Delayed-type Hypersensitivity Test Research Articles

Antioxidant and immunomodulatory activities of ethanol extracts from Syzygium cumini L. Skeels and Pogostemon cablin Benth

Syzygium cumini and Pogostemon cablin are mostly cultivated in tropical climates for culinary and perfumery purposes, yet their potential medicinal properties remain underreported. The aim of this study was to examine the antioxidant and immunomodulatory activities of ethanol extracts from S. cumini (EESC) and P. cablin (EEPC). Reflux extraction was carried out using 96% ethanol on the collected plant specimens to produce EESC and EEPC. Secondary metabolites of each extract were identified using gas chromatography-mass spectrometry (GC-MS). The extracts were measured for total phenol and flavonoid levels and 2,2-diphenyl-1-picrylhydrazyl (DPPH) scavenging activity. The immunomodulatory activity test was carried out in vivo by assessing several parameters, including the phagocytic index via the carbon clearance method, organ indices, antibody titers, and delayed-type hypersensitivity (DTH) response, using sheep red blood cells (SRBC) as antigens. The extracts were also examined for their anti-inflammatory activity in acute and chronic inflammation models. In the DPPH antioxidant test, EESC and EEPC had IC50 values of 12.33 µg/mL and 182.17 µg/mL, respectively. Both extracts showed immunosuppressant activity, marked by a phagocytic index of <1. EESC yielded lower organ indices for the liver (p=0.025 at 200 mg/kg BW), spleen (p=0.028 at 100 mg/kg BW), and thymus (p=0.032 at 200 mg/kg BW) compared to the control group. For EEPC, lower organ indices were observed in the liver at 100 mg/kg BW (p=0.005) and 200 mg/kg BW (p=0.031). In the primary antibody titer and DTH tests, both EESC and EEPC showed immunosuppressant activity at 200 and 400 mg/kg BW (p<0.05). The extracts suppressed both the innate and adaptive immune systems. Both EEPC (p=0.004) and EESC at 100 mg/kg BW (p=0.03) significantly reduced serum TNF-α levels. In conclusion, EESC and EEPC have the potential as immunosuppressive and anti-inflammatory agents.

501 The effect of dietary camelina, flaxseed, and canola oil supplementation on skin fatty acid profile, inflammation and immune responses in healthy adult horses

Abstract As n-6 and n-3 fatty acids (FA) have pro- and anti-inflammatory effects, respectively, supplemental oils have the potential to impact inflammation, the immune response, and skin and coat health. This study aimed to evaluate the effects of camelina (CAM), in comparison to canola (OLA) and flaxseed (FLX) oil on the skin FA profile, inflammation, and the immune response in horses. Horses [n = 24; 14.2 ± 4.8 yr; 518 ± 63 kg body weight (BW)] were acclimated to sunflower oil over 4 wk. Groups were balanced by location, age, weight, sex, and breed and randomly assigned one of the three treatments (CAM, OLA, FLX). Horses were fed their assigned oil at a dose of 0.37 g/kg BW for 16 wk. Skin samples were collected on wk 16, and the skin FA composition was analyzed by gas chromatography. On wk 10 and 12, keyhole limpet hemocyanin (KLH) was administered intramuscularly to each horse to stimulate a systemic immune response. Blood was collected on wk 14 and 16 for the evaluation of plasma KLH immunoglobulin G (IgG) concentrations using a modified ELISA protocol and results were presented as optical density units. On wk 12, after the first KLH injection, a delayed-type hypersensitivity (DTH) test was performed by intradermally injecting KLH and the area of swelling was measured using the software ImageJ at 0.5-, 1-, 8-, 24-, 48-, 72-, and 96-h post-injection. Data were analyzed using the GLIMMIX procedure of SAS considering the effects of treatment, time, and their interaction as fixed, and the effects of horse and location as random. Repeated measures on horses were modelled using a spatial power covariance structure. Significance was declared at P ≤ 0.05 and when fixed effects were significant, means were separated using Tukey-Kramer adjustments. Immune and inflammatory responses were produced by the KLH injections; however, there were no differences among treatment groups regarding KLH antibody and DTH response (P > 0.44). The percentages of palmitic (P = 0.02) and stearic acid (P = 0.03) in the skin at wk 16 were greater in FLX as compared with CAM but neither differed from the OLA. The percentage of linoleic acid (P = 0.01) was greater in CAM than OLA but neither differed from FLX at wk 16. The percentage of all other FA was similar among treatment groups (P > 0.06). These results suggest that CAM is comparable to FLX and OLA regarding support of an immune and inflammatory response, and the oil FA composition was not reflected in the skin FA profile after feeding for 16 wk as hypothesized. In conclusion, at the dose provided, CAM supports a similar immune and inflammatory response to KLH as FLX and OLA; however, the lack of incorporation of FA from the diet into the skin should be further explored.

