bluuebunny/crossref_metadata_2025_split huggingface dataset:
Discrete modeling of macrophage differentiation
Authors: Butchy Adam A , Miskov-Zivanov Natasa 
Published: January 2023

Abstract
The process through which macrophages differentiate into either an M1 or M2 state is thought to be well elucidated. Literature suggests that only a handful of receptors and their ligands play a role in deciding macrophage fate. These receptors share many signaling pathways and work together to create either an M1 or M2 phenotype. While these phenotype generalizations are broad, the M1 and M2 states have been used as placeholders to describe drastically different metabolic phenotypes.
To determine if this complex process could be controlled by a handful of ligands and receptors, we used discrete modeling—a modeling method that uses Boolean logic and a simple notation that qualitatively describe a system—to create a model of the differentiation pathway as it is currently understood. From there, we began testing the model, seeing if we could elicit either M1 or M2 states by altering the levels of cytokines and chemokines in the model to simulate a macrophage being exposed to different stimuli. According to literature, M1 activation is generally elicited by: LPS, RAGE, and Interferon gamma; and M2 activation by: CSF-1, IL-13, and IL-4. In order to determine if a phenotype was successfully observed, we defined M1 activation as high expression of TNF-alpha, IL-12 and CD-48; and M2 activation as high expression of IL-10, TGF-beta and CD-36.
With this model, we were able to simulate the process of M1 and M2 differentiation using changes in ligand concentrations to elicit the metabolic phenotypes of each activation state. This means that the current understanding of the differentiation pathway is sufficient to build functional differentiation models. This opens up opportunities to explore the broad metabolic variation within activation states.
MicroRNA-30 modulates metabolic inflammation by regulating Notch signaling in adipose tissue macrophages.
Authors: Miranda Kathryn , Nagarkatti Prakash , Nagarkatti Mitzi 
Published: January 2023

Abstract
Macrophages are innate immune cells that play integral roles in maintenance of adipose tissue homeostasis. Visceral obesity stimulates pro-inflammatory macrophages to infiltrate into adipose tissue causing chronic low-grade inflammation that can lead to insulin resistance and cardiometabolic disorders. Notch signaling is elevated in obesity and serves as a form of communication between macrophages and adipocytes. Blockade of this pathway promotes adipose tissue browning while reducing inflammation and therefore, has therapeutic potential. MicroRNAs (miRNA, miR) are non-coding RNAs that bind the 3′UTR of target mRNAs to repress their translation and miRNA based therapies are presently being developed for clinical purposes. In the current study, we identified differentially expressed miRNAs in adipose tissue macrophages (ATMs) between lean and obese mice by microarray. Array analysis and PCR validation revealed miRNAs -30a-5p, -30c-5p, and -30e-5p were downregulated in obese ATMs suggesting the miR-30 family plays an important role in macrophage phenotype. Pathway analysis demonstrated that miR-30 targeted Notch signaling genes including Delta-like ligand-4 (DLL4), a previously identified therapeutic target for cardiometabolic disorders. It was noted that DLL4 expression was increased on obese ATMs and in vitro miRNA transfection studies further demonstrated that miR-30 modulates Notch signaling-induced inflammation. These data demonstrate for the first time that the miR-30 family may play a critical role in the regulation of DLL4-mediated Notch signaling in ATMs, thereby modulating metabolic inflammation. (Supported in part by NIH grants P01AT003961, R01AT006888, R01ES019313, R01MH094755 and P20GM103641).
The HMGCR inhibitor Fluvastatin induces apoptosis and autophagy in primary and neoplastic mast cells
Authors: Barnstein Brian O , Paez Patrick A , Kolawole Elizabeth Motunrayo , Ryan John J 
Published: January 2023

Abstract
Mast cells are best known for their role in inflammatory and allergic disease. Mast cell leukemia is a rare subtype of acute myelogenous leukemia, typically expressing the c-Kit mutation, D816V, that yields factor-independent growth. Understanding and treating mast cell hyperplasia and leukemia could be beneficial to both inflammatory and malignant diseases. Recently the family of statin drugs, widely employed as HMG CoA Reductase (HMGCR) inhibitors to lower serum cholesterol, have also been found to be immunosuppressive. However, statin effects on mast cell survival are not known. We report that Fluvastatin induces apoptosis in mouse bone marrow-derived mast cells (BMMC) and P815 mastocytoma cells in a dose- and time-dependent manner. Apoptosis was evidenced by DNA fragmentation and caspase activation. Importantly, the mastocytoma line P815, which expresses mutant c-Kit, was readily killed by Fluvastatin, with an IC50 of 5–10μM. P815 cell death was marked by c-Kit upregulation preceding mitochondrial membrane instability and Caspase-9 activation. Chemical blockade of autophagy reduced Fluvastatin-mediated apoptosis in primary mast cells, but augmented apoptosis in transformed mast cells. These data suggest that the HMGCR pathway may be targeted to suppress mast cell survival, and that its combination with autophagy inhibitors may be useful for treating mast cell neoplasias.
A20 regulates NF-κB activation through K48 linked polyubiquitination of NEMO
Authors: Lee Younglang , Lee Eries , Zilberman-Rudenko Jevgenia , Chen Yong , Gucek Marjan , Siegel Richard M. , Hanson Eric Paul 
Published: January 2023

