Palmitoylation landscapes across human cancers reveal a role of palmitoylation in tumorigenesis.

BACKGROUND: Protein palmitoylation, which is catalyzed by palmitoyl-transferase and de-palmitoyl-transferase, plays a crucial role in various biological processes. However, the landscape and dynamics of protein palmitoylation in human cancers are not well understood. METHODS: We utilized 23 palmitoyl-acyltransferases and seven de-palmitoyl-acyltransferases as palmitoylation-related genes for protein palmitoylation analysis. Multiple publicly available datasets were employed to conduct pan-cancer analysis, examining the transcriptome, genomic alterations, clinical outcomes, and correlation with c-Myc (Myc) for palmitoylation-related genes. Real-time quantitative PCR and immunoblotting were performed to assess the expression of palmitoylation-related genes and global protein palmitoylation levels in cancer cells treated with Myc depletion or small molecule inhibitors. Protein docking and drug sensitivity analyses were employed to predict small molecules that target palmitoylation-related genes. RESULTS: We identified associations between palmitoylation and cancer subtype, stage, and patient survival. We discovered that abnormal DNA methylation and oncogenic Myc-driven transcriptional regulation synergistically contribute to the dysregulation of palmitoylation-related genes. This dysregulation of palmitoylation was closely correlated with immune infiltration in the tumor microenvironment and the response to immunotherapy. Importantly, dysregulated palmitoylation was found to modulate canonical cancer-related pathways, thus influencing tumorigenesis. To support our findings, we performed a proof-of-concept experiment showing that depletion of Myc led to reduced expression of most palmitoylation-related genes, resulting in decreased global protein palmitoylation levels. Through mass spectrometry and enrichment analyses, we also identified palmitoyl-acyltransferases ZDHHC7 and ZDHHC23 as significant contributors to mTOR signaling, DNA repair, and immune pathways, highlighting their potential roles in tumorigenesis. Additionally, our study explored the potential of three small molecular (BI-2531, etoposide, and piperlongumine) to modulate palmitoylation by targeting the expression or activity of palmitoylation-related genes or enzymes. CONCLUSIONS: Overall, our findings underscore the critical role of dysregulated palmitoylation in tumorigenesis and the response to immunotherapy, mediated through classical cancer-related pathways and immune cell infiltration. Additionally, we propose that the aforementioned three small molecule hold promise as potential therapeutics for modulating palmitoylation, thereby offering novel avenues for cancer therapy.

DFNA5 regulates immune cells infiltration and exhaustion.

BACKGROUND: DFNA5 (GSDME) belongs to Gasdermin familily that is involved in a variety of cancers and triggers cell pyroptosis after chemical treatment. However, the relationship in DFNA5 between prognosis and immune cells in diverse cancers has been receiving little attention. Tumor immune cells infiltration and exhaustion may associate with patients prognosis. The roles of DFNA5 in tumor immune cells infiltration and exhaustion have not been clarified. METHODS: The expression level of DFNA5 was determined by the Tumour Immune Estimation Resource and the Oncomine database. Then the impacts of DFNA5 in prognosis were assessed by Kaplan-Meier plotter and ULACAN. The correlations between DFNA5 and tumour-infiltrating lymphocytes were explored by TIMER. In addition, the relationships in the expression levels of DFNA5 and typical genes combination of tumour-infiltrating lymphocytes were analysed by GEPIA and TIMER. In this study, we screened the chemokine and immune related proteins interacted with DFNA5 using TurboID system to explore the instantaneous or weak interactions. RESULTS: DFNA5 significantly influences the prognosis in different cancers according to The Cancer Genome Atlas (TCGA). The expression levels of DFNA5 showed positive correlations to the infiltration of macrophages, CD8 + T cells, CD4 + T cells in liver hepatocellular carcinoma (LIHC), colon adenocarcinoma (COAD), and lung adenocarcinoma (LUAD). DFNA5 expression displayed obvious correlations with multiple lymphocytes gene makers in COAD, LIHC and LUAD. DFNA5 expression has effects on the prognosis of liver hepatocellular carcinoma and LUAD. DFNA5 upregulated the expression levels of PDCD1 and CD274 in a dose-dependent manner. Chemokine and immune related proteins interact with DFNA5. CONCLUSIONS: These results indicate that DFNA5 is related to patient prognosis and immune cells, consisting of macrophages, CD4 + T cells, and CD8 + T cells, in diverse cancers. In addition, DFNA5 expression might contribute to the regulation of T cell exhaustion, tumour-associated macrophages (TAMs), and Tregs in COAD, LIHC and LUAD. DFNA5 may regulate immune infiltration via EIF2AK2. IFNGR1 was related to the functions of PD-L1 expression and PD-1 checkpoint pathway. These results indicate that DFNA5 levels may be act as a prognostic factor and predict the degrees of immune cells infiltration in LIHC and LUAD.

