bluuebunny/crossref_metadata_2025_split huggingface dataset:
Magnetic bead-based 28-plex immunoassay for the quantification of rhesus and cynomolgus monkey cytokines (144.34)
Authors: Brodey Mary , Liu Nina , Huuskonen Jarkko , Beernink Hans 
Published: January 2023

Abstract
Non-human primates possess high degrees of homology with humans, and thus are crucial model systems in a number of areas of biomedical research (neuroscience, endocrinology, reproductive physiology, cardiovascular physiology, infectious diseases, vaccine and drug efficacy and safety). Monitoring complex cytokine profiles has proven to be invaluable in these various areas. Using the Luminex® xMAP® platform, Life Technologies has developed a 28-plex magnetic bead-based cytokine immunoassay for the analysis of rhesus monkey (Macaca mulatta) and cynomolgus monkey (Macaca fascicularis) samples. Validated sample types include serum, plasma, and conditioned tissue culture medium. Analytes include: EGF, eotaxin, FGF basic, G-CSF, GM-CSF, HGF, IFN-γ, IL-1β, IL-1RA, IL-2, IL-4, IL-5, IL-6, IL-8, IL-10, IL-12 p40/p70, IL-15, IL-17, I-TAC, MDC, MIF, MIG, MIP-1α, MIP-1β, MCP-1, RANTES, TNF-α, and VEGF. This magnetic-bead based assay is compatible with robotic washing systems, and thus may be fully automated. Total incubation time is 3.5 hours. The utility of the assay is demonstrated by determining cytokine profiles obtained with LPS-stimulated or PMA plus A23187-stimulated peripheral blood mononuclear cells (PBMCs) from rhesus monkeys and cynomolgus monkeys. Linearity of dilution experiments with tissue culture medium and serum produce R2 values of 0.98 and greater, while sample recovery routinely falls between 80 and 120%. Intra-assay and inter-assay precision is ≤10% CV.
Donor Tr1 cells potently expand in vivo and prevent GVHD after TLI/ATS non-myeloablative conditioning and allogeneic bone marrow transplantation (145.17)
Authors: Pillai Asha , van der Merwe Marie , Ong Taren , Vogel Peter , George Tracy 
Published: January 2023

Abstract
Murine total lymphoid irradiation (TLI) + anti-thymocyte serum (ATS) has been well-translated to human transplantation (Lowsky et al, NEJM 2005). Though host TLI/ATS treatment allows post-transplant donor Foxp3+ Treg expansion, &gt;90% of donor CD4+CD25+ cells are Foxp3neg. We now show that the latter are Tr1-type regulatory T cells. BALB/c (H-2d) hosts receiving TLI/ATS + bone marrow + splenocytes (BMT) from C57BL/6 (H-2b) donors had 90% 60-day survival without GVHD (n=10). BALB/c hosts given 800 cGy or 400 cGy total body irradiation (TBI)/ATS+BMT had 100% 30-day GVHD death (n=10). TLI/ATS+BMT hosts had a 10-fold increase in H-2Kb+CD4+CD25+ cell absolute number in day 6 host spleen versus TBI/ATS controls (p&lt; 0.05). Using Foxp3-GFP transgenic donors, &gt;90% of the H-2Kb+CD4+CD25+ cells after TLI/ATS+BMT are Foxp3(GFP)neg. Supernatant assay on day 6 sorted TLI/ATS+BMT H-2Kb+CD4+CD25+, 72-hour PMA/ionomycin stimulated cells revealed potent IL-10, low IL-2, IL-5, and IFN-γ, and no IL-4 or TNF-α (p&lt; 0.05) secretion. Day 6 TLI/ATS+BMT Tr1 donor cells were potent suppressors of donor-host MLR. Donor CD25 cell depletion pre-TLI/ATS+BMT caused day 6 donor CD8+ T cell-mediated GVHD and loss of donor Tr1 cells, indicating that Tr1 cell expansion protects from GVHD post-TLI/ATS+BMT. We are characterizing a host APC-dependent mechanism for this Tr1 expansion. Our findings delineate a novel mechanism and strategy for in vivo Tr1 expansion for tolerance induction and adoptive immunotherapy.
Stem cell mobilization with G-CSF induces Th-17 differentiation and promotes scleroderma (145.19)
Authors: MacDonald Kelli , Olver Stuart , Kuns Rachel , Varelias Antiopi , Raffelt Neil , Don Alistair , Markey Kate , Wilson Yana , Smyth Mark , Iwakura Yoichiro , Tocker Joel , Clouston Andrew , Hill Geoff 
Published: January 2023

