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Background: There is an increasing interest in targeted application of probiotic bacteria for prevention and treatment of airway diseases, including allergies. Here, we investigated the beneficial effects of preventive intranasal treatment with probiotics Lactobacillus rhamnosus GG and L. rhamnosus GR‐1 in a mouse model of allergic asthma. Methods: Lactobacillus rhamnosus was administered intranasally eight times on days 1‐4 and 8‐11 at 5 × 108 CFU/dose, followed by a 2‐week asthma induction protocol with birch pollen extract on alternating days. Effects of preventive treatment were analyzed based on serum antibody levels, bronchoalveolar lavage cell counts, lung histology, lung cytokine levels, and airway hyperreactivity. Colonization and translocation of L. rhamnosus were assessed by bacterial cell counts in nasal mucosa, fecal samples, cervical lymph nodes, and blood. Binding of fluorescent L. rhamnosus to fixed murine nasal mucosal cells and airway macrophages was visualized by fluorescence microscopy. Results: Transient colonization of the murine upper airways by L. rhamnosus GG was demonstrated and was approximately ten times higher compared to L. rhamnosus GR‐1. Marked binding of fluorescent L. rhamnosus GG to murine nasal mucosal cells and airway macrophages was visualized. Preventive treatment with L. rhamnosus GG (but not L. rhamnosus GR‐1) resulted in a significant decrease in bronchoalveolar lavage eosinophil counts, lung interleukin‐13 and interleukin‐5 levels, and airway hyperreactivity. A tendency toward a decrease in serum Bet v 1‐specific immunoglobulin G1 was likewise observed. Conclusion: Intranasally administered L. rhamnosus GG prevents the development of cardinal features of birch pollen‐induced allergic asthma in a strain‐specific manner. Intranasal Lactobacillus rhamnosus GG (but not L. rhamnosus GR‐1) pretreatment prevents the development of airway inflammation and hyperreactivity in a mouse model of birch pollen‐induced asthma, without observable side effects. Effects of intranasally administered L. rhamnosus GG are linked to a significant decrease in lung eosinophilia and lung IL‐13 and IL‐5 levels. L. rhamnosus GG adheres well to isolated murine nasal mucosa cells and airway macrophages, and can transiently colonize the murine upper airways. This is less pronounced for L. rhamnosus GR‐1. [ABSTRACT FROM AUTHOR]

Despite the broad use of single-cell/nucleus RNA sequencing in plant research, accurate cluster annotation in less-studied plant species remains a major challenge due to the lack of validated marker genes. Here, we generated a single-cell RNA sequencing atlas of soil-grown wheat roots and annotated cluster identities by transferring annotations from publicly available datasets in wheat, rice, maize, and Arabidopsis. The predictions from our orthology-based annotation approach were next validated using untargeted spatial transcriptomics. These results allowed us to predict evolutionarily conserved tissue-specific markers and generate cell type-specific gene regulatory networks for root tissues of wheat and the other species used in our analysis. In summary, we generated a single-cell and spatial transcriptomics resource for wheat root apical meristems, including numerous known and uncharacterized cell type-specific marker genes and developmental regulators. These data and analyses will facilitate future cell type annotation in non-model plant species., Competing Interests: Declaration of interests The authors declare no competing interests., (Copyright © 2025 The Author(s). Published by Elsevier Inc. All rights reserved.)

Spatially resolved omics technologies are transforming our understanding of biological tissues. However, the handling of uni- and multimodal spatial omics datasets remains a challenge owing to large data volumes, heterogeneity of data types and the lack of flexible, spatially aware data structures. Here we introduce SpatialData, a framework that establishes a unified and extensible multiplatform file-format, lazy representation of larger-than-memory data, transformations and alignment to common coordinate systems. SpatialData facilitates spatial annotations and cross-modal aggregation and analysis, the utility of which is illustrated in the context of multiple vignettes, including integrative analysis on a multimodal Xenium and Visium breast cancer study., Competing Interests: Competing interests: J.M. holds equity in Glencoe Software, which builds products based on OME–NGFF. F.J.T. consults for Immunai, Inc., Singularity Bio B.V., CytoReason Ltd, Cellarity and Omniscope and has ownership interest in Dermagnostix GmbH and Cellarity. O.S. is a paid consultant of Insitro, Inc., S.K.S. is a consulting scientific cofounder for Digital Biology, Inc. The remaining authors declare no competing interests., (© 2024. The Author(s).)

