bluuebunny/crossref_metadata_2025_split huggingface dataset:
The regulatory function of lpr TCRαβ+ CD4-CD8- double negative T cells requires autocrine interferon gamma (168.24)
Authors: Juvet Stephen , Han Mei , Kim Edward , Vanama Ramesh , Joe Betty , Jeon Caroline , Zhao Fei , Amash Alaa , Adeyi Oyedele , Zhang Li 
Published: January 2023

Abstract
Mice and humans with Fas mutations develop lymphoproliferative (lpr) disease and accumulate a large number of lymphocytes including TCRαβ+ CD4-CD8- non-NK double negative (DN) T cells. In these settings, CD4+ T cells and B cells are implicated in causing lupus-like autoimmunity; in contrast, the function of the DN T cells is largely unknown. DN T cells in other situations have been shown to secrete interferon gamma (IFNγ) and possess FasL-dependent regulatory functions. Therefore, we sought to determine whether lpr DN T cells can exert regulatory effects on syngeneic T cells and to what extent these phenomena might depend on IFNγ secretion. We found that lpr DN T cells produce IFNγ after antigen stimulation and can suppress and kill syngeneic Fas+ CD4+ T cells in responding to alloantigens. Lpr DN T cells deficient in either IFNγ or its receptor exhibited reduced suppressive and cytotoxic function in vitro and a decreased ability to suppress CD4+ T cells in vivo during graft-versus-host disease (GVHD). FasL-deficient DN T cells from gld mice also failed to suppress GVHD. These findings demonstrate that lpr DN T cells are able to suppress Fas+ syngeneic CD4+ T cells in vitro and in vivo. Expression of both FasL and the IFNγ receptor on lpr DN T cells is critical to their regulatory function. These studies have revealed a previously unrecognized IFNγ-IFNγ receptor autocrine loop that is central to DN T cell mediated immune suppression.
Differential gene expression and suppressive properties of ex vivo-generated vs. naturally-occurring regulatory T-cells (168.25)
Authors: Karlsson Fridrik , Gray Laura , Zhang Songlin , Grisham Matthew 
Published: January 2023

Abstract
We have recently reported a novel ex vivo method to convert large numbers of CD4+Foxp3- T-cells to induced regulatory T-cells (iTregs; CD4+Foxp3+) via their incubation with plate-bound CD3 mAb, IL-2, TGFβ and retinoic acid (RA) for 4 days. These iTregs possess enhanced immune-suppressive activity in vitro and in vivo when compared to naturally-occurring Tregs (nTregs; CD4+Foxp3+). The objectives of this study were to: a) quantify and compare mRNA transcripts for genes known to be associated with Treg function using flow-purified iTregs and nTregs obtained from Foxp3GFP reporter mice and b) evaluate the stability of iTreg-mediated suppression in vitro and in vivo. We found 8 genes associated with iTregs that were significantly up-regulated when compared with nTregs with CCL4 (132-fold), Granzyme A (18-fold) and Runx1 (14-fold) showing the most dramatic increases. We also observed significant downregulation of 19, Treg-associated genes with IL-18 (600-fold), CD7 (300-fold), and IL-7 (100-fold) showing the most dramatic reductions in iTregs vs. nTregs. In addition, we found that when ex vivo-generated iTregs were “rested” by culturing iTregs for an additional 3 days in the absence of CD3 mAb, TGFβ and RA but in the presence of IL-2, Foxp3 expression, proliferative activity and in vitro suppressive activity were maintained. Unexpectedly, we found that these cells were more potent than nTregs in reversing preexisting colitis in a mouse model of chronic gut inflammation.
BCL6 in control of the Th2 inflammatory activity of regulatory T cells (168.3)
Authors: Sawant Deepali , Sehra Sarita , Kaplan Mark , Dent Alexander 
Published: January 2023

Abstract
The transcriptional repressor Bcl6 is a critical arbiter of T helper cell fate, promoting the follicular helper (Tfh) lineage while repressing other T helper cell lineages. Bcl6-deficient (Bcl6-/-) mice develop a spontaneous and severe Th2-type inflammatory disease early in life, thus warranting assessment of the role of Bcl6 in Treg cell function. Bcl6-/- Tregs displayed enhanced suppressor function in vitro and were competent at suppressing Th1-mediated colitogenic inflammation in vivo. In contrast, Bcl6-/- Treg cells were uniquely defective in their ability to control Th2-type airway inflammation, and in fact, actively exacerbated the disease. Bcl6 deficiency promoted the expression of Th2 cytokines by Tregs, in part due to increased transcriptional activity of Gata3, and also due to increased expression of microRNA-21 (miR-21). Ectopic expression of miR-21 in naïve T cells in vitro induced polarization towards Th2, while antagomir-mediated miR-21 repression was able to specifically suppress Th2 differentiation. Our results define novel roles for Bcl6 in regulating Treg gene expression and in dictating the ability of Treg cells to control Th2 inflammation. Thus, Bcl6 represents a crucial regulatory layer in the Treg functional program.
Column-free isolation of untouched CD4+CD127lowCD49d- human regulatory T cells in 45 minutes (168.4)
Authors: Kokaji Andy , MacDonald G. , Thomas Terry , Fairhurst Maureen 
Published: January 2023

