Lymphocyte Apoptosis Research Articles

MONOCYTIC MYELOID-DERIVED SUPPRESSOR CELL EXPANSION AFTER CARDIAC SURGERY WITH CARDIOPULMONARY BYPASS INDUCES LYMPHOCYTE DYSFUNCTION.

Cardiac surgery with cardiopulmonary bypass (CPB) is associated with an immune paresis that predisposes to the development of postoperative infections and sepsis. Among factors responsible for CPB-induced immunosuppression, circulating myeloid-derived suppressor cells (MDSCs) have been found to induce early lymphocyte apoptosis and lymphocyte proliferation inhibition. However, the mechanisms involved are not fully understood. In this study, we found that the main lymphocyte subsets decreased significantly 24 h after cardiac surgery with CBP. As expected, cardiac surgery with CPB induced a monocytic MDSC expansion associated with an increased T-cell apoptosis and decreased proliferation capacity. Noteworthy, granulocytic MDSCs remain stable. Myeloid-derived suppressor cell depletion restored the ability of T-cell to proliferate ex vivo . After CPB, indoleamine 2,3-dioxygenase activity and IL-10 plasma level were increased such as programmed death-ligand 1 monocytic expression, whereas plasma level of arginine significantly decreased. Neither the inhibition of indoleamine 2,3-dioxygenase activity nor the use of anti-programmed death-ligand 1 or anti-IL-10 blocking antibody restored the ability of T-cell to proliferate ex vivo . Only arginine supplementation restored partially the ability of T-cell to proliferate.

Analysis of annexin V levels and cytokine status in patients with acute myocardial infarction

Aim. Study of annexin V levels and cytokine status in patients with acute myocardial infarction. Materials and methods. We examined 27 patients with Q-forming myocardial infarction who were admitted 12–24 hours after the development of the clinic, and 20 individuals in the control group. The lipid spectrum was determined using an automatic analyzer Humalizer-3000 (Human, Germany). Immunological studies were performed on a flow cytofluorimeter Navios (Beckman Coulter, USA). Lymphocyte subpopulations were evaluated using monoclonal antibodies produced by Immunotech (Coulter Corporation, USA). A set of ANNEXIN V-FITC/7 AAD (Beckman Coulter, USA) was used to isolate cells that had undergone apoptosis. Isolation of peripheral blood mononuclears was performed by centrifugation on a density gradient Ficol-hypac (Pharmacia Fine Chemicals, Sweden), interleukin-1β was determined using the Vector-Best test system (Vector-Best, Russia), and the tumor necrosis factor was determined using the RayBio Human TNF-alpha ELISA Kit (USA). Statistical data processing was performed using the program package Statistica 10.0. Results. In patients with acute myocardial infarction on the background of dyslipidemia (total cholesterol and LDL cholesterol increased, cholesterol-HDL reduced) showed a significant decrease in the total number of T-lymphocytes (СD 3+) (p < 0.05), T-suppressor (СD 8+) (p < 0.01), B-cells number (СD 19+) and the ratio of CD 4+/CD 8+ lymphocytes increased (p < 0.05). In patients with myocardial infarction there was a significant increase of apoptotic cells and рro-inflammatory cytokines involved in apoptosis (IL-1β and TNF-α) in the blood compared with the control. Conclusion. Increased spontaneous apoptosis of peripheral blood lymphocytes in patients with acute myocardial infarction indicates the systemic nature of apoptosis in acute coronary pathology, due to increased annexin-dependent apoptosis of T-lymphocytes as a result of immune disorders, which leads to activation of the cytokine system with an increased concentration of рro-inflammatory mediators (TNF-α, IL-1β). The identified dependencies require further study with the prospect of developing predictive algorithms for myocardial infarction outcomes based on indicators of annexin V and pro-inflammatory cytokines.

