Delayed-type Hypersensitivity Response Research Articles

Histamine releasing factor and elongation factor 1 alpha secreted via malaria parasites extracellular vesicles promote immune evasion by inhibiting specific T cell responses

Protozoan pathogens secrete nanosized particles called extracellular vesicles (EVs) to facilitate their survival and chronic infection. Here, we show the inhibition by Plasmodium berghei NK65 blood stage-derived EVs of the proliferative response of CD4+ T cells in response to antigen presentation. Importantly, these results were confirmed in vivo by the capacity of EVs to diminish the ovalbumin-specific delayed type hypersensitivity response. We identified two proteins associated with EVs, the histamine releasing factor (HRF) and the elongation factor 1α (EF-1α) that were found to have immunosuppressive activities. Interestingly, in contrast to WT parasites, EVs from genetically HRF- and EF-1α-deficient parasites failed to inhibit T cell responses in vitro and in vivo. At the level of T cells, we demonstrated that EVs from WT parasites dephosphorylate key molecules (PLCγ1, Akt, and ERK) of the T cell receptor signalling cascade. Remarkably, immunisation with EF-1α alone or in combination with HRF conferred a long-lasting antiparasite protection and immune memory. In conclusion, we identified a new mechanism by which P.berghei-derived EVs exert their immunosuppressive functions by altering T cell responses. The identification of two highly conserved immune suppressive factors offers new conceptual strategies to overcome EV-mediated immune suppression in malaria-infected individuals.

Targeted tumor-derived cellular immunotherapy in advanced breast cancer patients induces initial immune responses and tumor regression.

6 Background: SV-BR-1-GM is a GM-CSF secreting breast cancer cell line which also expresses HLA class I & II antigens. In a previous clinical trial, a partial response of widely metastatic breast cancer was seen in a patient who allele-matched SV-BR-1-GM at HLA-DRB3. Here we report regression of metastatic breast cancer and efficacy analysis with immunologic correlates in subjects of a Phase I/IIa trial of SV-BR-1-GM in patients with advanced breast cancer. Methods: 24 patients with recurrent and/or metastatic breast cancer refractory to standard therapies have been dosed with SV-BR-1-GM in a Phase I/IIa trial. Patients who have undergone at least one cycle of SV-BR-1-GM therapy were analyzed. Patients first received low-dose cyclophosphamide, then intradermal injection of SV-BR-1-GM (20-40x106 cells/four sites) with interferon-α injected into inoculation sites (10,000 IU/site) ~2 & 4 days subsequently. Immunologic responses were measured by delayed type hypersensitivity (DTH) after inoculation and IgG titers of antibodies against SV-BR-1 were measured in sera samples collected prior to and after the first dose of SV-BR-1-GM by flow cytometry. Results: 24 patients have been inoculated with SV-BR-1-GM cells with no unexpected adverse events. None of the patients exhibited immediate hypersensitivity to SV-BR-1. DTH response was recorded in 18 subjects till date, of these, 61% exhibited DTH to cell inoculations. The patient with the most marked DTH response, 01-002, also had a clinical response with regression of multiple lung metastases. Two other patients had evidence of tumor regression (tumor regression is matched SV-BR-1-GM at least at one HLA allele). Sera samples from 6 patients were evaluated and found to contain anti-SV-BR-1 antibodies. Conclusions: SV-BR-1-GM in this regimen appears to be safe and well-tolerated. Initial analysis of SV-BR-1-GM showed significant DTH and antibody responses. HLA matching improves the response rate and is being evaluated as a predictor of response. Clinical trial information: NCT03066947.