Abstract
The clinical shift toward utilizing G-CSF mobilized stem cell grafts has resulted in a striking increase in chronic GVHD after transplantation although the mechanisms involved are unclear. Comparison of cytokine expression following TCR activation of splenocytes from naïve and G-CSF treated B6 or BALB/c donors (low and high responders respectively) showed little effect of G-CSF on Th1 or Th2 cytokine production. In contrast, IL-17 production was dramatically enhanced in response to G-CSF in both strains. G-CSF induced IL-17 production occurred in both CD4 and CD8 conventional T cells and by using relevant knock-out mice or blocking reagents we demonstrated that this was independent of IL-6, TGF-beta or IL-23 signalling. However, the induction of IL-17 by G-CSF was completely dependent on IL-21 signalling and G-CSF induced IL-21 generation in CD4 T cells independent of IL-17 itself. We utilized multiple models of GVHD using G-CSF mobilized B6 or BALB/c wild-type or IL-17 deficient donors, in both MHC matched and mismatched settings. Surprisingly, IL-17 was critical for the induction of sclerodermatous chronic GVHD occurring after transplant using either donor strain. Importantly, IL-17 controlled the dramatic sequestration of macrophages into skin that coincided with the fibrogenic response. This study provides a logical explanation for the propensity of allogeneic stem cell transplantation to invoke sclerodermatous GVHD and suggests a therapeutic strategy for intervention.
Cytotoxic T Lymphocytes (CTL) specific for CMV, adenovirus, and EBV can be generated from naive T cells for adoptive immunotherapy (145.22)
Authors: Hanley Patrick , Cruz Conrad Russell , Savoldo Barbara , Leen Ann , Decker William , Moldren Jeffrey , Liu Hao , Gee Adrian , Demmler Gail , Heslop Helen , Rooney Cliona , Dotti Gianpietro , Brenner Malcolm , Rodgers John , Shpall Elizabeth , Bollard Catherine 
Published: January 2023

Abstract
Adoptive transfer of virus-specific T cells can effectively restore anti-viral immunity after stem cell transplant. However, standard protocols for eliciting virus-specific T cells from adult seropositive donors have been ineffective when applied to cord blood (CB) or virus-naïve adult donors. We tested alternative techniques for eliciting primary T cell responses in vitro. Culturing APC transduced with an Ad5f35pp65 vector and cytokines IL-2,7,12 and 15, we generated T cells specific for multiple viruses from CB and CMV-seronegative (CMVneg) donors. 9 CB-derived T cell lines contained 87% (range 81-94) CD8+ and 26% (12-40) CD4+ T cells. Specificity was determined in 51Cr release and IFN-γ ELISPOT assays against CMVpp65, adenovirus, and EBV targets. Results showed mean spot forming cells (SFC) following incubation with CMVpp65 (209;range 45-694), Adhexon (74;0-128), and EBV (157;23-291) targets. Further, CMVpp65-specific T cells expanded from 7 CMVneg adult donors showed mean 142(38-410) SFC to CMVpp65. CB and CMVneg-derived T cells derived only from naïve (CD45RA+/CCR7+) T cells and recognized "unconventional" CMVpp65 epitopes as identified by overlapping pp65 peptide pools using IFN-γ ELISPOT, suggesting an inherent difference between naïve and memory T cell responses to CMV. Virus-specific T cells can now be generated from naïve sources and could potentially be used clinically in these highest risk patients after transplant.
Identification of a novel immunosuppressive DC subpopulation in tolerized heart allografts (145.23)
Authors: Diao Jun , Zhao Jun , Mikhailova Anastassia , Cattral Mark 
Published: January 2023