ПУТИ РЕАЛИЗАЦИИ АПОПТОЗА ЛИМФОЦИТОВ ЧЕЛОВЕКА, ИНДУЦИРОВАННОГО ПЕРОКСИДОМ ВОДОРОДА, И ИХ РЕГУЛИРОВАНИЕ

The changes in marker indicators of apoptotic cell death were studied: the level of functional activity of initiating caspases-8, -9, -12, membrane death receptors Fas, intracellular reactive oxygen species, cytosolic calcium, mitochondrial potential of peripheral blood lymphocytes of donors induced by exposure to hydrogen peroxide at final concentrations of 10-6 and 10-5 mol/l. Activation of initiating caspase-8 associated with membrane death receptors Fas was detected after exposure to hydrogen peroxide lymphocytes at a concentration of 10-6 mol/l. There was a significant increase in the functional activity of initiating caspase-9, the level of Fas receptors and intracellular reactive oxygen species, as well as a decrease in the concentration of cytosolic calcium and the value of the mitochondrial membrane potential 1 and 2 hours after modification of cells with hydrogen peroxide at a concentration of 10-5 mol/l compared with those for intact immunocytes. It was found that apoptosis of lymphocytes under the influence of exogenous hydrogen peroxide is realized with the participation of receptor-mediated caspase, mitochondrial (with activation of caspase-9) and p53-dependent mechanisms. The protective effect of resveratrol (10-6 mol/l) on lymphocytes after exposure to hydrogen peroxide (10-5 mol / l), which is due to its ability to reduce the level of reactive oxygen species in intact and modified hydrogen peroxide immunocytes, as well as deactivate hydrogen peroxide.

Immune dysregulation associated with co-occurring germline CBL and SH2B3 variants

BackgroundCBL syndrome is a RASopathy caused by heterozygous germline mutations of the Casitas B-lineage lymphoma (CBL) gene. It is characterized by heterogeneous clinical phenotype, including developmental delay, facial dysmorphisms, cardiovascular malformations and an increased risk of cancer development, particularly juvenile myelomonocytic leukemia (JMML). Although the clinical phenotype has been progressively defined in recent years, immunological manifestations have not been well elucidated to date.MethodsWe studied the genetic, immunological, coagulative, and clinical profile of a family with CBL syndrome that came to our observation after the diagnosis of JMML, with homozygous CBL mutation, in one of the members.ResultsVariant analysis revealed the co-occurrence of CBL heterozygous mutation (c.1141 T > C) and SH2B3 mutation (c.1697G > A) in two other members. Patients carrying both mutations showed an ALPS-like phenotype characterized by lymphoproliferation, cytopenia, increased double-negative T-cells, impaired Fas-mediated lymphocyte apoptosis, altered cell death in PBMC and low TRECs expression. A coagulative work-up was also performed and showed the presence of subclinical coagulative alterations in patients carrying both mutations.ConclusionIn the reported family, we described immune dysregulation, as part of the clinical spectrum of CBL mutation with the co-occurrence of SH2B3.

Evaluation of the antiviral effect of four plant polysaccharides against duck circovirus

Recently, outbreaks of duck circovirus (DuCV) are frequently occurring worldwide due to secondary infections caused by post infection-induced immunosuppression. Due to a lack of preventive drugs and vaccines, the waterfowl industry losses are ever increasing. In this study, we extracted Astragalus polysaccharides (APS), pine pollen polysaccharides (PPPS), Aloe vera polysaccharides (AVE), and Ficus carica polysaccharides (FCPS) from Astragalus, pine pollen, aloe, and F. carica leaves, respectively. We randomly divided 150 one-day-old Cherry Valley ducks into five groups, which were inoculated with the DuCV solution and orally administered APS, PPPS, AVE, FCPS, and phosphate buffer saline (PBS), respectively. We collected the duck immune organs and serum samples at 8, 16, 24, 32, 40, and 48 days post-infection (dpi). Using clinical symptom analysis, molecular biology experiments, and serological experiments, we proved that plant polysaccharides could (a) improve the duck immunity, (b) reduce the viral load, and (c) mitigate DuCV-induced damage to immune organs, with both APS and PPPS having significant effects. Moreover, we detected viral load and cytokines within the first 8 dpi. Since the body's innate immunity could inhibit viral replication within five days of virus infection, 1–5 dpi was the best treatment time. Among the four polysaccharides showing in vitro anti-apoptotic activity, APS and PPPS significantly inhibited the DuCV infection-induced apoptosis of peripheral blood lymphocytes. Overall, since our findings show APS and PPPS having significant anti-DuCV effects both in vivo and in vitro, they can be promising candidates for preventing DuCV infection in ducks.

