Spleen Cells Research Articles

Gaucher’s Disease – What Should You Know

Gaucher’s Disease (GD) has a special place among ultra-rare diseases, which is a disease that occurs in less than onein 50 000 persons. The disease is caused by a hereditary autosomal deficiency of the lysosomal enzyme glucocerebrosidase. The pathognomonic feature of GD is the presence of storage cells or gaucher cells in spleen, liver and other organs and tissues. There are several clinical types of gaucher disease, differing in manifestation, prognosis and treatment. It is very important to diagnose the disease early so that the correct treatment can be implemented. Nowadays, it is not uncommon for general practitioners and specialists to make the wrong diagnosis due to the low probability and lack of knowledge of the disease, leading to a worsening patient's condition. A proper examination consists of a medical history, physical examination and targeted radiological examinations, which are of particular importance in the diagnosis due to changes in the body that are not otherwise detectable, but which already offer a chance to make a diagnosis. The latest treatment is based on enzyme replacement therapy, but unfortunately the prognosis in some cases is still unfavourable. This is why early diagnosis and research to find an effective treatment are so important.

Developmental Vitamin D Deficiency and the Vitamin D Receptor Control Hematopoiesis.

Vitamin D status, the vitamin D receptor (VDR), and the ability to produce active vitamin D [1,25(OH)2D, regulated by Cyp27b1] regulate fetal and adult hematopoiesis. Transgenic reporter mice that express the tdTomato RFP as an indication of Vdr expression were used to identify immune cells that express the Vdr. Vdr/tdTomato+ hematopoietic progenitors were identified as early as embryonic day (E)15.5, establishing that these cells have expressed the Vdr and are vitamin D targets. Maternal vitamin D deficiency [D-; serum 25(OH)D < 20 ng/ml] or Vdr knockout or Cyp27b1 knockout resulted in embryos with fewer fetal progenitors. Vdr/tdTomato+ expression was found to increase with age in CD8+ T cells and innate lymphoid cells (ILCs)1 and ILC3, suggesting that initial Vdr expression in these cells is dependent on environmental factors immediately postbirth. In adult tissues, the frequencies of mature T cells and ILCs as well as Vdr/tdTomato expression were reduced by D-. Maternal D- resulted in fewer progenitors that expressed Vdr/tdTomato+ at E15.5 and fewer Vdr/tdTomato+ immune cells in the adult spleen than offspring from D+ mice. We challenged D- mice with H1N1 influenza infection and found that D- mice were more susceptible than D+ mice. Treating D- mice with vitamin D restored Vdr/tdTomato+ expression in splenic T cells and partially restored resistance to H1N1 infection, which shows that developmental D- results in lingering effects on Vdr expression in the adult immune system that compromise the immune response to H1N1 infection. Vitamin D and the Vdr regulate hematopoiesis in both fetal and postnatal phases of immune cell development that impact the immune response to a viral infection.

Prebiotic inulin controls Th17 cells mediated central nervous system autoimmunity through modulating the gut microbiota and short chain fatty acids

ABSTRACT Multiple sclerosis (MS) is an autoimmune disease characterized by inflammatory demyelination occurring in the central nervous system (CNS). Inulin is a common prebiotic that can improve metabolic disorders by modulating the gut microbiota. However, its capacity to affect CNS autoimmunity is poorly recognized. Experimental autoimmune encephalomyelitis (EAE) is a classical mouse model of MS. Herein, we found that oral administration of inulin ameliorated the severity EAE in mice, accompanied by reductions in inflammatory cell infiltration and demyelination in the CNS. These reductions were associated with decreased proportion and numbers of Th17 cells in brain and spleen. Consistent with the findings, the serum concentrations of IL-17, IL-6, and TNF-α were reduced in inulin treated EAE mice. Moreover, the proliferation of auto-reactive lymphocytes, against MOG35-55 antigen, was attenuated ex vivo. Mechanistically, inulin treatment altered the composition of gut microbiota. It increased Lactobacillus and Dubosiella whereas decreased g_Prevotellaceae_NK3B31_group at the genus level, alongside with elevated concentration of butyric acid in fecal content and serum. In vitro, butyrate, but not inulin, could inhibit the activation of MOG35-55 stimulated lymphocytes. Furthermore, fecal microbiota transplantation assay confirmed that fecal contents of inulin-treated normal mice had an ameliorative effect on EAE mice. In contrast, antibiotic cocktail (ABX) treatment diminished the therapeutic effect of inulin in EAE mice as well as the reduction of Th17 cells, while supplementation with Lactobacillus reuteri restored the amelioration effect. These results confirmed that the attenuation of inulin on Th17 cells and inflammatory demyelination in EAE mice was dependent on its modulation on gut microbiota and metabolites. Our findings provide a potential therapeutic regimen for prebiotic inulin supplementation in patients with multiple sclerosis.

