Number Of Splenocytes Research Articles

Effect of Pulsed Electromagnetic Field Stimulation on Splenomegaly and Immunoglobulin E Levels in 2,4-Dinitrochlorobenzene-Induced Atopic Dermatitis Mouse Model

Effects of pulsed electromagnetic fields (PEMF) on immunological factors in a 2,4-dinitrochlorobenzene (DNCB)-induced atopic dermatitis (AD) model were investigated. Hairless mice were randomly assigned to control, acetone and olive oil solution-treated (AOO), PEMF 15 Hz, PEMF 75 Hz, and sham groups (n = 5 each). AOO solution was used to dissolve DNCB. Both PEMF and sham groups were exposed to similar DNCB doses, causing similar AD symptoms. After AD induction for five weeks, only the PEMF groups were exposed to PEMF stimulations (15 Hz, 75 Hz, and 15 mT) inside the solenoid coil, for two weeks. In both groups, splenomegaly was observed, as AD was induced by hyperimmune reactions caused by DNCB sensitization. However, splenomegaly did not occur in the PEMF-exposed groups, and spleen weight decreased similarly to that of the control. Hence, the total splenocytes in the PEMF group were similar to those in the control group, whereas the sham group showed three times the number of splenocytes compared with the PEMF group. The serum immunoglobulin E levels did not significantly change in the PEMF group; however, they increased more than fourfold in the sham group. These results demonstrate that PEMF stimulation ameliorated the abnormal symptoms caused by hyperimmune reactions.

ОЦЕНКА ИММУНОМОДУЛИРУЮЩЕГО ДЕЙСТВИЯ СУХОГО ЭКСТРАКТА ТРАВЫ TRIFOLUM PRATENSE L.

The purpose of the study was to determine the optimal concentration of dry extract of the herb Trifolium pratense L. to reduce the immunosuppressive effect of methotrexate on the lymphoid organs of mice. Materials and methods. The immunomodulatory effect of the dry extract of the herb Trifolium pratense L. was assessed by the ability to bring the weight and cellularity of the lymphoid organs of the immune system of mice when exposed to methotrexate to the level of intact animals. Experimental studies were performed on 129 mature outbred white male mice. Immunomodulatory activity was assessed by determining the mass of the thymus and spleen, the number of thymocytes, splenocytes and myelokaryocytes. Results. A study of the immunomodulatory properties of the dry extract of the herb Trifolium pratense L. revealed that this phytosubstance in the selected dosage of 50 mg/kg did not change the weight and number of cells in the thymus and spleen of mice, including the number of bone marrow cells, while restoring the weight and cellularity of the immune system organs of mice exposed to methotrexate, which is comparable to the effect of the immunomodulator «Immunal». Conclusion. Dry extract of the herb Trifolium pratense L. at an optimal dosage of 50 mg/kg can reduce the immunosuppressive effect of methotrexate, which may be due to the high content of phenolic compounds. This herbal medicine can be considered as an immunomodulatory herbal remedy.

Anti-tumoral Immunity and Chemo-preventive Effectiveness of Herbal Extracts of Curcumin, Ginger, Clove and Amygdaline in Ehrlich Ascites Carcinoma-Challenging Mice.

Due to its systemic toxicity, traditional chemotherapy of tumors is being taken into consideration. Herbal therapy, containing phytochemical polyphenol derivatives such as Curcumin (Cur), Ginger (Gin), Cloves (Clov) and Amygdaline (Amyg), is one of the numerous complementary and alternative approaches as an anti-cancer therapy and holds great promise for cancer chemo-prevention with fewer side effects. The current study was designated to assess anti-tumoral immunity and anti-cancer and chemo-preventive effectiveness of herbal extracts of Cur, Ginger, Clov and Amyg in Ehrlich Ascites Carcinoma (EAC)-challenging mice. Chemo-preventive efficacy of herbal extracts of Cur, Gin, Clov and Amyg were analyzed in vivo by examination of the apoptosis rate of EAC tumor cells by flow cytometry. The total numbers of EAC cells, splenocytes counts and leucocytes count with their differentials relative % in peripheral blood (PB) of EACchallenging mice were investigated. EAC-challenging mice treated with herbal extracts of Cur, Gin, Clov and Amyg showed a marked decline in EAC tumor cell count and a noticeable increase in apoptosis rate of EAC tumor cells, a remarkable decrease in serum level of cancer antigen 125 (CA-125) with an obvious increase in the number of splenocytes comparing to that in EAC-challenging mice treated with PBS alone. Moreover, the data indicated an insignificant change in the total leucocytes count and their differentials relative % of eosinophil, neutrophils, monocytes and lymphocytes in EAC-challenging mice treated with Cur and Amyg, but these parameters were markedly increased in EAC-challenging mice injected with Gin and Clov compared to that in EAC-challenging mice treated with PBS alone. To conclude, the herbal extracts of Cur, Gin, Clov and Amyg may have anti-tumoral immunity and anti-cancer potency and potential to reduce the resistance to cancer conventional chemotherapy and exert cancer chemo-protective approaches with low adverse effects. Further research is necessary to determine the regimen's toxicity on various tissues and organs and to connect the diagnostic and therapeutic approaches used in the regimen's biomedical use.

