Plaque-forming Cell Assay Research Articles

Immunotoxicity of 2-Acetyl-4-tetrahydroxybutylimidazole in BALB/c mice with different vitamin B6 nutritional statuses

Caramel color is a widely used food pigment, and 2-Acetyl-4-tetrahydroxybutylimidazole (THI) is a by-products of Class III caramel color. Some studies have shown that THI can reduce the number of peripheral blood lymphocytes. However, the comprehensive mechanism of THI immunotoxicity requires further study. In this study, the effects of THI on lymphocyte count, humoral immunity, cellular immunity and nonspecific immunity were determined and the effect of the nutritional status of VB6 on THI immunotoxicity was evaluated. Female BALB/c mice were divided into 3 groups and fed chow containing different doses of VB6: VB6-normal (6 mg/kg VB6), VB6-deprived (0.5 mg/kg VB6) or VB6-enhanced (12 mg/kg VB6) feed. Each group was further divided into 4 subgroups and treated with THI (0.5, 2.5 or 12.5 mg/kg bw) or the solvent control by gavage for 30 days. The thymic cortical thickness was measured with ViewPoint; the proportions of major immune cells and T cells in peripheral blood and tissues were detected via flow cytometry; the transformation and proliferation abilities of T and B cells were detected via T and B lymphocyte proliferation assays; NK cell activity was assessed via lactate dehydrogenase assays; humoral immune function was assessed via plaque-forming cell assays; and the immune function of T lymphocytes was assessed via delayed type hypersensitivity assays. The results showed that compared with those in the corresponding control group, the white blood cell count and lymphocyte count decreased significantly in all the VB6-deprived groups, in the 2.5 and 12.5 mg/kg VB6 groups, and in the 12.5 mg/kg VB6-enhanced group. With increasing THI dose, the thymic cortical layer became thinner. In the thymus, THI increased the proportions of CD3+ T cells and mature CD8+ T cells and decreased the proportions of immature double-positive, double-negative T cells and CD69-expressing lymphocytes. The proportions of naïve T cells and Tcm (central memory T) cells related to homing decreased. The proportion of mature T cells in the spleen decreased significantly. The proliferation of T cells stimulated by ConA decreased after THI exposure. VB6-deficient mice were more sensitive to THI immunotoxicity, and supplementation with VB6 had a certain protective effect on these mice. The results of the PFC and NK cell activity assays indicated that THI exposure might not affect humoral immune or innate immune function.

Effects of 2-acetyl-4-tetrahydroxybutylimidazole exposure during gestation and lactation on maternity and offspring immune function in Balb/c mice

2-Acetyl-4-tetrahydroxybutylimidazole (THI), a by-product of Class Ⅲ caramel color, is generally recognized to cause lymphopenia in mammals. However, it remains unknown whether THI exposure during gestation and lactation causes damage to the immune system of offspring. In this study, pregnant Balb/c mice were gavaged with 0, 0.5, 2.5 and 12.5 mg/kg THI from gestation day (GD) 6 to postanal day (PND) 21, after which we treated another batch of dams from GD6 to PND21 and the offspring for 3 weeks after weaning with 0, 2, 10, 50 mg/L THI in drinking water respectively, and investigated the immunological anomalies of dams and offspring. The results showed that lymphopenia was observed in dams but not in weaning pups on PND21, which were exposed to THI during gestation and lactation. 2 mg/L THI and 2.5 mg/kg THI began to cause a remarkable reduction of the numbers of white blood cells and lymphocytes in dams. Besides both the cellular and the humoral immune response was not affected in weaning pups, which were measured by plaque-forming cell (PFC) assay and delayed-type hypersensitivity (DTH) assay respectively. Furthermore, THI could be detected in the plasma of dams with a dose-dependent manner, but not in that of both female and male weaning pups. In both male and female offspring being treated with 10 and 50 mg/L THI for another 3 weeks after weaning, lymphocytopenia was observed and T lymphocytes including CD4+ and CD8+ cells were significantly reduced in their spleens except lymph nodes. 10 and 50 mg/L THI treatment increased CD4+ and CD8+ single positive cells in thymus of female and male weaning mice. Mitogen-induced proliferation ability of T cells in the spleen and lymph nodes was impaired in female weaning mice exposed 50 mg/L THI, while male weaning mice treated with 10 and 50 mg/L THI showed impairment in the spleen but not lymph nodes. Based on the results in this study, no observed adverse effect level (NOAEL) for 3-week THI treatment in weaning mice was considered to be 2 mg/L (0.30 mg/kg bw for female mice and 0.34 mg/kg bw for male mice). And NOAEL for THI treatment in dams might be set to 0.5 mg/kg bw/day. Collectively from the perspective of NOAEL, offspring are not more sensitive than dams or adult mice.