Abstract
Activation of the NF-κB family of transcription factors is related to diverse biological processes, such as development, immune responses, and inflammation. As a result, mutations affecting the NF-κB signaling pathway cause syndromes characterized by immunodeficiency and autoinflammaotory disease. Recently, we identified the C-terminus of NEMO to be a regulator of NF-κB signaling, as a class of NEMO mutations lacking the C-terminus (ΔCT-NEMO) confer gain-of-function properties to the IKK complex due to impaired recruitment of A20 to the TNFR. Here, we investigated details by which A20 recruitment to the TNFR suppresses NF-κB signaling. The C-terminus of NEMO has been previously identified to preferentially recognize K63-linked polyubiquitin that in its unanchored form can mediate NEMO/A20 interaction. We found that the IKK complex containing ΔCT-NEMO forms but not other mutant forms of NEMO exhibit impaired K63 polyubiquitin binding ability. In addition to unanchored K63-lilnked polyubiquitin, we hypothesized that direct polyubiquitination of A20 could enable a functional interaction between A20 and NEMO. Using mass spectroscopy, we identified 4 lysine residues on A20 to be ubiquitination sites. Mutation of these lysines to arginine impaired A20’s ability to suppress TNF induced NF-κB activation. We noted stabilization of NEMO in A20 deficient cells following TNF stimulation. Biochemical studies in Jurkat and 293 cells revealed A20 dependent K48 linked polyubiquitination of NEMO at the TNFR following receptor stimulation. Our data suggest a mechanism of NF-κB regulation in which A20 is directly K63 polyubiquitinated to functionally interacts with NEMO which it targets by K48 linkages proteasomal degradation.
Identification of Nuclear Matrix Protein 4 (Nmp4) as a novel regulator of innate immunity
Authors: Bidwell Joseph , Morris David L , Adaway Michele , Huang Shengping , Shao Yu , Wek Ronald C , Sun Jie 
Published: January 2023

Abstract
Mesenchymal stem/progenitor cells (MSPCs) have immunomodulatory activity. We previously observed that osteoprogenitors and CD8+T cells are increased in the bone marrow (BM) of mice lacking the transcription factor Nmp4. Transcriptome analysis of MSPCs from Nmp4−/− and wild type (WT) mice revealed significant changes in genes related to innate immune responses, including reduced expression of Tlr3 and Mda5 in the null MSPCs. Loss of Nmp4 increased mRNA translation, ribosomal biogenesis, and Gadd34 expression in MSPCs; collectively, these processes likely promote the unfolded protein response (UPR), expanding the processing capacity of the endoplasmic reticulum. Therefore, we hypothesized that disabling Nmp4 would improve innate immune cell function. To test this, Nmp4−/− mice (C57BL6) were infected intranasally with influenza A/PR8 (H1N1; 150pfu). Despite having similar virus load in the lung, H1N1-infected Nmp4−/− mice lost less body weight than WT mice post infection and had improved survival. Infected Nmp4−/− mice had fewer infiltrating neutrophils (PMNs; CD11b+/Ly6G+) and monocytes (CD11b+/SiglecFNeg/Ly6GNeg/Ly6C+) in bronchoalveolar lavage (BAL) but more BAL macrophages (CD11c+/SiglecF+); CD4+ and CD8+ T cells were not different between genotypes. Ex vivo, Nmp4-deficiency did not alter the differentiation of BM marrow-derived macrophages; however, Chil3/Ym1 and Alox15 mRNA expression was reduced in Nmp4−/− macrophages polarized with IL-4/IL-13, suggesting cell-autonomous regulation of M2 macrophage activation by Nmp4. Our data point to Nmp4 as a novel regulator of innate immunity and influenza response in mice.
Discovery of two meningeal macrophage populations with differential roles during homeostasis and inflammation
Authors: Rua Rejane , Johnson Kory , McGavern Dorian B 
Published: January 2023