Comprehensive analysis of differentially expressed mRNA and circRNA in Ankylosing spondylitis patients' platelets.

Ankylosing spondylitis (AS) is a chronic inflammatory disease significantly decreasing the quality of life. Platelets play an important and active role in the development of AS. Accumulating evidence demonstrated platelets contain diverse RNA repository inherited from megakaryocytes or microvesicles. Platelet RNAs are dynamically affected by pathological conditions and could be used as diagnostic or prognostic biomarkers. However, the role of the platelet RNAs in AS is elusive. In this study, we compared mRNA and circRNA profiles in platelets between AS patients and healthy controls using RNA sequencing and bioinformatic analysis, and found 4996 mRNAs and 2942 circRNAs were differently expressed. The significantly over-expressed mRNAs in AS patients are involved in platelet activity, gap junction, focal adhesion, rap1 and toll and Imd signaling pathway. The previous identified platelet-derived immune mediators such as P2Y1, P2Y12, PF4, GPIbα, CD40L, ICAM2, CCL5 (RANTES), TGF-ß (TGF-ß1 and TGF-ß2) and PDGF (PDGFB and PDGFA) are also included in these over expressed mRNAs, implying these factors may trigger inflammatory cascades and promote the development of AS. Additionally, we found two down-regulated circRNA (circPTPN22 and circFCHSD2) from the intersection analyses of platelets and spinal ligament tissues of AS patients. The circRNA-miRNA-mRNA regulatory network of these two circRNAs was constructed, and the target mRNAs were enriched in Th17 cell differentiation, inflammatory bowel disease, cell adhesion molecules, cytokine-cytokine receptor interaction, Jak-STAT and Wnt signaling pathway, all these pathways participate in the bone remodeling and pro/anti-inflammatory immune regulation in AS. Then, qRT-PCR was performed to validate the expression of selected key mRNAs and circRNAs and the results demonstrated that the expression levels of P2Y12, GPIbα, circPTPN22 and circFCHSD2 were consistent with the sequencing analysis. In addition, the high expression of five predicted miRNAs interacting with circPTPN22 and circFCHSD2 were also detected in AS by qRT-PCR. Taken together, our study presents a comprehensive overview of mRNAs and circRNAs in platelets in AS patients and offers new insight into the mechanisms of platelet involving in the pathogenesis of AS. The mRNAs and circRNAs identified in this study may serve as candidates for diagnosis and targeted treatment of AS.

Injured liver-released miRNA-122 elicits acute pulmonary inflammation via activating alveolar macrophage TLR7 signaling pathway.

Hepatic injury is often accompanied by pulmonary inflammation and tissue damage, but the underlying mechanism is not fully elucidated. Here we identify hepatic miR-122 as a mediator of pulmonary inflammation induced by various liver injuries. Analyses of acute and chronic liver injury mouse models confirm that liver dysfunction can cause pulmonary inflammation and tissue damage. Injured livers release large amounts of miR-122 in an exosome-independent manner into the circulation compared with normal livers. Circulating miR-122 is then preferentially transported to mouse lungs and taken up by alveolar macrophages, in which it binds Toll-like receptor 7 (TLR7) and activates inflammatory responses. Depleting miR-122 in mouse liver or plasma largely abolishes liver injury-induced pulmonary inflammation and tissue damage. Furthermore, alveolar macrophage activation by miR-122 is blocked by mutating the TLR7-binding GU-rich sequence on miR-122 or knocking out macrophage TLR7. Our findings reveal a causative role of hepatic miR-122 in liver injury-induced pulmonary dysfunction.

