablation or shRNAmir-mediated knockdown of Mdr1 in Th17 and Th1 cells results in local dysfunction in the ileum, and these cells transfer Crohn’s disease-like ileitis in Rag1−/− hosts. Mdr1 enforces Th17 and Th1 cell survival and limits pro-inflammatory cytokine (TNFα, IFNγ) expression in the presence of conjugated bile acids (CBAs), which are actively reabsorbed through the ileal mucosa as a function of enterohepatic bile acid circulation. Accordingly, genetic or pharmacologic blockade of ileal CBA reabsorption restores Mdr1-deficient Th17 and Th1 cell homeostasis in ilea of transferred Rag1−/− hosts and rescues ileitis. In addition, MDR1 loss-of-function is evident in both ileitis-prone (SAMP1/YitFc) mice, and a subset of ileal Crohn’s disease patients. These data indicate that coordinated, local and druggable interactions between mucosal TH cells and mucosa-associated bile acids in the ileum contribute to intestinal immune homeostasis.
Contraction of intestinal effector T cells by retinoic acid-induced purinergic receptor P2X7
Authors: Hashimoto-Hill Seika , Friesen Leon , Kim Myunghoo , Kim Chang H
Published: January 2023

Vitamin A is a key regulator of immunity and immune tolerance. Here we report that the vitamin A metabolite retinoic acid regulates the population size of intestinal effector T cells by inducing purinergic receptor P2X7 expression. RA induces P2X7 expression by direct binding to an intragenic enhancer of the P2rx7 gene in CD4+T cells, rendering them highly susceptible to apoptosis triggered by P2X7 activation in response to extracellular ATP and NAD. Utilizing P2X7-null mice, we found that this tolerogenic mechanism is critical for reining in the expansion of Th1 and Th17 effector T cell populations in the intestine where the concentrations of extracellular ATP/NAD and retinoic acid are high. We demonstrated that P2X7 activation can ameliorate colitis by inducing the apoptosis of inflammatory effector T cells. We conclude that the RA-P2X7 axis is a novel immune homeostatic mechanism by which the size of potentially inflammatory effector T cell populations is tightly controlled to prevent inflammatory diseases.
Short chain fatty acids differentially regulate differentiation and function of Th1 and Th17 cells in the intestines
Authors: Chen Liang , Sun Mingming , Wu Wei , Chen Feidi , Huang Xiangsheng , Xiao Yi , Yao Suxia , Cong Yingzi
Published: January 2023

The mechanisms by which gut microbiota regulate host immune responses are still not completely understood. Short chain fatty acids (SCFA), mainly acetate (C2), propionate (C3), and butyrate (C4), the metabolites produced solely by gut microbiota, have been shown to promote development of Treg cells, and possibly other T cell subsets, in the intestines. However, the mechanisms involved remain unclear. In this report, we investigated how SCFA regulate development and function of microbiota antigen-specific T cells. We treated CBir1 transgenic T cells, specific for an immunodominant microbiota antigen CBir1 flagellin, with SCFA under various T cell polarization conditions to determine how they regulate T cell differentiation, and transferred those T cells into RAG−/− mice to determine their function in induction of colitis. Our results demonstrated that C2, C3 and C4 all promoted T cell production of IFN-γ and IL-10 when cultured under neutral and Th1 conditions. Under Th17 conditions C2, C3 promoted, while C4 inhibited, T cell IL-17 production at early stage of cell divisions. However, all three SCFA promoted T cell IL-10 production. Mechanistically, SCFA activation of mTOR pathway but not inhibition of HDAC or binding GPR43 mediated their effects on Th1 and Th17 cells. Functionally, transfer of C2- and C4-treated Th1 and Th17 cells induced less severe colitis in RAG−/− mice compared to untreated T cells, probably through induction of IL-10 by those T cells. Collectively, our study indicated that SCFA differentially regulate T cells differentiation and function in the intestines, which could contribute the maintenance of intestinal homeostasis.
IL-33 regulates the IgA-microbiota axis to restrain IL-1α dependent colitis and tumorigenesis
Authors: Malik Ankit , Sharma Deepika , Zhu Qifan , Karki Rajendra , Guy Clifford , Vogel Peter , Kanneganti Thirumala-Devi
Published: January 2023

