Elevated Expression Of IL-1β Research Articles

Dysregulated Airway Host Defense in Hyper IgE Syndrome due to STAT3 Mutations.

Hyper IgE syndrome (STAT3-HIES), also known as Job's syndrome, is a rare immunodeficiency disease typically caused by dominant-negative STAT3 mutations. STAT3-HIES syndrome is characterized by chronic pulmonary infection and inflammation, suggesting impairment of pulmonary innate host defense. To identify airway epithelial host defense defects consequent to STAT3 mutations that, in addition to reported mutant STAT3 immunologic abnormalities, produce pulmonary infection. STAT3-HIES sputum was evaluated for biochemical/biophysical properties. STAT3-HIES excised lungs were harvested for histology; bronchial brush samples were collected for RNA sequencing and in vitro culture. A STAT3-HIES-specific mutation (R382W), expressed by lentiviruses, and a STAT3 knockout, generated by CRISPR/Cas9, were maintained in normal human bronchial epithelia under basal or inflammatory (IL1β) conditions. Effects of STAT3 deficiency on transcriptomics, and epithelial ion channel, secretory, antimicrobial, and ciliary functions were assessed. Mucus concentrations and viscoelasticity were increased in STAT3-HIES sputum. STAT3-HIES excised lungs exhibited mucus obstruction and elevated IL1β expression. STAT3 deficiency impaired CFTR-dependent fluid and mucin secretion, inhibited expression of antimicrobial peptides, cytokines, and chemokines, and acidified airway surface liquid at baseline and post-IL1β exposure in vitro. Notably, mutant STAT3 suppressed IL1R1 expression. STAT3 mutations also inhibited ciliogenesis in vivo and impaired mucociliary transport in vitro, a process mediated via HES6 suppression. Administration of a γ-secretase inhibitor increased HES6 expression and improved ciliogenesis in STAT3 R382W mutant cells. STAT3 dysfunction leads to multi-component defects in airway epithelial innate defense, which, in conjunction with STAT3-HIES immune deficiency, contributes to chronic pulmonary infection.

Melatonin counteracts polyethylene microplastics induced adreno-cortical damage in male albino rats

There are various substances that can disrupt the homeostatic mechanisms of the body, defined as endocrine-disrupting chemicals (EDCs). The persistent nature of microplastics (MPs) is a cause for concern due to their ability to accumulate in food chains and widespread use, making their toxic effects particularly alarming. The potential of MPs for disrupting the endocrine system was observed in multiple tissues. Moreover, the adrenal gland is known to be extremely sensitive to EDCs, while with the effect of MPs on the adrenal gland has not previously been studied. This study aimed to highlight the potential polyethylene microplastics (PE-MPs) induced adreno-toxic effects rather than exploring the implicated mechanisms and concluding if melatonin (Mel) can afford protection against PE-MPs induced adreno-toxicity. To fulfill the goal, six groups of rats were used; control, Mel, PE-MPs (3.75 mg/kg), PE-MPs (15 mg/kg), PE-MPs (3.75 mg/kg) +Mel, and PE-MPs (15 mg/kg) +Mel. PE-MPs induced toxic changes in the adrenal cortex, which was evident by increased adrenal weight, histopathological examination, and ultrastructural changes detected by electron microscope. A reduction in serum cortisol and an increase in serum adrenocorticotropic hormone resulted from the adreno-toxic effects of PE-MPs. Mechanisms may include the reduction of steroidogenesis-related genes, as PE-MPs drastically reduce mRNA levels of StAR, Nr0b1, Cyp11A1, as well as Cyp11B1. Also, oxidative stress that results from PE-MPs is associated with higher rates of lipid peroxidation and decreased superoxide dismutase and glutathione. PE-MPs inflammatory effect was illustrated by elevated expression of IL-1β and NF-ķB, detected by immunohistochemical staining, in addition to increased expression of caspase-3 and mRNA of Bax, markers of proapoptotic activity. The impacts of PE-MPs were relatively dose-related, with the higher dose showing more significant toxicity than the lower one. Mel treatment was associated with a substantial amelioration of PE-MPs-induced toxic changes. Collectively, this study fills the knowledge gap about the MPs-induced adrenal cortex and elucidates various related toxic mechanisms. It also supports Mel’s potential protective activity through antioxidant, anti-inflammatory, anti-apoptotic, and gene transcription regulatory effects.

Transcriptional profiling of a fungal granuloma reveals a low metabolic activity of Paracoccidioides brasiliensis yeasts and an actively regulated host immune response.

