bluuebunny/crossref_metadata_2025 huggingface dataset:
The landscape of immune responses and persistent viral reservoirs in systemic and lymphoid tissues in rhesus macaques infected by a newly developed SHIV-CH848
Authors: XU HUANBIN , Ziani Widade , Wang Xiaolei , Veazey Ronald S.
Published: January 2023

Background

The chimeric simian/human immunodeficiency viruses (SHIVs), carrying HIV envelope glycoproteins from transmitted founder (T/F) viruses, are valuable as a challenge viruses in nonhuman primate models for validating HIV-1 vaccines, assessing viral reservoirs and latency, and developing functional “cure” strategies in HIV infection. This study comprehensively investigated immunological changes, viral replication and reservoir size in deep tissues of SHIV-CH848-infected rhesus macaques on antiretroviral therapy (cART).

Material & Method

A total of 10 rhesus macaques (Macaca mulatta, RMs) were intravenously (i.v.)inoculated with SHIV-CH848, containing vpu-env(gp140) sequence from T/F HIV-1 subtype C strain CH848. At 3 months post SHIV infection, 5 animals received cART for 6 months, and remaining 5 animals as untreated controls. Blood, lymph node and rectal biopsies were collected to examine plasma viral load, cell-associated viral RNA/DNA and host immune responses.

Results

SHIV-CH848 inoculation resulted in rapid viral replication, reaching peak viremia at 14dpi. SHIV-CH848 primary infection significantly depleted peripheral and mucosal CD4+ T cells. Also, broadly neutralizing antibodies were also not elicited up to 15 months post infection. Despite gradual recovery of CD4+ T cells after cART, the levels of integrated viral DNA in tissues remained stable throughout 6-months of cART, and rapid viral rebound was observed after ART interruption.

Conclusions

These findings suggest that the new SHIV strain, SHIV-CH848, is able to maintain persistent viral replication in macaques, and establish long-term viral reservoirs/latency in tissues, serving as a valuable model for HIV research.
N6-methyladenosine modification enables viral RNA to escape recognition by RNA sensor RIG-I
Authors: Lu Mijia , Zhang Zijie , Xue Miaoge , Zhao Boxuan Simen , Harder Olivia , Li Anzhong , Liang Xueya , Gao Thomas Z , Feng Zongdi , Niewiesk Stefan , Peeples Mark E. , He Chuan , Li Jianrong
Published: January 2023

Internal N6-methyladenosine (m6A) modification of RNA is one of the most abundant modifications in eukaryotic cells as well as in viruses. However, the biological role(s) of RNA m6A in virus-host interaction remains elusive. Using human metapneumovirus (hMPV), a medically important non-segmented negative-sense RNA virus as a model, we demonstrate that m6A serves as a molecular marker for innate immune discrimination of self from nonself RNAs. We show that hMPV RNAs are m6A methylated and that viral m6A methylation promotes hMPV replication and gene expression. Inactivating these m6A sites with synonymous mutations resulted in m6A deficient recombinant hMPVs that induced significantly higher expression of type I interferon (IFN) that restricted viral replication. Notably, the induction of type I IFN by m6A-deficient rhMPVs and virion RNA was dependent on the cytoplasmic RNA sensor RIG-I, not MDA5. Using RIG-I pull down and RIG-I limited digestion assay, we mechanistically demonstrated that m6A-deficient virion RNA exposed to cytoplasm has much high accessibility to the binding of RIG-I, which allows RIG-I’s translocation along the RNA strand and subsequential conformational change, oligomerization, activation of IFN signaling cascade, and then even higher RIG-I expression during anti-virus status. Furthermore, m6A-deficient rhMPVs triggered higher type I IFN in vivo and were significantly attenuated in the lower respiratory tract yet retained high immunogenicity in cotton rats. Collectively, our results highlight that (i) virus acquires m6A in their RNAs as a means of mimicking cellular RNA to avoid the detection by innate immunity; and (ii) viral m6A RNA can serve as a novel target to attenuate hMPV for vaccine purposes.
A Brazilian Zika virus isolate preferentially induces T follicular helper cell responses while suppressing Th1 immunity
Authors: Pardy Ryan D , Gentile Maria E , King Irah L , Richer Martin J
Published: January 2023

Zika virus (ZIKV) is a mosquito-borne pathogen that caused a series of increasingly severe outbreaks in Micronesia, French Polynesia, and South and Central America. Recent work from our laboratory has shown that, compared to a pre-epidemic isolate (ZIKV-CDN), a Brazilian ZIKV isolate (ZIKV-BR) possesses a novel capacity to suppress antigen-specific CD8 T cell responses, resulting in sustained infection. However, it is unknown whether ZIKV-BR also modulates CD4 T cell immunity. Thus, we investigated the CD4 T cell response to infection with each ZIKV isolate. Our data demonstrate that the CD4 T cell response to ZIKV-BR is reduced in magnitude compared to the response induced by ZIKV-CDN. Further, CD4 T cells are less polarized to the Th1 subtype, express less T-Bet, and are functionally impaired, as they produce less IFN-γ following ex vivo restimulation. Although we observed no alterations in the Th2, Th17 or T regulatory cell compartments, we observed a striking accumulation of PD-1hiCXCR5+ T follicular helper (Tfh) cells 10 days post-infection with ZIKV-BR. This response correlated with an enhanced germinal center B cell response, and increased detection of germinal center formation by confocal microscopy. Future studies will aim to determine the mechanism through which ZIKV-BR infection enhances the Tfh response, and the implications of promoting Tfh responses while dampening Th1 responses for antiviral immunity. Together, our data suggest that contemporary ZIKV strains have evolved to modulate CD4 T cell responses and this could provide a model for interrogating the signals required for Tfh development.
Risk assessment test for MAR ASD: a subtype of Autism
Authors: Celis Nora Alexandra Ramirez , Van de Water Judy
Published: January 2023

Background

Autism spectrum disorder (ASD) is an important health issue and affects 1 in 59 children in the US. Prior studies found that maternal autoantibody related autism (MAR ASD) is thought to be responsible for ~20% of ASD cases. We aimed to create a robust assay to identify ASD-specific autoantibody patterns of reactivity against the previously identified brain antigens CRMP1, CRMP2, GDA, LDHA, LDHB, NSE, STIP1, and YBOX1 in the blood of mothers whose children have ASD and determine their relationship with ASD behavioral outcome.