Abstract
Regulatory T cells (Tregs) are a subset of lymphocytes that play a key role in maintaining immune homeostasis. The isolation of highly purified Tregs is essential for advancing this exciting field of research. FOXP3 remains the best marker for the identification of Tregs, but its intracellular localization currently precludes its use for the isolation of viable human Tregs. To overcome these technical difficulties, researchers have exploited the expression of other surface markers to isolate Treg populations. Typically, the expression of CD127 inversely correlates with FOXP3 expression, while CD49d, is expressed on the majority of pro-inflammatory effector cells but is absent on Tregs. Removal of CD127high cells from the CD4+ T-cell population enriches for Tregs with high levels of FOXP3 expression. Additional depletion of CD49d expressing cells further removes contaminating IFNγ and IL-17 secreting cells. Together this strategy permits selective isolation of highly enriched Tregs. We have developed a new kit for the negative enrichment of highly purified human Tregs. Our new EasySep™ CD4+CD127lowCD49d- kit isolates untouched Tregs by by column-free magnetic separation in only 45 minutes. Mean CD4+ T-cell purity is 94.3% +/- 5.5% of which 65.5% +/- 11.7% express high levels of FOXP3 (n=7). This new product will add to the array of powerful yet convenient tools available for the isolation of functional highly purified human Tregs.
Unique function of B cells in immune regulation through CD86-mediated maintenance of Foxp3 (168.6)
Authors: Ray Avijit , Basu Sreemanti , Dittel Bonnie 
Published: January 2023

Abstract
In recent years, B cells have emerged as an important player in the immune regulatory network. Using the animal model of multiple sclerosis, experimental autoimmune encephalomyelitis (EAE), we observed a lack of resolution of disease in B cell-deficient (μMT) mice. Whereas, their WT counterparts recovered, indicating a regulatory role for B cells in central nervous system (CNS) autoimmunity. CD4+Foxp3+ T regulatory cells (Tregs) also mediate resolution of EAE. Absence of CD80/CD86 (B7) on B cells correlated with late emergence of Tregs in the CNS during EAE, suggesting an interaction between B cells and Tregs through B7 molecules. Anti-CD20 mediated B cell depletion disrupted this interaction resulting in significant reduction in the number of Tregs and like μMT, manifested in unresolved EAE. Interestingly, in vitro studies identified a unique role for B cells in maintaining Treg Foxp3 expression, which was enhanced upon CD86 upregulation following anti-IgM activation. Adoptive transfer of B cells into μMT mice resulted in partial restoration of Treg numbers leading to resolution of EAE. When deficient in CD86, B cells had a significantly reduced ability to restore Treg numbers in μMT. These data identify a distinct role for B cells in immune regulation through CD86-mediated interaction with Tregs and maintenance of Foxp3. This novel immune regulatory property of B cells may be harnessed to develop therapies for autoimmune diseases.
Evidence for NK cell subset licensing in anti-viral and anti-leukemic responses after syngeneic hematopoietic stem cell transplantation in mice (169.10)
Authors: Sungur Can , Alvarez Maite , Tang-Feldman Yajarayma , Lanier Lewis , Pomeroy Claire , Murphy William 
Published: January 2023

