<Emphasis Type="Italic">In vivo</Emphasis> Proinflammatory Cytokine Production by CD-1 Mice in Response to Equipotential Doses of <Emphasis Type="Italic">Rhodobacter capsulatus</Emphasis> PG and <Emphasis Type="Italic">Salmonella enterica</Emphasis> Lipopolysaccharides

The capacities of relatively nontoxic lipopolysaccharide (LPS) from Rhodobacter capsulatus PG and highly potent LPS from Salmonella enterica serovar Typhimurium to evoke proinflammatory cytokine production have been compared in vivo. Intravenous administration of S. enterica LPS at a relatively low dose (1 mg/kg body weight) led to upregulation of TNF-α, IL-6, and IFN-γ production by non-sensitized CD-1 mice. LPS from R. capsulatus PG used at a four-times higher dose than that from S. enterica elicited production of almost the same amount of systemic TNF-α; therefore, the doses of 4 mg/kg LPS from R. capsulatus PG and 1 mg/kg LPS from S. enterica were considered to be approximately equipotential doses with respect to the LPS-dependent TNF-α production by CD-1 mice. Rhodobacter capsulatus PG LPS was a weaker inducer of the production of TNF-α, IL-6, and IFN-γ, as compared to the equipotential dose of S. enterica LPS. Administration of R. capsulatus PG LPS before S. enterica LPS decreased production of IFN-γ, but not of TNF-α and IL-6, induced by S. enterica LPS. Rhodobacter capsulatus PG LPS also suppressed IFN-γ production induced by S. enterica LPS when R. capsulatus PG LPS had been injected as little as 10 min after S. enterica LPS, but to a much lesser extent. Rhodobacter capsulatus PG LPS did not affect TNF-α and IL-6 production induced by the equipotential dose of S. enterica LPS. In order to draw conclusion on the endotoxic activity of particular LPSs, species-specific structure or arrangement of the animal or human immune systems should be considered.     

Dynamics of antagonistic potency of <Emphasis Type="Italic">Rhodobacter capsulatus</Emphasis> PG lipopolysaccharide against endotoxin-induced effects

The dynamics of antagonistic potency of lipopolysaccharide (LPS) isolated from Rhodobacter capsulatus PG on the synthesis of proinflammatory (TNF-α, IL-1β, IL-8, IL-6, IFN-γ) and antiinflammatory (IL-10, IL-1Ra) cytokines induced by highly stimulatory endotoxins from Escherichia coli or Salmonella enterica have been studied. Using human whole blood, we have shown that R. capsulatus PG LPS inhibited most pronouncedly the endotoxin-induced synthesis of TNF-α, IL-1β, IL-8, and IL-6 during the first 6 h after endotoxin challenge. Similarly, the endotoxin-induced release of IFN-γ was abolished by R. capsulatus PG LPS as well (24 h). In contrast to the above-mentioned cytokines, the relatively weak antagonistic activity of R. capsulatus PG LPS against endotoxin-triggered production of IL-6 and IL-8 was revealed. Since R. capsulatus PG LPS displays more potent antagonistic activity against deleterious effects of S. enterica LPS than those of E. coli LPS in the cases of such cytokines as IL-1β (6 and 24 h), IL-6 and IL-8 (4 h), we conclude that the effectiveness of protective action of antagonist is mostly determined by the primary lipid A structure of the employed agonist.     

Usefulness of selected laboratory markers in ulcerative colitis

Introduction

This study aimed to compare the accuracy of selected laboratory markers in assessing disease activity in patients with ulcerative colitis (UC). The analysis included serum IL-2, IL-4, IL-6, IL-10, IL-17, TNF-α, IFN-γ, hsCRP, peripheral regulatory T cells, as well as fecal calprotectin and lactoferrin.

Patients and methods

A group of 45 adults with UC was enrolled in the study. Disease activity was assessed using the Mayo endoscopic index, while for clinical activity scoring, the Clinical Activity Index (CAI) was used. Concentrations of markers investigated were estimated by means of flow cytometry and enzyme-linked immunosorbent assays: the results were correlated with both indices.

