bluuebunny/crossref_metadata_2025 huggingface dataset:
Brain-Thymus communication as a novel immunosuppressive mechanism in the GL261 model of Glioblastoma
Authors: Ayasoufi Katayoun , Khadka Roman H. , Jin Fang , Malo Courtney Sarah , Desai Nidhi , Johnson Aaron J.
Published: January 2023

Glioblastoma multiforme (GBM) is incurable and among the most lethal of cancers. Immunotherapy for GBM is hampered by the fact that GBM patients are severely immunosuppressed with dangerously low T cell counts. Importantly, immunotherapy for GBM will be ineffective without defining the mechanism of immunosuppression. Given that GBM does not metastasize, we investigated the possibility of neuro-immune interactions using the GL261 glioma model in C57BL/6 mice. C57BL/6 mice harboring GL261 gliomas have abnormal thymuses which are acutely and significantly involuted compared to healthy control mice. Glioma bearing mice also have significant thymic atrophy which increases with tumor burden. Further structural analysis of the thymus in glioma bearing mice revealed disrupted thymic epithelial cells and reduced EPCAM+ cell expression of MHC II. Thymic subsets in tumor bearing mice were also developmentally skewed with increased generation of single positive T cells and a block at double negative 2 (DN2) to DN3 transition. Peripherally, MHCII expression was also significantly decreased in blood derived cells in glioma bearing mice. This correlated with an overall reduction in total CD4 T cell numbers. The results of this study demonstrate the GL261 model is recapitulating the immune suppression observed in GBM patients. These results also support the presence of a brain-thymus interaction that could negatively impact the development of immunotherapy strategies to treat GBM.
Therapeutically targeting B7-H3 via chimeric antigen receptors and bispecific killer cell engagers in non-small cell lung cancer
Authors: Zhao Qi , Liu Jie , Yang Shuo
Published: January 2023

Non-small-cell lung cancer (NSCLC) is the most common type of lung cancer. The B7 family antigens, such as PD-1 and its receptor B7-H1 (PD-L1), are considered therapeutic targets in NSCLC. However, little is known about the significance of other B7 family members. In this study, we determine the expression level of B7-H3 protein as a new B7 family protein in cell lines and tumor tissues from NSCLC patients by immunohistochemistry and quantitative RT-PCR. By screening in a phage displaying human antibody library, we have identified fully human antibodies specific for B7-H3 antigen. To explore B7-H3 as a potential target in NSCLC, two forms of antibody therapeutics, chimeric antigen receptor (CAR) T cells and bispecific killer cell engager (BiKE) based on one anti-B7-H3 antibody have been developed. As results, CAR T cells targeting B7-H3 demonstrated the cytotoxicity against NSCLC cells in vitro and in vivo. Meanwhile, BiKE treatment was demonstrated to significantly inhibit cell growth by inducing natural killer cells. The cytotoxicity of two agents will be further investigated in the PDX mouse models. This study suggests B7-H3 as a promising therapeutics target in NSCLC.
Antibodies targeting the intracellular liver stage malaria parasite after infection can be a potent means of reducing parasite liver burden
Authors: Sack Brandon K. , Behet Marije , Mikolajczak Sebastian , Cardamone Hayley , Nguyen Thao , Flannery Erika , Vaughan Ashley M. , Oliver Brian , Vigdorovich Vladimir , Carbonetti Sara , Sather Noah , Scholzen Anja , Sauerwein Robert , Kappe Stefan H.I
Published: January 2023

Malaria is one of the oldest and deadliest diseases known to humans, and in 2017 an estimated 216 million people were infected with 445,000 succumbing to disease. Antibodies which block the skin-to-liver stages of malaria infection continue to be pursued due to their potential to stop infection prior to the progression to the disease and transmission-causing blood stages. Phase III clinical trials with a vaccine targeting the major sporozoite surface protein (circumsporozoite protein) have provided suboptimal efficacy in the field which must be improved before large scale use for malaria prevention and eradication. New antibody targets will likely be needed in order to provide complete, sterilizing protection. Thus far, the search has largely been limited to targeting surface and secreted proteins of the sporozoite, merozoite or gametocyte stages.

Here, we present data demonstrating that antibodies which instead target the intracellular liver stage parasite after hepatocyte infection can be a potent means of limiting parasite liver infection in vivo. This was done in a rodent malaria model by targeting two different proteins found on the parasitophorous vacuole membrane that forms the border between parasite and host cytoplasm—indicating antibodies are entering the hepatocyte to mediate their effect. This phenomenon was also demonstrated using the most common human malaria species Plasmodium falciparum in humanized liver chimeric mice where human antibodies targeting the parasite periphery were able to potently reduce liver burden and parasite liver growth between days 5–6. These data present a new class of antibody targets for malaria and strongly argue for the inclusion of intracellular antigens in novel vaccine formulations.
Chimeric antigen receptors against HIV: Bispecific CD4-based designs for enhanced breadth/potency and minimal immunogenicity
Authors: Hajduczki Agnes , Danielson David T. , Scanlan Aaron W. , Berger Edward A.
Published: January 2023

