Effects of arbutin on glucose uptake by glucose transporter 4 (GLUT4) and its cytoprotective properties in L6 skeletal muscle cell line.

It has been well known that oxidative stress and increased intracellular reactive oxygen species (ROS) have a pivotal role in disrupting the insulin signaling pathways leading to cellular insulin resistance. In this study, we evaluated arbutin's effects on glucose uptake by GLUT4 and cytoprotective properties in the L6 skeletal muscle cell line. The effect of arbutin and tertiary butyl hydrogen peroxide (t-BHP) on glucose uptake in cultured L6 cells was investigated by flow cytometry. We also evaluated gene expression levels of GLUT1 and GLUT4 in the L6 cells by quantitative real-time polymerase chain reaction analysis. The results from the study demonstrated that the optimum ROS generation occurred 3 h after 100 µM t-BHP treatment and pretreatment with arbutin (500 and 1000 µM) significantly inhibited the t-BHP induced ROS generation (p < .05). Our result indicated that 3 h pretreatment of L6 cells with 1000 µM of arbutin before 50 µM t-BHP significantly increased glucose uptake than the 50 µM t-BHP alone group (p < .05). Our findings may suggest that an increase in the uptake of 2-NBDG by L6 cells with arbutin pretreatment can be associated with increased expression of GLUT4 and GLUT1 under oxidative stress.

A Large Retrospective Study of Epidemiological Characteristics of COVID-19 Patients in the North of Iran: Association between SARS-CoV-2 RT-PCR Ct Values with Demographic Data.

Objectives: To avoid worsening from mild, moderate, and severe diseases and to reduce mortality, it is necessary to identify the subpopulation that is more vulnerable to the development of COVID-19 unfavorable consequences. This study aims to investigate the demographic information, prevalence rates of common comorbidities among negative and positive real-time reverse-transcriptase polymerase chain reaction (rRT-PCR) patients, and the association between SARS-CoV-2 cycle threshold (Ct) at hospital admission, demographic data, and outcomes of the patients in a large population in Northern Iran. Methods: This large retrospective cross-sectional study was performed from 7 March to 20 December 2020. Demographic data, including gender, age, underlying diseases, clinical outcomes, and Ct values, were obtained from 8,318 cases suspected of COVID-19, who were admitted to four teaching hospitals affiliated to Babol University of Medical Sciences (MUBABOL), in the north of Iran. Results: Since 7 March 2020, the data were collected from 8,318 cases suspected of COVID-19 (48.5% female and 51.5% male) with a mean age of 53 ± 25.3 years. Among 8,318 suspected COVID-19 patients, 3,250 (39.1%) had a positive rRT-PCR result; 1,632 (50.2%) patients were male and 335 (10.3%) patients died during their hospital stay. The distribution of positive rRT-PCR revealed that most patients (464 (75.7%)) had a Ct between 21 and 30 (Group B). Conclusion: Elderly patients, lower Ct, patients having at least one comorbidity, and male cases were significantly associated with increased risk for COVID-19-related mortality. Moreover, mortality was significantly higher in patients with diabetes, kidney disease, and respiratory disease.

Pathophysiological roles of chronic low-grade inflammation mediators in polycystic ovary syndrome.

Polycystic ovary syndrome (PCOS) is the most common hormonal imbalance disease in reproductive-aged women. Its basic characteristics are ovulatory dysfunction and ovarian overproduction of androgens that lead to severe symptoms such as insulin resistance, hirsutism, infertility, and acne. Notwithstanding the disease burden, its underlying mechanisms remain unknown, and no causal therapeutic exists. In recent years, further studies showed that inflammation processes are involved in ovulation and play a key role in ovarian follicular dynamics. Visceral adipose tissue can cause inflammatory response and maintenance of the inflammation state in adipocytes by augmented production of inflammatory cytokines, monocyte chemoattractant proteins, and recruitment of the immune cell. Therefore, the PCOS can be related to a low-grade inflammation state and inflammatory markers. Investigating the inflammatory processes and mediators that contribute to the commencement and development of PCOS can be a critical step for better understanding the pathophysiology of the disease and its treatment through inhibition or control of related pathways. In the present review, we discuss the pathophysiological roles of chronic low-grade inflammation mediators including inflammasome-related cytokines, interleukin-1ß (IL-1ß), and IL-18 in PCOS development.