Next-generation PFAS 6:2 fluorotelomer sulfonate reduces plaque formation in exposed white-footed mice.

6:2 fluorotelomer sulfonate (6:2 FTS) has been used as a replacement for legacy per- and polyfluoroalkyl substances (PFAS). We assessed reproductive and developmental effects in a human-wildlife hybrid animal model based on the association of adverse effects linked to legacy PFAS with these sensitive life stages. In this study, white-footed mice were exposed orally to 0, 0.2, 1, 5, or 25 mg/kg-day 6:2 FTS for 112 days (4 weeks premating exposure plus at least 4 weeks mating exposure). Pregnancy and fertility indices were calculated, and litter production, total litter size, live litter size, stillbirths, litter loss, average pup weight, and pinna unfolding were assessed. Sex steroid and thyroid hormone serum levels were assessed. Body weight, histopathology, and immune function were also assessed in this study. Reproductive endpoints were not significantly altered in response to 6:2 FTS. Spleen weight increased in male mice dosed with 6:2 FTS. Immune function determined via a plaque-forming cell (PFC) assay was decreased in both male and female mice in the 2 highest doses. A low benchmark dose was calculated based on PFCs as the critical effect and was found to be 2.63 and 2.26 mg/kg-day 6:2 FTS in male and female mice, respectively. This study characterizes 6:2 FTS as being potentially immunotoxic with little evidence of effect on reproduction and development; furthermore, it models acceptable levels of exposure. These 2 pieces of information together will aid regulators in setting environmental exposure limits for this PFAS currently thought to be less toxic than other PFAS.

Cigarette Smoke Exposure Inhibits Early Phase of Antibody Production through Inhibition of Immune Functions in Alveolar Macrophage

Background::Cigarette smoke (CS) is inhaled into the lung. Alveolar macrophage (AM) is known to play an important role in the lung immune system. However, the relationship between AM functions and antibody production by CS is not fully investigated.Objective::Therefore, we investigated the effects of AM from CS exposed mice on antibody production. Mice were exposed to 20 cigarettes/day for 10 days. AM were obtained by broncho-alveolar lavage. Antibody production was analyzed by plaque-forming cell assay using seep red blood cell (SRBC) as antigen.Methods::B cell proliferation was analyzed by 3H-thymidine incorporation. Phagocytic activity using fluorescein isothiocyanate-labeled SRBC and expressions of surface antigens on AM were analyzed by flow cytometry. Cytokines and NF-κB mRNA expressions of AM were analyzed by RTPCR.Results and Discussion:Antibody production was decreased at the induction phase, but not at the expression phase by AM from smoked mice (SM) compared with non-smoked mice (NSM). B cell proliferation was decreased by cigarette extracts dose-dependently. Phagocytic activity of AM was decreased in SM compared with NSM. Expression of surface antigens on AM was decreased in SM compared with NSM. Cytokines or NF-κB mRNA expressions of AM were decreased in SM compared with NSM.Conclusion::These results suggest that the inhibition of antibody production by cigarette smoking is caused by the inhibition of phagocytosis and expressions of surface antigens associated with antigen presentation. Such inhibition of AM functions may increase the risk of bacterial and virus infections.

