bluuebunny/crossref_metadata_2025_split huggingface dataset:
T cell receptor knockout efficiency utilizing an engineered meganuclease is influenced by stimulation conditions.
Authors: Brown Audrey E , MacLeod Daniel T. , Martin Aaron J. , Wetzel Keith J. , Pham Christina D. , Antony Jeyaraj , McCreedy Bruce , Bartsevich Victor V. , Nicholson Michael G. , Smith Jeff , Jantz Derek 
Published: January 2023

Abstract
Recent reports describing allogeneic chimeric antigen receptor (CAR) T cell therapies have generated tremendous excitement over the possibility of a universal CAR T platform. We are utilizing a highly engineered meganuclease designed to target the T cell receptor (TCR) alpha chain to eliminate expression of the TCR, preventing graft versus host disease following adoptive transfer. Meganuclease cleavage efficiency is highly dependent on stimulation of T cells prior to delivery of the meganuclease. To determine optimal activation conditions for TCR knockout and cell yield, T cells were activated with Dynabeads® Human T cell Activator CD3/CD28, Immunocult™ CD3/CD28 and CD3/CD28/CD2 T Cell Activators, and MACS® GMP TransAct CD3/CD28 reagent prior to nucleofection. T cells were stained prior to and following nucleofection to identify cell activation markers and cytokine receptors (CD69, IL2R, IL7R, IL15R, IL21R). Knockout efficiency was determined by staining for CD3 expression. Our results indicate that Dynabead-stimulated T cells expressed the highest levels of activation markers at 18 hours post-stimulation, and that Dynabeads® and Immunocult™ CD3/CD28/CD2 reagents both promote robust T cell proliferation and support the highest TCR knockout efficiency following our protocol. Furthermore, expression of cytokine receptors varies both during stimulation and post-nucleofection, suggesting an optimal cocktail of cytokines may promote T cell proliferation. Future studies will focus on further optimization of T cell stimulation conditions that may promote TCR knockout and expansion of T cells post-nucleofection.
Development and characterization of Th17-related U-PLEX® assays
Authors: Wallowitz Mikhael L , Ning Qian , Shrestha Arun , Shelburne Chris , Liu Pu , Stewart David , Wolfert Robert , Oberoi Pankaj , Wohlstadter Jacob N 
Published: January 2023

Abstract
Th17-related cytokines mediate a host’s defense mechanisms against various infections and play a crucial role in crosstalk between the immune system and affected tissues. Here we describe the development and characterization of multiplexed Th17-related immunoassays on MSD’s flexible U-PLEX platform.
Monoclonal and polyclonal antibodies were conjugated with biotin and/or SULFO-TAG™ and screened as capture and/or detection reagents. A number of analytical parameters were used to select antibody pairs, and assays were developed by optimizing antibody concentrations, calibrator curves, specificity, matrix tolerance, and assay robustness. Assay performance was verified to be compatible with other U-PLEX assays by running Th17-related multiplexes on the U-PLEX platform with controls and samples.
Calibration curves showed expected signals, sensitivity, precision, and accuracy. Control samples for the assays had CVs &lt; 10% within runs and &lt; 25% between runs. Sensitivities were &lt; 1 pg/mL for the majority of the assays. Dilution linearity and spike recovery studies demonstrated acceptable matrix tolerance and accurate quantification for most of the assays tested across all matrices (typically between 75% – 125%). Cross-reactivity between assays was shown to be typically &lt; 0.5%. Results demonstrated a strong correlation between samples measured on U-PLEX multiplex and streptavidin singleplex panels with r2 values &gt; 0.95 and slopes between 0.8–1.2.
Over 20 Th17-related assays for human and mouse were developed for the U-PLEX platform. These assays enable the measurement of multiple proteins on biological matrices that are relevant to a wide range of life science applications and pre-clinical or clinical studies.
Target specific siRNA-conjugate therapy for experimental autoimmune myasthenia gravis
Authors: Huda Ruksana 
Published: January 2023