Indomethacin attenuates neuroinflammation and memory impairment in an STZ-induced model of Alzheimer's like disease.

Objective: Non-steroidal anti-inflammatory drugs (NSAIDs) exposure might be considerably associated with a decreased risk of Alzheimer's disease (AD). Therefore, we conducted an experiment to investigate the impact of indomethacin (IND) on inflammasome as a key player of neuroinflammation.Methods: The Alzheimer's-like condition was induced by streptozotocin (STZ) in rats. IND was injected intraperitoneally 1 d prior to STZ administration and resumed with 2 d interval up to 60 d. Morris water maze (MWM) was utilized to assess learning and memory. The expression level of genes that contribute to the inflammasome pathway was measured using real-time polymerase chain reaction (PCR). To authenticate the obtained outcomes, immunostaining for caspase-1, interleukin-1ß (IL-1ß), and phosphorylated tau (p-Tau) protein was conducted.Results: Behavioral experiments indicated that IND treatment was able to improve learning and memory performance (p<.05). A significant decrease in C-terminal caspase recruitment domain [CARD] domain-containing protein 4 (NLRC4), nucleotide-binding oligomerization domain [NOD]-like receptor protein 3 (NLRP3), IL-1ß, and apoptosis-associated speck-like protein containing CARD (ASC) mRNA expression was recorded in IND administered group compared with the STZ group (p<.05). Furthermore, expression levels of IL-18 and caspase-1 in the hippocampus of IND-treated group tended to decrease. Immunostaining evaluations showed that few positive cells for caspase-1, IL-1ß, and p-Tau protein in IND treated animals, whereas the number of positive cells was considerably increased in STZ treated animals (p<.05).Conclusion: It could be deduced that IND improves neuroinflammation and memory impairment in AD through decreasing IL-1ß and caspase-1 that are associated with suppression of NLRC4 and NLRP3 inflammasome genes. This holds the potential to introduce valuable targets in the field for successful combat against AD.

Contribution of inflammasome complex in inflammatory-related eye disorders and its implications for anti-inflammasome therapy.

Inflammasome complex is regarded as a major molecular regulator that exerts a significant function in caspase-1 activation and consequently, the development of cytokines like interleukin-1ß (IL-1ß) and interleukin-18 (IL-18). The secretion of these cytokines may induce inflammation. The role of inflammasomes in the pathologic process of eye-related illnesses like glaucoma, age-related macular degeneration (AMD), and diabetic retinopathy has been well studied over the past decade. However, the detailed pathogenic mechanism of inflammasomes in these retinal diseases is still unknown. Therefore, further investigation and understanding various aspects of inflammasome complexes as well as their pivotal roles in the immunopathology of human ocular illnesses are essential. The present review aims to describe the significant involvement of inflammasomes in the immunopathology of important inflammatory retinal illnesses, including glaucoma, age-related macular degeneration (AMD), and diabetic retinopathy focusing on anti-inflammasome therapy as a promising approach in the treatment of inflammation-related eye diseases.

Diagnostic methods for Helicobacter pylori infection: ideals, options, and limitations.

