Single-cell profiling identifies mechanisms of inflammatory heterogeneity in chronic rhinosinusitis.

The heterogeneous cellular microenvironment of human airway chronic inflammatory diseases, including chronic rhinosinusitis (CRS) and asthma, is still poorly understood. Here, we performed single-cell RNA sequencing (scRNA-seq) on the nasal mucosa of healthy individuals and patients with three subtypes of CRS and identified disease-specific cell subsets and molecules that specifically contribute to the pathogenesis of CRS subtypes. As such, ALOX15+ macrophages contributed to the type 2 immunity-driven pathogenesis of one subtype of CRS, eosinophilic CRS with nasal polyps (eCRSwNP), by secreting chemokines that recruited eosinophils, monocytes and T helper 2 (TH2) cells. An inhibitor of ALOX15 reduced the release of proinflammatory chemokines in human macrophages and inhibited the overactivation of type 2 immunity in a mouse model of eosinophilic rhinosinusitis. Our findings advance the understanding of the heterogeneous immune microenvironment and the pathogenesis of CRS subtypes and identify potential therapeutic approaches for the treatment of CRS and potentially other type 2 immunity-mediated diseases.

TSPO deficiency exacerbates acute lung injury via NLRP3 inflammasome-mediated pyroptosis.

BACKGROUND: Acute respiratory distress syndrome (ARDS) is a common cause of respiratory failure in many critically ill patients. Although inflammasome activation plays an important role in the induction of acute lung injury (ALI) and ARDS, the regulatory mechanism of this process is still unclear. When cells are stimulated by inflammation, the integrity and physiological function of mitochondria play a crucial part in pyroptosis. However, the underlying mechanisms and function of mitochondrial proteins in the process of pyroptosis are largely not yet known. Here, we identified the 18-kDa translocator protein (TSPO), a mitochondrial outer membrane protein, as an important mediator regulating nucleotide-binding domain, leucine-rich repeat, and pyrin domain-containing protein 3 (NLRP3) inflammasome activation in macrophages during ALI. METHODS: TSPO gene knockout (KO) and lipopolysaccharide (LPS)-induced ALI/ARDS mouse models were employed to investigate the biological role of TSPO in the pathogenesis of ARDS. Murine macrophages were used to further characterize the effect of TSPO on the NLRP3 inflammasome pathway. Activation of NLRP3 inflammasome was preformed through LPS + adenosine triphosphate (ATP) co-stimulation, followed by detection of mitochondrial membrane potential, reactive oxygen species (ROS) production, and cell death to evaluate the potential biological function of TSPO. Comparisons between two groups were performed with a two-sided unpaired t -test. RESULTS: TSPO- KO mice exhibited more severe pulmonary inflammation in response to LPS-induced ALI. TSPO deficiency resulted in enhanced activation of the NLRP3 inflammasome pathway, promoting more proinflammatory cytokine production of macrophages in LPS-injured lung tissue, including interleukin (IL)-1ß, IL-18, and macrophage inflammatory protein (MIP)-2. Mitochondria in TSPO -KO macrophages tended to depolarize in response to cellular stress. The increased production of mitochondrial damage-associated molecular pattern led to enhanced mitochondrial membrane depolarization and pyroptosis in TSPO -KO cells. CONCLUSION: TSPO may be the key regulator of cellular pyroptosis, and it plays a vital protective role in ARDS occurrence and development.

Genetic Deletion of Thorase Causes Purkinje Cell Loss and Impaired Motor Coordination Behavior.

