Early rhombic lip Protogenin+ve stem cells in a human-specific neurovascular niche initiate and maintain group 3 medulloblastoma.

We identify a population of Protogenin-positive (PRTG+ve) MYChigh NESTINlow stem cells in the four-week-old human embryonic hindbrain that subsequently localizes to the ventricular zone of the rhombic lip (RLVZ). Oncogenic transformation of early Prtg+ve rhombic lip stem cells initiates group 3 medulloblastoma (Gr3-MB)-like tumors. PRTG+ve stem cells grow adjacent to a human-specific interposed vascular plexus in the RLVZ, a phenotype that is recapitulated in Gr3-MB but not in other types of medulloblastoma. Co-culture of Gr3-MB with endothelial cells promotes tumor stem cell growth, with the endothelial cells adopting an immature phenotype. Targeting the PRTGhigh compartment of Gr3-MB in vivo using either the diphtheria toxin system or chimeric antigen receptor T cells constitutes effective therapy. Human Gr3-MBs likely arise from early embryonic RLVZ PRTG+ve stem cells inhabiting a specific perivascular niche. Targeting the PRTGhigh compartment and/or the perivascular niche represents an approach to treat children with Gr3-MB.

Gallic acid attenuates murine ulcerative colitis by promoting group 3 innate lymphocytes, affecting gut microbiota, and bile acid metabolism.

Gallic acid (GA), a plant phenol that is widely distributed in fruits and vegetables, and exhibits a protective role against ulcerative colitis (UC). UC is an inflammatory disease characterized by immune response disorders. However, the role and mechanism of action of GA in gut immunity remain unknown. Here, we observed that GA treatment improved enteritis symptoms, decreased the concentrations of cytokines TNF-α, IFN-γ, IL-6, IL-17A, and IL-23, increased the concentrations of cytokines IL-10, TGF-ß and IL-22, and increased the proportion of group 3 innate lymphoid cells (ILC3) in mesenteric lymph nodes and lamina propria. However, GA did not upregulate ILC3 or impair UC in antibody-treated sterile mice. Notably, transplantation of fecal bacteria derived from GA-treated UC mice, instead of UC mice, increased ILC3 levels. Therefore, we analyzed the gut microbiota and related metabolites to elucidate the mechanism promoting ILC3. We determined that GA treatment altered the diversity of the gut microbiota and activated the bile acid (BA) metabolic pathway. We evaluated three BAs, namely, UDCA, isoalloLCA, and 3-oxoLCA that were significantly upregulated after GA treatment, improved UC symptoms, and elevated the proportion of ILC3 in vivo and in vitro. Collectively, these data indicate that GA attenuates UC by elevating ILC3 proportion, regulating the gut microbiota, and impacting BA metabolism. Additionally, we highlight the modulatory effects of BAs on ILC3 for the first time. Our findings provide novel insights into the multiple roles of GA in alleviating UC and provide a mechanistic explanation that supports the dietary nutrition in UC therapy.

Washed microbiota transplantation promotes homing of group 3 innate lymphoid cells to the liver via the CXCL16/CXCR6 axis: a potential treatment for metabolic-associated fatty liver disease.

Despite the observed decrease in liver fat associated with metabolic-associated fatty liver disease (MAFLD) in mice following fecal microbiota transplantation, the clinical effects and underlying mechanisms of washed microbiota transplantation (WMT), a refined method of fecal microbiota transplantation, for the treatment of MAFLD remain unclear. In this study, both patients and mice with MAFLD exhibit an altered gut microbiota composition. WMT increases the levels of beneficial bacteria, decreases the abundance of pathogenic bacteria, and reduces hepatic steatosis in MAFLD-affected patients and mice. Downregulation of the liver-homing chemokine receptor CXCR6 on ILC3s results in an atypical distribution of ILC3s in patients and mice with MAFLD, characterized by a significant reduction in ILC3s in the liver and an increase in ILC3s outside the liver. Moreover, disease severity is negatively correlated with the proportion of hepatic ILC3s. These hepatic ILC3s demonstrate a mitigating effect on hepatic steatosis through the release of IL-22. Mechanistically, WMT upregulates CXCR6 expression on ILC3s, thereby facilitating their migration to the liver of MAFLD mice via the CXCL16/CXCR6 axis, ultimately contributing to the amelioration of MAFLD. Overall, these findings highlight that WMT and targeting of liver-homing ILC3s could be promising strategies for the treatment of MAFLD.