Chimeric antigen receptor (CAR) technology has potential to achieve durable (life-long?) control of HIV in the absence of antiretroviral drugs, i.e. a “functional cure”. Thus in targeting the HIV-1 Env glycoprotein expressed on the surface of virus-producing cells, CAR design must strive not only for high potency and HIV specificity, but also to approach the ideal qualities of inescapablilty and non-immunogenicity. Our strategy involves bispecific targeting domains consisting of an extracellular region of human CD4, the primary HIV receptor, attached to a second motif that recognizes a distinct highly conserved region on the otherwise highly variable Env. The second motif is intended both to enhance CAR potency, and to prevent the CD4 from acting as an HIV entry receptor on CAR-expressing CD8+ T cells. We have focused on second motifs predicted to lack the immunogenicity associated with antibody-based CARs. A particularly favorable construct contains CD4 linked to the carbohydrate recognition domain of human mannose-binding lectin (MBL), which recognizes the highly conserved oligomannose patch on gp120 (Ghanem et al., Cytotherapy). In another variant, CD4 is linked to the N-terminal portion of the CCR5 co-receptor. We found that the bispecific CARs broadly suppressed infection by genetically diverse HIV-1 isolates, with significantly greater potency than a monospecifc CD4 CAR. Similar results were obtained using rhesus macaque PBMCs and a rhesus version of the CD4-MBL CAR. These results set the stage for testing the activities of the CD4-based bispecific CARs in relevant animal models including rhesus macaques and humanized mice.
Efficacy of immunotherapy approaches in metastatic and non-metastatic lung cancer models
Authors: Bommireddy Ramireddy , Menon Ashwathi P. , Munoz Luis E. , Huang Lei , Kim Janet , Kumar Jay , Lakshmanan Ramana , Ramalingam Suresh S. , Selvaraj Periasamy
Published: January 2023

Lung cancer is the leading cause of cancer related deaths accounting for approximately 30% of all cancer related mortalities affecting both men and women. Currently, there is no cure for the common non-small cell lung cancer (NSCLC) thus development of efficacious therapies is urgently needed. In this study, we investigate the efficacy of tumor membrane-based vaccine immunotherapy and immune checkpoint blockade antibodies in an aggressive, non-metastatic, immune checkpoint inhibitor resistant lung cancer model (LL/2) and a highly metastatic lung cancer (CMT-167) model. LL/2 and CMT-167 tumor tissues were harvested from C57BL/6 mice and processed to generate tumor membrane vesicles (TMVs). TMVs were then mixed with adjuvants and used for immunization in conjunction with immune checkpoint inhibitors. Tumor growth was monitored every 3 days. Survival was assessed using a Kaplan-Meier survival curve and significance determined using a Log-rank test for comparison analysis. Metastasis of CMT-167 was assessed by clonogenic assay. TMV alone, or in combination with adjuvants, anti-PD-1 or anti-CTLA-4 antibodies neither decreased the tumor growth nor improved the survival of mice challenged with LL/2 tumor cells. However, anti-PD-1 antibody delayed the tumor growth, improved the survival and reduced pulmonary metastasis in mice challenged with CMT-167 tumor cells. These models provide a valuable tool in analyzing the mechanism of interaction of immune cells with tumors that lead to either tumor regression or resistance to immunotherapies.
Potentiating effects of high-molecular weight fucoidan-agaricus mix (CUA) feeding on tumor vaccination
Authors: Miyazaki Yoshiyuki , Iwaihara Yuri , Nakamizo Masaji , Takeuchi Shugo , Takeuchi Hideaki , Tachikawa Daisuke
Published: January 2023

Fucoidan is a series of sulfated polysaccharides derived from brown algae that mainly consist of L-fucose. Various biological activities of fucoidan such as anti-cancer and immune modulatory effects were reported, and our investigation with animal experiments and human trials demonstrated fucoidan from Cladosiphon okamuranus and Undaria pinnatifida effectively augmented anti-tumor immunity in combination with Agaricus blazei mycelium extract. In this study, we evaluated dietary effects of the two types of high-molecular weight fucoidans and the agaricus extract mix (CUA) on achievement of effective tumor vaccination with a tumor antigen gp70 expressed on colon-26 tumor cell line. Balb/c mice were immunized with 0.05 mg gp70 peptide emulsified in complete freund’s adjuvant and intake 1% CUA containing AIN93G diet for 4 weeks. This procedure totally enhanced systemic immune function because splenocytes from the vaccinated mice extensively proliferate in response to concanavalin A-stimulation. The NK cell activity and gp70 peptide-stimulated IFN-gamma production in splenocytes from the vaccinated mice were tended to augment by the CUA feeding. On the other hands, the CUA feeding potentiated the killing activity to colon-26 carcinoma of draining lymph node (LN) cells from the vaccinated mice in association with increase of gp70-specific CD8-positive T cell population. Furthermore, the expressions of MHC class II molecule (I-A/I-E) on CD11c-positive and F4/80-positive populations of LN cells from the vaccinated mice were elevated by the intake of CUA. These results suggested that the CUA feeding potentially support effective induction of anti-tumor immune function by vaccination with tumor antigen peptides.
Depletion of islet resident macrophages protects mice from type 1 diabetes
Authors: Carrero Javier A. , McCarthy Derrick , Ferris Stephen , Wan Xiaoxiao , Hu Hao , Zinselmeyer Bernd , Vomund Anthony , Unanue Emil
Published: January 2023

