bluuebunny/crossref_metadata_2025_split huggingface dataset:
CRISPR/Cas9 Gene Editing of MIR155HG in Primary Human T Cells to Prevent Acute Graft-Versus-Host Disease
Authors: Neidemire-Colley Lotus M , Khanal Shrijan , Braunreiter Kara M , Snyder Katiri Jenna , Gao Yandi , Kalyan Sonu , Kararoud Meisam Naeimi , Duszynsk Molly , Chi Mengna , Malik Punam , Choe Hannah , Garzon Ramiro , Ranganathan Parvathi 
Published: August 2023

Abstract

Introduction
Acute graft-versus-host disease (GVHD) mediated by donor alloreactive T cells is the leading cause of non-relapse mortality in patients post-allogeneic hematopoietic cell transplantation. Here we genetically engineered donor T cells by genomic deletion of microRNA-155 host gene (MIR155HG) to prevent GVHD.

Methods
Using CRISPR/Cas9 genome editing, we targeted exon 3 (Ex3) of MIR155HG. Genomic deletion was evaluated by genomic PCR and RNA expression of mature miR-155 by qRT-PCR. In xenogeneic acute GVHD experiments, NSG mice were irradiated and transplanted with miR-155 Ex3 edited or control T cells to assess functionality in vivo. Histological analysis of target tissues (skin and liver) was performed at end of study. Weekly flow cytometric evaluation of human donor T cells in peripheral blood was performed. MOLM-13 leukemic target cells were cultured with miR-155 Ex3 edited or control CD8+ effector T cells for 72 hours and cytotoxic function was assessed by CD107a degranulation, intracellular IFN-γ production and leukemic cell death by flow cytometry.

Results
Ex3 deletion was confirmed by genomic PCR resulting in downregulation of mature miR-155 expression. Recipients of Ex3 deleted human donor T cells display lower GVHD clinical scores, lower average histopathological scores and significantly improved survival compared to mice receiving control T cells. Ex3 T cells displayed potent CTL function comparable to control T cells resulting in leukemic cell death.

Conclusion
CRISPR/Cas9 deletion of miR-155 in donor T cells protects against development of lethal GVHD while maintaining beneficial anti-leukemic effect. Our studies present genomic editing of donor T cells as a novel strategy to prevent GVHD.

Funding for this project was provided through The Ohio State University Leukemia Research Program to PR and RG, NIH R01CA252469 to PR, and NIH T32CA090223 fellowship to KB.
Sentinel flaps correlate with pro-inflammatory and immunoregulatory mediators of human face transplantation rejection
Authors: Crisler William J , Win Thet Su , Barrera Victor , Ho-Sui Shannan , Clark Rachael A 
Published: August 2023

Abstract
Early diagnosis and treatment of immune rejection is critical for face transplants (vascularized composite allografts, VCA). VCA rejection is diagnosed by histologic evaluation of skin biopsies, yet repeated biopsies are a considerable aesthetic problem. Sentinel flaps, fasciocutaneous radial forearm flaps, have been used for remote-site rejection monitoring, but the concordance between tissues during rejection remains unclear. With access to the world’s largest cohort of VCA patients at a single center worldwide, we studied 14 paired skin biopsies from sentinel flaps (SF) and facial allografts (FA) during rejection and non-rejection. We previously identified 15 pro-inflammatory genes (including GZMB, HLA-DPB1, IFNG) and 15 immunoregulatory genes (including SOCS1, CTLA4, IDO1) upregulated in VCA rejection. SF and FA had an average correlation coefficient of 0.88 and 0.76 for pro- and anti-inflammatory gene sets, respectively. During rejection, there was no significant difference in expression of the 15 pro-inflammatory genes between tissue type (SF versus FA), and only one immunoregulatory gene (PDCD1) was significantly different. Across all samples, there were 64 differentially expressed genes (DEG) between rejection and non-rejection, 13 DEG between tissue types, and only 2 DEG when statistically accounting for differences in tissue type driven by rejection status (log2FC&gt;1, p&lt;0.05). High throughput sequencing (HTS) of the CDR3 of TCR-β genes from 4 matched pairs during rejection identified a mean of 9.5 expanded shared T cell clones, indicating that shared antigens drove rejection in both tissues. Together, these data indicate that SF could serve as effective remote-site monitoring for face transplant rejection.
NIH T32 Training Grant 2T32AR007098-47 Dept. of Dermatology, Harvard Medical School RT170025 (P.I. Rachael A. Clark) Department of Defense - congressionally Directed Medical Research Programs; "T cells and Rejection in Vascularized Composite Allografts).
Interleukin 18 (IL-18) Suppresses Thymus Regeneration
Authors: Granadier David , Cooper Kirsten , Acenas Dante , Kinsella Sinéad , Evandy Cindy , Iovino Lorenzo , deRoos Paul , Hernandez Vanessa , Shannon-Sevillano Steve , Kousa Anastasia , Dudakov Jarrod 
Published: August 2023