Sex differences in donor T cell targeting of host splenocyte subpopulations in acute and chronic murine graft-vs.-host disease: implications for lupus-like autoimmunity.

This study sought to compare in vivo sex differences in either a Th1-dominant CTL response or a Tfh-mediated lupus-like antibody response using the parent-into F1 murine model of acute or chronic GVHD respectively. In acute GVHD we observed no significant sex differences in the hierarchy of donor CD8 CTL elimination of splenocyte subsets. B cells were the most sensitive to elimination in both sexes; however, the male response was significantly stronger. Sex differences in chronic GVHD were more widespread; females exhibited significantly greater numbers of total splenocytes and host CD4 Tfh cells, B cells and CD8 T cells consistent with reports of greater female autoantibody production in this model. The more potent male CTL response in acute GVHD conflicts with reports of greater female CTL responses following infections or vaccines and may reflect the absence of exogenous innate immune stimuli in this model.

Effect on the splenocyte function of weaned piglets induced by continuous lipopolysaccharide injections.

When piglets are exposed to pathogens for a long period, the immune system organs, among them the spleen, play a major role in combating the stress caused by those pathogens. In the present study, the effect on splenocyte function was investigated in a model of weaned piglets in which stress was induced by multiple low doses of lipopolysaccharide (LPS). Forty-eight 28-day-old piglets were divided into two groups: the LPS group and the control group. During the experimental period of thirteen days, the LPS group was intraperitoneally injected with LPS (100 μg/kg) once per day, and the control group was injected with the same volume of 0.9% sterile saline. On the 1st, 5th, 9th and 13th days, the piglets' spleens were collected for isolating splenocytes. The proliferation ability of splenocytes was evaluated by the cell-counting-kit 8 method. Flow cytometry was used to detect cell cycle stage and apoptosis, and the nitric oxide level of cell supernatant was also tested. In the experimental group, the proliferation ability of splenocytes was enhanced, the proportion of cells in the G0/G1 phase was smaller, and cells were promoted to the S and G2/M phases. Meanwhile, apoptosis was suppressed and nitric oxide release upregulated. The results were significantly different between the LPS group and the control group on the 5th and 9th days. The difference between the results of one group and those of the other suggest that after the 5th LPS injection, multiple low doses of LPS activated splenocytes and restored the number of splenocytes, which maintained and possibly enhanced the regulation of the immune function of the spleen.

B cell-derived IL-10 is an autocrine survival factor and maintains immune homeostasis in the spleen

Abstract IL-10 is a key anti-inflammatory cytokine and IL-10 KO mice develop colitis. Mice with a global IL-10 KO or constitutive B cell-specific IL-10 KO (CD19-Cre.IL-10fl/fl) have otherwise normal B cells. Surprisingly, we found that within 14-21 days, mice with a tamoxifen (Tam) -induced B cell specific deletion of IL-10R (hCD20-ERT2-Cre.IL-10Rfl/fl) had a 50% decrease in splenic B cell number vs. Cre-controls. B cell frequency was unchanged due to a concomitant fall in both T cells and innate cells resulting in a 58% fall in splenocyte number. Plasma cells, CD4+ and CD8+ T cells, and neutrophils were disproportionately decreased, resulting in a 20-40% reduction in frequency. A similar but somewhat less severe loss of B cells/splenocytes was seen in Tam-treated hCD20-ERT2-Cre.IL-10fl/fl mice. Adoptive transfer competition studies in immunized µMT hosts treated with Tam revealed a selective loss of hCD20-ERT2-Cre.IL-10fl/fl vs. CD19-Cre.IL-10fl/fl B cells. Furthermore, by day 21, acute loss of B cell IL-10 or IL-10R resulted in ~20% weight loss and colon length, consistent with colitis (pathology pending). In conclusion, IL-10 is an autocrine B cell survival factor and acute loss of B cell IL-10 or IL-10R is associated with a marked contraction of B cells and associated collapse in T cells and innate cells in the spleen, resulting in immune dysregulation and colitis. This requirement for B cell IL-10 has not been previously recognized because it is compensated for developmentally.