Abstract
The meningeal and perivascular spaces of the central nervous system (CNS) are inhabited by specialized macrophages. Under steady state conditions, we discovered two populations: immature macrophages (MHC-II−) and mature macrophages (MHC-II+). Microarray analyses revealed that IL-10 and TGFb were upstream regulators of the immature macrophage transcriptome, which included stem cell-specific genes. These data suggest that immature macrophages represent local progenitors maintained by anti-inflammatory cytokines within the meninges. Interestingly, in naïve mice MHC-II+ macrophages were enriched upon aging and upregulated inflammatory genes, suggesting age-based maturation. To better understand the dynamics of these macrophages, we triggered CNS inflammation by inoculating mice with lymphocytic choriomeningitis virus (LCMV). Both myeloid populations were infected by the virus, and intravital imaging studies revealed that they were targeted by infiltrating virus-specific CD8+ T cells, which promoted their depletion. Following viral clearance, myeloid repopulation of the meninges was derived largely from infiltrating monocytes that engrafted this CNS niche and adopted a transcriptomic signature of mature resident meningeal macrophages. In stark contrast, sterile depletion of meningeal macrophages without infection induced massive local proliferation of immature macrophages that transformed into mature macrophages and repopulated the meninges. This occurred in the absence of peripheral monocyte engraftment. Collectively, these data indicate that the CNS meninges are inhabited by two macrophage populations with a differential ability to repopulate the niche based on the inflammatory milieu.
A viral immunoevasin controls innate immunity by targeting the prototypical natural killer cell receptor family (NK1.1/NKR-P1)
Authors: Carlyle James R , Berry Richard , Rahim Mir Munir A , Reichel Johanna J , Popovic Branka , Tanaka Miho , Fu Zhihui , Balaji Gautham R , Lau Timothy NH , Tu Megan M , Kirkham Christina L. , Mahmoud Ahmad B , Mesci Aruz , Krmpotic Astrid , Allan David S.J. , Makrigiannis Andrew P , Jonjic Stipan , Rossjohn Jamie , Aguilar Oscar A 
Published: January 2023

Abstract
Natural killer (NK) cells play a key role in innate immunity by detecting alterations in self and non-self ligands via paired NK cell receptors (NKR). Despite identification of numerous NKR-ligand interactions, physiological ligands for the prototypical NK1.1 orphan receptor remain elusive. Here, we identify a viral ligand for the inhibitory and activating NKR-P1 (NK1.1) receptors. This murine cytomegalovirus (MCMV)-encoded protein, m12, restrains NK effector function by directly engaging the inhibitory NKR-P1B receptor. However, m12 also interacts with the activating NKR-P1A/C receptors to counterbalance m12 decoy function. Structural analyses reveal that m12 sequesters a large NKR-P1 surface area via a “polar claw” mechanism. Polymorphisms in, and ablation of, the viral m12 protein and host NKR-P1B/C alleles impact NK cell responses in vivo. Thus, we identify the long-sought foreign ligand for this key immunoregulatory NK cell receptor family and reveal how it controls the evolutionary balance of immune recognition during host-pathogen interplay.
Regulation of immune gene expression by the zinc finger DNA binding protein, CNBP/ZNF9
Authors: Chen Yongzhi , Jiang Zhaozhao , Caffrey Daniel R. , Shruti Sharma , Fitzgerald Katherine A. 
Published: January 2023

Abstract
An inducible program of inflammatory gene expression is a hallmark of antimicrobial defenses. This response is controlled by a collaboration involving signal-dependent activation of transcription factors, transcriptional co-regulators, and chromatin-modifying factors. Here we have identified a highly conserved Zinc finger DNA binding protein CNBP (also called ZNF9) upregulated in myeloid cells exposed to lipopolysaccharide. CNBP resides primarily in the cytosol and upon TLR4 engagement, CNBP translocate to the nucleus. To investigate the functional consequences of these events, we generated mice lacking CNBP and characterized the role of CNBP in controlling the inducible transcriptional program using a combination of RNA-sequencing and multiplex gene expression analysis (Nanostring). In response to an array of signals such as LPS, CNBP-deficient macrophages were impaired in their ability to induce important immune genes including IL12p40 and IL6 amongst others. CNBP-deficient cells showed normal activation of TBK1 and p65. However, the nuclear translocation of c-Rel that is critical for the inducible expression of IL-12p40 was significantly compromised in cells lacking CNBP. Furthermore, Chromatin Immunoprecipitation experiments revealed that CNBP itself bound the IL12 promoter and c-Rel binding to this promoter was compromised in CNBP-deficient macrophages. Lastly, ectopic expression of wild type CNBP but not a mutant form that could not be phosphorylated together with c-Rel led to synergistic activation of an IL12 promoter driven reporter gene. Collectively, these findings identify CNBP as a novel DNA binding protein that acts as a transcriptional regulator controlling the inflammatory program in macrophages.
Sterile type 2 inflammation triggered by microparticles is abrogated by inhibition of IL33 through blockade of Bruton’s tyrosine kinase
Authors: Mishra Pankaj Kumar , Palma Mark J , Millman Ariel , Hallab Nadim J , Beebe Kathleen , Benevenia Joseph , Gause William C 
Published: January 2023