The small intestinal crypts harbor secretory Paneth cells (PCs) which express bactericidal peptides that are crucial for maintaining intestinal homeostasis. Considering the diverse environmental conditions throughout the course of the small intestine, multiple subtypes of PCs are expected to exist. We applied single-cell RNA-sequencing of PCs combined with deep bulk RNA-sequencing on PC populations of different small intestinal locations and discovered several expression-based PC clusters. Some of these are discrete and resemble tuft cell-like PCs, goblet cell (GC)-like PCs, PCs expressing stem cell markers, and atypical PCs. Other clusters are less discrete but appear to be derived from different locations along the intestinal tract and have environment-dictated functions such as food digestion and antimicrobial peptide production. A comprehensive spatial analysis using Resolve Bioscience was conducted, leading to the identification of different PC's transcriptomic identities along the different compartments of the intestine, but not between PCs in the crypts themselves.

To better understand intrinsic resistance to immune checkpoint blockade (ICB), we established a comprehensive view of the cellular architecture of the treatment-naive melanoma ecosystem and studied its evolution under ICB. Using single-cell, spatial multi-omics, we showed that the tumor microenvironment promotes the emergence of a complex melanoma transcriptomic landscape. Melanoma cells harboring a mesenchymal-like (MES) state, a population known to confer resistance to targeted therapy, were significantly enriched in early on-treatment biopsies from non-responders to ICB. TCF4 serves as the hub of this landscape by being a master regulator of the MES signature and a suppressor of the melanocytic and antigen presentation transcriptional programs. Targeting TCF4 genetically or pharmacologically, using a bromodomain inhibitor, increased immunogenicity and sensitivity of MES cells to ICB and targeted therapy. We thereby uncovered a TCF4-dependent regulatory network that orchestrates multiple transcriptional programs and contributes to resistance to both targeted therapy and ICB in melanoma., Competing Interests: Declaration of interests J.-C.M., F.R., J.P., and D.P. are authors on a patent application related to this work., (Copyright © 2023 The Authors. Published by Elsevier Inc. All rights reserved.)

Purpose: Exposure to low concentrations of toluene diisocyanate (TDI) leads to immune-mediated chemical-induced asthma. The role of the adaptive immune system has already been thoroughly investigated; nevertheless, the involvement of innate immune cells in the pathophysiology of chemical-induced asthma is still unresolved. The aim of the study is to investigate the role of innate lymphoid cells (ILCs) and dendritic cells (DCs) in a mouse model for chemical-induced asthma., Methods: On days 1 and 8, BALB/c mice were dermally treated (20 μL/ear) with 0.5% TDI or the vehicle acetone olive oil (AOO; 2:3). On days 15, 17, 19, 22 and 24, the mice received an oropharyngeal challenge with 0.01% TDI or AOO (1:4). One day after the last challenge, airway hyperreactivity (AHR) to methacholine was assessed, followed by an evaluation of pulmonary inflammation and immune-related parameters, including the cytokine pattern in bronchoalveolar lavage fluid, lymphocyte subpopulations of the lymph nodes and their ex vivo cytokine production profile, blood immunoglobulins and DC and ILC subpopulations in the lungs., Results: Both DC and ILC2 were recruited to the lungs after multiple airway exposures to TDI, regardless of the prior dermal sensitization. However, prior dermal sensitization with TDI alone results in AHR and predominant eosinophilic airway inflammation, accompanied by a typical type 2 helper T (Th2) cytokine profile., Conclusions: TDI-induced asthma is mediated by a predominant type 2 immune response, with the involvement of adaptive Th2 cells. However, from our study we suggest that the innate ILC2 cells are important additional players in the development of TDI-induced asthma., Competing Interests: There are no financial or other issues that might lead to conflict of interest., (Copyright © 2021 The Korean Academy of Asthma, Allergy and Clinical Immunology · The Korean Academy of Pediatric Allergy and Respiratory Disease.)