Abstract
The role of dendritic cells (DC) in the maintenance of allograft tolerance is unclear. We analyzed the composition and function of DC infiltrates in tolerized murine heart allografts. Methods. B6 donor splenocytes (Day -7) plus anti-CD4 antibody (Day -1, 2 5 and 7) were injected into BALB/c recipients of B6 heart allografts; mice with heart grafts beating &gt; 80 days were considered tolerant. Syngenic heart grafts (B6 to B6) were used as control. Results: We found that graft-infiltrating Lin-CD11c+MHCII+ cDC contained a novel Gr-1+ cDC subpopulation. B220+CD11c+MHCII+ plasmacytoid DC (pDC) were undetectable. In contrast to allograft Gr-1- cDC, which resemble typical immature tissue cDC, Gr-1+ cDC express lower levels of MHC class II and CD11c, adhere to culture plates, exhibit resistance to maturation stimuli, and stimulate allogeneic lymphocytes poorly. Gr-1+ cDC are distributed uniquely in tolerant allografts but not syngenic grafts or acutely rejecting allografts. Gr-1+cDC are absent in lymphoid tissues. Our previous studies have shown that the cytokine milieu of tumors skews pre-cDC differentiation towards Gr-1+ cDC. We found that pre-cDC exist in heart allografts. It remains to be determined whether tolerized hearts promote pre-cDC to differentiate into Gr-1+ cDC in situ. Conclusion: Tolerized allografts contain an immunosuppressive DC subpopulation that may contribute to the maintenance of long-term allograft survival.
Modulation of influenza-specific humoral recall responses by CD154 blockade and cyclosporine A (145.28)
Authors: Azimzadeh Agnes , Schroeder Carsten , Zhang Tianshu , Ramirez-Medina Karina , Avon Chris , Pasetti Marcella , Pierson N.III Richard 
Published: January 2023

Abstract
Background: Current methods of immunosuppression are broadly inhibitory and thus associated with an increased risk of opportunistic infections. Here we compare the effect of CD154 blockade (αCD154) and Cyclosporine A (CsA) therapy on secondary responses to influenza, when the first immunization was performed before therapy, as might be done in transplant wait-listed patients. Methods. Sixteen cynomologus monkeys were immunized with influenza vaccine in absence of immunosupression and administered a secondary boost immunization with either αCD154 mAb (IDEC-131), CsA, or no treatment. Ab responses against three influenza antigens (H1N1, H3N2, B) were measured by ELISA. An average fold increase &gt;1.4 was considered positive. Results. 14 of 16 animals responded to the primary immunization (I) performed in absence of therapy. In untreated controls, 4/5 animals mounted a strong secondary response (II). With αCD154, 4/6 animals showed mild to strong IgM and IgG, and 2/6 demonstrated either IgM or IgG responses. In contrast, in CsA-treated animals, only 2/5 produced IgM without IgG, and 3/5 mounted low titers of IgG but no IgM. Conclusions. Relative to CsA, αCD154 only moderately suppressed secondary Ab responses to flu antigens. This suggests that αCD154 treatment spares important facets of adaptive immunity against opportunistic infections, potentially conferring a clinically significant advantage over conventional immunosuppression with respect to common pathogens
Detection and ex vivo expansion of dormant anti-viral CTL precursors isolated from recipients of unrelated umbilical cord blood transplant (UCBT). (145.31)
Authors: Szabolcs Paul , Marti Luciana , Leen Ann , Baker Jennifer , Janetzky Sivia 
Published: January 2023