Tissue resident macrophages are found at birth in the islets of Langerhans of all strains of mice examined. To examine the macrophage’s role in islet homeostasis and diabetogenesis, we depleted the cells using a monoclonal antibody against the CSF-1 receptor. Depletion of islet macrophages in the C57BL/6 mouse strain for 2–6 weeks did not affect multiple parameters of homeostasis including glucose tolerance, insulin content, or whole islet transcriptome. In contrast, depletion of the islet macrophages in 3 week old diabetes-prone NOD mice led to multiple changes in the progression of disease. First, there was a reduction in CD4 T cells and dendritic cells that infiltrate islets at 4–6 weeks of age. Second, there was a reduction in the presentation of insulin-derived peptides to T cells by dispersed islet cells. Third, the development of autoimmune diabetes was significantly reduced. The protection from diabetes induced by anti-CSF-1 receptor antibody was effective even when treatment was started at 10 weeks of age, a time where NOD mice have significant infiltration in their islets. Despite the protection from diabetes, mice treated with anti-CSF-1 receptor antibody at both 3 and 10 weeks of age harbored extensive leukocytic infiltration into islets when examined at 20–40 weeks of age. Treatment with an anti-PD-1 antibody led to immediate diabetes progression in all of the long-term protected NOD mice. These results show that depletion of islet-resident macrophages limited the islet leukocytic infiltration in the early phase of diabetogenesis and also permitted the establishment of a long-lived protective regulatory milieu late in the disease.
Transcriptomic meta-analysis reveals signatures of chronic inflammation in the classical monocyte population
Authors: Golden Jackelyn , Richardson Brian , Seth Divya , Cartwright Michael , Sekaly Rafick-Pierre , McCormick Thomas S. , Cooper Kevin D. , Cameron Cheryl M. , Cameron Mark J.
Published: January 2023

Individuals with chronic diseases are reported to have increased classical monocytes (CD14++CD16neg) which can be activated by an infectious agent and/or inflammatory milieu. Both psoriasis and HIV are considered chronic inflammatory diseases and affected individuals display alterations in monocyte phenotype and function. Moreover, individuals with either psoriasis or HIV demonstrate a significantly increased risk of developing cardiovascular disease (CVD). We sorted classical monocytes from psoriatic and healthy controls and compared them to classical monocytes from PLHIV individuals (elite controllers (EC) and non-controllers (NC, cART suppressed). Using RNA-Seq, we identified significant differentially expressed genes (DEG, p<0.05) in psoriasis monocytes (164 DEGs; compared to controls) and in HIV+ classical monocytes from ECs (540 DEGs; compared to NC). We then performed a meta-analysis of the psoriasis transcriptome to the EC-HIV+ transcriptome, revealing a common set of DEGs comprising a unique gene signature involving cellular stress, chemokines, adhesion, and the clotting cascade. We further analyzed the common DEGs via pathway analysis (p & false discovery rate<0.05) and STRING analysis to reveal a common dysregulated network of DEG between psoriasis and HIV. Importantly, we identified a focused network of DEG that may hallmark chronic inflammation in monocyte phenotypes. Therefore, our transcriptional meta-analysis identified candidate biomarkers that may underlie common pathologic mechanisms in psoriasis and HIV and serve as highly refined targets to treat not only primary disease, but also associated comorbidities (e.g. CVD) related to inflammation.
Intravenous anesthetic propofol and its analogue flopofol attenuated 5-lipoxygenase function and leukotriene B4 production
Authors: Yuki Koichi , Koutsogiannaki Sophia
Published: January 2023

Anesthesia is a routine clinical practice in surgical and interventional procedures, provided by using volatile and intravenous anesthetics. A growing literature suggests that anesthetics affect immune cellular function, and it is of great importance to understand the mechanism given an increased number of immunocompromized patients undergo anesthesia in current practice. Propofol is a very common intravenous anesthetic. We found that propofol attenuated the production of leukotriene B4 (LTB4), a major neutrophil chemoattractant, in addition to other leukotrienes in our experimental sepsis model. Because leukotrienes are produced via 5-lipoxgenase (5-LOX) activity, we hypothesized that propofol would attenuate 5-LOX activity. Our in vitro experiments using N-formyl-methionyl-leucyl-phenylalanine (fMLP)-stimulated neutrophils and 5-LOX transfected HEK cells showed that propofol attenuated 5-LOX activity. Using photoactivable propofol called azi-Pm, we showed that propofol directly bound to 5-LOX at functionally active sites. Flopofol is a propofol analogue with the substitution of 1-hydroxyl group to fluorine, and previously shown to lack anesthetic effect. In contrast, this analogue also attenuated 5-LOX activity and reduced LTB4 production, suggesting the possibility to use anesthetic as a prototype of immunomodulator. In addition, the impact of propofol in neutrophil function needs to be delineated in clinical scenario.
Influenza A virus infection causes chronic lung disease linked to IL-13 and mucin expression at sites of viral remnants
Authors: Keeler Shamus P. , Agapov Eugene , Hinojosa Michael E. , Letvin Adam N. , Wu Kangyun , Holtzman Michael J.
Published: January 2023

Clinical and experimental observations suggest that the development of chronic obstructive lung disease is linked to respiratory viral infection. However, there is no experimental model that establishes the long-term aspect of this relationship using a human pathogen with high level viral replication. Here we show that influenza A virus (IAV) infection achieves million-fold increases in viral load in the lung and dose-dependent severity of acute illness followed by long-term persistence of negative and positive strand viral remnants and similarly dose-dependent development of chronic lung disease. The disease persists for at least 6 months and is manifested by focal areas of bronchiolization and mucus production that contain increased levels of IAV remnants along with mucin Muc5ac and Il13 mRNA expression. These disease manifestations and airway hyper-reactivity are attenuated with loss of IL-13 production or signaling (using Il13 or Stat6 deficient mice). These deficiencies also cause reciprocal increases in l17a mRNA and neutrophils in the lung, however, post-viral mucus production and hyper-reactivity are unchanged with combined IL-13–IL-17a deficiency compared to IL-13 deficiency or with STAT6-IL-17a deficiency compared to STAT6 deficiency. The results establish the capacity of a potent human respiratory virus to produce chronic lung disease at sites of viral remnants, presumably reflecting locations of viral replication that eventually translate to IL-13-driven mucus production. Since highly infectious mouse parainfluenza virus causes similar disease, the findings also implicate high-level viral replication and severity of infection as key determinants of hyper-secretory lung diseases such as asthma and COPD.
MARCH1 plays an indispensable role in the development of airway allergic immunity and asthma
Authors: Castellanos Carlos A. , Ren Xin , Huang Xiaozhu , Shin Jeoung-Sook
Published: January 2023