Granulomas are important immunological structures in the host defense against the fungus Paracoccidioides brasiliensis, the main etiologic agent of Paracoccidioidomycosis (PCM), a granulomatous systemic mycosis endemic in Latin America. We have performed transcriptional and proteomic studies of yeasts present in the pulmonary granulomas of PCM aiming to identify relevant genes and proteins that act under stressing conditions. C57BL/6 mice were infected with 1x106 yeasts and after 8- and 12-weeks of infection, granulomatous lesions were obtained for extraction of fungal and murine RNAs and fungal proteins. Dual transcriptional profiling was done comparing lung cells and P. brasiliensis yeasts from granulomas with uninfected lung cells and the original yeast suspension used in the infection, respectively. Mouse transcripts indicated a lung malfunction, with low expression of genes related to muscle contraction and organization. In addition, an increased expression of transcripts related to the activity of neutrophils, eosinophils, macrophages, lymphocytes as well as an elevated expression of IL-1β, TNF-α, IFN-γ, IL-17 transcripts were observed. The increased expression of transcripts for CTLA-4, PD-1 and arginase-1, provided evidence of immune regulatory mechanisms within the granulomatous lesions. Also, our results indicate iron as a key element for the granuloma to function, where a high number of transcripts related to fungal siderophores for iron uptake was observed, a mechanism of fungal virulence not previously described in granulomas. Furthermore, transcriptomics and proteomics analyzes indicated a low fungal activity within the granuloma, as demonstrated by the decreased expression of genes and proteins related to energy metabolism and cell cycle.

18F]GE-180-PET and Post Mortem Marker Characteristics of Long-Term High-Fat-Diet-Induced Chronic Neuroinflammation in Mice.

Obesity is characterized by immoderate fat accumulation leading to an elevated risk of neurodegenerative disorders, along with a host of metabolic disturbances. Chronic neuroinflammation is a main factor linking obesity and the propensity for neurodegenerative disorders. To determine the cerebrometabolic effects of diet-induced obesity (DIO) in female mice fed a long-term (24 weeks) high-fat diet (HFD, 60% fat) compared to a group on a control diet (CD, 20% fat), we used in vivo PET imaging with the radiotracer [18F]FDG as a marker for brain glucose metabolism. In addition, we determined the effects of DIO on cerebral neuroinflammation using translocator protein 18 kDa (TSPO)-sensitive PET imaging with [18F]GE-180. Finally, we performed complementary post mortem histological and biochemical analyses of TSPO and further microglial (Iba1, TMEM119) and astroglial (GFAP) markers as well as cerebral expression analyses of cytokines (e.g., Interleukin (IL)-1β). We showed the development of a peripheral DIO phenotype, characterized by increased body weight, visceral fat, free triglycerides and leptin in plasma, as well as increased fasted blood glucose levels. Furthermore, we found obesity-associated hypermetabolic changes in brain glucose metabolism in the HFD group. Our main findings with respect to neuroinflammation were that neither [18F]GE-180 PET nor histological analyses of brain samples seem fit to detect the predicted cerebral inflammation response, despite clear evidence of perturbed brain metabolism along with elevated IL-1β expression. These results could be interpreted as a metabolically activated state in brain-resident immune cells due to a long-term HFD.

Blocking DCIR mitigates colitis and prevents colorectal tumors by enhancing the GM-CSF-STAT5 pathway.

Dendritic cell immunoreceptor (DCIR; Clec4a2), a member of the C-type lectin receptor family, plays important roles in homeostasis of the immune and bone systems. However, the intestinal role of this molecule is unclear. Here, we show that dextran sodium sulfate (DSS)-induced colitis and azoxymethane-DSS-induced intestinal tumors are reduced in Clec4a2-/- mice independently of intestinal microbiota. STAT5 phosphorylation and expression of Csf2 and tight junction genes are enhanced, while Il17a and Cxcl2 are suppressed in the Clec4a2-/- mouse colon, which exhibits reduced infiltration of neutrophils and myeloid-derived suppressor cells. Granulocyte-macrophage colony-stimulating factor (GM-CSF) administration ameliorates DSS colitis associated with reduced Il17a and enhanced tight junction gene expression, whereas anti-GM-CSF exacerbates symptoms. Furthermore, anti-NA2, a ligand for DCIR, ameliorates colitis and prevents colorectal tumors. These observations indicate that blocking DCIR signaling ameliorates colitis and suppresses colonic tumors, suggesting DCIR as a possible target for the treatment of these diseases.

Urinary Tract Infections Impair Adult Hippocampal Neurogenesis.