Thorase belongs to the AAA+ ATPase family, which plays a critical role in maintaining cellular homeostasis. Our previous work reported that Thorase was highly expressed in brain tissue, especially in the cerebellum. However, the roles of Thorase in the cerebellum have still not been characterized. In this study, we generated conditional knockout mice (cKO) with Thorase deletion in Purkinje cells. Thorase cKO mice exhibited cerebellar degenerative diseases-like behavior and significant impairment in motor coordination. Thorase deletion resulted in more Purkinje neuron apoptosis, leading to Purkinje cell loss in the cerebellum of Thorase cKO mice. We also found enhanced expression of the inflammatory protein ASC, IL-1ß, IL-6 and TNF-α in the Thorase cKO cerebellum, which contributed to the pathogenesis of cerebellar degenerative disease. Our findings provide a better understanding of the role of Thorase in the cerebellum, which is a theoretical basis for Thorase as a therapeutic drug target for neurodegenerative diseases.

Anti-PD-1 antibody armored γδ T cells enhance anti-tumor efficacy in ovarian cancer.

γδ T cells have the unique ability to detect a wide range of tumors with low mutation burdens, making them attractive candidates for CAR-T-cell therapy. Unlike αß T cells and other immune cells, γδ T cells are superior in MHC non-restriction, selective cell recruitment, and rapid activation. However, clinical trials have shown limited clinical benefits, and the adoptive transplantation of γδ T cells has often fallen short of expectations. We hypothesized that the limited effectiveness of γδ T cells in eradicating tumor cells may be attributed to the inhibitory tumor microenvironment induced by the suppressive PD-1/PD-L1 axis. Herein, we constructed novel armored γδ T cells capable of secreting humanized anti-PD-1 antibodies, referred to as "Lv-PD1-γδ T cells. Lv-PD1-γδ T cells showed improved proliferation and enhanced cytotoxicity against tumor cells, resulting in augmented therapeutic effects and survival benefits in ovarian tumor-bearing mice. These engineered cells demonstrated a prolonged in vivo survival of more than 29 days, without any potential for tumorigenicity in immunodeficient NOD/SCID/γ null mice. We also found that Lv-PD1-γδ T cells exhibited excellent tolerance and safety in humanized NOD/SCID/γ null mice. With attenuated or eliminated immunosuppression and maximized cytotoxicity efficacy by the local secretion of anti-PD1 antibodies in tumors, Lv-PD1-γδ T cells can serve as a promising "off-the-shelf" cell therapy against cancers.

Serum metabolomic analysis reveals disorder of steroid hormone biosynthesis in patients with idiopathic inflammatory myopathy.

Idiopathic inflammatory myopathy (IIM) is a heterogeneous group of autoimmune diseases with various clinical manifestations, treatment responses, and prognoses. According to the clinical manifestations and presence of different myositis-specific autoantibodies (MSAs), IIM is classified into several major subgroups, including PM, DM, IBM, ASS, IMNM, and CADM. However, the pathogenic mechanisms of these subgroups remain unclear and need to be investigated. Here, we applied MALDI-TOF-MS to examine the serum metabolome of 144 patients with IIM and analyze differentially expressed metabolites among IIM subgroups or MSA groups. The results showed that the DM subgroup had lower activation of the steroid hormone biosynthesis pathway, while the non-MDA5 MSA group had higher activation of the arachidonic acid metabolism pathway. Our study may provide some insights into the heterogeneous mechanisms of IIM subgroups, potential biomarkers, and management of IIM.

Joint together: The etiology and pathogenesis of ankylosing spondylitis.

Spondyloarthritis (SpA) refers to a group of diseases with inflammation in joints and spines. In this family, ankylosing spondylitis (AS) is a rare but classic form that mainly involves the spine and sacroiliac joint, leading to the loss of flexibility and fusion of the spine. Compared to other diseases in SpA, AS has a very distinct hereditary disposition and pattern of involvement, and several hypotheses about its etiopathogenesis have been proposed. In spite of significant advances made in Th17 dynamics and AS treatment, the underlying mechanism remains concealed. To this end, we covered several topics, including the nature of the immune response, the microenvironment in the articulation that is behind the disease's progression, and the split between the hypotheses and the evidence on how the intestine affects arthritis. In this review, we describe the current findings of AS and SpA, with the aim of providing an integrated view of the initiation of inflammation and the development of the disease.