Amelioration of Cyclophosphamide Induced Immunosupression by the Hydro-Alcoholic Extract of Gymnema Sylvestre Leaves in Albino Rats

Cyclophosphamide (CP), the most commonly used anti-neoplastic agent causes immunosuppression and toxic effects on various organs that are the limiting factors of cancer treatment. It can be hypothesized that addition of new immunopotentiating agents with detoxification properties would have beneficial role in cancer therapy. Many researchers have proved that, if certain plant products are combined with cancer chemotherapeutic agents, reduce toxicities and improve tumour response. In Ayurveda, Gymnema sylvestre is commonly used for diabetes, obesity and asthma. Also it possesses anti-inflammatory, astringent and digestive properties. Reports on the immunostimulatory activity of Gymnema sylvestre leaves are available from some in vitro and in vivo experiments. With this background the present study was undertaken to evaluate the potential beneficial role of hydro-alcoholic extract of Gymnema sylvestre leaves (GSE) on cyclophosphamide induced immunnosupression in rats. In this experiment, five groups (n=6 in each) of wistar albino rats were randomly divided to receive drugs and vehicle orally for 21 days. Gr I and II received vehicle. Gr III, IV and V were administered with Levamisole 50 mg/kg, GSE 25mg/kg and GSE 50 mg/kg respectively. Except Gr I rats, all rats were injected intraperitoneally with Cyclophosphamide (100mg/kg) on day 9th and 16th of drug treatment. The effects on various organ weights, rise in Haemagglutination titre to Sheep RBC Antigen, delayed type of hypersensitivity (DTH) response to Sheep RBC, percentage of neutrophil adhesion to nylon fibre and phagocytic index from carbon clearance test were evaluated. Humoral and cellular immunity were measured from HA titre and DTH response respectively. It has been observed that, GSE 50 mg/kg significantly increased the antibody titre, percentage neutrophil adhesion and phagocytic index in CP induced immunosuppressed rats. It also restored the CP induced changes in organ weights and the DTH response at 24 and 48 hours of antigen challenge. But these effects were not comparable to that of Levamisole. Our study shows that Gymnema sylvestre reduced the CP induced immunotoxicities and therefore, it could be a safe supplement to cyclophosphamide chemotherapy.

Effects of Exercise on Stress-induced Attenuation of Vaccination Responses in Mice.

To investigate the effects of acute eccentric exercise (ECC) and voluntary wheel exercise training (VWR) on antibody and cell-mediated immune responses to vaccination in chronically stressed mice. We hypothesized that both ECC and VWR would attenuate chronic stress-induced reductions in vaccination responses. Mice were randomized into four groups: control (CON), stress (S)-ECC, S-VWR, and S-sedentary (SED). Stressed groups received chronic restraint stress for 6 h·d, 5 d·wk for 3 wk. After the first week of stress, S-ECC were exercised at 17 m·min speed at -20% grade for 45 min on a treadmill and then intramuscularly injected with 100 μg of ovalbumin (OVA) and 200 μg of alum adjuvant. All other groups were also vaccinated at this time. Stress-VWR mice voluntarily ran on a wheel for the entire experiment. Plasma was collected before, and at 1, 2, and 4 wk postvaccination. Enzyme-linked immunosorbent assay was performed to analyze anti-OVA IgG and IgM antibodies. After 3 wk of chronic stress, all mice were injected with OVA into the ear to determine the delayed-type hypersensitivity. We found that chronic restraint stress significantly reduced body weight and caused adrenal hypertrophy. We also found both S-ECC and S-VWR groups had significantly elevated anti-OVA IgG (P < 0.05), whereas no significant differences between the two exercise groups. Neither S-ECC nor S-VWR altered anti-OVA IgM or delayed-type hypersensitivity responses compared with S-SED group. Acute eccentric exercise and voluntary exercise training alleviated the chronic stress-induced anti-OVA IgG reductions in vaccination responses.

Pam3CSK4 adjuvant given intranasally boosts anti-Leishmania immunogenicity but not protective immune responses conferred by LaAg vaccine against visceral leishmaniasis