Clostridium, Bacteroides and Prevotella associates with increased fecal metabolites Trans-4-Hydroxy-L-proline and Genistein in active pulmonary tuberculosis patients during anti-tuberculosis chemotherapy with isoniazid-rifampin-pyrazinamide-ethambutol (HRZE).

Purpose: To investigated the changes of gut microbiome and fecal metabolome during anti-tuberculosis chemotherapy with isoniazid (H)-rifampin (R)-pyrazinamide (Z)-ethambutol (E). Patients and methods: (1) In this study, we recruited 168 stool specimens from 49 healthy volunteers without M. tuberculosis (Mtb), 30 healthy volunteers with latently infected by Mtb, 41 patients with active tuberculosis (ATB), 28 patients with 2-month HRZE treatment and 20 patients with 2-month HRZE followed by 4-month HR treatment. (2) We used 16S rRNA sequencing and an untargeted Liquid Chromatograph Mass Spectrometer-based metabolomics to investigate the changes of gut microbiome and the alteration of fecal metabolome, respectively, during anti-TB chemotherapy. Results: Mtb infection can reduce the diversity of intestinal flora of ATB patients and change their taxonomic composition, while the diversity of intestinal flora of ATB patients were restored during anti-TB chemotherapy. Especially, family Veillonellacea and Bateroidaceae and their genera Veillonella and Bacteroides significantly increased in the gut microbiota during anti-TB chemotherapy. Additionally, Mtb infection dynamically regulates fecal metabolism in ATB patients during anti-TB chemotherapy. Interestingly, the altered abundance of fecal metabolites correlated with the altered gut microbiota, especially the change of gut Clostridium, Bacteroides and Prevotella was closely related to the change of fecal metabolites such as Trans-4-Hydroxy-L-proline and Genistein caused by Mtb infection or anti-TB chemotherapy. Conclusion: Anti-TB chemotherapy with HRZE can disrupt both gut microbiotas and metabolome in ATB patients. Some specific genera and metabolites are depleted or enriched during anti-TB chemotherapy. Therefore, revealing potential relevance between gut microbiota and anti-TB chemotherapy will provide potential biomarkers for evaluating the therapeutic efficacy in ATB patients. Supplementary Information: The online version contains supplementary material available at 10.1007/s12088-022-01003-2.

Photothermal and Enhanced Photocatalytic Therapies Conduce to Synergistic Anticancer Phototherapy with Biodegradable Titanium Diselenide Nanosheets.

Nanomaterial-based photothermal and photocatalytic therapies are effective against various types of cancers. However, combining two or more materials is considered necessary to achieve the synergistic anticancer effects of photothermal and photocatalytic therapy, which made the preparation process complicated. Herein, the authors describe simple 2D titanium diselenide (TiSe2 ) nanosheets (NSs) that can couple photothermal therapy with photocatalytic therapy. The TiSe2 NSs are prepared using a liquid exfoliation method. They show a layered structure and possess high photothermal conversion efficiency (65.58%) and good biocompatibility. Notably, upon near-infrared irradiation, these NSs exhibit good photocatalytic properties with enhanced reactive oxygen species generation and H2 O2 decomposition in vitro. They can also achieve high temperatures, with heat improving their catalytic ability to further amplify oxidative stress and glutathione depletion in cancer cells. Furthermore, molecular mechanism studies reveal that the synergistic effects of photothermal and enhanced photocatalytic therapy can simultaneously lead to apoptosis and necrosis in cancer cells via the HSP90/JAK3/NF-κB/IKB-α/Caspase-3 pathway. Systemic exploration reveals that the TiSe2 NSs has an appreciable degradation rate and accumulates passively in tumor tissue, where they facilitate photothermal and photocatalytic effects without obvious toxicity. Their study thus indicates the high potential of biodegradable TiSe2 NSs in synergistic phototherapy for cancer treatment.

