IL-10 Deficient Mice Research Articles

Interleukin-10 Promotes Porcine Circovirus Type 2 Persistent Infection in Mice and Aggravates the Tissue Lesions by Suppression of T Cell Infiltration

Interleukin (IL)-10, as a key anti-inflammatory cytokine, increases during porcine circovirus type 2 (PCV2) infection, but the role of IL-10 in the process remains to be defined. In the present study, using an IL-10 deficient mice model, we found that IL-10 deficiency prevented the reduction of splenic lymphocytes (CD45+ cells) induced by PCV2 and promoted CD4+ and CD8+ T cell infiltration in lungs through inducting more T cell chemokines (CCL3, CXCL9, and CXCL10). Simultaneously, PCV2 infection induced a significant increase of pro-inflammatory cytokines and PCV2-specific antibodies in IL-10 deficient mice than in wild-type mice, resulting in a lower viral load in lung and a milder lung lesion in IL-10 deficient mice relative to wild-type mice. Moreover, the amounts of pulmonary CD4+ and CD8+ T cells were all inversely correlated with the lung lesions, as well as the viral load of PCV2. These results demonstrate that PCV2 infection employs IL-10 to block the transfer of T cells to the lungs of mice, and IL-10 attenuates the production of pro-inflammatory cytokines and PCV2-specific antibodies. The lack of T cell infiltration, pro-inflammatory cytokines, and PCV2-specific antibodies promote PCV2 replication, leading to a more severe lung lesion in mice.

The commensal Escherichia coli CEC15 reinforces intestinal defences in gnotobiotic mice and is protective in a chronic colitis mouse model

Escherichia coli is a regular inhabitant of the gut microbiota throughout life. However, its role in gut health is controversial. Here, we investigated the relationship between the commensal E. coli strain CEC15 (CEC), which we previously isolated, and the intestine in homeostatic and disease-prone settings. The impact of CEC was compared to that of the probiotic E. coli Nissle 1917 (Nissle) strain. The expression of ileal and colonic genes that play a key role in intestinal homeostasis was higher in CEC- and Nissle-mono-associated wild-type mice than in germfree mice. This included genes involved in the turnover of reactive oxygen species, antimicrobial peptide synthesis, and immune responses. The impact of CEC and Nissle on such gene expression was stronger in a disease-prone setting, i.e. in gnotobiotic IL10-deficient mice. In a chronic colitis model, CEC more strongly decreased signs of colitis severity (myeloperoxidase activity and CD3+ immune-cell infiltration) than Nissle. Thus, our study shows that CEC and Nissle contribute to increased expression of genes involved in the maintenance of gut homeostasis in homeostatic and inflammatory settings. We show that these E. coli strains, in particular CEC, can have a beneficial effect in a chronic colitis mouse model.

Editing of the gut microbiota reduces carcinogenesis in mouse models of colitis-associated colorectal cancer.

Chronic inflammation and gut microbiota dysbiosis, in particular the bloom of genotoxin-producing E. coli strains, are risk factors for the development of colorectal cancer. Here, we sought to determine whether precision editing of gut microbiota metabolism and composition could decrease the risk for tumor development in mouse models of colitis-associated colorectal cancer (CAC). Expansion of experimentally introduced E. coli strains in the azoxymethane/dextran sulfate sodium colitis model was driven by molybdoenzyme-dependent metabolic pathways. Oral administration of sodium tungstate inhibited E. coli molybdoenzymes and selectively decreased gut colonization with genotoxin-producing E. coli and other Enterobacteriaceae. Restricting the bloom of Enterobacteriaceae decreased intestinal inflammation and reduced the incidence of colonic tumors in two models of CAC, the azoxymethane/dextran sulfate sodium colitis model and azoxymethane-treated, Il10-deficient mice. We conclude that metabolic targeting of protumoral Enterobacteriaceae during chronic inflammation is a suitable strategy to prevent the development of malignancies arising from gut microbiota dysbiosis.

Composition of the Intestinal Microbiota Determines the Outcome of Virus-Triggered Colitis in Mice.

