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Frontiers in Immunology 2023Considering the similarities between swine and humans, it is a logical consequence to use swine as a translational model in research and drug development, including...
Considering the similarities between swine and humans, it is a logical consequence to use swine as a translational model in research and drug development, including non-clinical safety. Here, we compared the reactivity of peripheral blood mononuclear cells (PBMCs) from humans and minipigs under the influence of different compounds . We conducted a flow cytometry-based proliferation assay that focused on the T-cell response to three different stimuli: concanavalin A (ConA), phytohemagglutinin-L (PHA-L), and staphylococcal Enterotoxin B (SEB). Furthermore, four approved immunosuppressive drugs-abatacept, belatacept, rapamycin, and tofacitinib-which are used for the treatment of rheumatoid arthritis or rejection in transplant recipients, were combined with the different stimuli. This allowed us to study the effect of suppressive drugs in comparison with the different stimuli in both species. We examined proliferating T cells (CD3) and investigated the presence of TCR-αβ and TCR-γδ T cells. Differences in the response of T cells of the two species under these various conditions were evident. CD4 T cells were more activated within humans, whereas CD8 T cells were generally more abundant in swine. The effectiveness of the used humanized antibodies is most likely related to the conserved structure of CTLA-4 as abatacept induced a much stronger reduction in swine compared with belatacept. The reduction of proliferation of rapamycin and tofacitinib was highly dependent on the used stimuli. We further investigated the effect of the immunosuppressive compounds on antigen-specific restimulation of pigs immunized against porcine circovirus 2 (PCV2). Treatment with all four compounds resulted in a clear reduction of the proliferative response, with rapamycin showing the strongest effect. In conclusion, our findings indicate that the effectiveness of suppressive compounds is highly dependent on the stimuli used and must be carefully selected to ensure accurate results. The results highlight the importance of considering the response of T cells in different species when evaluating the potential of an immunomodulatory drug.
Topics: Humans; Swine; Animals; Swine, Miniature; Abatacept; Leukocytes, Mononuclear; CD8-Positive T-Lymphocytes; Immunosuppressive Agents; Sirolimus; Receptors, Antigen, T-Cell
PubMed: 38264655
DOI: 10.3389/fimmu.2023.1327776 -
Cureus Dec 2023In the last couple of decades, much progress has been made in studying bacteria living in humans. However, there is much more to learn about bacteria immune cell...
In the last couple of decades, much progress has been made in studying bacteria living in humans. However, there is much more to learn about bacteria immune cell interactions. Here, we show that anaerobic bacteria do not grow when cultured overnight with human cells under atmospheric air. Air contains about 18% oxygen, which inhibits the growth of these bacteria while supporting the cultivation of human cells. The bacteria cultured with human peripheral blood mononuclear cells (PBMCs) inflamed with phytohemagglutinin (PHA) greatly increased the production of proinflammatory cytokines like tumor necrosis factor-alpha (TNFα) while inhibiting the production of monocyte chemoattractant protein-1 (MCP-1), an important chemokine.
PubMed: 38222203
DOI: 10.7759/cureus.50586 -
Proteome Science Jan 2024Tepary bean (Phaseolus acutifolius A. Gray) is one of the five species domesticated from the genus Phaseolus with genetic resistance to biotic and abiotic stress. To...
BACKGROUND
Tepary bean (Phaseolus acutifolius A. Gray) is one of the five species domesticated from the genus Phaseolus with genetic resistance to biotic and abiotic stress. To understand the mechanisms underlying drought responses in seed storage proteins germinated on water and polyethylene glycol (PEG-6000) at -0.49 MPa, we used a proteomics approach to identify potential molecular target proteins associated with the low water potential stress response.
METHODS
Storage proteins from cotyledons of Tepary bean seeds germinated at 24, 48 and 72 h on water and PEG-6000 at -0.49 MPa were analyzed by one-dimensional electrophoresis (DE) with 2-DE analysis and shotgun mass spectrometry. Using computational database searching and bioinformatics analyses, we performed Gene Ontology (GO) and protein interactome (functional protein association network) String analyses.