Simultaneous deletion of ORMDL1 and ORMDL3 proteins disrupts immune cell homeostasis.

ORMDL3 is a prominent member of a family of highly conserved endoplasmic reticulum resident proteins, ORMs (ORM1 and ORM2) in yeast, dORMDL in Drosophila and ORMDLs (ORMDL1, ORMDL2, and ORMDL3) in mammals. ORMDL3 mediates feedback inhibition of de novo sphingolipid synthesis. Expression levels of ORMDL3 are associated with the development of inflammatory and autoimmune diseases including asthma, systemic lupus erythematosus, type 1 diabetes mellitus and others. It has been shown that simultaneous deletions of other ORMDL family members could potentiate ORMDL3-induced phenotypes. To understand the complex function of ORMDL proteins in immunity in vivo, we analyzed mice with single or double deletions of Ormdl genes. In contrast to other single and double knockouts, simultaneous deletion of ORMDL1 and ORMDL3 proteins disrupted blood homeostasis and reduced immune cell content in peripheral blood and spleens of mice. The reduced number of splenocytes was not caused by aberrant immune cell homing. A competitive bone marrow transplantation assay showed that the development of Ormdl1-/-/Ormdl3-/- B cells was dependent on lymphocyte intrinsic factors. Highly increased sphingolipid production was observed in the spleens and bone marrow of Ormdl1-/-/Ormdl3-/- mice. Slight, yet significant, increase in some sphingolipid species was also observed in the spleens of Ormdl3-/- mice and in the bone marrow of both, Ormdl1-/- and Ormdl3-/- single knockout mice. Taken together, our results demonstrate that the physiological expression of ORMDL proteins is critical for the proper development and circulation of lymphocytes. We also show cell-type specific roles of individual ORMDL family members in the production of different sphingolipid species.

Treatment with lysophosphatidic acid improves glomerulonephritis through the suppression of macrophage activation in a murine model of systemic lupus erythematosus.

Several therapeutic agents have been developed and used for the clinical treatment of systemic lupus erythematosus (SLE). In cases where SLE is accompanied by severe organ failures, such as neuropsychiatric lupus erythematosus (NPSLE) and acute onset of lupus nephritis, the use of potent immunosuppressive drugs, such as cyclophosphamide, is necessary. However, potent immunosuppressive drugs are known to increase infection risks. Thus, the development of therapeutic agents with novel mechanisms is urgently required. Previously, we reported that treatment with lysophosphatidic acid (LPA) prevents depression-like behaviours by suppressing microglial activation in MRL/lpr mice. In this study, we examined whether the treatment with LPA improves glomerulonephritis by affecting systemic immunity in MRL/lpr mice. Eighteen-week-old MRL/lpr mice were treated with a vehicle or LPA for 3 weeks. After treatment, the glomerular inflammation and damage parameters were compared between the 2 groups. Moreover, we examined the effects of LPA on immune cells by flow cytometry using isolated splenocytes. LPA treatment in MRL/lpr mice significantly reduced the daily urinary albumin content and suppressed the CD68-positive cells and Periodic acid-Schiff (PAS)-positive areas in the glomeruli. The treatment also suppressed plasma anti-dsDNA antibodies and inflammatory cytokines in MRL/lpr mice. Although LPA did not significantly affect the total number of splenocytes, the treatment significantly reduced CD11b+Ly6G-Ly6C- cells (mature macrophages), as well as CD11b+Ly6G-Ly6C-CD68+ cells (activated mature macrophages). These results suggest that LPA may improve glomerulonephritis by suppressing macrophage activation in MRL/lpr mice.

The mechanisms underlying alcohol-induced decreased splenic size: A network meta-analysis study.