Abstract
Two potentially fatal viruses ,CMV and adenovirus can be detected 2-4 weeks after UCBT. Infection may trigger in vivo priming of the infused na ve T cells despite immunosuppressive (IS)medications. We hypothesized that 1) threshold numbers of CTL precursors can be identified that may influence outcome 2) CTL precursors may reside below the level of detection during IS. Methods: Monocytes infected with an adenovirus vector encoding CMV pp65 (Ad5f35pp65) were used as APC in the presence of IL7. Antiviral CTLp was quantitated by IFNg ELISPOT among 1x10e5 responders after overlapping peptide pool stimulation. Results: 8 infected patients at a median age of 9.9 years have been studied before day 100 . Only 1/3 patients with adenovirus exhibited viremia while all 5 patients with CMV had viremia. In freshly drawn blood, 1/8 patients had any detectable reactivity. However, after 9-12 days of stimulation we could enumerate CTLp in 7/8 patients. For adenovirus, the median hexon-specific response was 52 SFC, and the penton-specific CTLp frequency was 13 SFC. CMV pp65-specific response was 6.7 SFC. No responses were found against CMV EI-1 demonstrating critical role for antigenic restimulation. All patients survive beyond 6 months. Conclusion: To our knowledge these are the first data demonstrating that dormant virus-specific CTLp are present already in the first 3 months in infected UCBT recipients. Although these rare events are below detection ex vivo expansion permits quantitation.
High-mobility group box1 contributes to kidney ischemia reperfusion injury through TLR4 signaling (145.38)
Authors: Wu Huiling , Ma Jin , Wang Peng , Wyburn Kate , Chadban Steven 
Published: January 2023

Abstract
Kidney ischemia reperfusion injury (IRI) is TLR4 dependent. HMGB1 is a TLR4 endogenous ligand expressed within IRI kidney. We aimed to determine 1) whether HMGB1 contributes to kidney IRI; 2) whether this is via a direct HMGB1-TLR4 interaction; 3). therapeutic potential of neutralizing antibody to HMGB1 in kidney IRI. Methods: Wild-type (WT) and TLR4-/- mice received anti-HMGB1 Ab, rHMGB1, or appropriate controls by ip injection 1 hour before 22 min ischemia or immediately after reperfusion. Results: Mice pretreated with anti-HMGB1 Ab were protected against kidney IRI, with lower serum creatinine and less tubular damage versus control mice (p&lt;0.05-0.001). Tubulo-interstitial neutrophil (day 1, p&lt;0.001) and macrophage (day 5, p &lt;0.001) infiltration were less in anti-HMGB1 Ab treated mice versus controls. IL6 & TNF-a mRNA levels were reduced in anti-HMGB1 Ab treated IRI kidneys (p&lt;0.05-0.001). Conversely, administration of rHMGB1 post reperfusion exacerbated kidney IRI in WT mice. TLR4-/- mice were protected against kidney IRI, a finding that was unchanged by administration of either anti-HMGB1 Ab or rHMGB1. As with pretreatment, treatment of neutralizing Ab to HMGB1 after reperfusion also protected against kidney IRI. Conclusion: Endogenous HMGB1 is a mediator of kidney damage following IRI, which may operate through the TLR4 pathway. Administration of a neutralizing antibody to HMGB1, either prior to or soon after IRI, afforded significant protection.
STAT1-deficient bone marrow prevents GVHD by modulating plasmacytoid DCs while preserving vaccine-mediated responses to tumor (145.40)
Authors: Capitini Christian , Larabee Shannon , Qin Haiying , Song Young , Khan Javed , Mackall Crystal , Fry Terry 
Published: January 2023

Abstract
A barrier to using donor lymphocyte infusions (DLI) after allogeneic blood and marrow transplantation (alloBMT) is induction of graft versus host disease (GVHD). We have previously demonstrated that gamma interferon (IFNg) receptor-/- bone marrow abrogates GVHD after T cell depleted (TCD) alloBMT, however the mechanism remains elusive. Since the IFNg receptor signals through STAT1, we hypothesized that STAT1-/- bone marrow could also prevent GVHD. Using TCD minor histocompatibility antigen-mismatched alloBMT with delayed DLI and tumor vaccines, we demonstrate that STAT1-/- whole bone marrow protects against a leukemic and solid tumor challenge as efficiently as wild type marrow, but STAT1-/- marrow prevents GVHD-associated weight loss. Using a Cre-Flox system, we generated mice that had STAT1 selectively deleted in either CD11c+ (Dendritic cells, DCs) or lysozyme+ (macrophage/neutrophils) cells, but as marrow donors neither prevented GVHD, implying other STAT1-/- cell subsets are critical. Prior to DLI administration, an increase in splenic plasmacytoid DCs in recipients of STAT1-/- marrow is observed. Microarray analysis of pDCs from recipients of STAT1-/- marrow shows marked upregulation of S100A8 and S100A9, molecules typically associated with myeloid-derived suppressor cells. Taken together, STAT1-/- marrow replicates the effects of IFNg receptor-/- marrow in terms of preventing GVHD and improved efficacy of DLI and vaccines, but also has immunomodulatory effects on pDCs.
TRAF6, a new member of the proximal signaling complex recruited by BAFFR and TACI in B lymphocytes (34.1)
Authors: Hildebrand Joanne , Bishop Gail 
Published: January 2023