Simple SummaryUrinary tract infections are associated with features of cognitive decline and memory deficits, where the underlying correlation or mechanism is still not clear. In this study, we investigate the effect of urinary tract infections on cognitive functions in rodents and whether it is associated with adult hippocampal neurogenesis, a process that is detrimental for memory formation. We have shown that urinary tract infection affects the time spent exploring a novel arm in the Y-maze test. This was accompanied with a decrease in the proliferation of neural stem cells at an early time point post infection and a persistent decrease in neurogenesis at a later time point (34 days). We also detected decreased levels of neurotrophic factors important for neurogenesis and an elevated expression of interleukin 1β in the hippocampus. Treatment with either anti-inflammatory drugs or anti-biotics does not recover proliferation of neural stem cells. Here, we present hippocampal neurogenesis as a possible contributor to cognitive changes associated with urinary tract infections. Given the significant increase in urinary tract infection occurrence, it is important to address some of the detrimental effects that such an infection can have at the level of the brain.Previous studies have suggested a link between urinary tract infections (UTIs) and cognitive impairment. One possible contributing factor for UTI-induced cognitive changes that has not yet been investigated is a potential alteration in hippocampal neurogenesis. In this study, we aim to investigate the effect of UTI on brain plasticity by specifically examining alterations in neurogenesis. Adult male Sprague Dawley rats received an intra-urethral injection of an Escherichia coli (E. coli) clinical isolate (108 CFU/mL). We found that rats with a UTI (CFU/mL ≥ 105) had reduced proliferation of neural stem cells (NSCs) at an early time point post infection (day 4) and neurogenesis at a later time point (day 34). This was associated with the decreased expression in mRNA of BDNF, NGF, and FGF2, and elevated expression of IL-1β in the hippocampus at 6 h post infection, but with no changes in optical intensity of the microglia and astrocytes. In addition, infected rats spent less time exploring a novel arm in the Y-maze test. Treatment with an anti-inflammatory drug did not revert the effect on NSCs, while treatment with antibiotics further decreased the basal level of their proliferation. This study presents novel findings on the impact of urinary tract infections on hippocampal neurogenesis that could be correlated with cognitive impairment.

JAK2/STAT3 Axis Intermediates Microglia/Macrophage Polarization During Cerebral Ischemia/Reperfusion Injury

Background: Subtypes of microglia/macrophage regulate the inflammation in the opposite direction during ischemic stroke. JAK2/STAT3 signaling pathway participates in the development of stroke-related inflammation via ischemic stimulation. However, the relationship between JAK2/STAT3 pathway and microglia/macrophage phenotype transformation is unclear. Methods: This study established a transient middle cerebral artery occlusion (tMCAO) model in male STAT3f/f and STAT3f/f LysMcre+ mice and evaluated the neurological deficit on the 3rd day using Longa score. The brains were stained by TTC to determine the infarction volume. Western blotting and QPCR were used to determine the expression of JAK2/STAT3 pathway and microglia/macrophage-related markers. Immunofluorescence staining was used to detect the levels of polarization-related indexes. QPCR also assessed the effect of STAT3 knockout on inflammatory factors in the infarction. Moreover, established the OGD/R model using BV2 cells to further verify the role of STAT3 on microglia/macrophage polarization. Results: For the conditioned STAT3-KO mice, the infarction was significantly increased after MCAO, accompanied by the aggravation of neurological deficit. Higher expression of iNOS and CD16/32 than Arg-1, Ym-1, and CD206 in vivo and in vitro, and decreased p-STAT3/STAT3 ratio in STAT3f/f LysMcre+ mice, while the p-JAK2/JAK2 ratio increased. In addition, increased M1/M2 ratio and elevated expression of IL-1β, IL-6, TNF-α with STAT3 deletion, as well as increased CD68+/iNOS+ cell numbers. Conclusion: Collectively, these results reveal that JAK2/STAT3 signaling pathway regulates the microglia/macrophage polarization (skewing toward the M2 polarization) during the CIRI, thus alleviating brain damage. Therefore, approaches targeting JAK2/STAT3 activation are promising therapies for ischemic stroke.