Organ weight change is widely accepted as a measure of toxicologic pathology. We and other groups have shown that excessive alcohol exposure leads to decreased spleen weight in rodents. This study explores the mechanisms underlying alcohol-induced splenic injury through a network meta-analysis. QIAGEN Ingenuity Pathway Analysis (IPA) and Mammalian Phenotype (MP) Ontology were used to identify alcohol-related molecules associated with the small spleen phenotype. Search Tool for the Retrieval of Interacting Genes/Proteins (STRING) and IPA bioinformatics tools were then used to analyze the biologic processes and enriched signaling pathways engaging these molecules. In addition, the "downstream effects analysis" algorithm was used to quantify alcohol's effects. IPA identified 623 molecules affected by alcohol and a Venn diagram revealed that 26 of these molecules overlapped with those associated with the MP Ontology of small spleen. The 26 molecules are TGFB1, CASP8, MTOR, ESR1, CXCR4, CAMK4, NFKBIA, DRD2, BCL2, FAS, PEBP1, TRAF2, ATM, IGHM, EDNRB, MDM2, GLRA1, PRF1, TLR7, IFNG, ALOX5, FOXO1, IL15, APOE, IKBKG, and RORA. Some of the 26 molecules were also associated with the MP Ontology of abnormal white pulp and red pulp morphology of the spleen, abnormal splenic cell ratio, decreased splenocyte number, abnormal spleen physiology, increased splenocyte apoptosis, and reduced splenocyte proliferation. STRING and IPA "Core Analysis" showed that these molecules were mainly involved in pathways related to cell apoptosis, proliferation, migration, and immune responses. IPA's "Molecular Activity Predictor" tool showed that concurrent effects of activation and inhibition of these molecules led to decreased spleen size by modulating apoptosis, proliferation, and migration of splenocytes. Our network meta-analysis revealed that excessive alcohol exposure can damage the spleen through a variety of molecular mechanisms, thereby affecting immune function and human health. We found that alcohol-mediated splenic atrophy is largely mediated by increased apoptosis signaling, migration of cells, and inhibition of splenocyte proliferation.

Effect of the ati Gene Deletion on the Pathogenicity and Immunogenicity of the Vaccinia Virus.

Among the nonvirion proteins of the vaccinia virus (VACV), a 94-kDa long protein is most abundantly present; the protein is a truncated form of the 150-kDa A-type inclusion (ATI) protein of the cowpox virus encoded by the ati gene. This VACV protein does not form intracellular ATIs, being as it is a major immunogen upon infection/immunization of humans or animals with the VACV. Antibodies specific to this protein are not virus-neutralizing. The present study focused on the effect of the production of this nonstructural major immunogenic VACV protein on the manifestation of pathogenicity and immunogenicity of the virus in the BALB/c mouse model of infection. In order to introduce a targeted deletion into the VACV LIVP genome, the recombinant integration/deletion plasmid pΔati was constructed and further used to generate the recombinant virus LIVPΔati. The pathogenicity of the VACV LIVP and LIVPΔati strains was studied in 3-week-old mice. The mice were intranasally infected with the viruses at a dose of 107 pfu; 50% of the animals infected with the parent LIVP strain died, while infection with the LIVPΔati strain led to the death of only 20% of the mice. Intradermal vaccination of mice aged 6- weeks with the LIVPΔati virus statistically significantly increased the production of VACV-specific IgG, compared to that after intradermal vaccination with VACV LIVP. Meanwhile, no differences were noted in the cell-mediated immune response to the vaccination of mice with VACV LIVP or LIVPΔati, which was assessed by ELISpot according to the number of splenocytes producing IFN-γ in response to stimulation with virus-specific peptides. Intranasal infection of mice with lethal doses of the cowpox virus or the ectromelia virus on day 60 post-immunization with the studied VACV variants demonstrated that the mutant LIVPΔati elicits a stronger protective response compared to the parent LIVP.

Effect of tetrapeptide Acetyl-(D-Lys)-Lys-Arg-Arg-amide on immunological and biochemical parameters of Wistar rats using passive smoking models