Methods

We obtained plasma enrolled in the CHARGE study from mothers of children diagnosed with ASD (n=481) and control mothers of typically developing children (TD, n=377). Maternal plasma was probed against the eight antigens by ELISA and WB. We then determined patterns of reactivity that had a strong association with ASD by permutation analysis.

Results

We found several ASD specific patters of reactivity (positives for several antigens) in 16% of the ASD samples and 0% in the TD group. From those, 4 patterns of reactivity were statistically significant using Fisher’s exact test 0.05% showing a significant association of having autoantibodies against a combination of those antigens and a higher risk of having a child with MAR ASD. Also, preliminary studies indicate a positive correlation between having reactivity to multiple antigens and higher Autism Severity Scores (ADOS composite).

Conclusion

We developed the first risk assessment test to predict ASD with 100% specificity based on reactivity to different combinations of brain antigens. We are currently working on the development of multiple MAR ASD animal models.
Insights into the role of Epstein-Barr virus infection in multiple sclerosis using a novel humanized mouse model of disease
Authors: Allanach Jessica R. , Hardman Blair K , Fettig Naomi M , Mouat Isobel , Gu Yu , Jean-Baptiste Virginie , Shanina Iryna , Osborne Lisa C , Vorobeychik Galina , Horwitz Marc S
Published: January 2023

Multiple sclerosis (MS) is an autoimmune disease of the central nervous system without a clear cause. Epstein-Barr virus (EBV) is proposed to contribute to the pathogenesis of MS through unknown mechanisms. Evidence for a role of EBV infection in MS comes from both epidemiological and experimental studies, however, due to its narrow host tropism, there are currently few suitable animal models of MS that incorporate EBV infection. Recent advancements in humanized mouse modelling has enabled direct infection for the study of EBV-associated malignancies. We therefore chose to evaluate the role of EBV infection in humanized mice with experimental autoimmune encephalomyelitis (EAE), a widely used model of MS. Immunocompromised mice were engrafted with peripheral blood mononuclear cells (PBMC) derived from individuals with relapsing MS (RRMS) or from matched healthy EBV seropositive or seronegative donors. We observed that HuPBMC mice induced with EAE developed ascending paralysis, weight loss and signs of discomfort consistent with classical EAE models. Further, HuPBMC EAE showed significant T cell infiltration of both the brain and spinal cord, notably of IFN gamma-expressing CD4 and CD8 T cells, resulting in spinal cord and cerebellar demyelination. HuPBMC EAE mice derived from EBV seropositive donors developed earlier disease onset with more severe clinical symptoms compared to EBV seronegative donor-derived mice. We also observed increase disease severity among mice derived from RRMS patients compared to healthy controls. With continued improvement and characterization of this novel humanized EAE model, additional environmental and genetic risk factors can be evaluated in a system with human immune-mediated pathology.
Macrophage cannabinoid 1 receptor promotes obesity-induced insulin resistance and inflammation
Authors: Han Ji Hye , Kim Wook
Published: January 2023

NLRP3 inflammasome plays a crucial role in immune responses, promoting the maturation of proinflammatory cytokines interleukin 1β (IL-1β) and IL-18, and has been associated with a variety of inflammatory conditions, including obesity and type 2 diabetes. Our recent studies have figured that inhibition of cannabinoid 1 receptor (CB1R) improves metabolic abnormalities and suppresses macrophage infiltration into white adipose tissue, activation of NLRP3 inflammasome, and production of proinflammatory cytokines. Macrophages are implicated in this process. To test the role of CB1R, we established mice with selective CB1R-defeciency in macrophages (MacCB1R KO). We investigate that feeding a Western-type diet to control and MacCB1R KO mice caused similar obesity, but only mice with MacCB1R KO remained insulin sensitive. The protection of mice with macrophage-specific CB1R-deficiency against insulin resistance was associated with reduced tissue infiltration by macrophages. Immunophenotypic parameters have been proved that CB1R is involved for pro-inflammatory macrophage polarization. These studies demonstrated that CB1R in macrophages is associated to establish obesity-induced insulin resistance and inflammation.
The role of Interleukin-22 and its receptor Interleukin-22Rα on the regulation of inflammatory responses in the brain
Authors: Lee DaHae , Kim Yejin , Jo Hyejung , Kang Jae Seung
Published: January 2023