Abstract
Myasthenia gravis (MG) is an autoimmune disease of muscle weakness caused by Acetylcholine receptor- autoantibody and complement mediated damage of neuro-muscular junction (NMJ). We have recently shown a greater advantage of using siRNA over other conventional therapeutic tools in an experimental mouse model of MG (EAMG). However, the major drawbacks of siRNA therapy is reduced efficacy due to the lack of target restricted delivery, where it is necessary to suppress pathogenic response arising predominantly from a particular cell/tissue type. In this study, we used a novel strategy of cell specific new formulation of siRNA or siRNA conjugate delivery to pathogenic cells in EAMG. Our preliminary work shows successful preparation of siRNA conjugate and its functional efficacy over siRNA only treatment in an in vitro rodent cell line. Successful completion of the study may provide a solution for the siRNA or monoclonal antibody therapy hurdle and a new direction to ultimately explore a highly effective and safe therapy for patients with MG.
Anti-MAdCAM-1 therapy does not affect immune surveillance in the central nervous system
Authors: Stuve Olaf , D'Haens Geert R.A.M. , Reinisch Walter , Vermeire Severine , Vogelsang Harald , Allez Matthieu , Desreumaux Pierre , Van Gossum Andre , Baumgart Daniel , Hassan-Zahraee Mina , Cheng John , Yanhua Zhang , Sandborn William , Kaminski Annamarie , Ahmad Alaa , Pradhan Vivek , Cataldi Fabio , Clare Robert , Gorelick Ken 
Published: January 2023

Abstract

Background
Blocking integrins can be associated with an increased risk of progressive multifocal encephalopathy (PML). MAdCAM-1 is an adhesion molecule that is not constitutively expressed in healthy CNS and considered mostly gut-selective. However, MAdCAM-1 is upregulated in choroid plexus epithelium during experimental autoimmune encephalomyelitis (EAE. PF-00547659 is a fully human mAb that is highly selective for MAdCAM-1. The purpose of this study was to investigate its effect on cellular elements of immune surveillance in the CNS and blood of patients with Crohn’s disease (CD).

Methods
All methods were tested in a control cohort of volunteers with CD (Cohort 1). Study patients (Cohort 2) had CD and prior treatment with anti-TNF and immunosuppressant therapies. Patients underwent a lumbar puncture (LP1) followed by 3 monthly injections of 225 mg PF-00547659. After the last dose, a LP2 was performed. CSF was analyzed within 24h by multi-parameter flow cytometry.

Results
In Cohort 1, the lymphocyte subset percentages were the same at both time points. Cohort 2 consisted of 30 patients in whom 2 LPs were performed. Their pre-treatment lymphocyte numbers were lower than those of subjects who had stopped immunosuppressants, but otherwise results were the same. CSF lymphocytes (cells per mL) shown as geometric means (CV%). LP1: 37 patients, total lymphocytes 391 (140%), CD3+/CD4+ cells 246 (152%), CD3+/CD8+ cells 100 (126%), CD4:CD8 ratio 2.46 (57%). LP2: 30 patients, total lymphocytes 513 (137%), CD3+/CD4+ cells 332 (143%), CD3+/CD8+ cells 123 (125%), CD4:CD8 ratio 2.69 (56%).

Conclusion
This is the first evidence that a therapeutic dose of anti-MAdCAM-1 mAb does not affect the number and composition of leukocytes in the CNS.
A predictive disease-specific protein biomarker fingerprint for clinical relapses and glucocorticoids treatment of multiple sclerosis
Authors: Raphael Itay , Chase Carol , Gomez-Rivera Francisco , Forsthuber Thomas G 
Published: January 2023