Abstract
The cytokines BAFF (B-cell activating factor of the TNF family, BLyS) and APRIL (a proliferation-inducing ligand) activate several major signaling cascades responsible for B-cell survival and homeostasis. Made primarily by myeloid cells, BAFF binds and activates three cell membrane receptors; BCMA (B-cell maturation antigen), TACI (transmembrane activator and CAML interactor), and BAFF-R (BAFF Receptor, BR3), while APRIL binds TACI and BCMA. Studies of genetically altered mice demonstrate that TACI and BCMA perform niche roles in B-cell Ig isotype switching and plasma cell maintenance. In contrast, BAFF-R deletion in mice results in greater than 90% loss of mature B-cells, revealing it as an essential mediator of B-cell maturation and survival beyond the immature transitional (T1) stage. TNF receptor family proteins utilize characteristic combinations of TNF receptor associated factors (TRAFs) to bridge receptor activation with a multitude of downstream signaling pathways. To date, studies of TRAF recruitment by TACI and BAFFR and their roles in signaling have been limited, and TRAF3 was the only TRAF thought to associate with these receptors. Here we present the novel finding that TRAF6 is directly recruited by TACI and BAFFR in B-cells, and that it forms an essential link to the initiation of the canonical NF-kB pathway and other pro survival signals.
Molecular cloning of human IL-16 gene and its expression in Escherichia Coli (34.17)
Authors: Si Chuanping , Li Shuling , Gao Huijie , Fu Jia 
Published: January 2023

Abstract
Objective: To clone human IL-16 gene, and construct its prokaryotic expression vector, and express it in E. coli DH5. Methods: The total RNA was extracted from peripheral blood mononuclear cells stimulated by histamine. And hIL-16 cDNA was amplified by using RT-PCR and cloned into pUC18 T-vector. After the sequence of hIL-16 cDNA was confirmed, the cDNA was inserted into prokaryotic expression vector pMAL-C2. The recombinant expression plasmid pMAL-IL-16 was identified by endonucleases digestion and PCR, and then transformed into E. coli DH5, human IL-16 expressed in DH5 was identified by SDS-PAGE and Western blotting method. Results: Obtained cDNA of hIL-16 was identical with that published on Genebank, and the prokaryotic expression vector pMAL-IL-16 was constructed correctly. Recombinant protein was expressed in E.coli when induced with IPTG and had a molecular weight of 60kD, wich is consistent with theoretical calculation. Conclusion: The hIL-16 cDNA was cloned and the prokaryotic expression plasmid pMAL-IL-16 was constructed successfully. The E. coli DH5 that stably expressed hIL-16 was obtained, which provide experimental basis for the further studying of functional activities and application of hIL-16. [Key words] Human interleukin-16, Molecular cloning, Prokaryotic expression
DBM1285 suppresses LPS-induced TNF-α production by blocking p38 MAPK/MK2 signaling pathway in macrophages (35.6)
Authors: Park Song-Kyu , Choi In Young , Kang Jong Soon , Han Sang-Bae , Lee Chang Woo , Lee Kiho , Kim Hwan Mook 
Published: January 2023