Immunomodulation Induced in BALB/c Mice after Subacute Exposure to Hydroalcoholic Extract of Artimisia Dracunculus.

Tarragon, with the scientific name of Artemisia dracunculus, is a perennial herbaceous plant with a wide spectrum of pharmacologic properties. In the current investigation, BALB/c mice were used to examine the immunomodulatory effects of hydroalcoholic extract of tarragon (HET). Mice were treated with hydroalcoholic extract of Artimisia dracunculus (HET) at two doses (250 and 500 mg/kg) for 14 days. The host hematological parameters, spleen cellularity histopathology, hemagglutination titer assay (HA), delayed-type hypersensitivity (DTH) responses, IFN-γ and IL-4 levels produced by spelenocytes, and the proliferation of lymphocytes were assayed. HET at a high dose significantly could increase the number of white blood cells and lymphocytes compared to the control group. The lymphocyte proliferation in exposure to PHA significantly increased in the HET group at both doses compared to the control group, whilst this index in the presence of LPS increased significantly for the 500 mg/kg-HET group only. Moreover, in the HA and DTH tests, HET significantly increased the proliferation of lymphocytes as compared with the control group. Furthermore, HET significantly increased the amount of IFN-γ parallel to a decrease in the level of IL-4 in compared to the control group. Based on our findings, HET has potent immunostimulant characteristics. More investigation into tarragon's potential to be used in the treatment of disorders caused by a weakened immune response should be conducted.

Results of a phase I/IIa trial of SV-BR-1-GM inoculation with low-dose cyclophosphamide and interferon alpha (Bria-IMT) in metastatic breast cancer

ABSTRACT This Phase I/IIa open-label, single-arm clinical trial addressing advanced, refractory, metastatic breast cancer was conducted at six medical centers in the United States. We repeated inoculations with irradiated SV-BR-1-GM, a breast cancer cell line with antigen-presenting activity engineered to release granulocyte-macrophage colony-stimulating factor (GM-CSF), with pre-dose low-dose cyclophosphamide and post-dose local interferon alpha. Twenty-six patients were enrolled; 23 (88.5%) were inoculated, receiving a total of 79 inoculations. There were six Grade 4 and one Grade 5 adverse events noted (judged unrelated to SV-BR-1-GM). Disease control (stable disease [SD]) occurred in 8 of 16 evaluable patients; 4 showed objective regression of metastases, including 1 patient with near-complete regressions in 20 of 20 pulmonary lesions. All patients with regressions had human leukocyte antigen (HLA) matches with SV-BR-1-GM; non-responders were equally divided between matching and nonmatching (p = .01, Chi-squared), and having ≥2 HLA matches with SV-BR-1-GM (n = 6) correlated with clinical benefit. Delayed-type hypersensitivity (DTH) testing to candida antigen and SV-BR-1-GM generated positive responses (≥5 mm) in 11 (42.3%) and 13 (50%) patients, respectively. Quantifying peripheral circulating tumor cells (CTCs) and cancer-associated macrophage-like cells (CAMLs) showed that a drop in CAMLs was significantly correlated with an improvement in progression-free survival (PFS; 4.1 months vs. 1.8 months, p = .0058). Eight of 10 patients significantly upregulated programmed cell death ligand 1 (PD-L1) on CTCs/CAMLs with treatment (p = .0012). These observations support the safety of the Bria-IMT regimen, demonstrate clinical regressions, imply a role for HLA matching, and identify a possible value for monitoring CAMLs in peripheral blood.

The scar-reducing effect of a novel chitosan gel: an in vivo study.