Results

The study demonstrated that both fecal markers, i.e. calprotectin (r = 0.880, P<0.001) and lactoferrin (r = 0.799, P<0.001) correlated closely with the Mayo endoscopic score, and might be used to evaluate the severity of UC in the clinical setting. The correlation of these markers with CAI was also significant, with r = 0.831 for calprotectin (P<0.001) and r = 0.672 for lactoferrin (P<0.05). As for the other markers investigated, only IL-6 (r = 0.598, P<0.001), IL-17A (r = 0.587, P<0.005), and TNF-α (r = 0.701, P<0.001) correlated closely with the Mayo endoscopic index. The correlation of the markers with CAI was also significant, though weaker, with r = 0.525 for IL-6 (P<0.001), r = 0.587 for IL-17A (P<0.05), and r = 0.624 for TNF-α (P<0.001).

Discussion

Despite the fact, that UC is generally considered to be an IL-13-driven, Th2-like type of disease, markers of inflammation such as serum interleukin (IL)-6, IL-17, TNF-α, fecal calprotectin and lactoferrin might be useful in assessing disease activity.

     

Genetic,Immune-Inflammatory,and Oxidative Stress Biomarkers as Predictors for Disability and Disease Progression in Multiple Sclerosis

The aim of this study was to evaluate the TNFβ NcoI polymorphism (rs909253) and immune-inflammatory, oxidative, and nitrosative stress (IO&NS) biomarkers as predictors of disease progression in multiple sclerosis (MS). We included 212 MS patients (150 female, 62 male, mean (±standard deviation (SD)) age?=?42.7?±?13.8 years) and 249 healthy controls (177 female, 72 male, 36.8?±?11 years). The disability was measured the Expanded Disability Status Scale (EDSS) in 2006 and 2011. We determined the TNFβ NcoI polymorphism and serum levels of interleukin (IL)-6, tumor necrosis factor (TNF)-α, interferon (IFN)-γ, IL-4, IL-10, and IL-17, albumin, ferritin, and plasma levels of lipid hydroperoxides (CL-LOOH), carbonyl protein, advanced oxidation protein products (AOPPs), nitric oxide metabolites (NOx), and total radical-trapping antioxidant parameter (TRAP). The mean EDSS (±SD) in 2006 was 1.62?±?2.01 and in 2011 3.16?±?2.29, and disease duration was 7.34?±?7.0 years. IL-10, TNF-α, IFN-γ, AOPP, and NOx levels were significantly higher and IL-4 lower in MS patients with a higher 2011 EDSS scores (≥3) as compared with those with EDSS?<?3. The actual increases in EDSS from 2006 to 2011 were positively associated with TNF-α and IFN-γ. Increased IFN-γ values were associated with higher pyramidal symptoms and increased IL-6 with sensitive symptoms. Increased carbonyl protein and IL-10 but lowered albumin levels predicted cerebellar symptoms. The TNFB1/B2 genotype decreased risk towards progression of pyramidal symptoms. Treatments with IFN-β and glatiramer acetate significantly reduced TNF-α but did not affect the other IO&NS biomarkers or disease progression. Taken together, IO&NS biomarkers and NcoI TNFβ genotypes predict high disability in MS and are associated with different aspects of disease progression. New drugs to treat MS should also target oxidative stress pathways.     

Analysis of Influence of Baicalin Joint Resveratrol Retention Enema on the TNF-α, SIgA,IL-2, IFN-γ of Rats with Respiratory Syncytial Virus Infection

Explore the influence of baicalin joint resveratrol retention enema on TNF-α, SIgA, IL-2, and IFN-γ of rats with respiratory syncytial virus (RSV) infection. The 60 SD rats were randomly divided into normal group, model group, baicalin group, resveratrol group, joint group, and ribavirin group. For model group, baicalin group, resveratrol group, joint group, and ribavirin group, rats were given RSV virus suspension intranasally for 3 days, and model group was not given administration. Baicalin group, resveratrol group, joint group, and ribavirin group were, respectively, given baicalin 100 mg/kg/day, resveratrol 30 mg/kg/day, baicalin joint resveratrol, and ribavirin 1 g/kg/day retention enema. After continuously given administration 7 days, rats were measured in serum TNF-α, IL-2, IFN-γ levels and SIgA levels in bronchoalveolar lavage fluid. Model group, TNF-α, IL-2, IFN-γ, and SIgA were significantly higher than the normal group (P < 0.05); Baicalin group, resveratrol group, ribavirin group, TNF-α, IL-2, IFN-γ, and SIgA were significantly higher than the model group (P < 0.05); TNF-α, IL-2 between baicalin group, resveratrol group, ribavirin group, have no significant difference (P > 0.05); Baicalin group, resveratrol group, joint group, IFN-γ, and SIgA were significantly higher than the ribavirin group (P < 0.05); Joint group TNF-α, IL-2, IFN-γ, and SIgA were significantly higher than baicalin group, resveratrol group, and ribavirin group (P < 0.05). Baicalin joint resveratrol retention enema can increase RSV infection model in rats serum TNF-α, IL-2, IFN-γ levels and SIgA levels in bronchoalveolar lavage fluid, which may anti-virus through this mechanism.     