Median Tissue Culture Infectious Dose 50 (TCID50) Assay to Determine Infectivity of Cytopathic Viruses.

The emergence of zoonotic viruses like severe acute respiratory syndrome coronavirus (SARS-CoV), Middle East respiratory syndrome coronavirus (MERS-CoV), and SARS-CoV-2 have significantly impacted global health and economy. The discovery of other viruses in wildlife reservoir species present a threat for future emergence in humans and animals. Therefore, assays that are less reliant on virus-specific information, such as neutralization assays, are crucial to rapidly develop diagnostics, understand virus replication and pathogenicity, and assess the efficacy of therapeutics against newly emerging viruses. Here, we describe the discontinuous median tissue culture infectious dose 50 (TCID50) assay to quantitatively determine the titer of any virus that can produce a visible cytopathic effect in infected cells.

Intestinal CXCR6+ ILC3s migrate to the kidney and exacerbate renal fibrosis via IL-23 receptor signaling enhanced by PD-1 expression.

Group 3 innate lymphoid cells (ILC3s) regulate inflammation and tissue repair at mucosal sites, but whether these functions pertain to other tissues-like the kidneys-remains unclear. Here, we observed that renal fibrosis in humans was associated with increased ILC3s in the kidneys and blood. In mice, we showed that CXCR6+ ILC3s rapidly migrated from the intestinal mucosa and accumulated in the kidney via CXCL16 released from the injured tubules. Within the fibrotic kidney, ILC3s increased the expression of programmed cell death-1 (PD-1) and subsequent IL-17A production to directly activate myofibroblasts and fibrotic niche formation. ILC3 expression of PD-1 inhibited IL-23R endocytosis and consequently amplified the JAK2/STAT3/RORγt/IL-17A pathway that was essential for the pro-fibrogenic effect of ILC3s. Thus, we reveal a hitherto unrecognized migration pathway of ILC3s from the intestine to the kidney and the PD-1-dependent function of ILC3s in promoting renal fibrosis.

Breast Cancer With HER2 Immunohistochemical Score 2 and Average HER2 Signals/Cell 6 or More and HER2/CEP17 Ratio Less Than 2 ('ISH Group 3'): A Multiinstitutional Cohort Analysis Emphasizing Outcome and Molecular Subtype.

Breast cancer (BC) with average human epidermal growth factor receptor 2 (HER2) signals/cell ≥6 and HER2/chromosome enumeration probe 17 (CEP17) ratio <2 (in situ hybridization [ISH] group 3) is very rare, accounting for 0.4% to 3.0% of cases sent for the dual-probe ISH assay. Although such patients are currently eligible for treatment with HER2-targeted therapy, their characteristics and outcomes remain poorly understood. Sixty-two BCs with equivocal HER2 immunohistochemical score (2+) and reflex ISH group 3 results were identified across 4 institutions. Available clinicopathologic characteristics, MammaPrint and BluePrint molecular results, and follow-up information were retrospectively analyzed. Most BCs with HER2 equivocal immunohistochemical and ISH group 3 results were histologic grade 2 or 3 (100%), estrogen receptor (ER) positive (90.3%), with an average HER2 signals/cell of 7.3. Molecular profiles revealed that 80% (16/20) of tumors were luminal subtypes, and HER2 molecular subtype was identified in 10% of tumors (2/20). Twelve (19.4%) out of 62 patients developed local recurrence and/or distant metastasis with a median follow-up of 50 months. One (10%) of 10 patients achieved pathologic complete response after neoadjuvant chemotherapy. Forty-nine (79%) out of 62 patients completed anti-HER2 agents, and exploratory analysis showed no statistically significant difference in disease outcomes between patients who completed anti-HER2 treatment and those who did not. Univariate analysis revealed advanced clinical stage, and ER/progesterone receptor negativity was associated with unfavorable disease outcomes, and exploratory multivariate analysis demonstrated that clinical stage was the most significant factor associated with disease outcomes in the studied population. These findings increase our understanding of this rare, but clinically important HER2 category. Large-scale prospective randomized studies are needed to further evaluate the role of perioperative HER2-targeted therapy in this patient population.