Abstract
Wear debris microparticle release associated with total joint arthroplasty may contribute to harmful inflammation, osteolysis and implant failure. We examined the role of different molecular signaling pathways in microparticle-mediated inflammation. Intraperitoneal inoculation of cobalt chrome (CoCr) in mice induced a robust innate type 2 response with increases in IL4, IL5, IL13, IL33, M2 macrophages, eosinophils and neutrophils compared to controls given vehicle only. Blockade of the SYK signaling pathway with the inhibitor BAY 61-3606 or the down stream BTK signaling pathway with Ibrutinib abrogated the CoCr-mediated innate type 2 inflammation. Further studies in CBA/Nxid mice, a genetic BTK loss of function mutant also showed decreases in type 2 inflammation, confirming a critical role for BTK in CoCr mediated sterile type 2 inflammation. Inoculation of JH−/−, B cell deficient mice, or depletion of T cells by anti-CD4 (GK1.5) antibody, had no impact on CoCr mediated type 2 inflammation. We further demonstrated that the major source of IL33 was macrophages and intra-peritoneal administration of recombinant IL33 rescued the CoCr mediated type 2 response after BTK blockade. These studies indicate that the type 2 immune response caused by solid particles is abrogated by blockade of BTK signaling pathways and that exogenous IL33 is sufficient to rescue this response. These studies elucidate the nature of wear debris-mediated type 2 sterile inflammation and provide potential targets for future therapy.
Downregulation of the O-GlcNAc signaling promotes activation of the innate immune response in microbial sepsis
Authors: Li Xinghui , Gong Wei , Li Lupeng , Wen Haitao 
Published: January 2023

Abstract
Sepsis is the most common cause of death in intensive care units. Acute systemic inflammatory responses mediate morbidity in microbial sepsis. Studies of immune system metabolism have recently identified the tight link between metabolic reprogramming and immune system activation. A robust increase in glucose metabolism is observed in the initial phase of sepsis, indicating the involvement of glucose metabolism in the pathogenesis of sepsis. However, the role of individual glucose metabolism pathways in the regulation of septic inflammation is poorly understood. The hexosamine synthesis pathway (HBP) is a unique glucose metabolism pathway leading to protein modification by the O-linked b-N-acetylglucosamine (O-GlcNAc), a process known as O-GlcNAcylation. Here we report a critical role of HBP-associated O-GlcNAc signaling in modulating septic inflammation. Activation of TLR4 signaling results in decreased HBP flux activity and protein O-GlcNAcylation in macrophages. Decreased O-GlcNAc signaling is also observed in peritoneal macrophages following cecal ligation and puncture (CLP)-induced experimental sepsis, indicating that decreased O-GlcNAc signaling may exacerbate septic inflammation. Myeloid-specific deletion (OgtΔmye) of O-GlcNAc transferase (OGT), the key enzyme mediating protein O-GlcNAcylation, leads to increased activation of the NF-κB signaling and cytokine production upon the stimulation with various TLR agonists. Furthermore, OgtΔmye mice show increased susceptibility to LPS-induced endotoxin shock and CLP-induced sepsis. These findings identify a novel link between glucose metabolism and innate immune activation and suggest OGT as a therapeutic target for treating microbial sepsis.
Tissue resident memory B cells established in lungs in allergic asthma
Authors: Turner Damian L , Verter Jacob , Turner Ronak , Cao Minwei 
Published: January 2023