Recombinant flagellin-PAL fusion protein of <Emphasis Type="Italic">Legionella pneumophila</Emphasis> induced cell-mediated and protective immunity against bacteremia in BALB/c mice

We report a new recombinant fusion protein composed of full-length Legionella pneumophila flagellin A and peptidoglycan-associated lipoprotein (PAL), rFLA-PAL, capable of inducing protective immunity against L. pneumophila. The recombinant protein was over expressed in Escherichia coli strain BL21 (DE3) using pET-28a (+) expression vector (pET28a-flaA-pal) and purified by Ni²⁺ exchange chromatography. Immunological properties of rFLA-PAL were assessed in a mouse model. Female BALB/c mice, immunized with rFLA-PAL, exhibited a rapid increase in serum antibody concentration against each of its protein portions. Furthermore, a strong activation of both innate and adaptive cell-mediated immunity was observed as indicated by antigen-specific splenocyte proliferation, IFN-γ and IL-12 production, and early production of TNF-α in the serum and in splenocyte cultures which were separately assessed against PAL and FLA. BALB/c mice were challenged with a lethal dose of L. pneumophila intravenously. In a 10-days follow-up after intravenous lethal challenge with L. pneumophila, a 100% survival rate was observed for mice immunized with rFLA-PAL, same as for those immunized with a sublethal dose of L. pneumophila. Based on the potent immune responses observed in mice immunized with rFLA-PAL, this recombinant fusion protein could be a potential vaccine candidate against the intracellular pathogen L. pneumophila.     

Enzyme-Linked Immunosorbent Spot (ELISpot) monitoring of cytokine-producing cells for the prediction of acute rejection in renal transplant patients

The purpose of this study was to evaluate T-cell immunity markers using serial post-transplantation monitoring of cytokine-producing cells during the first post-transplant months for the prediction of acute rejection and potentially chronic rejection of kidney allograft. We followed 57 kidney allograft recipients for meanly 3 years post-transplantation. Blood samples were collected pre-transplant, 2, 4 and 12 weeks post-transplant. The frequencies of IL-10-, IL-17- and IFN-γ-producing cells were determined in all time-points using ELISPOT assay. The results of ELISpot monitoring and levels of IL-23 and TGF-β were compared between recipients with acute (n = 12) or chronic rejection episodes and patients with stable graft function (n = 45). In all post-transplant time-points, significantly high frequencies of IFN-γ- and IL-17-producing cells and low frequency of IL-10-producing cells were observed in rejection group versus patients with stable graft function (P<0.0001). TheROCcurve analysis for determining the reliability of cytokine-producing cells for the prediction of acute rejection revealed that AUC was 0.046 for IL-10 (P<0.001), 0.927 for IL-17 (P<0.001) and 0.929 for INF-γ-producing cells (P<0.001). Our results indicate that analyzing the frequencies of INF-γ/IL-10/IL-17-producing cells may define a reliable panel for the prediction of acute rejection within the first post-transplant year which could also be applicable for the prediction of chronic rejection episodes.     

Structural and functional characterization of a novel immunomodulatory glycoprotein isolated from ajowan (<Emphasis Type="Italic">Trachyspermum ammi</Emphasis> L.)