Helicobacter pylori (H. pylori) resides in the stomach, colonizes gastric epithelium, and causes several digestive system diseases. Several diagnostic methods utilizing invasive or non-invasive techniques with varying levels of sensitivity and specificity are developed to detect H. pylori infection. Selection of one or more diagnostic tests will depend on the clinical conditions, the experience of the clinician, cost, sensitivity, and specificity. Invasive methods require endoscopy with biopsies of gastric tissues for the histology, culture, and rapid urease test. Among non-invasive tests, urea breath test and fecal antigen tests are a quick diagnostic procedure with comparable accuracy to biopsy-based techniques and are methods of choice in the test and treatment setting. Other techniques such as serological methods to detect immunoglobulin G antibodies to H. pylori can show high accuracy as other non-invasive and invasive biopsies, but do not differentiate between current or past H. pylori infections. Polymerase chain reaction (PCR) is an emerging option that can be categorized as invasive and non-invasive tests. PCR method is beneficial to detect H. pylori from gastric biopsies without the need for the cultures. There is no other chronic gastrointestinal infection such as H. pylori with a set of comparable diagnostic methodologies. Despite the availability of multiple diagnostic methods, it remains unclear on the choice of any one method as the gold standard for detecting H. pylori infection, especially in epidemiological studies. In this work, we review the principal diagnostic methods used to detect H. pylori infection and their advantages and disadvantages, and applications in clinical practice.

Iranian propolis efficiently inhibits growth of oral streptococci and cancer cell lines.

BACKGROUND: Propolis is a natural bee product with a wide range of biological activities that are related to its chemical composition. The present study investigated the quantification of quercetin (Q) in Ardabil ethanol extract of propolis (AEEP), and then compared its anti-bacterial, anti- biofilm and cytotoxic effects on cancer and normal cell lines. METHOD: In the present study, the chemical composition of AEEP was determined through the high-performance liquid chromatography (HPLC). The AEEP and its main component, quercetin (Q), were evaluated in vitro against 57 oral streptococci by a broth micro-dilution method. The biofilm formation was assessed through the crystal violet staining and MTT assays. The impact of AEEP and Q anti-proliferative effect were evaluated on the fibroblast as normal and cancer cell lines (KB and A431). RESULTS: The Q concentration in the composition of AEEP was 6.9% of all its components. The findings indicated that the AEEP and Q were efficient against the cariogenic bacteria and were able to inhibit the S.mutans biofilm adherence at a sub-MIC concentration. Moreover, electron micrographs indicated the inhibition of biofilms compared to control biofilms. In addition, the AEEP and Q indicated a dose-dependent cytotoxic effect on A431 and KB cell lines. On the contrary, they had no cytotoxic effect on fibroblast cells. CONCLUSION: The results indicated that the synergistic impact of main components of AEEP was related to the inhibition of the cancer cell proliferation, cariogenic bacteria and oral biofilm formation. It may play a promising role in the complementary medicine and, it is suggested to be used as food additives.

Association of TNF-α but not IL-1ß levels with the presence of Helicobacter pylori infection increased the risk of peptic ulcer development.

Peptic ulcer is a lesion in the mucosa of the digestive tract affecting many people all around the world. Recent investigations have indicated that produced inflammatory cytokines such as TNF-α and IL-1ß in response to gastric infection by Helicobacter pylori play an important role in the development of peptic ulcer. With regard to the significance of these cytokines in peptic ulcer development and the high prevalence of this disease in the developing countries, this study aimed to investigate the association of TNF-α and IL-1ß with peptic ulcer in the presence of H. pylori. This case-control study enrolled 61 patients with peptic ulcer disease (PUD) as cases and 59 people without peptic ulcer (NPUD) as controls. Blood samples and endoscopic biopsies were collected. H. pylori infection was confirmed by using rapid urease test (RUT), specific IgG measurement and histopathological examination. Then, IL-1ß and TNF-α levels were evaluated using enzyme linked immunosorbent assay (ELISA). The seropositivity of H. pylori was 62.5% in the studied population, while by considering RUT and histopathological examination along with specific-IgG antibody, H. pylori infection decreased to 56.7%. In addition, H. pylori infection was significantly (OR = 0.37; 95% CI = 0.17-0.82; P = .02) associated with peptic ulcer development. The TNF-α level in PUD and infected H. pylori subjects was significantly higher than that of control and un-infected H. pylori individuals. However, no significant difference of IL1ß level was observed between PUD and control groups as well as between H. pylori infected and un-infected individuals. Interestingly, IL-1ß level in PUD patients without H. pylori infection was significantly higher than infected ones. Moreover, a significant correlation between specific-IgG antibody with TNF-α level was observed. Taken together, our results showed that increased level of TNF-α could probably play pivotal role in pathogenesis of peptic ulcer in the presence of H. pylori infection. These findings also highlighted the importance of IL-1ß in the absence of H. pylori infection in peptic ulcer development.