Scar tissue formation, as a normal part of wound healing, initiates in the proliferation phase, continues after the remodelling phase, and may cause an unpleasant appearance or disruption in normal functioning. This study investigated the effects of a topical gel on acute wound healing and reducing scars in a rat model. ChitoScar (ChitoTech Company, Iran), a commercial scar-reducing gel based on chitosan, was analysed for antibacterial and antiviral activity through a quantitative suspension test. Its cytotoxic effect was investigated, and then irritation and delayed-type hypersensitivity tests were carried out on rabbits through direct application of the gel. Furthermore, the effect of the chitosan-based gel on wound healing and scar tissue formation was studied in rats with an acute wound in two groups: the treatment group (topical application of the chitosan-based gel); and the control group (without treatment). Histopathological examination was carried out based on the inflammatory cells, collagen fibre, keratinocytes and fibroblasts. Analysis revealed that the chitosan-based gel had no cytotoxicity and caused no erythema, oedema, local or other systemic adverse response. Wound healing occurred earlier in the treatment group, which was a result of a significant increase in re-epithelialisation, angiogenesis, fibroblast population and collagen fibre thickness (p<0.05). In the treatment group, wounds healed completely after 21 days and scars totally disappeared after 28 days, while in the control group, wound healing remained incomplete with distinct scar tissue. The results demonstrated the positive effect of the chitosan-based gel on the duration and quality of the wound healing process, as well as minimising the scar tissue formation in this in vivo study.

Propolis Silver Nanoparticles as an Adjuvant in Immunization of Rats with Citrobactor Freundii Antigens.

This study aimed to examine the impact of Citrobacter freundii killed whole cell sonicated antigen (KWCSAg) alone and in combination with propolis nanoparticles on humoral immunoglobulin (IgG) and cellular immune responses of rats. The ELISA interleukin 4 (IL4) and IgG, delayed-type hypersensitivity (DTH) skin test, and phagocytosis activity tests were used in this study. In total, 45 rats were divided into five groups of 9 rats. The first group received a 1,000 μg\ml dose of KWCSAg-CF. The second group received an injection of 1,000 μg/ml of KWCSAg-CF antigen and 30 mg/ml of propolis AgNPs. The third group received an injection of 1,000 μg/ml of KWCSAg-CF antigen along with 10 mg/ml of propolis AgNPs. The fourth group was subjected to 30 mg/ml of propolis AgNPs. One ml of phosphate-buffered saline (pH 7.2) was injected into the fifth group (the negative control group). The rats received booster injections of the same antigens after 14 days. Blood was obtained from them to detect immunoglobulin and interleukin 4 (IL-4) on days 21, 28, 32, 46, 50, and 60 following the injection. The second group showed the most significant rise in IL-4 and IgG concentration, followed by the third group, the first group, and the fourth group, compared to the negative control group (fifth group). In all immunized groups, the DTH test results demonstrated an increase in the means of induration with significant differences (P<0.05) of the concentrated antigen after 24 and 48 h, and subsequently a decrease after 72 h, compared to the negative control group. At 48 h after the concentrated antigen was indurated, the second group displayed the most significant increase in diameter.

Oncolytic Newcastle disease virus effects on immune response - a new issue in cancer treatment.

Millions of people are diagnosed with cancer each year, and fighting it puts a heavy financial burden on communities and governments. Numerous advances have been made in the field of cancer; one of the newest methods is using oncolytic viruses. This study aimed to evaluate the effect of oncolytic Newcastle disease virus wild-type strains (NDV-WTS) on the immune system. Forty mice were divided into four groups (10 animals in each group). The control group received phosphate buffered saline, and experimental group 1 (NDV-WTS 1), experimental group 2 (NDV-WTS 2), and experimental group 3 (NDV-WTS 3) received 10-1, 10-2, and 10-3 titers of Newcastle virus on 0, 14th, and 28th days. On the 31st day, 100 µL of Newcastle virus was injected into the left footpads of animals. After 48 hours, delayed-type hypersensitivity (DTH) reactions were measured. On the 33rd day, peritoneal macrophages were isolated. Then proliferation of the cells was measured by the methyl-thiazolyl-tetrazolium (MTT) test. Neutral red uptake and respiratory burst of peritoneal macrophages were also assessed. Data were analyzed using statistical software SPSS, version 19. The results of the DTH test showed that footpad swelling in control, NDV-WTS 1, NDV-WTS 2, and NDV-WTS 3 groups were 23.5%, 23.5%, 23.6% and 23.6%. No significant differences were seen between the groups in this regard (P &gt; 0.05). A negative nitroblue tetrazolium (NBT) reduction test as an indicator of macrophage's respiratory burst, showed no significant difference between the groups (P &gt; 0.05). The neutral red uptake assay and MTT test showed no significant differences between the groups (P &gt; 0.05). The results of this study showed that NDV-WTS in doses of 10-1, 10-2, and 10-3 have no adverse effects on healthy normal cells.

An arginase1- and PD-L1-derived peptide-based vaccine for myeloproliferative neoplasms: A first-in-man clinical trial.