Immune and microbiome modulatory effects of Limosilactobacillus fermentum NCDC 400 in an immunocompromised mouse model

The present study was aimed to assess and validate the safety and functional efficacy of an indigenous probiotic strain Limosilactobacillus fermentum NCDC 400 (hereafter, LFN400) in an immunocompromised murine model. The study included four groups; a normal control (NC) group without immune suppression; an experimental model control (MC) with immune suppression induced via intraperitoneal cyclophosphamide (Cy) administration; and two MC groups orally administered with either low dose (LD) or high dose (HD) of LFN400 at dose 108 and 1010 CFU/mouse/day, respectively, for 15-days. Both control groups received normal saline as placebo control. LFN400 improved specific experimental characteristics including hematological and serum biochemical markers. Compared to MC group, LFN400-fed groups showed markedly (P < 0.05) decreased arrays of detrimental caecal enzymes. We did not observe instances of bacterial translocation of LFN400 from gut to bloodstream or extra-intestinal organs. LFN400 intake significantly (P < 0.05) enhanced spleen cell differentiation, immune and oxidative stress markers, and restored Cy-induced histopathological changes in multiple tissues, including the spleen. There was no genotoxic effect of LFN400 on bone marrow cells. Although not statistically significant, LFN400 feeding moderately increased gut microbiome diversity, supporting the growth of beneficial saccharolytic microorganisms and reducing the presence of pathobionts. The findings demonstrate that the probiotic strain LFN400 possesses in vivo safety and immunomodulatory potency and thus should be considered a potential candidate for future human clinical studies.

N-formylmethionine-leucyl-phenylalanine protects against irradiation-induced damage to hematopoiesis and intestines

BackgroundIonizing radiation (IR), including radiotherapy, can exert lasting harm on living organisms. While liposaccharide (LPS) offers resistance to radiation damage, it also induces toxic responses. Thankfully, an LPS analogue called N-formylmethionine-leucyl-phenylalanine (fMLP) holds the potential to mitigate this toxicity, offering hope for radiation protection.MethodsSurvival of C57BL/6 mice exposed to IR after administration with fMLP/LPS/WR-2721 or saline was recorded. Cell viability and apoptosis assay of bone marrow (BMC), spleen and small intestinal epithelial (HIECs) cells were tested by Cell Counting Kit-8 (CCK-8) and flow cytometry assay. Tissue damage was evaluated by Hematoxilin and Eosin (H&E), Ki-67, and TUNEL staining. RNA sequencing was performed to reveal potential mechanisms of fMLP-mediated radiation protection. Flow cytometry and western blot were performed to verify the radiation protection mechanism of fMLP on the cell cycle.ResultsThe survival rates of C57BL/6 mice exposed to ionizing radiation after administering fMLP increased. fMLP demonstrated low toxicity in vitro and in vivo, maintaining cell viability and mitigating radiation-induced apoptosis. Moreover, it protected against tissue damage in the hematopoietic and intestinal system. RNA sequencing shed light on fMLP’s potential mechanism, suggesting its role in modulating innate immunity and cell cycling. This was evidenced by its ability to reverse radiation-induced G2/M phase arrests in HIECs.ConclusionfMLP serves as a promising radioprotective agent, preserving cells and radiosensitive tissues from IR. Through its influence on the cell cycle, particularly reversing radiation-induced arrest in G2/M phases, fMLP offers protection against IR’s detrimental effects.

Intranasal Immunization with a Recombinant Adenovirus Encoding Multi-Stage Antigens of Mycobacterium tuberculosis Preferentially Elicited CD8+ T Cell Immunity and Conferred a Superior Protection in the Lungs of Mice than Bacillus Calmette-Guerin.