Integrative genomic expression analysis reveals stable differences between lung cancer and systemic sclerosis.

BACKGROUND: The incidence and mortality of lung cancer are the highest among all cancers. Patients with systemic sclerosis show a four-fold greater risk of lung cancer than the general population. However, the underlying mechanism remains poorly understood. METHODS: The expression profiles of 355 peripheral blood samples were integratedly analyzed, including 70 cases of lung cancer, 61 cases of systemic sclerosis, and 224 healthy controls. After data normalization and cleaning, differentially expressed genes (DEGs) between disease and control were obtained and deeply analyzed by bioinformatics methods. The gene ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway enrichment analysis were performed online by DAVID and KOBAS. The protein-protein interaction (PPI) networks were constructed from the STRING database. RESULTS: From a total of 14,191 human genes, 299 and 1644 genes were identified as DEGs in systemic sclerosis and lung cancer, respectively. Among them, 64 DEGs were overlapping, including 36 co-upregulated, 10 co-downregulated, and 18 counter-regulated DEGs. Functional and enrichment analysis showed that the two diseases had common changes in immune-related genes. The expression of innate immune response and response to virus-related genes increased significantly, while the expression of negative regulation of cell cycle-related genes decreased notably. In contrast, the expression of mitophagy regulation, chromatin binding and fatty acid metabolism-related genes showed distinct trends. CONCLUSIONS: Stable differences and similarities between systemic sclerosis and lung cancer were revealed. In peripheral blood, enhanced innate immunity and weakened negative regulation of cell cycle may be the common mechanisms of the two diseases, which may be associated with the high risk of lung cancer in systemic sclerosis patients. On the other hand, the counter-regulated DEGs can be used as novelbiomarkers of pulmonary diseases. In addition, fat metabolism-related DEGs were consideredto be associated with clinical blood lipid data.

Deficiency of NEIL3 Enhances the Chemotherapy Resistance of Prostate Cancer.

Acquired treatment resistance is an important cause of death in prostate cancer, and this study aimed to explore the mechanisms of chemotherapy resistance in prostate cancer. We employed castration-resistant prostate cancer (CRPC), neuroendocrine prostate cancer (NEPC), and chemotherapy-resistant prostate cancer datasets to screen for potential target genes. The Cancer Genome Atlas (TCGA) was used to detect the correlation between the target genes and prognosis and clinical characteristics. Nei endonuclease VIII-like 3 (NEIL3) knockdown cell lines were constructed with RNA interference. Prostate cancer cells were treated with enzalutamide for the androgen deprivation therapy (ADT) model, and with docetaxel and cisplatin for the chemotherapy model. Apoptosis and the cell cycle were examined using flow cytometry. RNA sequencing and western blotting were performed in the knockdown Duke University 145 (DU145) cell line to explore the possible mechanisms. The TCGA dataset demonstrated that high NEIL3 was associated with a high T stage and Gleason score, and indicated a possibility of lymph node metastasis, but a good prognosis. The cell therapy models showed that the loss of NEIL3 could promote the chemotherapy resistance (but not ADT resistance) of prostate cancer (PCa). Flow cytometry revealed that the loss of NEIL3 in PCa could inhibit cell apoptosis and cell cycle arrest under cisplatin treatment. RNA sequencing showed that the knockdown of NEIL3 changes the expression of neuroendocrine-related genes. Further western blotting revealed that the loss of NEIL3 could significantly promote the phosphorylation of ATR serine/threonine kinase (ATR) and ATM serine/threonine kinase (ATM) under chemotherapy, thus initiating downstream pathways related to DNA repair. In summary, the loss of NEIL3 promotes chemotherapy resistance in prostate cancer, and NEIL3 may serve as a diagnostic marker for chemotherapy-resistant patients.

Biodegradable Bi2 O2 Se Quantum Dots for Photoacoustic Imaging-Guided Cancer Photothermal Therapy.