Porcine circovirus type 2 induces apoptosis by exosomal miR-125a-5p targeting Bcl-2 in porcine lymphocytes

In order to investigate the apoptosis triggered by porcine circovirus type 2 (PCV2) in lymphocytes and the underlying mechanism, the levels of apoptosis and the expression levels of miRNA were examined by flow cytometry, Western blotting and real-time PCR (qPCR). The mimics or inhibitors of miR-125a-5p, an apoptosis-related miRNA, were transfected into PK-15 cells, and the apoptosis rate was examined upon overexpression or inhibition of mir-125a-5p. The target gene of mir-125a-5p was predicted by bioinformatics method, and the regulation of mir-125a-5p on the target gene was analyzed by luciferase reporter assay. The expressions of Bcl-2, Bax, cytochrome C and caspase-3 were detected by Western blotting. The results showed that exosomes secreted by PK-15 cells infected with PCV2 significantly increased the lymphocyte apoptosis rate, which was dose-dependent in certain concentration range. The expression of miR-125a-5p was dramatically increased. The apoptosis rate was increased significantly in the cells transfected with miR-125a-5p. It was predicted that there were binding sites of miR-125a-5p at Bcl-2 3'UTR by TargetScan. The luciferase activity of wild-type pmir-Bcl-2 3'UTR was inhibited significantly by miR-125a-5p mimics, but that of mutant pmir-Bcl-2 3'UTR was not changed. By Western blotting, Bcl-2 was reduced significantly, while Bax, cytochrome C and caspase-3 increased significantly, and the ratio of Bcl-2/Bax was significantly decreased. These results showed that PCV2 up-regulated the expression of miR-125a-5p through exosomes, then inhibited the expression of Bcl-2 at both mRNA and protein level, activated mitochondrial apoptosis pathway and induced apoptosis in lymphocytes.

Effects of anti-stress agents on the growth performance and immune function in broiler chickens with vaccination-induced stress

ABSTRACT The aim of this study was to evaluate the effects of anti-stress agents on the growth performance and immune function of broilers under immune stress conditions induced by vaccination. A total of 128, 1-day-old Arbor Acres broilers were randomly divided into four groups. Group normal control (NC) was the control group. Group vaccination control (VC), T 0.5%, and T 1% were the treatment groups, which were nasally vaccinated with two doses of the Newcastle disease virus (NDV) vaccine. The chicks in groups T 0.5% and T 1% were fed conventional diets containing 0.5% and 1% anti-stress agents. Thereafter, these broilers were slaughtered on 1, 7, 14, and 21 days post-vaccination. The results indicated that anti-stress agents could significantly reduce serum adrenocorticotropic hormone (ACTH) (P < 0.01) and cortisol (CORT) (P < 0.05) levels, and improve the growth performance (P < 0.05) and immune function of broilers (P < 0.05); However, the levels of malondialdehyde (MDA) (P < 0.05) were decreased, and the decreased total antioxidant capacity (T-AOC) (P < 0.01) levels mediated by vaccination were markedly improved. In addition, anti-stress agents could attenuate apoptosis in spleen lymphocytes (P < 0.01) by upregulating the ratio of Bcl-2 to BAX (P < 0.01) and downregulating the expression of caspase-3 and -9 (P < 0.01), which might be attributed to the inhibition of the enzymatic activities of caspase-3 and -9 (P < 0.05). In conclusion, anti-stress agents may improve growth performance and immune function in broilers under immune-stress conditions. RESEARCH HIGHLIGHTS Investigation of effects and mechanism of immune stress induced by vaccination. Beneficial effect of anti-stress agents on growth performance, immune function, oxidative stress, and regulation of lymphocyte apoptosis. Demonstration of the effects of apoptosis on immune function in the organism.

Activated T cells are the cellular source of IL-22 that enhances proliferation and survival of lymphocytes in Nile tilapia

As a pleiotropic cytokine mainly secreted by CD4+ T cells, interleukin (IL)-22 plays an important role in immune regulation and infection elimination. Despite IL-22 homologues have been identified in non-mammal, whether and how IL-22 participates in the adaptive immune response of early vertebrates have not been fully addressed. In this study, we identified an evolutionarily conserved IL-22 from Nile tilapia Oreochromis niloticus (defined as OnIL-22), proved by its properties regarding sequence, gene structure, functional domain, tertiary structure and phylogeny. IL-22 was broadly expressed in lymphoid-related tissues of tilapia, and with relatively higher levels in skin, gill, intestine and liver. The expression of OnIL-22 in spleen lymphocytes was markedly induced at the adaptive immune stage after Streptococcus agalactiae infection. Moreover, once lymphocytes were activated by PMA plus ionomycin or T-cell specific mitogen PHA in vitro, OnIL-22 expression was obviously up-regulated at both mRNA and protein levels. These results thus suggest that activated T cells produce IL-22 to take part in the adaptive immune response of tilapia. Furthermore, treatment of lymphocytes with recombinant OnIL-22 increased the expression of genes related to proliferation and survival, and further promoted the proliferation and reduced the apoptosis of lymphocytes during bacterial infection or T-cell activation. These cellular effects of IL-22 seem to be associated with JAK1/STAT3 axis downstream of IL-22, because IL-22 application not only elevated the mRNA expression of JAK1 and STAT3, but also enhanced their phosphorylation in lymphocytes. Altogether, we suggest that activated T cells produce IL-22 to promote lymphocyte proliferation and survival probability via JAK1/STAT3 signaling pathway, thus participating in adaptive immune response of Nile tilapia. Our study therefore provides helpful perspective for understanding the function and mechanism of adaptive immune system in teleost.