TSPO Deficiency Exacerbates GSDMD-Mediated Macrophage Pyroptosis in Inflammatory Bowel Disease.

BACKGROUND: the 18-kDa translocator protein (TSPO) is a mitochondrial outer membrane protein, and its expression tends to increase in response to inflammatory stimulation, rapidly. However, the role of TSPO in inflammation and pyroptosis is not yet clear. Here, we identified TSPO as a novel key regulator of pyroptosis. (2) Methods: TSPO knockout and DSS induced mouse inflammatory bowel disease (IBD) models were employed to assess the roles of TSPO in the pathogenesis of IBD. Primary peritoneal macrophages from TSPO knockout mice were applied to evaluate the mechanism of TSPO in cell pyroptosis. CONCLUSIONS: in response to inflammatory injury, TSPO expression is rapidly upregulated and provides a protective function against GSDMD-mediated pyroptosis, which helps us better understand the biological role of TSPO and a novel regulatory mechanism of the pyroptosis process.

ATPase Thorase Deficiency Causes α-Synucleinopathy and Parkinson's Disease-like Behavior.

Parkinson's disease (PD) is one of the most common neurodegenerative diseases and is pathologically characterized by α-synucleinopathy, which is harmful to dopaminergic neurons. However, the underlying mechanisms and pathogenesis of PD remain unclear. The AAA + ATPase Thorase was identified as being essential for neuroprotection and synaptic plasticity by regulating the AMPA receptor trafficking. Here, we found that conditional knockout of Thorase resulted in motor behaviors indicative of neurodegeneration. Genetic deletion of Thorase exacerbated phenotypes of α-synucleinopathy in a familial PD-like A53T mouse model, whereas overexpression of Thorase prevented α-syn accumulation in vivo. Biochemical and cell cultures studies presented here suggest that Thorase interacts with α-syn and regulates the degradation of ubiquitinated α-syn. Thorase deficiency promotes α-syn aggregation in primary cultured neurons. The discoveries in this study provide us with a further understanding of the pathogenesis of α-synucleinopathies including PD.

Genome Instability and Long Noncoding RNA Reveal Biomarkers for Immunotherapy and Prognosis and Novel Competing Endogenous RNA Mechanism in Colon Adenocarcinoma.

Background: Long noncoding RNAs (lncRNAs) crucially modulate DNA damage responses/repair in cancer cells. However, the underlying regulatory role of genome integrity and its clinical value in colon adenocarcinoma (COAD) remains unclear. This study links genome instability to lncRNA using computational biology techniques, in attempt to propose novel biomarkers of immunotherapy outcome, and investigated a potential competing endogenous RNA (ceRNA) as a molecular regulatory mechanism. Methods: TCGA-COAD patients were divided into genome unstable (GU)-like and genome stable (GS)-like clusters via hierarchical clustering to predict immunotherapy outcomes. Multivariate Cox model was established to predict the overall survival rate in COAD patients. Additionally, SVM and LASSO algorithms were applied to obtain hub lncRNAs. A novel genome instability-related ceRNA network was predicted with the Starbase 2.0 database. To better understand how these genes fundamentally interact during tumor progression and development, the mutation analysis and single-gene analysis for each gene was performed. Results: In contrast to those in the GS-like cluster, GU-like-cluster patients demonstrated a higher tumor mutational burden (TMB)/microsatellite instability (MSI), DNA polymerase epsilon (POLE) mutation rate, and immune checkpoint expression, all indicate a greater predictive power for response rate for immunotherapy. The novel prognostic signature demonstrated an outstanding predictive performance (AUC > 0.70). The genes in the genome insatiability-related ceRNA network (including four axes: AL161772.1-has-miR-671-5p (hsa-miR-181d-5p, has-miR-106a-5p)-NINL, AL161772.1-has-miR-106a-5p-TNFSF11, AC124067.4-hsa-miR-92b-3p (hsa-miR-589-5p)-PHYHIPL, and BOLA3-AS1-has-miR-130b-3p-SALL4) were identified as critical regulators of tumor microenvironment infiltration, cancer stemness, and drug resistance. qPCR was performed to validate the expression patterns of these genes. Furthermore, the MSI-high proportion was greater in patients with mutated type than in those with the wild type according to all four target genes, indicating that these four genes modulate genomic integrity and could serve as novel immunotherapy biomarkers. Conclusion: We demonstrated that genome instability-related lncRNA is a novel biomarker for immunotherapy outcomes and prognosis. A novel ceRNA network that modulates genomic integrity, including four lncRNA-miRNA-mRNA axes, was proposed.