Interleukin (IL)-22 is a member of IL-10 family, is a potent mediator of inflammatory responses. It is produced by activated CD4+ T cells and natural killer (NK) cells and takes effect on non-hematopoietic cells mainly stromal and epithelial cells. And, IL-22 receptor consists of IL-22Rα and IL-10Rβ. It is known that IL-22Rα expression is restricted to non-hematopoietic cells in the skin, pancreas, intestine, liver, lung and kidney. Even though IL-22 is mainly involved in the development of inflammatory responses, but there is a no report regarding their roles on inflammatory responses in the brain. In the present study, I investigated the role of IL-22 and its receptor on inflammatory responses in the brain using mouse microglia cell line, BV2 and mouse hippocampal neuronal cell line, HT22. When BV2 and HT22 were treated with rIL-22 (20 ng/ml), expression of cyclooxygenase (COX)-2, was increased and its expression is followed by the increased of PGE2 production. Next, I examined whether production of pro-inflammatory cytokines, such as such as IL-6 and TNF-α, and pro-inflammatory chemokines by IL-22 treatment. As a result, I confirmed that IL-6, TNF-α, and MIG/CXCL9 are remarkably increased. Taken together, IL-22α is spontaneously expressed on the cells, especially microglia and neuron and it is closely involved in the development of inflammatory responses in the brain.
Role of C-C Motif Chemokine Ligand 20 in the Regulation of Epidermal γδ T Cells
Authors: Bshara-Corson Savannah , Gonzalez Isaac , Jameson Julie M
Published: January 2023

The epidermis provides a barrier that protects against environmental damage and infection by pathogens. Epidermal T cells and keratinocytes work together to maintain skin homeostasis upon damage. During tissue repair, epidermal γδ T cells become activated to secrete cytokines, chemokines and growth factors to initiate inflammatory responses. Previous work in our lab has shown that during obesity and type 2 diabetes, Chemokine Receptor 6 (CCR6) is downregulated by murine epidermal T cells. The ligand for CCR6, C-C Motif Chemokine Ligand 20 (CCL20), is produced by keratinocytes and is responsible for cellular migration. Little is known about how CCR6 and CCL20 ligation affects epidermal γδ T cells. In this study, murine epidermal γδ T cells were harvested and stimulated in the absence or presence of CCL20 to discover how CCL20 affects activation and function of the cells. Epidermal γδ T cells that have been stimulated with anti-CD3ɛ and CCL20 downregulate their T cell receptor and upregulate CD25 showing cell activation. Interestingly, CCL20 costimulates T cell receptor stimulation to activate epidermal γδ T cells. Immunofluorescent microscopy was used to visualize the impact of CCL20 on activated cells verses non-activated cells in situ. Epidermal γδ T cells treated with CCL20 did not migrate within or out of the epidermis unlike previous studies on dermal γδ T cells. Flow cytometric analysis and Luminex assays were performed to determine which cytokines and chemokines are produced in response to CCL20. We show that CCL20 skews epidermal γδ T cells to produce pro-inflammatory cytokines and chemokines. Future studies will determine how CCL20 and CCR6 ligation affects murine models with obesity and type 2 diabetes.
CXCL13-CXCR5 signaling and its co-expression network are associated with colorectal cancer cell growth and poor prognosis
Authors: Nunez Sha’Kayla K , Griffen Tiara L , Young Corey D , Wu Ji Yuan , Davis Jennifer , Kopetz Scott , Lillard James W
Published: January 2023

The heterogeneity of colorectal cancer (CRC) and its microenvironment has molecular traits of immune cells, and is driven by dysregulated WNT, RAS-RAF-MAPK, TGF-β, and PI3K-AKT pathways. CXCL13-CXCR5 signaling has been shown to induce PI3K- and Ras-dependent activation of MAPK/ERK1-2 to promote cancer cell survival, invasion, and growth. The hypothesis tested in this study is CXCL13-CXCR5 signaling and co-expression networks mediate CRC progression, cell growth, and survival. RNAseq data from CRC cases, along with matched controls, were acquired from The Cancer Genome Atlas (TCGA). Weighted gene co-expression network analysis (WGCNA) elucidated gene networks that significantly correlated with CRC poor prognosis. CXCL13, CXCR5, and other co-expressed genes were identified in the WGCNA designated brown module (M3). The canonical pathways and their upstream regulators of this module were analyzed using Ingenuity Pathway Analysis (IPA). M3 hub genes XCL1, CCL19, PIK3AP1, FLT3, and WNT1 were significantly and positively correlated with PI3K, WNT, ERK1 and ERK2 activation, which promotes cell proliferation and survival. Next, we measured the CRC cell line growth effected by CXCL13 using a XTT assay. In confirmation, CXCL13 treatment of CRC cell lines significantly increased CRC cell line proliferation, but had no direct influence on cell survival. Our findings strongly support CXCL13-CXCR5 signaling and its co-expression networks activates important pathways associated with CRC progression and tumor growth.
PD-1/PD-L1 signaling regulates the thymic niche size of agonistly selected T cells
Authors: Moran Amy , Emlen Ester , Caruso Breanna
Published: January 2023

Effective peripheral T cell responses rely upon a diverse T cell repertoire, generated during thymic selection. Signaling through the TCR establishes thresholds for thymocyte selection in order to create this repertoire. Because co-stimulation modifies TCR signaling, it may contribute to the regulation of T cell development by determining signal strength thresholds that direct thymocytes into specific lineages. Programmed death-1 (PD-1), a member of the B7/CD28 family of costimulatory/inhibitory receptors, has been implicated in the development of thymocytes because it is expressed in CD4− CD8− double-negative thymocytes during TCRb rearrangement. Further, the ligands for PD-1, PD-L1 and PD-L2, are expressed on thymic epithelial cells. PD-L2 is restricted to the medulla whereas PD-L1 is expressed throughout the thymus. Here, we show that PD-1 modulates agonist selection, a process in which some high-affinity thymocytes will mature into unconventional T cell lineages (e.g. regulatory T cells, NKT cells). Transgenic mice lacking either PD-L1 or PD-1 have increased numbers of agonistly selected T cells compared to wildtype animals. Interestingly, loss of PD-L1 increases the selection of conventional and agonistly selected T cells whereas loss of PD-1 limits the phenotype to agonist lineages. These data demonstrate that PD-1, PD-L1, and possibly PD-L2 interactions, potentially within the thymic microenvironment in which they are expressed, play a role in determining T cell fate and shaping the final T cell repertoire.
Vacuolar ATPase a2-subunit is a multifocal regulator of T cell development
Authors: Peterson Theodore V , Reynolds Joseph M
Published: January 2023