Abstract
Despite extensive research, multiple sclerosis (MS) remains a disease that lacks a definitive prognostic test to predict imminent disease relapses. Thus, patients may undergo years of unnecessary treatments. Additionally, current treatments for MS vary significantly in efficacy between individual patients, and thus there is a critical need to develop biomarkers for treatment efficacy and resistance. To address these issues, we recently developed a high-throughput quantitative proteomics method to measure changes in proteome expression over the course of the preclinical experimental autoimmune encephalomyelitis (EAE) model. Interestingly, using the EAE model we revealed characteristic CNS-specific protein expression waves prior to the onset of clinical symptoms. Moreover, we have identified key proteins with altered expression that correlated with the therapeutic efficacy of glucocorticoid treatment. Bioinformatics analysis revealed candidate protein biomarkers to predict treatment efficacy and clinical disease course. Importantly, these proteins could be detected in serum and expression trajectories analysis identified a strong correlation of the CNS proteome to their levels in serum. Prospective studies in the EAE model using these candidate protein biomarkers showed their effectiveness in predicting clinical disease and treatment responses. Our studies suggest the utility for establishing homologous protein biomarkers in human MS patients. Finally, our work investigating the CNS proteome over the course of EAE may provide novel insights and molecular targets for disease mechanisms and treatments of MS.
Identifying biomarkers for monitoring progression of multiple sclerosis
Authors: Chase Carol Ann , Raphael Itay , Forsthuber Thomas 
Published: January 2023

Abstract
There are currently no reliable methods for assessing the progression of multiple sclerosis (MS) from the relapsing-remitting to the secondary progressive form. This gap in knowledge hinders the ability for therapeutic intervention and ultimately results in continued relapses and physiological deterioration. To begin to address the urgent need for biomarkers of progressive MS we investigated proteome changes over the disease course of progressive experimental autoimmune encephalomyelitis (EAE) in NOD mice as a preclinical model of the disease. Our lab has pioneered a novel high-throughput quantitative proteomic technique which we used to quantify expression levels of central nervous system (CNS) proteins over the course of monophasic EAE in C57.BL/6 mice. We utilized bioinformatics tools to prioritize key proteins whose expression level correlated specifically with the progressive phase of disease in the NOD EAE model. Importantly, we were able to detect corollary changes in these CNS-specific proteins in the serum, pointing to a minimally invasive means of monitoring disease progress and measuring drug efficacy. Our studies will provide a proof-of-concept for identifying homologous human biomarkers to guide treatment in individual patients. Furthermore, our results may provide insights into mechanisms that contribute to disease pathology and offer additional therapeutic targets for slowing the progression of MS.
Metabolic Reprograming of Alloantigen-Activated T Cells after Hematopoietic Cell Transplantation
Authors: Yu Xue-Zhong , Mehrotra Shikhar , Chi Hongbo , Nguyen Hung D. 
Published: January 2023

Abstract
T cells are the driving force in the induction of graft-versus-host disease (GVHD); yet little is known about T-cell metabolism in response to alloantigens after hematopoietic cell transplantation (HCT). The current study demonstrates that donor T cells undergo reprograming after allogeneic HCT by switching from fatty acid β-oxidation (FAO) and pyruvate oxidation via the TCA cycle to aerobic glycolysis, thereby increasing dependence upon glutaminolysis and the pentose phosphate pathway. Glycolysis is required for optimal function of alloantigen activated T-cells and induction of GVHD, as inhibition of glycolysis by targeting mTORC1 or pfkfb3 ameliorates GVHD mortality and morbidity. Thus, donor T cells use glycolysis as the predominant metabolic process after allogeneic HCT, which can be a valid therapeutic target for the control of GVHD.
Transient depletion of donor CD4+ T cells early after HCT allows recipient tissue PD-L1 to tolerize infiltrating CD8+ T cells and to prevent acute GVHD while preserving GVL effect
Authors: song qingxiao , Ni Xiong , Cassady Kaniel , Deng Ruishu , Jin Hua , Zhang Mingfeng , Forman Stephen , Martin Paul , Wang Jianmin , Zeng Defu 
Published: January 2023