Membrane associated RING-CH-1 (MARCH1) is an E3 ubiquitin ligase expressed mostly by antigen presenting cells (APCs) of the hematopoietic lineage. MARCH1 ubiquitinates surface MHCII and CD86 targeting these molecules for lysosomal degradation. MARCH1-mediated MHCII ubiquitination is crucial for thymic regulatory T cell generation. Whether MARCH1 plays a role in airway allergic inflammation remains unknown. We interrogated the function of MARCH1 in a well-established house dust mite (HDM) driven allergic inflammation mouse model of asthma. Despite displaying higher levels of MHCII and CD86 in APCs, MARCH1-deficient mice were incapable of developing allergic airway inflammation comprising of lung eosinophil accumulation, Th2 cell production of IL-4 and IL-13 cytokines, serum IgE responses, airway hyper-reactivity and mucus production. MARCH1-deficient mice failed to elicit IL-4 production from naïve CD4+ T cells in the lung draining lymph node during HDM sensitization. In addition, MARCH1-deficient mice did not effectively expand effector Th2 cells in the lungs after HDM challenge. Importantly, ablation of MARCH1 expression protected HDM sensitized mice from developing airway inflammation and hyper-production of IgE following re-exposure to HDM. These findings indicate that MARCH1 plays a crucial role in the initiation and progression of allergic airway inflammation, raising the possibility that modulating MARCH1 function might be therapeutically beneficial in asthma treatment.
A novel role of human lung endothelial cells in allergic airway disease by producing and responding to IL-33
Authors: Howard Chanie L. , Decker Donna C. , Swanson Ivy Aneas , Blaine Kelly M. , Nobrega Marcelo A. , Sperling Anne I.
Published: January 2023

Allergic asthma affects more than 300 million people worldwide and is characterized by airway hypersensitivity and eosinophilia. A prevalent feature of allergic asthma is increased serum IL-33 levels, and asthma GWAS have demonstrated that SNPs in the IL-33 locus are significantly associated with disease. While IL-33 and its downstream type 2 responses have been extensively studied in murine models of asthma, much less is known about the regulation of human IL33. Analysis of the mouse and human IL33 loci reveals little genomic conservation between the two species in non-coding regions. We generated a novel BAC transgenic mouse strain containing the human IL-33 locus with a fluorescent reporter to interrogate the regulation and expression of human IL-33. Surprisingly, the mice expressed human IL-33 primarily in endothelial cells, whereas murine IL-33 was expressed primarily in epithelial cells. These results mirror the expression profiles of IL-33 in primary human lung cells from the LungGENS database, thus demonstrating that our novel mouse model faithfully replicates human IL-33 expression. To understand how human IL-33 in the lung is regulated and expressed during inflammation, we examined BAC transgenic mice challenged with either house dust mite extract (HDM) or poly(I:C). In contrast to murine IL-33, expression of human IL-33 was reduced during allergic inflammation. We tested whether IL-33 is able to autoregulate itself via a negative feedback loop. Indeed, IL-33 administration downregulated the expression of human IL-33 in lung endothelial cells. Together, these data emphasize a distinct and novel role in humans for lung endothelial cells in allergic airway disease by producing and responding to IL-33.
Estrogen contributes to sex differences in M2-polarization during asthma
Authors: Keselman Aleksander M. , Fang Jessie , White Preston B. , Heller Nicola M.
Published: January 2023

Asthma is a chronic inflammation of the airways that exhibits sex differences, affecting mostly boys in childhood and women in adulthood. Alveolar macrophages have emerged as major mediators of allergic lung inflammation. We hypothesized that alveolar macrophages respond to estrogen by enhancing M2 responses and thereby contribute to sex differences in asthma. We found that macrophages from female mice exhibited increased expression of canonical M2 markers like Ym1 and Arg1 in response to IL-4. Signaling through ERα further enhanced IL-4-induced M2 gene expression in macrophages from female mice. The promoters of these genes contained increased abundance of transcriptionally active modifications like H3Ac and H3K4Me3 in macrophages from female mice. Using an OVA model of allergic lung inflammation, we found that female mice exhibited enhanced M2-polarization in vivo after allergen challenge. M2-polarization was impaired in ovariectomized (OVx) mice and but was restored with estrogen replacement. Further, we established a mixed bone marrow chimera model whereby irradiated mice were reconstituted with a 1:1 mix of bone marrow from CD45.1+ WT and CD45.2+ LysMCREERαflox/flox mice. In these mice, ERα-sufficient alveolar macrophages exhibited enhanced M2-polarization compared to ERα−/− alveolar macrophages. ERα−/−cells were largely retained in the bone marrow, suggesting that E2 regulates leukocyte trafficking during lung inflammation. Together these data suggest that sex and hormonal factors contribute to sex differences in macrophage responses during asthma. Understanding the role of estrogen signaling in M2-polarization is paramount for identifying novel therapeutic targets to better treat women with asthma.
The loss of DGK protects against allergic airway inflammation and airway hyperresponsiveness
Authors: Singh Brenal , Lu Wen , Schmidt-Paustian Amanda , Kambayashi Taku
Published: January 2023