T cells are required for nearly all aspects of the adaptive immune response, and as such, their development into functional maturity is a complex and tightly controlled process. Developing thymocytes must coordinate a host of transcriptional, metabolic, and proliferative changes as they mature, and disturbances in this process underlie a number of diseases and syndromes, including cancers, immunodeficiencies, and autoimmunity. Interestingly, our recent work has identified an essential role for the a2-subunit isoform of the vacuolar ATPase (a2V) – a highly conserved intracellular proton pump – in orchestrating critical changes at multiple points of murine αβ T cell development. Cre-recombinase mediated deletion of a2V restricted to the hematopoietic compartment or immature thymocytes leads to a profound peripheral lymphopenia specifically confined to CD4+ and CD8+ αβ T cells, a condition originating from several deficiencies in intrathymic T cell development. Thymuses of a2V-deficient mice are reduced in size and cellularity, observations temporally correlated with the timing of a2V deletion. These deficiencies are in part tied to perturbations within the Notch1 signaling pathway, and obstructions at the β-selection checkpoint and a near complete failure of positive selection suggest further deficiencies in proliferative and TCR-mediated pathways, and therefore a broad and multifaceted role for a2V during early T cell development. Importantly, these data support a recent focus on V-ATPase as a chemotherapeutic target, and furthermore suggest that a2V may be a specific and efficacious target for combating proliferative potential of autoreactive or malignant T cells.
Single cell RNAseq analysis reveals profound abnormalities in the distribution and diversity of thymic epithelial cells in Rag1 mutant mice
Authors: Pala Francesca , Martins Andrew , Oguz Cihan , Lack Justin , Tsang John , Notarangelo Luigi D , Bosticardo Marita
Published: January 2023

The thymus contains a heterogeneous population of stromal cells which orchestrate T cell development and selection. Recent works highlighted the complexity of thymic stromal cells, and medullary thymic epithelial cells (mTECs) in particular. However, the developmental origin, hierarchy, and function of these subpopulations remain ill-defined. Moreover, the characterization of cortical TECs (cTECs) has been largely restricted to the adult thymus. Here, we compared TEC distribution and gene expression in wild-type (WT) and in mice carrying Rag1 hypomorphic mutations observed in patients with immune deficiency and immune dysregulation.

Single cell RNA-seq analysis of TECs isolated from adult Rag1 mutant mice revealed an excess of cTECs, which segregated in different clusters. The mTEC compartment, albeit reduced, showed a similar distribution of previously described subsets (mTEC I–IV), suggesting perturbation of mTEC development rather than mTEC differentiation into functional subsets. To address whether such abnormalities of cTEC and mTEC abundance and subset distribution in Rag1 mutant mice may reflect defects in TEC development, we extended scRNA-seq analysis to TECs from WT mice of neonatal age. A similar pattern was observed in TECs of adult Rag1 mutant mice and WT mice of neonatal age, indicating altered TEC development when Rag1 activity is perturbed. This defect correlates with a decreased number of double-positive thymocytes, suggesting that impaired lymphostromal cross-talk in the thymus of Rag1 mutant mice (and patients with similar defects) is associated with abnormalities of TEC composition which may contribute to altered immune tolerance that is often observed in these conditions.
Meningeal lymphatic dysfunction exacerbates traumatic brain injury pathogenesis
Authors: Bolte Ashley C , Hurt Mariah E , Smirnov Igor , Dutta Arun B , Kovacs Michael A , McKee Celia A , Natale Nick , Ennerfelt Hannah E , Nguyen Bao H , Frost Elizabeth L , Lammert Catherine E , Kipnis Jonathan , Lukens John R
Published: January 2023

Traumatic brain injury (TBI) has emerged as a leading cause of death and disability. Despite being a growing medical issue, the biological factors that promote central nervous system (CNS) pathology and neurological dysfunction following TBI remain poorly characterized. Recently, the meningeal lymphatic system was identified as a critical mediator of drainage from the CNS. In comparison to other peripheral organs, our understanding of how defects in lymphatic drainage from the CNS contribute to disease is limited. It is still unknown how TBI impacts meningeal lymphatic function and whether disruptions in this drainage pathway are involved in driving TBI pathogenesis. Here we demonstrate that even mild forms of brain trauma cause severe deficits in meningeal lymphatic drainage that can last out to at least two weeks post-injury. To investigate a mechanism behind impaired lymphatic function in TBI, we examined how increased intracranial pressure (ICP) influences the meningeal lymphatics, as increased ICP commonly occurs in TBI. We demonstrate that increased ICP is capable of provoking meningeal lymphatic dysfunction. Moreover, we show that pre-existing lymphatic dysfunction mediated by targeted photoablation before TBI leads to increased neuroinflammation and cognitive deficits. These findings provide new insights into both the causes and consequences of meningeal lymphatic dysfunction in TBI and suggest that therapeutics targeting the meningeal lymphatic system may offer strategies to treat TBI.
Characterization of the microglia in three non-overlapping lysosomal storage diseases
Authors: cougnoux antony , Drummond Rebecca , Fellmeth Mason , Navid Fatemeh , Collar Amanda , Iben James , Cawley Niamh , Wassif Christopher , Pickel James , Kulkarni Ashok , Lionakis Michail , Porter Forbes
Published: January 2023