The use of adjuvants in vaccine formulations is a well-established practice to improve immunogenicity and protective immunity against diseases. Previously, we have demonstrated the feasibility of intranasal vaccination with the antigen of killed Leishmania amazonensis promastigotes (LaAg) against experimental leishmaniasis. In this work, we sought to optimize the immunogenic effect and protective immunity against murine visceral leishmaniasis conferred by intranasal delivery of LaAg in combination with a synthetic TLR1/TLR2 agonist (Pam3CSK4). Intranasal vaccination with LaAg/PAM did not show toxicity or adverse effects, induced the increase of delayed-type hypersensitivity response and the production of inflammatory cytokines after parasite antigen recall. However, mice vaccinated with LaAg/PAM and challenged with Leishmania infantum presented significant reduction of parasite burden in both liver and spleen, similar to those vaccinated with LaAg. Although LaAg/PAM intranasal vaccination had induced higher frequencies of specific CD4+ and CD8+ T cells and increased levels of IgG2a antibody isotype in serum, both LaAg and LaAg/PAM groups presented similar levels of IL-4 and IFN-y and decreased production of IL-10 when compared to controls. Our results provide the first evidence of the feasibility of intranasal immunization with antigens of killed Leishmania in association with a TLR agonist, which may be explored for developing an effective and alternative strategy for vaccination against visceral leishmaniasis.

Study of Clinical Profile and Immunological Dysfunction in One Hundred Patients of Recurrent Dermatophytosis

Background: An alarming increase in number of patients with chronic and recurrent dermatophytosis has invoked the need to study immunological parameters of the host in addition to traditional pathogens studies. This study was done to evaluate delayed type of hypersensitivity (DTH) response by estimating TH1 cells along with TH17 cells and immediate hypersensitivity (IH) response by estimating TH2 cells population in peripheral blood along with intradermal trichophytin skin test in patients with recurrent dermatophytosis. Methods: In this case-control study, 100 patients with recurrent dermatophytosis and 50 controls were included. Flow-cytometric evaluation of TH1, TH17 and TH2 cells in peripheral blood along with intradermal trichophytin extract test was done. Dermatophytes pathogens were isolated on culture and antifungal drug susceptibility test was done. Relevant risk factors for recurrent disease were analysed and serum IgE levels were estimated. Findings: TH1 cells were reduced in 72.6 % cases vs 52 % controls (p=0.01) and TH17 cells were reduced in 49.4% cases vs 26 % controls (p=0.006). Intradermal trichophytin test demonstrated negative DTH response in 100 % of cases (p=0.0001). Positive IH response was observed in 91% cases vs 68% controls (p=0.0001). T.mentagrophytes complex (95.8%) was the most common species isolated followed by T.rubrum (4.16%). The risk factors associated were family history of tinea, sharing of towels and clothes and corticosteroids use in recent past (p= 0.007, p=0.009, p=0.0001). Serum IgE was elevated in 83.15 % cases (p=0.01). Interpretations:Our study demonstrated impaired DTH response and reduced TH1 and TH17 cell population in patients with recurrent dermatophytosis. Funding Statement: The authors declare: None . Declaration of Interests: The authors declare: None. Ethics Approval Statement: The study protocol was approved by the Institutional Ethics Committee (Intramural) of the Postgraduate Institute of Medical Education and Research, Chandigarh.

Sex Differences in Mouse Popliteal Lymph Nodes

Females have more robust immune responses than males, well-illustrated by the degree of inflammation elicited during delayed-type hypersensitivity (DTH) responses. Here, we have investigated underlying sex differences that may contribute to differential footpad DTH responses using wildtype and four core genotypes (FCG) mice and popliteal lymphnode cellularity and gene expression. DTH responses in XX and XY FCG females showed no role for almost all genes expressed on sex chromosomes. After then filtering-out genes differentially expressed between XX and XY females, only one gene was sexually differentially expressed in wildtype mice, glycosylation-dependent cell adhesion molecule 1 (Glycam1), expressed 7-fold higher in females. Glycam1 facilitates leukocyte entry through high endothelial venules. Consistent with greater Glycam1 expression, female nodes contained twice as many cells. While females had more memory T cells in their nodes, males had a higher percentage of T regulatory cells. This sexual dimorphism in wildtype animals manifested pre-pubertally, was enhanced post-pubertally, and was eliminated by castration. The formation of male gonads is determined by the expression of Sry. Sry overexpression, which does not affect testosterone levels, produced an exaggerated male phenotype. We conclude that Sry expression through formation of the male gonad indirectly negatively impacts the potential for local inflammation.