Effects of an environmentally relevant PCB-mixture on immune function, clinical chemistry, and thyroid hormone levels in adult female B6C3F1 mice

ABSTRACT Polychlorinated biphenyls (PCBs) have been assessed for immunotoxicity; however, humans and wildlife are exposed to multiple PCBs environmentally. Therefore, the aim of this study was to examine the effects of a complex 37 PCB congener mixture identified in blubber specific to dolphins residing in the estuarine waters of Charleston, South Carolina. Immunotoxicity was determined in adult female B6C3F1 mice by assessing lymphocyte proliferation, splenic and thymic immunophenotypes, and IgM production. Mice were exposed via oral gavage to the PCB-mixture (0, 1.8, 3.6, 7.1, or 14.3 mg/kg/day) for 28 days to yield a targeted total administered dose (TAD) 0, 50, 100, 200, or 400 mg/kg. Significant increased liver weight occurred at the highest treatment. IgM production was suppressed compared to control for all treatments. Numbers of thymic CD4+/CD8+, CD4-/CD8-, and CD4+/CD8- cells were not altered, but numbers of thymic CD4-/CD8+ cells were significantly increased in the highest treatment. Lymphocyte proliferation was not markedly affected by any treatment. The numbers of splenic CD4/CD8 T-cells or MHCII+ cells were not significantly changed. Humoral immunity using the plaque-forming cell assay for determining the specific IgM antibody-forming cell response appeared to be the most sensitive endpoint affected. As the lowest concentration tested resulted in decreased IgM production and total and free thyroxine (T4) serum levels a NOAEL was not identified. The calculated ED50 for suppression of IgM production was 2.4 mg/kg/day.

A soluble FcγR homolog inhibits IgM antibody production in ayu spleen cells.

Classical Fc receptors (FcRs) mediate the binding to and recognition of the Fc portion of antibodies and play an important role during immune responses in mammals. Although proteins similar to soluble FcRs have been identified in fish, little is known about the role of such proteins in fish immunity. Here, we cloned a cDNA sequence encoding a soluble Fc receptor for an immunoglobulin G (FcγR) homolog from ayu (Plecoglossus altivelis) (PaFcγRl). The predicted protein was composed of two immunoglobulin C2-like domains but lacked a transmembrane segment and a cytoplasmic tail. The PaFcγRl transcripts were distributed at low levels in all tested tissues, but significantly increased after Vibrio anguillarum infection. The PaFcγRl protein was expressed in the head kidney, trunk kidney, and neutrophils. Recombinant PaFcγRl (rPaFcγRl) was secreted when transfected into mammalian cells and the native protein was also detected in serum upon infection. rPaFcγRl was also demonstrated to bind to ayu IgM, as assessed by cell transfection. Suppressive activity of the recombinant mature protein of PaFcγRl (rPaFcγRlm) on in vitro anti-sheep red blood cell (SRBC) responses was detected by a modified hemolytic plaque forming cell assay. In conclusion, our study revealed that PaFcγRl is closely involved in the negative regulation of IgM production in the ayu spleen.

Immunomodulatory activities of Acacia catechu, a traditional thirst quencher of South India

BackgroundAcacia catechu has been widely used in Ayurveda for treating many diseases. Its heartwood extract is used in asthma, cough, bronchitis, colic, diarrhea, dysentery, boils, skin afflictions, sores and for stomatitis. The decoction of heartwood is used for drinking purpose in southern part of India especially in Kerala. ObjectiveThe current study was carried out to evaluate immunomodulatory effects of heartwood extracts of A. catechu in Swiss albino mice. Material and methodsIn vivo immunomodulatory activity was analyzed by hemagglutinating antibody (HA) titer, plaque forming cell assay and delayed type hypersensitivity (DTH). In vitro immunomodulatory potential of the extracts was studied using peritoneal macrophages and splenocytes from mice. Effect of extracts on phagocytic activity of macrophages was analyzed by nitroblue tetrazolium (NBT) reduction assay and cellular lysosomal enzyme assay. Anti-inflammatory activity was studied by nitric oxide (NO) assay and production of TNF-α and IL-10. ResultsA dose dependent increase in antibody titer was observed with extracts treatment. Treatment with extracts produced an enhancement in the number of antibody producing cells in the spleen. DTH reaction was significantly decreased with extracts treatment. An increased phagocytic response was shown by peritoneal macrophages on treatment with the extracts as evidenced by its effect on NBT reduction and cellular lysosomal enzyme activity. The extracts inhibited the release of pro-inflammatory cytokine TNF-α and the production of NO. IL-10 production was significantly increased after extract treatment. ConclusionThe results of the present study indicate the immunomodulatory effects of A. catechu extracts on humoral, cell mediated and non-specific immune functions.