Ajowan (Trachyspermum ammi L.) spice has been used in food preparations and also as a traditional medicine in Ayurveda. Although a number of pharmacological activities have been attributed to ajowan, its role in immunomodulation is not known. The main objective of the present study is to examine the macromolecular immunomodulatory components. Macrophage activation was studied by nitric oxide (NO) release, phagocytosis and secretion of pro-inflammatory cytokines as the markers. Ethanol precipitate (fractional) of ajowan aqueous extract was subjected to conventional chromatography (Q Sepharose followed by Bio-Gel P-100). One of the proteins (30.7 kDa; ajowan glycoprotein or Agp) showed effective mitogenic activity towards splenocytes. Agp is a O-linked glycoprotein with the glycans contributing to one-third of the molecular mass. It has a high content of glutamic acid, serine, aspartic acid and proline whereas galactose (45.7%), arabinose (34.5%), glucose (7%), mannose (5%) and xylose (4%) are the constituent sugars. Secondary structure analysis indicated that Agp contains 79% α-helices and 21% random coil. Internal sequencing of the tryptic peptides did not show homology with the existing proteins in the database (BLAST). Agp at 1 μg/mL induced proliferation of B-cell enriched murine splenocytes and activated macrophages in releasing NO and promoted phagocytosis (p < 0.01). RAW 264.7 cells produced pro-inflammatory cytokines (IL-12, TNF-α and IFN-γ) at 1 μg/mL Agp (p < 0.01). Deproteinized Agp (dpAgp) failed to elicit activation of murine immune cells, whereas deglycosylated Agp (20 kDa; dgAgp) showed compromised efficiency. This is the first report of an immunomodulatory protein from ajowan.     

Changes of Chemerin Production in Obese Patients with Different States of Carbohydrate Metabolism

Chemerin is an adipose tissue mediator involved in the regulation of many processes, including lipogenesis, inflammatory responses, etc. The role of chemerin in the development of insulin resistance still needs better understanding. The aim of the study was to investigate chemerin production in obese patients with different states of carbohydrate metabolism. The study included 155 patients with diagnosed obesity and 34 patients with overweight. The control group 1 consisted of 43 conditionally healthy donors without obesity. For comparison of the research data on evaluation of tissue-specific mRNA expression of the genes IL-6, TNF-α, RARRES2, (encoding IL-6, TNF-α, and chemerin, respectively) control group 2 consisting of 30 non-obese was also included into this study. The relative level of mRNA expression of the genes IL-6, TNF-α and RARRES2 (encoding IL-6, TNF-α and chemerin, respectively) was carried out using real time PCR. Concentrations of IL-6, TNF-α, and chemerin were measured in serum/plasma using an enzymelinked immunosorbent assay (ELISA). Significant differences were found in the plasma level of chemerin and tissue-specific patterns of RARRES2 gene expression in obese patients; these changes depended on the degree of obesity and the state of carbohydrate metabolism. Opposite associations between RARRES2 gene expression and expression TNF-α and IL-6 genes have been recognized in adipose tissues of different localization: in obese patients (BMI ≤ 40 kg/m²) without type 2 diabetes mellitus (DM2) they were negative, while in obese patients with DM2 diabetes they were positive. The recognized correlations between the plasma content of chemerin and the expression level of its gene in biopsies with various parameters of carbohydrate and lipid metabolism, and proinflammatory molecules indicate chemerin involvement in metabolic and immune processes in obesity.     

Intralesional uridine-5′-triphosphate (UTP) treatment induced resistance to <Emphasis Type="Italic">Leishmania amazonensis</Emphasis> infection by boosting Th<Subscript>1</Subscript> immune responses and reactive oxygen species production

Leishmania amazonensis is the etiologic agent of cutaneous leishmaniasis, an immune-driven disease causing a range of clinical symptoms. Infections caused by L. amazonensis suppress the activation and function of immune cells, including macrophages, dendritic cells, and CD4⁺ T cells. In this study, we analyzed the course of infection as well as the leishmanicidal effect of intralesional UTP treatment in L. amazonensis-infected BALB/c mice. We found that UTP treatment reduced the parasitic load in both footpad and lymph node sites of infection. UTP also boosted Th1 immune responses, increasing CD4⁺ T cell recruitment and production of IFN-γ, IL-1β, IL-12, and TNF-α. In addition, the role of UTP during innate immune response against L. amazonensis was evaluated using the air pouch model. We observed that UTP augmented neutrophil chemoattraction and activated microbicidal mechanisms, including ROS production. In conclusion, our data suggested an important role for this physiological nucleotide in controlling L. amazonensis infection, and its possible use as a therapeutic agent for shifting immune responses to Th1 and increasing host resistance against L. amazonensis infection.     