The OTX2 Gene Induces Tumor Growth and Triggers Leptomeningeal Metastasis by Regulating the mTORC2 Signaling Pathway in Group 3 Medulloblastomas.

Medulloblastoma (MB) encompasses diverse subgroups, and leptomeningeal disease/metastasis (LMD) plays a substantial role in associated fatalities. Despite extensive exploration of canonical genes in MB, the molecular mechanisms underlying LMD and the involvement of the orthodenticle homeobox 2 (OTX2) gene, a key driver in aggressive MB Group 3, remain insufficiently understood. Recognizing OTX2's pivotal role, we investigated its potential as a catalyst for aggressive cellular behaviors, including migration, invasion, and metastasis. OTX2 overexpression heightened cell growth, motility, and polarization in Group 3 MB cells. Orthotopic implantation of OTX2-overexpressing cells in mice led to reduced median survival, accompanied by the development of spinal cord and brain metastases. Mechanistically, OTX2 acted as a transcriptional activator of the Mechanistic Target of Rapamycin (mTOR) gene's promoter and the mTORC2 signaling pathway, correlating with upregulated downstream genes that orchestrate cell motility and migration. Knockdown of mTOR mRNA mitigated OTX2-mediated enhancements in cell motility and polarization. Analysis of human MB tumor samples (N = 952) revealed a positive correlation between OTX2 and mTOR mRNA expression, emphasizing the clinical significance of OTX2's role in the mTORC2 pathway. Our results reveal that OTX2 governs the mTORC2 signaling pathway, instigating LMD in Group 3 MBs and offering insights into potential therapeutic avenues through mTORC2 inhibition.

PUMC-MB1 is a novel group 3 medulloblastoma preclinical model, sensitive to PI3K/mTOR dual inhibitor.

PURPOSE: Medulloblastoma (MB), a common and heterogeneous posterior fossa tumor in pediatric patients, presents diverse prognostic outcomes. To advance our understanding of MB's intricate biology, the development of novel patient tumor-derived culture MB models with necessary data is still an essential requirement. METHODS: We continuously passaged PUMC-MB1 in vitro in order to establish a continuous cell line. We examined the in vitro growth using Cell Counting Kit-8 (CCK-8) and in vivo growth with subcutaneous and intracranial xenograft models. The xenografts were investigated histopathologically with Hematoxylin and Eosin (HE) staining and immunohistochemistry (IHC). Concurrently, we explored its molecular features using Whole Genome Sequencing (WGS), targeted sequencing, and RNA sequecing. Guided by bioinformatics analysis, we validated PUMC-MB1's drug sensitivity in vitro and in vivo. RESULTS: PUMC-MB1, derived from a high-risk MB patient, displayed a population doubling time (PDT) of 48.18 h and achieved 100% tumor growth in SCID mice within 20 days. HE and Immunohistochemical examination of the original tumor and xenografts confirmed the classification of PUMC-MB1 as a classic MB. Genomic analysis via WGS revealed concurrent MYC and OTX2 amplifications. The RNA-seq data classified it within the Group 3 MB subgroup, while according to the WHO classification, it fell under the Non-WNT/Non-SHH MB. Comparative analysis with D283 and D341med identified 4065 differentially expressed genes, with notable enrichment in the PI3K-AKT pathway. Cisplatin, 4-hydroperoxy cyclophosphamide/cyclophosphamide, vincristine, and dactolisib (a selective PI3K/mTOR dual inhibitor) significantly inhibited PUMC-MB1 proliferation in vitro and in vivo. CONCLUSIONS: PUMC-MB1, a novel Group 3 (Non-WNT/Non-SHH) MB cell line, is comprehensively characterized for its growth, pathology, and molecular characteristics. Notably, dactolisib demonstrated potent anti-proliferative effects with minimal toxicity, promising a potential therapeutic avenue. PUMC-MB1 could serve as a valuable tool for unraveling MB mechanisms and innovative treatment strategies.