The development of a tuberculosis (TB) vaccine is imperative. Employing multi-stage Mycobacterium tuberculosis (Mtb) antigens as targeted antigens represents a critical strategy in establishing an effective novel TB vaccine. In this investigation, we evaluated the immunogenicity and protective efficacy of a recombinant adenovirus vaccine expressing two fusion proteins, Ag85B-ESAT6 (AE) and Rv2031c-Rv2626c (R2), derived from multi-stage antigens of Mtb via intranasal administration in mice. Intranasal delivery of Ad-AE-R2 induced both long-lasting mucosal and systemic immunities, with a preferential elicitation of CD8+ T cell immunity demonstrated by the accumulation and retention of CD8+ T cells in BALF, lung, and spleen, as well as the generation of CD8+ TRM cells in BALF and lung tissues. Compared to subcutaneous immunization with Bacillus Calmette-Guerin (BCG), Ad-AE-R2 provided superior protection against high-dose intratracheal BCG challenge, specifically within the lungs of mice. Our findings support the notion that empowering T cells within the respiratory mucosa is crucial for TB vaccine development while highlighting targeting CD8+ T cell immunity as an effective strategy for optimizing TB vaccines and emphasizing that eliciting systemic memory immunity is also vital for the successful development of a TB mucosal vaccine. Furthermore, our results demonstrate that the BCG challenge serves as a convenient and efficient method to evaluate candidate vaccine efficacy.

Lespedeza homoloba enhances the immunosuppressive milieu of adipose tissue and suppresses fasting blood glucose.

Immune cells migrate to hypertrophied adipocytes and release proinflammatory cytokines, leading to adipocyte dysfunction and diabetes. Numerous species of Lespedeza, which are members of the plant family Fabaceae and distributed primarily in temperate Asia and North America, exhibit binding to peroxisome proliferator-activated receptor (PPAR) γ, a target nuclear receptor for treating diabetes. Therefore, the present study aimed to determine which species of Lespedeza plants exert an anti-inflammatory effect in adipose tissue and suppression of blood glucose increase through PPARγ ligand and radical scavenging activity. PPARγ binding and DPPH radical scavenging assays of L. homoloba (LH), L. thunbergii (LT), L. maximowiczii (LM) and L. thunbergii (LT) were performed. LH and LT showed significant ligand activity towards PPARγ and notable radical scavenging activity. LH exhibited a stronger DPPH radical scavenging activity than LT and thus was measured adiponectin secretion from 3T3-L1-derived adipocytes and IL-10 secretion from murine splenocytes. LH increased the adiponectin and the IL-10 secretions. In flow cytometric analysis, BALB/c male mice administered LH exhibited an increase in regulatory T cells (Tregs) and cytotoxic T lymphocyte-associated protein (CTLA)-4+ Tregs as well as a decrease in T helper (Th)17, Th17/Treg ratio and CD8+ and CD4+ T cells in subcutaneous adipose tissue. Conversely, in the spleen, LH decreased Tregs and increased Th17 cells, Th17/Treg ratio and CD4+ and CD8+ T cells. These findings indicated that LH activated immunoreaction in the spleen and Treg cells that migrate to subcutaneous adipose tissue may suppress inflammation. In fasting blood glucose and adiponectin assays, LH-exposed mice exhibited suppression of fasting glucose levels. Therefore, LH may prevent type 2 diabetes by suppressing adipose tissue inflammation.

Abstract P389: Immunogenic Peptides Responsible for Experimental Hypertension Contribute to Systemic Lupus Erythematosus