Abstract
Prevention of graft-versus-host-disease (GVHD) while preserving graft-versus-leukemia/lymphoma (GVL) effect remains the Holy Grail for allogeneic hematopoietic cell transplantation (HCT). Purified donor CD4+ T cells induce strong but CD8+ T cells induce little acute GVHD (aGVHD); however, the mechanisms remain unknown. Recipient tissue expression of PD-L1 (B7H1) down-regulates aGVHD via simultaneous PD-L1 interactions with CD80 and PD-1 on donor T cells, but whether this mechanism impacts on GVL effect remains unclear. In the current studies, with MHC-mismatched HCT model of C57BL/6 donor to BALB/c recipient, we observed that 1) although sorted wild-type (WT) CD8+T cells induced no signs of aGVHD, purified PD-1−/− CD8+ T cells induced lethal aGVHD; 2) while WTCD8+ T or very low-dose (0.1×106) CD4+ T cells alone induced little signs of aGVHD, co-transplantation of them induced lethal aGVHD; 3) depletion of CD4+ T cells by anti-CD4 mAb early after HCT prevented aGVHD in wild-type but not in PD-L1−/− recipients, and the GVHD prevention is associated with increase of serum IFN-γ, upregulation of host tissue PD-L1, upregulation of CD80 and PD-1 by CD8+ T cells, as well as increase of apoptosis and anergy of tissue infiltrating CD8+ T cells; 4) interestingly, the depletion of donor CD4+ T cells augmented donor CD8+T cell proliferation without increasing anergy or apoptosis in the lymphoid tissue and preserved strong GVL effect. These results indicate that depletion of donor CD4+ T cells early after HCT leads to increasing susceptibility of donor CD8+ T cells to host tissue PD-L1-mediated apoptosis and anergy in GVHD target tissues but not in the lymphoid tissues, such that GVHD is prevented and GVL effect is preserved.
Inhibition of Endogenous Memory CD8 T Cell Infiltration into Cardiac Allografts Subjected to Prolonged Cold Ischemic Storage is Required to Promote Long-Term Graft Survival Naoki Kohei, Hidetoshi
Authors: Fairchild Robert L , Kohei Naoki , Tsuda Hidetoshi , Tanaka Toshiaki , Iida Shoichi , Abe Toyofumi , Kish Danielle , Ayasoufi Kathy , Valujskikh Anna , Fairchild Robert L 
Published: January 2023

Abstract
Endogenous memory CD8 T cells infiltrate MHC-mismatched cardiac allografts and produce IFN-γ in response to donor class I MHC within 24 hrs after graft reperfusion in mice. We have reported that prolonged cold ischemic graft storage (CIS) provokes intense inflammation within hours after allograft reperfusion and promotes CTLA-4Ig-resistant endogenous memory CD8 T cell rejection of the allograft by day 28 post-transplant (whereas CTLA-4Ig prolongs survival of allografts subjected to minimal CIS &gt; day 60). We tested strategies inhibiting endogenous memory CD8 T cell infiltration into cardiac allografts subjected to prolonged CIS to promote long-term allograft survival. Peri-transplant anti-LFA-1 mAb and anti-CD154 mAb treatment of recipients of allografts subjected to minimal CIS prolonged survival of 60% of allograft &gt; 100 days. In contrast, this treatment prolonged only 20% of allografts subjected to prolonged CIS beyond day 80 post-transplant, with rejection accompanied by the appearance of high titers of donor-specific antibody (DSA &gt;10,000 vs. &lt;100 in long-term survivors). Use of a new regimen: peri-transplant recipient treatment with anti-LFA-1 mAb, anti-TNFa mAb, and anti-CD154 mAb plus additional doses of anti-CD154 mAb on days 14 and 16 post-transplant promoted long-term survival of 60% allografts subjected to prolonged CIS past day 100 post-transplant. Recipients with surviving allografts accepted donor skin allografts, but rejected third-party skin allografts. These studies indicate long-term survival of allografts subjected to prolonged CIS by using strategies that include reagents inhibiting endogenous memory CD8 T cell infiltration into the allografts and reagents that inhibit the production of DSA.
Understanding the effects of an inflammatory milieu on the development of mesenchymal stem cells: implication for treating inflammatory diseases
Authors: Sherman Lauren S. , Ponzio Nicholas M , Rameshwar Pranela 
Published: January 2023