An Evaluation of Type 1 Interferon Related Genes in Male and Female-Matched, SARS-CoV-2 Infected Individuals Early in the COVID-19 Pandemic.

SARS-CoV-2 infection has claimed just over 1.1 million lives in the US since 2020. Globally, the SARS-CoV-2 respiratory infection spread to 771 million people and caused mortality in 6.9 million individuals to date. Much of the early literature showed that SARS-CoV-2 immunity was defective in the early stages of the pandemic, leading to heightened and, sometimes, chronic inflammatory responses in the lungs. This lung-associated 'cytokine storm' or 'cytokine release syndrome' led to the need for oxygen supplementation, respiratory distress syndrome, and mechanical ventilation in a relatively high number of people. In this study, we evaluated circulating PBMC from non-hospitalized, male and female, COVID-19+ individuals over the course of infection, from the day of diagnosis (day 0) to one-week post diagnosis (day 7), and finally 4 weeks after diagnosis (day 28). In our early studies, we included hospitalized and critically care patient PBMC; however, most of these individuals were lymphopenic, which limited our assessments of their immune integrity. We chose a panel of 30 interferon-stimulated genes (ISG) to evaluate by PCR and completed flow analysis for immune populations present in those PBMC. Lastly, we assessed immune activation by stimulating PBMC with common TLR ligands. We identified changes in innate cells, primarily the innate lymphoid cells (ILC, NK cells) and adaptive immune cells (CD4+ and CD8+ T cells) over this time course of infection. We found that the TLR-7 agonist, Resiquimod, and the TLR-4 ligand, LPS, induced significantly better IFNα and IFNγ responses in the later phase (day 28) of SARS-CoV-2 infection in those non-hospitalized COVID-19+ individuals as compared to early infection (day 0 and day 7). We concluded that TLR-7 and TLR-4 agonists may be effective adjuvants in COVID-19 vaccines for mounting immunity that is long-lasting against SARS-CoV-2 infection.

GLP-1 receptor agonists alleviate colonic inflammation by modulating intestinal microbiota and the function of group 3 innate lymphoid cells.

Glucagon-like peptide-1 receptor agonists (GLP-1RAs), which are drugs used for treating type 2 diabetes, have been reported to exert anti-inflammatory effects on inflammatory bowel disease (IBD), the mechanism of which remains elusive. Here, we report that GLP-1RAs ameliorate dextran sulfate sodium (DSS)-induced colitis in both wild-type and T/B-cell-deficient mice through modulating group 3 innate lymphoid cells (ILC3s), a subset of innate lymphoid cells that regulate intestinal immunity. GLP-1RAs promote IL-22 production by ILC3, and the protective effect of GLP-1RAs on DSS-induced colitis was abrogated in ILC3-deficient RORgtgfp/gfp mice. Furthermore, the treatment effect of GLP-RAs on colitis, as well as the generation of IL-22-producing ILC3s by GLP-RAs, is dependent on the gut microbiota. GLP-1RAs increase the abundance of Firmicutes and Proteobacteria in the gut, particularly beneficial bacteria such as Lactobacillus reuteri, and decrease the abundance of enteropathogenic Staphylococcus bacteria. The untargeted gas chromatography (GC)/liquid chromatography (LC)-mass spectrometry (MS) of faecal metabolites further revealed enrichment of N,N-dimethylsphingosine (DMS), an endogenous metabolite derived from sphingosine, in the GLP-1RA-treated group. Strikingly, DMS ameliorates colitis while promoting intestinal IL-22-producing ILC3s. Taken together, our findings show that GLP-1RAs exert a therapeutic effect on colitis possibly by regulating the microbiota-DMS-IL-22+ILC3 axis, highlighting the potential beneficial role of GLP-RAs in inflammatory intestinal disorders with diabetes complications.