Catechin rich butanol fraction extracted from Acacia catechu L. (a thirst quencher) exhibits immunostimulatory potential

Acacia catechu L., (Fabaceae) named as “catechu” is a plant, the decoction of heartwood of which is daily consumed as thirst quencher by a good percentage of the population in South India. The plant is mainly distributed in India and other Asian countries. It has been used in Indian traditional medicine for the treatment of asthma, bronchitis, colic, diarrhea, boils, skin afflictions, sores and stomatitis. The present investigation was aimed to study the immunomodulatory effects of different fractions of ethanol extract of A. catechu heartwood and HPLC analysis of the active fraction. Three fractions namely, butanol, chloroform and ethyl acetate were prepared from ethanol extract of A. catechu heartwood. Each of these fractions was assessed for its immunomodulatory activity. In vivo immunomodulatory activity was analyzed by sheep red blood cells (SRBC) specific hemagglutinating antibody titer, plaque-forming cell assay and delayed type hypersensitivity (DTH) reaction in Swiss albino mice. In vitro immunomodulating potential of the fractions was studied using murine peritoneal macrophages and splenocytes. Non-specific immune functions such as phagocytosis (nitroblue tetrazolium reduction assay and cellular lysosomal enzyme assay), nitric oxide (NO) production and cytokine release (TNF-α and IL-10) were studied in macrophages. In addition, splenocyte proliferation was also studied. In the in vivo experiments, butanol and chloroform fractions showed an increase in antibody titer dose-dependently. At higher dose (400 mg/kg b. w.) treatment the butanol fraction produced an enhancement in the number of plaque-forming cells (antibody producing cells) in the spleen. SRBC induced DTH reaction was significantly increased with butanol fraction in a dose-dependent manner. Peritoneal macrophages showed an increased phagocytic response on treatment with butanol fraction (100 μg/mL) as evidenced by its effect on nitroblue tetrazolium reduction and cellular lysosomal enzyme activity. All three fractions inhibited the production of NO and the release of TNF-α. Interleukin-10 production was significantly increased after treatment with butanol fraction. High-performance liquid chromatography analysis of the butanol fraction showed the presence of high concentration of catechin. The results suggested that butanol fraction of ethanol extract of A. catechu heartwood had immunomodulatory effects on non-specific, humoral, and cell-mediated immune functions. This study may be useful in validating the rationality of daily consumption of decoction of A. catechu and also its use in traditional medicine system. The study also suggests the possible use of A. catechu in the immunostimulatory herbal preparations.

Developmental immunotoxicity is not associated with the consumption of transgenic Bt rice TT51 in rats

TT51 is a transgenic strain of Bt rice generated by fusing a synthetic CryAb/Ac gene into MingHui rice. In this study, rats from F0, F1, and F2 generations were fed a diet with 60% TT51 rice, MingHui rice, or nominal-origin rice. The study focused on developmental immunotoxicity in F1 and F2 offspring after long-term consumption of TT51. A wide range of immunological parameters was monitored in this two-generation study on reproductive toxicity. The experiments were performed on F1 and F2 offspring at postnatal days 21 and 42. No adverse clinical effects were observed in any of the experimental groups. In addition, histopathology observations and immunotoxicity tests, including hematological indicators, spleen lymphocyte subsets, natural killer cell activity, lymphoproliferative response, and plaque-forming cell assay, revealed no significant difference between the groups. These results indicated that developmental immunotoxicity was not associated with a diet of transgenic Bt rice TT51, compared to the parental MingHui rice.