Introduction: Hypertension and systemic lupus erythematosus (SLE) lead to modification of self-peptides by isolevuglandin (isoLG) lipid oxidation products. These self-peptides are presented by class 1 major histocompatibility complexes and drive CD8 + T cell activation. We have identified self-peptides from the sodium/glucose cotransporter2 (SGLT2); cadherin 16; Kelch domain-containing protein 7A and solute carrier family 23 that are IsoLG-adducted and cause CD8 + T cell activation in both ang II and L-NAME/high salt models of hypertension. These prime hypertension in response to low dose ang II. Hypothesis: We hypothesized that the same immunogenic peptides responsible for experimental hypertension contribute to systemic lupus erythematosus. Methods: A fluorescent probe made of recombinant IgG and the murine class 1 MHC H2-D b was loaded with either unadducted SGLT2 peptide or an isoLG-adducted SGLT2 peptide. This probe was used in flow cytometry to detect CD8 + T cells specific for this peptide antigen in 12-week-old C57Bl/6 and female B6SLE1.2.3 mice. Results: The unadducted SGLT2 probe recognized &lt; 1% of CD8 + T cells (Figure). In contrast, ang II-infusion caused a striking increase in IsoLG-SGLT2 specific CD8 + T cells in C57Bl/6 aortas (Figure panels A and D). In C57Bl/6 very few CD8 + T cells recognizing IsoLG-SGLT2 were observed in the spleen (Panel B and E). Importantly, from 5 to 20% of CD8 + T cells in spleens of female B6SLE1.2.3 mice recognized the IsoLG-SGLT2 peptide (Figure panels C and E). Image 1. Conclusions: These findings strongly suggest that IsoLG-adducted peptides responsible for ang II-induced hypertension contribute to SLE and suggest that these conditions share similar immunogenicity.

IL-18 primes T cells with an antigen-inexperienced memory phenotype for proliferation and differentiation into effector cells through Notch signaling.

Recent studies have revealed that a subset of CD8+ T cells exhibit innate features and can be activated by cytokines. However, the precise mechanisms underlying the proliferation and differentiation of these cells remain unclear. Here, we demonstrated that CD44highCD8+ T cells in the mouse spleen express functional interleukin-18 (IL-18) receptors, whereas CD44lowCD8+ T cells do not. In response to IL-18 stimulation, these cells activated various metabolic pathways, upregulated the expression of surface molecules, such as c-Kit (CD117), CD25, and PD-1, and induced progression through the G1/S phase in the cell cycle. IL-18-primed cells, expressing a high-affinity receptor for IL-2, exhibited robust proliferation in response to IL-2 and underwent differentiation into effector cells. The splenic CD44highCD8+ T cells exhibited high expression levels of CD122, CD62L, CCR7, and CXCR3, along with CD5, indicating their potential for migration to the lymph nodes, where they could undergo expansion and terminal differentiation into effector cells. Additionally, in a tumor model, administration of IL-18 increased the accumulation of CD8+ T cells in both the lymph nodes and tumors. It is noteworthy that stimulation of CD44highCD8+ T cells with IL-18 upregulated the Notch-1 receptor and c-Myc. Moreover, inclusion of γ-secretase inhibitors attenuated the effect of IL-18 on both proliferation and interferon-γ production in the cells. These results demonstrate that IL-18 primes CD44highCD122highCXCR3highCD62LhighCD8+ T cells for expansion and differentiation into effector cells in a Notch signaling-dependent manner.

Sepsis induced dysfunction of liver type 1 innate lymphoid cells

BackgroundSepsis is a life-threatening condition triggered by uncontrolled immune responses to infection, leading to widespread inflammation, tissue damage, organ dysfunction, and potentially death. The liver plays a crucial role in the immune response during sepsis, serving as a major site for immune cell activation and cytokine production. Liver type 1 innate lymphoid cells (ILCs) consist of NK cells and ILC1s. They maintain the local immune microenvironment by directly eliminating target cells and secreting cytokines. However, the specific roles and pathological changes of liver-resident NK cells and ILC1s during sepsis remain poorly understood.ResultsThis study aims to investigate the pathological changes of NK cells and ILC1s, which might contribute the dysfunction of liver. Sepsis mouse model was established by cecal ligation and puncture (CLP). Mouse immune cells from liver were isolated, and the surface makers, gene expression profiles, cytokine response and secretion, and mitochondrial function of NK (Natural Killer) cells and ILC1s (Innate Lymphoid Cell 1) were analyzed. A significant decrease in the number of mature NK cells was observed in the liver after CLP. Furthermore, the secretion of interferon-gamma (IFN-γ) was found to be reduced in spleen and liver NK cells when stimulated by IL-18. Mitochondrial activities in both liver NK cells and ILC1 were found to be increased during sepsis, suggesting an enhanced metabolic response in these cells to combat the infection. However, despite this heightened activity, liver NK cells exhibited a decreased level of cytotoxicity, which might impact their ability to target infected cells effectively. RNA sequencing supported and provided the potential mechanisms for the proinflammatory effects and exhaustion like phenotypes of liver NK cells.ConclusionsSepsis induces dysfunction and exhaustion-like phenotypes in liver NK cells and ILC1, which might further impair other immune cells and represent a potential therapeutic target for sepsis.