Abstract
The thymus is highly sensitive to acute injury, particularly to cytoreductive conditioning given pre-hematopoietic stem cell transplant (HCT). The thymus is capable of a remarkable degree of regeneration, however its reparative capacity and T cell productivity decline with age. This leaves HCT recipients vulnerable to relapse of malignancy and opportunistic infection – the leading causes of post-HCT mortality – during a prolonged period of lymphopenia. Better understanding the endogenous mechanisms by which thymus regeneration is regulated may inform therapeutic interventions to improve T cell reconstitution in these patients. Here, we identify the inflammatory cytokine interleukin 18 (IL-18) as a suppressant of thymic function and propose its targeting to improve immune reconstitution.
We posit that distinct forms of acute injury (i.e., irradiation, chemotherapy, infection and stress) lead to caspase-1 activation within the thymus and subsequently increased active IL-18. Our preliminary data suggests that IL-18 receptor (IL-18R) expressing natural killer (NK) cells are activated following acute damage in an IL-18 dependent fashion. Importantly, depletion of this NK1.1+ IL-18R+ population improves thymus regeneration, suggesting that IL-18 induces an inflammatory response in organ-resident NK cells capable of impairing thymus repair.
Our findings implicate IL-18 as a novel regulator of thymic regeneration and identify NK cells as likely mediators in its pathway. Furthermore, we suggest that IL-18 can be therapeutically targeted to improve T cell reconstitution following cytoreductive therapy and HCT.
Supported by grants from NIH (R01 HL145276, P01 AG052359)
Expanded T regulatory cells fail to control germinal center responses in the lung lymph nodes of pulmonary fibrosis patients
Authors: Velez Tania Estefania , Yoon Young me , Hollinger Maile K , Blaine Kelly M , Lee Cathryn T , Strek Mary E , Guzy Robert , Sperling Anne I 
Published: August 2023

Abstract
Idiopathic pulmonary fibrosis (IPF) is the most common form of interstitial lung diseases (ILD), which is characterized by progressive scarring of the lung. While immune suppression is a viable therapeutic option for many ILDs, it is detrimental in IPF. Nevertheless, enlargement of lung-draining lymph nodes (LLN) is prevalent and predicts survival outcomes in all ILD patients, including IPF, suggesting that there is also an immune component to IPF pathogenesis. To test this hypothesis, we investigated the cellular composition of the LLN in IPF and non-IPF ILD patients. All ILD patients’ LLN had increased germinal centers (GC), implicating that immune responses are not being resolved. As T regulatory cells (Tregs) are key in the control and resolution of immune responses, we investigated the phenotype of Tregs in the LLN of both IPF and non-IPF ILD patients to reveal their role in lung fibrosis. Tregs were expanded in LLN of all ILD patients compared to controls. Interestingly, there was a predominance of GATA3-expressing Tregs in ILD patients’ LLN. However, Tregs of ILD patients expressed significantly elevated levels of ICOS and CD137 compared to controls, indicating recent activation. Surprisingly, CD25 expression was significantly lower on Tregs from ILD patients’ LLN. Sc-RNAseq confirms similar Treg phenotypes in the lungs of IPF patients. In a mouse model of bleomycin-induced pulmonary fibrosis, Tregs expand and display a similar activated phenotype. Preliminary experiments suggest that GATA3 expression in Tregs is detrimental in this model. Together, these data indicate that there are unresolved GC responses in the LLN of all ILD patients, including IPF, and suggest that Tregs are ineffectual at controlling the GC response.
American Heart Association Postdoctoral Fellowship, Award number (835938)
1 Liver + 4 Diseases = n Drugs
Authors: Dell Garrison Jeffrey , Steele Hope , Cheng Jason Liang , Breckenridge Joey Emery , Willicut Ashley , Ghastine Andrew , Culberson Erica , Herro Rana 
Published: August 2023