Lysosomal storage diseases (LSDs) are a large family of inherited disorders characterized by abnormal endolysosomal accumulation of cellular material due to catabolic enzyme or transporter deficiencies. Depending on the affected metabolic pathway, these diseases manifest with somatic or central nervous system signs and symptoms. Neuroinflammation is a hallmark feature of LSDs with neurologic involvement such as mucolipidosis type IV (Mcoln1) or Niemann-Pick disease, type C1 (Npc1), but not Fabry disease (Gla). We investigated the properties of microglia from these three disorders using mouse models and a combination of flow cytometric, RNA sequencing, biochemical and immunofluorescence analyses. Stored material is present in the microglia of the different mice however only in Niemann-Pick disease, type C1 did we observe that neuroinflammation increases with disease progression. Fabry disease mice do not have neurological manifestation, and this is reflected by the minor changes in microglial properties. Lastly, mucolipidosis type IV mice have an early gliosis which decreases with disease progression. Comparison among microglia transcriptome profiles form these 3 diseases, and other disorders where transcriptome data is available, showed similarities with other neurodegenerative diseases.

The phenotypic similarities between microglia from rare monogenic disorders, where the primary metabolic disturbance is known, and common neuroinflammatory diseases may provide novel insights for therapeutic intervention.
The role of myeloid cell-specific ten eleven translocase 2 (TET2) in regulation of neuroinflammation
Authors: Munie Ashley N , Segal Benjamin
Published: January 2023

Multiple Sclerosis (MS), is an inflammatory demyelinating disease of the central nervous system (CNS) believed to be autoimmune in etiology. Analysis of legions from MS patients, as well as studies in Experimental Autoimmune Encephalomyelitis (EAE), a murine model of MS, have indicated that myeloid cells play a critical role in the pathogenesis of CNS autoimmunity. Recently, Ten-Eleven Translocation 2 (TET2) was identified as a susceptibility locus for MS. TET2 is a Fe(III), α-ketoglutarate dependent enzyme that catalyzes the oxidation of methylated cytosine to 5′-hydroxymethyl cytosine (5hmC), a stable epigenetic mark. There is growing evidence that TET2 acts as a negative regulator of myeloid cell activation. Further, TET2 and 5hmC are reduced in peripheral blood mononuclear cells of MS patients compared with healthy volunteers. The objective of the current study is to investigate the hypothesis that reductions of TET2 expression and activity in myeloid cells is a critical step in the development of pro-inflammatory myeloid cells during CNS autoimmunity. Our laboratory has found that Tet2 transcripts levels and 5hmC content are reduced in CNS-infiltrating myeloid cells at the peak of EAE compared with the onset or late stages. Adoptive transfer of encephalitogenic CD4+ Th17 cells into Tet2−/+ mice lead to an exacerbated clinical course compared with their Tet2+/+ counterparts. Our data suggests that TET2 expression and activity is dysregulated in myeloid cells and limits neuroinflammation and chronic neurological disability. Interventions that promote Tet2 expression in myeloid cells may ameliorate EAE and, by extension, MS.
Age-Dependent Synthesis of Diverse Autoantibodies in the 3xTg-AD Model of Alzheimer’s Disease
Authors: Ma Donglai , Song Wei , Kapadia Minesh , Fahnestock Margaret , Sakic Boris
Published: January 2023

Elevated levels of serum autoantibodies are a well-documented phenomenon in patients with Alzheimer’s disease (AD), but the nature of the autoimmunity is unclear. Triple-transgenic (3xTg-AD) mice are a well-established model that develops AD-like pathology. We observed that recent cohorts of male 3xTg-AD mice exhibit autoimmune manifestations, yet lack plaque/tangle pathology. In the current study, we identified the time course and type of autoantibodies produced during the progression and attenuation of systemic autoimmunity. Tissue samples were collected from 3xTg-AD and wild type (WT) males and females at 2, 6 and 12 months of age. Serum autoantibodies were measured by both indirect immunofluorescence and 16-antigen line immunoassay. To assess immunosuppression, serum samples from a separate cohort of mice chronically exposed to cyclophosphamide were also assessed. Several autoantibody types increased in frequency and concentration in aging 3xTg-AD mice, particularly in adult males. The most prevalent were anti-nuclear autoantibodies, particularly those targeting the nucleosomes. Among the panel of autoantibodies measured, reactivity to a single antigen (nucleosomes) was observed in the majority of samples. A few mice showed reactivity to other autoantigens, such as histones, dsDNA, RNP/Sm, AMA-M2 and rib. P. Reactivity to all autoantigens was abolished in mice exposed to the immunosuppressant cyclophosphamide. The emergence of circulating autoantibodies similar to classical autoimmune diseases (such as lupus) suggests an important, yet unreported aspect of pathology in this AD model. Future studies are aimed to elucidate the origin and role of the autoantibodies in the context of phenotype loss in 3xTg-AD males.
Linking environmental exposures to molecular regulation of inflammation via epigenetic histone signatures in an exacerbation prone pediatric asthma cohort
Authors: Knapp Jennifer R , Harmacek Laura D , Crooks James , Tu Ting-Hui Clair , Magallon Roman E , Leach Sonia M , Danhorn Thomas , Schiltz Allison , Voelker Dennis R , Siebold Max A , Thornburg Jonathan W , Fingerlin Tasha E. , Liu Andrew W , O’Connor Brian P.
Published: January 2023

