bluuebunny/crossref_metadata_2025_split huggingface dataset:
Differential kinetics of CD4+ and CD8+ T cell responses: implications for the assessment of immune responses
Authors: Hokey David A , Graves Andrew J , O’Dee Dawn M , Higgins Adam , Hooper Idris , Schaeffer Jacqueline , Walker Barry 
Published: January 2023

Abstract
Intracellular cytokine staining (ICS) to detect cellular immune responses to infection or vaccination has become routine. Laboratories develop and optimize ICS assays independently and there are many variations of the assay in use today. However, little is known regarding the underlying biology that results in the variable outcomes following assay optimization. The results from the different assays can range from highly similar (slight changes in magnitude) to incredibly different, such as large changes in magnitude or a change in the responding cell types detected. These large differences prompted our investigation into the underlying reasons for the discrepancies in reported data for clinical trials. Here, we used leukapheresis samples from CMV-reactive volunteers to examine viability over time, the effect of costimulation, and response kinetics. The data show that golgi transport inhibitors diminish cell viability after 12 hours of stimulation, emphasizing the importance of viability markers particularly during long stimulations. Costimulatory antibodies increase responses but at the expense of sensitivity, given the increase in background. Interestingly, we found that CD4+ and CD8+ T cells exhibit differential kinetics in their response to CMVpp65 peptides. CD8+ T cells were largely exhausted by 6 hours following the initiation of stimulation, while CD4+ T cells continue to make cytokines well past 20 hours of stimulation. These data have implications on the design of ICS assays and the interpretation of data generated by ICS. Furthermore, the data provide a basis for understanding the differences in ICS assays and provide insight into the kinetics of CD4+ and CD8+ T cell responses.
Tumor blood vessel-associated antigens, DLK-1 and DLK-2, as targets for anti-cancer immunotherapy
Authors: Fabian Kellsye Paula , Storkus Walter 
Published: January 2023

Abstract
Tumors induce pathological angiogenesis that results in a tumor vascular network that is highly abnormal. These aberrant blood vessels support tumor growth and progression and also immunosuppression by promoting a hypoxic and high interstitial fluid pressure-environment that impedes immune trafficking to the tumor bed. Hence, normalization of tumor blood vessels is essential in ensuring success of anti-cancer therapy. We have previously shown that pericytes in tumor blood vessels of murine renal cell carcinoma (RENCA) express the Notch antagonist delta-like homolog-1 (DLK1). DLK1-targeted vaccination of RENCA-bearing mice resulted in vascular normalization and CD8+ T cell-dependent inhibition of tumor growth. Here we show that DLK2, a homolog of DLK1, is upregulated in RENCA pericytes after DLK-1 vaccine-induced removal of DLK1 in the tumor microenvironment (TME). Using recombinant lentiviruses and adenoviruses encoding full-length DLK1 and DLK2, we demonstrate that coordinate vaccination against these two antigens leads to superior antigen-specific CD8+ T cell-induced tumor growth inhibition than vaccination with either antigen alone. We also show that DLK-1 and DLK-2 coordinate vaccination shifts the TME to a more “immunocompetent” phenotype by normalizing tumor vasculature and decreasing the immunosuppressive cell population. We conclude that specific vaccination targeting DLK-1 and DLK-2 is a promising approach for the treatment of vascularized tumors.
Toll-like receptor ligands in conjunction with agonist anti-OX40 mAb immunotherapy possess differential capacity to revive anergic self-reactive CD8 T cells
Authors: Kasiewicz Melissa J. , Farhad Mohammad , Redmond William L. 
Published: January 2023