Influence of Helicobacter pylori virulence factors CagA and VacA on pathogenesis of gastrointestinal disorders.

Helicobacter Pylori (H. pylori) is a gram-negative bacteria infecting numerous people all over the world. It has been established that H. pylori play an important role in pathogenesis of gastritis, peptic ulcer and gastric cancer. Pathogenic features of this bacterium are mainly attributes to the existence of pathogenic islands (PAI) genes. The most known genes in these islands are cytotoxin-associated gene A (CagA) and vacuolating cytotoxin gene (VacA). Most studies demonstrated various frequency of CagA and VacA in patient with peptic ulcer or gastritis in different countries. This variation in CagA and VacA frequency may be due to the capability of this bacterium to be genetically versatile and can alter the expression of these genes with geographic diversity. Although H. pylori infection is not usually associated with any clinical symptoms, but sometimes leads to inflammation in gastrointestinal system and resulted in peptic ulcer and gastric cancer. In this regard, this review will illustrate the importance of Helicobacter pylori in pathogenesis of gastrointestinal disorders with focusing on CagA and VacA virulence factors.

Zika virus: epidemiology, clinical aspects, diagnosis, and control of infection.

Zika virus (ZIKV) is an emerging pathogen of huge public health significance to human beings. Although majority of infections are benign with self-limiting symptoms, the recent outbreak has established an association with the increased incidence of some congenital anomalies such as microcephaly. In other words, due to the large extent of the virus and mosquito vectors, the infection has become a thoughtful health problem for human societies, though now, there are no antiviral therapies or vaccines against this virus. In spite of extensive research carried out by scientists, not so much information has been gathered about this viral infection. In the current review, we prepared an overview of the remarkable progress made in understanding about the epidemiology, immunology, clinical presentation, and diagnosis methods of ZIKV infection.

Helicobacter pylori cagL amino acid polymorphism D58E59 pave the way toward peptic ulcer disease while N58E59 is associated with gastric cancer in north of Iran.

The cagL protein of Helicobacter pylori involving in pathogenesis of gastroduodenal disorders. Therefore, the current study was conducted to determine the cagL amino acid polymorphisms in patients with gastric diseases. One hundred gastric biopsies were collected from gastritis, peptic ulcer (PUD) and gastric cancer (GC) patients and were screened for cagL using polymerase chain reaction (PCR). Also, sequence variations of the cagL were assessed via sequence translation. The cagL geneopositivity was 71.6% in patients were infected with H. pylori. The cagL from PUD indicated a higher rate of D58 amino acid sequence polymorphism than those of the GC and gastritis (P < 0.05). The D58 polymorphism showed an increased risk of PUD up to 6.5-fold (95% CI: 1.2-35.7). This position was occupied with amino acid N58 in GC. The E59 polymorphism was more frequently found in PUD and GC than gastritis patients. Additionally, presence of Q62 and N122 significantly observed in PUD and GC, whereas I60 was detected in PUD patients. Our results demonstrated that presence of the D, I, Q and N at position 58, 60, 62 and 122, respectively increased the risk of peptic ulcer. However, amino acid N, M, Q and N at the same position alongside V134 increased the risk of gastric cancer.

Expression of a Chimeric Allergen with High Rare Codons Content in Codon Bias-Adjusted Escherichia coli: Escherichia coli BL21 (DE3)-Codon Plus RIL as an Efficient Host.