Abstract
The burden placed on the global population by the instance of liver diseases is significant and there exists a great need for accurate diagnostic method alternatives to invasive biopsy. Here we conduct a comparative histopathological and transcriptomic analyses of 4 models of liver diseases to identify distinct biological pathways which aide in understanding the mechanisms of each disease and provide insight into biomarkers for diagnosing disease, monitoring disease progression, and for evaluating future therapeutic efficacy. The NASH model is metabolically induced by high fat/high fructose diet, whereas the three other models are hepatotoxicants, and involve Allyl Alcohol (AA), 4,4′-Methylenedianiline (MDA), or Bromobenzene (BB). Analysis of liver sections stained with Hematoxylin and Eosin and Masson’s trichrome revealed distinct pathological differences between each of the models. These differences were further confirmed by gene set enrichments and pathway analyses. This transcriptomic comparative study outlines distinct pathways important for inflammation, steatosis and fibrosis of the liver that are unique to each model in addition to common pathways contributing to disease pathogenesis. It further validates the histopathological features characteristic of each disorder.
Negative regulation of intestinal IgA by inflammatory cues in absence of infection
Authors: Sung Chi Chang , Barbet Gaetan 
Published: August 2023

Abstract
Pathogen-specific IgA elicited at mucosal site is associated with a better immune protection upon vaccination. Although some effective mucosal vaccines are available and approved, the majority of current vaccines are administered intramuscularly or subcutaneously and were developed irrespectively of a particular site of infection. Vaccine adjuvants are selected in their capacity to elicit a strong inflammatory response. However, few studies show the effectiveness of FDA-approved adjuvants to mucosal vaccines. Here, we unveil the role of inflammatory pathways in their negative regulation of IgA production in murine intestines. We are currently studying and identifying the molecular and cellular pathways involved in the production of protective IgA antibodies in the intestines in absence of particular inflammatory cues. Indeed, in absence of some inflammatory signals, we observed an increase of free IgAs and IgA-bound bacteria in the intestines of mice in steady state. IgA levels were normal in the serum and the concentration of IgG antibodies were also normal in both the intestines and serum suggesting a skewed toward IgAs restricted to the intestines. Furthermore, we could show that this IgA production is for the most part T-dependent and involves the formation of germinal centers localized in the Peyer’s Patches. We are now developing vaccine models to elicit potent IgA-mediated protection in the aim to use these finding at our advantage to promote pathogen-specific intestinal responses through new designs of mucosal vaccines. We believe that developing mucosal vaccines that elicit a protective immunity through natural routes of infection hold the promise to act better, faster and safer against worldwide pathogenic threat.
RWJF Grant Project # 826399 Moderna Research Fellowship
TFR cell-mediated regulation of IgA and gut microbiota in food allergy
Authors: Abdi Abdullahi M , Dent Alexander L 
Published: August 2023

