Novel report of the phytochemical composition from Fraxinus excelsior pollen grains

In this research, we investigated the phytochemical profiles for two models of aqueous (Aq) and methanolic (Me) pollen extracts of F. excelsior from three pollination periods from hermaphrodite flowers (H) of polygamous and male flowers of pure male (M) in order to identify their constituent compounds. Pollens of both phenotypes H and M were collected during three pollination periods and were analyzed by light and scanning electron microscopy (LM/SEM). The total phenolic content (TPC) and flavonoids content (TFC) was measured using Folin-Ciocalteu and aluminum chloride (AlCl3) methods, respectively. Antioxidant activities were evaluated using ferric reducing antioxidant power (FRAP) and scavenging free radical DPPH• and ABTS•+. GC-FID and GC/MS were used to identify the chemical composition of essential oils. There was a significant difference (P < 0.001) between the means of TPC for M and TFC for the H. Comparison of H and M antioxidant activities showed that DPPH (IC50) to be (2.977 ± 0.117 μM) during the second pollination period of M and (4.877 ± 0.021 μM) for first period of H. The majority of the compounds identified were linalool (35.42%) from the monoterpenoides in H and Delta-cadinene (43.22%) belonging to the sesquiterpenes in M. We concluded that there is a significant difference between the H and M compounds in pollen at different periods

Allergen‐specific immunotherapy by recombinant Der P1 allergen‐derived peptide‐based vaccine in an allergic mouse model

Abstract Aim Increased IgE levels have made house dust mite allergens one of the most frequent causes of allergies worldwide. Treatment reduces the IgE antibodies and types two cytokines, namely interleukin‐4 (IL‐4) and IL‐13. Although existing treatments significantly reduce IgE or IL‐4/IL‐13, they are very costly. This study aimed to construct a recombinant protein derived from rDer p1 peptides in the form of an immunotherapy approach and to measure the response of IgE and IgG antibodies. Methods The proteins were isolated, purified, and evaluated using the SDS‐PAGE and Bradford test and confirmed by using Western blot. To evaluate immunotherapy efficiency, 24 BALB/C mice were sensitized intraperitoneally with house dust mites (HDM) adsorbed to Aluminum hydroxide (Alum) and randomly divided into four groups of six: control sensitized, HDM extract, rDer p1, and DpTTDp vaccine. To immunization, four groups of random mice were each treated with phosphate‐buffered saline, 100 μg of rDer p1 protein, DpTTDp, or HDM extract, every 3 days. Direct ELISA determined HDM‐specific IgG and IgE subclasses. Data were analyzed in SPSS and Graph pad prism software. Values of p < .05 were considered significant. Results After immunization of mice, the rDer P1 and recombinant vaccine like HDM extract increased IgG antibody titer and decreased IgE‐dependent reactivity in allergic mice to rDer P1. Also, the levels of inflammatory IL‐4 and IL‐13 cytokines as allergic stimulants decreased. Conclusion The use of present available recombinant proteins is considered a viable, cost‐effective, and long‐term option for providing effective HDM allergy immunotherapy vaccines without side effects

Exhaustion of T lymphocytes in the tumor microenvironment : Significance and effective mechanisms

T lymphocytes play crucial roles in adaptive immune responses to tumors. However, due to different tolerance mechanisms and inhibitory effects of the tumor microenvironment (TME) on T cells, responses to tumors are insufficient. In fact, cellular and molecular suppressive mechanisms repress T cell responses in the TME, resulting in senescent, anergic and exhausted lymphocytes. Exhaustion is a poor responsive status of T cells, with up-regulated expression of inhibitory receptors, decreased production of effective cytokines, and reduced cytotoxic activity. Low immunogenicity of tumor antigens and inadequate presentation of tumor-specific antigens results in inappropriate activation of naive T lymphocytes against tumor antigens. Moreover, when effector cytotoxic T cells enter TME, they encounter a complicated network of cells and cytokines that suppress their effectiveness and turn them into exhausted T cells. Thus, the mechanism of T cell exhaustion in cancer is different from that in chronic infections. In this review we will discuss the main components such as inhibitory receptors, inflammatory cells, stromal cells, cytokine milieu as well as environmental and metabolic conditions in TME which play role in development of exhaustion. Furthermore, recent therapeutic methods available to overcome exhaustion will be discussed