Oil-producing enterprise

Arterial hypertension is an urgent health problem worldwide causing an increase in temporary and permanent disability, invalidity and mortality due to cardiovascular diseases. Researchers recognize the multifactorial nature of arterial hypertension, but environmental factors are of particular potential importance. The working conditions at oil production enterprises are characterized by a more pronounced influence of these factors which may predispose for early development of disadaptation disorders, functional changes in immune and humoral regulation, and, finally, for increased risk of cardiovascular diseases in people engaged in oil production. The aim of the present work was to study the features of immunity and humoral risk factors of arterial hypertension in hypertensive employees at an oil-producing enterprise. To this purpose, a comparative analysis of lymphocyte subpopulations (CD3+CD4+, CD3+CD25+), markers of apoptosis (CD3+CD95+, TNFR, p53, Bax, Annexin V-FITC+7AAD), phagocytic activity of leukocytes (absolute phagocytosis index), and the levels of vascular humoral factors (nitric oxide and homocysteine) was performed in employees of an oil production enterprise exposed to adverse production factors. The observational group consisted of employees with established episodes of increased blood pressure. A comparison group consisted of individuals without clinical manifestations of cardiovascular disease. As a result of the clinical and laboratory examination of employees at the oil-producing enterprise with arterial hypertension, some functional changes in immune regulation were revealed. This group was characterized by a significantly (p 0.05) decreased CD3+CD4+, and CD3+CD25+ lymphocyte contents, along with increased levels of regulatory CD127 lymphocytes against the comparison group (p 0.05). The workers at an oil production enterprise with arterial hypertension are characterized by decreased (p 0.05) phagocytic activity of peripheral blood leukocytes using the criteria of absolute phagocytosis. We found some signs of inhibited lymphocyte apoptosis (p 0.05), i.e., a decrease in CD95+, TNFR, and p53 over the background values, as well as increased Bax levels over the comparison group (p 0.05). However, the content of TNFR and p53 significantly (p 0.05) exceeded the reference level, regardless of previous arterial hypertension episodes. Development of the high blood pressure episodes among the employees at oi-producing plant showed a significant association (p 0.05) with elevated levels of homocysteine and nitric oxide concentrations which are known to induce endothelial dysfunction, atherogenesis and, hence, a persistent increase in blood pressure.

MTOR Modulates the Endoplasmic Reticulum Stress-Induced CD4+ T Cell Apoptosis Mediated by ROS in Septic Immunosuppression.

Introduction When sepsis attacks the body, the excessive reactive oxygen species (ROS) production can result to endoplasmic reticulum stress (ERS) and eventually cause lymphocyte apoptosis. The mammalian target of rapamycin (mTOR) is essential for regulating lymphocyte apoptosis; we hypothesized that it mediates CD4+ T cell apoptosis during ROS-related ERS. Method We, respectively, used ROS and ERS blockers to intervene septic mice and then detected ERS protein expression levels to verify the relationship between them. Additionally, we constructed T cell-specific mTOR and TSC1 gene knockout mice to determine the role of mTOR in ROS-mediated, ERS-induced CD4+ T cell apoptosis. Results Blocking ROS significantly suppressed the CD4+ T cell apoptosis associated with the reduction in ERS, as revealed by lower levels of GRP78 and CHOP. ERS rapidly induced mTOR activation, leading to the induction of CD4+ T cell apoptosis. However, mTOR knockout mice displayed reduced expression of apoptotic proteins and less ER vesiculation and expansion than what was observed in the wild-type sepsis controls. Conclusion By working to alleviate ROS-mediated, ERS-induced CD4+ T cell apoptosis, the mTOR pathway is vital for CD4+ T cell survival in sepsis mouse model.