Exposure to Solar UVR Suppresses Cell-Mediated Immunization Responses in Humans: The Australian Ultraviolet Radiation and Immunity Study

Animal and human studies show that exposure to solar-simulated UVR is immunomodulatory. Human studies that used natural sun exposure and controlled for confounding are rare. We immunized 217 healthy adults (age range= 18-40 years) with a T-cell-dependent antigen, keyhole limpet hemocyanin, and measured personal clothing-adjusted UVR exposure (for 5 days before and after immunization), lifetime cumulative UVR exposure, serum 25-hydroxyvitamin D concentration at immunization, and potential confounding factors. We tested cellular and humoral immune responses in relation to UVR exposure. The delayed-type hypersensitivity response to keyhole limpet hemocyanin recall challenge was lower in individuals with higher personal clothing-adjusted UVR exposure on the day before immunization (P= 0.015) and during intervals spanning the day before to 2-3 days after immunization. There was an incremental increase in T helper type 17 cells (as a proportion of CD4+ T cells) from preimmunization to postimmunization in the high, compared with the low, personal clothing-adjusted UVR exposure group (0.31% vs. -0.39%, P= 0.004). Keyhole limpet hemocyanin-specific antibody titers were not associated with acute or cumulative UVR exposure or serum 25-hydroxyvitamin D levels. Higher UVR exposure at antigen sensitization was associated with a reduced delayed-type hypersensitivity response and altered T helper type 17 kinetics. This has implications for the effectiveness of vaccinations and susceptibility to infections that rely on cell-mediated immune responses.

English

The isolation, characterization and effects of lactobacillus spp. from fresh palm wine was undertaken in this study. The aim of this study was to isolate and investigate the in vivo effects of the isolates on innate and cellular immune system. The result of the phenotypic and genotypic characterizations showed the presence of Lactobacillus brevis, Lactobacillus paracasei subsp. Tolerans, Lactobacillus paracasei and Lactobacillus yonginensis, which are members of Lactobacillus spp., isolated. The effects of the isolates on innate and cellular immunity were investigated, using two models namely: in vivo leukocytes mobilization rate (LMR) and delayed type hypersensitivity response (DTHR). The different isolates had very significant effect on cellular immunity as represented by their ability to stimulate delayed type hypersensitivity response in treated rats. They caused a percentage increase of 300, 600, 600, 500 and 650% for L. brevis, L. paracasei subsp. Tolerans, L. paracasei, L. yonginensis and positive control respectively when compared with the negative control. Similarly, the various isolates also affected significantly the innate immune system through their marked influence on the total leukocyte count according to the result of the experiment. They were able to produce a percentage increase of 213, 204, 188.6, and 152.5% for L. paracasei, L. yonginensis, L. paracasei subsp. Tolerans and L. brevis respectively; while the positive control produced 83.8% increase when compared with the negative control. The result of this study showed that the four lactobacillus spp. isolated from fresh palm wine significantly affected the innate and cellular component of the immune system positively. Key words: Delayed type, hypersensitivity response, innate immunity, cellular immunity, leukocyte mobilization.

Serine protease inhibitors rich Coccinia grandis (L.) Voigt leaf extract induces protective immune responses in murine visceral leishmaniasis

Leishmaniasis is a parasite-mediated tropical disease affecting millions of individuals worldwide. The available antileishmanial chemotherapeutic modalities exhibit adverse toxicity, exorbitant price and advent of drug-resistant parasites. Hence, plant-derived products are an alternative preference for the emergence of novel and effective antileishmanial agents that rejuvenate the host immunity with limited toxicity. The present work is complementary to our previous report that revealed the in vitro antileishmanial and immunomodulatory activity of Coccinia grandis (L.) Voigt leaf extract (Cg-Ex) rich in serine protease inhibitors. Thus, preliminary objectives of the study were to elucidate the leishmanicidal activity and host effector mechanism in Leishmania donovani infected BALB/c mice treated with Cg-Ex. Oral administration of Cg-Ex significantly reduced the spleen and liver parasite burden at dose-dependently. The parasite elimination was associated with generation of ROS and NO that are interrelated with up-regulation of disease-suppressing Th1 cytokines and down-regulation of disease-promoting Th2 cytokines at both protein and mRNA level. Moreover, Cg-Ex augmented the delayed-type hypersensitivity (DTH) response and serum IgG2a level which are correlated with the diminution of parasite burden with no hepatic and renal toxicity. Additionally, histological analysis of spleen depicted the improvement of structural disorganization of white and red pulp after Cg-Ex treatment. Therefore, our intriguing findings have presented the first indication of in vivo antileishmanial efficacy through activation of pro-inflammatory immune responses of the host by a natural plant leaf extract (Cg-Ex) containing serine protease inhibitors which could have a role as a potential immunomodulator against visceral leishmaniasis.

