Type 2 cytokines in the thymus activate Sirpα+ dendritic cells to promote clonal deletion.

The thymus contains a diversity of dendritic cells (DCs) that exist in defined locations and have different antigen-processing and -presenting features. This suggests that they play nonredundant roles in mediating thymocyte selection. In an effort to eliminate SIRPα+ classic DC2 subsets, we discovered that a substantial proportion expresses the surface lectin, CD301b, in the thymus. These cells resemble the CD301b+ type 2 immune response promoting DCs that are present in the skin-draining lymph nodes. Transcriptional and phenotypic comparison to other DC subsets in the thymus revealed that thymic CD301b+ cDCs represent an activated state that exhibits enhanced antigen processing and presentation. Furthermore, a CD301b+ cDC2 subset demonstrated a type 2 cytokine signature and required steady-state interleukin-4 receptor signaling. Selective ablation of CD301b+ cDC2 subsets impaired clonal deletion without affecting regulatory T cells (Treg cells). The T cell receptor α repertoire sequencing confirmed that a cDC2 subset promotes deletion of conventional T cells with minimal effect on Treg cell selection. Together, these findings suggest that cytokine-induced activation of DCs in the thymus substantially enforces central tolerance.

Myeloid cells activate iNKT cells to produce IL-4 in the thymic medulla.

Interleukin-4 (IL-4) is produced by a unique subset of invariant natural killer T (iNKT) cells (NKT2) in the thymus in the steady state, where it conditions CD8+ T cells to become "memory-like" among other effects. However, the signals that cause NKT2 cells to constitutively produce IL-4 remain poorly defined. Using histocytometry, we observed IL-4-producing NKT2 cells localized to the thymic medulla, suggesting that medullary signals might instruct NKT2 cells to produce IL-4. Moreover, NKT2 cells receive and require T cell receptor (TCR) stimulation for continuous IL-4 production in the steady state, since NKT2 cells lost IL-4 production when intrathymically transferred into CD1d-deficient recipients. In bone marrow chimeric recipients, only hematopoietic, not stromal, antigen-presenting cells (APCs), provided such stimulation. Furthermore, using different Cre-recombinase transgenic mouse strains to specifically target CD1d deficiency to various APCs, together with the use of diphtheria toxin receptor (DTR) transgenic mouse strains to deplete various APCs, we found that macrophages were the predominant cell to stimulate NKT2 IL-4 production. Thus, NKT2 cells appear to encounter and require different activating ligands for selection in the cortex and activation in the medulla.

ARTC2.2/P2RX7 Signaling during Cell Isolation Distorts Function and Quantification of Tissue-Resident CD8+ T Cell and Invariant NKT Subsets.

Peripheral invariant NKT cells (iNKT) and CD8+ tissue-resident memory T cells (TRM) express high levels of the extracellular ATP receptor P2RX7 in mice. High extracellular ATP concentrations or NAD-mediated P2RX7 ribosylation by the enzyme ARTC2.2 can induce P2RX7 pore formation and cell death. Because both ATP and NAD are released during tissue preparation for analysis, cell death through these pathways may compromise the analysis of iNKT and CD8+ TRM Indeed, ARTC2.2 blockade enhanced recovery of viable liver iNKT and TRM The expression of ARTC2.2 and P2RX7 on distinct iNKT subsets and TRM is unclear, however, as is the impact of recovery from other nonlymphoid sites. In this study, we performed a comprehensive analysis of ARTC2.2 and P2RX7 expression in iNKT and CD8+ T cells in diverse tissues, at steady-state and after viral infection. NKT1 cells and CD8+ TRM express high levels of both ARTC2.2 and P2RX7 compared with NKT2, NKT17, and CD8+ circulating memory subsets. Using nanobody-mediated ARTC2.2 antagonism, we showed that ARTC2.2 blockade enhanced NKT1 and TRM recovery from nonlymphoid tissues during cell preparation. Moreover, blockade of this pathway was essential to preserve functionality, viability, and proliferation of both populations. We also showed that short-term direct P2RX7 blockade enhanced recovery of TRM, although to a lesser degree. In summary, our data show that short-term in vivo blockade of the ARTC2.2/P2RX7 axis permits much improved flow cytometry-based phenotyping and enumeration of murine iNKT and TRM from nonlymphoid tissues, and it represents a crucial step for functional studies of these populations.

Post-traumatic Acetabulum Open Reduction Internal Fixation in a Patient with a BMI of 77: A Case Report.