Abstract
Previous work has shown that the induction of CD8 T cell anergy to a prostate-specific self-antigen can be reversed following ligation of the TNFR family member OX40 (CD134) with an agonist anti-OX40 mAb plus vaccination. The provision of the Toll-like receptor 4 (TLR4) ligand bacterial lipopolysaccharide (LPS) further enhanced T cell responses following combined anti-OX40/vaccine immunotherapy. Although this approach facilitated increased CD8 T cell infiltration into the prostate, these cells were unable to elicit significant destruction of the prostate tissue. In the current study, we examined the extent to which co-administration of select TLR ligands would affect the ability of anti-OX40/vaccine therapy to revive prostate-specific CD8 T cells rendered anergic in vivo. These data revealed a differential capability of TLR agonists in conjunction with anti-OX40/vaccine immunotherapy to generate robust effector CD8 T cells capable of trafficking into normal prostate epithelia and inducing tissue-specific damage. These data provide the framework for translational studies to evaluate the efficacy of administering OX40 agonists with tumor-specific vaccination and TLR ligand adjuvants to maximize T cell-mediated therapeutic responses.
Efficacy of an oral recombinant attennated Salmonella vaccine encoding cjaA in turkeys colonized with Campylobacter jejuni
Authors: Sylte Matthew Jon , Looft Torey P. , Chandra Lawrance C. , Bearson Shawn M. , Bearson Bradely L. , Susta Leonardo 
Published: January 2023

Abstract
Campylobacter jejuni spp. jejuni, (C. jejuni) is a commensal bacterium in the lower gastrointestinal tract of poultry (e.g., chickens and turkeys), where it causes no clinical disease. Consumption of C. jejuni-contaminated poultry products is the most likely route of transmission to humans. Strategies previously attempted in chickens to reduce intestinal C. jejuni colonization include administration of probiotics, non-antibiotic feed additives, vaccines and bacteriophages. These interventions have not been evaluated in turkeys. Recombinant attenuated Salmonella vaccines (RASV) encoding C. jejuni cjaA are reported to reduce cecal C. jejuni colonization in chickens. The goal of this study was to evaluate the efficacy of a RASV encoding cjaA in turkey poults experimentally colonized with C. jejuni. The cjaA gene from C. jejuni was cloned into RASV strain χ8133 (RASV-cjaA). A control RASV χ8133 (RASV-control) was generated lacking a cjaA insert. At different dates of age, poults received either one or two doses of RASV-cjaA. Other poults received 2 doses of RASV-control. Intestinal colonization by RASV-cjaA was evaluated and was detectable up to 6 days after oral inoculation. Poults were orally challenged with C. jejuni and euthanized at days 3, 7, 14 and 28 days post-challenge to evaluate cecal colonization (cfu/g content) and histopathological changes. Poults vaccinated with 2 doses of RASV-cjaA showed significant reduction (P&lt;0.01) in C. jejuni cecal colonization and histopathological lesion scoring, as compared to those receiving a single dose of RASV-cjaA or RASV-control. These data demonstrate that oral RASV-cjaA significantly reduced C. jejuni colonization in turkeys, which may enhance food safety.
The role of IRF7 and NF-κB pathways in the induction of antiviral responses in chicken tracheal epithelial cells following exposure to TLR3 and 4 ligands
Authors: Barjesteh Neda , Abdelaziz Khaled Taha , Sharif Shayan 
Published: January 2023

Abstract
With recent outbreaks of avian influenza viruses (AIV), there is a need for better understanding of host responses in the respiratory system to AIV in order to develop and improve prophylactic procedures for control of AIV. Toll-like receptors (TLRs) are molecules in the immune system that rapidly induce antiviral responses in the host. Activation of TLRs by their ligands leads to a variety of cellular responses, including the production of interferons (IFNs). However, there is little information about induced antiviral responses in chicken trachea following TLR ligand treatment. We cultured primary chicken tracheal epithelial cells (cTECs) to determine antiviral responses induced by TLR3 and 4 ligands. These cells were treated with polyI:C and LPS from Escherichia coli 026:B6, TLR3 and 4 ligands respectively, before AIV infection. We also used BX795 and celastrol, inhibitors of IRF7 and NF-κB pathways, respectively, in order to gain insight into the mechanisms involved in the induced antiviral responses. Our results showed that treatment of cTECs with TLR ligands reduced the replication of AIV in cTECs. Treatment of cTECs with polyI:C resulted in highest reduction of viral replication in cTECs followed by treatment of cTECs with LPS. BX795 increased AIV replication in polyI:C and LPS treated cells while celastrol increased AIV replication in polyI:C treated cells. Overall, these results provided evidence that cTECs are able to mount antiviral responses against AIV after TLR ligand stimulation. Moreover, we determined the role of IRF7 and NF-κB signalling pathways in the induction of antiviral responses. Studies are underway to examine the ability of cTECs to produce type I IFNs and the effect of other cells, such as macrophages, on cTECs.
Host genetic factors and miRNA-linked dendritic cell responses associated with the outcome of treatment response in HIV-1/HCV co-infected individuals
Authors: Jain Pooja , Sehgal Mohit , Narang Rubina , Ginwala Rashida , Zeremski Marija , Talal Andrew , Elrod Elizabeth , Grakoui Arash , Li Qi-Ging , Philip Ramila , Khan Zafar 
Published: January 2023