Stimulation of murine immune response by Clerodendrum infortunatum

Background: Clerodendrum infortunatum is a medicinal plant found especially in the Sub-Himalayan regions of West Bengal. Various tribal communities of this region have been using these plants as a source of natural medicine. Objective: Medicinal values of C. infortunatum are well established, but the immunomodulatory properties have never been studied. Therefore, we wanted to investigate various immunomodulatory activities of this plant. Materials and Methods: Several parameters such as plaque-forming cell assay, hemagglutination titer, phagocytic activity of macrophages, and inhibition of lipopolysaccharide (LPS) were performed. Moreover, effects of C. infortunatum on the weight of various organs and total serum protein, albumin, and globulin levels were also determined. The phytochemical fingerprints of C. infortunatum extracts were obtained from Fourier transform infrared and gas chromatography-mass spectrometry analysis. Results: The macrophage stimulation and plaque-forming cell numbers (using sheep red blood cell as antigen) were significantly proliferated and found optimum at 100 and 250 mg/kg doses of leaf and root extracts simultaneously. The extent of stimulation in mural humoral immunity was in the order of leaf > root > stem. The anti-inflammatory activity of C. infortunatum was confirmed by the inhibition of LPS-induced nitric oxide synthesis by macrophages. Conclusion: It can be concluded that 70% hydromethanolic extract of C. infortunatum has profound immunomodulation potentials in the murine model, stimulating both humoral and innate form of the immune system. Abbreviations used: PFC: Plaque-forming cell assay; HA titer: Hemagglutination titer; LPS: Lipopolysaccharide; FTIR: Fourier transform infrared; GC-MS: Gas chromatography-mass spectrometry; SDS: Sodium dodecyl sulfate; sRBC: Sheep red blood cell; PBS: Phosphate buffer saline; PC: Phagocytic capacity; PI: Phagocytic index; NaCl: Sodium chloride; CaCl2: Calcium chloride; NBT: Nitro blue tetrazolium; DMSO: Dimethyl sulfoxide; KOH: Potassium hydroxide; H2SO4: Sulfuric acid; HOCl: Hypochlorous acid; H2O2: Hydrogen peroxide; EDTA: Ethylenediaminetetraacetic acid.

Anti-inflammatory and immunomodulatory activities of Inula cappa roots (Compositae).

Background The medicinal properties of Inula cappa, a perennial shrub, are ascribed to its roots. In this article, we study the anti-inflammatory and immunomodulatory activities of I. cappa root extracts based on its utilization in traditional medicine-treatment of rheumatoid arthritis, menoxenia, fever, jaundice and many others. Methods Anti-inflammatory and immunomodulatory activities of I. cappa extracts were studied. The anti-inflammatory activity was determined by carrageenan-induced rat paw edema and cotton pellet-induced granulation method while the immunomodulatory activity was estimated by phagocytic assay method, hemagglutinating antibody (HA) titer assay, delayed-type hypersensitivity assay method, plaque forming cell assay and determination of immunoprophylatic activity. Results The methanolic extract showed maximum reduction in the rat paw edema and showed significant inhibition of the cotton pellet-induced granulomas in rats. The methanolic extract also showed potential immunomodulatory activity in all the assays performed. Two sesquiterpenes, isoalantolactone and germacranolide were also isolated from the methanolic extract. Conclusions The present study supports the evidence that the roots of I. cappa can be used as a potent anti-inflammatory and immunomodulatory agent.

Discovery and Pharmacological Characterization of Novel Quinazoline-Based PI3K Delta-Selective Inhibitors.

Inhibition of the lipid kinase PI3Kδ is a promising principle to treat B and T cell driven inflammatory diseases. Using a scaffold deconstruction-reconstruction strategy, we identified 4-aryl quinazolines that were optimized into potent PI3Kδ isoform selective analogues with good pharmacokinetic properties. With compound 11, we illustrate that biochemical PI3Kδ inhibition translates into modulation of isoform-dependent immune cell function (human, rat, and mouse). After oral administration of compound 11 to rats, proximal PD markers are inhibited, and dose-dependent efficacy in a mechanistic plaque forming cell assay could be demonstrated.