Inflammatory bowel diseases (IBD) affect over 5 million individuals in the industrialized world, with an increasing incidence rate worldwide. IBD also predisposes affected individuals to development of colorectal cancer, which is a leading cause of cancer-related deaths in adults. Mutations in genes encoding molecules in the IL-33 signaling pathway are associated with colitis and colitis-associated cancer (CAC), but how IL-33 modulates gut homeostasis is unclear. Here, we show that Il33-deficient mice are highly susceptible to colitis and CAC. IL-33 promotes IgA production in the intestine, which is important for maintaining gut microbial homeostasis. Consequently, Il33-deficient mice develop a dysbiotic microbiota characterized by increased level of mucolytic and colitogenic bacteria. In response to DSS administration, this microbial landscape promotes the release of IL-1α, which acts as a critical driver of colitis and CAC. Consequently, reconstitution of symbiotic microbiota or IL-1α ablation markedly ameliorates colitis susceptibility in Il33−/− animals. Our results demonstrate that IL-33 promotes IgA production to maintain gut microbial homoeostasis, and restrain IL-1α dependent colitis and CAC. This study therefore highlights modulation of IL-33, IgA, IL-1α and the microbiota as potential therapeutic targets in the treatment of IBD and CAC.
Effect of TGF-β on Antigen-specific CD8+ T cell Response in Colitis
Authors: Demel Erika L , Mishra Shruti , Liu Yong , Liu Xun , Ma Chaoyu , Dube Peter , Zhang Nu
Published: January 2023

Inflammatory Bowel Disease (IBD) is a chronic inflammatory disease of the gastrointestinal tract primarily including Crohn’s Disease (CD) and Ulcerative Colitis (UC). IBD affects 1.6 million Americans, and the worldwide incidence of IBD is growing in areas where the disease was previously infrequent. Currently, genetics, intestinal microbiota, environment, and dysregulation of the intestinal immune response are thought to be responsible for the development of IBD. Increased understanding of the immune response in IBD can be used to develop and improve treatments for patients. Much work has been done on the role of CD4+ T helper cells in IBD. The role of CD8+ T cells has been less heavily investigated despite noted differences in CD8+ T cells between UC and CD patients and healthy subjects.

In order to follow antigen-specific CD8+ T cell response during IBD, we developed a mouse strain in which the expression of model antigen Ova was specifically induced in the large intestine epithelium upon tamoxifen treatment (i.e. CDX2-cre/ERT2 Rosa26-Ova mice). By examining adoptively transferred OT-1 T cells, large intestine-specific CD8+ T cell response was determined during DSS induced colitis. Interestingly, the response of control OT-1 T cells was largely restricted to the large intestine while TGF-β unresponsive OT-1 T cells underwent dramatic expansion at both gut-associated and systemic sites. The functional consequence of TGF-β-controlled expansion of CD8+ T cells in colitis is currently under investigation.
Targeted delivery of microbial metabolite, urolithin A protects from chemically (DSS or TNBS) induced colitis in pre-clinical models
Authors: Singh Rajbir , Hegde Bindu , Von Baby Becca , Sadeep C , Kotla Niranjan , Chandrasekar Bhargavi , Marepally Srujan , Bodduluri Haribabu , Vemula Praveen K , Jala Venkatakrishna R
Published: January 2023