Development of a high-throughput screening platform to identify new therapeutic agents for Medulloblastoma Group 3.

Pediatric brain tumors (PBTs) represent about 25 % of all pediatric cancers and are the most common solid tumors in children and adolescents. Medulloblastoma (MB) is the most frequently occurring malignant PBT, accounting for almost 10 % of all pediatric cancer deaths. MB Group 3 (MB G3) accounts for 25-30 % of all MB cases and has the worst outcome, particularly when associated with MYC amplification. However, no targeted treatments for this group have been developed so far. Here we describe a unique high throughput screening (HTS) platform specifically designed to identify new therapies for MB G3. The platform incorporates optimized and validated 2D and 3D efficacy and toxicity models, that account for tumor heterogenicity, limited efficacy and unacceptable toxicity from the very early stage of drug discovery. The platform has been validated by conducting a pilot HTS campaign with a 1280 lead-like compound library. Results showed 8 active compounds, targeting MB reported targets and several are currently approved or in clinical trials for pediatric patients with PBTs, including MB. Moreover, hits were combined to avoid tumor resistance, identifying 3 synergistic pairs, one of which is currently under clinical study for recurrent MB and other PBTs.

Intranasal B5 promotes mucosal defence against Actinobacillus pleuropneumoniae via ameliorating early immunosuppression.

Actinobacillus pleuropneumoniae (APP) is an important pathogen of the porcine respiratory disease complex, which leads to huge economic losses worldwide. We previously demonstrated that Pichia pastoris-producing bovine neutrophil ß-defensin-5 (B5) could resist the infection by the bovine intracellular pathogen Mycobacterium bovis. In this study, the roles of synthetic B5 in regulating mucosal innate immune response and protecting against extracellular APP infection were further investigated using a mouse model. Results showed that B5 promoted the production of tumour necrosis factor (TNF)-α, interleukin (IL)-1ß, and interferon (IFN)-ß in macrophages as well as dendritic cells (DC) and enhanced DC maturation in vitro. Importantly, intranasal B5 was safe and conferred effective protection against APP via reducing the bacterial load in lungs and alleviating pulmonary inflammatory damage. Furthermore, in the early stage of APP infection, we found that intranasal B5 up-regulated the secretion of TNF-α, IL-1ß, IL-17, and IL-22; enhanced the rapid recruitment of macrophages, neutrophils, and DC; and facilitated the generation of group 3 innate lymphoid cells in lungs. In addition, B5 activated signalling pathways associated with cellular response to IFN-ß and activation of innate immune response in APP-challenged lungs. Collectively, B5 via the intranasal route can effectively ameliorate the immune suppression caused by early APP infection and provide protection against APP. The immunization strategy may be applied to animals or human respiratory bacterial infectious diseases. Our findings highlight the potential importance of B5, enhancing mucosal defence against intracellular bacteria like APP which causes early-phase immune suppression.

Gut microbiota-derived 12-ketolithocholic acid suppresses the IL-17A secretion from colonic group 3 innate lymphoid cells to prevent the acute exacerbation of ulcerative colitis.