Arginase-1 (ARG1) and Programed death ligand-1 (PD-L1) play a vital role in immunosuppression in myeloproliferative neoplasms (MPNs) and directly inhibit T-cell activation and proliferation. We previously identified spontaneous T-cell responses towards PD-L1 and ARG1 derived peptide epitopes in patients with MPNs. In the present First-in-Man study we tested dual vaccinations of ARG1- derived and PD-L1-derived peptides, combined with Montanide ISA-51 as adjuvant, in patients with Janus Kinase 2 (JAK2) V617F-mutated MPN. Safety and efficacy of vaccination with ARG1- derived and PD-L1-derived peptides with montanide as an adjuvant was tested in 9 patients with MPN The primary end point was safety and toxicity evaluation. The secondary end point was assessment of the immune response to the vaccination epitope (www.clinicaltrials.gov identifier NCT04051307). The study included 9 patients with JAK2-mutant MPN of which 8 received all 24 planned vaccines within a 9-month treatment period. Patients reported only grade 1 and 2 vaccine related adverse events. No alterations in peripheral blood counts were identified, and serial measurements of the JAK2V617F allelic burden showed that none of the patients achieved a molecular response during the treatment period. The vaccines induced strong immune responses against both ARG1 and PD-L1- derived epitopes in the peripheral blood of all patients, and vaccine-specific skin-infiltrating lymphocytes from 5/6 patients could be expanded in vitro after a delayed-type hypersensitivity test. In two patients we also detected both ARG1- and PD-L1-specific T cells in bone marrow samples at the end of trial. Intracellular cytokine staining revealed IFNγ and TNFγ producing CD4+- and CD8+- T cells specific against both vaccine epitopes. Throughout the study, the peripheral CD8/CD4 ratio increased significantly, and the CD8+ TEMRA subpopulation was enlarged. We also identified a significant decrease in PD-L1 mRNA expression in CD14+ myeloid cells in the peripheral blood in all treated patients and a decrease in ARG1 mRNA expression in bone marrow of 6 out of 7 evaluated patients. Overall, the ARG1- and PD-L1-derived vaccines were safe and tolerable and induced strong T-cell responses in all patients. These results warrant further studies of the vaccine in other settings or in combination with additional immune-activating treatments.

Evaluation of the immunotoxicity and allergenicity of a new intranasal influenza vector vaccine against tuberculosis carrying TB10.4 and HspX antigens.

A new vaccine candidate TB/FLU-05E has been developed at the Smorodintsev Research Institute of Influenza (Russia). The vaccine is based on the attenuated influenza strain A/PR8/NS124-TB10.4-2A-HspX that expresses mycobacterial antigens TB10.4 and HspX. This article describes the results of preclinical immunotoxicity and allergenicity studies of the new vector vaccine TB/FLU-05E against tuberculosis. The experiments were conducted on male CBA mice, С57/black/6 mice, and guinea pigs. The vaccine candidate was administered intranasally (7.7 lg TCID50/animal and 8.0 lg TCID50/animal) twice at a 21-day interval. The immunotoxic properties of the vaccine were assessed in mice according to the following parameters: spleen and thymus weight and their organ-to-body weight ratio, splenic and thymic cellularity, hemagglutination titer assay, delayed-type hypersensitivity test, and phagocytic activity of peritoneal macrophages. Histological examination of the thymus and spleen and white blood cell counts were also performed. Allergenicity of the vaccine was assessed in guinea pigs using conjunctival and general anaphylaxis reaction tests. The results showed that double immunization with the TB/FLU-05E vaccine did not affect the phagocytic activity of peritoneal macrophages, cellular and humoral immunity after immunization with a heterologous antigen (sheep red blood cells), or the organ-to-body weight ratio of immunocompetent organs (thymus and spleen). The vaccine candidate demonstrated no allergenic properties. According to the results of this study, the TB/FLU-05E vaccine is well-tolerated by the immune system and demonstrates no immunotoxicity or allergenicity.