Abstract
Mesenchymal stem cells (MSCs), multipotent cells found in various adult tissues, are an attractive source of cells for cellular therapy and regenerative medicine. Reasons include their ease to expand, plasticity to generate cells of all germ layers, reduced ethical concerns, and ability to be available as ‘off the shelf’ cells for immediate use in transplantation. MSCs can respond differently to varying microenvironments to perform distinct immune functions. The microenvironment can also affect the developmental state of MSCs. Better understanding of how the microenvironment influences MSC multipotency is crucial for effective translational use of these cells in the clinic. This study tested the hypothesis that the changes in an inflammatory microenvironment will influence MSC function. To study these effects, an in vitro model of GvHD was developed by culturing MSCs in a three-dimensional model – rather than in a traditional monolayer – under physiological conditions. The GvHD model was generated based on a modified two-way mixed lymphocyte reaction. The cells were then assessed for phenotype, proliferation, and viability. Following growth in suspension, the MSCs continued to express an MSC phenotype. Proliferation and viability studies suggested that, although the cells were proliferating, they underwent cell death at a quicker rate than their two-dimensional culture counterparts. When cultured in the GvHD model, MSCs exhibited increased viability as compared to culture in the three-dimensional system alone. This in vitro GvHD model will elucidate factors within the inflammatory milieu that alter MSC multipotency. Identifying these factors will allow for more controlled and effective clinical use of MSCs.
Evaluation of T-Track® CMV to assess the functionality of Cytomegalovirus-reactive cell-mediated immunity in hemodialysis patients
Authors: Boeckl Katharina , Banas Bernhard , Böger Carsten , Lückhoff Gerhard , Krüger Bernd , Barabas Sascha , Starke Astrid , Bendfeldt Hanna , Schemmerer Mathias , Köstler Josef , Wagner Ralf , Deml Ludwig , Leicht Joachim , Krämer Bernhard K. 
Published: January 2023

Abstract

Background
Impairment of Cytomegalovirus (CMV)-specific cell-mediated immunity (CMI) by immunosuppressive therapy is a major cause for uncontrolled CMV replication and related clinical complications.

Objective
The aim of the study was to assess the suitability of the immune monitoring tool T-Track® CMV to determine CMV-reactive CMI in hemodialysis patients.

Study Design
Positive and negative agreement of T-Track® CMV with CMV-serology have been examined in a cohort of 124 hemodialysis patients of whom 67 (54%) revealed a positive CMV serostatus and compared with that of QuantiFERON®-CMV and a mixture of 6 CMV iTAG tetramers.

Results
Positive T-Track® CMV, QuantiFERON®-CMV and CMV tetramer results were obtained in 60/67, 45/62 and 40/52 CMV-seropositive hemodialysis patients resulting in 89.6%, 72.6% and 76.9% positive agreement with CMV-serology, respectively. Positive T-Track® CMV responses were dominated by pp65-reactive cells (58/67 (86.6%)), whereas IE-1 responsive cells were observed in 33/67 (49.3%) CMV-seropositive patients. T-Track® CMV, QuantiFERON®-CMV and CMV tetramers showed 78.9% (45/57), 87.3% (48/55) and 93.3% (42/45) negative agreement with serology respectively. Notably, T-Track® CMV-positive tests in seronegative patients were mostly caused by IE1-reactive cells at low spot counts.