RESULTS
Comparative analysis showed that the effect of PEG-6000 on root growth was parallel to that on germination. Based on the SDS‒PAGE protein banding patterns and 2-DE analysis, ten differentially abundant seed storage proteins showed changes in storage proteins, principally in the phaseolin and lectin fractions. We found many proteins that are recognized as drought stress-responsive proteins, and several of them are predicted to be intrinsically related to abiotic stress. The shotgun analysis searched against UniProt's legume database, and Gene Ontology (GO) analysis indicated that most of the seed proteins were cytosolic, with catalytic activity and associated with carbohydrate metabolism. The protein‒protein interaction networks from functional enrichment analysis showed that phytohemagglutinin interacts with proteins associated with the degradation of storage proteins in the cotyledons of common bean during germination.
CONCLUSION
These findings suggest that Tepary bean seed proteins provide valuable information with the potential to be used in genetic improvement and are part of the drought stress response, making our approach a potentially useful strategy for discovering novel drought-responsive proteins in other plant models.
PubMed: 38195472
DOI: 10.1186/s12953-023-00225-6 -
European Journal of Clinical... Mar 2024Impaired T-cell responses to mitogens and high T-cell activation marker (TAM) expression on Mycobacterium tuberculosis-specific T-cells characterize immunopathology in...
Impaired T-cell responses to mitogens and high T-cell activation marker (TAM) expression on Mycobacterium tuberculosis-specific T-cells characterize immunopathology in patients with tuberculosis (TB). In a study of patients with TB (n = 60) and asymptomatic contacts (controls, n = 37), we found that TB patients had higher CD38 T-cell proportions specific for M. tuberculosis protein (PPD), yet total proportions of PPD-specific T-cells were comparable. Notably, both activated (CD38) and total IFN-γ T-cells from TB patients had lower mitogen (phytohemagglutinin, PHA)-induced responses. This impaired mitogen response improved the classification efficacy of the TAM-TB assay, especially employing the PPD/PHA-induced T-cell ratio.
Topics: Humans; Mitogens; Tuberculin; Tuberculosis; Mycobacterium tuberculosis; T-Lymphocytes; Antigens, Bacterial
PubMed: 38167987
DOI: 10.1007/s10096-023-04741-3 -
Asia Pacific Allergy Dec 2023White bean allergy is uncommon and rarely reported. Herein, we report a case of white bean allergy in a patient with Down syndrome. A 7-year-old girl with Down syndrome...
White bean allergy is uncommon and rarely reported. Herein, we report a case of white bean allergy in a patient with Down syndrome. A 7-year-old girl with Down syndrome experienced allergic symptoms twice after eating white bean and visited our hospital for a food allergy investigation. An ImmunoCAP assay revealed a white bean-specific IgE (13.4 kU/L) in the patient's serum. In addition, her skin prick test result was positive. Moreover, ingestion of 2 g of boiled white beans in an oral food challenge test induced intermittent cough, desaturation, generalized urticaria, abnormal sleep, and mild hypotension. Thus, we diagnosed the patient with white bean allergy. We performed western blotting and mass spectrometric analysis and detected the following allergens: Phytohemagglutinin, group 3 late embryogenesis abundant protein, lipoxygenase, and legumin. In addition, we detected several candidate allergenic proteins for the first time. White bean, runner bean, or azuki bean was considered the primary source of sensitization because although immunoblotting inhibition tests revealed that the abovementioned beans inhibited other legumes, soybean, which she tolerates, showed little inhibition of the other legumes. However, we could not confirm whether the patient could ingest legumes other than soybean or white bean because her family did not wish to continue with further testing. This is the first report of a case of systemic allergic reactions to white bean in a child with Down syndrome. Further studies are needed to identify white bean allergens and understand the relationship between Down syndrome and white bean allergy.
PubMed: 38094097
DOI: 10.5415/apallergy.0000000000000111 -
Immunobiology Jan 2024Multipotent mesenchymal stromal cells (MSCs) have demonstrated a pronounced immunosuppressive activity, the manifestation of which depends on the microenvironmental...