Abstract
HIV-1/HCV co-infection is a significant burden on global economy and public health. PEGylated interferon and ribavarin remain the essential components of anti-HCV treatment. We investigated the host genetic and immunological correlates of successful treatment response in conjunction with mechanisms by which PEG-IFN is able to clear the virus in a cohort of HIV-1/HCV co-infected individuals undergoing IFN/RBV treatment. First we demonstrate that functional state of DCs before/during therapy influences the treatment outcome. Upon genotyping IFNL3 polymorphisms rs12979860, rs4803217 and ss469415590, we found rs12979860 to be a better predictor of treatment outcome. Next, we compared the expression of forty-six IFN-stimulated genes prior to and after the treatment and observed that pre-treatment levels of several ISGs were higher in SVRs compared to NRs. In continuation, we identified miRNAs whose expression can be regulated by IFN in periphery. miRNA expression patterns in mDCs and pDCs in response to IFN-α were examined and observed miR-221 downregulation via IFN induced STAT3 inhibition in both. Using in-silico approaches followed by experimental validation, CDKN1C, CD54 and SOCS1 were identified as miR-221 targets. Moreover, miR-221 overexpression in mDCs enhanced their secretion of proinflammatory cytokines IL-6 and TNF-α but reduced the secretion of anti-inflammatory cytokine IL-10. These observations were extended and correlated with those obtained with patients’ PBMCs, hepatocytes and kupffer cells from HCV infected individuals as well as individuals with alcoholic cirrhosis. In summary, these studies demonstrated the role of IFN-α/miR-221 axis in HCV pathogenesis and response to IFN-based treatments.
The survival of monocytes infected by human cytomegalovirus is dependent on a unique signalsome induced during entry.
Authors: Peppenelli Megan A , Arend Kyle C , Cojohari Olesea , Moorman Nathaniel J , Chan Gary C 
Published: January 2023

Abstract
HCMV infection of immunocompromised individuals often leads to multi-system organ failure. The development of multi-system organ failure is dependent on the ability of HCMV to spread to peripheral organs, which is mediated by blood monocytes. We have previously shown HCMV to extend the short 48hr lifespan of monocytes. Mechanistically, HCMV upregulated cellular myeloid leukemia sequence 1 (Mcl-1) and heat shock protein 27 (HSP27), to block the proteolytic cleavages necessary for the formation of active caspase 3. We now show that, compared to other myeloid survival factors, HCMV infection more efficiently upregulates Mcl-1 and the only survival factor to rapidly induce HSP27. These observations, suggest a unique virus specific mechanism of induction; thus, we examined receptor: ligand signaling events, since viral anti-apoptotic proteins are not expressed until differentiation into macrophages is complete. We determined that HCMV-induced EGFR/PI3K/AKT signaling increased the transcription of both Mcl-1 and HSP27. Yet, protein expression of Mcl-1 and HSP27 is increased via the gH/aVb3/src and the gB/EGFR pathways, respectively. To address this discrepancy between transcripts and protein we evaluated the effects of HCMV on the translation of Mcl-1 and HSP27. Indeed, we found, in contrast to myeloid survival factors, HCMV stimulated the translation of Mcl-1 and HSP27. Overall, these data indicate that inhibition of apoptosis by HCMV is through the unique signaling regulation of Mcl-1 and HSP27; ensuring short-lived monocyte survival past the normal 48hr viability checkpoint, a key event necessary for viral dissemination.
Transcriptional regulation in the development of CD8ab+ iNKT cells in a humanized mouse model
Authors: Yuan Weiming , Wen Xiangshu , Li Michelle , Xiong Ran , Bosselut Remy , Rao Ping 
Published: January 2023