Epidemiological data suggests that consumption of diets rich in phytochemicals are protective in inflammatory bowel diseases (IBDs). However, the beneficial effects are not uniform among individuals and attributed to variations in gut microbiota, and altered capacity to generate certain metabolites. Urolithin A (UroA) (3,8-dihydroxybenzo[c]chromen-6-one) is a microbial metabolite, derived from ellagic acid and ellagitannins, major poly phenolic components in berries and pomegranates. Here, we examined therapeutic applications of UroA and mechanisms of action in IBDs. Our studies suggested that UroA significantly reduced LPS induced inflammatory mediators (e.g., IL-6, TNF-alpha and IL-12) as well as ROS production in mouse bone marrow-derived macrophages (BMDMs), dendritic and THP1 cells. Most importantly, UroA also reduced LPS induced systemic inflammation in mouse models. Next, we examined therapeutic applications of UroA in dextran sodium sulphate (DSS)-induced, 2,4,6-Trinitrobenzenesulfonic acid (TNBS)-induced colitis models. Treatment with UroA significantly reduced both acute and chronic DSS-induced colitis as well as TNBS induced colitis in mouse models. Most importantly, delivery of UroA utilizing inflammation targeting oral nano-particles (single treatment regimen) effectively mitigated the colitis in both models. In summary, these results highlight even presence of single microbial metabolite at right location at right time will have significant beneficial effects to protect from adverse inflammatory activities.
Canonical Wnt Signaling in CD11c+ antigen presenting cells controls inflammation-associated colon carcinogenesis and commensal homeostasis
Authors: Swafford Daniel J , Shanmugam Arulkumaran , Ranganathan Punithavathi , Manicassamy Santhakumar
Published: January 2023

The intestine contains high levels of the Wnt family of ligands, and the Wnt-signaling pathway is essential for intestinal homeostasis. Thus, aberrant Wnt/β-catenin-signaling may result in inflammatory bowel disease (IBD) and IBD-associated colon carcinogenesis. Although past studies have focused on this signaling pathway’s role in intestinal stem cell proliferation and epithelial cell maintenance, its role in shaping mucosal immune responses and commensal homeostasis in the gut remains unknown. In the present study, using a mouse model of colitis-associated colorectal cancer, we demonstrated that canonical Wnt-signaling in intestinal CD11c+ antigen presenting cells (APCs) controls intestinal inflammation and suppresses inflammation-driven colon carcinogenesis by imparting an anti-inflammatory phenotype. Genetic deletion of Wnt co-receptors, low-density lipoprotein receptor-related protein 5 and 6 (LRP5/6), in CD11c+ APCs in mice (LRP5/6ΔAPC mice) caused enhanced intestinal inflammation with increased tumor numbers in the colon due to increased production of pro-inflammatory cytokines and decreased expression of immune regulatory factors resulting from commensal dysbiosis. Mechanistically, loss of LRP5/6-mediated signaling in intestinal APCs resulted in enhanced effector T cell differentiation and decreased regulatory T cell differentiation. Furthermore, our study demonstrates that in LRP5/6ΔAPC mice conditional activation of β-catenin in intestinal APCs or in vivo blockade of IL-6 results in reduced intestinal inflammation with decreased colon tumor burden. These results reveal a mechanism by which intestinal APCs control gut inflammation and commensal homeostasis via canonical Wnt signaling.
The effects of obesity on the efficacy of immunotherapy.
Authors: Dudzinski Stephanie O , Beckermann Kathryn E , Rathmell Jeffrey C , Giorgio Todd D
Published: January 2023

Immunotherapy is a revolutionary treatment that harnesses the immune system to fight cancer. However, only a small fraction of patients responds to these treatments. Differences in an individual’s immune system and/or the tumor microenvironment may explain why immunotherapy is effective in only some patients. Obesity is an important cancer risk factor that is known to impair protective immunity, which may impact the effectiveness of immunotherapy. However, the impact of obesity on immunotherapy efficacy has never been investigated. We thus hypothesized that obesity reduces immunotherapy efficacy.