Intestinal microbiota dysbiosis and metabolic disruption are well-known as the primary triggers of ulcerative colitis (UC). However, their role in regulating the group 3 innate lymphoid cells (ILC3s), which are essential for intestinal health, remains unexplored during the development of disease severity. Here, our results showed that the microbiota structure of patients with severe UC (SUCs) differed from those with mild UC (MiUCs), moderate UC (MoUCs), and healthy controls (HCs). Microbes producing secondary bile acids (SBAs) and SBAs decreased with the aggravation of UC, and a strong positive correlation existed between them. Next, fecal microbiota transfer was used to reproduce the human-derived microbiota in mice and decipher the microbiota-mediated inflammatory modulation during an increase in disease severity. Mice receiving SUC-derived microbiota exhibited enhancive inflammation, a lowered percentage of ILC3s, and the down-regulated expressions of bile acid receptors, including vitamin D receptor (VDR) and pregnane X receptor (PXR), in the colon. Similar to clinical results, SBA-producing microbes, deoxycholic acids (DCA), and 12-ketolithocholic acids (12-KLCA) were diminished in the intestine of these recipients. Finally, we compared the therapeutic potential of DCA and 12-KLCA in preventing colitis and the regulatory mechanisms mediated by ILC3s. 12-KLCA but not DCA represented a strong anti-inflammatory effect associated with the higher expression of VDR and the lower secretion of IL-17A from colonic ILC3s. Collectively, these findings provide new signatures for monitoring the acute deterioration of UC by targeting gut microbiota and bile acid metabolism and demonstrate the therapeutic and preventive potential of a novel microbiota-derived metabolite, 12-KLCA.

MYC overexpression and SMARCA4 loss cooperate to drive medulloblastoma formation in mice.

Group 3 medulloblastoma is one of the most aggressive types of childhood brain tumors. Roughly 30% of cases carry genetic alterations in MYC, SMARCA4, or both genes combined. While overexpression of MYC has previously been shown to drive medulloblastoma formation in mice, the functional significance of SMARCA4 mutations and their suitability as a therapeutic target remain largely unclear. To address this issue, we combined overexpression of MYC with a loss of SMARCA4 in granule cell precursors. Both alterations did not increase proliferation of granule cell precursors in vitro. However, combined MYC overexpression and SMARCA4 loss successfully induced tumor formation in vivo after orthotopic transplantation in recipient mice. Resulting tumors displayed anaplastic histology and exclusively consisted of SMARCA4-negative cells although a mixture of recombined and non-recombined cells was injected. These observations provide first evidence for a tumor-promoting role of a SMARCA4 deficiency in the development of medulloblastoma. In comparing the transcriptome of tumors to the cells of origin and an established Sonic Hedgehog medulloblastoma model, we gathered first hints on deregulated gene expression that could be specifically involved in SMARCA4/MYC driven tumorigenesis. Finally, an integration of RNA sequencing and DNA methylation data of murine tumors with human samples revealed a high resemblance to human Group 3 medulloblastoma on the molecular level. Altogether, the development of SMARCA4-deficient medulloblastomas in mice paves the way to deciphering the role of frequently occurring SMARCA4 alterations in Group 3 medulloblastoma with the perspective to explore targeted therapeutic options.

Prognostic significance of molecular subgroups in survival outcome for children with medulloblastoma in Malaysia.