Asthma is a chronic allergic inflammatory airway disease that is caused by aberrant immune responses to inhaled allergens, which leads to airflow obstruction driven in part by increased sensitivity of airway smooth muscle to contractile agonists, a process known as airway hyperresponsiveness (AHR). The inflammation of allergic asthma is driven by type 2 cytokines released by Th2 CD4+ T cells and group 2 innate lymphoid cells (ILC2) in the lung. Here, we report that targeting DAG kinase zeta (DGKζ), a negative regulator of DAG-mediated cell signaling, protects against allergic asthma. DGKζ knockout (KO) mice exhibited reduced type 2 airway inflammation and were completely resistant to AHR induction in a mouse model of allergic asthma. Surprisingly, the mechanism by which DGKζ protected against airway inflammation and AHR were separable. Targeted deletion of DGKζ in T cells led to decreased type 2 inflammation with no attenuation of AHR. In contrast, conditional deletion of DGKζ in airway smooth muscle cells led to decreased AHR despite no changes in airway inflammation. Importantly, the pharmacological inhibition of DGK provided protection against airway inflammation and AHR in mice, and also reduced bronchoconstriction of human airways. Together, our data suggest that DGKζ is potentially a novel therapeutic target for allergic asthma. Moreover, the targeted deletion of DGKζ reveals that the inflammatory and AHR components of allergic asthma are not as interdependent, as generally believed.
TRIM37 regulates allergic airway inflammation in a murine model of asthma
Authors: Chang Hung-Chih , Lu Jean , Chen Hui-Chen
Published: January 2023

Patients with asthma, an allergic airway inflammation disease caused by dysregulation of T helper cell type 2 (TH2), have dramatically increased in the past decades and cost of care is more than 11.3 billion per year. Evidence reveal that activation of pattern recognition receptor (PRR) signaling in both hematopoietic and non-hematopoietic compartment are critical for the development of allergic airway inflammation. TRIM37 is the only member of Tripartite motif (TRIM) family protein that contains TRAF domain, studies indicated that TRIM37 might involve in regulating PRR-, TNFR- and cytokine-mediated signaling pathways suggesting its important role in regulating immune responses. However, whether there is functional impact of TRIM37 on the development of asthma remains to be examined. Using immunohistochemistry staining of lung tissues from naïve and allergen-induced inflamed mice, positive staining of TRIM37 was observed in airway epithelial cells. Interestingly, the expression level was further augmented when lung was inflamed. Functional analysis of TRIM37 indicated that TRIM37 was involved in regulating TLR-mediated signaling in airway epithelial cells, as production of pro-inflammatory cytokines was reduced in response to TLR agonists when TRIM37 was knocked-down in the cells. Furthermore, when TRIM37 were knocked-down by intratracheally delivering lentivirus carrying TRIM37shRNA into the sensitized mice before challenge showed that knockdown of TRIM37 expression in the lung during challenge diminished cardinal features of asthma. These findings suggested the involvement of TRIM37 in allergic airway inflammation.
Soluble LOX-1: A Potential biomarker for SLE and Cardiovascular Comorbidity
Authors: Sagar Divya , Gaddipati Ranjitha , Ongstad Emily , Rahman Saifur , Belkhodja Mehdi , Bhagroo Nicholas , An Ling-Ling , Hasni Sarfaraz , Siegal Richard , Sanjuan Miguel A. , Kolbeck Roland , Karathanasis Sotirios , Sims Gary P. , Gupta Ruchi
Published: January 2023

Cardiovascular disease is the leading cause of mortality among systemic lupus erythematosus (SLE) patients. LOX-1 is a scavenger receptor strongly implicated in plaque formation, progression and destabilization. It gets upregulated during vascular inflammation on endothelial cells, monocytes, macrophages, smooth muscle cells and platelets. Soluble LOX-1 (sLOX-1) can serve as a potential biomarker for identifying SLE patients with increased cardiovascular risk. In a 120 SLE patient cohort, circulating sLOX-1 levels were 2-fold higher compared to healthy controls, particularly in those14–40 years of age. Patients with high sLOX-1 had elevated proinflammatory HDL, oxLDL, CRP and SAA levels; known markers for vascular injury. High sLOX-1 patient HDL had higher cholesterol content and lower efflux capacity compared to healthy and low sLOX-1 HDL. Both patient derived monocytes and low-density granulocytes (LDGs) express significant levels of membrane LOX-1 compared to controls. Monocytes when conditioned with oxLDL and stimulated with immune complexes, release significant levels of sLOX-1 and cytokines through the LOX-1 receptor. SLE HDL failed to nuclear colocalize the inflammation resolution factor ATF3, in monocyte derived macrophages. Additionally, patient LDGs underwent rapid NETosis in the presence of immune complexes when primed with oxLDL, through the LOX-1 receptor. Disease modified lipoproteins in SLE function as LOX-1 ligand contributing to inflammatory immune activation adding to cardiovascular risk. Hence, sLOX-1 can be predictive of increased cardiovascular risk and LOX-1 receptor blockade may be a promising target for ameliorating cardiovascular damage in SLE.
Lupus nephritis: the roles of C1q and C3 in preventing antibody mediated injury
Authors: Skopelja-Gardner Sladjana , Peng YuFeng , Colonna Lucrezia , Sun Xizhang , Tanaka Lena , Salant David , Elkon Keith B.
Published: January 2023