Abstract
Natural killer (NK) cells are an initial defense against cancers and viruses and can be classified into subsets based on their expression of receptors. These subsets are considered “licensed” if inhibitory receptors are capable of binding host MHC I and “unlicensed” if not. Since in vivo data on licensing and subsets are limited, our studies aim to show licensing in tumor and viral models as well as elucidate the roles of NK subsets in a post-hematopoietic stem cell transplantation (HSCT) setting. We have recently observed that cells bearing the Ly49G2 inhibitory receptor predominated post-HSCT regardless of MHC I haplotype. Additionally, studies in resting mice indicated that the unlicensed subset is the major mediator of murine cytomegalovirus (MCMV) protection. Surprisingly, we have found that depleting unlicensed, Ly49G2+, NK cells from leukemia (C1498) bearing, H2b strain mice after syngeneic HSCT significantly improved survival of mice, while depleting the licensed subset, Ly49C/I+, decreased survival. Furthermore, depletion of the Ly49C/I+ subset resulted in significantly increased viral loads in the liver, spleen, and salivary glands in H2b strain mice infected with MCMV 14 days post-HSCT. Conversely, depletion of the unlicensed ly49G2+ subset resulted in only a minor increase in viral load. Collectively, these data support the importance of NK licensing in tumor and viral models post-HSCT and suggest a regulatory role for the unlicensed subsets.
Graft versus host disease is associated with overproduction of TNFα and not with interleukin 17 (169.14)
Authors: Moreno-Lafont Martha , Gutiérrez-Hoya Adriana , Montiel-Cervantes Laura , Vela-Ojeda Jorge , Rodríguez-Cortés Octavio , López-Santiago Rubén 
Published: January 2023

Abstract
Allogeneic transplantation of hematopoietic cells (HCT) is a therapy used to treat a number of malignant and nonmalignant diseases, including acute and chronic leukemia, myelomas, lymphomas, aplastic anemia, solid tumors and severe immunodeficiencies. Unfortunately, a main complication of this treatment is the development of graft versus host disease (GVHD). GVHD is characterized by events caused by the reaction of immunocompetent cells from the graft against host tissues, developing an inflammatory reaction. In this work we studied 6 patients undergoing HCT, and one patient undergoing syngeneic transplantation of hematopoietic stem cells; 6 healthy donors were included as controls. In contrast to studies in mice showing that proinflammatory cytokine Th17 correlated with the occurrence and severity of GVHD, we did not observe overproduction of IL-17 in any patient that developed GVHD, in comparison to IL-17 production in controls and before transplantation. Furthermore, patients whose lymphocytes did not respond to polyclonal activators were of poor prognosis, correlating with graft loss, infections by opportunistic microorganisms and relapse. Remarkably, patients with GVHD showed exacerbated TNFα production, which correlated with the occurrence and severity of GVHD. These data show the importance of the TNFα and its contribution to the widespread damage experienced by patients who develop GVHD, and the importance of the use of specific immunosuppression of TNFα.
Pretransplant conditioning is required for minor but not major mismatch graft-versus-host disease: a vital role for intestinal damage (169.15)
Authors: Nalle Sam , Joseph Nora , Savage Peter , Turner Jerrold 
Published: January 2023

Abstract
Graft-versus-host disease (GVHD) is a major complication of allogeneic bone marrow transplantation (BMT). Previous studies suggest that the magnitude of the pre-BMT conditioning influences the severity of GVHD. To test this, we developed minor mismatch (129→B6) and major mismatch (Balb/c→B6) models of GVHD. In lethally irradiated WT recipients, major mismatch caused severe GVHD that was fatal in all recipients by 24 days after BMT. Minor mismatch led to a milder GVHD, where 50% of recipients survived to 35 days after BMT. We examined the role of irradiation in these models by minor or major mismatch adoptive transfer (AT) into unconditioned immunodeficient recipients. Major mismatch AT alone resulted in prominent GVHD. In contrast, minor mismatch AT required irradiation conditioning to cause GVHD. Conditioning and minor mismatch AT lead to elevated IFNγ, TNF, and IL-6, and an increased ability of donor cells to kill allogeneic targets in vivo (63±7% vs. 26±6%) compared to AT alone. As the intestine is very sensitive to irradiation, it was possible intestinal damage from the irradiation conditioning was facilitating minor mismatch GVHD. When we induced transient colonic damage with dextran sodium sulfate prior to minor mismatch AT, we observed systemic target organ histopathology characteristic of GVHD. Thus, we conclude that pretransplant conditioning, particularly intestinal damage, is required for minor mismatch GVHD.
IL-17 expression is inversely correlated with luminal narrowing in transplant vasculopathy: potential involvement of eNOS induction in vascular endothelial cells (169.20)
Authors: Choy Jonathan , Enns Winnie , Lee Martin , Escalante Nichole , von Rossum Anna , McManus Bruce 
Published: January 2023