Environmental exposures, such as air pollution, are correlated with asthma susceptibility and airway inflammation. Epigenetic mechanisms can translate these exposures to regulation of molecular pathways that drive asthma pathogenesis. Indeed, specific epigenetic modifications have been linked to regulation of asthma-associated inflammation. Yet, there have been no studies linking environmental exposures to direct regulation of molecular mechanisms driving dysfunction of inflammation in pediatric asthma. Here we examined a cohort of exacerbation prone pediatric asthmatics tracking exposure through multiple air monitoring methods, including personalized vest monitors, stationary monitors, and Colorado DPHE monitors. We performed ChIP-seq examining histone modifications in PBMCs and integrated the epigenetic data with the environmental exposure data from the air pollution monitors. Our results statistically linked specific histone modifications in PBMCs to specific types of environmental exposures within the pediatric asthmatics. Annotation of these histone modifications to proximal genes identified a potential immune program regulating asthma inflammation. Concomitant examination of the PBMC transcriptome enabled functional analysis of epigenetic regulation of these immune genes. The resultant molecular profiles can be used to identify biomarkers for patient stratification and disease course prediction. Moreover, these studies identify a potential link between the molecular programming associated with environmental exposures and regulation of key biological pathways, which alter immune cell activity that ultimately can increase asthma exacerbation in children.
Human and murine eosinophils differ in their ability to synthesize key bioactive lipids involved in asthma
Authors: Archambault Anne-Sophie , Brassard Julyanne , Bernatchez Émilie , Martin Cyril , Di Marzo Vincenzo , Laviolette Michel , Boulet Louis-Philippe , Blanchet Marie-Renée , Flamand Nicolas
Published: January 2023

CONTEXT

High eosinophil (EOS) counts is a key feature of asthma. EOS notably affect the asthmatic response by the mean of their lipidome. Mouse models have been developed in the hope of defining new pharmacological targets to treat asthma. However, many pinpointed targets could not be translated into clinics, underscoring that key differences exist between mice and humans. Herein, we compared the ability of human (h) and mouse (m) EOS to synthesize key bioactive lipids from arachidonic acid (AA) and docosahexaenoic acid (DHA).

RESULTS

In response to AA, hEOS synthesized both LTC4 and LTB4 while mEOS only synthesized LTB4. The synthesis of the 15-lipoxygenase products 15-HETE and 12-HETE also differed, hEOS producing them at a 5:1 ratio while mEOS producing them at a 1:2 ratio. mEOS consistently synthetized more PGE2 than hEOS. As for DHA, mEOS produced ten times the levels of specialized pro-resolving mediators (resolvins, maresins and protectins) than hEOS. In contrast, hEOS produced important amounts of the endocannabinoid 2-AG and its congener 2-DHG, a biosynthetic pathway that was 10 – 50 times less important in mEOS.

CONCLUSIONS

Our data show that the lipidomes of hEOS and mEOS differ, notably by their production of leukotrienes, their opposite 15-HETE/12-HETE ratio, their ability to synthesize specialized pro-resolving mediators (mEOS > hEOS), and the inability of mEOS to synthesize anti-inflammatory endocannabinoids. We thus propose that in mouse models of asthma, the impact of LTB4 is likely overrated, the role of specialized pro-resolving mediators amplified, and the role of endocannabinoids underrated, compared to asthmatics.
The leukotriene B4 receptors contributes to neutrophil-dominant asthmatic airway inflammation via IL-1β and IL-17 production
Authors: Kwak DongWook , Park Donghwan , Jang Jae-Hyun , Kim Jae-Hong
Published: January 2023

The leukotriene B4 is one of the chemo-attractant molecule for neutrophils, its receptors, BLT1/2 may contribute to neutrophil-dominant airway inflammation. Therefore, we investigated whether BLT1/2 signaling was associated with neutrophil-dominant severe asthma. The role of BLT1/2 as well as 5-/12-lipoxygenase in neutrophil-dominant airway inflammation was investigated using pharmacological inhibitors in murine model. Blockade of BLT1/2 significantly suppressed airway inflammation including thickness of airway wall, recruitment of immune cells and IL-17 production in neutrophil-dominant airway inflammation. 5-/12-LOX, upstream of LTB4 receptors was also critical for neutrophil-dominant airway inflammation and synthesis of IL-17. Furthermore, inhibition of 5-/12-LOX, BLT1/2, suppressed NF-κB activation, thus attenuating airway inflammation and production of IL-17, suggesting that NF-κB lies downstream of BLT1/2 in neutrophilic airway inflammation. Our results suggest that ‘5-/12-LOX-BLT1/2-NF-κB’ cascade significantly contributes to neutrophil-dominant airway inflammation through IL-17 synthesis. What is interesting is IL-1β also acts important role in the neutrophil-dominant asthmatic condition with IL-17 production. The high-expression level of IL-1β which is mainly produced by NLRP3 inflammasome activation already reported clinically in the sputum of neutrophilic asthma patients. Consequentially, our studies suggest that the cooperative mechanism of IL-1β and IL-17 considerably acts to neutrophil-dominant airway inflammation and the leukotriene B4 receptors have possibilities to control the severity via regulation effect of IL-1β and IL-17.
Asthmatic airway inflammation was alleviated by limiting the level of lung extracellular ATP
Authors: Huang Yung-An , Kuo Ming-Ling
Published: January 2023