The intestinal microbiota is a complex ecosystem implicated in host health and disease. Inflammatory bowel disease (IBD) is a multifactorial chronic disorder of the gastrointestinal mucosa. Even though the exact mechanisms are still unknown, the intestinal microbiota is crucial in IBD development. We previously showed that murine norovirus (MNV) induces colitis in the Il10-deficient (Il10−/−) mouse model of IBD in a microbiota-dependent manner. Thus, in this study we analyzed whether distinct minimal bacterial consortia influence the outcome of MNV-triggered colitis in Il10−/− mice. Gnotobiotic Il10−/− mice associated with Oligo-Mouse-Microbiota 12 (OMM12) or Altered Schaedler Flora (ASF) developed little to no inflammatory lesions in the colon and cecum. MNV infection exacerbated colitis severity only in ASF-colonized mice, but not in those associated with OMM12. Four weeks after MNV infection, inflammatory lesions in ASF-colonized Il10−/− mice were characterized by epithelial hyperplasia, infiltration of inflammatory cells, and increased barrier permeability. Co-colonization of ASF-colonized Il10−/− mice with segmented filamentous bacteria (SFB) abolished MNV-induced colitis, whereas histopathological scores in SFB-OMM12-co-colonized mice stayed unchanged. Moreover, SFB only colonized mice associated with ASF. The SFB-mediated protective effects in ASF-colonized mice involved enhanced activation of intestinal barrier defense mechanisms and mucosal immune responses in the chronic and acute phase of MNV infection. SFB colonization strengthened intestinal barrier function by increasing expression of tight junction proteins, antimicrobial peptides and mucus. Furthermore, SFB colonization enhanced the expression of pro-inflammatory cytokines such as Tnfα, Il1β, and Il12a, as well as the expression of the regulatory cytokine Tgfβ. Altogether, our results showed that MNV-triggered colitis depends on the microbial context.

IL-10 producing B cells rescue mouse fetuses from inflammation-driven fetal death and are able to modulate T cell immune responses

Understanding the mechanisms leading to fetal death following maternal subclinical infections is crucial to develop new therapeutic strategies. Here we addressed the relevance of IL-10 secreting B cells (B10) in the maintenance of the immune balance during gestation. µMT females lacking mature B cells presented normal pregnancies, although their fetuses were smaller and their Treg pool did not expand as in B cell sufficient controls. Pregnant µMT females were more susceptible to LPS despite having less Treg; their fetuses died at doses compatible with pregnancy in WT animals. Adoptive transfer of IL-10 negative B effector cells or B cells from IL-10 deficient mice did not modify this outcome. The transfer of B10 cells or application of recombinant murine IL-10 reduced the fetal loss, associated with a normalization of Treg numbers and cytokine modulation at the feto-maternal interface. B cell-derived IL-10 suppressed the production of IL-17A and IL-6 by T cells and promoted the conversion of naïve cells into Treg. B10 cells are required to restore the immune balance at the feto-maternal interface when perturbed by inflammatory signals. Our data position B cells in a central role in the maintenance of the balance between immunity and tolerance during pregnancy.

Tumor necrosis factor stimulates fibroblast growth factor 23 levels in chronic kidney disease and non-renal inflammation

Fibroblast growth factor 23 (FGF23) regulates phosphate homeostasis, and its early rise in patients with chronic kidney disease is independently associated with all-cause mortality. Since inflammation is characteristic of chronic kidney disease and associates with increased plasma FGF23 we examined whether inflammation directly stimulates FGF23. In a population-based cohort, plasma tumor necrosis factor (TNF) was the only inflammatory cytokine that independently and positively correlated with plasma FGF23. Mouse models of chronic kidney disease showed signs of renal inflammation, renal FGF23 expression and elevated systemic FGF23 levels. Renal FGF23 expression coincidedwith expression of the orphan nuclear receptor Nurr1 regulating FGF23 in other organs. Antibody-mediated neutralization of TNF normalized plasma FGF23 and suppressed ectopic renal Fgf23 expression. Conversely, TNF administration to control mice increased plasma FGF23 without altering plasma phosphate. Moreover, in Il10-deficient mice with inflammatory bowel disease and normal kidney function, plasma FGF23 was elevated and normalized upon TNF neutralization. Thus, the inflammatory cytokine TNF contributes to elevated systemic FGF23 levels and also triggers ectopic renal Fgf23 expression in animal models of chronic kidney disease.

