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Effects of fatty acid mixtures on proliferation of peripheral blood mononuclear cells in dairy cows.Veterinary Research Communications Jun 2023This in vitro study was performed to assess the effects of three different mixtures of nonesterifed fatty acids (NEFA) on mitogen-driven proliferation of peripheral...
This in vitro study was performed to assess the effects of three different mixtures of nonesterifed fatty acids (NEFA) on mitogen-driven proliferation of peripheral blood mononuclear cells (PBMC) in dairy cows. Substantially, the three mixtures differed for n-6 to n-3 fatty acids (FA) ratio and were intended to mimic plasma NEFA composition of cows given fat supplements with different n-6 to n-3 FA ratio. PBMC from six Holstein heifers were cultured in media containing three different mixtures of oleic, palmitic, stearic, linoleic, palmitoleic, or linolenic acid at concentrations mimicking different degree of lipomobilisation. Proliferation of PBMC was stimulated by concanavalin A or pokeweed mitogen (PWM). Low concentrations of the three mixtures (62.5 and 125 µmol/l), did not affect the ability of PBMC to proliferate. Concentrations of the three mixtures mimicking medium-intense lipomobilisation (500, 1,000 and 1,500 µmol/l) impaired to the same extent proliferation of PBMC. The addition to cultures of the three mixtures at concentration of 250 µmol/l, impaired the proliferation only in PBMC stimulated with PWM. Even in this case, the three mixtures did not exert differential effects on PBMC proliferation. Present results reinforce the hypothesis that high concentrations of plasma NEFA play a role in the immunosuppression taking place in cows undergoing intense lipomobilisation, and authorize to suggest that under these conditions different composition of plasma NEFA in terms of different n-6 to n-3 FA ratio cannot prevent their negative effects on lymphocyte proliferation.
Topics: Cattle; Animals; Female; Fatty Acids; Leukocytes, Mononuclear; Fatty Acids, Nonesterified; Dietary Supplements; Cell Proliferation; Lactation; Diet
PubMed: 36446991
DOI: 10.1007/s11259-022-10024-7 -
Animal : An International Journal of... Feb 2024Gastrointestinal (GI) parasites cause significant production losses in grazing ruminants which can be mitigated by breeding animals resistant to disease. Lymphocyte...
Gastrointestinal (GI) parasites cause significant production losses in grazing ruminants which can be mitigated by breeding animals resistant to disease. Lymphocyte cytokine production and parasite-specific Immunoglobulin A (IgA) are adaptive immune traits associated with immunity to GI parasites. To explore the utility of these traits for selective breeding purposes, this study estimated the genetic parameters of the immune traits in sheep and assessed their relationship with disease and productivity traits. Whole blood stimulation assays were performed on 1 040 Scottish Blackface lambs at two months of age in 2016-2017. Blood was stimulated with either pokeweed mitogen (PWM), a non-specific activator of lymphocytes, and Teladorsagia circumcincta (T-ci) larval antigen to activate parasite-specific T lymphocytes. The type of adaptive immune response was determined by quantifying production of cytokines interferon-gamma (IFN-γ), interleukin (IL)-4, and IL-10, which relate to T-helper type (Th) 1, Th2 and regulatory T cell responses, respectively. Serum T-ci specific IgA was also quantified. Heritabilities were estimated for each immune trait by univariate analyses. Genetic and phenotypic correlations were estimated between different immune traits, and between immune traits vs. disease and productivity traits that were recorded at three months of age. Disease phenotypes were expressed as faecal egg counts (FEC) of nematode parasites (Strongyles and Nematodirus), faecal oocyst counts (FOC) of coccidian parasites, and faecal soiling score; production was measured as lamb live weight. Significant genetic variation was observed in all immune response traits. Heritabilities of cytokine production varied from low (0.14 ± 0.06) to very high (0.77 ± 0.09) and were always significantly greater than zero (P < 0.05). IgA heritability was found to be moderate (0.41 ± 0.09). Negative associations previously identified between IFN-γ production and FOC, and IL-4 production and strongyle FEC, were not evident in this study, potentially due to the time-lag between immune and parasitology measures. Instead, a positive genetic correlation was found between FOC and PWM-induced IFN-γ production, while a negative genetic correlation was found between FOC and T-ci induced IL-10. Live weight was negatively genetically correlated with IFN-γ responses. Overall, IFN-γ and IL-4 responses were positively correlated, providing little evidence of cross-regulation of Th1 and Th2 immunity within individual sheep. Furthermore, T-ci specific IgA was highly positively correlated with PWM-induced IL-10, indicating a possible role for this cytokine in IgA production. Our results suggest that while genetic selection for adaptive immune response traits is possible and may be beneficial for parasite control, selection of high IFN-γ responsiveness may negatively affect productivity.