Introduction: Advancements in genomic profiling led to the discovery of four major molecular subgroups in medulloblastoma (MB), which have now been incorporated into the World Health Organization classification of central nervous system tumors. The current study aimed to determine the prognostic significance of the MB molecular subgroups among children in Malaysia. Methods: We assembled MB samples from children <18 years between January 2003 and June 2017 from four pediatric oncology centers in Malaysia. MB was sub-grouped using 850k DNA methylation testing at German Cancer Research Centre, Heidelberg, Germany. Results: Fifty samples from patients diagnosed and treated as MB were identified. Two (4%) of the 50 patients' tumor DNA samples were insufficient for analysis. Of the remaining 48 patients, 41 (85%) samples were confirmed as MB, while for 7 (15%) patients, DNA methylation classification results were discrepant with the histopathological diagnosis of MB, with various other diagnoses. Of the 41 MB patients, 15 patients were stratified as standard-risk (SR), 16 patients as high-risk (HR), and ten as infants (age <3 years old). Molecular subgrouping of the whole cohort revealed four (14%) WNT, 11 (27%) SHH, 10 (24%) Group 3, and 16 (39%) Group 4. Treatment abandonment rates for older children and infants were 22.5% and 10%, respectively. After censoring treatment abandonment, for SR patients, the 5-year event-free survival (EFS) and overall survival (OS) were 43.1% ± 14.7% and 46.9 ± 15.6%, respectively, while in HR, 5-year EFS and OS were both 63.6% ± 14.5%. Infants had a 5-year EFS and OS of 55.6% ± 16.6% and 66.7% ± 15.7%, respectively. WNT tumors had the best 5y-OS, followed by Group 3, Group 4, and SHH in children ≥3 years old. In younger children, SHH MB patients showed favorable outcomes. Conclusion: The study highlights the importance of DNA methylation profiling for diagnostic accuracy. Most infants had SHH MB, and their EFS and OS were comparable to those reported in high-income countries. Due to the relatively small cohort and the high treatment abandonment rate, definite conclusions cannot be made regarding the prognostic significance of molecular subgroups of MB. Implementing this high-technology investigation would assist pathologists in improving the diagnosis and provide molecular subgrouping of MB, permitting subgroup-specific therapies.

Group 3 Innate Lymphoid Cells Exacerbate Lupus Nephritis by Promoting B Cell Activation in Kidney Ectopic Lymphoid Structures.

Group 3 innate lymphoid cells (ILC3s) represent a new population in immune regulation, yet their role in lupus nephritis (LN) remains elusive. In the present work, systemic increases in ILC3s, particularly in the kidney, are observed to correlate strongly with disease severity in both human and murine LN. Using MRL/lpr lupus mice and a nephrotoxic serum-induced LN model, this study demonstrates that ILC3s accumulated in the kidney migrate predominantly from the intestine. Furthermore, intestinal ILC3s accelerate LN progression, manifested by exacerbated autoimmunity and kidney injuries. In LN kidneys, ILC3s are located adjacent to B cells within ectopic lymphoid structures (ELS), directly activating B cell differentiation into plasma cells and antibody production in a Delta-like1 (DLL1)/Notch-dependent manner. Blocking DLL1 attenuates ILC3s' effects and protects against LN. Altogether, these findings reveal a novel pathogenic role of ILC3s in B cell activation, renal ELS formation and autoimmune injuries during LN, shedding light on the therapeutic value of targeting ILC3s for LN.

IL-18-induced HIF-1α in ILC3s ameliorates the inflammation of C. rodentium-induced colitis.

Group 3 innate lymphoid cells (ILC3s) are vital for defending tissue barriers from invading pathogens. Hypoxia influences the production of intestinal ILC3-derived cytokines by activating HIF. Yet, the mechanisms governing HIF-1α in ILC3s and other innate RORγt+ cells during in vivo infections are poorly understood. In our study, transgenic mice with specific Hif-1a gene inactivation in innate RORγt+ cells (RAG1KO HIF-1α▵Rorc) exhibit more severe colitis following Citrobacter rodentium infection, primarily due to the inability to upregulate IL-22. We find that HIF-1α▵Rorc mice have impaired IL-22 production in ILC3s, while non-ILC3 innate RORγt+ cells, also capable of producing IL-22, remain unaffected. Furthermore, we show that IL-18, induced by Toll-like receptor 2, selectively triggers IL-22 in ILC3s by transcriptionally upregulating HIF-1α, revealing an oxygen-independent regulatory pathway. Our results highlight that, during late-stage C. rodentium infection, IL-18 induction in the colon promotes IL-22 through HIF-1α in ILC3s, which is crucial for protection against this pathogen.

Pseudomonas aeruginosa strains belonging to phylogroup 3 frequently exhibit an atypical quorum sensing response: the case of MAZ105, a tomato rhizosphere isolate.