IL-33 promotes antiviral immunity and protects against congenital Zika disease in mice

Abstract Zika virus (ZIKV) infection during pregnancy can cause congenital Zika syndrome and other birth defects. The alarmin IL-33 can evoke strong antiviral immune responses during viral infection. However, whether IL-33 is involved in immune responses during ZIKV infection is not clear. In this study, we infected three-week-old wildtype (WT) and IL-33 knockout (KO) mice with ZIKV and analyzed the ZIKV-specific cytotoxic T lymphocyte (CTL) responses at 6 days post infection. We found that IL-33 deficiency resulted in decreased ZIKV-specific CTL responses as evidenced by lower numbers of IFN-g+TNF-a+ CD8+ T effectors. To determine if IL-33 contributes to anti-ZIKV immunity during pregnancy, we infected WT pregnant mice with ZIKV and found increased IL-33 in the placenta at a mid-gestational period (embryo E13.5) compared with that of uninfected ones. Likewise, elevated IL-33 expression was also observed in human trophoblast cell lines JEG-3 and HTR-8 upon ZIKV infection. To determine if IL-33 plays a protective role in congenital Zika disease, we infected pregnant WT and IL-33 KO mice by i.p. injection of ZIKV at E6.5 and found higher percentages of fetal demise and absorption in IL-33 KO mice compared with that in WT mice (16.7 % vs. 3.3 %) from necropsy on E13.5. In addition, the fetuses from IL-33 KO mice displayed smaller sizes than those of WT fetuses (55 mm2 vs. 48 mm2). Moreover, IL-33 deficient mice had lower numbers of virus-specific CD8+ T cells in the placenta, uterine and decidua. Importantly, IL-33 deficiency resulted in higher viral loads in both the placenta and fetus. In summary, our study highlights IL-33 as a key immune regulator in inducing anti-ZIKV CTL responses and protecting against congenital Zika disease.

Antibiotics exacerbated colitis by affecting the microbiota, Treg cells and SCFAs in IL10-deficient mice

Many studies have shown that antibiotic therapy can attenuate colitis in IL10-deficient (IL10−/−) mice. However, these results have indicated that antibiotics were more successful in preventing, rather than treating established colitis. Those antibiotic treatments attempted to only partially alter the intestinal microbiota and to not eliminate it completely. Therefore, we treated IL10−/− mice with the multiantibiotic regimen that was used to develop a pseudo-germ-free mouse model to determine whether multi-antibiotics attenuated or exacerbated colitis.We evaluated the colitis in IL10−/− mice receiving the antibiotic treatment versus those receiving the water control; furthermore, we investigated the gut microbiota, the intestinal immune cell proportions and the cytokine secretion. Surprisingly, the IL10-/- mice receiving the antibiotic treatment had more severe intestinal colitis and a swollen cecum than those receiving the water control. Moreover, the abundance of microbiota and content of short-chain fatty acids (SCFAs) in the colon were dramatically decreased. Additionally, the proportions of Treg cells and Th1 cells in the colons of IL10-/- mice were also decreased. The mechanism may be that the decrease in the microbiota leads to a decrease in the proportions of Treg cells and SCFAs, which are necessary to maintain intestinal homeostasis. All changes lead to further exacerbated colitis in IL10−/− mice with antibiotic treatment.

Protease Activity of Campylobacter jejuni HtrA Modulates Distinct Intestinal and Systemic Immune Responses in Infected Secondary Abiotic IL-10 Deficient Mice.

Even though human Campylobacter jejuni infections are progressively increasing worldwide, the underlying molecular mechanisms of pathogen-host-interactions are still not fully understood. We have recently shown that the secreted serine protease HtrA plays a key role in C. jejuni cellular invasion and transepithelial migration in vitro, and is involved in the onset of intestinal pathology in murine infection models in vivo. In the present study, we investigated whether the protease activity of HtrA had an impact in C. jejuni induced acute enterocolitis. For this purpose, we perorally infected secondary abiotic IL-10−/− mice with wildtype C. jejuni strain NCTC11168 (11168WT) or isogenic bacteria carrying protease-inactive HtrA with a single point mutation at S197A in the active center (11168HtrA−S197A). Irrespective of the applied pathogenic strain, mice harbored similar C. jejuni loads in their feces and exhibited comparably severe macroscopic signs of acute enterocolitis at day 6 postinfection (p.i.). Interestingly, the 11168HtrA−S197A infected mice displayed less pronounced colonic apoptosis and immune cell responses, but enhanced epithelial proliferation as compared to the 11168WT strain infected controls. Furthermore, less distinct microscopic sequelae in 11168HtrA−S197A as compared to parental strain infected mice were accompanied by less distinct colonic secretion of pro-inflammatory cytokines such as MCP-1, IL-6, TNF, and IFN-γ in the former as compared to the latter. Strikingly, the S197A point mutation was additionally associated with less pronounced systemic pro-inflammatory immune responses as assessed in serum samples. In conclusion, HtrA is a remarkable novel virulence determinant of C. jejuni, whose protease activity is not required for intestinal colonization and establishment of disease, but aggravates campylobacteriosis by triggering apoptosis and pro-inflammatory immune responses.