Topics: Sheep; Animals; Parasites; Interleukin-10; Interleukin-4; Genetic Profile; Sheep, Domestic; Phenotype; Cytokines; Immunoglobulin A; Scotland; Sheep Diseases; Parasite Egg Count; Feces
PubMed: 38232660
DOI: 10.1016/j.animal.2023.101061 -
Journal of Chromatography. B,... Aug 2021B lymphocytes ('B cells') are components of the human immune system with obvious potential for medical and biotechnological applications. Here, we discuss the isolation...
B lymphocytes ('B cells') are components of the human immune system with obvious potential for medical and biotechnological applications. Here, we discuss the isolation of primary human B cells from both juvenile and adult tonsillar material using a two-step procedure based on gradient centrifugation followed by separation on a nylon wool column as alternative to the current gold standard, i.e., negative immunosorting from buffy coats by antibody-coated magnetic beads. We show that the nylon wool separation is a low-cost method well suited to the isolation of large amounts of primary B cells reaching purities ≥ 80%. More importantly, this method allows the preservation of all B cell subsets, while MACS sorting seems to be biased against a certain B cell subtype, namely the CD27 B cells. Importantly, compared to blood, the excellent recovery yield during purification of tonsillar B cells provides high number of cells, hence increases the number of subsequent experiments feasible with identical cell material, consequently improving comparability of results. The cultivability of the isolated B cells was demonstrated using pokeweed mitogen (PWM) as a stimulatory substance. Our results showed for the first time that the proliferative response of tonsillar B cells to mitogens declines with the age of the donor. Furthermore, we observed that PWM treatment stimulates the proliferation of a dedicated subpopulation and induces some terminal differentiation with ASCs signatures. Taken together this indicates that the proposed isolation procedure preserves the proliferative capability as well as the differentiation capacity of the B cells.
Topics: Adult; B-Lymphocytes; Cell Culture Techniques; Cell Proliferation; Cell Separation; Cells, Cultured; Centrifugation; Child; Humans; Nylons; Palatine Tonsil; Pokeweed Mitogens
PubMed: 34325309
DOI: 10.1016/j.jchromb.2021.122853 -
Fish & Shellfish Immunology Jul 2019Pax5 (Paired Box 5), a nuclear transcription factor expressed in B cell specifically, is a key regulator for B cell activation. In this study, we cloned and identified a...
Pax5 (Paired Box 5), a nuclear transcription factor expressed in B cell specifically, is a key regulator for B cell activation. In this study, we cloned and identified a Pax5 gene (OnPax5) from Nile tilapia (Oreochromis niloticus), which has an open reading frame of 1278 bp, encoding deduced amino acid sequence of 425 residues. OnPax5 contains a conserved DNA-binding domain encoding the paired box, an octapeptide, a homeobox homology region, a transactivation and a repressor domain. OnPax5 is constitutively expressed in various analyzed tissues of tilapia, with a relatively high expression in lymphoid organs, including spleen (SPL), anterior kidney (AK), and thymus. What's more, OnPax5 is highly expressed in leukocytes especially in IgM lymphocytes sorted from peripheral blood (PBL), SPL and AK. When stimulated with lipopolysaccharide (LPS) in vivo, OnPax5 expression was significantly up-regulated in PBL, SPL and AK. Upon stimulation with LPS, pokeweed mitogen and mouse anti-OnIgM monoclonal antibody in vitro, the expression of OnPax5 was also significantly up-regulated in leukocytes from SPL and AK. Taken together, Pax5, the B cell lineage specific activator factor, might get involved in B cell activation in Nile tilapia.
Topics: Amino Acid Sequence; Animals; Base Sequence; Cichlids; Fish Diseases; Fish Proteins; Gene Expression Profiling; Gene Expression Regulation; Immunity, Innate; PAX5 Transcription Factor; Phylogeny; Sequence Alignment
PubMed: 31039440
DOI: 10.1016/j.fsi.2019.04.059 -
Frontiers in Immunology 2020Quinolinate (Quin) is a classic example of a biochemical double-edged sword, acting as both essential metabolite and potent neurotoxin. Quin is an important metabolite...
Quinolinate (Quin) is a classic example of a biochemical double-edged sword, acting as both essential metabolite and potent neurotoxin. Quin is an important metabolite in the kynurenine pathway of tryptophan catabolism leading to the synthesis of nicotinamide adenine dinucleotide (NAD). As a precursor for NAD, Quin can direct a portion of tryptophan catabolism toward replenishing cellular NAD levels in response to inflammation and infection. Intracellular Quin levels increase dramatically in response to immune stimulation [e.g., lipopolysaccharide (LPS) or pokeweed mitogen (PWM)] in macrophages, microglia, dendritic cells, and other cells of the immune system. NAD serves numerous functions including energy production, the poly ADP ribose polymerization (PARP) reaction involved in DNA repair, and the activity of various enzymes such as the NAD-dependent deacetylases known as sirtuins. We used highly specific antibodies to protein-coupled Quin to delineate cells that accumulate Quin as a key aspect of the response to immune stimulation and infection. Here, we describe Quin staining in the brain, spleen, and liver after LPS administration to the brain or systemic PWM administration. Quin expression was strong in immune cells in the periphery after both treatments, whereas very limited Quin expression was observed in the brain even after direct LPS injection. Immunoreactive cells exhibited diverse morphology ranging from foam cells to cells with membrane extensions related to cell motility. We also examined protein expression changes in the spleen after kynurenine administration. Acute (8 h) and prolonged (48 h) kynurenine administration led to significant changes in protein expression in the spleen, including multiple changes involved with cytoskeletal rearrangements associated with cell motility. Kynurenine administration resulted in several expression level changes in proteins associated with heat shock protein 90 (HSP90), a chaperone for the aryl-hydrocarbon receptor (AHR), which is the primary kynurenine metabolite receptor. We propose that cells with high levels of Quin are those that are currently releasing kynurenine pathway metabolites as well as accumulating Quin for sustained NAD synthesis from tryptophan. Further, we propose that the kynurenine pathway may be linked to the regulation of cell motility in immune and cancer cells.