TSPO deficiency accelerates amyloid pathology and neuroinflammation by impairing microglial phagocytosis.

Increasing evidence has placed inflammation and immune dysfunction at the center of the pathogenesis of Alzheimer's disease (AD). The mitochondrial protein translocator protein (18 kDa) (TSPO) is highly upregulated in microglia and astrocytes in response to inflammatory stimulation. However, the biological action of TSPO in the pathogenesis of AD has not been determined to date. In this study, we showed that TSPO expression was upregulated in brain tissues from AD patients and AD model mice. APP/PS1 mice lacking TSPO generated significantly higher levels of Aß1-40 and Aß1-42 peptides and more Aß plaques, as well as enhanced microglial activation, in the brain. TSPO-deficient microglia cultured in vitro showed a significant decrease in the ability to phagocytose Aß peptides or latex beads and generated more proinflammatory cytokines (TNF-α and IL-1ß) in response to Aß peptides. Our findings suggest that TSPO has protective functions against neuroinflammation and Aß pathogenesis in AD. TSPO may be a potential drug target for the development of drugs that have therapeutic or preventive effects in neuroinflammatory diseases.

The Function of the Metals in Regulating Epigenetics During Parkinson's Disease.

Parkinson's means Parkinson's disease, a chronic degenerative disease of central nervous system. The main area which is affected by this disease is motor system. Since it firstly founded by James Parkinson in his 1817 publication, nowadays, people still have lots of questions about this disease. This review mainly summarizes the epigenetics of Parkinson's. DNA methylation is one of the epigenetic mechanisms of Parkinson's. During the development of disease, global hypomethylation, and hypermethylation happen in different areas of patients. Another epigenetic mechanism is histone modification. People believe that some metals can induce Parkinson's disease by modulating epigenetic mechanisms. This review summarizes the relationships between different metals and Parkinson's disease. However, the specific roles of most metals in epigenetics are still unknown, which need further research.

The Involvement of Metals in Alzheimer's Disease Through Epigenetic Mechanisms.

Alzheimer's disease (AD) is the most frequent cause of dementia among neurodegenerative diseases. Two factors were hypothesized to be involved in the pathogenesis of AD, namely beta-amyloid cascade and tauopathy. At present, accumulating evidence suggest that epigenetics may be the missing linkage between genes and environment factors, providing possible clues to understand the etiology of the development of AD. In this article, we focus on DNA methylation and histone modification involved in AD and the environment factor of heavy metals' contribution to AD, especially epigenetic mechanisms. If we can integrate information together, and that may find new potential targets for the treatment.

TSPO deficiency induces mitochondrial dysfunction, leading to hypoxia, angiogenesis, and a growth-promoting metabolic shift toward glycolysis in glioblastoma.