High-Dimensional Characterization of IL-10 Production and IL-10-Dependent Regulation during Primary Gammaherpesvirus Infection.

IL-10 is a potent immunomodulatory cytokine produced by multiple cell types to restrain immune activation. Many herpesviruses use the IL-10 pathway to facilitate infection, but how endogenous IL-10 is regulated during primary infection in vivo remains poorly characterized. In this study, we infected mice with murine gammaherpesvirus 68 (γHV68) and analyzed the production and genetic contribution of IL-10 by mass cytometry (cytometry by time-of-flight) analysis. γHV68 infection elicited a breadth of effector CD4 T cells in the lungs of acutely infected mice, including a highly activated effector subset that coexpressed IFN-γ, TNF-α, and IL-10. By using IL-10 GFP transcriptional reporter mice, we identified that IL-10 was primarily expressed within CD4 T cells during acute infection in the lungs. IL10gfp-expressing CD4 T cells were highly proliferative and characterized by the expression of multiple coinhibitory receptors, including PD-1 and LAG-3. When we analyzed acute γHV68 infection of IL-10-deficient mice, we found that IL-10 limits the frequency of both myeloid and effector CD4 T cell subsets in the infected lung, with minimal changes at a distant mucosal site. These data emphasize the unique insights that high-dimensional analysis can afford in investigating antiviral immunity and provide new insights into the breadth, phenotype, and function of IL-10-expressing effector CD4 T cells during acute virus infection.

IL-33 Is a Negative Regulator of Vaccine-Induced Antigen-Specific Cellular Immunity.

The cytokine IL-33 is a well-established inducer of Th2 responses. However, roles for IL-33 in promoting CD8, Th1, and T regulatory cell responses have also emerged. In this study, the role of IL-33 as a regulator of particulate vaccine adjuvant-induced Ag-specific cellular immunity was investigated. We found that polymeric nanoparticles surpassed alum in their ability to enhance Ag-specific CD8 and Th1 responses. IL-33 was a potent negative regulator of both CD8+ T cell and Th1 responses following i.m. vaccination with Ag and nanoparticles, whereas the cytokine was required for the nanoparticle enhancement in Ag-specific IL-10. In contrast to the effect on cellular immunity, Ab responses were comparable between vaccinated wild-type and IL-33-deficient mice. IL-33 did not compromise alum-induced adaptive cellular immunity after i.m. vaccination. These data suggest that IL-33 attenuates the induction of cellular immune responses by nanoparticulate adjuvants and should be considered in the rational design of vaccines targeting enhanced CD8 and Th1 responses.

IL-24 contributes to skin inflammation in Para-Phenylenediamine-induced contact hypersensitivity

Para-Phenylenediamine (PPD) is an aromatic amine used in hair dyes and in temporary black henna tattoos, which is a frequent cause of allergic contact dermatitis (ACD). ACD is a skin inflammatory reaction characterized by modifications such as spongiosis, exocytosis and acanthosis. The aim of this study is to characterize the expression and the role of IL-20-related cytokines, including IL-19, IL-20, IL-22 and IL-24, in ACD. The expression of IL19, IL20, IL22 and IL24 is increased in affected skin from PPD allergic patients compared with uninvolved skin. In addition, the expression of these cytokines positively correlates with clinical symptoms. To assess their role in ACD, we set up a mouse model of PPD-induced allergic contact dermatitis and we showed that, in contrast to Il22-deficient mice, Il22ra1-, Il20rb- and Il24-deficient mice are partially protected against development of PPD-induced contact hypersensitivity. These mice have decreased ear thickening and less acanthosis compared with WT mice after PPD treatment. In addition, the absence of IL-22R, IL-20R2 or IL-24 affects the recruitment of neutrophils into the skin but not the total IgE production. Taken together, these results demonstrate the implication of IL-24 via the IL-20R type II receptor in the inflammatory process of ACD.