As new 2D layered nanomaterials, Bi2 O2 Se nanoplates have unique semiconducting properties that can benefit biomedical applications. Herein, a facile top-down approach for the synthesis of Bi2 O2 Se quantum dots (QDs) in a solution is described. The Bi2 O2 Se QDs with a size of 3.8 nm and thickness of 1.9 nm exhibit a high photothermal conversion coefficient of 35.7% and good photothermal stability. In vitro and in vivo assessments demonstrate that the Bi2 O2 Se QDs possess excellent photoacoustic (PA) performance and photothermal therapy (PTT) efficiency. After systemic administration, the Bi2 O2 Se QDs accumulate passively in tumors enabling efficient PA imaging of the entire tumors to facilitate imaging-guided PTT without obvious toxicity. Furthermore, the Bi2 O2 Se QDs which exhibit degradability in aqueous media not only have sufficient stability during in vivo circulation to perform the designed therapeutic functions, but also can be discharged harmlessly from the body afterward. The results reveal the great potential of Bi2 O2 Se QDs as a biodegradable multifunctional agent in medical applications.

Effect of Forsythiaside A on the RLRs Signaling Pathway in the Lungs of Mice Infected with the Influenza A Virus FM1 Strain.

Forsythiaside A, a phenylethanoid glycoside monomer extracted from Forsythia suspensa, shows anti-inflammatory, anti-infective, anti-oxidative, and antiviral pharmacological effects. The precise mechanism underlying the antiviral action of forsythiaside A is not completely clear. Therefore, in this study, we aimed to determine whether the anti-influenza action of forsythiaside A occurs via the retinoic acid-inducible gene-I-like receptors (RLRs) signaling pathway in the lung immune cells. Forsythiaside A was used to treat C57BL/6J mice and MAVS-/- mice infected with mouse-adapted influenza A virus FM1 (H1N1, A/FM1/1/47 strain), and the physical parameters (body weight and lung index) and the expression of key factors in the RLRs/NF-κB signaling pathway were evaluated. At the same time, the level of virus replication and the ratio of Th1/Th2 and Th17/Treg of T cell subsets were measured. Compared with the untreated group, the weight loss in the forsythiaside A group in the C57BL/6J mice decreased, and the histopathological sections showed less inflammatory damage after the infection with the influenza A virus FM1 strain. The gene and protein expression of retinoic acid-inducible gene-I (RIG-I), MAVS, and NF-κB were significantly decreased in the forsythiaside A group. Flow cytometry showed that Th1/Th2 and Th17/Treg differentiated into Th2 cells and Treg cells, respectively, after treatment with forsythiaside A. In conclusion, forsythiaside A reduces the inflammatory response caused by influenza A virus FM1 strain in mouse lungs by affecting the RLRs signaling pathway in the mouse lung immune cells.

Anti-influenza activity of berberine improves prognosis by reducing viral replication in mice.

Berberine, a natural isoquinoline alkaloid isolated from the berberis species, has a wide array of biological properties such as anti-inflammatory, antibacterial, antifungal, and antihelminthic effects. We evaluated the antiviral effect of berberine against influenza A/FM1/1/47 (H1N1) in vivo and in vitro. The results showed that berberine strongly suppressed viral replication in A549 cells and in mouse lungs. Meanwhile, berberine relieved pulmonary inflammation and reduced necrosis, inflammatory cell infiltration, and pulmonary edema induced by viral infection in mice when compared with vehicle-treated mice. Berberine suppressed the viral infection-induced up-regulation of TLR7 signaling pathway, such as TLR7, MyD88, and NF-κB (p65), at both the mRNA and protein levels. Furthermore, berberine significantly inhibited the viral infection-induced increase in Th1/Th2 and Th17/Treg ratios as well as the production of inflammatory cytokines. Our data provide new insight into the potential of berberine as a therapeutic agent for viral infection via its antiviral activity.

A novel reporter system for neutralizing and enhancing antibody assay against dengue virus.