Characterization of IgD and IgT with their expressional analysis following subtype II megalocytivirus vaccination and infection in rock bream (Oplegnathus fasciatus)

In this study, heavy chain genes of IgD and IgT were sequenced and characterized their gene expression in rock bream (Oplegnathus fasciatus). Rock bream (RB)-IgD cDNA is 3319 bp in length and encodes a leader region, variable domains, a μ1 domain, and seven constant domains (CH1–CH7). A membrane-bound (mIgT) and secretory form (sIgT) of RB-IgT cDNAs are 1902 bp and 1689 bp in length, respectively, and encode a leader region, variable domains, four constant domains (CH1–CH4) and C-terminus. Their predicted 3D-structure and phylogenetic relation were similar to those of other teleost. In healthy fish, RB-IgD and mIgT gene expressions were higher in major lymphoid organs and blood, while RB-sIgT gene was more highly expressed in midgut. IgT expressing cells were detected in melano-macrophage centers (MMC) of head kidney in immunohistochemistry analysis. Under immune stimulation in vitro, RB-IgD and IgT gene expressions were upregulated in head kidney and spleen cells by bovine serum albumin or a rock bream iridovirus (RBIV) vaccine. In vivo, their expressions were significantly upregulated in head kidney, blood, and gill upon vaccination. Especially, RB-mIgT gene expression in head kidney and blood was upregulated at day 3 after vaccination while upregulated at earlier time point of day 1 by challenge with RBIV. This may suggest that memory cells might be produced during the primary response by vaccination and rapidly proliferated by secondary immune response by viral infection. RB-sIgT gene expression was highly upregulated in peripheral blood in vaccinated fish after viral infection, indicating that IgT plays an important role in systemic immune response as well as mucosal immune system. Our findings provide information on the role of RB-IgT in adaptive immunity during vaccination and viral infection in the vaccinated fish.

Multi-epitope Vaccine Design against Grouper Iridovirus (GIV) Using Immuno-bioinformatics Approach

Grouper Iridovirus (GIV) infection induced cell death in grouper spleen cells and caused serious systemic diseases with more than 90% mortality. Therefore, effective strategies are critically needed to prevent economic losses and maintain the sustainability of grouper aquaculture. Using immuno-bioinformatics, this study aimed to create a multi-epitope vaccine (MEV) that would be effective against GIV. The GIV major capsid protein sequences were retrieved from the NCBI proteome database. Out of 284 epitopes, 17 CTL, 12 HTL, and 10 B-cell epitopes were predicted to be antigenic, non-allergenic, and non-toxic. 10 highly antigenic and overlapping epitopes were shortlisted. To generate full-length epitope vaccine candidates, the selected antigenic epitopes were fused with linkers and adjuvants. Four sets of different linker combinations (no linker, GGS, EAAK, GGGS, GPGPG, KK, and AAY) were tested and compared for their antigenicity, allergenicity, and toxicity using several servers. Molecular dynamics simulations with GROMACS were used on the modelled 3D structures to examine their stability. The results of vaccine candidate sequences screening and MD simulation predicted that the structure with GGS linker is relatively stable with a high antigenic index, non-allergenic, and non-toxic. The designed MEV in the present study could be a potential candidate for further vaccine production process against GIV.

Chemokine receptors are required for effector T-cell trafficking to GVHD tissues but not to bone marrow

AbstractIn allogeneic hematopoietic stem cell transplantation (allo-SCT), alloreactive donor T cells mediate the graft-versus-leukemia effect but also attack nonhematopoietic tissues, causing graft-versus-host disease (GVHD). Reducing alloreactive T-cell trafficking to GVHD target tissues while allowing their access to bone marrow (BM) and spleen, major sites of malignant hematopoiesis, is a rational strategy for reducing the GVHD risk when using alloreactive T cells as a therapeutic. Here, we show that effector T-cell (Teff) entry into BM and spleen in unmanipulated mice and in mice that received transplantation without alloreactive T cells is augmented by pertussis toxin (PTX)-sensitive chemokine receptor signaling. However, unexpectedly, in the presence of a GVH response, chemokines no longer draw T cells into BM and spleen but remain critical for their recruitment to GVHD target tissues. Consistent with this, PTX-treated Teff cells were as efficacious as untreated T cells in killing leukemia cells in BM and spleen in mice with a concurrent GVHD response. These results suggest a strategy to improve the safety of alloreactive T-cell therapeutics in treating leukemias in the context of an allo-SCT.