Background: Occupational asthma, induced by workplace exposures to low molecular weight agents such as toluene 2,4-diisocyanate (TDI), causes a significant burden to patients and society. Little is known about innate lymphoid cells (ILCs) in TDI-induced asthma. A critical regulator of ILC function is microRNA-155, a microRNA associated with asthma., Objective: To determine whether TDI exposure modifies the number of ILCs in the lung and whether microRNA-155 contributes to TDI-induced airway inflammation and hyperresponsiveness., Methods: C57BL/6 wild-type and microRNA-155 knockout mice were sensitised and challenged with TDI or vehicle. Intracellular cytokine expression in ILCs and T-cells was evaluated in bronchoalveolar lavage (BAL) fluid using flow cytometry. Peribronchial eosinophilia and goblet cells were evaluated on lung tissue, and airway hyperresponsiveness was measured using the forced oscillation technique. Putative type 2 ILCs (ILC2) were identified in bronchial biopsies of subjects with TDI-induced occupational asthma using immunohistochemistry. Human bronchial epithelial cells were exposed to TDI or vehicle., Results: TDI-exposed mice had higher numbers of airway goblet cells, BAL eosinophils, CD4 + T-cells and ILCs, with a predominant type 2 response, and tended to have airway hyperresponsiveness. In TDI-exposed microRNA-155 knockout mice, inflammation and airway hyperresponsiveness were attenuated. TDI exposure induced IL-33 expression in human bronchial epithelial cells and in murine lungs, which was microRNA-155 dependent in mice. GATA3 + CD3 - cells, presumably ILC2, were present in bronchial biopsies., Conclusion: TDI exposure is associated with increased numbers of ILCs. The proinflammatory microRNA-155 is crucial in a murine model of TDI asthma, suggesting its involvement in the pathogenesis of occupational asthma due to low molecular weight agents., Competing Interests: Conflict of interest: E.E. Blomme reports grants from Fund for Scientific Research in Flanders (projects G053516N and G0G2318N) and Ghent University (BOF19-GOA-008), during the conduct of the study. Conflict of interest: S. Provoost has nothing to disclose. Conflict of interest: E. Bazzan has nothing to disclose. Conflict of interest: H.P. Van Eeckhoutte has nothing to disclose. Conflict of interest: M.P. Roffel has nothing to disclose. Conflict of interest: L. Pollaris has nothing to disclose. Conflict of interest: A. Bontinck has nothing to disclose. Conflict of interest: M. Bonato has nothing to disclose. Conflict of interest: L. Vandenbroucke has nothing to disclose. Conflict of interest: F. Verhamme has nothing to disclose. Conflict of interest: G.F. Joos reports grants and personal fees from AstraZeneca and GlaxoSmithKline, personal fees from Bayer, Eureca and Teva, grants from Chiesi, outside the submitted work; all payments were made to his employer. Conflict of interest: M.G. Cosio has nothing to disclose. Conflict of interest: J.A.J. Vanoirbeek has nothing to disclose. Conflict of interest: G.G. Brusselle has nothing to disclose. Conflict of interest: M. Saetta reports grants from Laboratori Guidotti SpA and Chiesi Farmaceutici SpA, outside the submitted work. Conflict of interest: T. Maes reports grants from Ghent University (concerted research action BOF-GOA-008) and Fund for Scientific Research in Flanders (projects G053516N and G0G2318N), during the conduct of the study; personal fees for advisory board work from GlaxoSmithKline, outside the submitted work; and is shareholder of Oryzon Genomics and Mendelion Lifesciences SL., (Copyright ©ERS 2020.)

Background: Exposure to diisocyanates is an important cause of occupational asthma (OA) in the industrialized world. Since OA occurs after long-term exposure to diisocyanates, we developed a chronic mouse model of chemical-induced asthma where toluene diisocyanate (TDI) was administered at two different exposure sites., Objectives: Evaluating the effect of long-term respiratory isocyanate exposure - with or without prior dermal exposure- on sensitization, inflammatory responses and airway hyperreactivity (AHR)., Methods: On days 1 and 8, BALB/c mice were dermally treated (20 μl/ear) with 0.5% 2,4-toluene diisocyanate TDI or the vehicle acetone olive oil (AOO) (3:2). Starting from day 15, mice received intranasal instillations with 0.1% TDI of vehicle five times in a week, for five successive weeks. One day after the last instillation airway hyperreactivity (AHR) to methacholine was assessed, followed by an evaluation of pulmonary inflammation and structural lung changes. Immune-related parameters were assessed in the lungs (BAL and tissue), blood, cervical- and auricular lymph nodes., Results: Mice repeatedly intranasally exposed to TDI showed systemic sensitization and a mixed Th1/Th2 type immune response, without the presence of AHR. However, when mice are first dermally sensitized with TDI, followed by repeated intranasal TDI challenges, this results in a pronounced Th2 response and AHR., Conclusion: Dermal exposure to TDI determines airway hyperreactivity after repeated airway exposure to TDI., (Copyright © 2019 Elsevier B.V. All rights reserved.)