Tetrapeptides, the homologues of adrenocorticotropic hormone fragment (15-18), play a special role among the bioregulators of the immune system. These compounds have cerebroprotective, neuroprotective and antioxidant properties. However, no available studies concerned the immunotropic properties of tetrapeptides in the models with exposure to xenobiotics, thus making such research quite relevant. Our study concerned the effects of tetrapeptide Acetyl-(D-Lys)-Lys-Arg-Arg-amide (laboratory code KK1) on the immunological and biochemical parameters of 72 female Wistar rats exposed to passive smoking. The experimental animals were fumigated with tobacco smoke for 8 hours. Synthetic peptide KK1 was administered intranasally at a dose of 40 g/kg/day five times a day for 10 days. Cotinine was detected only in blood serum of rats from experimental groups, thus confirming a contribution of this tetrapeptide to the trend for normalization of some immunological parameters in experimental animals subjected to passive tobacco smoking, expressed as an increase in thymus mass and the number of splenocytes, and a decrease in the circulating immune complexes compared to the parameters of smoking rats of the group. We revealed that passive tobacco smoking in rats was accompanied by a general tendency to accumulation of iron, lead and nickel in peripheral blood. There was a marked increase in the concentration of cadmium, lead and cobalt in rats of the experimental group compared with the content of these trace elements in the liver of smoking animals injected with tetrapeptide KK1. The revealed shifts in immunological indices may be based, firstly, on hepatotoxic effect of ecotoxicants, The lymphoid lineage is mostly affected thus causing hypoplasia of the central and peripheral immunity organs. An evident sign of such pathology is a decreased cellularity of hematopoietic and lymphoid organs found in the present study. Secondly, the tobacco smoke components with prooxidant action may disrupt cellular redox homeostasis, causing damage to cell membranes, resulting in necrosis or apoptosis, thus explaining the revealed decrease in the number of thymocytes, splenic karyocytes and a decrease in the weight of organs. Thus, our results suggest usage of experimental passive smoking in order to evaluate efficiency of the tetrapeptides. Administration of Acetyl-(D-Lys)-Lys-Arg-Arg-amide peptide to the passively smoking rats is associated with tendency to normalize the mass of the thymus and spleen, the number of thymocytes and splenocytes, and a decrease in circulating immune complexes. Further studies are required to elucidate the effects of tetrapeptides upon the immune system.

Immunomorphological Evaluation of Pregnant Rats Exposed to Passive Smoking and their Offspring (Neuroendocrine Aspects)

The aim of the study was to comprehensively assess histological parameters of the hypothalamic neurosecretory and immune systems in pregnant rats exposed to passive smoking and their offspring.Material and methods. We studied morphological and immunological parameters of pregnant Wistar rats exposed to passive smoking and those of the control group, as well as their offspring. The obtained material was processed using histological, immunohistochemical, morphometric and immunological methods.Results. The results obtained demonstrated that in rats exposed to passive smoking, the sizes of neurosecretory cells (NSCs) of the supraoptic (SO) and paraventricular (PV) nuclei of the hypothalamus increased, the number of p53 positive NSCs increased, and bcl-2 protein expression decreased. Tobacco smoking caused formation of a proaptotic dominant in the neurosecretory cells of the supraoptic and paraventricular nuclei of the hypothalamus. Passive smoking led to a decreased body weight, a decreased number of thymocytes and myelokaryocytes in pregnant rats. In young rats born from the animals exposed to passive smoking, there was a slowdown in the processes of postnatal differentiation of the adrenal cortex (fascicular zone) with preservation of the extended zone of the fetal cortex. Notably, histo- and morphogenesis both in the organs of the primary (thymus) and secondary (spleen, lymph nodes) links of immunogenesis were delayed. In such young rats, a decreased body weight, thymus, number of thymocytes and splenocytes were recorded.Conclusion. Total results of the study evidence that passive smoking causes immunosuppressive changes in pregnant rats and their offspring combined with delayed postnatal histogenesis and proapoptotic manifestations in the nonapeptide-dergic hypothalamicpituitary adrenocortical system, which can be regarded as an unfavorable factor in the implementation of the neuroendocrine regulative mechanisms of adaptogenesis processes.

Antixenosis of Thodan (AAB), a resistant Musa cultivar to Odoiporus longicollis (Olivier) and characterization of larvicide molecules

Banana pseudostem borer (BPB) Odoiporus longicollis (Olivier) (Coleoptera, Curculionidae) is a serious pest of Musa cultivars. Experimental maintenance of larvae in the live pseudostem of cultivar Thodan, a resistant AAB Musa cultivar has resulted antixenosis followed by death of larvae within a week. Antixenosis was characterized by significant decrease of total hemocytes and sharp changes on the proportionate distribution of different types of hemocytes in larvae. As the number of plasmatocytes, prohemocytes, splenocytes and adipohemocytes decreased the number of granulocytes and oenocytes increased. Antixenosis also caused accumulation of 20-hydroxyecdysone (20E) and significant inhibition on the activities of trypsin like serine protease (TlSP) and phenoloxidase (PO). Phytochemical analysis of Thodan resulted characterization of three larvicides such as Betulinic acid (BA), Stigmasterol-3-O-glucoside (SOG) and Sulfoquinovosyl diacyl glycerol (SQDG), and the content in the pseudostem ranged 0.0027 to 0.007 per cent. All the three larvicides were highly toxic to the larvae with LD50 of 0.38 ppm for SOG, 0.41 ppm for SQDG and 0.83 ppm for BA. Simultaneous action of three larvicides in the live pseudostem resulted resistance in Thodan against infestation by BPB. Susceptible Nendran showed negligibly low content of SOG (0.0011%) and SQDG (0.0013%) but no detectable quantity of BA. Intoxication by all the three larvicides caused significant changes on the proportionate distribution of hemocytes, accumulation of 20-hydroxy ecdysone (20E) and inhibition of TlSP and PO, the enzymes involved in larval metamorphosis and cuticle sclerotisation. This study demonstrated that resistance of Thodan against O. longicollis is due to adverse effect of these larvicides on endocrine system, cuticle development and cytotoxicity of hemocytes. As these larvicide molecules are stable compounds, there is scope for them to be used as substitutes in place of deleterious insecticides for the management BPB.