Transcriptome profile of spleen tissues from locally-adapted Kenyan pigs (Sus scrofa) experimentally infected with three varying doses of a highly virulent African swine fever virus genotype IX isolate: Ken12/busia.1 (ken-1033)

BackgroundAfrican swine fever (ASF) is a lethal hemorrhagic disease affecting domestic pigs resulting in up to 100% mortality rates caused by the ASF virus (ASFV). The locally-adapted pigs in South-western Kenya have been reported to be resilient to disease and harsh climatic conditions and tolerate ASF; however, the mechanisms by which this tolerance is sustained remain largely unknown. We evaluated the gene expression patterns in spleen tissues of these locally-adapted pigs in response to varying infective doses of ASFV to elucidate the virus-host interaction dynamics.MethodsLocally adapted pigs (n = 14) were experimentally infected with a high dose (1x106HAD50), medium dose (1x104HAD50), and low dose (1x102HAD50) of the highly virulent genotype IX ASFV Ken12/busia.1 (Ken-1033) isolate diluted in PBS and followed through the course of infection for 29 days. The in vivo pig host and ASFV pathogen gene expression in spleen tissues from 10 pigs (including three from each infective group and one uninfected control) were analyzed in a dual-RNASeq fashion. We compared gene expression between three varying doses in the host and pathogen by contrasting experiment groups against the naïve control.ResultsA total of 4954 differentially expressed genes (DEGs) were detected after ASFV Ken12/1 infection, including 3055, 1771, and 128 DEGs in the high, medium, and low doses, respectively. Gene ontology and KEGG pathway analysis showed that the DEGs were enriched for genes involved in the innate immune response, inflammatory response, autophagy, and apoptosis in lethal dose groups. The surviving low dose group suppressed genes in pathways of physiopathological importance. We found a strong association between severe ASF pathogenesis in the high and medium dose groups with upregulation of proinflammatory cytokines and immunomodulation of cytokine expression possibly induced by overproduction of prostaglandin E synthase (4-fold; p < 0.05) or through downregulation of expression of M1-activating receptors, signal transductors, and transcription factors. The host-pathogen interaction resulted in induction of expression of immune-suppressive cytokines (IL-27), inactivation of autophagy and apoptosis through up-regulation of NUPR1 [5.7-fold (high dose) and 5.1-fold (medium dose) [p < 0.05] and IL7R expression. We detected repression of genes involved in MHC class II antigen processing and presentation, such as cathepsins, SLA-DQB1, SLA-DOB, SLA-DMB, SLA-DRA, and SLA-DQA in the medium and high dose groups. Additionally, the host-pathogen interaction activated the CD8+ cytotoxicity and neutrophil machinery by increasing the expression of neutrophils/CD8+ T effector cell-recruiting chemokines (CCL2, CXCL2, CXCL10, CCL23, CCL4, CXCL8, and CXCL13) in the lethal high and medium dose groups. The recovered pigs infected with ASFV at a low dose significantly repressed the expression of CXCL10, averting induction of T lymphocyte apoptosis and FUNDC1 that suppressed neutrophilia.ConclusionsWe provide the first in vivo gene expression profile data from locally-adapted pigs from south-western Kenya following experimental infection with a highly virulent ASFV genotype IX isolate at varying doses that mimic acute and mild disease. Our study showed that the locally-adapted pigs induced the expression of genes associated with tolerance to infection and repression of genes involved in inflammation at varying levels depending upon the ASFV dose administered.

Features of CD4+ lymphocytes apoptosis in children with cryptosporidiosis

The aim of this study was to identify the features of CD4+ lymphocytes apoptosis in children with cryptosporidiosis. Feces for the detection of cryptosporidium and venous blood for the study of lymphocytes apoptosis were the material of the study. Mononuclear leukocytes were isolated from venous whole blood and cultured in a complete culture medium. Cells were incubated for 24 hours with inducers of the receptor (TNFα) and mitochondrial pathways of apoptosis (dexamethasone).Cryptosporidia have been found in 35% of acute intestinal infections in children. The number of lymphocytes with cytoflurimetric signs of apoptosis in the group of cryptosporidiosis-positive patients did not differ from that in patients without cryptosporidiosis (p=0.421). There were no intergroup differences in the number of CD4+ lymphocytes expressing the Fas receptor on their surface (p=0.462). In cultures incubated in the presence of dexamethasone, a decrease in the number of apoptotically altered CD4+ lymphocytes was registered only in the group of cryptosporidiosispositive patients (p=0.028).The study showed that in cryptosporidiosis, the sensitivity of CD4+ cells to the induction of the mitochondrial pathway of apoptosis changes in favor of slowing down this variant of cell death.