Abstract
Invariant NKT (iNKT) cells are innate-like T cells showing potent anti-tumor function in conventional mouse models. In sharp contrast, iNKT cell ligands have had limited efficacy in human anti-tumor clinics, mostly due to the profound differences in the properties and compositions of iNKT cells between human and mice, in particular, the presence of CD8+ iNKT cells only in humans. To build more relevant in vivo models for studying human iNKT cells, we first developed a CD1d-humanized mouse model (hCD1d-KI) with human CD1d knocked in (Proc. Natl. Acad. Aci. USA 110: 2963–8, 2013). To further humanize the mouse model, we introduced human invariant TCRa chain (Va24Ja18) into the hCD1d-KI mice. Interestingly, we observed a substantial subset of iNKT cells expressing CD8ab in the new humanized mouse model (J. Immunol. 195:1459–69, 2015). The CD8ab+ iNKT cells show a strong Th1-biased cytokine response and potent cytotoxicity upon activation. The low binding of iNKT TCRs to human CD1d/lipid complex and high prevalence of Vb7 TCRb among these CD8+ iNKT cells strongly suggested a novel low avidity-based developmental program for the ontogeny of these iNKT cells which includes the suppression of the transcriptional factor, Th-POK. This suppression of Th-POK is essential for the development of CD8ab+ iNKT cells as these iNKT cells can not be detected upon forced expression of Th-POK early in T cell development. Our establishment of the new mouse model extensively humanized in CD1d/iNKT TCR system will greatly facilitate the investigation of in vivo functional properties of human iNKT cells as well as future design and optimization of iNKT cell ligands for diverse immunotherapies.
The Dendritic Cell MHC II Peptidome Derives from a Variety of Processing Pathways and Includes Peptides with a Broad Spectrum of HLA-DM Sensitivity
Authors: Stern Lawrence , Santambrogio Laura 
Published: January 2023

Abstract
The repertoire of peptides displayed in vivo by MHC II molecules derives from a wide spectrum of proteins produced by different cell types. While intracellular endosomal processing in dendritic cells and B cells has been characterized for few antigens the overall range of processing pathways responsible for generating the MHCII peptidome are currently unclear. To determine the contribution of non-endosomal processing pathways we eluted and sequenced over 3000 HLA-DR1-bound peptides presented in vivo by dendritic cells. Processing enzymes were identified by reference to a database of experimentally determined cleavage sites and experimentally validated for four epitopes derived from complement 3, collagen II thymosin b4 and gelsolin. We determined that self-antigens processed by tissue-specific proteases, including complement, matrix metalloproteases, caspases, and granzymes, and carried by lymph, contribute significantly to the MHC II self-peptidome presented by conventional dendritic cells in vivo. Additionally, presented peptides exhibited a wide spectrum of binding affinity and HLA-DM susceptibility. The results indicate that the HLA-DR1-restricted self-peptidome presented under physiological conditions derives from a variety of processing pathways. Non-endosomal processing enzymes add to the number of epitopes cleaved by cathepsins, altogether generating a wider peptide repertoire. Together with HLA-DM-dependent and HLA-DM-independent loading pathways, this ensures that a broad self-peptidome is presented by dendritic cells.
MHCII with full peptide occupancy is a substrate for HLA-DM catalyzed peptide exchange
Authors: Reyes-Vargas Eduardo , Barker Adam P , Zhou Zemin , He Xiao , Jensen Peter E 
Published: January 2023

Abstract
The major histocompatibility complex class II (MHCII) antigen-processing pathway is critical for adaptive immunity as the mechanism through which ligands are generated for recognition by CD4 T cells. HLA-DM edits the repertoire of peptides that are presented to T cells by catalyzing multiple rounds of peptide exchange on MHCII molecules. The catalytic potency of DM varies such that peptide exchange is markedly enhanced on some peptide-MHC (pMHC) complexes and not others. This capacity to differentially edit complexes influences specificity and immunodominance in T cell responses. The catalytic mechanism and rules that govern DM susceptibility are not fully understood. Evidence suggests that partial occupancy at the N-terminus of bound peptides is critical for complex sensitivity to DM. We employed a fluorescence polarization assay to independently assess the effective affinity and catalytic turnover of DM, components of the catalytic mechanism, for pMHC complexes in real-time kinetic measurements of DM activity. We found that a complex with an N-terminal truncated form of the immunodominate HA peptide, resulting in the loss of three H-bonds mediated by the P-2 and P-1 residues, showed an expected decrease in complex stability and increase in affinity for DM binding, but an unexpected increase in the catalytic turnover when compared to the unmodified HA-MHC complex. This result was even more pronounced when a conserved bidentate H-bonding residue near the P1 anchor was disrupted by site-directed mutagenesis on an HA-MHC complex. Together, this implies that pMHC interactions near the N-terminal segment of a peptide may be intact in the functionally active DM-pMHC catalytic complex and provide insights into the DM catalytic mechanism.
Targeting of B cell class-switching and plasma cell survival pathways by a Rab7 inhibitor suppresses autoantibody responses in murine lupus
Authors: Lou Zheng , Lam Tonika , Kulp Dennis , Wang Rui , Taylor Julia , Casali Paolo , Xu Zhenming 
Published: January 2023