IL-4 promotes stromal cell expansion and is critical for development of a type-2, but not a type 1 immune response.

IL-4 is critical for differentiation of Th2 cells and antibody isotype switching, but our work demonstrated that it is produced in the peripheral LN under both Type 2, and Type 1 conditions, raising the possibility of other functions. We found that IL-4 is vital for proper positioning of hematopoietic and stromal cells in steady state, and the lack of IL-4 or IL-4Rα correlates with disarrangement of both follicular dendritic cells and CD31+ endothelial cells. We observed a marked disorganization of B cells in these mice, suggesting that the lymphocyte-stromal cell axis is maintained by the IL-4 signaling pathway. This study showed that absence of IL-4 correlates with significant downregulation of Lymphotoxin alpha (LTα) and Lymphotoxin beta (LTβ), critical lymphokines for the development and maintenance of lymphoid organs. Moreover, immunization of IL-4 deficient mice with Type 2 antigens failed to induce lymphotoxin production, LN reorganization, or germinal center formation, while this process is IL-4 independent following Type 1 immunization. Additionally, we found that Type 1 antigen mediated LN reorganization is dependent on IFN-γ in the absence of IL-4. Our findings reveal a role of IL-4 in the maintenance of peripheral lymphoid organ microenvironments during homeostasis and antigenic challenge.

Interleukin-6 Contributes to the Development of Anemia in Juvenile CKD

IntroductionAnemia is a common complication of chronic kidney disease (CKD) in children; however, the role of inflammation in its pathogenesis remains incompletely understood.MethodsTo elucidate the role of interleukin (IL)-6 in renal anemia, we induced CKD by adenine diet in juvenile wild-type (WT) and IL-6 deficient (Il6KO) mice, and examined serum IL-6 and relevant parameters in children with CKD.ResultsWT-CKD mice developed anemia despite increases in serum erythropoietin and displayed low serum iron and elevated serum IL-6. IL-6 deficiency resulted in a significant improvement of red blood cell count and hemoglobin in CKD mice. This effect was associated with improvement of hypoferremia by Il6 deletion, likely mediated by hepcidin. However, correction of hypoferremia by oral iron supplementation in WT-CKD mice did not fully replicate the protective effects of Il6 deletion, suggesting an additional iron-independent role for IL-6 in CKD-anemia. Indeed, Il6 deletion mitigated the severity of renal fibrosis and alleviated relative erythropoietin insufficiency in CKD mice. Cytokine profiling in a pediatric CKD cohort demonstrated that of 10 cytokines (IL-1β, IL-2, IL-4, IL-6, IL-8, IL-10, IL-12, IL-13, tumor necrosis factor (TNF)-α, and interferon-γ), only IL-6 was significantly (inversely) associated with hemoglobin when adjusted for glomerular filtration rate (GFR). The association between IL-6 and hemoglobin in children with CKD remained significant after adjustment for CKD stage, iron therapy, and hepcidin.DiscussionIL-6 contributes to development of anemia in juvenile CKD, through mechanisms that include induction of hypoferremia, aggravation of renal fibrosis, and alteration of the erythropoietin axis. IL-6 appears to be a promising therapeutic target in the management of CKD-anemia.

IL-10 participates in the expansion and functional activation of CD8+ T cells during acute infection with Trypanosoma cruzi.