Investigations into the Immunotoxicity and Allergic Potential Induced by Topical Application of N-Butylbenzenesulfonamide (NBBS) in a Murine Model

N-Butylbenzene sulfonamide (NBBS) is a commonly used plasticizer found in numerous products. Due to its extensive use, lack of adequate toxicological data, and suspicion of toxicity based on the presence of structural alerts, it was nominated to the National Toxicology Program for comprehensive toxicological testing. The purpose of this study was to evaluate the potential for hypersensitivity and immune suppression following dermal exposure to NBBS using a murine model. NBBS tested negative in a combined irritancy/local lymph node assay (LLNA), classifying it as nonirritating and nonsensitizing. To estimate the immunosuppressive potential of NBBS, assays that assessed immunotoxicity were performed, including the immumnoglobulin (Ig) M response to T-cell-dependent antigen sheep red blood cells (SRBC), using the plaque-forming cell (PFC) assay and immune cell phenotyping. After a 28-d treatment with NBBS, mice exposed to the lowest concentration (25% NBBS) showed a significant increase in IgM-producing B cells in the spleen. No marked changes were identified in immune cell markers in the lymph node. In contrast to body weight, a significant elevation in kidney and liver weight was observed following dermal exposure to all concentrations of NBBS. These results demonstrate that dermal exposure to NBBS, other than liver and kidney toxicity, did not apparently induce immunotoxicity in a murine model.

The transferability from rat subacute 4-week oral toxicity study to translational research exemplified by two pharmaceutical immunosuppressants and two environmental pollutants with immunomodulating properties

Exposure to chemicals may have an influence on the immune system. Often, this is an unwanted effect but in some pharmaceuticals, it is the intended mechanism of action. Immune function tests and in depth histopathological investigations of immune organs were integrated in rodent toxicity studies performed according to an extended OECD test guideline 407 protocol. Exemplified by two immunosuppressive drugs, azathioprine and cyclosporine A, and two environmental chemicals, hexachlorobenzene and benzo[a]pyrene, results of subacute rat studies were compared to knowledge in other species particular in humans. Although immune function has a high concordance in mammalian species, regarding the transferability from rodents to humans various factors have to be taken into account. In rats, sensitivity seems to depend on factors such as strain, sex, stress levels as well as metabolism. The two immunosuppressive drugs showed a high similarity of effects in animals and humans as the immune system was the most sensitive target in both. Hexachlorobenzene gave an inconsistent pattern of effects when considering the immune system of different species. In some species pronounced inflammation was observed, whereas in primates liver toxicity seemed more obvious. Generally, the immune system was not the most sensitive target in hexachlorobenzene-treatment. Immune function tests in rats gave evidence of a reaction to systemic inflammation rather than a direct impact on immune cells. Data from humans are likewise equivocal. In the case of benzo[a]pyrene, the immune system was the most sensitive target in rats. In the in vitro plaque forming cell assay (Mishell–Dutton culture) a direct comparison of cells from different species including rat and human was possible and showed similar reactions. The doses in the rat study had, however, no realistic relation to human exposure, which occurs exclusively in mixtures and in a much lower range. In summary, a case by case approach is necessary when testing immunotoxicity. Improvements for the translation from animals to humans related to immune cells can be expected from in vitro tests which offer direct comparison with reactions of human immune cells. This may lead to a better understanding of results and variations seen in animal studies.

Immunomodulatory activity of Nerium indicum through inhibition of nitric oxide and cyclooxygenase activity and modulation of TH1/TH2 cytokine balance in murine splenic lymphocytes.

Nerium indicum is a medicinal plant which is used in the treatment of wide range of illness in different ethnopharmacological practices. We have studied the immunomodulatory activity of the 70% hydro-methanolic extract of N. indicum leaves (NILE) by studying plaque forming cell assay, haemagglutination titre, cell proliferation assay, inhibition of nitric oxide expression and cyclooxygenase activity, estimation of IL-2, IFN-γ, IL-4, IL-10, TNF-α and IgM levels. Furthermore, we have characterized NILE with Fourier Transform Infra-red (FTIR) spectroscopy. The results displayed that NILE possessed potent immunomodulatory activity by up-regulating IL-2, IFN-γ, IL-10 expression and down-regulating IL-4, TNF-α expression in vitro. NILE also stimulated the in vivo expression of immunoglobulin level in mouse. FTIR analysis revealed phytocompounds of diverse chemical nature present in NILE. Thus, the potent immunomodulatory activity of N. indicum may have serious implications in the treatment of inflammatory diseases in the future.