Imunomodulatory Effect of Red Tip Leaf’s (Syzygiummyrtifolium Walp.) Ethanol Extract on Male Rat

The pattern of modern life demands that everything must be done very fast and instant. The quality of the consumed food, air pollution, lack of exercise and stress can cause the body’s endurance to decline continuously. The Red Tip (Syzigium myrtifolium Walp.) has been researched to have a flavonoid compound that has benefits as an anti-inflammatory and have potentially as an immunomodulator. The purpose of this study is to prove the effect of immunomodulator on the activity of phagocytosis, the total number and the differential of leukosit, the antibody titer value, and delayed type hypersensitivity response. Group treatment to test the clearance of carbon divided into 7 groups, each consisting of 5 male mice including the group of CMC-Na 0.5 %, Imboost ® dose 22.5 mg/ kgBB, the red tip leaf’s ethanol extract with dose of 50, 100, 200, 400, and 800 mg/kg orraly distributed for 7 days, and on the 8th day the carbon suspension was given using i.v. The blood intake were done in certain times and then the absorbance was measured using UV-Vis spectrophotometer. The liver and spleen of the rats were taken and weighed. The treatment group for antibody titer was divided into 7 groups with each consisting of 5 male mice using the same treatment as above except positive control that has been used levamisole on 25 mg/kgBB dose that has been distributed orraly for 14 days. The non spesific immune respond utilizing carbon cleanses method by measuring the carbon elemination speed and total amount and leukocyte differential. The specific immune response testing using antibody titers was done based on glutination which was formed and the delayed type hypersensitivity testing based on the mice swollen foot. The research outcome shows that the distribution of the red tip leaf’s ethanol extract with dose of 100, 200, 400, and 800 mg/kgBB is increasing the phagocytosis activity significantly compared to CMC-Na 0.5% (p &lt; 0.05). The of the red tip leaf’s ethanol extract is increasing the total of leukocytes and neutrophils of the segment. The distribution of the red tip leaf’s ethanol extract with dose of 50, 100, 200, 400, and 800 mg/kgBB is increasing the formation of rat’s immune cell antibodies significantly compared to CMC-Na 0,5% (p &lt; 0,05).This research prove that the red tip leaf’s ethanol extract has an effect of imunostimulator againts the phagocytosis activity, the total sum and leukocytes differential along with antibody titer value and the delayed type hypersensitivity.&#x0D; &#x0D;

Abstract CT166: Injection site reactions correlate to delayed type hypersensitivity tests and suggest that GP2 reverses immune suppression of trastuzumab-treated HER2 positive patients in a phase IIb study evaluating HER2/neu peptide GP2 (GLSI-100) vs. GM-CSF after adjuvant trastuzumab in HER2 positive women with breast cancer

Abstract The authors did not submit an updated abstract. The original abstract should be considered final. Citation Format: Snehal S. Patel, David B. McWilliams, Mira S. Patel, Jaye Thompson, F. Joseph Daugherty. Injection site reactions correlate to delayed type hypersensitivity tests and suggest that GP2 reverses immune suppression of trastuzumab-treated HER2 positive patients in a phase IIb study evaluating HER2/neu peptide GP2 (GLSI-100) vs. GM-CSF after adjuvant trastuzumab in HER2 positive women with breast cancer [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2022; 2022 Apr 8-13. Philadelphia (PA): AACR; Cancer Res 2022;82(12_Suppl):Abstract nr CT166.

Baseline GP2 immune response as an independent prognostic factor in a phase IIb study evaluating HER2/neu peptide GP2 (GLSI-100) versus. GM-CSF alone after adjuvant trastuzumab in HER2-positive women with breast cancer.

e12519 Background: Delayed type hypersensitivity (DTH) skin tests to GP2 were conducted in the randomized, active-controlled, single-blinded, multicenter Phase IIb trial investigating GLSI-100 (GP2+GM-CSF) administered in the adjuvant setting to node-positive and high-risk node-negative breast cancer patients with tumors expressing HER2. Previously it has been reported that subjects with baseline response to GP2 by DTH were more likely to experience a recurrence sooner than those without a positive baseline response. It was of interest to determine if this response was independent of other known prognostic factors. Methods: Patients were randomized and received GLSI-100 (500 mcg GP2: 125 mcg GM-CSF) or control (GM-CSF only) via 6 intradermal injections every 3-4 weeks as part of the Primary Immunization Series (PIS) for 6 months and 4 booster injections every 6 months thereafter. GP2 DTH skin tests were placed at baseline and after the 6th dose. After 48-72 hours, the largest perpendicular diameters of induration were measured and the orthogonal mean was calculated. Induration of over 5mm was considered a positive response. The relationship between prognostic factors such as demographic and disease characteristics and the baseline DTH response were investigated by logistic regression. Results: The study enrolled 180 patients who were HER2 3+ positive and low HER2 expressors (1-2+). After 5 years of follow-up, the Kaplan-Meier estimated 5-year DFS rate in the 46 HER2 3+ patients treated with GLSI-100, if completing the PIS, was 100% versus 89.4% (95% CI:76.2, 95.5%) in the 50 patients treated with GM-CSF (p = 0.0338). Just under 23% of subjects experienced an immune response to GP2 by DTH at baseline. The incidence was similar in patients who were HER2/neu positive and those considered low expressors and was equally distributed across GLSI-100 and placebo-treated subjects. Known prognostic factors such as hormone receptor status, stage at presentation, node status, etc. were evenly distributed across subjects that experienced a baseline DTH response to GP2 and those who did not. Each prognostic factor was investigated in a logistic model to determine if it predicted baseline DTH immune status. The prognostic factors did not predict baseline DTH immune response in the logistic models. There appears to be no correlation between baseline immune response to GP2 and known prognostic factors. Conclusions: Baseline GP2 immune response as measured by the delayed-type hypersensitivity test may be an independent prognostic factor for recurrence. Knowledge of this GP2 immune response may identify a patient with increased risk of rapid recurrence. Clinical trial information: 00524277.