Pseudomonas aeruginosa is a widespread γ-proteobacterium and an important opportunistic pathogen. The genetically diverse P. aeruginosa phylogroup 3 strains are characterized by producing the pore-forming ExlA toxin and by their lack of a type III secretion system. However, like all strains of this species, they produce several virulence-associated traits, such as elastase, rhamnolipids and pyocyanin, which are regulated by quorum sensing (QS). The P. aeruginosa QS response comprises three systems (Las, Rhl and Pqs, respectively) that hierarchically regulate these virulence factors. The Pqs QS system is composed of the PqsR transcriptional factor, which, coupled with the alkyl-quinolones HHQ or PQS, activates the transcription of the pqsABCDE operon. The products of the first four genes of this operon produce HHQ, which is then converted to PQS by PqsH, while PqsE forms a complex with RhlR and stabilizes it. In this study we report that mutations affecting the Pqs system are particularly common in phylogroup 3 strains. To better understand QS in phylogroup 3 strains we studied strain MAZ105 isolated from tomato rhizosphere and showed that it contains mutations in the central QS transcriptional regulator, LasR, and in the gene encoding the PqsA enzyme involved in the synthesis of PQS. However, it can still produce QS-regulated virulence factors and is virulent in Galleria mellonella and mildly pathogenic in the mouse abscess/necrosis model; our results show that this may be due to the expression of pqsE from a different PqsR-independent promoter than the pqsA promoter. Our results indicate that using anti-virulence therapy based on targeting the PQS system will not be effective against infections by P. aeruginosa phylogroup 3 strains.

Inhaled therapies targeting prostacyclin pathway in pulmonary hypertension due to COPD: systematic review.

Background: Pulmonary hypertension due to chronic obstructive pulmonary disease (COPD) and interstitial lung disease (ILD) is classified as group 3 pulmonary hypertension. Inhaled treprostinil, a prostaglandin I2 analogue also known as prostacyclin, has recently been approved as a first drug for patients with pulmonary hypertension secondary to ILD. However, due to a lack of evidence, no therapies are currently approved for those with COPD-associated pulmonary hypertension. Thus, this systematic review aims to summarise the current evidence to assess the impact of inhaled prostaglandin I2 analogue use on the pulmonary hemodynamics, exercise function, lung function, and gas exchange in patients with pulmonary hypertension due to COPD. Methods: We systematically searched the electronic databases of Medline, Embase, Scopus and Cochrane from inception to 1 February 2023. Studies of adult patients with a confirmed diagnosis of COPD-associated pulmonary hypertension who received inhaled drugs targeting the prostacyclin pathway were included in the systematic review. Case reports, systematic reviews, conference abstracts with no full text, non-full-text articles, non-English manuscripts and book chapters were excluded from this systematic review. A risk-of-bias assessment was carried out for the studies included in this review, using two different Cochrane risk-of-bias tools for randomised and non-randomised clinical trials. Results: A total of four studies met our inclusion criteria and were included in this systematic review. The results of one prospective clinical trial showed an improvement in the pulmonary hemodynamics (e.g., cardiac index, cardiac output and mean pulmonary artery pressure) in response to inhaled prostacyclin use in patients with pulmonary hypertension secondary to COPD. However, the severity of dyspnoea, lung function, exercise capacity and gas exchange were not affected when inhaled prostacyclin was used for patients with COPD-related pulmonary hypertension. Conclusion: This systematic review demonstrated that although inhaled prostacyclin does not seem to improve COPD-related outcomes (e.g., lung function and exercise capacity), short-term use of inhaled prostacyclin has the potential to reduce mean pulmonary artery pressure and pulmonary vascular resistance without impairing ventilation-perfusion mismatch. Further studies with larger sample sizes are warranted. Systematic review registration: CRD42022372803, https://www.crd.york.ac.uk/prospero/display_record.php?RecordID=372803.