Topics: Animals; Biomarkers; Cell Movement; Gerbillinae; HSP90 Heat-Shock Proteins; Hippocampus; Immunity; Indoleamine-Pyrrole 2,3,-Dioxygenase; Inflammation; Kynurenine; Lipopolysaccharides; Liver; Male; Mice; Mice, Inbred C57BL; NAD; Pokeweed Mitogens; Poly(ADP-ribose) Polymerases; Quinolinic Acid; Rats; Spleen; Tryptophan
PubMed: 32153556
DOI: 10.3389/fimmu.2020.00031 -
Cytokine May 2021Immunological assays are the basis for many diagnostic tests for infectious diseases in animals and humans. Application in wildlife species, including the African...
Immunological assays are the basis for many diagnostic tests for infectious diseases in animals and humans. Application in wildlife species, including the African elephant (Loxodonta africana), is limited however due to lack of information on immune responses. Since many immunoassays require both identified biomarkers of immune activation as well as species-specific reagents, it is crucial to have knowledge of basic immunological responses in the species of interest. Cytokine gene expression assays (GEAs) used to measure specific immune responses in wildlife have frequently shown that targeted biomarkers are often species-specific. Therefore, the aim of this study was to identify elephant-specific cytokine biomarkers to detect immune activation and to develop a GEA, using pokeweed mitogen stimulated whole blood from African elephants. This assay will provide the foundation for the development of future cytokine GEAs that can be used to detect antigen specific immune responses and potentially lead to various diagnostic tests for this species.
Topics: Animals; Cytokines; Elephants; Gene Expression Regulation; Immunoassay
PubMed: 33548797
DOI: 10.1016/j.cyto.2021.155453 -
Journal of Neurotrauma May 2024Traumatic brain injury (TBI) is a common cause of morbidity and mortality in children. We have previously shown that TBI with a concurrent extracranial injury reliably...
Granulocyte- Macrophage Colony-Stimulating Factor Reverses Immunosuppression Acutely Following a Traumatic Brain Injury and Hemorrhage Polytrauma in a Juvenile Male Rat Model.
Traumatic brain injury (TBI) is a common cause of morbidity and mortality in children. We have previously shown that TBI with a concurrent extracranial injury reliably leads to post-injury suppression of the innate and adaptive immune systems. In patients with post-injury immune suppression, if immune function could be preserved, this might represent a therapeutic opportunity. As such, we examined, in an animal injury model, whether systemic administration of granulocyte macrophage colony-stimulating factor (GM-CSF) could reverse post-injury immune suppression and whether treatment was associated with neuroinflammation or functional deficit. Prepubescent male rats were injured using a controlled cortical impact model and then subjected to removal of 25% blood volume (TBI/H). Sham animals underwent surgery without injury induction, and the treatment groups were sham and injured animals treated with either saline vehicle or 50 μg/kg GM-CSF. GM-CSF was administered following injury and then daily until sacrifice at post-injury day (PID) 7. Immune function was measured by assessing tumor necrosis factor-α (TNF-α) levels in whole blood and spleen following stimulation with pokeweed mitogen (PWM). Brain samples were assessed by multiplex enzyme-linked immunosorbent assay (ELISA) for cytokine levels and by immunohistochemistry for microglia and astrocyte proliferation. Neuronal cell count was examined using cresyl violet staining. Motor coordination was evaluated using the Rotarod performance test. Treatment with GM-CSF was associated with a significantly increased response to PWM in both whole blood and spleen. GM-CSF in injured animals did not lead to increases in levels of pro-inflammatory cytokines in brain samples but was associated with significant increases in counted astrocytes. Finally, while injured animals treated with saline showed a significant impairment on behavioral testing, injured animals treated with GM-CSF performed similarly to uninjured animals. GM-CSF treatment in animals with combined injury led to increased systemic immune cell response in whole blood and spleen in the acute phase following injury. Improved immune response was not associated with elevated pro-inflammatory cytokine levels in the brain or functional impairment.
PubMed: 38623766
DOI: 10.1089/neu.2023.0169