Easy approach to detect cell immunity to COVID vaccines in common variable immunodeficiency patients.

Patients with primary antibody deficiencies, such as Common Variable Immunodeficiency (CVID), have some problems to assess immune response after coronavirus disease (COVID) vaccination. Cutaneous delayed-type hypersensitivity (DTH) has the potential to be used as a useful, simple, and cheaper tool to assess T-cell (T lymphocyte) function. Seventeen patients with CVID, a rare disease, received two doses of the mRNA-based Pfizer-BioNTech COVID-19 vaccine. Humoral Immune Response (HIR) was determined by measuring specific immunoglobulin G (IgG) antibodies, and Cellular Immune Response (CIR) was evaluated using an ex vivo interferon-gamma release assay (IGRA) and in vivo by DTH skin test. Two weeks after the second dose of the vaccine, 12 out of 17 CVID patients have high optical density (OD) ratios of specific anti-spike protein (S) IgG whereas five patients were negative or low. Ex vivo CIR was considered positive in 14 out of 17 S1-stimulated patients. Unspecific stimulation was positive in all 17 patients showing no T-cell defect. A positive DTH skin test was observed in 16 CVID patients. The only patient with negative DTH also had negative ex vivo CIR. The use of DTH to evaluate CIR was validated with an optimal correlation with the ex vivo CIR. The CIR after vaccination in patients with antibody deficiencies seems to have high precision and more sensitivity to antibodies-based methods in CVID. There is a remarkable correlation between cutaneous DTH and ex vivo IGRA after COVID vaccination. A COVID-specific skin DTH test could be implemented in large populations. Cutaneous delayed-type hypersensitivity has the potential to be used as a useful, simple, and cheaper tool to assess T-cell functioning.

IMMUNOSTIMULANT POTENTIAL OF OYSTER MUSHROOM NUGGET (Pleourotus ostreatus)

Oyster mushrooms are widely known as having high beta glucan content. Beta glucan is a water insoluble fiber that indicates an immunomodulator with increasing the activity of NK cells in the bloodstream which provides immune effect. Oyster mushroom nuggets are processed oyster mushrooms that can be proven containing the highest beta glucans. This study aims to observe the immunostimulant potential of oyster mushroom nuggets in BALB/c mice by analyzing its white blood cell profile, spleen and liver histopathology. The research method is conducted through orbital sinus of the rat's eye using a microhematocrit to count total leukocytes, lymphocytes, and granulocytes with the Delayed Type Hypersensitivity test. The 18mg/kg body weight oyster mushroom nugget can increase lymphocytes cells 7.2 ± 1.22 103/µl. It is significantly different from control group. Immunostimulatory activity also was proven with the widening of the white pulp of the spleen with the growth of young lymphocytes after being given oyster mushroom nuggets of 18 mg/kg body weight.

Safety profile of intravenous administration of live Pichia pastoris cells in mice.