Inhalation of commonly present irritants, such as chlorine and chlorine derivatives, can cause adverse respiratory effects, including irritant-induced asthma (IIA). We hypothesize that due to airway barrier impairment, exposure to hypochlorite (ClO - ) can result in airway hypersensitivity. C57Bl/6 mice received an intra-peritoneal (i.p.) injection of the airway damaging agent naphthalene (NA, 200 mg/kg body weight) or vehicle (mineral oil, MO). In vivo micro-computed tomography (CT) images of the lungs were acquired before and at regular time points after the i.p., Treatment: After a recovery period of 14 days an intranasal (i.n.) challenge with 0.003% active chlorine (in ClO - ) or vehicle (distilled water, H 2 O) was given, followed by assessment of the breathing frequency. One day later, pulmonary function, along with pulmonary inflammation was determined. Lung permeability was assessed by means of total broncho-alveolar lavage (BAL) protein content and plasma surfactant protein (SP)-D levels. In vivo micro-CT imaging revealed enlargement of the lungs and airways early after NA treatment, with a return to normal at day 14. When challenged i.n. with ClO - , NA-pretreated mice immediately responded with a sensory irritant response. Twenty-four hours later, NA/ClO - mice showed airway hyperreactivity (AHR), accompanied by a neutrophilic and eosinophilic inflammation. NA administration followed by ClO - induced airway barrier impairment, as shown by increased BAL protein and plasma SP-D concentrations; histology revealed epithelial denudation. These data prove that NA-induced lung impairment renders the lungs of mice more sensitive to an airway challenge with ClO - , confirming the hypothesis that incomplete barrier repair, followed by irritant exposure results in airway hypersensitivity.

Background: While the importance of the Th2 cytokine IL-13 as a central mediator of airway hyperreactivity (AHR) has been described in allergic protein-induced asthma, this has never been investigated in chemical-induced asthma., Objective: We examined the importance of IL-13 in a mouse model of chemical-induced AHR, using toluene-2,4-diisocyanate (TDI)., Methods: In a first set-up, wild type (WT) and IL-13 knockout (KO) C57Bl/6 mice were dermally treated on days 1 and 8 with 1% TDI or vehicle (acetone/olive oil) on both ears. On day 15, mice received an intranasal instillation with 0.1% TDI or vehicle. In a second set-up, WT mice sensitized with 1% TDI or vehicle, received i.v. either anti-IL-13 or control antibody prior to the intranasal challenge., Results: TDI-sensitized and TDI-challenged WT mice showed AHR to methacholine, in contrast to TDI-sensitized and TDI-challenged IL-13 KO mice, which also showed lower levels of total serum IgE. TDI-sensitized and TDI-challenged IL-13 KO mice had lower numbers of T-cells in the auricular lymph nodes. TDI-treated WT mice, receiving anti-IL-13, showed no AHR, in contrast to those receiving control antibody, despite increased levels of IgE. Anti-IL-13 treatment in TDI-treated WT mice resulted in lower levels of serum IL-13, but did not induce changes in T- and B-cell numbers, and in the cytokine production profile., Conclusion and Clinical Relevance: We conclude that IL-13 plays a critical role in the effector phase of chemical-induced, immune-mediated AHR. This implicates that anti-IL-13 treatment could have a beneficial effect in patients with this asthma phenotype.