Decorating and loading ghosts with allergens for allergen immunotherapy

More than 25% of the global population has IgE mediated allergic diseases. Allergen immunotherapy (AIT) is the only available form of treatment that alters the underlying mechanism of IgE-mediated allergic diseases. AIT is aimed at desensitizing allergic individuals by repeatedly administering disease-causing allergens over a long period of time. Despite its proven efficacy in numerous clinical trials, the effectiveness of AIT still suffers some drawbacks due to the quality of allergens used and in particular the unavailability of efficient allergen delivery systems. Several studies have demonstrated that bacterial ghosts (BG) systems can be used to display and deliver antigens to their targets for the management of diseases. However, there is no report documenting the use of BG systems for immunotherapy of IgE-mediated diseases so far. Thus, in this review, we intend to discuss the potentialities of BG systems for displaying and delivering allergens for future management of IgE-mediated diseases

The Induction of Metformin Inhibitory Effects on Tumor Cell Growth in Hypoxic Condition

It is aimed to evaluate the actual anti-cancerous effects of metformin on cancer cells in hypoxic condition. Non-cancerous cells (HEK293) and cancer cells (MCF-7) were cultured in both hypoxia and normoxia conditions and treated with different concentrations of metformin. The proliferation, apoptosis, and necrosis rate were assessed using MTT test and Annexin V assay. The S6K1 phosphorylation was assessed using western blotting. Zymography was used to measure the activity of metalloproteinase-9 (MMP-9). Metformin treatment inhibited proliferation of cancer cells in the optimal concentration of 10 mM under hypoxia condition, while it showed no effects on non-cancerous cell viability. The statistical analysis of MTT assay indicated that the pro-apoptotic function of metformin for cancer cells under hypoxia condition compared to normoxia was significant with different metformin concentrations (p<0.01). However, the effect of metformin treatments for non-cancerous cells under hypoxia condition compared to normoxia was not significant. Western-blot analysis indicated a significant decrease in S6K1 phosphorylation in cancer cells under hypoxia condition (p<0.05). Nevertheless, there was no considerable difference between normoxia and hypoxia conditions in non-cancerous cells. MMP-9 zymography analysis revealed that the highest inhibition of MMP-9 activity was observed in hypoxia condition by 20mM of metformin concentration only in cancer cell. The results indicate that in hypoxia condition metformin exerts its anti-cancerous function by inhibiting proliferation and tumor progression and inducing cell apoptosis more effectively than normoxia condition. In line with cancer cell conditions, most importantly hypoxic condition, metformin can be considered as a potential anti-cancerous drug

Expression of recombinant parvalbumin from wolf-herring fish and determination of its IgE-binding capability

In this study, we produced the recombinant form of parvalbumin from wolf-herring fish and determined its IgE reactivity. Parvalbumin cDNA was sub-cloned into pET28 and expressed in Escherichia coli BL-21. The immunoreactivities of the recombinant and native parvalbumins were compared, and the effect of calcium binding was determined by sera from 25 fish-allergic patients. ELISA and Western blotting confirmed similar IgE-reactivities of the recombinant and native proteins and confirmed that this phenomenon is highly dependent on calcium binding. The recombinant protein was 94.5% similar to carp parvalbumin (Cyp c1). Approximately 72% of patients reacted strongly with recombinant parvalbumin, 80% of them reacted with the native form and only 56% showed IgE reactivity with crude extract. Because the IgE-binding capacity of recombinant wolf-herring parvalbumin is retained and is highly similar to Cyp c1, the wild and hypoallergenic forms of this allergen could be used for diagnosis and immunotherapy of fish allergy, respectively

Construction of a recombinant B-cell epitope vaccine based on a der p1-derived hypoallergen: A bioinformatics approach

Aim: House dust mite (HDM) allergens are important elicitors of IgE-mediated allergies. This study was aimed at constructing and characterizing a recombinant fusion protein, DpTTDp, which was based on carrier-bound Der p 1-derived peptides for HDM allergen immunotherapy. Methods: Using the Immune Epitope Database (IEDB), we identified from Der p 1, a 34-mer hypoallergenic peptide. Two copies of the hypoallergen were then fused to a partial fragment of a tetanus toxoid molecule's N-and C terminus and expressed in Escherichia coli. After purification to homogeneity, the protein was evaluated for allergenicity and its ability to induce blocking antibodies upon immunization. Results: Upon immunization of mice, DpTTDp induced high levels of protective IgG-antibodies that blocked allergic patients' IgE reactivity to HDM. In addition, DpTTDp lacked relevant IgE-reactivity, induced low T-cell proliferation and IFN-γ in peripheral blood mononuclear cells of HDM-allergic patients' sera. Conclusion: The protein represents a promising HDM-allergy immunotherapy candidate vaccine. © 2018 Future Medicine Ltd