Lupus nephritis (LN) is a common and serious complication of systemic lupus erythematosus (SLE). While defective clearance of apoptotic cells (AC) and immune complexes (IC) is implicated in LN pathogenesis, the precise way in which different complement components impact kidney protection and injury is unknown. To address this, we created C57BL/6 mice with defective clearance of AC (Mfge8 deficiency) and anti-chromatin antibodies (sle1 interval) that were also deficient in either C1q [C1q Triple mutant (C1qTM)] or C3 (C3TM). Both C1q and C3 TM mice spontaneously developed anti-DNA antibodies and glomerulonephritis associated with glomerular IgG deposits after the age of 6 months. To dissociate the effects of complement on B cells versus effects on the kidney, we examined the effects of antibody mediated kidney injury (nephrotoxic nephritis, NTN) in mice deficient in the clearance of AC with complement deficiency [double knockout (DKO) (Mfge8 C1q or Mfge8 C3) mice]. In NTN, 50% of C1q DKO and 25% of C3 DKO, but no Mfge8 KO, died after NTN induction and the surviving C1q DKO mice developed severe proteinuria. Surprisingly, both spontaneous and NTN-induced nephritis in C1q TM or C1q DKO had strong glomerular C3 and C3d staining, while strong staining for the membrane attack complex (MAC) was observed in C3 TM or C3 DKO mice. These findings indicate that: 1) C1q and C3 independently protect against kidney injury separate from their effects on B cell function 2) In the presence of excess AC and low C1q, antibody mediated kidney injury is associated with C3 activation, presumably by the lectin or alternative pathway and 3) In the presence of excess AC and low C3, antibodies can cause MAC activation by pathways that remain to be determined.
Increased NKp44 (−) ILC3s in the peripheral blood of patients with Crohn’s disease
Authors: Li Jian , Drourr Joshua , Glover Sarah
Published: January 2023

Crohn’s disease (CD) is a complex chronic immune-mediated disorder of the human gastrointestinal tract. Emerging evidence indicates an essential role of the innate immunity in this process. Newly identified innate lymphoid cells (ILCs) are important components of the innate immunity. They act as the first line of defense to eliminate the invading pathogens. Previously, we have reported that IFN-γ-producing CD127 (+) ILC1 subset accumulated in the inflamed terminal ileum of CD patients at the expense of IL-22-producing NKp44 (+) ILC3s. This ILC phenotypic alteration was associated with the expansion of pathogenic Th17/Th1 subset among the diseased tissues. In order to further understand the contribution of ILCs to the pathogenesis of CD, we evaluated the aberrant ILCs’ responses in the peripheral blood of active CD patients in this study. We noticed that the frequency of IFN-γ-producing-CD127 (+) ILC1s was increased in the circulation of CD patients compared to donors. Furthermore, the frequency of NKp44 (−) CD117 (+) ILC3s was also significantly increased. Interestingly, they did not produce any of the common ILC cytokines: IL-17A, IL-22 and IFN-γ which is consistent with their ILC progenitor status in recent reports. In addition, almost half of the NKp44 (−) CD117 (+) ILC3s in the peripheral blood of CD patients were HLA-DR (MHC-II) positive which reveals their potential to interact and regulate systemic adaptive immunity, especially CD4 T cell responses. Different from ILC1s and ILC3s, the frequency of CRTH2 (+) ILC2s was decreased. More detailed analysis of the dysregulated innate and adaptive immune responses will help us to better understand the mechanism of CD pathogenesis and develop more novel therapeutic strategies.
A myeloid population potentially corresponding to myeloid monocytic suppressor cells (MoMDSCs) is increased in the blood of Rheumatoid Arthritis patients and associated with osteoclastogenesis
Authors: Macaubas Claudia , Gaertner Fabian , Yeh Chia-Lin , Rajasekaran Narendiran , Ilstad-Minnihan Alexandra , Nakamura Mary C. , Mellins Elizabeth D.
Published: January 2023

The development of erosive arthritis and joint damage are common and severe complications of rheumatoid arthritis (RA). Erosions are caused by excessive activity of osteoclasts (OCs), the only known bone resorbing cells. OCs are myeloid lineage cells and can be generated in vitro from CD14+ circulating myeloid cells, using RANKL and M-CSF. To characterize circulating myeloid lineage cells in RA, we performed mass cytometry (CyTOF) analysis of blood from 11 RA patients and 10 age and gender-matched healthy controls. Using CITRUS analysis, we found a myeloid cell cluster that is significantly elevated in RA patients compared to healthy subjects. The phenotype of this cluster is CD11b+, CD14+, CD16−, HLA-DR-/lo, CCR2hi, RANKhi, and phosphorylated (p)STAT3hi. The surface phenotype of this cluster is associated with monocytic myeloid derived suppressor cells (MoMDSCs). Analysis by conventional flow of a separate group of 14 RA patients showed that frequency of these myeloid cells correlates with number of blood osteoclasts precursors (OCP) based on the in vitro assay. Baseline level of pSrc and pJNK in these cells also correlated with OCP numbers. Both of these molecules are involved in downstream signaling cascades involved in osteoclastogenesis and osteoclast survival. We also found that intermediate (CD14++CD16+) monocytes are elevated in RA, as previously described, but these cells did not show a positive correlation with OCP numbers. These results indicate that a group of myeloid cells, potentially corresponding to MoMDSCs, are elevated in the blood of RA patients. Their phenotype (high RANK, phosphorylated Src, phosphorylated JNK) and frequency indicate that they may be involved in osteoclastogenesis, as described in mouse models.
Identification of perturbations in macrophage polarization in active systemic lupus erythematosus.
Authors: Labonte Adam C. , Bachali Prathyusha , Catalina Michelle , Robl Robert , Geraci Nicholas S. , Lipsky Peter , Grammer Amrie
Published: January 2023