Potent induction of TNF-α during interaction of immune effectors with oral tumors as a potential mechanism for the loss of NK cell viability and function

The inhibitory role of TNF-α on survival of naïve and IL-2 treated NK cells has been demonstrated in the past. However, its effect on the function of these cells against tumor cells, in particular against oral tumors has not been established. We investigated the significance of secreted TNF-α in death and functional loss of splenocytes and NK cells in ex-vivo cultures with oral tumors. Oral tumors trigger potent secretion of TNF-α by human and murine immune effectors. Absence of TNF-α increases the cytotoxic activity and secretion of IFN-γ by IL-2 treated splenocytes and NK cells in co-cultures with MOK L2D1+/p53?/? oral tumor cells. IL-2 treated splenocytes and NK cells from TNF-α ?/? mice survive and proliferate more when compared to cells from TNF-α +/+ mice. Cell death induced by F. nucleatum, an oral bacteria, in TNF-α ?/? splenocytes are considerably lower than that induced in TNF-α +/+ splenocytes where potent release of TNF-α is reproducibly observed. Addition of exogenous rTNF-α to IL-2 treated splenocytes and NK cells decreased survival and function of splenocytes and NK cells obtained from TNF-α ?/? mice against oral tumors. These findings suggest that potent induction of TNF-α during interaction of immune effectors with oral tumors and/or oral bacteria is an important factor in decreasing the function and survival of cytotoxic immune effectors. Strategies to neutralize TNF-α may be beneficial in the treatment of oral cancers.     

Immunostimulatory Effect of Enzyme-Modified <Emphasis Type="Italic">Hizikia fusiforme</Emphasis>in a Mouse Model In Vitro and Ex Vivo

Hizikia fusiforme, a brown seaweed, has been utilized as a health food and in traditional medicine. In this study, we investigated whether enzyme-modified H. fusiforme extracts (EH) have immunological effects compared with normal H. fusiforme extracts (NH). The effects of NH and EH on immune responses were investigated by assessing nitric oxide (NO) production, phagocytosis, and cytokine secretion in RAW 264.7 murine macrophages and mice. Also, fucosterol was evaluated to find the active component of NH and EH by addressing cytotoxicity test and NO production. Both of NH and EH significantly increased cell viability and NO synthesis. Tumor necrosis factor-α (TNF-α) expression was more induced by EH with LPS treatment. Phagocytic activity, as the primary function of macrophages, was markedly induced by EH treatment. Additionally, EH encouraged splenocyte proliferation and recovered the levels of cytokines IL-1β, IL-6, and TNF-α in mice. Finally, fucosterol increased NO production with no cytotoxicity, which means that fucosterol is an active component of EH. In conclusion, EH has the potential to modulate immune function and could offer positive therapeutic effect for immune system diseases.     

Effects of Hydrogen-Rich Saline on Hepatectomy-Induced Postoperative Cognitive Dysfunction in Old Mice

This study aims to investigate the protective effects and underlying mechanisms of hydrogen-rich saline on the cognitive functions of elder mice with partial hepatectomy-induced postoperative cognitive dysfunction (POCD). Ninety-six old male Kunming mice were randomly divided into 4 groups (n?=?24 each): control group (group C), hydrogen-rich saline group (group H), POCD group (group P), and POCD?+?hydrogen-rich saline group (group PH). Cognitive function was subsequently assessed using Morris water-maze (MWM) test. TNF-α and IL-1β levels were measured by enzyme-linked immunosorbent assay (ELISA) and immunohistochemistry, along with NF-κB activity determined by ELISA. The morphology of hippocampal tissues were further observed by HE staining. Learning and memory abilities of mice were significantly impaired at day 10 and day 14 post-surgery, as partial hepatectomy significantly prolonged the escape latency, decreased time at the original platform quadrant and frequency of crossing in group P when compared to group C (p?<?0.05). The surgery also increased the contents of TNF-α, IL-1β, and NF-κB activity at all time points after surgery (p?<?0.05). The introduction of hydrogen-rich saline (group PH) partially rescued spatial memory and learning as it shortened escape latency and increased time and crossing frequency of original platform compared to group P (p?<?0.05). Moreover, such treatment also decreased TNF-α and IL-1β levels and NF-κB activity (p?<?0.05). In addition, cell necrosis in the hippocampus induced by hepatectomy was also rescued by hydrogen-rich saline. Hydrogen-rich saline can alleviate POCD via inhibiting NF-κB activity in the hippocampus and reducing inflammatory response.     