Exacerbated Vascular Inflammation Following Traumatic Hemorrhage in Obese Rats

IntroductionObesity increases the risk for morbidity and mortality in trauma patients. These complications are associated with prolonged tissue hypoperfusion, hypercoagulability, and edema that reflect profound vascular damage. Traumatic hemorrhage acutely attenuates vascular responsiveness, but the degree of this dysfunction during the obese state is not known. Traumatic hemorrhage also increases circulating inflammatory cytokines; however, the local inflammatory response in vascular cells is unknown. We hypothesized that obesity potentiates trauma‐induced vascular inflammation and dysfunction.MethodsMale Sprague‐Dawley rats (~250g) fed either normal chow (NC; 13.5% kcal fat, n=20), moderate‐fat (MF; 33% kcal fat, n=20), or high‐fat (HF; 60% kcal fat, n=20) diet for 6‐8 weeks were assigned to a traumatic hemorrhage protocol or sham control. Under anesthesia, hemorrhage was induced by the laceration of a mesenteric artery and a Grade V splenic injury. Hypotension (mean arterial pressure = 30–40 mm Hg) was maintained for 30 minutes. Vasoconstrictor and vasodilator properties were assessed ex vivo in isolated mesenteric arteries pre‐ and post‐ hemorrhage. Carotid arteries collected after hemorrhage or sham control were flushed with trizol to collect endothelial cells (ECs). Gene expression was assessed in ECs and the remaining carotid artery (media + adventitia) cells for pro‐inflammatory interleukin (IL)‐1β, and IL‐6, pro‐oxidant NADPH oxidase 2 (NOX2), and α‐adrenergic receptor.ResultsIn NC rats, hemorrhage attenuated norepinephrine‐induced vasoconstriction and endothelium‐dependent dilation. This was accompanied by elevated IL‐1β expression in carotid artery ECs and elevated IL‐1β, IL‐6, NOX2, and α‐adrenergic receptor expression in the carotid artery media + adventitia post‐hemorrhage versus sham control. In MF and HF rats, norepinephrine‐induced vasoconstriction and endothelium‐dependent dilation did not change pre‐ to post‐hemorrhage. In sham animals, endothelial cell IL‐1β and NOX2 expression were higher in HF compared with NC rats. Similar to NC rats, the MF and HF rats had higher IL‐1β expression in carotid artery ECs, and IL‐1β, IL‐6, NOX2, and α‐adrenergic receptor expression in the carotid artery media + adventitia post‐hemorrhage compared with sham. The hemorrhage‐induced elevation of IL‐1β in carotid artery ECs and media + adventitia was greater in HF rats compared with NC and MF rats.ConclusionOur clinically‐relevant model of traumatic hemorrhage attenuates vascular responsiveness and induces vascular inflammation. The attenuated vascular responsiveness following hemorrhage is absent in MF and HF rats, while the elevated vascular inflammation persists. HF rats have an amplified arterial inflammatory response to hemorrhage, both in ECs and in cells of the media + adventitia. The augmented vascular inflammatory response to hemorrhage in HF rats may explain the worse outcomes in obese trauma patients.

Дегенеративная болезнь диска у молодых: цитокиновый профиль и факторы ангиогенеза

Back pain (BP), associated with the degenerative disc disease (DDD), poses a heavy social and economic burden due to early disability and indications to surgery, emerging in young adults. Pathophysiological basis of premature intervertebral disc (IVD) degeneration is being actively studied. The study was aimed to define the profiles of inflammatory cytokines in DDD, as well as their relationship to the structural spine diseases. The molecular genetic analysis of the mRNA gene abundance in patients with BP and herniated IVD after discectomy and healthy individuals was performed by the quantitative polymerase chain reaction method. High expression of TNFα, IL17 was revealed in the IVD tissues of the affected patients (p < 0.01); the levels of TNFα and IL1β correlated with the DDD severity (r = 0.301 and 0.37; p < 0.05). Elevated expression of IL1β, IL6 was found in peripheral white blood cells (p < 0.01); the levels of IL6 negatively correlated with Modic type 1 and 2 changes (r = –0.31; p < 0.05), and the levels of IL17 positively correlated with the IVD herniation in combination with erosions of the adjacent vertebral body endplates and Modic changes (r = 0.401; p < 0.05). The expression of VEGF-А in the IVD tissues and white blood cells negatively correlated with the DDD grades (r = –0.85; p < 0.001), indicating reduced vascularization in the terminal phase of the disease. The findings on DDD demonstrate the contribution of the local low-immune inflammation, coupled with the intense disc vascularization at the earlier stages, and associated with the reactive inflammation in vertebral bodies. The results are prerequisites for developing the anti-inflammatory and reparative therapy based on the DDD grade and the presence of Modic changes in young adults with BP.

Transcriptional response modules characterize IL-1β and IL-6 activity in COVID-19.

SummaryDysregulated IL-1β and IL-6 responses have been implicated in the pathogenesis of severe Coronavirus Disease 2019 (COVID-19). Innovative approaches for evaluating the biological activity of these cytokines in vivo are urgently needed to complement clinical trials of therapeutic targeting of IL-1β and IL-6 in COVID-19. We show that the expression of IL-1β or IL-6 inducible transcriptional signatures (modules) reflects the bioactivity of these cytokines in immunopathology modelled by juvenile idiopathic arthritis (JIA) and rheumatoid arthritis. In COVID-19, elevated expression of IL-1β and IL-6 response modules, but not the cytokine transcripts themselves, is a feature of infection in the nasopharynx and blood but is not associated with severity of COVID-19 disease, length of stay, or mortality. We propose that IL-1β and IL-6 transcriptional response modules provide a dynamic readout of functional cytokine activity in vivo, aiding quantification of the biological effects of immunomodulatory therapies in COVID-19.