CASE: We present the case of a 25-year-old man with body mass index of 77 who underwent open reduction internal fixation (ORIF) of a displaced fracture dislocation of the acetabulum after a high-speed motor vehicle accident. Remarkably, he achieved full weight-bearing with minimal hip pain and has returned to independent mobility and meaningful work. CONCLUSION: ORIF of an acetabular fracture in a patient with class III obesity presents many challenges. Positioning, surgical approach, fracture manipulation, and postoperative morbidity and mortality can be managed through interdisciplinary collaboration and preoperative communication.

Developmental origins and oncogenic pathways in malignant brain tumors.

Brain tumors such as adult glioblastomas and pediatric high-grade gliomas or medulloblastomas are among the leading causes of cancer-related deaths, exhibiting poor prognoses with little improvement in outcomes in the past several decades. These tumors are heterogeneous and can be initiated from various neural cell types, contributing to therapy resistance. How such heterogeneity arises is linked to the tumor cell of origin and their genetic alterations. Brain tumorigenesis and progression recapitulate key features associated with normal neurogenesis; however, the underlying mechanisms are quite dysregulated as tumor cells grow and divide in an uncontrolled manner. Recent comprehensive genomic, transcriptomic, and epigenomic studies at single-cell resolution have shed new light onto diverse tumor-driving events, cellular heterogeneity, and cells of origin in different brain tumors. Primary and secondary glioblastomas develop through different genetic alterations and pathways, such as EGFR amplification and IDH1/2 or TP53 mutation, respectively. Mutations such as histone H3K27M impacting epigenetic modifications define a distinct group of pediatric high-grade gliomas such as diffuse intrinsic pontine glioma. The identification of distinct genetic, epigenomic profiles and cellular heterogeneity has led to new classifications of adult and pediatric brain tumor subtypes, affording insights into molecular and lineage-specific vulnerabilities for treatment stratification. This review discusses our current understanding of tumor cells of origin, heterogeneity, recurring genetic and epigenetic alterations, oncogenic drivers and signaling pathways for adult glioblastomas, pediatric high-grade gliomas, and medulloblastomas, the genetically heterogeneous groups of malignant brain tumors. This article is categorized under: Gene Expression and Transcriptional Hierarchies > Gene Networks and Genomics Adult Stem Cells, Tissue Renewal, and Regeneration > Stem Cell Differentiation and Reversion Signaling Pathways > Cell Fate Signaling.

Constitutively Active MDA5 Proteins Are Inhibited by Paramyxovirus V Proteins.

Excessive interferon (IFN) production and signaling can lead to immunological and developmental defects giving rise to autoimmune diseases referred to collectively as "type I interferonopathies." A subset of these diseases is caused by monogenic mutations affecting proteins involved in nucleic acid sensing, homeostasis, and metabolism. Interferonopathic mutations in the cytosolic antiviral sensor MDA5 render it constitutively hyperactive, resulting in chronic IFN production and IFN-stimulated gene expression. Few therapeutic options are available for patients with interferonopathic diseases, but a large number of IFN evasion and antagonism strategies have evolved in viral pathogens that can counteract IFN production and signaling to enhance virus replication. To test the hypothesis that these natural IFN suppressors could be used to subdue the activity of interferonopathic signaling proteins, hyperactive MDA5 variants were assessed for susceptibility to a family of viral MDA5 inhibitors. In this study, Paramyxovirus V proteins were tested for their ability to counteract constitutively active MDA5 proteins. Results indicate that the V proteins are able to bind to and disrupt the signaling activity of these MDA5 proteins, irrespective of their specific mutations, reducing IFN production and IFN-stimulated gene expression to effectively suppress the hyperactive antiviral response.

Ultrasound Guided Intra-thymic Injection to Track Recent Thymic Emigrants and Investigate T Cell Development.

To track recent thymic emigrants (RTEs) or study T cell development in the thymus, intra-thymic injection of a cellular tag or precursor cells for various T cell lineages is often desired. However, the traditional surgical approach to expose the thymus for intra-thymic injection is time-consuming and can cause a high level of pain and stress to animals, which might disrupt immune homeostasis, potentially confounding the results. Here, we introduce an ultrasound guided intra-thymic injection procedure, which is non-surgical and minimally invasive to animals. This technique is relatively easy to learn and offers an efficient and accurate tool to track RTEs or perform intra-thymic transfer of various cell types to investigate their differentiation.

Sticky situations: cyanoacrylate exposures reported to a poison control system.

