bluuebunny/crossref_metadata_2025_split huggingface dataset:
Dynamic modulation of cortical actin at the immunological synapse controls lytic granule secretion in cytotoxic T lymphocytes
Authors: Kapnick Senta , Ritter Alex , Griffiths Gillian , Lippincott-Schwartz Jennifer , Schwartzberg Pamela 
Published: January 2023

Abstract
CD8+ cytotoxic T lymphocytes (CTLs) are critical for eliminating virally infected cells. Upon target recognition, CTLs reorganize their actin cytoskeleton to facilitate the polarized secretion of specialized granules containing lytic proteins that kill cells. Because a single CTL can kill multiple targets, granule secretion must be tightly regulated, ensuring only appropriate cells are killed, and preserving granules for serial killing. However, how CTLs regulate secretion remains unclear. Here, we used live confocal and TIRF microscopy to evaluate how cortical actin affects granule secretion at the membrane. While previous work has shown that clearance of actin at the synapse precedes granule secretion, we find that after granule fusion, cortical actin recovers and no further secretion is observed. Depolymerization of recovered actin led to resumed granule fusion, suggesting that recovered actin acts as a barrier to prevent further lytic granule secretion. We further show that CTLs from mice unable to secrete granules due to a Rab27a mutation that prevents granule fusion, failed to recover cortical actin at the synapse. This suggests that granule fusion itself triggers actin recovery in CTLs. Thus, cortical actin both regulates, and is regulated by, secretion. Finally, using fluorescent reporters, we correlated the clearance and recovery of actin with PIP2. PIP2 binds numerous actin regulatory proteins, suggesting phosphatidylinositol distribution in the membrane as a potential mechanism through which CTLs regulate the density of cortical actin during killing. Our work provides insight into actin-related mechanisms regulating CTL secretion that may serial killing capacity during immune responses.
The Limit of MHC Class II-driven Selection in Germinal Centers
Authors: Yeh Chen-Hao , Nojima Takuya , Kuraoka Masayuki , Kelsoe Garnett H. 
Published: January 2023

Abstract
B cells undergo iterative rounds of proliferation, hypermutation and selection in the germinal centers (GCs) to generate high affinity, protective antibody. It is now thought that GC B cells with higher affinity BCRs acquire more antigen for processing and presenting in association with MHC class II (MHCII) to T follicular helper (TFH) cells than lower affinity competitors. The resulting increase in T-cell help is hypothesized to be the major factor that drives affinity maturation. In this model for GC B cell selection, the density of peptide/MHCII (pMHCII) complexes available on the B-cell membrane is the major determinant of affinity-dependent selection. To quantify the limits of this model for GC selection, we generated short- and long-term chimeric mice with equal numbers of isogenic, mature B cells that differed by their capacity to express MHCII (MHCII+/+ and MHCII+/−). Separately, both B cell types generated virtually identical humoral responses, including affinity maturation, in response to NP-OVA. In competition, MHCII+/+ B cells were preferentially recruited to early GCs consistent with a critical role for pMHCII density in T:B collaboration. However, once established in GCs, no further competitive advantage was observed during the full course (24 days) of the GC response. During this period, both MHCII+/+ and MHCII+/− GC B cells comparably accumulated VH gene segment mutations and exhibited comparable rates of affinity maturation to the NP hapten. These data indicate that B-cell selection determined by MHCII density is more stringent prior to the establishment of GCs than in the GC itself. This observation may be relevant to our recent observation of the persistence of low affinity GC B cells.
IL-10 downregulates IL-4-induced IgE production through indirect effects on B cells
Authors: Lin Adora A , Nutman Thomas B 
Published: January 2023