Abstract
Allergic asthma is a disease driven by the adaptive immune system in the lungs. B cells play a critical role during the allergen sensitization process by producing IgE and possibly through antigen presentation. While most current research has focused on the role of CD4+ tissue resident memory (TRM) cells, it is not known whether TRM B cells exist within the lungs and what role they might play. To investigate this question, we sensitized mice to house dust mite (HDM) intranasally for three weeks to induce asthmatic symptoms. We in vivo labeled mice with circulating fluorescent anti CD45 mAb and assessed the lung B cells’ susceptibility to labeling. We found that B cells, that are resistant to in vivo labeling, are a major population of lung lymphocytes following HDM sensitization. We were further able to detect these non-circulating B cells in the lungs as late as 10 weeks after treatment in sensitized mice. Furthermore, these cells were found in the same anatomic locations as CD4+ TRM cells and underwent expansion during HDM challenge. These data suggest that B cells do form a tissue resident population in the lungs and that these cells are involved in allergic asthmatic responses. This resident memory population of B cells are also locally activated upon allergen rechallenge. Whether these B cells are reactivated by HDM-derived antigens or by T cell-derived cytokines is yet to be determined.
Induction of CXCR5+ follicular CD8 T cells by CD40L adjuvanted DNA/MVA vaccination is associated with enhanced control of pathogenic SHIV infection.
Authors: Velu Vijayakumar , Styles Tiffany , Reddy Pradeep Babu Jagadeesh , Hicks Sakeenah , Gangadhara Sailaja , Amara Rama Rao 
Published: January 2023

Abstract
A significant challenge to HIV eradication is the elimination of viral reservoirs in germinal center (GC) resident T follicular helper cells. Therapeutic interventions that target viral reservoirs should induce anti-viral CD8 T cells that home to GC. We and others recently described a unique population of CD8 T cells that express the chemokine receptor CXCR5 (required for homing to GC) during chronic HIV/SIV infections. A critical question that remains to be addressed is if follicular homing CD8 T cells can be induced by vaccination and if so how will they impact control of HIV. Here we show that vaccination of rhesus macaques (RM) with DNA/MVA (modified vaccinia Ankara) SHIV vaccine induces low levels of vaccine-specific CXCR5+ CD8 T cells in blood. However, adjuvanting the DNA vaccine with CD40L dramatically enhanced the induction of CXCR5+ SHIV-specific CD8 T cells up to 10-fold with a frequency of about 5% of total CD8. These CXCR5+ CD8 T cells expressed higher levels of cytolytic molecules perforin and granzyme-B with higher proliferative capacity (ki-67) compared to CXCR5-subset. Following a pathogenic SHIV infection, the CXCR5+ CD8 subset expanded rapidly. Importantly, the frequency of SHIV-specific CXCR5+ CD8 T cells post vaccination and post infection showed a significant inverse correlation with viral load (p=0.003, R=−0.66), these associations were not observed with CXCR5-CD8 T cells. These data demonstrate that CXCR5+ CD8 T cells can be induced by vaccination, their induction can be augmented by CD40L, and the vaccine-induced CXCR5+ CD8 T cells contribute to control of pathogenic immunodeficiency virus infection. These findings have important implications for developing HIV cure strategies.
CAR NK and CAR T cells versus CTL: factors limiting the sensitivity and efficiency of engineered cytolytic effectors
Authors: Mazzanti Nicholas William , Anikeeva Nadia , Sato Takami , Sykulev Yuri 
Published: January 2023

Abstract
Factors limiting highly sensitive and efficient responses of virus- and cancer-specific CTL have been extensively studied. However, the sensitivity and effectiveness of T cells transduced with CAR specific for target molecules on cancer cells have been less characterized. We endowed NK92 cells and T cells purified from human cord blood with second generation CAR specific for High Molecular Weight Melanoma Associated Antigen (HMW-MMA) and compared the ability of CAR-NK92 and CAR-T cells to kill skin melanoma and uveal melanoma cell lines in in vitro cytolytic assay. We also evaluated the ability of established human virus-specific CTL clones to kill the same target cells presenting cognate pMHC ligands. Skin melanoma cells were more sensitive to specific lysis than uveal melanoma cells. The extent of the specific lysis strongly depended on the level of HMW-MMA expression. CTL clones appeared to be more superior effectors than CAR-NK92 or CAR-T cells as they lysed the melanoma cells that displayed much lower levels of cognate pMHC ligands. CAR-NK92 cells proliferated more vigorously than NK92 cells and maintained the level of CAR expression in culture. In contrast, CAR-CD8+T cells exhibited lower proliferative capacity than the control T cells and were losing cell surface CAR expression. We are currently evaluating the extent of TCR vs CAR clustering and co-clustering of CAR with TCR at the cell surface. We are also comparing Ca2+ flux and granule release by CAR-NK92, CAR-CD8 T cells and CTL in response to CAR or TCR stimulation. Disparities in these parameters could account for differential sensitivity and efficiency of engineered and natural cytolytic effectors.