BACKGROUND: Dengue virus (DENV) still poses a global public health threat, and no vaccine or antiviral therapy is currently available. Antibody plays distinct roles in controlling DENV infections. Neutralizing antibody is protective against DENV infection, whereas sub-neutralizing concentration of antibody can increase DENV infection, termed antibody-dependent enhancement (ADE). Plaque-based assay represents the most widely accepted method measuring neutralizing or enhancing antibodies. RESULTS: In this study, a novel reporter virus-based system was developed for measuring neutralization and ADE activity. A stable Renilla luciferase reporter DENV (Luc-DENV) that can produce robust luciferase signals in BHK-21 and K562 cells were used to establish the assay and validated against traditional plaque-based assay. Luciferase value analysis using various known DENV-specific monoclonal antibodies showed good repeatability and a well linear correlation with conventional plaque-based assays. The newly developed assay was finally validated with clinical samples from infected animals and individuals. CONCLUSIONS: This reporter virus-based assay for neutralizing and enhancing antibody evaluation is rapid, lower cost, and high throughput, and will be helpful for laboratory detection and epidemiological investigation for DENV antibodies.

SIRT3 and SIRT4 double-genes remodeled the mitochondrial network to induce hepatocellular carcinoma cell line differentiation and suppress malignant phenotypes.

Tumor cells with damaged mitochondria undergo metabolic reprogramming, but gene therapy targeting mitochondria has not been comprehensively reported. In this study, plasmids targeting the normal hepatocyte cell line (L-O2) and hepatocellular carcinoma cell line were generated using three genes SIRT3, SIRT4, and SIRT5. These deacetylases play a variety of regulatory roles in cancer and are related to mitochondrial function. Compared with L-O2, SIRT3 and SIRT4 significantly ameliorated mitochondrial damage in HCCLM3, Hep3B and HepG2 cell lines and regulated mitochondrial biogenesis and mitophagy, respectively. We constructed double-gene plasmid for co-express SIRT3 and SIRT4 using the internal ribosome entry site (IRES). The results indicated that the double-gene plasmid effectively expressed SIRT3 and SIRT4, significantly improved mitochondrial quality and function, and reduced mtDNA level and oxidative stress in HCC cells. MitoTracker analysis revealed that the mitochondrial network was restored. The proliferation, migration capabilities of HCC cells were reduced, whereas their differentiation abilities were enhanced. This study demonstrated that the use of IRES-linked SIRT3 and SIRT4 double-gene vectors induced the differentiation of HCC cells and inhibited their development by ameliorating mitochondrial dysfunction. This intervention helped reverse metabolic reprogramming, and may provide a groundbreaking new framework for HCC treatment.

Identification and characterization of a linearized B-cell epitope on the pr protein of dengue virus.

The four serotypes of dengue virus (DENV) represent one of the major mosquito-borne pathogens globally; so far no vaccine or specific antiviral is available. During virion maturation, the pr protein is cleaved from its precursor form the prM protein on the surface of immature DENV by host protease. Recent findings have demonstrated that the pr protein not only played critical roles in virion assembly and maturation, but was also involved in antibody-dependent enhancement of DENV infection. However, the B-cell epitopes on the pr protein of DENV have not been well characterized. In this study, a set of 11 partially overlapping peptides spanning the entire pr protein of DENV-2 were fused with glutathione S-transferase and expressed in Escherichia coli. ELISA screening with murine hyperimmune antiserum against immature DENV identified the P8 peptide (57KQNEPEDIDCWCNST7¹) in the pr protein as the major immunodominant epitope. Fine mapping by truncated protein assays confirmed the 8-e peptide 57KQNEPEDI64 was the smallest unit capable of antibody binding. Importantly, the 8-e epitope reacted with sera from dengue fever patients. Site-directed mutagenesis revealed the asparagine residue at position 59 was important for epitope recognition. The 8-e epitope coincided well with the B-cell epitopes predicted by Immune Epitope Database analysis, and 3D structural modelling mapped the 8-e peptide on the surface of prM-E heterodimers. Overall, our findings characterized a linearized B-cell epitope on the pr protein of DENV, which will help to understand the life cycle of DENV and pathogenesis of dengue infections in human.