Chronic Alcohol Administration Induces Inflammasome Activation in Mouse Lungs

ObjectiveExcessive alcohol misusers are 2–4 times more likely to develop respiratory infections and acute respiratory distress syndrome as compared to non‐alcoholics. Alcohol misuse enhances oxidative stress in the lung, leading to impaired lung immunity. The molecular mechanisms by which this lung immune dysfunction occurs remain poorly defined. Inflammasome activation occurs in alcoholic steatohepatitis and functions in oxidative stress and host defense. Autophagy is critical for the engulfment and degradation of intracellular pathogens and regulates inflammasome activation. Thus, we hypothesized that autophagy prevents oxidative stress and inflammasome activation in the lung induced by alcohol.MethodsLungs were collected from littermate Atg5F/F control or LysM‐Cre‐Atg5F/F mice with a macrophage‐specific knockout of the autophagy gene Atg5 that were fed either a control or ethanol‐containing (5% vol/vol) Lieber‐DeCarli diet for 5 weeks. Markers of oxidative stress and inflammation were measured in the lungs from these mice. mRNA levels of NADPH oxidase 4 (Nox4) and transforming growth factor beta (TGFβ) were assessed by qRT‐PCR. Protein expression of NLRP3, caspase‐1 and ASC were measured by western blotting and interleukin (IL)‐1β was quantitated by ELISA.ResultsIn vivo studies demonstrated that alcohol: 1) increased the mRNA levels of Nox4 and TGFβ; 2) increased components of the inflammasome complex, namely NLRP3, ASC and caspase‐1; and 3) elevated IL‐1β expression, indicative of inflammasome activation. Knockdown of Atg5 enhanced inflammasome activation and exacerbated the oxidative stress and inflammatory effects of alcohol.ConclusionsMacrophage autophagy likely plays a critical role in regulating lung oxidative stress and inflammasome activation during chronic alcohol ingestion. Inactivation of the autophagic pathway exacerbates the deleterious effects of alcohol. Therapeutic targeting of autophagy or inflammasome activation pathways could mitigate alcohol's detrimental effects on the lung and improve lung host immune defense.Support or Funding InformationResearch Funding Source: Department of Veterans Affairs 2I01BX001786 (RTS), and National Institute of Alcohol Abuse and Alcoholism 4R00AA021803 (SMY) and 5R01AA022601 (MJC).This abstract is from the Experimental Biology 2019 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.

Genetics of COPA syndrome.

Inborn errors of immunity usually not only result in immunodeficiency but may also manifest as immune dysregulation in the form of autoinflammation, autoimmunity, or sometimes malignancy. One of the most recently discovered monogenic disorder of immune dysregulation is COPA syndrome. COPA syndrome is an inherited autoimmune disorder caused by mutations in COPA gene. COPA gene encodes for α subunit of the COP1 protein, which is involved in the reverse vesicular protein transport from Golgi apparatus to the endoplasmic reticulum (ER). The inheritance pattern of COPA syndrome is autosomal dominant, and the patients typically present with interstitial lung disease with pulmonary hemorrhage and subsequently develop arthritis. Immunological features involve autoantibody formation, elevated expression of IL-1β and IL-6, and increase in the number of Th17 cells. Molecular pathophysiology of COPA syndrome is not clearly understood. However, it is known that accumulation of unfolded proteins in ER leads to ER stress, which is an indirect result of aberrant vesicular transport of proteins from Golgi apparatus to ER and defective cellular autophagy. ER stress induces inflammation and is responsible for pathogenesis of a large number of chronic inflammatory diseases. Unfolded protein response process responds to improperly folded proteins and defends against stress in ER to ensure the fidelity of the protein folding. It maintains the expression of stress-response genes and causes initiation of inflammatory signaling pathways essential for the innate immunity. Mutation in COPA gene associated with defective protein sorting to ER has unearthed a new primary immunodeficiency disease with a unique clinical phenotype. This review highlights the clinical and molecular aspects of COPA syndrome.

Abstract P260: Utilizing Real-World Big Data in the Search for New Renoprotective Drugs

Background: The incidence of chronic kidney disease (CKD) has been increasing globally. Because CKD worsens morbidity and mortality, elucidation of its mechanism and discovery of novel therapeutic strategies are imperative. Purpose: We utilized real-world big data to search for potential novel drugs for the treatment of CKD. Method: Signal detection by reported odds ratios (RORs) was performed using the Food and Drug Administration Adverse Events Reporting System (FAERS), one of the largest global databases. We then analyzed the relevance between existing drugs and nephritis. Drugs with low reported rates of nephritis were identified as candidate drugs for CKD; their renoprotective effects were verified by basic research. Results: Analysis of the FAERS database revealed a significant inverse association between tubulointerstitial nephritis and factor Xa (FXa) inhibitors (ROR, 0.65; 95% CI, 0.49-0.85), and between nephritis and cholinesterase inhibitors (ROR, 0.23; 95% CI, 0.07-0.71). The FXa inhibitor (FXa-I) suppressed unilateral ureteral obstruction (UUO)-induced tubulointerstitial fibrosis (UUO, 5.20 ± 0.73%; UUO+FXa-I, 3.36 ± 0.32%; p < 0.05) and extracellular matrix expression (collagen I, collagen III, and fibronectin; p < 0.01). The FXa-I also attenuated UUO-induced macrophage infiltration (UUO, 9.59 ± 0.62%; UUO+FXa-I; 6.70 ± 0.98%; p < 0.05) and inflammatory molecule upregulation (monocyte chemoattractant protein 1 [MCP-1], interleukin 1 beta [IL-1β], and tumor necrosis factor alpha [TNF-α]; p < 0.01). Furthermore, the FXa-I significantly reduced the UUO-induced increase in plasma creatinine in mice (UUO, 0.163 ± 0.005 mg/dL; UUO+FXa-I, 0.136 ± 0.011 mg/dL; p < 0.05). The cholinesterase inhibitor significantly mitigated the increased expression of collagen I and MCP-1 in aristolochic acid-induced CKD mice ( p < 0.05). That also inhibited elevated expression of IL-1β and TNF-α in lipopolysaccharide-stimulated J774 mouse cells. Conclusion: We believe that the anti-inflammatory effects of FXa inhibitors and cholinesterase inhibitors make them potentially useful renoprotective drugs. Thus, real-world big data might be utilized for the discovery of drug repositioning candidates for the treatment of CKD.