SLE is characterized by abnormalities in B cell and T cell function, but the role of disturbances in the polarization of macrophages (Mφ) into functional subpopulations is unclear. Gene expression profiles were analyzed to identify differentially expressed (DE) genes between healthy controls and patients with either inactive or active SLE. While hundreds of DE genes were identified in B and T cells of active SLE patients, there were no DE genes found in B or T cells from patients with inactive SLE compared to healthy controls. In contrast, large numbers of DE genes were found in myeloid cells (MC) from both active and inactive SLE patients, and many of these were Mφ polarization genes (M1: STAT1, SOCS3; M2: STAT3, STAT6, CD163). Weighted gene correlation network analysis (WGCNA) was used to identify modules of genes associated with clinical activity in SLE patients; among these were disease activity-correlated modules containing polarization signatures of predominantly M1-associated genes. No disease activity-correlated modules were enriched in M2-associated genes. Pathway and upstream regulator analysis of DE genes from both active and inactive SLE MC were cross-referenced with high-scoring hits from the drug discovery Library of Integrated Network-based Cellular Signatures (LINCS) to identify new strategies to treat both stages of SLE. In summary, altered MC gene expression is characteristic of both active and inactive SLE, and disease activity is associated with an alteration in the MC signature towards the M1 phenotype. These data suggest that while B and T cell hyperactivity are associated with active SLE pathology, myeloid cells potentially direct flare-ups and remission by switching between M1- and M2-like polarization states.
Functions of GATA-3 Expression in Regulatory T Cells During Type 1 and Type 2 Inflammation
Authors: Gurram Rama Krishna , Zhu Jinfang (Jeff)
Published: January 2023

The transcription factor GATA-3 is not only responsible for effector CD4 T cell differentiation, but also essential for the homeostasis and stability of regulatory T cells (Tregs). Previous study from our group has shown that mice with both T-bet and GATA-3 deletion in Tregs developed severe autoimmune disease correlated with Tregs switching from Tregs to Th17-like cells. However, mice with only GATA3 deletion in Tregs were normal at steady state. To determine the role of GATA-3 in Tregs during inflammation, we used Th1 and Th2 mouse models. Our results suggest that GATA-3-deficient Tregs lose their Treg phenotype during a Th1 inflammation induced by scurfy fetal liver transplantation. However, in an asthma model GATA-3 (fl/fl) Foxp3 YFP Cre mice displayed reduced asthma symptoms, without affecting Th2 cell accumulation, or IL-4 and IL-13 cytokine production in the lung. Therefore, we propose that the expression of GATA-3 in Tregs may prevent Tregs from converting into other Th subsets during type 1 inflammation. On the other hand, GATA-3 expression in Tregs may contribute to Th2 responses by switching Tregs into Th2 cells under type 2 environment.
Early burst of glycolysis in microglia regulates mitochondrial dysfunction in oligodendrocytes under neuro-inflammation
Authors: Giri Shailendra , Suhail Hamid , Singh Jaspreet , Kumar Ashok , Rattan Ramandeep
Published: January 2023

Brain resident microglia plays a pathological role in the loss of oligodendrocyte’s function under neuroinflammation. The precise signaling mechanism by which microglial driven factor attenuates the myelin gene expression under such condition is poorly defined. Here we used bioenergetic approach to understand the microglia-oligodendrocyte crosstalk. We found that mixed glial cells under inflammatory stimulation using lipopolysaccharide plus interferon gamma (LI) had significantly enhanced glycolysis with drastic attenuation of mitochondrial respiration and myelin gene expression. Dissecting the mixed glial culture into pure microglia, astrocyte and oligodendrocytes (OLs) revealed that primary microglia was the main source of higher glycolysis, inflammatory cytokines, and nitric oxide (NO). An early burst of glycolysis in microglia in response to LI treatment caused the production of inflammatory cytokines and NO, which is mediated by PDK1-Akt signaling. Culturing of OLs with LI-conditioned microglial media (mCM) resulted in complete failure of mitochondrial respiration in OLsalong with decreased expression of myelin genes. We identified that nitric oxide produced by microglia is the key player in the loss of mitochondrial respiration in OLs and targeting either glycolysis, PDK1 or Akt in microglia resulted in significant recovery of mitochondrial function and myelin genes expression in OLs. In summary, we identified a potential therapeutic signaling cascade in microglia; an early burst of glycolysis mediated by PDK1 and Akt, responsible for hyper-production of NO, ensuing in the failure of mitochondrial respiration in oligodendrocytes, and leading in reduction of myelin genes expression.
Modulation of TLR2/1 by PAM3CSK4 to induce generation of immunosuppressive macrophages as a therapeutic approach for SLE
Authors: Horuluoglu Begum Han , Bayik Defne , Goguet Emilie , Tross Debra , Blanco Luz P. , Kaplan Mariana J. , Klinman Dennis M.
Published: January 2023

Systematic Lupus Erythematosus (SLE) is an autoimmune disease characterized by the overproduction of autoantibodies and the chronic inflammation of multiple organs. Early studies suggested that T and/or B cell abnormalities were critical to disease pathogenesis in lupus patients. More recent work shows that abnormalities in the innate immunity play a contributory if not central role. In that context, we find that SLE patients have significantly more CD14lowCD16+ monocytes compared to healthy controls.CD14lowCD16+ monocytes are a subset known as non-classical monocytes and display inflammatory characteristics upon activation. The frequency of inflammatory macrophages correlates with disease severity and is associated with reduced clearance of apoptotic cells and the overproduction of inflammatory cytokines.