Immunomodulatory Effects of Lepidium meyenii Walp. Polysaccharides on an Immunosuppression Model Induced by Cyclophosphamide.

Background Lepidium meyenii Walp. (Maca) has emerged as a functional plant food and traditional herb owing to its biological activities; Maca polysaccharides as an important active component of Maca have good immunomodulatory effect; however, studies on the immunomodulatory effect of Maca polysaccharides are mainly focused on macrophages; little attention has been devoted to the mechanisms and other immune cells. This study is aimed at investigating the immunomodulatory effects and mechanisms of Maca polysaccharides. Methods Sixty mice were divided into five groups, and the mice were injected with cyclophosphamide to establish an immunosuppression model except for those in the common group. The body weights were measured, as well as immune-related indices, such as organ indices, haematological parameters, lymphocyte cycle, and proliferation, cytokine, and protein expression levels. Results The weight loss and immune organ index decline caused by cyclophosphamide could be reversed by MP. Furthermore, MP increased WBC and HGB counts and reduced the ratio of G0/G1 phase obviously, increased the proportion of S phase and G2/M phase in peripheral blood lymphocytes, increased the counts of CD4+ T cells and the ratio of CD4+/CD8+, and reduced the inhibition rate of splenic lymphocytes. MP affected the production of cytokines by increasing IFN-γ, TNF-α, and IL-2 levels and by decreasing IL-4 levels. MP increased the mRNA expression of T-bet and the protein expression of Bcl-2 in the spleen and decreased the protein expression of caspase-3 and Bax. Conclusions Maca polysaccharides might be the basic material for Maca's immunomodulatory effect. The mechanism was perhaps related to inhibiting lymphocyte apoptosis and promoting the balance of Th1/Th2 cell subsets.

Wnt/β-Catenin Protects Lymphocytes from HIV-Mediated Apoptosis via Induction of Bcl-xL.

HIV infection mediates the apoptosis of lymphocytes, the mechanism of which is multifaceted. Here, we evaluated the role of Wnt/β-catenin signaling in HIV-associated T cell apoptosis, as Wnt/β-catenin regulates the transcriptional activity of genes impacting apoptosis. We specifically investigated the role of the Wnt/β-catenin pathway in the HIV-associated apoptosis of CD4+ T cells and CD4dimCD8bright T cells, a population that is infected by HIV. We found that the induction of β-catenin, via a 6-bromoindirubin-3-oxime (BIO), significantly rescued HIV-infected CD4+ and CD4dimCD8bright T cells from apoptosis by >40–50%. Further, a small-molecule inhibitor of the Wnt/β-catenin pathway (PNU-74654) reversed BIO-mediated protection from HIV-associated apoptosis. BIO also induced Bcl-xL, an anti-apoptotic protein, and a target gene of Wnt/β-catenin, in CD4+ and CD4dimCD8bright T cells by approximately 3-fold. Inhibiting Bcl-xL by WEHI-539 abrogated β-catenin-mediated apoptotic protection in infected CD4+ and CD4dimCD8bright T cells. Collectively, these findings demonstrate that engaging Wnt/β-catenin signaling in HIV-infected T cells protects them from HIV-associated apoptosis by inducing Bcl-xL.

Radioprotective Effect of Rosmarinus officinalis L (Rosemary) Essential Oil on Apoptosis, Necrosis and Mitotic Death of Human Peripheral Lymphocytes (PBMCs).

Background: Rosemary plant, with phenolic compounds, is known as an antioxidant herb and able to scavenge free radical agents in the biological environment; therefore, it is expected that the rosemary essential oil (R-EO) shows the radioprotective effect to protect individuals who are physically in contact with ionizing radiation.Objective: This study aimed to assess the radioprotective effect of R-EO on human peripheral blood mononuclear cells (PBMCs).Material and Methods: In this experimental study, the toxicity of the rosemary essential oil on PBMCs was assessed by the 3-(4, 5-dimethylthiazolyl-2)-2, 5-diphenyltetrazolium bromide (MTT) assay. The cells were irradiated at 0. 25 and 200 cGy using a 6 MV X-ray linear accelerator. The survival, apoptosis, necrosis, and survival enhancement factors of cells were analyzed by MTT and flow cytometry analyses with a non-toxic concentration of the rosemary essential oil (IC10).Results: Irradiation of cells in the presence of R-EO caused a significant increase in cell survival compared with the control in both 25 and 200 cGy radiation doses. Also, the percentages of apoptosis and necrosis of cells showed a significant decrease compared with the control.Conclusion: Rosemary essential oil as a natural and non-toxic compound could show favorable radioprotective effects in such a way that significantly increases the survival rate and decreases the percentage of apoptosis and necrosis of PBMCs.