Abstract
Control of allergic disease (and of helminth-associated pathology) is associated with decreased antigen-specific IgE and increased antigen-specific IgG4. Although IL-10 appears to contribute to this process, underlying mechanisms are largely unknown. This study explored how IL-10 differentially regulates human IgE and IgG4 production. PBMCs and highly purified total, naïve (CD27-), and memory (CD27+) B cells were cultured with combinations of IL-4, IL-10, and anti-CD40. Quantitative PCR was done for Cε and Cγ4 germline transcripts (GLTs). IgG and IgE isotypes were quantified from culture supernatants using multiplexed immunoassays. IL-4 induced the production of IgE and IgG4 by PBMCs (IgE: geo mean 16.2-fold above baseline; IgG4: geo mean 4.7-fold). IL-4 also increased expression of Cε and Cγ4 GLTs to 54.1-fold and 5.3-fold above baseline, respectively. Addition of IL-10 significantly (p&lt;0.01) reduced IL-4-induced IgE production to 4.6-fold and expression of Cε GLTs to 24.7-fold above baseline, without affecting IgG4 production or expression of Cγ4 GLTs. In cultures of isolated total B cells and B cell subsets, IL-10 had no significant effect on IL-4-induced IgE production; however, IL-10 increased IL-4-induced IgG4 production over 20-fold. IL-4-induced transcription of Cε and Cγ4 GLTs in isolated B cells was not affected by IL-10. Examination of IL-10 receptor (IL-10R) expression on B cell subsets showed increased frequencies of IL-10R+ memory B cells compared to naïve B cells. These results suggest that IL-10 acts indirectly through accessory cells to downregulate IL-4-induced production of IgE. IL-10 can act directly on B cells to upregulate IL-4-induced production of IgG4, likely downstream of germline transcription.
The LFA1 ligand, CD54, is not required for the differentiation of TFH cells or germinal center formation
Authors: Bowlin Marvin Q , Bradley John E , Zajac Allan J , Bullard Daniel C , Randall Troy D 
Published: January 2023

Abstract
In germinal centers (GC), B cells undergo cellular proliferation and affinity maturation, and ultimately differentiate into either high-affinity antibody-secreting cells (ASCs) or memory B cells. GC B cells present antigen to T follicular helper (TFH) cells in order to elicit survival and proliferation signals. The integrin, Lymphocyte Function-Associated Antigen 1 (LFA1), is required for this process, as mice lacking LFA1 or treated with LFA1 blocking antibodies fail to form GCs or TFH cells. We hypothesized that the counter-receptor for LFA1, ICAM-1 or CD54, would be similarly required for GC formation and TFH differentiation. To test this hypothesis, we infected CD54-deficient (KO) and wild-type (WT) mice with influenza virus (PR8) and enumerated ASCs and GC B cells on days 7, 9, and 14. We found that KO mice had more GC B cells than WT mice on days 9 and 14, but observed no difference in the number of ASCs or TFH cells. To determine if this result was intrinsic to the B cell lineage, we generated 50:50 (KO:WT) bone marrow chimeras. Following reconstitution, we infected the mice with PR8 and assayed donor-specific GC B cells and ASCs on days 9 and 14. On day 9, we found that GC B cells were highly enriched for KO cells, whereas the ASCs were highly enriched for WT cells. In contrast, TFH cells were represented equally in WT and KO populations. On day 14, we found that GC B cells were still highly enriched for KO cells, whereas there was no significant difference between WT and KO ASCs at this time. These results lead us to the surprising conclusion that, although the loss of LFA1 leads to the failure of TFH and GC responses, the loss of CD54 does not affect the TFH compartment and actually improves GC B cell responses.
Regulatory T cells facilitate Th17 responses to Bordetella pertussis and aid in resolution of whooping cough in mice.
Authors: Gates Hillary A , Smith Andrew A , Jakoush Tarek A , Silva Elvia , You Hana , Wu Victoria , Westfall Jessica , Segal-Abramson Tzvia 
Published: January 2023

Abstract
Whooping cough is a highly contagious respiratory disease caused by Bordetella pertussis (Bp). The prolonged nature of whooping cough has been associated with delayed Th17 responses, which are essential to resolve the infection. Regulatory T cells (Treg) are historically immunosuppressive, but have recently been demonstrated to facilitate proper immune responses during certain infections and to occasionally adapt proinflammatory phenotypes. Because Treg and Th17 cells are considered “brothers in arms,” we evaluated the role of Tregs during immune response to B. pertussis. Here, we test the hypothesis that Tregs are crucial for generating a proinflammatory response that resolves whooping cough. We show that homeostatic lung Tregs acquire a highly proliferative Th17 phenotype during Bp infection in mice and that the frequency of this Th17-like Tresg (T17reg) population raises in parallel with Th17 influx and expansion at late phases of infection. Our data suggests that the frequency of lung T17reg may be modulated by levels of IFNa and IL-2 in the lung. Depleting Foxp3+ cells throughout Bp infection resulted in decreased MHC class II expression on dendritic and endothelial cells, suggesting reduced APC maturation. Additionally, Treg knockout lead to increased lung IL-2 levels (which may diminish Th17 differentiation/accumulation) and compromised IL-17 secretion. Moreover, infected mice lacking Treg had an increased Bp load in the lungs. Our data support Tregs facilitating and assisting Th17 immune responses to Bp to help resolve the infection.
STAT2-mediated regulation of Macrophage Phenotype during Influenza and Bacterial Super-infection
Authors: Gopal Radha , Lee Benjamin , McHugh Kevin J. , Rich Helen E , Ramanan Krishnaveni , Mandalapu Sivanarayana , Clay Michelle E , Manni Michelle L , Robinson Keven M , Rangel-Moreno Javier , Alcorn John F 
Published: January 2023