Abstract
The role of IL-17 in transplant vasculopathy (TV), which is a leading reflection of chronic allograft rejection, remains poorly defined. The pathogenesis of TV involves luminal narrowing of allograft arteries caused by immune-driven thickening of the intima as well as insufficient compensatory vasodilation or remodeling. The vasodilatory process involves reduced expression and/or activity of the nitric oxide-producing enzyme eNOS by the endothelium. We examined the expression of IL-17 by immunohistochemistry in formalin-fixed, paraffin-embedded human specimens of TV (n=8). IL-17 was expressed by infiltrating leukocytes in the intima of arteries with TV and the majority of IL-17-positive cells were T cells (67.2 ± 11.0% of IL-17A-positive cells were also CD3-positive). Unexpectedly, the number of IL-17-positive cells (when normalized to total T cells) was inversely correlated with the amount of luminal narrowing (Pearson’s r = -0.709, p&lt;0.05), but was not correlated with the intima/media ratio (r = 0.069, p = NS). Because our findings suggest that IL-17 may be involved in compensatory vasodilation, the effect of IL-17 on eNOS expression was examined. IL-17 dose-dependently increased eNOS protein expression in cultured endothelial cells, as determined by Western blotting. These findings suggest that IL-17 may limit luminal narrowing in arteries with TV by contributing to vasodilation through induction of eNOS expression in the endothelium.
Acute rejection results in a unique signature of microRNAs (169.22)
Authors: Wei Liang , Wang Mouer , Harris Aleishia , Pham Betty , Martinez Olivia , Krams Sheri 
Published: January 2023

Abstract
MicroRNAs (miRNAs) are small non-coding RNA molecules that regulate the post-transcriptional expression of genes. Little is known about the roles of specific miRNAs in alloimmune responses. Groups of syngeneic and allogeneic recipients of heart transplants were sacrificed on day 7 post-transplant and heart grafts and spleens obtained. Graft infiltrating lymphocytes (GIL) and peripheral blood lymphocytes (PBL) were isolated on post-transplant day 5. MicroRNAS were profiled using TaqMan microfluidic cards. Forty-two miRNAs were increased and 32 were decreased (p&lt;0.05) in allogeneic heart grafts as compared to syngeneic. To determine if these miRNAs were expressed by lymphocytes or heart tissue, we profiled the GIL and determined that 22 miRNAs were increased and 52 decreased. There was less differential expression seen in spleen and PBL with 11 and 9 miRNAs changed respectively. Seventeen miRNAs were selected for validation by qPCR. The miRNAs, miR-7a and miR-182 were significantly increased in the heart graft, GIL, and PBL indicating that acute rejection results in specific profile of miRNAs in both the graft and the periphery. Further studies analyzing the targets of the miRNAs associated with graft rejection may lead to novel therapeutics and diagnostics.
Heme-oxygenase-1 in host tissues prevents visceral GVHD by regulating donor T cell expansion. (169.24)
Authors: Varelias Antiopi , Kreijveld Ellen , Olver Stuart , Koyama Motoko , Robb Renee , Raffelt Neil , Wilson Yana , Kuns Rachel , Don Alistair , Markey Kate , Anderson Greg , Clouston Andrew , MacDonald Kelli , Hill Geoff 
Published: January 2023

Abstract
Heme oxygenase-1 (HO-1), a ubiquitously expressed enzyme that degrades heme, has anti-inflammatory, anti-apoptotic and anti-proliferative actions yet its role in alloreactivity is unclear. Previously we demonstrated that HO-1 mRNA levels were elevated in recipient tissues of IFNγR-/- animals resistant to acute GVHD of the GI tract. Here we demonstrate that HO-1 expression in host tissues is critical for the suppression of acute GVHD as B6.HO-1-/- recipients of allogeneic BALB/c.WT grafts developed severe acute GVHD with rapid mortality (0% survival at d10) while WT mice survived longer term (median survival 42d). This was T cell mediated since mice transplanted with T cell depleted grafts showed no evidence of acute GVHD. Histological analysis revealed B6.HO-1-/- recipients developed severe acute GVHD of the GI tract and liver. TNF and IFNγ were elevated in the sera of B6.HO-1-/- recipients and systemic IFNγ but not TNF neutralization prevented hyperacute gut GVHD. The transplantation of grafts containing luciferase+ donor T cells demonstrated increased bioluminence signals in the mesenteric lymph node, spleen and GI tract of HO-1-/- recipients. Furthermore, the absolute number of donor CD4+ and CD8+ T cells co-producing the pro-inflammatory cytokines TNF and IFNγ were increased early after BMT in the lymph nodes of B6.HO-1-/- recipients, demonstrating that HO-1 within recipient tissue controls the acquisition and expansion of donor T cell effector function after BMT.
SRL-derived Tregs blocking alloresponses, but its generation is impaired by inflammatory cytokines (169.29)
Authors: Xu Luting , La Monica Giovanna , Leventhal Joseph , Gallon Lorenzo 
Published: January 2023