ID: 195: Identification of cytokine cell sources in a new mouse model of systemic juvenile idiopathic arthritis highlights innate immunity of this disorder

Objective: Systemic juvenile idiopathic arthritis (sJIA) is a rheumatic childhood disease, with distinctive systemic inflammatory features and a typical cytokine profile. The etiology of sJIA is largely unknown. Here, we aimed to elucidate the role of specific cell populations and cytokines in the pathogenesis, by means of a new mouse model of sJIA relying on the injection of complete Freund’s adjuvant (CFA) in interferon-gamma deficient (IFN-g KO) mice. Wild type (WT) mice developed a minor inflammation, indicating that IFN-g provides protection in the disease process. Methods: Cytokine mRNA levels were analyzed in organs and purified immune cell populations of CFA-challenged IFN-g KO and WT mice. Cytokine antagonists and cell depleting antibodies were administered to mice. Clinical, laboratory and immune features of sJIA were evaluated. Results: In the diseased IFN-g KO mice, elevated expression of IL 1β, IL-6, IL-17 and IL-22 were found in lymph nodes and – remarkably – also in lung tissue. Gamma-delta T cells were a major cell source for IL-17, and anti-IL-17 antibodies abrogated the disease. In the non-diseased WT mice, expression of IFN-g was almost exclusively found in NK, NKT and gamma-delta T cells. Transient neutralization of IFN-g as well as depletion of NK cells in WT mice both resulted in a fulminant sJIA-like disease. Conclusion: These data pinpoint innate immune cells and non-lymphoid organs as important sources of cytokines in CFA-induced animal models. The results also demonstrate that NK cells provide a regulatory function in the pathogenesis of sJIA, by production of IFN-g.

Central cyclooxygenase inhibition during the course of lung injury attenuates breathing frequency

Ventilatory pattern is a neurally integrated output that is altered by lung injury. Work in our lab has provided evidence that ventilatory pattern changes persist in situ, and are concurrent with elevated IL‐1β expression, astrogliosis and altered synaptic efficacy in the nucleus of the tractus solitarii (nTS), a brainstem region critical for viscerosensory integration. Most recently, we observed that microinjection of exogenous IL‐1β into the nTS of naïve rats evokes some of the ventilatory pattern changes observed during lung injury. Collectively, these findings suggest that lung injury exerts influence over the neural control of breathing in part through the action of neuroinflammation. We therefore hypothesized that inhibiting neuroinflammation during the course of lung injury would attenuate breathing frequency (FR) and reduce IL‐β expression in the nTS. To test this hypothesis, we continuously administered either vehicle or indomethacin (IND), a cyclooxygenase‐1/2 (COX‐1/2) inhibitor, into the right lateral ventricle of bleomycin lung‐injured rats (bleo) via Alzet pump. We plethysmographically recorded breathing pattern and densitometrically measured brainstem IL‐1β immunoreactivity (IR) at day 7 of lung injury. We observed a significant (P<0.05) attenuation of FR in bleo rats receiving IND (161±56bpm, n=10) as compared to bleo rats receiving vehicle (217±66bpm, n=10). The level of IL‐1β IR detected within the nTS of IND treated bleo rats was reduced. Taken together, our results indicate that ventilatory pattern changes, which occur in the setting of lung injury, are in part dependent upon the central action of COX‐1/2, and are associated with decreased IL‐1β expression in the nTS.