Abstract
TNF-α is a major inflammatory cytokine that plays an important role in the development of various inflammatory diseases. In this study, we report DBM1285 as a novel inhibitor of TNF-α production and a therapeutic candidate for RA treatment. DBM1285 concentration-dependently inhibited lipopolysaccharide (LPS)-induced TNF-α secretion in various cells of macrophage/monocyte lineage, including bone marrow macrophages, THP-1 cells and RAW 264.7 cells. However, LPS-induced mRNA expression of TNF-α was not affected by DBM1285 in these cells. Further studies demonstrated that the inhibitory effect of DBM1285 on TNF-α production might be mediated by post-transcriptional regulation through the modulation of the p38 mitogen activated protein kinase (MAPK)/MAPKAP kinase 2 (MK2) signaling pathway. We also confirmed that DBM1285 directly inhibits p38 MAPK enzymatic activity. In vivo administration of DBM1285 inhibited LPS-induced increase in serum level of TNF-α in mice. DBM1285 also suppressed progression of RA in an adjuvant-induced arthritis model. These results suggest that DBM1285 might be a promising therapeutic candidate for the treatment of RA.
Regulation of hematopoietic stem cell quiescence by TXNIP (36.1)
Authors: Choi Inpyo , Jeong Mira , Yoon Suk Ran 
Published: January 2023

Abstract
Hematopoietic stem cells (HSCs) are distinct cells that are predominantly located in the bone marrow microenvironment, which is referred to as the niche. HSC quiescence, activation, and mobilization are coordinately regulated in their niche by intrinsic and extrinsic mechanisms. Several genes, including those that encode transcriptional regulators, cell cycle regulatory proteins, tumor suppressors, and proto-oncogenes, intrinsically regulate the balance between HSC quiescence and activation. The vitamin D3 up-regulated protein 1 (VDUP1) is a 397 amino acid residue, 50-kDa protein that belongs to the arrestin family. We hypothesized that VDUP1 plays a pivotal role in controlling HSC cell cycle, migration, and BM niche interactions. The expression of VDUP1 is decreased during HSC activation. In VDUP1-/- mice, the LT-HSC population is decreased and exhausted, and its capacity to repopulate is rapidly lost. The VDUP1 deficiency reduced the osteopontin/integrin β1-mediated interaction between HSCs and the bone marrow niche and impaired homing and retention in the osteoblastic niche, resulting in mobilized HSCs. Thus, we propose that VDUP1 is essential for maintaining HSC quiescence and interactions with the BM niche.
Analysis of human immunoglobulin gene repertoire in NOD/scid IL2Rγ null mice engrafted with human cord blood derived hematopoietic stem cells (36.14)
Authors: Chang Hong , Biswas Subhabrata , Tallarico Aimee , Sarkis Phuong , Marasco Wayne 
Published: January 2023

Abstract
The humanized NOD/scid IL2rg-/- (hNSG) and similar mouse models hold promise as testing platforms for preclinical studies. Here, we analyzed the human immunoglobulin (IG) variable heavy and light chain gene repertoire from sorted single B-cells of hNSG mice engrafted with cord blood derived hematopoietic stem cells. Unique IGHV (339) and IGLV (364, kappa and lambda) gene segments were derived from major B cell subsets identified in a largely normal B cell development pathway. However, the hNSG mice expressed elevated CD5+ mature B-1-like cells in the periphery. In mice and humans, B-1 cells are the major source of autoantibodies. Although the hNSG mice expressed a highly diverse repertoire comparable to humans, detailed genetic analysis of IG gene segments from the mature B cell subset revealed the presence of long IGH-CDR3 regions with positive charges—a hallmark for auto/polyreactivity. For functional tests, several in vitro expressed scFv-Fc IG candidates, derived from cloned heavy and light chain cognate pairs of single B cells, tested positive in an anti-nuclear antibody assay for auto/polyreactivity and/or bound to HIVgp120 in ELISA assays. The natural induction during HIV-1 infection of broadly neutralizing antibodies (BnAbs) which display autoantibody-like characteristics may be curtailed by the checkpoint control mechanisms during B cell ontogeny. We propose the hNSG mouse can be a model platform to test this hypothesis and interrogate how BnAbs are formed.