Abstract
About two grams of secretory Immunoglobulin A (sIgA) are secreted daily into the intestine, where it serves as the first line of defense against pathogens and protects the host’s commensals by coating them, ensuring mucosal immune system homeostasis. Gut microbiota (Mb)-specific IgA can be generated via T cell-dependent (TD) or T cell-independent (TI) pathways. However, the TD route is required to generate high-affinity, pathogen-specific antibodies (Abs). T follicular helper (Tfh) cells are essential for germinal center (GC) development, promoting B cells expansion. Nevertheless, the role of T follicular regulatory (Tfr) cells in Mb-reactive IgA production is unclear. Unpublished results and a prior work demonstrated that deleting Tregs or Tfr cells in mice increased antigen-specific IgA titers, indicating that these cells negatively control IgA response. Thus, we hypothesized that TFR cells regulate gut Mb-reactive IgA via the regulation of high-affinity IgA production against gut Mb. To test this hypothesis, TFR cell-deficient Bcl6FC (Foxp3 cre-Bcl6 fl/fl) and WT mice were primed with peanut plus cholera toxin through oral gavage in a food allergy model. Then, we stained fecal Mb with anti-mouse IgA, sorted commensals into IgA +and IgA −populations and sequence 16S rRNA gene of the sorted bacteria (IgA-seq) to evaluate the Mb-reactive IgA response. Flow cytometry data showed significantly higher IgA-coated bacteria in Bcl6FC than in WT mice, showing TFR cells adversely modulate IgA response to gut Mb. Our current experiments will dissect if the higher Mb-reactive IgA secreted in the absence of TFR cells were non-specific and polyreactive or were specific towards certain members of gut commensals.
R21 AI172087 NIH
Synergy between anti-commensal antibodies and IL-10 in gut homeostasis and disease
Authors: Hsu Jung-Shan , Klocke Barbara J , Landuyt Ashley E , Muir Rachel Q , Jennings Melissa S , Molina Patrick A , Randall Troy D , Elson Charles O , Lee Goo , Maynard Craig L 
Published: August 2023

Abstract
To limit immune responses to gut-resident microbes, mucus and the epithelial barrier physically separate the host immune system from microbial communities. Immunoglobulin A (IgA) antibodies are known to help to regulate the colonization and functions of mucus-associated commensals. However, the mechanisms whereby other isotypes, including IgG, interface with other components of the immune system to promote tolerance of MA microbes are still being unraveled. Our lab previously showed that, in mice with impaired production of T-dependent (TD) anti-commensal antibodies, IgA- and IgG-mediated recognition of mucus-derived bacterial antigens is significantly reduced. Moreover, transient depletion of IL-10-producing cells from mice devoid of TD antibodies resulted in rapid onset colitis. Therefore, we hypothesized that anti-commensal antibodies synergize with IL-10 to promote immune homeostasis in the colon. Here, we show that, relative to wild type mice, there are increased levels of colonic IL-10-producing CD4 T cells in mice lacking both IgG and IgA (Aicda−/−) but not in mice lacking only IgA (Iga−/−). Additionally, transient depletion of IL-10-producing cells resulted in severe colitis in Aicda−/− but not Iga−/− mice. Importantly, this colitis developed despite elevated levels of IgM in the colon mucus layer of Aicda−/− mice. Furthermore, we observed elevated levels of IgG in the colon mucus layer of Iga−/− mice suggesting a potentially compensatory role for IgG in mice devoid of IgA. Collectively, our data suggest an important contribution for anti-commensal IgG, including in combination with IL-10, in promoting immune tolerance of border-dwelling microbes.
R01 AI162736, R21 DK118386
Endogenous retroviruses act as a tonic signal for Treg induction and oral tolerance development
Authors: Rivera Claudia A , Krishnamurthy Siddharth R , Belkaid Yasmine 
Published: August 2023

Abstract
Endogenous retrovirus (ERVs) result from accumulated germline infections by ancient exogenous retroviruses and constitute a sizable proportion of the mammalian genome. Recent findings support the idea that these elements control tissue immune threshold of activation and inflammation. Within this context, our lab has recently described that, in the skin, the beneficial immunity to the microbiota depends on endogenous retrovirus expression. How the crosstalk between ERVs and the microbiota controls tissue immunity at other barrier sites and how dysregulation of this dialogue impairs tolerance responses remain unknown. Our results reveal that antiretroviral (anti-RT) treatment (blocking the conversion of ERV into cDNA) impairs gut regulatory T cell homeostasis and differentiation and compromises the induction of oral tolerance to food antigens. Consistently, our data demonstrate that antiretroviral treatment impairs type I IFN signaling in mucosal dendritic cells involved in the induction of regulatory T cells. Further, our results show that epithelial ERV expression is highly conserved across tissues and seems to be only partially affected by the microbiota in the small intestine. Altogether, our results propose that ERV control immunoregulation within the gastrointestinal tract by acting as a tonic signal required for Treg induction and differentiation thereby promoting oral tolerance and preventing allergic responses.
This work was supported in part by intramural funds of NIAID, NIH. C.A.R is supported by Damon Runyon Fellowship program (DRG-2496-23).
Homeostatic intestinal immunity is dominated by microbiota-independent cytotoxic Th1 cells
Authors: Gine Eduardo Ansaldo , McFadden Taryn , Yong Daniel , Bouladoux Nicolas , Link Verena M , Corral Dan , Belkaid Yasmine 
Published: August 2023