Asthma is a chronic respiratory disease characterized by recurrently attacks of breathlessness and wheezing. ATP accumulation in bronchoalveolar lavage fluid (BALF) of asthmatic subjects indicates that extracellular ATP (eATP) and downstream responses are involved. The level of eATP is tightly controlled by its release and degradation in healthy tissues. Pannexin-1 is a channel contributing to ATP release, and CD39 is membrane bound apyrase contributing to ATP degradation. Cellular stress or inflammatory stimulation leads increased release of ATP and accompanied with the reduction of CD39. The recombinant adeno-associated virus (rAAV) vectors have been developed for efficient gene transfer with low toxic and difference tissue tropisms. Thus, we generated rAAV-CD39 and rAAV-s10Panx1 and examined their efficacy in OVA-mediated asthmatic mice model. The mice were sensitized intraperitoneally and challenged intratracheally with OVA and treated with the vectors three days before OVA challenge. At the end of procedure, some asthmatic and inflammatory cardinal features were examined. Elevated ATP level either in stimulated-culture supernatant and BALF of OVA-sensitized mice was determined. These two treatments downregulated the levels of ATP in BALF. Several asthmatic features, such as eosinophilia, Th2 cytokine production and AHR were decreased in the treatment groups. These treatments altered the T cell response by targeting the danger signal eATP and its metabolites. Taken together, these approaches are able to downregulate the asthma symptoms with the reduction of inflammatory responses in the lungs. Based on these results, eATP can be considered as a potential gene therapy target for asthma.
Clinical and immunological evaluation of cat-allergic asthmatics living with or without a cat
Authors: Wambre Erik , Bajzik Veronique , DeVeaux Michelle , Meier Pretty , Ruddy Marcella , DeBerg Hannah , Wang Claire Q , Farrington Mary , Radin Allen
Published: January 2023

Rationale

We aimed to evaluate the impact of daily cat exposure in cat allergic asthmatics by both clinical symptom and immunological measures.

Methods

Twenty adults with history of cat-induced asthma and rhinitis, positive serum cat dander-specific IgE (sIgE>0.35kU/L) and Skin Prick Test (SPT) were enrolled at a 1:1 ratio according to cat ownership. For comparison, cat extract- and Fel d1- specific basophil sensitivity test (BST), serum sIgE, sIgG4 and SPT were measured on Day 1 and 28; ambulatory spirometry and symptom measures were obtained daily. Feld1 and 4-reactive CD4+ T cells were profiled using a CD154 upregulation assay.

Results

Cat owners had higher clinical symptom scores & medication use and a trend toward lower FEV1 vs. those not living with cats. Significantly higher levels of cat dander sIgG4 were observed among cat owners, but no significant difference was observed for cat-dander sIgE or SPT. All subjects tested positive on BST to Fel d1 and cat dander. Cat-ownership was associated with reduced basophil sensitivity to Fel d1, but had positive BST to Fel d4 and 7. T-cell response to Fel d1 and 4 were differentially polarized, with Fel d1 responses strongly polarized toward Th2 in both groups. No significant correlation was observed between basophil and T cell responses against cat allergen components.

Conclusions

Cat allergic subjects living with a cat demonstrated reduced pulmonary function and greater clinical symptom severity, despite higher medication use and sIgG4. An immunological difference between cat-owners vs. non cat-owners was detected by basophil assay but not by T-cell response.
Development of allergic airway immunity depends on MARCH1-mediated ubiquitination of MHCII and CD86 in dendritic cells
Authors: Castellanos Carlos A , Ren Xin , Liang Hong-Erh , Laidlaw Brian J , Barczak Andrea , Rudolph Joshua , Eckalbar Walter L , Huang Xiaozhu , Cyster Jason G , Locksley Richard M , Erle David J , Ishido Satoshi , Shin Jeoung-Sook
Published: January 2023

Dendritic cells (DCs) play an essential role in the development of type 2 allergic immunity by presenting allergens to cognate naïve T cells inducing production of IL-4 thus priming type 2 T helper (Th2) cells. However, the molecular mechanisms underlying this function are poorly understood. We found that mice deficient in the expression of MARCH1 ubiquitin ligase in DCs were completely resistant to developing Th2 cell inflammation and eosinophilia upon airway exposure to house dust mite allergens. These mice exhibited normal expansion of allergen-specific CD4+ T cells and normal production of IFN-γ from the T cells but demonstrated a significant defect in the production of IL-4, suggesting a specific role of MARCH1 in DC priming of Th2 cells. Remarkably, mice lacking the ubiquitin acceptor amino acids of the MARCH1 substrates, MHCII and CD86, phenocopied MARCH1-deficient mice, indicating that MARCH1 promotes Th2 cell priming by mediating ubiquitination of MHCII and CD86. Ubiquitination of MHCII was important for CD11b+ DCs to transport respiratory allergens to the mediastinal lymph node whereas ubiquitination of CD86 as well as MHCII was involved in transcriptional reprograming of these DCs. In conclusion, dendritic cells depend on ubiquitination of MHCII and CD86 to condition themselves to effectively migrate to the draining lymph node and induce production of IL-4 from allergen-specific naïve CD4+ T cells.
Maternal Alpha-tocopherol Inhibits Development of Food Allergy Early in Life
Authors: Cook-Mills Joan M , Schroeder-Carter Lindsay M , Ramos-Solís Nicole
Published: January 2023

Food allergy can be life-threatening and has had a marked recent increase in prevalence. Interventions are critical to limit the development of food allergy early in life. We demonstrate that maternal dietary supplementation with α-tocopherol blocks development of food allergy. Briefly, female mice received diet supplemented with α-tocopherol or control chow starting at mating with males with skin barrier gene mutations. The pups, that are heterozygous for skin barrier mutations, received six cutaneous co-exposures to a food allergen and an environmental allergen. Oral challenge with food allergen induced anaphylaxis and increased inflammatory mediators in pups from mothers with control chow. Offspring anaphylaxis and expression of inflammatory mediators were blocked by maternal supplementation with α-tocopherol. In conclusion, maternal supplementation with α-tocopherol reduced neonate development of food allergy. These results have implications for supplementation of allergic mothers with α-tocopherol.
Perinatal androgen exposure impacts mast cell development and sex differences in mast cell-mediated pathophysiology
Authors: Moeser Adam James , Mackey Emily
Published: January 2023