Microbiota regulates the TLR7 signaling pathway against respiratory tract influenza A virus infection.

Although intestinal flora are crucial in maintaining immune homeostasis of the intestine, the role of intestinal flora in immune responses at other mucosal surfaces remains less clear. Here, we show that intestinal flora composition critically regulates the toll-like receptor 7 (TLR7) signaling pathway following respiratory influenza virus infection. TLR7 ligands rescued the immune impairment in antibiotic-treated mice. Intact microbiota provided signals leading to the expression of mRNA for TLR7, MyD88, IRAK4, TRAF6, and NF-κB at steady state. Significant changes in the composition of culturable commensal bacteria reduced the expression levels of components of the TLR7 signaling pathway. Our results reveal the importance of intestinal flora in regulating immunity in the respiratory mucosa through the upregulation of the TLR7 signaling pathway for the proper activation of inflammasomes.

Effects of Intestinal Microorganisms on Influenza-Infected Mice with Antibiotic-Induced Intestinal Dysbiosis, through the TLR7 Signaling Pathway.

BACKGROUND: Stability of intestinal flora is not only important for maintaining stable immune functions; it is also a key immune channel communicating the interaction between lung and intestine. In this study, probiotics and fecal microbiota transplantation (FMT) were used to regulate influenza-infected mice with antibiotic-induced intestinal dysbiosis and the effects of intestinal microorganisms on these mice were subsequently observed and evaluated. METHODS: Mice are housed in a normal environment with intranasal infection with influenza virus (FM1). Real-time quantitative polymerase chain reaction (RT-qPCR) was used to determine messenger RNA expression and lung viral replication of toll-like receptor 7 (TLR7), myeloid differentiation primary reaction 88 (MyD88) and nuclear factor κB (ss) p65 in the TLR7 signaling pathway. Western blotting is used to measure the expression levels of TLR7, MyD88, and NF-κB p65 proteins. Flow cytometry was used to detect the proportion of Th17/T regulated cells. RESULTS: Results showed that compared with the simple virus group, both diversity and species of intestinal flora in influenza-infected mice with antibiotic-induced intestinal dysbiosis were lower, in vivo viral replication was significantly increased, lung and intestinal tissues were seriously damaged, degree of inflammation increased, expression of the TLR7 signaling pathway increased, and the Th1/Th2:Th17/Treg ratio decreased. Probiotics and FMT effectively regulated intestinal flora, improved pathological lung changes and inflammation caused by influenza infection, and adjusted the TLR7 signaling pathway and the Th1/Th2:Th17/Treg ratio. This effect was not obvious in TLR7-⁣/- mice.In summary, by affecting the TLR7 signaling pathway, intestinal microorganisms reduced the inflammatory response in the lungs of influenza-infected mice with imbalances in antibiotic flora. CONCLUSIONS: By affecting the TLR7 signaling pathway, intestinal microorganisms reduced the inflammatory response in the lungs of influenza-infected mice with imbalances in antibiotic flora. In summary, damage to lung tissue and intestinal mucosa in influenza-infected mice with antibiotic-induced intestinal dysbiosis is more serious compared to simple virus-infected mice. Improving intestinal flora using probiotics or FMT can alleviate intestinal inflammation and improve pulmonary inflammation through the TLR7 signaling pathway.

Global genomic diversity and conservation of SARS-CoV-2 since the COVID-19 outbreak.

IMPORTANCE: Our results indicate that most severe acute respiratory syndrome coronavirus 2 genomes sampled from patients had a mutation rate ≤1.07 ‰ and genome-tail proteins (including S protein) were the main sources of genetic polymorphism. The analysis of the virus-host interaction network of genome-tail proteins showed that they shared some antiviral signaling pathways, especially the intracellular protein transport pathway.

Coordinated Priming of NKG2D Pathway by IL-15 Enhanced Functional Properties of Cytotoxic CD4+CD28- T Cells Expanded in Systemic Lupus Erythematosus.