Effects of Selenium Supplementation on Gene Expression Levels of Inflammatory Cytokines and Vascular Endothelial Growth Factor in Patients with Gestational Diabetes

Selenium is known to exert multiple beneficial effects including anti-inflammatory actions. The aim of the study was to evaluate the effects of selenium supplementation on gene expression levels of inflammatory cytokines and vascular endothelial growth factor (VEGF) in women with gestational diabetes (GDM). This randomized double-blind, placebo-controlled trial was carried out among 40 subjects diagnosed with GDM aged 18–40 years old. Subjects were randomly allocated into two groups to receive either 200 μg/day selenium supplements (n = 20) or placebo (n = 20) for 6 weeks. Gene expression of inflammatory cytokines and VEGF were assessed in lymphocytes of GDM women with RT-PCR method. Results of RT-PCR indicated that after the 6-week intervention, compared with the placebo, selenium supplementation downregulated gene expression of tumor necrosis factor alpha (TNF-α) (P = 0.02) and transforming growth factor beta (TGF-β) (P = 0.01), and upregulated gene expression of VEGF (P = 0.03) in lymphocytes of patients with GDM. There was no statistically significant change following supplementation with selenium on gene expression of interleukin (IL)-1β and IL-8 in lymphocytes of subjects with GDM. Selenium supplementation for 6 weeks in women with GDM significantly decreased gene expression of TNF-α and TGF-β, and significantly increased gene expression of VEGF, but did not affect gene expression of IL-1β and IL-8. Clinical trial registration number http://www.irct.ir: IRCT201612045623N95.     

Effect of influenza A virus and bacterial lipopolysaccharide on proliferation and expression of cytokines and other cellular factors in the endothelial cell line ECV-304

Viral infection and bacterial lipopolysaccharide (LPS) cause endothelial-cell dysfunction. The aim of the current study was to investigate the effect of influenza A virus and LPS from Escherichia coli on the proliferative activity and gene expression of cytokines and cellular factors (TNFα, TGFβ, IFN-γ, MMP-9, NF-κB, Rho A, eNOS, and iNOS) in human endothelial cells ECV-304. It was found that ECV-304 cells infected with very low infectious doses of influenza virus acquired the capacity for the long-term active proliferation (over eight passages). Addition of LPS from E. coli reduced the virus-stimulated cell proliferation. It was shown that influenza virus and LPS affected the gene expression of cytokine and other cellular factors. When endothelial cells were infected with influenza A virus in the presence of LPS, there was a significant increase in the expression of several genes and the expression pattern of certain genes was modified. Expression of MMP-9 gene inhibited by the virus and LPS separate exposure significantly increased during the first day after addition of the virus and LPS simultaneously. The same was true for the IFN-γ gene expression. TNFα gene was active only for 1–3 days whereas the expression of TGFβ, eNOS, iNOS, NF-κB and Rho A genes increased significantly on the fifth day, as it was observed with the cells treated with LPS only. Thus, the influenza A virus and LPS change the physiological state of endothelial cells. This occurred during various time periods (as well as at various degrees of viral infection) produced by different cellular factors and, possibly, involved different signaling pathways.     

Anti-viral Activities of <Emphasis Type="Italic">Oroxylum indicum</Emphasis> Extracts on Chikungunya Virus Infection

Chikungunya virus (CHIKV) is a re-emerging mosquito-borne alphavirus that poses a threat to human worldwide. Driven by the lack of approved medication and vaccination, research on anti-Chikungunya agents has received great attention. In an effort to determine potential inhibitor of CHIKV, this study aimed at investigating the potential anti-viral activity of Oroxylum indicum extracts towards CHIKV-infected Vero cells. The virucidal, pre- and post-treatment effects of O. indicum were evaluated, using the maximum non-toxic dose of O. indicum methanol and aqueous extracts as determined by cytotoxicity assay. The viral inhibitory effect was assessed by the morphological changes of Vero cells and further confirmed by plaque assay. Both methanol and aqueous extracts of O. indicum had similar cytotoxicity in Vero cells. Interestingly, the virucidal effect of O. indicum aqueous extract revealed a significant reduction on the viral titre (p < 0.05). The prophylactic effect of aqueous extract was demonstrated when the pre-treated cells exhibited a significant anti-CHIKV activity (p < 0.05). However, methanol extract of this plant exerted an anti-viral activity against CHIKV only to a certain extent. Therefore, the aqueous extract of this plant has a potential to inhibit the virus and acts as prophylactic agent against CHIKV. Further studies however are needed to substantiate the finding and to determine the important compound of this plant as well as the mechanism of action in treating CHIKV infection.     