Abstract
Barrier tissues such as the skin and gut form the first line of defense against pathogens and establish an immune dialogue with the resident microbiota. To that end, they harbor immune populations geared towards barrier function, tolerance, and repair. Of particular interest are the large numbers of T cells. Some are specific towards commensal microbes or recognize previously encountered pathogens. However, the origin, antigen specificity, and function of most barrier tissue-resident T cells remains elusive. We have identified a dominant CD4 T cell population in the small intestine lamina propria characterized by the expression of T-bet. Surprisingly, this population is present in both germ-free and conventionally reared (SPF) mice at homeostasis, and accounts for almost half of T cells at this site. Bulk and single-cell transcriptome analysis revealed that this population has cytotoxic potential. Furthermore, single-cell TCR sequencing showed parallel T cell repertoires in germ-free and SPF mice, suggesting shared antigen specificity. To explore the ontogeny of this population, we have identified clonally expanded TCRs and are generating low frequency retrogenic mice. Additionally, we have generated several T cell hybridomas from small intestine Th1 cells. Interestingly, preliminary results suggest that small intestinal Th1 TCRs exhibit some degree of self-reactivity, which may determine their development and differentiation in a foreign antigen-independent manner. Collectively, these studies will shed light on the function of a dominant intestinal T cell population and reveal novel insights into homeostatic tissue immune networks.
This research was supported by the Intramural Research Program of NIAID, NIH.
In neonatal mice, intestinal CD64+ CD206+ MHCII− macrophages do not recover one-week post-anti-CSF1 antibody injection, which affects key pro-angiogenic proteins
Authors: Managlia Elizabeth , Yan Xiaocai , De Plaen Isabelle 
Published: August 2023

Abstract
The small intestine of neonatal mice has abundant embryonically derived macrophages (MΦ) but only a small subset remains into adulthood. These MΦ play a role in fetal and neonatal development. In the intestine, they are juxtaposed to endothelial cells where they facilitate postnatal vascular expansion. Postnatal recruitment of bone-marrow-derived blood monocytes complicates the identification and study of the embryonically derived compartment since adult monocytes differentiate into intestinal MΦ of similar phenotype. Whether the different lineages have unique roles in neonates needs further investigation. To explore the role of intestinal MΦ during the neonatal period, we depleted neonatal MΦ by injecting 1-day old pups with anti-CSF1 antibodies and allowed the intestine to repopulate with monocyte-derived MΦ for a week.
Flow cytometry on small intestinal tissue revealed that at D8, 2.77% (± 0.32) of intestinal MΦ were CD64+ CD206+ MHCII− in IgG injected pups compared to 0.18% (± 0.05) in anti-CSF1 injected pups (p=0.001). These data show that postnatal bone-marrow derived MΦ do not reconstitute the intestinal CD64+ CD206+ MHCII− MΦ, which are abundant at birth but not detected in adults. We determined by western blot that intestinal Vegfr2 and Igf1r protein expression, which we have found to be critical for neonatal intestinal microvascular development, were decreased in 8-day old pups injected with anti-CSF1 2.1-fold (p=0.047) and 1.5-fold (p=0.15) respectively when compared to IgG injected littermates. Taken together, these results suggest that CD64+ CD206+ MHCII− MΦ may be embryonically derived and their transient presence may be important for neonatal microvascular development.
NIH DK116568 (IDP)