Knockdown of Interleukin-1 Receptor Type-1 on Endothelial Cells Attenuated Stress-Induced Neuroinflammation and Prevented Anxiety-Like Behavior

Interleukin-1β (IL-1β) is an inflammatory cytokine that plays a prominent role in stress-induced behavioral changes. In a model of repeated social defeat (RSD), elevated IL-1β expression in the brain was associated with recruitment of primed macrophages that were necessary for development of anxiety-like behavior. Moreover, microglia activation and anxiety-like behavior associated with RSD did not occur in IL-1 receptor type-1 knock-out (IL-1R1(KO)) mice. Therefore, the objective of this study was to examine the role of IL-1 signaling in RSD-induced macrophage trafficking to the brain and anxiety-like behavior. Initial studies revealed that RSD did not increase circulating myeloid cells in IL-1R1(KO) mice, resulting in limited macrophage trafficking to the brain. In addition, IL-1R1(KO) bone marrow-chimera mice showed that IL-1R1 expression was essential for macrophage trafficking into the brain. To differentiate cellular mediators of stress-induced IL-1 signaling, endothelial-specific IL-1R1 knock-down (eIL-1R1kd) mice were used. Both wild-type (WT) and eIL-1R1kd mice had increased circulating monocytes, recruitment of macrophages to the brain, and altered microglia activation after RSD. Nonetheless, RSD-induced expression of IL-1β, TNF-α, and IL-6 mRNA in brain CD11b(+) cells was attenuated in eIL-1R1kd mice compared with WT. Moreover, anxiety-like behavior did not develop in eIL-1R1kd mice. Collectively, these findings demonstrated that there was limited RSD-induced priming of myeloid cells in IL-1R1(KO) mice and disrupted propagation of neuroinflammatory signals in the brain of eIL-1R1kd mice. Furthermore, these data showed that transduction of IL-1 signaling by endothelial cells potentiates stress-induced neuroinflammation and promotes anxiety-like behavior.

Interleukin-1β Promotes Ovarian Tumorigenesis through a p53/NF-κB-Mediated Inflammatory Response in Stromal Fibroblasts

Cancer has long been considered a disease that mimics an “unhealed wound,” with oncogene-induced secretory activation signals from epithelial cancer cells facilitating stromal fibroblast, endothelial, and inflammatory cell participation in tumor progression. However, the underlying mechanisms that orchestrate cooperative interaction between malignant epithelium and the stroma remain largely unknown. Here, we identified interleukin-1β (IL-1β) as a stromal-acting chemokine secreted by ovarian cancer cells, which suppresses p53 protein expression in cancer-associated fibroblasts (CAFs). Elevated expression of IL-1β and cognate receptor IL-1R1 in ovarian cancer epithelial cells and CAFs independently predicted reduced overall patient survival, as did repressed nuclear p53 in ovarian CAFs. Knockdown of p53 expression in ovarian fibroblasts significantly enhanced the expression and secretion of chemokines IL-8, growth regulated oncogene-alpha (GRO-α), IL-6, IL-1β, and vascular endothelial growth factor (VEGF), significantly increased in vivo mouse xenograft ovarian cancer tumor growth, and was entirely dependent on interaction with, and transcriptional up-regulation of, nuclear factor-kappaB (NF-κB) p65. Our results have uncovered a previously unrecognized circuit whereby epithelial cancer cells use IL-1β as a communication factor instructing stromal fibroblasts through p53 to generate a protumorigenic inflammatory microenvironment. Attenuation of p53 protein expression in stromal fibroblasts generates critical protumorigenic functionality, reminiscent of the role that oncogenic p53 mutations play in cancer cells. These findings implicate CAFs as an important target for blocking inflammation in the tumor microenvironment and reducing tumor growth.

Abstract B59: The dual effect of therapy-induced IL-1β expression on tumor progression: Role of tumor-associated macrophages

Abstract A major obstacle in clinical oncology is that tumors eventually acquire resistance to therapy. The mechanism by which resistance arises is usually associated with changes made in tumor cells in response to the therapy, however, little is known about the effect of the host which may also contribute to tumor resistance. Previous studies from our lab suggest that the induction of host cellular and molecular events following therapy may promote tumor re-growth, angiogenesis and metastasis. In this study we uncovered the possible role IL-1β - a proinflammatory cytokine - has on the growth of primary tumor and its impact on metastasis following paclitaxel (PTX) chemotherapy. We found that plasma and spleen from PTX-treated mice exhibited elevated expression of IL-1β. Furthermore, plasma from PTX-treated mice induced tumor cell invasion, an effect which was abrogated in plasma from mice treated with PTX and Anakinra, an IL-1 receptor antagonist. Consequently, mice inoculated with Lewis lung carcinoma (LLC) cells pre-exposed to plasma from mice treated with the combined therapy survived longer than mice inoculated with tumor cells pre-exposed to plasma from mice treated with PTX or Anakinra monotherapy. We next determined whether treatment efficacy of PTX is enhanced by the combination of PTX and Anakinra using 4T1 metastatic breast cancer model in mice. Tumors treated with the combined therapy were significantly smaller than tumors treated with PTX or Anakinra monotherapy. Surprisingly, although the primary tumors were smaller, such mice succumb to metastasis earlier than mice in any of the other treated groups. Furthermore, an increased number of M2 macrophages colonizing 4T1 primary tumors treated with the combined therapy was observed when compared to their number in PTX treated tumors, indicating a possible pro-metastatic role of M2 macrophages in post treatment IL-1β deprived microenvironment. Parallel results were observed when LLC bearing IL-1β deficient mice, were treated with PTX. Taken together, these findings demonstrate that although treatment outcome is enhanced following the blockade of host-induced IL1β at the primary tumor level, at the metastatic level it may serve as a double-edge sward and can lead to metastatic spread. Citation Format: Tali Voloshin, Dror Alishekevitz, Valeria Miller, Lika Isakov, Rotem Bril, Moran Benhar, Roney Apte, Yuval Shaked. The dual effect of therapy-induced IL-1β expression on tumor progression: Role of tumor-associated macrophages. [abstract]. In: Proceedings of the AACR Special Conference on Tumor Invasion and Metastasis; Jan 20-23, 2013; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2013;73(3 Suppl):Abstract nr B59.