BACKGROUND: The ligands of mitochondrial translocator protein (TSPO) have been widely used as diagnostic biomarkers for glioma. However, the true biological actions of TSPO in vivo and its role in glioma tumorigenesis remain elusive. METHODS: TSPO knockout xenograft and spontaneous mouse glioma models were employed to assess the roles of TSPO in the pathogenesis of glioma. A Seahorse Extracellular Flux Analyzer was used to evaluate mitochondrial oxidative phosphorylation and glycolysis in TSPO knockout and wild-type glioma cells. RESULTS: TSPO deficiency promoted glioma cell proliferation in vitro in mouse GL261 cells and patient-derived stem cell-like GBM1B cells. TSPO knockout increased glioma growth and angiogenesis in intracranial xenografts and a mouse spontaneous glioma model. Loss of TSPO resulted in a greater number of fragmented mitochondria, increased glucose uptake and lactic acid conversion, decreased oxidative phosphorylation, and increased glycolysis. CONCLUSION: TSPO serves as a key regulator of glioma growth and malignancy by controlling the metabolic balance between mitochondrial oxidative phosphorylation and glycolysis.1. TSPO deficiency promotes glioma growth and angiogenesis.2. TSPO regulates the balance between mitochondrial oxidative phosphorylation and glycolysis.

Profiling the Gene Expression and DNA Methylation in the Mouse Brain after Ischemic Preconditioning.

Ischemic preconditioning (IPC) is a phenomenon in which a short-term sublethal ischemic exposure induces tolerance to a subsequent lethal ischemic insult; however, the detailed mechanism underlying IPC-induced neuroprotection remains obscure. Here, we applied middle cerebral artery occlusion, a preconditioning ischemic insult mouse model, to investigate the molecular mechanism underlying cerebral IPC. RNA sequencing and whole-genome bisulfite sequencing were performed to explore the gene expression profile and DNA methylation changes after cerebral IPC treatment. In this study, we identified 636 differentially expressed genes enriched for several pathways that were partially overlapping or interconnected in terms of similar gene function. The involvement of several genes in IPC-induced neuroprotection was first reported. Genes induced by IPC, including Arid5a, Nptx2 and Stc2, demonstrated a neuroprotective effect against oxygen-glucose deprivation induced neurotoxicity in vitro. Thus, our findings provide new insights into IPC signaling pathways and offer a novel therapeutic strategy towards stroke.

Stationary phase persister/biofilm microcolony of Borrelia burgdorferi causes more severe disease in a mouse model of Lyme arthritis: implications for understanding persistence, Post-treatment Lyme Disease Syndrome (PTLDS), and treatment failure.

Although most patients with Lyme disease can be cured with a 2-4 week antibiotic therapy, about 10-20% of patients continue to suffer prolonged persistent symptoms, a condition called post-treatment Lyme disease syndrome (PTLDS). The cause for PTLDS is unclear and hotly debated. Borrelia burgdorferi develops morphological variants under stress conditions but their significance is not clear. Here we isolated the biofilm-like microcolony (MC) and planktonic (spirochetal form and round body) (SP) variant forms from the stationary phase culture of B. burgdorferi and showed that the MC and SP variant forms were not only more tolerant to the current Lyme antibiotics but also caused more severe arthritis in mice than the log phase spirochete form (LOG). We propose to divide the persistent Lyme disease into two categories: (1) early development of persistent disease from inoculation with persister/biofilm at the beginning of infection introduced by tick bites, or Type I persistent disease (i.e., PTLDS); and (2) late development of persistent disease due to initial infection not being diagnosed or treated in time such that the infection develops into late persistent disease, or Type II persistent disease. Importantly, we show that the murine infection caused by LOG could be eradicated by ceftriaxone whereas the persistent infection established with MC could not be eradicated by doxycycline (Doxy), ceftriaxone (CefT), or vancomycin (Van), or Doxy+CefT or Van+CefT, but could only be eradicated by the persister drug combination daptomycin+doxycycline+ceftriaxone. We conclude that varying levels of persistence and pathologies of Borrelia infection and the corresponding different treatment responses are mostly dictated by the heterogeneous B. burgdorferi variant forms inoculated at the time of tick bites. These findings may have broad implications for understanding pathogenesis and treatment of not only persistent Lyme disease but also other persistent infections in general and call for studies to evaluate if treatment of persistent infections with persister drug combination regimens is more effective than the current mostly single-antibiotic monotherapy.