We previously found that the TLR2/1 agonist PAM3CSK4 (PAM3) can inhibit the generation of M1- while promoting the generation of immunosuppressive M2-like macrophages. This work examines whether PAM3 can shift the M1:M2 macrophage ratio to restore balance in lupus patients. Even though phenotypically, PAM3 did not seem to induce as many as CD163+ (a functional marker of M2) macrophages like M-CSF did, there was no significant difference in the endocytosis ability of the CD163+ macrophages induced by PAM3 compared M-CSF induced ones. The in vivo activity of PAM3 was investigated in the NZB/W murine model of lupus. Preliminary results indicate that, PAM3 injection increases the survival and decreases proteinuria in mice sera. These findings suggest that PAM3 may promote the polarization of SLE monocytes into immunosuppressive macrophages and thus could represent a novel approach to the therapy of this autoimmune disease.
SLAMF9 promotes inflammation and resistance to Salmonella infection
Authors: Wilson Timothy James , Clare Simon , Lyons Paul , Dougan Gordon , Smith Ken G. C.
Published: January 2023

SLAMF9 is an uncharacterized member of the CD2/SLAM family of cell surface receptors. Unlike other members of the SLAM-family, it contains no phosphorylated tyrosines or recognizable signaling motifs, and neither homotypic nor heterotypic interactions between SLAMF9 and other members of the SLAM family have yet been observed. Through generation of Slamf9−/− mice and novel antibody reagents, we have determined that SLAMF9 is expressed on plasmacytoid dendritic cells, B1 cells, and select populations of tissue-resident mononuclear phagocytes. RNA interference experiments in human THP-1 monocytes indicate that SLAMF9 is important in the production of pro-inflammatory cytokines following stimulation with TLR agonists, and infection of Slamf9−/− mice demonstrate the importance of SLAMF9 in clearance of systemic Salmonella Typhimurium infection.
Cerebrospinal fluid neopterin and CXCL13 are suitable biomarkers for staging and detection of treatment failure in a non-human primate model of human African trypanosomiasis
Authors: Maranga Dawn , Ngotho Maina , Mwadime Victor , Olouch George , Adino Thomas , Kagira John , Waema Maxwel , Bieler Sylvain , Ndung’u Joseph
Published: January 2023

Background

Follow-up of treated human African trypanosomiasis (HAT) patients has been a routine practice for confirming cure and early detection of treatment failure.

Objective

The aim of this study was to verify suitability of neopterin and CXCL13 as disease stage biomarkers and monitors of treatment efficacy in a vervet monkey model of HAT.

Methods

Six monkeys were infected with Trypanosoma brucei. Late stage disease was induced by sub-curative treatment with diminazene aceturate (DA) from 28 days post-infection (dpi). Upon relapse, the animals were treated curatively with melarsoprol (Mel B) from 82 dpi. Blood and cerebrospinal fluid (CSF) samples were collected weekly and assessed for parasitosis, as well as tested for neopterin and CXCL13 by ELISA, for a period of 39 weeks.

Results

Neopterin concentrations in serum increased rapidly after infection in early disease (stage 1), peaking at 14 dpi. Levels then dropped rapidly to pre-infection levels by 35 dpi. In contrast, there was a marginal increase in CSF neopterin during early infection, with a minor peak at 21 dpi. Serum CXCL13 increased rapidly from 7 dpi, peaking at 28 dpi. Levels dropped upon sub-curative treatment with DA. CXCL13 concentrations in CSF also increased gradually in early stage disease and continued to rise after sub-curative treatment with DA. Curative treatment with Mel B resulted in its gradual decline, reaching pre-infection levels 105 days later.

Conclusions

CSF neopterin and CXCL13 peak levels coinciding with time of relapses demonstrates the potential of these biomarkers in staging and detecting treatment failure. Moreover, the rapid fall in CSF neopterin after curative treatment confirms its great potential for use in development of a test of cure.
Peptide-Specific Engagement of Cerebrovascular Endothelial Cells Promotes Dysfunctional Calcium Signaling During Experimental Cerebral Malaria
Authors: Manglani Monica , Silva Alexander , McGavern Dorian
Published: January 2023

Cerebral malaria is a potentially fatal complication associated with Plasmodium falciparum infection. Disease pathology includes blood brain barrier (BBB) breakdown, microvascular hemorrhage, edema and brainstem herniation. Studies in rodents (referred to as experimental cerebral malaria) have revealed that parasite-specific CD8+ T cells induce this pathology via engagement of cerebrovascular endothelial cells (ECs). In this study, we sought novel insights into the mechanisms underlying alterations in BBB structure and function during the development of ECM. We developed a novel transgenic reporter system that allowed us to simultaneously monitor calcium signaling in cerebrovascular ECs and parasite-specific CD8+ T cells. We then used intravital two-photon microscopy (TPM) to monitor BBB dynamics in real-time at the peak of disease. Within venous networks, we observed a significant increase in the magnitude and propagation of EC calcium signaling. Calcium wave propagation along lengthy stretches of cerebrovascular endothelium was associated with regions of parasite-specific CD8+ T cell engagement (both stable and dynamic). Recognition of cognate peptide-MHC on luminal and abluminal EC surfaces resulted in calcium fluxes within parasite-specific CD8+ T cells, which was followed temporally by calcium wave propagation in the associated vasculature. Importantly, this pathophysiological response was completely eliminated by depletion of T cells. We postulate that T cell-mediated induction of calcium waves along cerebral blood vessels promotes noncytopathic disruption of the BBB. Therapeutic manipulation of this activity might help restore BBB homeostasis and prevent neurological complications during ECM.