Tim-3 Blockade Decreases the Apoptosis of CD8+ T Cells and Reduces the Severity of Sepsis in Mice

IntroductionThe T cell immunoglobulin and mucin domain 3 (Tim-3) mediated immunosuppressive pathway has been shown to play an essential role in the development of sepsis. However, the influence of Tim-3 blockade during sepsis and the possible effects on T cells’ function remains largely unknown. Our study investigates the role of Tim-3 in cecal ligation and puncture (CLP)-induced sepsis in mice. MethodsSepsis was induced in C57BL/6 male mice via CLP. The expression of Tim-3 in CD8+ T cells after CLP challenge was measured. A dose of 50 μg anti-Tim-3 antibodies was injected intraperitoneally 30 min after surgery. Postoperative survival, bacterial clearance in the blood and peritoneal lavage fluid, cytokine secretion in the blood, and lung and liver histology were evaluated. In addition, the apoptosis of immune cells in the spleen and thymus was examined, respectively. ResultsTim-3 expression was elevated in the splenic CD8+ T cells of septic mice. At the early stage of CLP-induced sepsis, blocking Tim-3 with anti-Tim-3 antibodies reduced the severity of sepsis. The anti-Tim-3 antibodies alleviated the morphology of lung and liver injuries in septic mice. The anti-Tim-3 antibodies also reduced the severity of the inflammatory responses and lymphocyte apoptosis in septic mice. ConclusionsAnti-Tim-3 antibodies might be a potential therapeutic strategy for sepsis.

Toddler With Fever and Limp.

Toddler With Fever and Limp.

Regulating effects of triptriolide on the proliferation and apoptosis of splenic lymphocyte in vivo and in vitro

ObjectiveTriptriolide (T11) is isolated from the Chinese herb Tripterygium wilfordii Hook F (TwHF), which has been used as an immunosuppressive reagent for many years in China, while the role of this compound in TwHF is still unknown. In this study, we try to investigate the immune regulating effects of T11 on the proliferation and apoptosis of splenic lymphocyte in vivo and in vitro. Methodswe pretreated the BALB/c mice with T11 (2.8, 14, 28 mg/kg) and prednisone acetate (PA, 1.3 mg/kg, positive control) for 7 days before lipopolysaccharides (LPS, 3 mg/kg) stimulation. Then the histopathologic change of spleen was examined by H&E method, and the apoptosis of splenic lymphocyte was assessed by flow cytometry method. In addition, we further studied the inhibition effect of T11 (50, 100, 150, 200 µg/mL) on the proliferation and activity of splenic lymphocyte, which was with or without the stimulation of concanavalin A (Con A, 5 µg/mL) and lipopolysaccharides (LPS, 5 µg/mL) in vitro, respectively. ResultsWe found that T11 not only can restore the pathogenesis process in spleen, and boost the splenic lymphocyte apoptosis in vivo, but also can inhibit the splenic lymphocyte proliferation and affect its viability in vitro. ConclusionThe results indicated that T11 might modulate the immune function of LPS-induced BALB/c mice by inhibiting the proliferation and inducing the apoptosis of splenic lymphocyte. Therefore, T11 could be a novel immunomodulator in autoimmune disease treatment in clinical.

Electroacupuncture at "Zusanli"(ST36) protects intestinal mucosal immune barrier by suppre-ssing apoptosis of intestinal lymphocytes and regulating expression of Bcl-2 and Bax in sepsis rats