Exendin-4 Exacerbates Burn-Induced Mortality in Mice by Switching to Th2 Response

IntroductionTo determine if Exendin-4 could be a therapeutic agent for burn-induced hyperglycemia. Materials and methodsMale Balb/c mice received a bolus of Exendin-4 intraperitoneally immediately after 15% total body surface area scald injury. Tail glucose levels were recorded and T-cell functions were analyzed at 4 h and 24 h postburn (pb). Pancreatic pathology was observed consecutively. The secretions of cytokines were detected in serum, spleen, and lung. Apoptosis of splenic CD3+ T-cells was examined by terminal deoxynucleotidyl transferase-mediated dUTP nick-end labeling and flow cytometry. ResultsAlthough Exendin-4 could attenuate burn-induced hyperglycemia in mice at 4 h pb, it accelerated their survival dose dependently with progressive depletion of splenocyte number. T-cell function underwent two-phasic changes following Exendin-4 treatment. Compared to placebo mice, T-cell from Exendin-4–treated mice was manifested with increased proliferation, while decreased IL-2 secretion and lower ratio of IL-4/IFN-γ at 4 h pb. However, at 24 h pb, it showed decreased proliferation, while increased IL-2 secretion and higher ratio of IL-4/IFN-γ. Exendin-4 could elicit higher circulating IL-6 and IL-10 levels at 4 h pb, which were pronounced in the lung at 24 h pb. In the meanwhile, severe inflammation could be found in the pancreas. At 24 h pb, the numbers of terminal deoxynucleotidyl transferase-mediated dUTP nick-end labeling or caspase-3 positive cells and the apoptosis of CD3+ T-cells were significantly increased in the spleens of Exendin-4 mice relative to placebo mice. ConclusionsThese data support a pathogenic role of Exendin-4 signaling during thermal injury, warning against its clinical application in acute insults.

LNP-CpG ODN-adjuvanted varicella-zoster virus glycoprotein E induced comparable levels of immunity with Shingrix™ in VZV-primed mice

Latent varicella-zoster virus (VZV) may be reactivated to cause herpes zoster, which affects one in three people during their lifetime. The currently available subunit vaccine Shingrix™ is superior to the attenuated vaccine Zostavax® in terms of both safety and efficacy, but the supply of its key adjuvant component QS21 is limited. With ionizable lipid nanoparticles (LNPs) that were recently approved by the FDA for COVID-19 mRNA vaccines as carriers, and oligodeoxynucleotides containing CpG motifs (CpG ODNs) approved by the FDA for a subunit hepatitis B vaccine as immunostimulators, we developed a LNP vaccine encapsulating VZV-glycoprotein E (gE) and CpG ODN, and compared its immunogenicity with Shingrix™ in C57BL/6J mice. The results showed that the LNP vaccine induced comparable levels of gE-specific IgG antibodies to Shingrix™ as determined by enzyme-linked immunosorbent assay (ELISA). Most importantly, the LNP vaccine induced comparable levels of cell-mediated immunity (CMI) that plays decisive roles in the efficacy of zoster vaccines to Shingrix™ in a VZV-primed mouse model that was adopted for preclinical studies of Shingrix™. Number of IL-2 and IFN-γ secreting splenocytes and proportion of T helper 1 (Th1) cytokine-expressing CD4+ T cells in LNP-CpG-adjuvanted VZV-gE vaccinated mice were similar to that of Shingrix™ boosted mice. All of the components in this LNP vaccine can be artificially and economically synthesized in large quantities, indicating the potential of LNP-CpG-adjuvanted VZV-gE as a more cost-effective zoster vaccine.