INTRODUCTION: Cyanoacrylate (Super Glue®) exposures are commonly reported to poison control centers, but little has been published in the medical literature regarding these exposures. We sought to characterize cyanoacrylate exposures reported to a poison control system. METHODS: We performed a retrospective review of a poison system's database for all cases of single-substance human exposure to cyanoacrylate-containing products from 2005 to 2015. Data collected included age, gender, route of exposure, clinical effects, treatments recommended and medical outcome. RESULTS: There were a total of 893 patients, 505 (56.6%) of which were female. Patient ages ranged from 6 months to 88 years with a median of 11 years. The vast majority of exposures (n = 871, 97.5%) were unintentional, but a small number of exposures (n = 22, 2.5%) were due to intentional misuse (such as trying to stop a bleeding cut) or malicious intent (such as purposefully gluing a person's eyes shut as a prank). Routes of exposure included: ingestion, n = 337 (37.7%); ocular, n = 322 (36.1%); dermatologic, n = 285 (31.9%); inhalation, n = 16 (1.8%); nasal, n = 1 (0.1%); and otic, n = 1 (0.1%); some patients had multiple routes of exposure. Treatments recommended by the poison center included irrigation (n = 411), petroleum jelly (n = 143), mineral oil (n = 131), topical antibiotic ointment (n = 82), peanut butter (n = 6), acetone (n = 4) and WD-40® (n = 2). A total of 657 patients (73.6%) were managed on-site, while 236 (26.4%) were seen in a health care facility. Among all exposures, effects were classified as none (n = 287), minor (n = 529) and moderate (n = 77). No major effects or deaths were reported. CONCLUSIONS: In this case series, the majority of cases occurred in children and most exposures did not result in significant morbidity. Notably, there was wide variation in terms of recommended treatments; further study is needed to determine the optimal treatment method and to standardize poison center recommendations for treating patients with cyanoacrylate exposures.

A retrospective review of isolated duloxetine-exposure cases.

BACKGROUND: Duloxetine is a serotonin norepinephrine reuptake inhibitor (SNRI) approved in the US for the treatment of major depression, generalized anxiety, fibromyalgia, diabetic peripheral neuropathy, and chronic musculoskeletal pain. Given the limited published information regarding human overdoses to this medication, our goal was to characterize such exposures. METHODS: We retrospectively reviewed a state poison system's database for all single agent exposures to duloxetine from 2004-2011. Data collected included age, gender, circumstances surrounding exposure, symptoms, and outcome. Patients with co-ingestants, confirmed non-exposure, unknown outcomes, or other coding errors were excluded. RESULTS: 159 cases were identified. 106 were included for review. Of 61 pediatric and adolescent cases (0-19 years old) identified, 53 involved unintentional overdose. Three patients experienced symptoms and none were admitted. All intentional ingestions(8) were seen in the emergency department, two patients experienced symptoms. No intentional ingestion was admitted for medical care. Fifty-one adult cases were included for review. Four adult patients were admitted following intentional duloxetine overdose with resolution of symptoms within 24 hours. Three adults were evaluated in a HCF following non-self-harm exposures to duloxetine. None of these patients were admitted. The remaining 15 adult patients with non-self-harm exposures were safely managed at home. CONCLUSION: The majority of non-self-harm duloxetine-exposed adult and pediatric/adolescent patients were safely managed at home and when evaluated in a healthcare facility, did not require further hospitalization. Intentional pediatric/adolescent and adult duloxetine exposures were managed in a healthcare facility but rarely resulted in further hospitalization, serious morbidity, or mortality.

Development of genome-wide DNA polymorphism database for map-based cloning of rice genes.

DNA polymorphism is the basis to develop molecular markers that are widely used in genetic mapping today. A genome-wide rice (Oryza sativa) DNA polymorphism database has been constructed in this work using the genomes of Nipponbare, a cultivar of japonica, and 93-11, a cultivar of indica. This database contains 1,703,176 single nucleotide polymorphisms (SNPs) and 479,406 Insertion/Deletions (InDels), approximately one SNP every 268 bp and one InDel every 953 bp in rice genome. Both SNPs and InDels in the database were experimentally validated. Of 109 randomly selected SNPs, 107 SNPs (98.2%) are accurate. PCR analysis indicated that 90% (97 of 108) of InDels in the database could be used as molecular markers, and 68% to 89% of the 97 InDel markers have polymorphisms between other indica cultivars (Guang-lu-ai 4 and Long-te-pu B) and japonica cultivars (Zhong-hua 11 and 9522). This suggests that this database can be used not only for Nipponbare and 93-11, but also for other japonica and indica cultivars. While validating InDel polymorphisms in the database, a set of InDel markers with each chromosome 3 to 5 marker was developed. These markers are inexpensive and easy to use, and can be used for any combination of japonica and indica cultivars used in this work. This rice DNA polymorphism database will be a valuable resource and important tool for map-based cloning of rice gene, as well as in other various research on rice (http://shenghuan.shnu.edu.cn/ricemarker).