Abstract
B cell class switch DNA recombination (CSR) and generation/maintenance of plasma cells underpin the production of pathogenic IgG autoantibodies in lupus. To address whether these two B cell lineage-specific processes can be targeted to inhibit the autoantibody response in lupus, we used a small molecule compound to specifically block the activity of the small GTPase GTPase, as prompted by our previous findings that Rab7 plays a B cell-intrinsic role in antibody responses. Inhibition of Rab7, which was upregulated in lupus B cells, suppressed pathogenic autoantibody production, arrested disease development and extended the life-span in MRL/Fas and Sle1/Sle2/Sle3 lupus-prone mice. The reduction in class-switched autoantibody response occurred despite normal numbers and functions of T cells, dendritic cells and macrophages, but was associated with significantly decreased numbers of IgG-expressing B cells and plasma cells. Accordingly, B cell class-switching and plasma cell survival, but not B cell differentiation into plasma cells, were inhibited by the Rab7 inhibitor through suppression of NF-kB activation, as revealed by impairment in NF-kB-dependent gene expression. Conversely, CSR and plasma cell survival could be rescued by an IKKb constitutively active mutant. These findings suggest a small molecule that targets two important B cell differentiation stages to effectively blunt autoantibody responses in lupus.
Supported by NIH grants AI 079705 and AI 105813 and the Alliance for Lupus Research (ALR) grant ALR 295955 to P.C.
Endogenous dsRNA and dsDNA sensing is increased in transitional B cells in BXD2 autoimmune-prone mice
Authors: Mountz John D , Hamilton Jennie Ann , Wu Qi , Yang PingAr , Luo Bao , Liu Shanrun , Li Jun , Hsu Hui-Chen 
Published: January 2023

Abstract
Enhanced nucleic acid-sensing (NS) pathways has been associated with development of autoimmune disease. Increased type I IFN signaling is identified in lupus prone BXD2 mouse B cells and there is a predominant production of autoantibodies against RNA binding autoantigens in BXD2 mice. Determination of the expression of Usp18 and Oasl2 in early CD93+ transitional B cells (T1 and T2) revealed a surprising finding that early transitional B cell exhibited elevated type I IFN signature gene expression, compared to CD93− mature B cells. Consistent with this, there is also elevated expression of NS pathway genes in early transitional B cells. The most highly expressed were the dsRNA sensing Rig1 and transcription factors Irf3 and Irf7, which were expressed at higher levels compared to pDCs. Interestingly, although Usp18 and Oasl2 levels were undetectable in BXD2-Ifnar−/− transitional B cells, dsRNA sensing gene expression were highly expressed suggesting an IFNAR-independent up-regulation of dsRNA NS pathway in transitional B cells. There was lower expression of Mda5 and Pkr5, and genes involved in DNA sensing including Zbp1 and Irf9. Stimulation of B cells with a RIG1 agonist, poly(I:C)-LMW/LyoVec leads to significantly elevated induction of NS pathway genes in both WT and IFNAR deficient transitional B cells from BXD2 mice. These results suggest that transitional B cells have an intrinsic enhanced nucleic acid-sensing pathway, allowing them to rapidly react to nucleic acid stimulation. Dysregulation of this pathway as a result of apoptotic cell clearance defects can act together with type I IFN stimulation to lead to survival and maturation of B cells that by-pass tolerance checkpoint 2 and produce anti-nucleic acid autoantibodies in lupus.