Central effects of &lt;i&gt;ex vivo&lt;/i&gt; caffeine-modulated immune cells in the mechanisms of editing depressive-like behavior

Depression is a serious medical and social problem due to its high prevalence, involvement of people of working age and lack of highly effective therapy. Social stressors contribute to the prevalence of depression. The COVID-19 pandemic and the associated rules of social distancing, military clashes, and a deteriorating economic situation can lead to a painful “breakdown” of socio-biological adaptation mechanisms and contribute to an increase in the prevalence of depressive disorders, which, according to WHO forecasts, may take second place in the world by 2030, which leads to interest in studying this problem and finding new effective approaches to therapy. Decreased cognitive function in depressive disorders is caused by neuroinflammatory and neurodegenerative changes. The latter are predominantly recorded in the hippocampus, numerous changes in the plasticity of which have been observed both in patients with clinical depression and in rodent models of depression. There is also a sufficient amount of data on the significant role of immune cells and their cytokines in depression, including in the development of behavioral phenotype. We have previously shown that spleen cells of depressive-like mice after ex vivo treatment with caffeine, a psychoactive drug with a wide range of immunomodulatory properties, change their functional activity and, after intravenous administration to syngeneic depressive-like recipients, have editing depressive-like behavior effect. The purpose of this work was to investigate the central effects of caffeine-modulated spleen cells in the mechanisms of editing depressive-like behavior. It was found that in depressive-like recipients after transplantation of syngeneic caffeine-modulated splenocytes, there is an increase in the density of neurons in the CA1 and CA3 zones of the hippocampus, accompanied by BDNF level increase in the hippocampus and prefrontal cortex against the background of a decrease of a number of pro-inflammatory (IL-1β, IL-6, IFNγ and TNFα) and increased of anti-inflammatory (IL-10 and IL-4) cytokines in brain structures pathogenetically significant for the state of depression. The mechanisms of the identified structural and functional changes in the recipient’s brain after the caffeine-modulated splenocytes transplantation are discussed, including their possible direct influence, confirmed by visualization of transplanted cells in the brain parenchyma of depressed-like recipients.

Spontaneous cytokine production by the cells from peripheral blood, lymph nodes, spleen, thymus and skin in a murine model of acute psoriasis-like dermatitis

Imiquimod-induced model of psoriasis in mice is used for the studies in psoriasis. Imiquimod administered as skin application triggers a complex multi-stage process simulating psoriatic inflammation of the skin, accompanied by changes in production of cytokines. The aim of this work was a comprehensive study of their contents in cell supernatants isolated from skin, blood, from central and peripheral lymphoid organs and the effect of imiquimod on these parameters. Forty-six C57BL/6 mice were divided into two groups: (1) control (n = 22), and (2) experimental (n = 24). Development of experimental pathology was evaluated by van de Fits et al. The mice from experimental group were treated with a cream containing 5% imiquimod (62.5 mg/ cm2/ day/mouse) for 7 days. On the 7th day of experiment, the animals were subject to blood sampling, extraction of spleen, lymph nodes, thymus, and skin biopsies were also made. To obtain cell suspensions, the tissues of spleen, lymph nodes, and thymus were crushed in a homogenizer, and then passed through cell filters (50 mcm pore size). The method of spontaneous migration was used to isolate skin cells. The cultures of blood, spleen, lymph nodes, thymus, and skin cells were incubated for 24 hours in RPMI-1640 followed by assessment of spontaneous cytokine production in supernatants. The cytokine level (IFNγ, IL-1β, IL-5, IL-6, IL-10, IL- 17, TNFα, IL-10) was determined using the murine Th1/Th2/Th17 Panel test system. Our study revealed that the cells isolated from blood and lymphoid organs synthesize increased amounts of pro-inflammatory cytokines: IL-1 and IL-17. The most pronounced changes in the cytokine profile are observed in cell supernatants from blood, skin and spleen cultures.