Conclusion
T-Track® CMV represents a sensitive assay format, enabling functional assessment of the network of clinically relevant subpopulations of CMV-responsive cells. Thus, T-Track® CMV may represent a valuable tool to identify patients at increased risk for CMV-related clinical complications and may help to guide personalized antiviral and immunosuppressive therapy.
Defining the optimal plasmacytoid dendritic cell graft source for hematopoietic stem cell transplantation
Authors: Hassan Mojibade , Lin Kaifeng , Li Jingxia , Harris Wayne , Waller Edmund K 
Published: January 2023

Abstract

Purpose
Allogeneic hematopoietic stem cell transplantation (HSCT) can cure patients with blood disorders and malignancies. Stem cell source can be bone marrow (BM) or peripheral blood mobilized by granulocyte-colony stimulating factor (G-CSF). Plasmacytoid dendritic cells (pDC) reduce the incidence and severity of GvHD. The BMT CTN 0201 study indicates overall survival, relapse, and the incidence of acute GvHD are similar in patients that receive BM versus G-CSF mobilized peripheral blood (G-PB) grafts, but increased incidence of chronic GvHD in patients who receive G-PB grafts. To explore the effect of graft source on the functional properties of pDC, we compared the effect of G-CSF mobilized pDC to BM pDC in the regulation of GvHD in a murine allogeneic HSCT model.

Methods
1) Expression of surface markers on BM and G-CSF treated pDC were assessed by flow cytometry. 2) We performed an HSCT from C57BL/6 mice into B10.BR recipients. Groups include HSC only, HSC + T cells, and HSC + T cells + BM pDC or G-CSF mobilized pDC. Hematopoietic chimerism, GvHD, and overall survival were assessed.

Results
1) BM pDC express CCR9 (marker for tolerogenic pDC) and CD62L at a higher frequency, suggesting enhanced homing to the gut, thymus, and lymph nodes. G-CSF mobilized pDC express maturational markers MHC II and CD86 at a higher frequency, suggesting effectiveness in activating T cells. 2) Mice that received transplants consisting of only HSC rejected the transplant. Mice that received transplants with T cells had greater than 90% donor hematopoietic cells. GvHD more than 40 days post transplant was highest in mice that received G-CSF mobilized pDC. Survival without pDC was under 60%. Groups that received pDC had an overall survival of greater than 80%.
IDH Mutant Gliomas Escape Natural Killer Cell Immune Surveillance by Downregulation of NKG2D Ligand Expression
Authors: Rao Aparna , Zhang Xiaoran , Deibert Christopher , Sette Paola , Grandi Paola , Amankulor Nduka 
Published: January 2023

Abstract

Background
Diffuse gliomas are fatal primary brain tumors that are poorly immunogenic. The basis for insufficient anti-tumor immunity in diffuse gliomas is not understood. Mutations in isocitrate dehydrogenases (IDH1 and IDH2) promote diffuse glioma formation through epigenetic reprogramming of a number of genes, including immune-related genes. Here, we identify epigenetic dysregulation of natural killer (NK) cell ligand genes as significant contributors to immune escape in glioma.

Methods
We analyzed the TCGA database for immune gene expression patterns in IDH mutant or wild-type gliomas and identified differentially expressed immune genes. NKG2D ligands expression levels and NK cell-mediated lysis were measured in patient-derived glioma stem cells and in genetically engineered astrocytes. Finally, we assessed the impact of hypomethylating agent Decitabine as a potential NK cell sensitizing agent in IDH mutant cells.

Results
All IDH mutant glioma stem-like cell lines exhibited significantly lower expression of NKG2D ligands compared with IDH wild-type cells. Consistent with these findings, IDH mutant glioma cells and astrocytes were resistant to NK cell-mediated lysis. The hypomethylating agent Decitabine increased NKG2D ligand expression, and restored NK- mediated lysis of IDH mutant cells in an NKG2D-dependent manner.

Conclusions
IDH mutant glioma cells acquire resistance to NK cells through epigenetic silencing of NKG2D ligands ULBP1 and ULPB3. Decitabine-mediated hypomethylation restores ULBP1 and ULBP3 expression in IDH mutant glioma cells and may provide a clinically useful method to sensitize IDH mutant gliomas to NK cell-mediated immune surveillance in patients with IDH mutated diffuse gliomas.