Inhibins regulate peripheral regulatory T cell induction through modulation of dendritic cell function.

We have previously reported that the absence of inhibins results in impaired dendritic cell (DC) maturation and function, leading to decreased T cell activation and diminished delayed‐type hypersensitivity responses. Here, we investigated the role of inhibins in peripheral regulatory T cell (Treg) induction in vitro and in vivo. Inhibin deficient (Inhα−/−) mice showed an increased percentage of peripherally induced Tregs in colonic lamina propria and mesenteric lymph nodes, compared to Inhα+/+ mice, which correlated with increased expression of PD‐L1 in CD103+ and CD8α+ DCs. Lipopolysaccharide‐stimulated bone marrow‐derived and ex vivo spleen‐ and lymph node‐purified CD11c+ Inhα−/− DCs induced higher Tregs in vitro. Moreover, in vivo anti‐DEC205‐ovalbumin (OVA) DC targeting of mice with adoptively transferred OVA‐specific T cells showed enhanced induced peripheral Treg conversion in Inhα−/− mice. These data identify inhibins as key regulators of peripheral T cell tolerance.

Assessment of antimicrobial and immunomodulatory activities of termite associated fungi, Termitomyces clypeatus R. Heim (Lyophyllaceae, Basidiomycota)

BackgroundTermitomyces clypeatus (T. clypeatus) is an edible mushroom specie which grows in various areas in Cameroon. It is indicated by several healers in treatment of several infectionsand is frequently used for this purpose. However, no study has been reported on its antibacterial and immunomodulatory activities which were the aim of this study.MethodsDisc diffusion method was used to appreciate the bactericidal activity against 4 bacteria and 2 yeast species. The immunomodulatory activities were assessed in mice, where the extract was administered by gavage and as supplement in the feed. The treatment was done for 10 days. Delayed type hypersensitivity test was carried out to assess the cell mediated immune response while the effect on humoral immunity was evaluated using hemagglutination assay and mice lethality test.. The body weight was also recorded in mice used for delayed hypersensitivity.ResultsIt was found that extract of T. clypeatus highly inhibited the growth of bacteria and yeast at different ratios compared to the medium (P < 0.05). The extract of T. clypeatus has reversed the immunosuppressed effects of dexamethasone on antibody formation. In addition, it significantly decreased of the mice lethality rate in mice infected by Pasteurella multocida (P < 0.05). Administration of T. clypeatus also significantly increased the delayed type hypersensitivity response in healthy and dexamethasone immunosuppressed mice (P < 0.05), but it significantly reduced the body weight of mice after 10 days.ConclusionThe results provided basic information demonstrating the antibacterial activity and immunostimulatory activity of T. clypeatus on both cell-mediated and humoral immunity.

Immunomodulatory effect of Cynara scolymus (artichoke) in rats.