Multipotent mesenchymal stromal cells (MSCs) have demonstrated a pronounced immunosuppressive activity, the manifestation of which depends on the microenvironmental factors, including O level. Here we examined the effects of MSCs on transcriptomic profile of allogeneic phytohemagglutinin-stimulated peripheral blood mononuclear cells (PBMCs) after interaction at ambient (20%) or "physiological" hypoxia (5%) O. As revealed with microarray analysis, PBMC transcriptome at 20% O was more affected, which was manifested as differential expression of more than 300 genes, whereas under 5% O 220 genes were changed. Most of genes at 20% O were downregulated, while at hypoxia most of genes were upregulated. Altered gene patterns were only partly overlapped at different O levels. A set of altered genes at hypoxia only was of particular interest. According to Gene Ontology a part of above genes was responsible for adhesion, cell communication, and immune response. At both oxygen concentrations, MSCs demonstrated effective immunosuppression manifested as attenuation of T cell activation and proliferation as well as anti-inflammatory shift of cytokine profile. Thus, MSC-mediated immunosuppression is executed with greater efficacy at a "physiological" hypoxia, since the same result has been achieved through a change in the expression of a fewer genes in target PBMCs.
Topics: Humans; Transcriptome; Leukocytes, Mononuclear; Mesenchymal Stem Cells; Cell Communication; Hypoxia; Cells, Cultured; Cell Proliferation
PubMed: 38091798
DOI: 10.1016/j.imbio.2023.152766 -
Conservation Physiology 2023Infectious diseases are a major driver of the global amphibian decline. In addition, many factors, including genetics, stress, pollution, and climate change can...
Infectious diseases are a major driver of the global amphibian decline. In addition, many factors, including genetics, stress, pollution, and climate change can influence the response to pathogens. Therefore, it is important to be able to evaluate amphibian immunity in the laboratory and in the field. The phytohemagglutinin (PHA) assay is an inexpensive and relatively non-invasive tool that has been used extensively to assess immunocompetence, especially in birds, and more recently in amphibians. However, there is substantial variation in experimental methodology among amphibian PHA studies in terms of species and life stages, PHA doses and injection sites, and use of experimental controls. Here, we compile and compare all known PHA studies in amphibians to identify knowledge gaps and develop best practices for future work. We found that research has only been conducted on a limited number of species, which may not reflect the diversity of amphibians. There is also a lack of validation studies in most species, so that doses and timing of PHA injection and subsequent swelling measurements may not effectively evaluate immunocompetence. Based on these and other findings, we put forward a set of recommendations to make future PHA studies more consistent and improve the ability to utilize this assay in wild populations, where immune surveillance is greatly needed.
PubMed: 38090122
DOI: 10.1093/conphys/coad090 -
International Journal of Pharmaceutics Jan 2024Immunological safety of nanofibers remains poorly reported within the scientific literature and lacks specific in vitro testing models distinct from those used to test...
Immunological safety of nanofibers remains poorly reported within the scientific literature and lacks specific in vitro testing models distinct from those used to test nanoparticles. To address the challenges of currently used conventional setups being described in the literature, we developed a novel in vitro model for nanofiber mats immunogenicity testing, which enables standardization of tested surface area, excludes nanofiber mat edges, and ensures stable contacts of cells with nanofibers during the experiment. The effect of nanofibers was assessed on peripheral blood mononuclear cells (PBMCs) by measuring their metabolic activity using MTS cell proliferation assay, where key performance parameters, i.e. cell number, phytohemagglutinin-L (PHA-L) concentration, incubation time and cell lysis were optimized. Repeatability of results obtained with non-activated and PHA-L-activated PBMCs in contact with differently thick polycaprolactone nanofiber mats was compared using both models. Our model provided more reproducible results with lower variability, exhibiting its higher reliability and accuracy than the conventional one. Furthermore, results showed the presence of thicker mats resulted in reduced metabolic activity and PBMC proliferation without any observed cytotoxicity, providing additional insights into their non-immunogenic characteristics. The developed model enables more accurate biological assessment that can support new guidelines for in vitro nanofiber testing and formulation.