Lycopene Alleviates Chronic Stress-Induced Spleen Apoptosis and Immunosuppression via Inhibiting the Notch Signaling Pathway in Rats.

Chronic stress induction in immunosuppression and splenocyte apoptosis is commonly associated with increased susceptibility to various diseases. Lycopene (LYC) is a member of the carotenoid family with immune restoration and anti-apoptotic function. However, little is known about the mechanisms underlying the protective roles of LYC against spleen injury induced by chronic stress. Herein, male Wistar rats were undergoing chronic restraint stress and/or administered LYC (10 mg/kg) for 21 days. The effective model establishment was validated by open-field tests and levels of corticosterone in serum. Histopathological staining observation displayed that LYC could reduce chronic stress-induced spleen structure damage. Furthermore, LYC treatment significantly reduced the number of apoptotic-positive splenocytes caused by chronic stress via the death receptor apoptotic pathway. We detected the interleukin 4 and interferon γ levels in serum and spleen to determine the ratio of Th1 and Th2 and found that LYC can alleviate the immunosuppression induced by chronic stress. Notably, western blot and real-time polymerase chain reaction indicated that LYC can reduce the expression of the Notch-pathway-related proteins and mRNA in rats exposed to chronic stress. Further study of the potential mechanisms by adding the Notch pathway inhibitor DAPT revealed that LYC alleviates the structure damage, apoptosis, and immunosuppression caused by chronic stress via the suppression of the Notch pathway. Overall, this study presents a strong rationale to target LYC as a treatment strategy to relieve chronic stress-induced spleen injury.

TMOD-14. CONJOINED CLASS I AND II EPITOPES ENHANCE NEOANTIGEN-TARGETED ACTIVE IMMUNOTHERAPY

Abstract INTRODUCTION Cancer vaccines involve the administration of tumor-associated antigens into the host to generate anti-tumor T cell responses. Glioblastoma (GBM) is a disease with poor prognosis. GBM has a limited number of immunotherapeutic targets due to low mutational load, and is also highly heterogeneous; targeting a single antigen leads to antigen escape and tumor growth. METHODS VMDK mice were subcutaneously implanted with 750,000 SMA560 cells and on days 1 and 8 post implantation, mice were treated with 15 nmol of the universal helper epitope, P30, conjoined to the MHCI-restricted neoepitopes Odc1MHCI-P30 or Topbp1MHCI-P30. Human CD27 (hCD27) transgenic mice were intracranially implanted with CT2A-Odc1, followed by anti-CD27 and 15 nmol of Odc1MHCI-P30. B16.OVA or B16.F10 tumor cells were intracranially implanted in hCD27 mice and received SIINFEKL-P30 or Trp2-P30 conjoined peptides. Tumor growth, survival, or IFNγ secretion of splenic or tumor-infiltrating cells was assessed. RESULTS Unlike Odc1MHCI mixed with P30, conjoined Odc1MHCI-P30 had equivalent immunogenicity and anti-tumor efficacy to that observed with native long Odc1 peptide. Native long peptide of Topbp1 did not elicit an antitumor response, yet Topbp1MHCI-P30 caused an increase in numbers of IFNγ-secreting splenocytes and a decrease in tumor growth and similar to that seen with Odc1MHCI-P30 . Anti-CD27 treatment significantly increased numbers of IFNγ secreting splenocytes in Odc1MHCI-P30 vaccinated hCD27 mice, and the use of anti-CD27 with Odc1MHCI-P30 achieved long-term survival in 90% of tumor bearing hCD27 mice. Anti-CD27 synergized with SIINFEKL-P30 and Trp2-P30 to significantly improve survival after administration of these peptides. CONCLUSIONS Our work shows that poorly immunogenic neoantigens can be conjoined to P30 and used to generate an anti-tumor response in mouse models of GBM, and anti-tumor responses of conjoined peptides can be enhanced with anti-CD27 treatment. Together, these data demonstrate the efficacy of neoantigen vaccines for the treatment of heterogeneous GBM.