Background: Pulmonary function measurements are important when studying respiratory disease models. Both resistance and compliance have been used to assess lung function in mice. Yet, it is not always clear how these parameters relate to forced expiration (FE)-related parameters, most commonly used in humans. We aimed to characterize FE measurements in four well-established mouse models of lung diseases., Method: Detailed respiratory mechanics and FE measurements were assessed concurrently in Balb/c mice, using the forced oscillation and negative pressure-driven forced expiration techniques, respectively. Measurements were performed at baseline and following increasing methacholine challenges in control Balb/c mice as well as in four disease models: bleomycin-induced fibrosis, elastase-induced emphysema, LPS-induced acute lung injury and house dust mite-induced asthma., Results: Respiratory mechanics parameters (airway resistance, tissue damping and tissue elastance) confirmed disease-specific phenotypes either at baseline or following methacholine challenge. Similarly, lung function defects could be detected in each disease model by at least one FE-related parameter (FEV 0.1 , FEF 0.1 , FVC, FEV 0.1 /FVC ratio and PEF) at baseline or during the methacholine provocation assay., Conclusions: FE-derived outcomes in four mouse disease models behaved similarly to changes found in human spirometry. Routine combined lung function assessments could increase the translational utility of mouse models.

Asthma may be induced by chemical sensitisers, via mechanisms that are still poorly understood. This type of asthma is characterised by airway hyperreactivity (AHR) and little airway inflammation. Since potent chemical sensitisers, such as toluene-2,4-diisocyanate (TDI), are also sensory irritants, it is suggested that chemical-induced asthma relies on neuro-immune mechanisms.We investigated the involvement of transient receptor potential channels (TRP) A1 and V1, major chemosensors in the airways, and mast cells, known for their ability to communicate with sensory nerves, in chemical-induced AHR.In vitro intracellular calcium imaging and patch-clamp recordings in TRPA1- and TRPV1-expressing Chinese hamster ovarian cells showed that TDI activates murine TRPA1, but not TRPV1. Using an in vivo model, in which an airway challenge with TDI induces AHR in TDI-sensitised C57Bl/6 mice, we demonstrated that AHR does not develop, despite successful sensitisation, in Trpa1 and Trpv1 knockout mice, and wild-type mice pretreated with a TRPA1 blocker or a substance P receptor antagonist. TDI-induced AHR was also abolished in mast cell deficient Kit(Wsh) (/Wsh) mice, and in wild-type mice pretreated with the mast cell stabiliser ketotifen, without changes in immunological parameters.These data demonstrate that TRPA1, TRPV1 and mast cells play an indispensable role in the development of TDI-elicited AHR., (Copyright ©ERS 2016.)

Both 2,4-toluene diisocyanate (TDI) and 4,4-methylene diphenyl diisocyanate (MDI) can cause occupational asthma. In this study, we optimized our mouse model of chemical-induced asthma in the C57Bl/6 mice strain using the model agent TDI. Furthermore, we validated MDI in this mouse model and investigated whether cross-reactivity between TDI and MDI is present. On days 1 and 8, C57Bl/6 mice were dermally treated (20 µl/ear) with 3 % MDI, 2 % TDI or the vehicle acetone olive oil (AOO) (3:2). On day 15, they received a single oropharyngeal challenge with 0.04 % MDI, 0.01 % TDI or the vehicle AOO (4:1). One day later, airway hyperreactivity (AHR) and pulmonary inflammation in the bronchoalveolar lavage (BAL) were assessed. Furthermore, total serum IgE levels, lymphocyte subpopulations in auricular lymph nodes and cytokine levels in supernatants of lymphocytes were measured. Both dermal sensitization with TDI or MDI resulted in increased total serum IgE levels along with T and B cell proliferation in the auricular lymph nodes. The auricular lymphocytes showed an increased release of both Th2 and Th1 cytokines. Mice sensitized and challenged with either TDI or MDI showed AHR, along with a predominant neutrophil lung inflammation. Mice sensitized with MDI and challenged with TDI or the other way around showed no AHR, nor BAL inflammation. Both TDI and MDI are able to induce an asthma-like response in this mouse model. However, cross-reactivity between both diisocyanates remained absent.