Co-exposure of bisphenol A and selenium deficiency induces pyroptosis via ROS/NLRP3 pathway in chicken spleen

Bisphenol A (BPA) is widely applied in plastic products, which will produce immunotoxicity to organisms after spilling in the environment, and become a kind of endocrine disruptor. Selenium (Se) is an essential trace element and plays an important role in maintaining redox homeostasis and immune function. BPA exposure and Se deficiency often occur together in livestock and poultry farming, however, studies on the effects of joint exposure on chicken immunotoxins have not been reported. Therefore, this study established a chicken spleen and MDCC-MSB1 cell model under the combined effects of BPA exposure or/and Se deficiency. Transcriptomic analysis showed that BPA exposure and/or Se deficiency induced differential enrichment of positive regulatory pathways such as NLRP3 inflammatory complex assembly, inflammatory response and cellular oxidative stress response. In the -Se+BPA group, pathological damage was significantly increased, Se content decreased, BPA accumulation, oxidative stress and pyroptosis. Meanwhile, the roles and mechanisms of oxidative stress and pyroptosis in BPA exposure or/and Se deficiency-induced splenic tissue injury were investigated by using IF and qRT-PCR methods. The results showed that joint BPA exposure with Se deficiency resulted in more significant changes in the above outcomes than one of them. The oxidative stress inhibitor NAC effectually reduced Se deficiency and BPA-induced oxidative stress and pyroptosis, further suggests that oxidative stress mediated Se deficiency or/and BPA-induced pyroptosis. This study revealed that BPA exposure and Se deficiency induced spleen pyroptosis in chickens via the ROS/NLRP3 pathway. These results provide the theoretical basis for the toxicity of BPA in poultry and enrich the toxicological mechanism of combined exposure of Se deficiency and environmental toxins.

Antipolyphosphate monoclonal antibodies derived from autoimmune mice

BackgroundInorganic polyphosphates (polyPs) are linear chains of phosphates that accelerate blood clotting. Targeting polyP in vivo has been shown to reduce thrombosis. ObjectivesTo identify and characterize anti-polyP monoclonal antibodies that could be used as analytical tools and as antithrombotic agents. MethodsHybridomas were prepared from spleen cells from autoimmune NZBWF1/J female mice and screened for anti-polyP antibodies. Antibodies that bound polyP using enzyme-linked immunosorbent assay and pull-down assays were further characterized with plate binding, surface plasmon resonance, and plasma-based clotting assays. Antithrombotic potential was evaluated in a murine ferric chloride–induced carotid artery thrombosis model. ResultsOf 4 antibodies that bound polyP in our pull-down assay, 2 (PP2069 and PP2099) were available for further characterization. While analyzing these anti-polyP antibodies, we found secretory leukocyte peptidase inhibitor (SLPI) to be a common contaminant of these antibodies and that SLPI binds polyP. We removed SLPI quantitatively from our purified immunoglobulin G. Both PP2069 and PP2099 immunoglobulin G displayed high affinity for polyP but also bound to other polyanions such as DNA, heparin, and certain other glycosaminoglycans, indicating limited specificity. Both antibodies inhibited polyP-initiated plasma clotting in vitro. When tested in vivo in a mouse thrombosis model, however, neither PP2069 nor PP2099 exhibited a significant antithrombotic effect. ConclusionAutoimmune mice spontaneously produce antibodies against polyP. The 2 examples of anti-polyP monoclonal antibodies studied here not only bound to polyP with high affinity but also cross-reacted with DNA and heparin. Neither antibody protected against thrombosis in a mouse model, but they might have some utility for in vitro studies of polyP.

Proinflammatory Effect of Membrane Vesicles Derived from Enterococcus faecalis at Neutral and Alkaline pH