IL-10 is a pleiotropic cytokine with immunoregulatory functions affecting various cell types. In a model of experimental infection with the protozoan Trypanosoma cruzi (T. cruzi), we found increased morbidity and lower parasite control in IL-10 deficient mice (IL-10 KO) compared to wild-type (WT) mice. Despite enhanced Mϕ function and dendritic cell activation, IL-10 KO mice were more susceptible to infection. The kinetics of T cells in spleen and peripheral blood revealed that infected IL-10 KO mice failed to increase the number of spleen and circulating total CD8+ T cells, a phenomenon observed from the second week of infection in WT mice. Total CD8+ T cells from IL-10 KO mice exhibited diminished proliferation, cytotoxic potential and IFN-γ production than their WT counterparts and T. cruzi-specific CD8+ T cells displayed reduced in vivo cytotoxicity. The absence of IL-10 selectively affected expansion, survival, and increased PD-1 expression of CD8+ T cells without altering these same parameters on CD4+ T cells. Increased inhibitory receptors expression and down-modulation of T-bet by CD8+ T cells from IL-10 KO infected mice were compatible with a T cell exhaustion phenotype. Collectively, these findings reveal that during acute infection, IL-10 plays a previously unrecognized stimulatory role on CD8+ T cells, the most relevant lymphocyte population for the control of intracellular T. cruzi stages. A clear knowledge of the underlying mechanisms that drive effector functions of cytotoxic T cells is critical to understand pathogen persistence and rational design of prophylactic strategies against T. cruzi.

PO-305 An 8-week, low carbohydrate, high fat, ketogenic diet enhanced exercise capacity through improved ketolysis and lipolysis in mice

PO-305 An 8-week, low carbohydrate, high fat, ketogenic diet enhanced exercise capacity through improved ketolysis and lipolysis in mice

Transcriptional profiling of irritant contact dermatitis (ICD) in a mouse model identifies specific patterns of gene expression and immune-regulation

BackgroundIrritant contact dermatitis (ICD) is a cutaneous inflammatory response to a variety of triggers that requires no sensitization and accounts for up to 80% of occupational dermatitis cases. IL-6 has been alternately associated with both allergic and irritant dermatitis and is closely linked to skin wound healing, therefore making it an ideal candidate to investigate in the mechanism of ICD. ResultsDespite being a well-known pro-inflammatory cytokine, IL-6 deficient (IL-6KO) mice show much more severe ICD than controls. Transcriptome analysis was employed to examine irritant-exposed and control skin samples from C57BL/6 and IL-6KO mice. Over 1900 transcripts were found differentially modulated between C57 (1184 total) and IL-6KO (802 total) mice with the magnitude of expression significantly disparate. Overall gene ontology revealed metabolic and cellular enriched functional processes but numerous pro-inflammatory and immune associated genes (Cxcl2, Cxcl3, Cxcl5, Acod, Hamp, c-Lectins, for example), keratin associated genes (Krt6b and various Krtaps), and members of the Sprr and Lce family, which promote skin barrier integrity and keratinocyte functions, were also differentially modulated. ConclusionsThe altered expression of these genes may provide a potential mechanism to explain the increased ICD severity in IL-6-deficient mice. Overall, this study offers new insight into the pathogenesis of ICD, indicates new mediators/biomarkers that may influence the variability of responses to irritants and provides potential targets for therapeutic development.

Giardia Infection of the Small Intestine Induces Chronic Colitis in Genetically Susceptible Hosts.

The lumen-dwelling protozoan Giardia is an important parasitic cause of diarrheal disease worldwide. Infection can persist over extended periods with minimal intestinal inflammation, suggesting that Giardia may attenuate host responses to ensure its survival, although clearance eventually occurs in most cases. IL-10 is an anti-inflammatory regulator critical for intestinal homeostasis and controlling host responses to bacterial exposure, yet its potential role in coordinating antiprotozoal host defense in the intestine is not known. In this study, we found that murine infection with the natural enteric pathogen Giardia muris induced a transient IL-10 response after 2-4 wk at the primary site of infection in the upper small intestine, but parasite colonization and eradication were not affected by the absence of the cytokine in gene-targeted mice. However, IL-10 was critical for controlling infection-associated immunological sequelae in the colon because severe and persistent diarrhea and colitis were observed in IL-10-deficient mice within 1-2 wk postinfection but not in uninfected littermate controls. Inflammation was characterized by epithelial hyperplasia, neutrophil and macrophage expansion, and Th1 induction and could be prevented by blockade of IL-12/IL-23 p40 but not depletion of CD11c+ dendritic cells. Furthermore, the intestinal microbiota underwent characteristic shifts in composition and was required for disease because antibiotics and loss of TLR signaling in MyD88-deficient mice protected against colitis. Together, our data suggest that transient infection by a luminal and seemingly noninflammatory pathogen can trigger sustained colitis in genetically susceptible hosts, which has broader implications for understanding postinfectious syndromes and other chronic intestinal inflammatory conditions.