Abstract
Influenza is a common contagious respiratory illness that affects people worldwide. Secondary bacterial infection is a major complication during influenza infection that causes significant morbidity and mortality. We have previously shown that influenza induces type I interferon (IFN)-mediated inhibition of Type17 immune responses, resulting in exacerbation of bacterial burden during influenza and Staphylococcus aureus super-infection in mice. Here, we investigated the role of the signaling mediator, STAT2, downstream of type I IFN signaling in both influenza infection and bacterial super-infection. Influenza-infected STAT2−/− mice had increased weight loss, increased viral burden, and increased inflammatory responses when compared to influenza-infected WT mice. Despite an exaggerated inflammatory response to influenza infection alone, influenza and methicillin resistant S. aureus (MRSA) super-infected STAT2−/− mice had lower bacterial burden, morbidity, and mortality than WT mice. STAT2−/− mice had an increased expression of markers associated with M1 (classical) and M2 (alternative) macrophages in lung during influenza or influenza-bacterial super infection compared to WT mice. Next, we found neutralization of IFNγ or Arginase1 decreased bacterial clearance in STAT2−/− mice in influenza-bacterial super-infection, suggesting that both M1 and M2 phenotype mediate host defense during influenza-bacterial super-infection. These data demonstrate a clear role for type I IFN induced STAT2 signaling in suppression of macrophage function during influenza, MRSA super-infection. Further, these studies reveal novel mechanistic insight into the roles of macrophage subpopulations in pulmonary host defense.
Blockade of type I interferon signaling reverses HIV-1 induced immune dysfunction and reduces HIV-1 reservoirs
Authors: Cheng Liang , Ma Jianping , Li Jingyun , Li Dan , Li Guangming , Yu Haisheng , Zhang Liguo , Su Lishan 
Published: January 2023

Abstract
Type I interferons (IFN-I) are critical for controlling virus infections, but they also contribute to impaired host immunity and virus persistence. Although IFN-I inhibit HIV-1 replication in vitro and in vivo, persistent IFN-I induction is correlated with immune dysfunction and disease progression in HIV-1 infected patients and in SIV infected monkeys. Moreover, despite efficient suppression of HIV-1 replication with combined antiretroviral therapy (cART), low levels of IFN-I signaling persist in some individuals, which may impede immune recovery and foster viral persistence. We report here studies using a monoclonal antibody to block IFN-α/β receptor (α-IFNAR) signaling in humanized mice (hu-mice) persistently infected with HIV-1. We found that IFNAR blockade rescued human T cells both in number and function despite elevated HIV-1 replication. We further discovered that, in HIV-1-infected mice treated with cART, blocking IFNAR fully reversed HIV-1-induced immune hyper-activation and rescued anti-HIV-1 immune responses. Finally, IFNAR blockade in the presence of cART reduced the size of HIV-1 reservoirs in lymphoid tissues and delayed HIV-1 rebound post-cART cessation. Our results suggest that blocking IFNAR may provide a novel approach to enhance immune recovery and to reduce HIV-1 reservoirs in those who sustain elevated IFN-I signaling during suppressive cART.
Ly6Chi monocytes mediate control of acute alphavirus infection by MAVS-dependent production of type I IFN
Authors: Haist Kelsey Cornell , Burrack Kristina S , Diamond Michael S , Morrison Thomas E 
Published: January 2023

Abstract
Chikungunya virus (CHIKV) and Ross River virus (RRV) are mosquito-borne alphaviruses that cause epidemics of debilitating musculoskeletal disease. Monocytes have been implicated in the pathogenesis of these infections, but their specific roles are less clear. To assess the role of monocytes in alphavirus pathogenesis, we used CCR2-DTR transgenic mice to deplete CCR2+ cells by administering diphtheria toxin (DT). DT-treated CCR2-DTR+ mice showed reduced weight gain and more severe disease following infection and had fewer Ly6Chi monocytes and NK cells in circulation compared to DT-treated WT mice. Also, depletion of CCR2+ cells, but not NK cells alone, restored virulence and increased viral loads in the muscle tissue of mice infected with RRV encoding attenuating mutations in nsP1 to levels seen in monocyte-depleted mice infected with fully virulent RRV. A similar increase in disease severity and viral loads was observed in DT-treated CCR2-DTR+;Rag1−/− mice infected with the nsP1 mutant virus, confirming that these effects are independent of adaptive immunity. In vitro, culture of Ly6Chi monocytes with RRV-infected cells resulted in induction of IFNα gene expression in monocytes that was MAVS-dependent. IFNα expression in monocytes also required new virus particle production from infected cells. Furthermore, monocytes sorted from the circulation and muscle of infected mice had elevated expression of Irf7. Finally, viral loads of the attenuated nsP1 mutant virus were equivalent to those of WT RRV in the muscle tissue of MAVS−/− mice. Collectively, these data suggest that MAVS-dependent production of type I IFN by monocytes is critical for control of acute alphavirus infection and that determinants in nsP1 counteract this response.
Missense SNP in fcer2 (CD23) results in increased transcription factor activity in human B cells
Authors: Chan Marcia , Gigliotti Nicole , Rezaiekhaligh Mo 
Published: January 2023