γδ T Cells Provide Protective Function in Highly Pathogenic Avian H5N1 Influenza A Virus Infection.

Given the high mortality rate (>50%) and potential danger of intrapersonal transmission, highly pathogenic avian influenza (HPAI) H5N1 epidemics still pose a significant threat to humans. γδ T cells, which participate on the front line of the host immune defense, demonstrate both innate, and adaptive characteristics in their immune response and have potent antiviral activity against various viruses. However, the roles of γδ T cells in HPAI H5N1 viral infection remain unclear. In this study, we found that γδ T cells provided a crucial protective function in the defense against HPAI H5N1 viral infection. HPAI H5N1 viruses could directly activate γδ T cells, leading to enhanced CD69 expression and IFN-γ secretion. Importantly, we found that the trimer but not the monomer of HPAI H5N1 virus hemagglutinin (HA) proteins could directly activate γδ T cells. HA-induced γδ T cell activation was dependent on both sialic acid receptors and HA glycosylation, and this activation could be inhibited by the phosphatase calcineurin inhibitor cyclosporin A but not by the phosphatidylinositol 3-kinase (PI3-K) inhibitors wortmannin and LY294002. Our findings provide a further understanding the mechanism underlying γδ T cell-mediated innate and adoptive immune responses against HPAI H5N1 viral infection, which helps to develop novel therapeutic strategies for the treatment of H5N1 infection in the future.

MicroRNA-146a Overexpression Impairs the Positive Selection during T Cell Development.

MicroRNAs play crucial roles in modulating immune system. miR-146a, a potent feedback suppressor of NF-κB signaling, was shown to limit the innate immune response and myelopoiesis in a knockout mouse model. Here, we observed high lymphopoiesis demonstrated as mild splenomegaly and severe lymphadenopathy in a miR-146a transgenic mouse model. Overexpression of miR-146a resulted in enhanced proliferation and reduced apoptosis of T cells. More activated CD4+ T cells or effector memory T cells were observed in transgenic mice even under physiological conditions. Importantly, as one of the key steps to generate central tolerance, the positive selection of thymocytes is impaired in transgenic mice, resulting in more CD4+CD8+ double-positive thymocytes but fewer CD4+CD8- and CD4-CD8+ single-positive thymocytes. The maturation of selected CD4-CD8+ thymocytes was also impaired, leading to more severe loss of CD4-CD8+ than CD4+CD8- thymocytes in thymus of transgenic mice. Gene expression profiling analysis identified nine positive selection-associated genes, which were downregulated in transgenic mice, including genes encoding major histocompatibility complex class I/II molecules, IL-7 receptor α chain, and Gimap4, whose downregulation may contribute to the impairment of positive selection. Gimap4 was verified as a novel target of miR-146a. These findings further extend our understanding of the function of miR-146a in T cell biology and identify a novel regulatory mechanism underlying the positive selection during T cell development.

Global characterization of differential gene expression profiles in mouse Vγ1+ and Vγ4+ γδ T cells.

Peripheral γδ T cells in mice are classified into two major subpopulations, Vγ1+ and Vγ4+, based on the composition of T cell receptors. However, their intrinsic differences remain unclear. In this study, we analyzed gene expression profiles of the two subsets using Illumina HiSeq 2000 Sequencer. We identified 1995 transcripts related to the activation of Vγ1+ γδ T cells, and 2158 transcripts related to the activation of Vγ4+ γδ T cells. We identified 24 transcripts differentially expressed between the two subsets in resting condition, and 20 after PMA/Ionomycin treatment. We found that both cell types maintained phenotypes producing IFN-γ, TNF-α, TGF-ß and IL-10. However, Vγ1+ γδ T cells produced more Th2 type cytokines, such as IL-4 and IL-5, while Vγ4+ γδ T cells preferentially produced IL-17. Our study provides a comprehensive gene expression profile of mouse peripheral Vγ1+ and Vγ4+ γδ T cells that describes the inherent differences between them.