Pichia pastoris has been widely used to produce antigenic proteins aimed to integrate subunit vaccines. Moreover, increasing interest in large-scale vaccine production at the lowest cost is rapidly focusing in the development of yeast surface display (YSD) systems for delivery of antigens. In this scenario, the safety of live yeast administration must be warranted, however, such information is very scarce. Here, we assess the intravenous administration (i.v.) of live P. pastoris cells in order to trace dissemination in BALB/c mice and to evaluate the immune response raised against the yeast compared to the well-defined pathogen Candida albicans. Our results demonstrate dissemination of P. pastoris to the heart, kidney, and spleen, but it is quickly eliminated during the first 48 h postinfection (hpi), with persistence in the liver along with mild mononuclear (MN) and polymorphonuclear (PMN) infiltrate, which was resolved at 144 hpi. In vivo delayed-type hypersensitivity test (DTH) or in vitro antigenic stimulation of mice splenocytes demonstrate that transient infection of P. pastoris did not induce a cell-mediated immune response nor increase the level of circulating IgG or IgM. These results demonstrate the innocuous profile of P. pastoris and support its use as a safe delivery system for vaccine development.

Calcium phosphate adjuvanted nanoparticles of outer membrane proteins of Salmonella Typhi as a candidate for vaccine development against Typhoid fever.

Introduction. The conventional adjuvants used in vaccines have limitations like induction of an imbalanced Th1 and Th2 immune response. To overcome this limitation, novel adjuvants and newer forms of existing adjuvants like calcium phosphate nanoparticles are being tested.Hypothesis/Gap Statement. Calcium phosphate adjuvanted outer membrane proteins vaccine may work as an efficient, safe and cost effective vaccine against Salmonella Typhi.Aim. Our goals were to evaluate the potential of calcium phosphate nanoparticles as an adjuvant using outer membrane proteins (Omps) of Salmonella Typhi as antigens for immune response, with montanide (commercially available adjuvant) as control, and its toxicity in rats.Methodology. Calcium phosphate adjuvanted outer membrane proteins nanoparticles were synthesized and characterized. The efficacy of vaccine formulation in mice and toxicity assay were carried out in rats.Results. The calcium phosphate nanoparticles varying in size between 20-50 nm had entrapment efficiency of 41.5% and loading capacity of 54%. The calcium phosphate nanoparticle-Omps vaccine formulation (nanoparticle-Omps) induced a strong humoral immune response, which was significantly higher than the control group for the entire period of study. In the montanide-Omps group the initial very high immune response declined steeply and then remained steady. The immune response induced by nanoparticle-Omps did not change appreciably. The cell mediated immune response as measured by lymphocyte proliferation assay and delayed type hypersensitivity test showed a higher response (P<0.01) for the nanoparticles-Omps group as compared to montanide-Omps group. The bacterial clearance assay also showed higher clearance in the nanoparticles-Omps group as compared to montanide-Omps group (approx 1.4%). The toxicity analysis in rats showed no difference in the values of toxicity biomarkers and blood chemistry parameters, revealing vaccine formulation was non-toxic in rats.Conclusion. Calcium phosphate nanoparticles as adjuvant in vaccines is safe, have good encapsulation and loading capacity and induce a strong cell mediated, humoral and protective immune response.

Исследование раздражающего и аллергизирующего действия кормовой добавки на основе солей гуминовых кислот

The article presents the results of studies on the assessment of the locally irritating and allergenic effect of a feed additive based on humic acid salts obtained by extraction of brown coal, enriched with macro- and microelements. The study of the local irritant effect was carried out on guinea pigs by skin application and rabbits by the conjunctival test method. It was found that the feed additive does not cause irritation when instilled in its native form on the conjunctiva of the eye of rabbits and when applied once to the skin of guinea pigs. Studying the allergenic properties of the feed additive in mice, a delayed-type hypersensitivity test was used. When taking into account the reaction index when comparing the indicators of the experimental and control groups of mice, the reaction index of the mice of the experimental group was slightly higher than that of the control group, but the differences are unreliable, which allows us to conclude that there are no pronounced sensitizing properties of the feed additive.

A phase II study of chemotherapy in combination with telomerase peptide vaccine (GV1001) as second-line treatment in patients with metastatic colorectal cancer.