Fractalkine receptor (CX3CR1) deficiency sensitizes mice to the behavioral changes induced by lipopolysaccharide

BackgroundInteractions between fractalkine (CX3CL1) and fractalkine receptor (CX3CR1) regulate microglial activation in the CNS. Recent findings indicate that age-associated impairments in CX3CL1 and CX3CR1 are directly associated with exaggerated microglial activation and an impaired recovery from sickness behavior after peripheral injection of lipopolysaccharide (LPS). Therefore, the purpose of this study was to determine the extent to which an acute LPS injection causes amplified and prolonged microglial activation and behavioral deficits in CX3CR1-deficient mice (CX3CR1-/-).MethodsCX3CR1-/- mice or control heterozygote mice (CX3CR1+/-) were injected with LPS (0.5 mg/kg i.p.) or saline and behavior (i.e., sickness and depression-like behavior), microglial activation, and markers of tryptophan metabolism were determined. All data were analyzed using Statistical Analysis Systems General Linear Model procedures and were subjected to one-, two-, or three-way ANOVA to determine significant main effects and interactions.ResultsLPS injection caused a prolonged duration of social withdrawal in CX3CR1-/- mice compared to control mice. This extended social withdrawal was associated with enhanced mRNA expression of IL-1β, indolamine 2,3-dioxygenase (IDO) and kynurenine monooxygenase (KMO) in microglia 4 h after LPS. Moreover, elevated expression of IL-1β and CD14 was still detected in microglia of CX3CR1-/- mice 24 h after LPS. There was also increased turnover of tryptophan, serotonin, and dopamine in the brain 24 h after LPS, but these increases were independent of CX3CR1 expression. When submitted to the tail suspension test 48 and 72 h after LPS, an increased duration of immobility was evident only in CX3CR1-/- mice. This depression-like behavior in CX3CR1-/- mice was associated with a persistent activated microglial phenotype in the hippocampus and prefrontal cortex.ConclusionsTaken together, these data indicate that a deficiency of CX3CR1 is permissive to protracted microglial activation and prolonged behavioral alterations in response to transient activation of the innate immune system.

Two populations of alternatively activated M2 macrophages (TLR4+ vs. Tim-3+) regulate inflammation and fibrosis in the heart during coxsackievirus-induced myocarditis (137.20)

Abstract Previously we showed that Toll-like receptor 4 (TLR4) and T cell Ig mucin-3 (Tim-3) cross-regulate each others expression on CD11b+ cells, with TLR4 increasing and Tim-3 reducing cardiac inflammation. We show here that TLR4 and Tim-3 are expressed on GR1+F4/80+IL-4R+ alternatively activated M2 macrophages from the heart during acute myocarditis, but are not expressed on GR1-F4/80+IL-4R- classically activated M1 macrophages. GR1+ cells also express the mannose receptor (CD206), caspase-1, IL-1β and IL-10. In gonadectomized male mice, M2 cells express significantly more Tim-3 and significantly less IL-1β with significantly reduced inflammation. In contrast, complement receptor 1/2 (CR1/2) deficient mice have a significantly increased TLR4+casp-1+IL-1β+ M2 population. Elevated expression of IL-1β by M2 macrophages in the heart leads to significantly increased myocarditis, fibrosis and heart failure in CR1/2 deficient mice. These data indicate that different M2 populations have contrasting roles on the development of heart disease: TLR4+casp-1+IL-1β+ M2 are associated with increased inflammation and fibrosis, while Tim-3+ M2 with decreased inflammation. Deciphering how M2 macrophage populations interact with autoreactive cells and antibodies will be critical in understanding the pathogenesis of autoimmune disease. Supported by NIH grants R01 HL087033 (DF) and R01 HL67290 (NRR).