Abstract
Asthma is one of the most common chronic conditions in children and more than 6 million children in the US are affected. Furthermore, asthma is the third ranking cause of hospitalizations among children younger than 15. Genetics appears to play a role in both predisposition and treatment resistance of asthma. CD23 is expressed on several cell types and has several roles in IgE-mediated immunity, including antigen focusing and presentation to Th2 cells; mediating production of pro-inflammatory cytokines by monocytes; transporting IgE and allergen-IgE complexes through gastrointestinal epithelium; and participating in both positive- and negative-feedback mechanisms in regulation of IgE synthesis by B cells. Single nucleotide polymorphisms (SNPs) in fcɛr2 encoding the low affinity IgE receptor CD23 (FcɛRII) are associated with increased severe asthma exacerbation and increased serum IgE levels. One SNP (C3750T, rs2228137) encodes a nonsynonymous amino acid change (R62W). Stable human B cell transfectants were established expressing either wild type CD23b-R62 or CD23b-R62W. Stimulation through CD23 caused differences in activation of proteins phosphorylated on tyrosine as well as kinetic differences in transcription factor activity. Peripheral blood was obtained from asthmatic patients genotyped for rs2228137. A comprehensive transcriptome analysis of B cells stimulated through CD23 was performed. Nineteen genes were differentially expressed between subjects homozygous for the wild type allele (cc) and those homozygous for the SNP allele (tt). These data suggest that the rs2228137 SNP may influence asthma severity through prolonged activation of B cells favoring increased IgE production.
The complex allergen house dust mite (HDM) acts directly on macrophages to stimulate actin remodeling
Authors: Keegan Achsah Dorsey , Rennoll Sherri A. , Qi Xiulan , Pedra Joao , Vogel Stefanie N , Gao Hongjuan 
Published: January 2023

Abstract
Asthma is a chronic inflammatory disease of the lungs that results in narrowing of the airways, making it difficult to breathe. Dust is a common trigger of allergy and asthma. House dust mites (HDM) are the main cause of dust allergies. Despite the fact that macrophages are present in abundant amounts within the lung and are one of the first cells exposed to HDM upon inhalation, little is known about the direct effects of HDM on macrophages. Here we demonstrate that HDM can directly act on bone marrow-derived macrophages (BMM) leading to actin remodeling. BMM treated with HDM demonstrated alterations in F-actin (based on Phalloidan staining), changes in shape, and increases in size significantly. We also observed alterations in the phosphorylation of cofilin and slingshot, two proteins that regulate actin polymerization. BMM demonstrated constitutive phosphorylation of cofilin with little phosphorylated slingshot at baseline. However, HDM treatment diminished cofilin phosphorylation while increasing the phosphorylation of slingshot. Since HDM is known to contain LPS, important for HDM-induced allergic inflammation in vivo, we examined the contribution of LPS to actin remodeling. Actin cytoskeleton changes were still evident in BMM treated with HDM in the presence of polymyxin B (PMB) or in Tlr4−/− BMM, although the average cell size was reduced compared to HDM alone. The HDM-induced changes in phosphorylation of cofilin and slingshot were unaffected by PMB or Tlr4 deficiency. These results suggest that HDM induces actin remodeling in macrophages by a mechanism that is largely LPS/TLR4-independent. These observations provide an opportunity to identify novel therapeutic targets for allergic asthma.
IL-33 elicits IL-5 dependent eosinophilia in vivo
Authors: Johansson Kristina Britt Charlotte , Malmhäll Carina , Ramos-Ramírez Patricia , Rådinger Madeleine 
Published: January 2023

Abstract

Introduction
Interleukin (IL)-33 plays important roles in the immunopathogenesis of allergy and asthma. It acts by promoting IL-5 and IL-13 secretion from IL-33 responsive type 2 innate lymphoid cells (ILC2s) or directly activates eosinophils and mast cells. IL-5 is crucial in eosinophilic inflammation by controlling differentiation and survival of eosinophils in the bone marrow as well as release and recruitment of eosinophils to the airways. While ILC2s have emerged as important producers of IL-5 in airways, the cellular source of IL-5 in the bone marrow is less explored. In this study we tested the hypothesis that IL-33 induced airway eosinophilia is an IL-5 dependent process in which ILC2s produce IL-5 locally in the bone marrow.