Topics: Nanofibers; Leukocytes, Mononuclear; Reproducibility of Results; Nanoparticles
PubMed: 38086494
DOI: 10.1016/j.ijpharm.2023.123696 -
Fish & Shellfish Immunology Jan 2024Cell-mediated cytotoxicity is a complex immune mechanism that involves the release of several killing molecules, being perforin (PRF) one of the most important effector...
Cell-mediated cytotoxicity is a complex immune mechanism that involves the release of several killing molecules, being perforin (PRF) one of the most important effector players. Perforin is synthesized by T lymphocytes and natural killer cells in mammals and responsible for the formation of pores on the target cell membrane during the killing process. Although perforin has been extensively studied in higher vertebrates, this knowledge is very limited in fish. Therefore, in this study we have identified four prf genes in European sea bass (Dicentrarchus labrax) and evaluated their mRNA levels. All sea bass prf genes showed the typical and conserved domains of its human orthologue and were closely clustered by the phylogenetic analysis. In addition, all genes showed constitutive and ubiquitous tissular expression, being prf1.9 gene the most highly expressed in immune tissues. Subsequently, in vitro stimulation of head-kidney (HK) cells with phytohemagglutinin, a T-cell activator, showed an increase of all prf gene levels, except for prf1.3 gene. European sea bass HK cells increased the transcription of prf1.2 and prf1.9 during the innate cell-mediated cytotoxic activity against xenogeneic target cells. In addition, sea bass infected with nodavirus (NNV) showed a similar expression pattern of all prf in HK and brain at 15 days post-infection, except for prf1.3 gene and in the gonad. Finally, the use of a polyclonal antibody against PRF1.9 showed an increase of positive cells in HK, brain and gonad from NNV-infected fish. Taken together, the data seem to indicate that all prf genes, except prf1.3, appear to be involved in the European sea bass immunity, and probably in the cell-mediated cytotoxic response, with PRF1.9 playing the most important role against nodavirus. The involvement of the PRFs and the CMC activity in the vertical transmission success of the virus is also discussed.
Topics: Humans; Animals; Bass; Phylogeny; Perforin; Fish Diseases; Mammals
PubMed: 38000653
DOI: 10.1016/j.fsi.2023.109244 -
The Journal of Experimental Medicine Jan 2024Lymphocyte-specific protein tyrosine kinase (LCK) is essential for T cell antigen receptor (TCR)-mediated signal transduction. Here, we report two siblings homozygous...
Lymphocyte-specific protein tyrosine kinase (LCK) is essential for T cell antigen receptor (TCR)-mediated signal transduction. Here, we report two siblings homozygous for a novel LCK variant (c.1318C>T; P440S) characterized by T cell lymphopenia with skewed memory phenotype, infant-onset recurrent infections, failure to thrive, and protracted diarrhea. The patients' T cells show residual TCR signal transduction and proliferation following anti-CD3/CD28 and phytohemagglutinin (PHA) stimulation. We demonstrate in mouse models that complete (Lck-/-) versus partial (LckP440S/P440S) loss-of-function LCK causes disease with differing phenotypes. While both Lck-/- and LckP440S/P440S mice exhibit arrested thymic T cell development and profound T cell lymphopenia, only LckP440S/P440S mice show residual T cell proliferation, cytokine production, and intestinal inflammation. Furthermore, the intestinal disease in the LckP440S/P440S mice is prevented by CD4+ T cell depletion or regulatory T cell transfer. These findings demonstrate that P440S LCK spares sufficient T cell function to allow the maturation of some conventional T cells but not regulatory T cells-leading to intestinal inflammation.
Topics: Infant; Humans; Animals; Mice; CD28 Antigens; CD4-Positive T-Lymphocytes; Immunologic Deficiency Syndromes; Lymphocyte Specific Protein Tyrosine Kinase p56(lck); Receptors, Antigen, T-Cell; Inflammation; Lymphopenia
PubMed: 37962568
DOI: 10.1084/jem.20230927