Abstract
Subpopulations of T cells play important roles in promoting transplant rejection or in maintaining transplantation tolerance. Little is known about the impact of immunosuppressive drugs calcineurin inhibitor Tacrolimus (Tac) and mTOR inhibitor Sirolimus (SRL) on the generation/expansion and functions of allospecific T cell subpopulations. Alloreactive CD4 T cells generated with naive T cells and allogeneic APCs were re-stimulated with autologous APCs and anti-CD3 in the presence of SRL/Tac and cytokines. Tac significantly blocked the productions of IFNgamma (Th1) and IL-17 (Th17) while SRL had minimal effect. FOXP3 expression (Treg) was markedly increased in SRL compared to Tac. These also confirmed at transcriptional levels by determining the T-bet, RORgt, and FOXP3 mRNA expressions. When used together, Tac at 2-5ng/ml with SRL at 2.5-10ng/ml achieved the maximal effect in inhibiting the productions of IFNgamma and IL-17 while maintaining a high level of FOXP3 expression. SRL-derived Tregs expressed normal Treg surface markers, were anergic to allostimulation, and functioned normally and specifically suppressed proliferation of allogenic effector T cells and Th1 and Th17 responses. When TNFalpha, IL-6, IL-1beta were added with SRL, there was a markedly decrease of SRL-induced Tregs. SRL significantly decreased DNA methyltransferase1, and thus increased stability of FOXP3 expression, which in part accounted for increased Treg conversion in alloreactive CD4 T cells.
Regulatory T cell therapy combined with donor-specific T cell deletion promotes long-term survival of islet allografts (169.32)
Authors: Lee Karim , Nguyen Vinh , Miao Gang , Scholz Hanne , Brennan Todd , Kang Sang-Mo , Tang Qizhi 
Published: January 2023

Abstract
Although Tregs are vital for peripheral tolerance, adoptive transfer of Tregs is insufficient in inducing long-term allograft survival in lymphoreplete hosts without preconditioning or adjunct immunosuppression. We hypothesize that reducing donor-reactive T cells may create a therapeutic window for Tregs to induce allograft tolerance without additional immunosuppression. We preconditioned recipient mice using donor-specific transfusion followed by cyclophosphamide treatment to selectively kill proliferating donor-reactive cells. This approach deleted 70-90% donor-reactive T cells, but failed to prolong islet allograft survival. Addition of Tregs with direct donor reactivity prior to transplantation led to long-term survival of BALB/c islets in 70% of C57BL/6 recipients. Equal numbers of polyclonal Tregs significantly prolonged graft survival, but 90% of the recipients rejected their grafts by 100 days. Increasing the numbers of polyclonal Tregs was able to confer long-term graft protection. Similar strategy of combining donor-reactive T cell deletion and polyclonal Tregs did not protect C3H islets in autoimmune NOD recipients. Further addition of Tregs specific for islet autoantigen was required to prolong graft survival in NOD mice. Together, these results demonstrate a clinically viable approach to harness the tolerogenic property of Tregs for inducing long-term allograft acceptance without additional immunosuppression.
Inhibition of Activated Protein C Attenuated Experimental Autoimmune Encephalomyelitis (171.13)
Authors: Alabanza Leah , Bynoe Margaret 
Published: January 2023

Abstract
Activated Protein C (APC) is an anticoagulant that is involved in the complex two-way interactions between the coagulation and immune systems. APC is widely known to be anti-inflammatory; most notably it has been clinically developed for treatment of severe sepsis. The anti-inflammatory properties of APC include conferring endothelial barrier protection, thus limiting the extravasation of immune cells into tissues. We investigated whether APC can affect the progression of experimental autoimmune encephalomyelitis (EAE), the mouse model for multiple sclerosis. The major pathological component of EAE is blood brain barrier (BBB) dysfunction resulting in the extravasation of autoreactive immune cells into the central nervous system (CNS). We observed that inhibition of endogenous APC during EAE resulted in considerable immune cell infiltration in the brain. Interestingly, however, these mice exhibited mild to no outward signs of EAE as assessed by paralysis severity. The anti-APC treated mice also had significantly higher percentage of CD4+Foxp3+ cells both in the CNS and in the periphery compared to controls. Moreover, the frequency of cells characterized as myeloid derived suppressor cells are increased in anti-APC treated EAE mice. Our study shows that lowered APC levels during EAE can lead to divergent effects on EAE pathogenesis, specifically leading to increased BBB dysfunction while simultaneously inducing immunological conditions that are non-pathogenic in EAE.