AMPA-ergic regulation of amyloid-\u03b2 levels in an Alzheimer\u2019s disease mouse model

BackgroundExtracellular aggregation of the amyloid-β (Aβ) peptide into toxic multimers is a key event in Alzheimer’s disease (AD) pathogenesis. Aβ aggregation is concentration-dependent, with higher concentrations of Aβ much more likely to form toxic species. The processes that regulate extracellular levels of Aβ therefore stand to directly affect AD pathology onset. Studies from our lab and others have demonstrated that synaptic activity is a critical regulator of Aβ production through both presynaptic and postsynaptic mechanisms. AMPA receptors (AMPA-Rs), as the most abundant ionotropic glutamate receptors, have the potential to greatly impact Aβ levels.MethodsIn order to study the role of AMPA-Rs in Aβ regulation, we used in vivo microdialysis in an APP/PS1 mouse model to simultaneously deliver AMPA and other treatments while collecting Aβ from the interstitial fluid (ISF). Changes in Aβ production and clearance along with inflammation were assessed using biochemical approaches. IL-6 deficient mice were utilized to test the role of IL-6 signaling in AMPA-R-mediated regulation of Aβ levels.ResultsWe found that AMPA-R activation decreases in ISF Aβ levels in a dose-dependent manner. Moreover, the effect of AMPA treatment involves three distinct pathways. Steady-state activity of AMPA-Rs normally promotes higher ISF Aβ. Evoked AMPA-R activity, however, decreases Aβ levels by both stimulating glutamatergic transmission and activating downstream NMDA receptor (NMDA-R) signaling and, with extended AMPA treatment, acting independently of NMDA-Rs. Surprisingly, we found this latter, direct AMPA pathway of Aβ regulation increases Aβ clearance, while Aβ production appears to be largely unaffected. Furthermore, the AMPA-dependent decrease is not observed in IL-6 deficient mice, indicating a role for IL-6 signaling in AMPA-R-mediated Aβ clearance.ConclusionThough basal levels of AMPA-R activity promote higher levels of ISF Aβ, evoked AMPA-R signaling decreases Aβ through both NMDA-R-dependent and -independent pathways. We find that evoked AMPA-R signaling increases clearance of extracellular Aβ, at least in part through enhanced IL-6 signaling. These data emphasize that Aβ regulation by synaptic activity involves a number of independent pathways that together determine extracellular Aβ levels. Understanding how these pathways maintain Aβ levels prior to AD pathology may provide insights into disease pathogenesis.

Overt Increase of Oxidative Stress and DNA Damage in Murine and Human Colitis and Colitis-Associated Neoplasia.

Patients with inflammatory bowel disease (IBD) have a higher risk of developing colitis-associated-cancer (CAC); however, the underlying processes of disease progression are not completely understood. Here, the molecular processes of inflammation-driven colon carcinogenesis were investigated using IL10-deficient mice (IL10 KO). IL10 KO mice were euthanized after development of colitis and dysplasia. IHC was performed for markers of colitis-induced DNA damage (CIDD): oxidative DNA lesions (8-oxoG), double-strand breaks (DSB; γH2AX). and DSB repair. MSI, LOH (Trp53, Apc), and global methylation (CIMP) were assessed on microdissected tissue. Comet assay for DNA damage, immunofluorescence, and immunoblotting were performed on intestinal organoids from wild-type (WT) and IL10 KO mice. Sequential biopsies and surgical specimens from IBD and CAC patients were used for IHC analysis. Severity of inflammation correlated with number of dysplasia. 8-oxoG and γH2AX-positive cells were significantly increased in inflamed and dysplastic areas along with activation of DSB repair. The amount of positively stained cells strongly correlated with degree of inflammation (8-oxoG: R = 0.923; γH2AX: R = 0.858). Neither CIMP, MSI nor LOH was observed. Enhanced DSBs in IL10 KO organoids were confirmed by comet assay and increased expression of γH2AX. Human clinical specimens exhibited significantly higher γH2AX and 8-oxoG in IBD, dysplasia, and CAC compared with normal mucosa. These data indicate that inflammation-driven colon carcinogenesis in IL10 KO mice and IBD patients is associated with oxidative DNA damage and overt presence of DSB. Mol Cancer Res; 16(4); 634-42. ©2018 AACR.