IntroductionThe present study explored the proinflammatory impact of Enterococcus faecalis membrane vesicles (MVs) derived from culture medium at pH levels of 7.4 and 9.0. MethodsE. faecalis MVs were obtained by centrifugation and purified by size-exclusion chromatography. Proteomic analyses were performed on E. faecalis MVs to investigate their components. THP-1 macrophages were exposed to E. faecalis MVs, and the inflammatory cytokines and proteins were evaluated using enzyme-linked immunosorbent assay and immunoblotting. The inflammatory cytokines in the serum of mice with intraperitoneal injection of E. faecalis MVs were evaluated by enzyme-linked immunosorbent assay, and immunophenotyping of spleen cells was investigated with flow cytometry. ResultsProteomic analysis revealed 196 proteins in E. faecalis MVs obtained under neutral and alkali conditions; 110 proteins were up-regulated, and 79 proteins were down-regulated by alkaline pH. E. faecalis MVs induced secretion of inflammatory factors interleukin (IL)-1β, IL-6, and tumor necrosis factor alpha in a concentration-dependent manner. Immunoblotting revealed that E. faecalis MVs increased expression of pro–IL-1β, nuclear factor kappa Bp65, and Toll-like receptor 2. In vivo studies demonstrated that E. faecalis MVs significantly promoted secretion of IL-1β in mouse serum, whereas inflammatory cells were activated in the spleen. E. faecalis MVs obtained at a pH of 9.0 showed stronger proinflammatory effects than those obtained under neutral pH. ConclusionsE. faecalis produces MVs that carry specific proteins associated with virulence factors, and these MVs can promote inflammation in vitro and in vivo. E. faecalis MVs obtained under alkaline conditions have a stronger proinflammatory effect.

Ficus hirta Vahl. alleviate LPS induced apoptosis via down-regulating of MiR-411 in orange-spotted grouper (Epinephelus coioides) spleen cell

Ficus hirta Vahl. (FhV) has been shown to have antimicrobial and antiviral efficacy. To further ascertain the pharmacological properties of FhV., and to search for alternatives to antibiotics. An in vitro experiment was carried out to evaluate what influence FhV. would have on LPS-induced apoptosis. In this study, Fas, an apoptosis receptor, was cloned, which included a 5′-UTR of 39 bp, an ORF of 951 bp, a protein of 316 amino acids, and a 3′-UTR of 845 bp. EcFas was most strongly expressed in the spleen tissue of orange-spotted groupers. In addition, the apoptosis of fish spleen cells induced by LPS was concentration-dependent. Interestingly, appropriate concentrations of FhV. alleviated LPS-induced apoptosis. Inhibition of miR-411 further decreased the inhibitory effect of Fas on apoptosis, which reduced Bcl-2 expression and mitochondrial membrane potential, enhanced the protein expression of Bax and Fas. More importantly, the FhV. could activate miR-411 to improve this effect. In addition, luciferase reporter assays showed that miR-411 binds to Fas 3'-UTR to inhibit Fas expression. These findings provide evidence that FhV. alleviates LPS-induced apoptosis by activating miR-411 to inhibit Fas expression and, therefore, provided possible strategies for bacterial infections in fish.

Follicular Helper T Cells in Peyer's Patches and Galactose- Deficient Iga1 Contribute to Iga Nephropathy.

Common primary glomerulonephritis with aberrant mucosal immunity is IgA nephropathy (IgAN). T follicular helper (TFH) cells are essential in regulating B cell differentiation. Peyer's patches (PPs) are the main site where IgA+ plasmablasts differentiate. Our study aimed to investigate the TFH cell's potential contribution to the etiology of IgA nephropathy. In PPs from IgAN mouse models, the ratio of the TFH cell, B220+IgA+, B220+IgM+, and B220-IgA+ lymphocytes were assessed. Then, we used Western blot to assess the expression of Bcl-6, Blimp- 1, and IL-21 proteins in PPs and used RTPCR to assess the expression of IL-21 and TGF-β1 mRNA. TFH cells coculture with spleen cells to measure the degree of IL-21 and the ratio of activation marker CD69 on the TFH cells. Naive B cells (CD27-IgD+) from children suffering from IgAN were cultured with TFH cell-related cytokines. The supernatant was detected to assess the excretion of galactose-deficient IgA1 (Gd-IgA1). IgAN mice developed noticeably increased degrees of IL-21 and CD69 on TFH cells than controls did, as well as higher percentages of B220+IgA+, B220+IgM+, B220+IgA+, TGF- β1, and IL-21 mRNA and Bcl-6, IL-21 proteins in PPs. The Gd-IgA1 level in the supernatant and IgAN- positive children's serum were noticeably higher than those of the healthy controls (P < 0.05). PPs provide the microenvironment to induce the production of IgA-secreting plasmablasts. TFH cells may be a key moderator to induce B cell differentiation into IgAsecreting plasmablasts and produce Gd-IgA1, which plays a significant part in IgAN's pathogenesis. It could be a new therapeutic target in the future.