Systemic lupus erythematosus (SLE) is a systemic autoimmune disorder, and numerous aberrations of T cell responses have been reported and were implicated in its pathophysiology. Recently, CD4-positive T cells with cytotoxic potential were shown to be involved in autoimmune disease progression and tissue damage. However, the effector functions of this cell type and their potential molecular mechanisms in SLE patients remain to be elucidated. In this study, we find that cytotoxic CD4+CD28- T cells are expanded in SLE patients with flow cytometry analysis, and the percentage of CD4+CD28- T cells positively correlates with the Systemic Lupus International Collaborating Clinics/ACR Damage Index (SDI). Furthermore, our study suggests that interleukin-15 (IL-15) promotes the expansion, proliferation, and cytotoxic function of CD4+CD28- T cells in SLE patients through activation of the Janus kinase3-STAT5 pathway. Further study indicates that IL-15 not only mediates the upregulation of NKG2D, but also cooperates with the NKG2D pathway to regulate the activation of the phosphatidylinositol 3-kinase (PI3K)/protein kinase B (AKT) pathway. Together, our study demonstrated that proinflammatory and cytolytic CD4+CD28- T cells expand in SLE patients. The pathogenic potential of these CD4+CD28- T cells is driven by the coupling of the IL-15/IL-15R signaling pathway and the NKG2D/DAP10 signaling pathway, which may open new avenues for therapeutic intervention to prevent SLE progression.

Metabolic profiling of patients with different idiopathic inflammatory myopathy subtypes reveals potential biomarkers in plasma.

Idiopathic inflammatory myopathy (IIM) are heterogeneous autoimmune diseases that primarily affect the proximal muscles. IIM subtypes include dermatomyositis (DM), polymyositis (PM), and anti-synthetase syndrome (ASS). Metabolic disturbances may cause irreversible structural damage to muscle fibers in patients with IIM. However, the metabolite profile of patients with different IIM subtypes remains elusive. To investigate metabolic alterations and identify patients with different IIM subtypes, we comprehensively profiled plasma metabolomics of 46 DM, 13 PM, 12 ASS patients, and 30 healthy controls (HCs) using UHPLC-Q Exactive HF mass spectrometer. Multiple statistical analyses and random forest were used to discover differential metabolites and potential biomarkers. We found that tryptophan metabolism, phenylalanine and tyrosine metabolism, fatty acid biosynthesis, beta-oxidation of very long chain fatty acids, alpha-linolenic acid and linoleic acid metabolism, steroidogenesis, bile acid biosynthesis, purine metabolism, and caffeine metabolism are all enriched in the DM, PM, and ASS groups. We also found that different subtypes of IIM have their unique metabolic pathways. We constructed three models (five metabolites) to identify DM, PM, ASS from HC in the discovery and validation sets. Five to seven metabolites can distinguish DM from PM, DM from ASS, and PM from ASS. A panel of seven metabolites can identify anti-melanoma differentiation-associated gene 5 positive (MDA5 +) DM with high accuracy in the discovery and validation sets. Our results provide potential biomarkers for diagnosing different subtypes of IIM and a better understanding of the underlying mechanisms of IIM.

Targeting VCP potentiates immune checkpoint therapy for colorectal cancer.

Immune checkpoint blockade therapies are still ineffective for most patients with colorectal cancer (CRC). Immunogenic cell death (ICD) enables the release of key immunostimulatory signals to drive efficient anti-tumor immunity, which could be used to potentiate the effects of immune checkpoint inhibitors. Here, we showed that inhibition of valosin-containing protein (VCP) elicits ICD in CRC. Meanwhile, VCP inhibitor upregulates PD-L1 expression and compromises anti-tumor immunity in vivo. Mechanistically, VCP transcriptionally regulates PD-L1 expression in a JAK1-dependent manner. Combining VCP inhibitor with anti-PD1 remodels tumor immune microenvironment and reduces tumor growth in mouse models of CRC. Addition of oncolytic virus further augments the therapeutic activity of the combination regimen. Our study shows the molecular mechanism for regulating PD-L1 expression by VCP and suggests that inhibition of VCP has the potential to increase the efficacy of immunotherapy in CRC.