Effects of short-term oral combined exposure to environmental immunotoxic chemicals in mice

People are constantly exposed to environmental chemicals through contact with the atmosphere or by ingestion of food. Therefore, when conducting safety assessments, the immunotoxic effects of combinations of chemicals in addition to toxicities produced by each chemical alone should be considered. The objective of the studies reported here were to demonstrate the combined effects of three well-known environmental immunotoxic chemicals -- methoxychlor (MXC), an organochlorine compound; parathion (PARA), an organophosphate compound; and piperonyl butoxide (PBO), an agricultural insecticide synergist -- by using a short-term oral exposure method. Seven-week-old Balb/cAnN mice received daily oral exposure to either one or two of the environmental immunotoxic chemicals for 5 consecutive days. On Day 2, all mice in each group were immunized with sheep red blood cells (SRBC), and their SRBC-specific IgM responses were analyzed by using an enzyme-linked immunosorbent assay and plaque-forming cell assay. T- and B-cell counts in the mouse spleens were also assessed via surface antigen expression. Mice that received MXC + PARA and PBO + MXC treatment showed marked decreases in SRBC-specific IgM production and T- and B-cell counts compared with those in mice that received vehicle control or the corresponding individual test substance. This suggests that simultaneous exposure to multiple environmental chemicals increases the immunotoxic effects of the chemicals compared to individual exposure.

Stimulation of Murine Immune Response by the Tubers of Dioscorea alata L. of North-Eastern Region of India

Dioscorea alata is a staple food in south-eastern Asia and Africa and also widely consumed in the North-eastern region of India. Its medicinal value is well established, but the immunomodulatory property has never been studied on animal model. Therefore, we have investigated the immunomodulatory activity of D. alata by performing plaque forming cell assay, haemagglutination titer, stimulation of phagocytosis assay, inhibition of lipopolysaccharide (LPS) induced nitric oxide (NO) synthesis, carbon-clearance test, in vitro cell adhesions assay and various cell counts. Effect of D. alata on various organ weight and total serum protein, albumin and globulin levels were also determined. At 100 mg/kg dose the macrophages were highly stimulated (P < 0.001) to proliferate and the plaque forming cell number was also found to be optimum (P < 0.001) at the same dose. Elimination of foreign material from the system was found to be highest at 250 mg/kg dose. Both at 100 and 250 mg/kg doses, the phagocytic activity of the macrophages was increased significantly (P < 0.001). Anti-inflammatory activity of D. alata was proved by the inhibition of LPS induced NO synthesis by macrophages. Therefore, it can be concluded that 70 % hydro-methanolic extract of D. alata has profound immunostimulating potential in the murine model, stimulating both humoral and innate arms of the immune system.

Modulatory Effect of a Unani Formulation (Jawarish amla sada) on Cyclophosphamideinduced Toxicity in Tumour Bearing Mice

Aims: Our aim was to study the modulatory effect of a Unani herbal formulation Jawarish amla sada against cyclophosphamide-induced toxicity in tumour bearing mice. Study Design: Non randomized control study. Place and Duration of Study: The study was conducted at the Department of Medical Elementology and Toxicology, Jamia Hamdard, New Delhi during 2008-10. Methodology: Study was conducted in Swiss albino mice divided in five groups (n=6). Animals were challenged with Ehrlich’s ascites tumour cells (1x10 cells). Cyclophosphamide (50 mg/kg body weight), an alkylating anticancer drug that especially affects humoral immune functions, was injected intraperitoneally in a single dose. The protective effect of Unani drug Jawarish amla sada (250 mg/kg body weight) was studied in tumour bearing animals treated with cyclophosphamide. Immune function assessment Research Article British Journal of Medicine & Medical Research, 2(3): 454-468, 2012 455 test such as plaque forming cell assay (PFC) and biochemical parameters such as activities of antioxidant enzymes and reduced glutathione were measured in mice. Results: Jawarish amla sada significantly modulated the immunosuppressive effect of cyclophosphamide as compared to the group treated with cyclophosphamide. Jawarish amla sada also protected activities of antioxidant enzymes such as catalase, glutathione peroxidase, glutathione reductase and glutathione S-transferase and significantly restored level of reduced glutathione in liver and kidney of tumour bearing mice exposed to cyclophosphamide. Similar protective effect of Jawarish amla sada was observed against elevated lipid peroxidation in these tissues. Conclusion: Jawarish amla sada showed potential to provide protection against toxic effects of cyclophosphamide in tumour bearing mice. The mechanism of action of the drug may be attributed to various antioxidants fortified in this herbal Unani formulation, which is used in the traditional system of medicine in Indian subcontinent against several liver ailments.