We studied the effects of obesity on the tumor microenvironment using a diet-induced obesity mouse model and immunoresponsive cell line. MC38 colon cancer cells were injected subcutaneously in C57BL/6 mice fed a high-fat or control diet. The tumors were collected three weeks post-injection and processed into single cell suspensions for flow cytometry. Samples were stained for helper T cells, cytotoxic T cells, and for macrophages with CD11b and F4/80, where CD206 and CD86 identified M2 and M1 macrophages, respectively. Initial experiments revealed a higher concentration of cytotoxic T cells in lean mice, which matches studies conducted in different cell lines. We will investigate the role of immune cell and cytokine differences further. In future experiments, we have engineered biocompatible nanoparticles that can modulate immune cells and shift the tumor microenvironment using siRNA, potentially improving immunotherapy efficacy in obesity. These initial studies will lay the foundation for future research and have important translational implications in improving the efficacy of immunotherapy treatment in obese patients.
Commensal dysbiosis alters the tumor microenvironment in luminal breast cancer
Authors: Buchta Claire M , Bostic Raegan R , Kearney John E , Rutkowski Melanie R
Published: January 2023

Many factors, including diet, race, and recurrent use of antibiotics, have been associated with an adverse outcome in breast cancer. These extrinsic factors also significantly impact the composition of commensal microbes. This is relevant, as a recent clinical trial demonstrated that patients with advanced hormone receptor+breast cancer had commensal dysbiosis, or significantly reduced composition of commensal microbes, supporting previous studies demonstrating that commensal microbes influence both the systemic immune environment and breast cancer progression. Dysbiosis was additionally shown to promote liver fibrosis in a mouse model of nonalcoholic fatty liver disease. As collagen deposition in mammary tissue is known to associate with breast cancer aggressiveness and metastasis, we hypothesized that dysbiosis enhances metastatic disease in luminal breast cancer through systemic changes to the immune environment, ultimately altering the extracellular matrix of tumors and mediating metastatic disease. To investigate this, we utilized a transplantable model of luminal HR+breast cancer where tumor fragments are transplanted into recipients with or without established commensal dysbiosis. We demonstrate that although the growth of primary tumors is not affected, commensal dysbiosis increases collagen deposition and macrophage infiltration in transplanted tumor fragments. Systemically, dysbiosis amplifies the production of CCL2, a chemokine associated with macrophage infiltration and metastatic potential. These data suggest that commensal dysbiosis enhances collagen deposition in breast tumors and influences the tumor microenvironment, potentially promoting metastatic disease.
Investigating the molecular mechanism of CRISPR-Cas adaptation of Streptococcus thermophilus
Authors: Louis Edwin Pierre , Wei Yunzhou , Terns Michael P
Published: January 2023

Bacteria are constantly challenged by viral infection. In order to combat potentially lethal viruses, bacteria possess adaptive immune defenses called CRISPR-Cas (Clustered Regularly Interspaced Short Palindromic Repeats-CRISPR-Associated) systems. These systems are RNA-based defense systems that memorize past viral infections to protect prokaryotes like bacteria from future infections. The acquisition of short DNA sequences (spacers) from previously encountered invaders allows bacteria to generate immunity. These systems capture spacers and incorporated them into their CRISPR loci to initialize immunity. Once integrated, spacers are then transcribed and processed into small RNA molecules called crRNAs, which are eventually bound by Cas proteins and form targeting complexes capable of cleaving viral DNA to achieve immunity. Genetic evidence indicates that Cas1, Cas2, Cas9 and Csn2 are each essential for CRISPR adaptation in the Type II-A system of Streptococcus thermophilus. However, the role of each of these proteins in adaptation is not clear. We have systematically constructed single cas gene deletion strains. We aim to detect and analyze low level of adaptation events in these genetic backgrounds, and in turn gain insight into the specific roles of each of these proteins. We also plan to apply biochemical approaches using purified proteins to investigate the molecular mechanistic steps of adaptation.