Radium-223 dichloride causes transient changes in natural killer cell population and cytotoxic function

Rationale Natural killer (NK) cells play an important role in both the innate and adaptive arms of the immune system. While previous studies have demonstrated the effects of ionizing radiation on cytotoxic function of NK cells, little is known about how a chronic exposure to high LET alpha particles emitted by radionuclides will affect both NK population size and function. This study was conducted to determine the effects of 223RaCl2 on splenic NK cell population size and function in Swiss Webster mice. Methods Swiss Webster mice were administered intravenously with 0, 50, or 600 kBq/kg 223RaCl2. Spleens were harvested at 5, 12, and 19 days post-administration. The numbers of splenocytes per spleen were enumerated and flow cytometry was used to assess changes in the distribution of splenocyte subpopulations of B, CD4 and CD8 T lymphocytes, and NK cells. NK functional activity was quantified using YAC-1 target cells and the 51Cr-release assay. Results The total number of splenocytes was unaffected by 223RaCl2. However, significant changes in the distribution of splenocyte subpopulations were observed for NK cells and CD8 T lymphocytes. NK functional activity was enhanced substantially relative to controls at 12 days post-administration, but decreased markedly by day 19. Conclusion NK functional activity is both diminished and enhanced by 223RaCl2 depending on both administered activity and time post-administration. These results suggest that there may be an optimum window of time to combine the 223RaCl2-induced antitumor NK cell response with other cancer therapies that elicit immune activation.

Effect of Carbon Black Nanoparticle on Neonatal Lymphoid Tissues Depending on the Gestational Period of Exposure in Mice.

Introduction: Particulate air pollution, containing nanoparticles, enhances the risk of pediatric allergic diseases that is potentially associated with disruption of neonatal immune system. Previous studies have revealed that maternal exposure to carbon black nanoparticles (CB-NP) disturbs the development of the lymphoid tissues in newborns. Interestingly, the CB-NP-induced immune profiles were observed to be different depending on the gestational period of exposure. It is important to identify the critical exposure period to prevent toxic effects of nanoparticles on the development of the immune system. Therefore, the present study was aimed to investigate the effect of CB-NP on the development of neonatal lymphoid tissues in mice, depending on the gestational period of exposure. Methods: Pregnant ICR mice were treated with a suspension of CB-NP (95 μg/kg body weight) by intranasal instillation; the suspension was administered twice during each gestational period as follows: the pre-implantation period (gestational days 4 and 5), organogenesis period (gestational days 8 and 9), and fetal developmental period (gestational days 15 and 16). The spleen and thymus were collected from offspring mice at 1, 3, and 5-days post-partum. Splenocyte and thymocyte phenotypes were examined by flow cytometry. Gene expression in the spleen was examined by quantitative reverse transcription-polymerase chain reaction. Results: The numbers of total splenocytes and splenic CD3−B220− phenotype (non-T/non-B lymphocytes) in offspring on postnatal day 5 were significantly increased after exposure to CB-NP during the organogenesis period compared with other gestational periods of exposure and control (no exposure). In contrast, expression levels of mRNA associated with chemotaxis and differentiation of immune cells in the spleen were not affected by CB-NP exposure during any gestational period. Conclusion: The organogenesis period was the most susceptible period to CB-NP exposure with respect to lymphoid tissue development. Moreover, the findings of the present and previous studies suggested that long-term exposure to CB-NP across multiple gestational periods including the organogenesis period, rather than acute exposure only organogenesis period, may more severely affect the development of the immune system.

Cordyceps sinensis aqueous extract regulates the adaptive immunity of mice subjected to 60 Co γ irradiation.

Cordyceps sinensis (CS) is a traditional Chinese medicine that is known for treating various diseases, and particularly for exerting therapeutic effects in immune disorders. The adaptive immunoregulatory effects of CS aqueous extract (CSAE) on γ-irradiated mice have not been reported previously. The study aimed to evaluate the therapeutic effects of CSAE in mice immunosuppressed by irradiation. We observed that CSAE administration significantly increased body weight and spleen index, as well as the number of white blood cells, lymphocytes, and platelets in peripheral blood, T and B lymphocytes in spleen tissue, and total serum immunoglobulins in irradiated mice, whereas total serum pro-inflammatory cytokine levels were decreased. Collectively, CSAE maintained the structural integrity of spleen tissue and repaired its damage in irradiated mice as shown by hematoxylin and eosin staining, and decreased the number of terminal deoxynucleotidyl transferase-mediated dUTP nick end labeling-positive splenocytes. Mechanistically, CSAE upregulated Bcl-2, and downregulated Bax and cleaved caspase-3 in spleen of irradiated mice. However, there were no significant differences in red blood cells and neutrophils in different groups. The results revealed that CSAE had protective effects against irradiation-induced immunosuppression, which was likely associated with an antiapoptotic effect and the regulation of adaptive immunity.