Modulatory Effect of a Unani Formulation (Jawarish amla sada) on Cyclophosphamide-induced Toxicity in Tumour Bearing Mice

Aims: Our aim was to study the modulatory effect of a Unani herbal formulation Jawarish amla sada against cyclophosphamide-induced toxicity in tumour bearing mice. Study Design: Non randomized control study. Place and Duration of Study: The study was conducted at the Department of Medical Elementology and Toxicology, Jamia Hamdard, New Delhi during 2008-10. Methodology: Study was conducted in Swiss albino mice divided in five groups (n=6). Animals were challenged with Ehrlich’s ascites tumour cells (1x10 cells). Cyclophosphamide (50 mg/kg body weight), an alkylating anticancer drug that especially affects humoral immune functions, was injected intraperitoneally in a single dose. The protective effect of Unani drug Jawarish amla sada (250 mg/kg body weight) was studied in tumour bearing animals treated with cyclophosphamide. Immune function assessment Research Article British Journal of Medicine & Medical Research, 2(3): 454-468, 2012 455 test such as plaque forming cell assay (PFC) and biochemical parameters such as activities of antioxidant enzymes and reduced glutathione were measured in mice. Results: Jawarish amla sada significantly modulated the immunosuppressive effect of cyclophosphamide as compared to the group treated with cyclophosphamide. Jawarish amla sada also protected activities of antioxidant enzymes such as catalase, glutathione peroxidase, glutathione reductase and glutathione S-transferase and significantly restored level of reduced glutathione in liver and kidney of tumour bearing mice exposed to cyclophosphamide. Similar protective effect of Jawarish amla sada was observed against elevated lipid peroxidation in these tissues. Conclusion: Jawarish amla sada showed potential to provide protection against toxic effects of cyclophosphamide in tumour bearing mice. The mechanism of action of the drug may be attributed to various antioxidants fortified in this herbal Unani formulation, which is used in the traditional system of medicine in Indian subcontinent against several liver ailments.

Chemotypical variations in Withania somnifera lead to differentially modulated immune response in BALB/c mice

Withania somnifera (Ashwagandha) is a plant with known ethnomedicinal properties and its use in Ayurvedic medicine in India is well documented. The present investigation reports on immunomodulatory efficacy of aqueous-ethanol extracts of roots of three selected Withania somnifera chemotypes designated as NMITLI 101R, NMITLI 118R and NMITLI 128R. Each chemotype was administered 10-100 mg/kg orally to BALB/c mice once daily for 14 days. The immunomodulatory consequences were recorded by determining the humoral immune response with the help of hemagglutination, plaque forming cell assay and cellular response by measuring delayed type hypersensitivity reaction. Additionally, other immune parameters such as proliferation of T and B cells, intracellular and secreted Th1 and Th2 cytokines along with modulation in ROS production by peritoneal macrophages were monitored after feeding with lower doses (3-30 mg/kg/day) of these three chemotypes individually. NMITLI 101R incited both humoral and cellular immune response in terms of higher number of antibody producing cells and enhanced foot pad swelling at the 10mg dose as also dose dependent B and T cell proliferations. Levels of intracellular and secreted cytokines post-NMITLI 101R treatment illustrated generation of mixed Th1/Th2 response that remained more polarized towards Th1. This chemotype also generated maximum reactive oxygen species. NMITLI 118R provoked comparatively reduced immune response in all humoral and cellular parameters at lower doses but induced highly polarized Th1 cytokine response. In contrast, NMITLI 128R led to enhanced antibody production with minimal cellular response demonstrating marginally Th2 dominance at a lower dose. Taken together, it may therefore be concluded that there were distinct modulation in the immune response exhibited by the three chemotypes of Withania somnifera and NMITLI 101R appeared to possess a better immunostimulatory activity than the other chemotypes at lower doses.