Cynara scolymus is a plant used both as food as well as medicinal plant worldwide. Cynarin is one of the main active principles of the plant, and it is also present in species such as Echinacea purpurae, which is known to have immunomodulatory activity. Thus, the objective of this study is to evaluate the immune effects of C.scolymus in rats. Rats were treated with 1.0-, 2.0-, or 4.0-g/kg body weight of C.scolymus extract for 28days. Haemogram, serum biochemistry, lymphoid organs weight, and their cell phenotypes were evaluated. Macrophages and neutrophils oxidative burst, specific humoral immune response, and the delayed-type hypersensitivity (DTH) were studied. No changes in the haemogram, biochemical profile, antibody titers, lymphoid organs, and in their cellularities were observed. An increase in the basal activity of reactive oxygen species from male's macrophage was observed. There was a suppression of the DTH response in both gender when treated with the highest dose of C.scolymus. This study is the first in the literature that revealed an immunosuppressive effect of C.scolymus. We also verified that the doses of artichoke extract here employed did not cause general toxicity.

THE EFECT OF IMMUNOMODULATOR BY EXTRACT ETHANOL OF HERBA BINARA (ARTEMISIA VULGARIS L.) TOWARD THE RESPONSE OF DELAYED-TYPE HYPERSENSITIVITY IN RAT MALE

Immunomodulator is a compound that can increaase the imuno system. One of the plants that have immunomodulator’s activity is Herba Binara (Artemisia vulgaris L.). the purpose of this research was to test the effect of immunomodulator by extract of herba binara ethanol on rat male. The activity of immunomodulator was determined by using digital pletysmometer by measuring the differences between the last leg swelling’s volume and the first leg swelling’s volume. The treatment group were divided into 5 groups. Each group consistof 5 rats CMC-Na 0,5% (negative control), Imboost 32,5 mg/kgBW (positive control), dose of EEHB 50, 100, 200 mg/kgBW, and bacteria E.coli as antigen. The results slowed that distribution of EEHB dose 200 mg/kgBW can give the effect of immunostimulant by swelling enthancement compared by CMC-Na 0,5 %. EEHB 200 mg/kgBW that have activity comparable with Imboost 32,5 mg/kgBW. Thus, it is concluded that of herba binara extract has immunomodulator effects on delayed-type hypersensitivity response of rat male.

Vaccination with whole-cell killed or recombinant leishmanial protein and toll-like receptor agonists against Leishmania tropica in BALB/c mice.

One strategy to control leishmaniasis is vaccination with potent antigens alongside suitable adjuvants. The use of toll-like receptor (TLR) agonists as adjuvants is a promising approach in Leishmania vaccine research. Leishmania (L.) tropica is among the less-investigated Leishmania species and a causative agent of cutaneous and sometimes visceral leishmaniasis with no approved vaccine against it. In the present study, we assessed the adjuvant effects of a TLR4 agonist, monophosphoryl lipid A (MPL) and a TLR7/8 agonist, R848 beside two different types of Leishmania vaccine candidates; namely, whole-cell soluble L. tropica antigen (SLA) and recombinant L. tropica stress-inducible protein-1 (LtSTI1). BALB/c mice were vaccinated three times by the antigens (SLA or LtSTI1) with MPL or R848 and then were challenged by L. tropica. Delayed-type hypersensitivity (DTH), parasite load, disease progression and cytokines (IL-10 and IFN-γ) responses were assessed. In general compared to SLA, application of LtSTI1 resulted in higher DTH, higher IFN-γ response and lower lymph node parasite load. Also compared to R848, MPL as an adjuvant resulted in higher DTH and lower lymph node parasite load. Although, no outstanding ability for SLA and R848 in evoking immune responses of BALB/c mice against L. tropica infection could be observed, our data suggest that LtSTI1 and MPL have a better potential to control L. tropica infection and could be pursued for the development of effective vaccination strategies.

MAVS deficiency induces gut dysbiotic microbiota conferring a proallergic phenotype

Prominent changes in the gut microbiota (referred to as "dysbiosis") play a key role in the development of allergic disorders, but the underlying mechanisms remain unknown. Study of the delayed-type hypersensitivity (DTH) response in mice contributed to our knowledge of the pathophysiology of human allergic contact dermatitis. Here we report a negative regulatory role of the RIG-I-like receptor adaptor mitochondrial antiviral signaling (MAVS) on DTH by modulating gut bacterial ecology. Cohousing and fecal transplantation experiments revealed that the dysbiotic microbiota of Mavs-/- mice conferred a proallergic phenotype that is communicable to wild-type mice. DTH sensitization coincided with increased intestinal permeability and bacterial translocation within lymphoid organs that enhanced DTH severity. Collectively, we unveiled an unexpected impact of RIG-I-like signaling on the gut microbiota with consequences on allergic skin disease outcome. Primarily, these data indicate that manipulating the gut microbiota may help in the development of therapeutic strategies for the treatment of human allergic skin pathologies.