Changes in Immunity,Expression of some Immune-Related Genes of Shabot Fish, <Emphasis Type="Italic">Tor grypus</Emphasis>, Following Experimental Infection with <Emphasis Type="Italic">Aeromonas hydrophila</Emphasis>: Effects of Autochthonous Probiotics

In this study, the effects of orally administrated two native probiotics (Lactobacillus plantarum and Lactobacillus delbrueckii ssp. bulguricus), isolated from the intestine of Shabot fish, Tor grypus, on some immune response parameters and immune-related genes expression against Aeromonas hydrophila in T. grypus were evaluated. Four hundred and eighty juveniles weighing 45?±?10 g were randomly divided into four groups (with three replications) and fed with the experimental diet containing 5?×?10⁷ cfu g^?1 of L. plantarum (G1), Lactobacillus bulgaricus (G2), Lactobacillus casei (G3), and a control diet (without probiotics) for 60 continuous days. At the end of the dietary treatments, fish were challenged with a lethal concentration of A. hydrophila (5?×?10⁸ CFU ml^?1) via intra peritoneal (i.p) injection. Blood and head kidney samples were taken from six fish in each treatment before challenging and 6, 12, 24, and 48 h and also 7 days after injection. The results showed that lysozyme, complement, bactericidal, and NBT activity of probiotic-treated groups were significantly elevated (P?<?0.05). The IL-8, IL-1β, and TNF-α gene expressions were significantly higher in all probiotic-treated groups (P?<?0.05). Meanwhile, a high direct correlation was observed between serum immune parameters and expression of immune-related genes (P?<?0.0001); furthermore, the highest correlation (R ²?=?0.634, P?<?0.0001) was recorded between IL-1β expression and NBT activity. It can be concluded that not only two native probiotics strains stimulate serum immune responses parameters and immune-related gene expression in T. grypus, but also a high correlation was seen among these indices. The study suggests that gastrointestinal colonization is preferred for host specificity as the strain previously derived from shabot fish displayed better colonization than the non-indigenous bacteria strain such as L. casei. Therefore, these native probiotics bacteria can be accounted as suitable candidates to immune stimulation in fish.     

A critical role for regulatory T cells in driving cytokine profiles of Th17 cells and their modulation of glioma microenvironment

IL-17A, produced by Th17 cells, may play a dual role in antitumor immunity. Using the GL261-glioma model, we investigated the effects of Th17 cells on tumor growth and microenvironment. Th17 cells infiltrate mouse gliomas, increase significantly in a time-dependent manner similarly to Treg and do not express Foxp3. To characterize the direct effects of Th17 cells on GL261 murine gliomas and on tumor microenvironment, we isolated IL-17-producing cells enriched from splenocytes derived from naïve (nTh17) or glioma-bearing mice (gTh17) and pre-stimulated in vitro with or without TGF-β. Spleen-derived Th17 cells co-expressing IL-17, IFN-γ and IL-10, but not Treg marker Foxp3, were co-injected intracranially with GL261 in immune-competent mice. Mice co-injected with GL261 and nTh17 survived significantly longer than gTh17 (P < 0.006) and gliomas expressed high level of IFN-γ and TNF-α, low levels of IL-10 and TGF-β. In vitro IL-17 per se did not exert effects on GL261 proliferation; in vivo gliomas grew equally well intracranially in IL-17 deficient and wild-type mice. We further analyzed relationship between Th17 cells and Treg. Treg were significantly higher in splenocytes from glioma-bearing than naïve mice (P = 0.01) and gTh17 produced more IL-10 than IFN-γ (P = 0.002). In vitro depletion of Treg using PC61 in splenocytes from glioma-bearing mice causes increased IL-17/IFN-γ cells (P = 0.007) and decreased IL-17/IL-10 cells (P = 0.03). These results suggest that Th17 polarization may be induced by Treg and that Th17 cells in gliomas modulate tumor growth depending on locally produced cytokines.