To observe the effect of electroacupuncture (EA) at "Zusanli" (ST36) on apoptosis of intestinal T lymphocytes, translocation of intestinal bacteria and expression of intestinal Bcl-2 and Bax proteins and intestinal mucosal immune barrier in sepsis rats, so as to explore its underlying mechanism in relieving sepsis. SD rats were randomly divided into sham operation (n=6), model (n=15), non-meridian and non-acupoint (non-acupoint, n=15) and acupoint EA(n=15) groups by using random number table method. The sepsis model was established by using cecal ligation and perforation(CLP) method. EA (2 Hz, 2 mA) was applied to bilateral ST36 or non-acupoint for 30 min one hour after modeling, once every day for 3 days. The rats' general conditions and fatality rate in 3 days after modeling were recorded. The liver, spleen and mesenteric lymph nodes were taken for bacterial culture to detect the translocation rate of intestinal bacteria. The small intestinal tissue was taken for observing histopathological changes (Chiu's score: 0-5 points) after HE staining, and for determining the expression levels of Bcl-2 and Bax proteins using Western blot. The intestinal mucosa was sampled for detecting the apop-tosis (apoptotic index) of lymphocytes by using terminal deoxynucleoitidyl transferase-mediated deoxyuridine triphosphate biotin nick end labeling (TUNEL) assay, and the counts of CD4+ and CD8+T cells using flow cytometry. The contents of IL-4 in the small intestine and that of secretory IgA (sIgA) in the small intestinal mucus were determined by using ELISA. After modeling, of the 15 rats in each of the 3 groups, 7, 7 and 2 in the model, non-acupoint and EA groups were dead in the first 3 days, with the fatality rate being 46.67% (7/15), 46.67% (7/15) and 13.33% (2/15), respectively (being obviously lower in the EA group than in the former two groups, P<0.05). Compared with the sham operation group, the incidence of intestinal bacterial translocation, apoptotic index, Chiu's score, and Bax expression were significantly increased (P<0.05), and the percentages of CD4+ and CD8+T cells, IL-4 and sIgA contents and Bcl-2 expression considerably decreased (P<0.05) in the model group. In comparison with the model group, modeling-induced increase of incidence of bacterial translocation, apoptotic index and Bax expression, and decrease of percentages of CD4+ and CD8+T cells, IL-4 and sIgA contents and Bcl-2 expression were reversed (P<0.05) in the EA group. EA at ST36 can reduce death rate and intestinal bacteria translocation incidence in sepsis rats, which may be related to its functions in regulating the expression of intestinal Bcl-2 and Bax proteins and inhibiting the apoptosis of intestinal mucosal T lymphocytes, thereby protecting the immune barrier function of intestinal mucosa to reduce the intestinal permeability.

Fumonisin B1 Induces Immunotoxicity and Apoptosis of Chicken Splenic Lymphocytes

Fumonisin B1 (FB1), produced by Fusarium, is among the most abundant and toxic mycotoxin contaminations in feed, causing damages to the health of livestock. However, the mechanisms of FB1 toxicity in chickens are less understood. As splenic lymphocytes play important roles in the immune system, the aim of this study was to investigate the immunotoxic effects and mechanisms of FB1 on chicken splenic lymphocytes. In the present study, the chicken primary splenic lymphocytes were harvested and treated with 0, 2.5, 5, 10, 20 and 40 μg/mL FB1. Then, the cell proliferation, damage, ultrastructure, inflammation and apoptosis were evaluated. Results showed that the proliferation rate of splenic lymphocytes was decreased by FB1 treatments. The activity of lactate dehydrogenase (LDH) was increased by FB1 treatments in a dose-dependent manner, implying the induction of cell damage. Consistently, the ultrastructure of splenic lymphocytes showed that FB1 at all the tested concentrations caused cell structure alterations, including nuclear vacuolation, mitochondrial swelling and mitochondrial crest fracture. Besides, immunosuppressive effects of FB1 were observed by the decreased concentrations of interleukin-2 (IL-2), IL-4, IL-12 and interferon-γ (IFN-γ) in the cell culture supernatant. Furthermore, apoptosis was observed in FB1-treated cells by flow cytometry. The mRNA expressions of apoptosis-related genes showed that the expression of Bcl-2 was decreased, while the expressions of the P53, Bax, Bak-1, and Caspase-3 were increased with FB1 treatment. Similar results were found in the concentrations of apoptosis-related proteins in the cell supernatant by ELISA assay. Moreover, regression analysis indicated that increasing FB1 concentration increased LDH activity, concentrations of Bax, Bak-1 and mRNA expression of Bak-1 linearly, increased M1 area percentage quadratically, decreased concentration of IFN-γ, mRNA expression of Bcl-2 linearly, and decreased concentrations of IL-2 and IL-4 quadratically. Besides, regression analysis also showed reciprocal relationships between IL-12 concentration, Caspase-3 mRNA expression and increasing FB1 concentration. The increasing FB1 concentration could decrease IL-12 concentration and increase Caspase-3 mRNA expression. Altogether, this study reported that FB1 induced the immunotoxicity of chicken splenic lymphocytes and caused splenic lymphocytes apoptosis by the Bcl-2 family-mediated mitochondrial pathway of caspase activation.