Evaluation of Immunomodulatory activity of Diosgenin in rats

Background: The immune system is intrinsic to health. Modulation of the immune responses to alleviate the diseases by using herbal plants has been of interest for many years. Diosgenin, a naturally occurring steroid saponin mainly present in the seeds of fenugreek (Trigonella foenum graecum) and in the root tubers of wild yams (Dioscorea villosa). Activation of specific and nonspecific immunity results in stimulation of immune response. Diosgenin has the positive effects on both specific and nonspecific immunity. Aim: To study the immunomodulatory activity of Diosgenin in albino wistar rats. Method: The suspension of Diosgenin was given orally at the dosage level of 50, 100 and 150 mg/kg for 21 days in a rat. The immunomodulatory activity on specific and non-specific immunity was studied by heamagglutination antibody (HA) titer, delayed type hypersensitivity (DTH) response and carbon clearance test. Immunosuppression in a rat was induced by using Cyclophosphamide (100 mg/kg, p.o.). Sheep red blood cells (SRBCs) were used as antigen (0.1ml 20% SRBCs). Result: Diosgenin exhibited significant increase in the production of antibody titer in response to SRBC antigen. A significant increase in both primary and secondary HA titer was observed in immunosuppressed group treated with Diosgenin when compared with negative control. A significant increase in the DTH response was observed in immunosuppressed animals treated with Diosgenin, pre-sensitized with SRBCs antigen. Diosgenin exhibited significant increase in phagocytic index against control group, indicating the stimulation of the reticuloendothelial system. Conclusion: The study indicates that Diosgenin triggers stimulatory effect on specific and nonspecific immune response. The immunostimulant effect of Diosgenin could be attributed due to its saponin glycoside.

Altered monocyte phenotypes but not impaired peripheral T cell immunity may explain susceptibility of the elderly to develop tuberculosis.

Tuberculosis (TB) is the leading killer due to a single infectious disease worldwide. With the aging of the global population, the case rate and deaths due to TB are highest in the elderly population. While general immunosenescence associated with old age is thought to contribute to the susceptibility of the elderly to develop active TB disease, very few studies of immune function in elderly individuals with Mycobacterium tuberculosis (M.tb) infection or disease have been performed. In particular, impaired adaptive T cell immunity to M.tb is one proposed mechanism for the elderly's increased susceptibility primarily on the basis of the decreased delayed type hypersensitivity response to tuberculin-purified protein derivative in the skin of elderly individuals. To investigate immunological reasons why the elderly are susceptible to develop active TB disease, we performed a cross-sectional observational study over a five year period (2012-2016) enrolling participants from 2 age groups (adults: 25-44 years; elderly: 65 and older) and 3 M.tb infection statuses (active TB, latent TB infection, and healthy controls without history of M.tb infection). We hypothesized that impaired peripheral T cell immunity plays a role in the biological susceptibility of the elderly to TB. Contrary to our hypothesis, we observed no evidence of impaired M.tb specific T cell frequency or altered production of cytokines implicated in M.tb control (IFN-γ, IL-10) in peripheral blood in the elderly. Instead, we observed alterations in monocyte proportion and phenotype with age and M.tb infection that suggest their potential role in the susceptibility of the elderly to develop active TB. Our results suggest a potential link between the known widespread low-grade systemic inflammation of old age, termed "inflammaging," with the elderly's specific susceptibility to developing active TB. Moreover, our results highlight the need for further research into the biological reasons why the elderly are more susceptible to disease and death from TB, so that public health systems can be better equipped to face the present and future problem of TB in an aging global population.