Background: GV1001 is a human telomerase peptide vaccine that induces a CD4/CD8 T-cell response against cancer cells, thereby affording an immunological anti-tumor effect. Here, we evaluated the efficacy and safety of GV1001 in combination with chemotherapy in patients with metastatic colorectal cancer who had failed first-line chemotherapy.Methods: This multicenter, non-randomized, single-arm phase II study recruited recurrent or metastatic colorectal cancer patients with measurable disease who had failed first-line chemotherapy. Patients received GV1001 and chemotherapy concomitantly based on a pre-established schedule. Cytotoxic chemotherapy and targeted agents (bevacizumab, cetuximab, or aflibercept) were allowed to be used at the discretion of the investigator. The primary endpoint was the disease control rate; secondary endpoints were the objective response rate, progression-free survival, overall survival, and safety outcomes. The baseline serum eotaxin level (a potential predictive biomarker of GV1001) was analyzed. To determine whether an adequate immune response had been induced, a delayed-type hypersensitivity test and a T-cell proliferation test were performed.Results: From May 13, 2015 to October 13, 2020, 56 patients with recurrent or metastatic colorectal cancer treated in seven hospitals of South Korea were enrolled. The median patient age was 64 years (range, 29-82 years); 67.9% were men. Of all patients, 66.1% had left-side colorectal cancer and the RAS mutation was present in 25%. The disease control rate and the objective response rates were 90.9% (95% confidence interval [CI]: 82.4-99.4%) and 34.1% (95% CI, 20.1-48.1%), respectively. The median progression-free survival was 7.1 months (95% CI, 5.2-9.1 months) and the median overall survival was 12.8 months (95% CI, 9.9-15.8 months). The most common all-grade adverse events were neutropenia (48.2%), nausea (26.8%), neuropathy (25.0%), stomatitis (21.4%), and diarrhea (21.4%). Immune response analysis showed that no patient had positive delayed-type hypersensitivity test results; antigen-specific T-cell proliferation was observed in only 28% of patients. The baseline eotaxin level was not associated with any efficacy outcome.Conclusion: Although no clear GV1001-specific immune response was observed, the addition of GV1001 vaccination to chemotherapy was tolerable and associated with modest efficacy outcomes.

Clinical and immunological study of specific Mycobacterium bovis-BCG transfer factor in lambs

This study was designed to prepare and evaluate the specific Mycobacterium bovis (M. bovis) Bacillus Calmette-Guerin (BCG) transfer factor (TF) from spleen cells of two lambs vaccinated subcutaneously S/C with BCG vaccine twice 2 weeks’ intervals, similarly one lamb injected with normal saline used as a donor of non-specific TF. The lambs were clinically examined at zero time and along 3 days post each immunization, then, delayed type hypersensitivity (DTH) was checked post 4 weeks. TF was obtained from spleen cells of the lambs, inspected for sterility; safety.TF was evaluated in 4 groups (five in each) of lambs 3-4 months old. The first group (recipient) was injected S/C with 2 ml of specific BCG-TF, similarly the 2nd, 3rd and 4th groups were injected with non-specific TF, BCG and normal saline respectively. The clinical signs and DTH test were checked as described above, only the specific TF recipient and BCG vaccinated lambs gave a positive DTH test. The serum concentration of inerlukin-17 (IL-17), interferon-gamma (IFN-γ) and migration inhibitory factor (MIF) cytokines were measured by Enzyme-linked immunosorbent assay (ELISA), which revealed a highly significant increase in the TF recipient lambs in comparison with the control and vaccinated groups. It concluded that the prepared specific M. bovis-BCG transfer factor was effective to induce cell mediated immunity via DTH and increasing the serum concentration of cytokines in recipient lambs.

Immunity response of mice infected with laser irradiated protoscoleces of Echinococcus granulosus

The present study investigated the effect of the Neodymium- Doped Yttrium Aluminum Garnet (Nd: Y3AL5G12) Laser eradiation 1000 mw, against infection with secondary hydatid disease in BALB/c mice by inoculating the animals with protoscoleces of Echinococcus granulosus exposed to laser irradiation, for various periods 40,60,120 and 240 minutes, with viability of 70%,57%,50% and 40%, respectively, contrasted to the control set (mice inoculated with unexposed protoscoleces) along three months, depending on the evaluation of the total count of WBCs and assessment of acquired cell- mediated immunity, demonstrated by delayed type hypersensitivity test (DTH). The results showed considerable increase (p<0.01) in the total WBCs counting, in mice injected with irradiated protoscoleces, up to 13962 cell/cm3, in comparison to the control group 5420 cell/cm3, after 120 minutes, two months post infection. Moreover, there was considerable excess (p<0.01) in the foot pad thickness in treated mice, 2.94 mm after 120 minute (24hour post antigen injection), two months post infection, 2.86mm after 120 minutes (24 hours post antigen injection), 1 month post infection and 2.64 mm, 120 minutes (24 hours post antigen injection), three months post infection, compared with the control group 1.18mm, 1.80mm and 1.72mm, respectively. Results revealed that irradiation of Echinococcus granulosus protoscoleces with laser had a significant influence on the production of WBCs and stimulation of delayed-type hypersensitivity reaction in mice.