Methods
IL-5 production by bone marrow ILC2s was analyzed by intracellular flow cytometry in a murine model of IL-33 induced airway eosinophilia.

Results
Intranasal IL-33 administration resulted in eosinophil infiltration in airways and increased eosinophils in bone marrow. It was accompanied by a dramatic induction of eotaxin-2 in airways, indicating eosinophil recruitment to the tissue. IL-5+ ILC2s as well as expression of the IL-33 receptor on ILC2s increased in the bone marrow in response to IL-33 administration. Importantly, airway and bone marrow eosinophils were greatly reduced in mice pre-treated with anti-IL-5 antibodies followed by intranasal IL-33.

Conclusion
Our results demonstrate that IL-33 elicits IL-5 dependent eosinophilia in vivo, where ILC2s represent a local source of IL-5 in the bone marrow.
mTORC1 in B cells regulates antibody responses and promotes mitochondrial and metabolic fitness
Authors: McLetchie Shawna , Raybuck Ariel , Cho Sung Hoon , Lin Jingxin , Boothby Mark Robin 
Published: January 2023

Abstract
B lymphocytes migrate among different micro-anatomic sites for diversification, selection, and eventual differentiation into antibody-secreting plasma cells. Emerging evidence supports the premise that aspects of the nutrient milieu vary within lymphoid micro-environments. However, the role of B cell-intrinsic metabolic programs in regulating B cell differentiation and antibody response quality remain unclear. We now show that the amino acid-sensing mTOR complex 1 (mTORC1) is essential for induction of Bcl6 and IRF4, key transcriptional regulators of germinal center and plasma cell fates. mTORC1 also enhances B cell proliferation upon exposure to antigen, increases the rate of somatic hyper-mutation, and is essential for generating high-affinity class switched antibodies. We also find that AMP-activated kinase (AMPK), an intracellular energy sensor, promotes plasma cell differentiation and antibody production. Collectively, these findings suggest mechanisms by which mTORC1 activity is critical for the germinal center reaction and producing class-switched, high-affinity antibodies, and indicate that AMPK can regulate the development and functional properties of terminally differentiated plasma cells.
Supported by NIH R01 AI113292, HL106812, and NCI T32CA009592-29
HLA-F and receptor function on B cells and monocytes in the inflammatory response.
Authors: Geraghty Daniel , Lee Ni , Wang Kevin L , Ishitani Akiko 
Published: January 2023

Abstract
HLA-F is expressed as an open conformer (OC) independent of bound peptide or β2-microglobulin upon surface upregulation on activated immune cells, binding and stabilizing MHC class I (MHCI) in the open conformer form. Functional roles for HLA-F and MHCI as open conformers include HLA-F and MHCI OC as ligands for KIR3DL2 and activating KIRs, and HLA-F as a cofactor with MHCI in a novel antigen acquisition pathway operating in both HLA class I and II antigen presentation. In addition, our work recently detected a unique conformational form of HLA-F expressed in resting B cells, with HLA-F specific mAb 6A4, distinguishing CD27− and CD27+ B cells. Expression of HLA-F on CD14+ monocytes as detected by 4B4 revealed a second unique conformational form. In addition to the ligand and cofactor roles, a third role for HLA-F as a receptor on B cells and monocytes was established. When purified B cells were subjected to specific mAb treatment, upregulation and secretion of IgA and IgM was observed. A receptor function was also detected upon treatment of purified monocytes with specific mAb, where cytokine and chemokine release marking the M1 and M2 monocyte differentiation pathways was observed. Treatment with mAb also resulted in monycyte differentiation into a macrophage and dendritic cell phenotype. When intracellular glycosylation of HLA-F was examined biochemically, the distinct alternate forms detected by anti-HLA-F mAbs 3D11, 6A4 and 4B4 appeared to mark HLA-F transit in different intracellular pathways that were potentially consistent with the different functional roles for HLA-F. Our results indicate HLA-F may play a heretofore unrecognized but highly central role in related inflammatory immune functions and dysfunctions.