Muscular dystrophies are a heterogeneous group of myopathies, characterized by muscle weakness and degeneration, without curative treatment. Mesoangioblasts (MABs) have been proposed as a potential regenerative therapy. To improve our understanding of the in vivo behavior of MABs and the effect of different immunosuppressive therapies, like cyclosporine A or co-stimulation-adhesion blockade therapy, on cell survival noninvasive cell monitoring is required. Therefore, cells were transduced with a lentiviral vector encoding firefly luciferase (Fluc) and the human sodium iodide transporter (hNIS) to allow cell monitoring via bioluminescence imaging (BLI) and small-animal positron emission tomography (PET). Non-H2 matched mMABs were injected in the femoral artery of dystrophic mice and were clearly visible via small-animal PET and BLI. Based on noninvasive imaging data, we were able to show that co-stim was clearly superior to CsA in reducing cell rejection and this was mediated via a reduction in cytotoxic T cells and upregulation of regulatory T cells., (Copyright © 2015 The Authors. Published by Elsevier Inc. All rights reserved.)

Methylisothiazolinone (MI), a widely used chemical preservative in industrial and household products, and cosmetics, has been associated with allergic contact dermatitis. However, the asthmogenic capacity of MI is currently unknown. In this study, we investigated the capacity of MI to elicit asthma-like responses in a validated mouse model. On days 1 and 8, mice (C57Bl/6 and BALB/c) were dermally treated with MI or vehicle on each ear. On day 15, mice received a single intranasal challenge with MI or vehicle. Immediately after the challenge, the early ventilatory response was measured using a double chamber plethysmograph. One day later, airway hyperreactivity, pulmonary inflammation and immune-related parameters were assessed. Dermal treatment with MI in both C57Bl/6 and BALB/c mice induced increased T- and B-cell proliferation in the auricular lymph nodes, along with IFN-γ production and limited increases in total serum IgE, confirming dermal sensitization. An airway challenge with MI led to an early ventilatory response (decreased breathing frequency), indicative for acute sensory irritation. However, 24h later no allergic respiratory response (no airway hyperreactivity (AHR) nor pulmonary inflammation) was found in either mouse strains. Our study indicates that MI can be classified as a strong dermal sensitizer and irritant, but not an asthmogen after initial dermal sensitization, followed by an airway challenge., (Copyright © 2015 Elsevier Ireland Ltd. All rights reserved.)

Aim: To evaluate the 1-year outcome of Minimally Invasive Single Implant Treatment (M.I.S.I.T.) based on ridge preservation and contour augmentation in patients with a high aesthetic risk profile., Materials and Methods: Periodontally healthy non-smoking patients with a failing tooth in the anterior maxilla (15-25) were selected. All were in need of a single implant and demonstrated high risk for aesthetic complications given mid-facial recession at the failing tooth and/or a buccal bone dehiscence and/or a thin-scalloped gingival biotype. Patients without mid-facial recession (NRG) received flapless tooth extraction and ridge preservation using a collagen-enriched bovine-derived xenograft, flapless installation of a bone condensing implant with variable-thread design (4-6 months later), a provisional screw-retained crown and connective tissue graft (CTG) inserted in the buccal mucosa (3 months later) and a permanent crown (3 months later). Patients with mid-facial recession (RG) were treated similarly, yet they received a CTG at the time of ridge preservation. All patients were treated by an experienced periodontist using a microsurgical approach. Primary outcome variables included papillary and mid-facial recession. Clinical parameters, pink (PES) and white aesthetic score (WES) were considered secondary outcome variables., Results: Fifty patients (25 females, 25 males; mean age 39, range 19-81; 42 in NRG and 8 in RG) met the selection criteria and consented to the treatment. Forty-seven could be examined at 12 months and all implants survived. Mean bone loss amounted to 0.48 mm (range 0.00-1.80) at 12 months. Papillary recession was minimal in both groups at 12 months (mean ≤ 0.3 mm), as was mid-facial recession in the NRG (mean 0.1 mm). Twelve patients in the NRG even demonstrated coronal migration of the mucosal margin following CTG and needed adaptation of the provisional crown to induce soft tissue retraction. Due to CTG at the time of ridge preservation in the RG, mid-facial soft tissue gain amounted to 0.9 mm at 12 months, hereby eliminating 2/3rd of the initial recession. PES and WES were favourable pointing to 10.9/14 and 8.2/10 respectively., Conclusion: This short-term prospective study offers a proof of principle of M.I.S.I.T. in patients with a high aesthetic risk profile., (© 2015 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd.)