Glycolytic Bias in mTORC2 Deficient Dendritic Cells Potentiates Antigen-Specific Immunity and Accelerates Graft Rejection

Background/Hypothesis The mechanistic target of rapamycin (mTOR) is known to function in two discrete complexes: mTOR complex 1 (mTORC1) and mTORC2. The function of mTORC1 in dendritic cells (DCs) has been studied extensively using rapamycin (RAPA) as an inhibitor. RAPA inhibition of mTORC1 prevents DC maturation, leading to decreased T effector cell proliferation and increased regulatory T cell (Treg) differentiation. Our group has recently demonstrated that mTORC2 deletion in DCs leads to both an enhanced pro-inflammatory DC phenotype and Th1/Th17 allogeneic T cell polarization and proliferation. However, the underlying mechanism has not been resolved. In addition, the role of mTORC2 in DCs in the context of transplantation has not been defined. We hypothesized that ablation of mTORC2 in DCs would alter metabolic activity, resulting in augmented antigen-specific T cell responses and accelerated graft rejection. Methods To assess the role of mTORC2 in DCs in transplantation, we used models of non-MHC mismatched skin transplantation in which either the graft donor or recipient was deficient in mTORC2 specifically in conventional CD11c+ DC (TORC2DC-/-). Graft survival was monitored; Banff rejection scoring was performed by a blinded pathologist. T cell infiltration and collagen degradation in the graft were determined by immunohistochemistry. To ascertain whether skin-resident TORC2DC-/- could augment inflammatory responses, we performed a cell-mediated delayed-type hypersensitivity assay. As mTORC2 has been implicated in cytoskeletal dynamics, we measured DC migration into secondary lymphoid tissue. To elucidate the role of mTORC2 in regulating DC metabolism we analyzed glycolytic capacity and mitochondrial respiratory activity of wild-type (WT) DC and TORC2DC-/- using a Seahorse XF Bioanalyzer. Mitochondrial mass and activity were determined via flow cytometric analysis of MitoTracker Green and TMRE uptake, respectively. ATP production was assessed using a luciferase-based luminescence assay. Results/Conclusions We demonstrate for the first time, that TORC2DC-/- deficiency in either skin graft donors or recipients accelerates immune-mediated rejection. TORC2DC-/- mice also exhibit enhanced T cell and inflammatory monocytic infiltration in the course of delayed-type hypersensitivity responses. These effects are not due to differential migration of TORC2DC-/- to secondary lymphoid tissue. TORC2DC-/- utilize an altered metabolic program, wherein glycolytic function is enhanced as compared to WT DCs. This metabolic phenotype corresponds with increased viability of TORC2DC-/- after stimulation, which may allow these TORC2DC-/- to persist in secondary lymphoid tissue longer than WT DCs. These findings reveal a novel role for mTORC2 in regulating DC immunometabolism, and may provide a basis for therapeutic targeting of DC metabolism to regulate immune responses in transplantation. NIH T32 AI74490.

Immunomodulatory effects of silymarin after subacute exposure to mice: A tiered approach immunotoxicity screening.

Silymarin is a flavonoid complex extracted from the Silybum marianum plant with a wide range of pharmacological and biochemical effects. In the present study, the immunomodulatory effects of silymarin were investigated in BALB/c mice. Silymarin was administered daily by intraperitoneal injection at doses of 50, 100 and 150 mg/kg for 14 consecutive days. Following the exposure, host hematological parameters, spleen cellularity and histopathological examination, as well as delayed-type hypersensitivity (DTH) responses, hemagglutination titers (HA), splenocyte cytokine production and lymphocyte proliferation assay were studied in all of the test groups of animals. The results showed that the low dose of silymarin (50 mg/kg) could stimulate both cellular and humoral immune functions in the treated hosts. In addition, silymarin at 100 mg/kg appeared to impact on DTH responses and lymphoproliferation. Based on the finding here, it would seem that silymarin has efficient immunostimulant properties. As a recommendation, the application of silymarin along with acupuncture technique (herbal acupuncture) can be thought as a good plan to modulate and enhance the immune system for the management of several immunodeficiency disorders. However, further studies are required to demonstrate this hypothesis.