Mast cell (MC)-associated diseases exhibit a sex bias with females often exhibiting more severe symptoms. Further, these sex biases exist in both prepubertal children and adults indicating that mechanisms other than adult sex hormones are involved. In previous studies, we showed that, compared with female MCs, male MCs store and release less histamine which coincided with reduced severity of MC-mediated anaphylaxis, and that these sex differences emerge prior to puberty. The early life mechanisms responsible for the emergence of sex differences in MC-associated disease remain poorly understood. Here we tested the hypothesis that perinatal androgen exposure, a major sexual organization event in males, drives sex differences in MC-mediated pathophysiology. Adult female mice that were previously exposed to high levels of androgens in the perinatal period (E16-PN7) via perinatal testosterone propionate administration had reduced IgE-mediated serum histamine levels and hypothermic responses compared with control females. In contrast, perinatal exposure of males to the endocrine disruptor DEHP, with known anti-androgenic effects, exacerbated IgE-mediated histamine levels and hypothermia. MC-deficient sash mice engrafted with BMMCs from adult perinatally androgenized females, exhibited reduced serum histamine and hypothermia responses compared with WT BMMC engrafted sash controls. Together, these data demonstrate that androgen exposure during the perinatal period shapes MC phenotype and MC-mediated anaphylaxis and thus could be an important determinant of differences in later life MC disease risk between the sexes.
Double Deletion of C5aR1 and C5aR2 during Streptococcal Pneumonia
Authors: Walachowski Sarah , Dudek Markus , Eshkind Leonid , Tansi Vanessa , Mizgerd Joseph P. , Bosmann Markus
Published: January 2023

The complement system is essential for the clearance of encapsulated bacteria such as Streptococcus pneumoniae (Spn), recognized as a major pathogen of pneumonia. Yet, it is not entirely clear whether C5 and its activation product C5a have beneficial or detrimental effects on the outcome of Spn infection. C5a binds to its two homologous C5a receptors (C5aR1 and C5aR2). The extent of functional overlap and role distribution between C5aR1 and C5aR2 remains enigmatic. A critical barrier to allow studies on the role distributions of C5aRs has been the absence of a mouse strain with dual deletion of both receptors - which are encoded as adjacent, linked genes on mouse chromosome 7.

CRISPR/Cas9 guided gene editing was combined with zygote/pronucleus microinjections in C57BL/6J mice to generate a C5aR1/C5aR2 double-knockout strain (C5aR1/2−/−). PCR-based screening and Sanger sequencing confirmed a 12.6 kB deletion of the coding regions of C5aR1 and C5aR2 of the new mouse strain, C57BL/6J-Del( 7C5ar2-C5ar1)1Bosm. Phenotyping of C5aR1/2−/− mice was followed by functional studies including intra-tracheal injections of recombinant mouse C5a or intranasal injections of Spn TIGR4. The airway influx of Ly6G+SiglecF- neutrophils by C5a was abrogated in C5aR1/2−/− mice. Surprisingly, the dual genetic absence of C5a receptors was associated with a higher build-up of neutrophil numbers in alveolar spaces after Spn TIGR4 infection. In a transcriptome-wide screen, FACS-sorted myeloid cells from infected lungs of double and single knockout mice were subjected to RNA-sequencing. Identifying novel differentially expressed genes in the C5aR1/2−/− mice during Spn infection will help to uncover potential synergisms and redundancies of C5aR1 and C5aR2.
Tspan2 is involved in anti-infectious immune responses and CNS autoimmunity
Authors: Scheu Stefanie , Ruland Christina , Lahl Katharina , Zimmer Andreas , Werner Hauke , Förster Irmgard , Sparwasser Tim , Alferink Judith
Published: January 2023

Tetraspanin proteins regulate critical cellular processes in CNS development and immune defence against pathogens. Through lateral association within membrane domains or binding of interaction partners these proteins are involved in pathogen recognition, proliferation and trafficking of immune cells, as well as cell differentiation and motility in the CNS. The tetraspanin family member Tspan2 was originally identified in oligodendrocyte lineage cells. It has been implicated in immunoregulation in the CNS as Tspan2−/− mice show increased microglia and astrocyte activation in the absence of inflammatory stimuli.

A role of Tspan2 in CNS inflammation and anti-infectious immune responses remains to be elucidated. In newly generated Tspan2/DTR/EGFP reporter mice, we detected Tspan2 expression predominantly in mature oligodendrocytes in the CNS and an enhanced clinical severity of experimental autoimmune encephalomyelitis upon immunization with MOG35–55 peptide in Tspan2−/− mice. In peripheral lymphoid organs, Tspan2 expression was found in neutrophils and the majority of dendritic cells (DCs). Tspan2−/− DCs showed reduced uptake of zymosan as compared to wild-type (WT) DCs indicating that Tspan2 affects fungal recognition by DCs. In the absence of Tspan2 neutrophils exhibited a reduced accumulation of reactive oxygen species (ROS) and enhanced migration to inflammatory sites in thioglycollate-induced peritonitis as compared to WT neutrophils. LPS challenge in vivo induced increased production of the proinflammatory cytokines TNF, IL-1β, and IL-6 in the spleens of Tspan2−/− as compared to WT mice. These data provide first evidence that Tspan2 regulates anti-infectious immune responses as well as CNS autoimmunity.