The expression of heterologous proteins in Escherichia coli (E. coli) is importantly affected by codon bias. Hence, the aim of the current study was to determine which codon bias-adjusted E. coli strain is sufficient for expression of a chimeric allergen coded by high rare codon content. To investigate the expression level, a chimeric protein of Chenopodium album (C. album) was used as an appropriate model. An expression construct was assembled and was transformed to four strains of codon bias-adjusted E. coli including origami, BL21 (DE3), BL21 (DE3)-codon plus RIL, and Rosetta. The level of expression and solubility of the chimeric allergen was analyzed by SDS-PAGE. In addition, the allergenicity of chimeric allergen was determined using immunoblotting. Our results showed that the chimeric allergen was expressed at high level in E. coli BL21 (DE3)-codon plus RIL and Rosetta. In detail, this recombinant allergen was isolated from soluble fraction in the codon bias-adjusted strains of E. coli BL21 (DE3)-codon plus RIL and Rosetta. Moreover, some lower molecular weight proteins were observed in Rosetta, which could be related to inappropriate expression or broken compartments of the chimeric allergen. The immunoblotting assay confirmed that the IgE-specific immune reactivity of our chimeric allergen expressed in BL21 (DE3)-codon plus RIL was significantly higher than the other strains. Our results showed that the expression of the chimeric allergen with high rare codons content in a codon bias-adjusted strain E. coli BL21 (DE3)-codon plus RIL improves the quality and solubility of the heterologous protein production.

Crocin attenuates the lipopolysaccharide-induced neuroinflammation via expression of AIM2 and NLRP1 inflammasome in an experimental model of Parkinson's disease.

The underlying mechanisms of inflammasome activation and the following dopaminergic neuron loss caused by chronic neuroinflammation remain entirely unclear. Therefore, this study aimed to investigate the impact of crocin on the inflammasome complex within an experimental model of Parkinson's disease (PD) using male Wistar rats. PD was induced by the stereotaxic injection of lipopolysaccharide (LPS), and crocin was intraperitoneally administrated one week before the lesion, and then treatment continued for 21 days. Open field (OF) and elevated plus maze tests were applied for behavioral assays. Furthermore, hematoxylin and eosin (H&E) and immunostaining were performed on whole brain tissue, while dissected substantia nigra (SN) was used for immunoblotting and real-time PCR to evaluate compartments involved in PD. The time spent in the center of test was diminished in the LPS group, while treatment with 30 mg/kg of crocin significantly increased it. H&E staining showed a significant increase in cell infiltration at the site of LPS injection, which was ameliorated upon crocin treatment. Notably, crocin-treated animals showed a reduced number of caspase-1 and IL-1ß positive cells, whereas the number of positive cells was increased in the LPS group (P < 0.05). A significant decrease in tyrosine hydroxylase (TH) expression was also found in the LPS group, while crocin treatment significantly elevated its expression. IL-1ß, IL-18, NLRP1, and AIM2 genes expression significantly increased in the LPS group. On the other hand, treatment with 30 mg/kg of crocin significantly downregulated the expression levels of these genes along with NLRP1 (P < 0.05). In summary, our findings suggest that crocin reduces neuroinflammation in PD by diminishing IL-1ß and caspase-1 levels, potentially by inhibiting the expression of AIM2 and NLRP1 genes.

Immunotherapy with a recombinant hybrid molecule alleviates allergic responses more efficiently than an allergenic cocktail or pollen extract in a model of chenopodium album allergy.

BACKGROUND: The aim of this study is to assess the therapeutic potential of a recombinant hybrid molecule (rHM) alongside an allergenic cocktail from recombinant wild-type allergens as well as pollen extract on Chenopodium album allergy, using a BALB/c mouse model. METHODS: The BALB/c mice had already been sensitized to C. album via intraperitoneal injections of alum-adsorbed allergenic cocktail and immunotherapy procedure was followed by subcutaneous injections of the rHM, allergenic cocktail and pollen extract at weekly intervals. Humoral immune responses were determined via measurement of specific antibodies in serum. Splenocytes of immunized mice were stimulated in vitro and then proliferation responses, cytokine secretion and mRNA expression of genes involved in immunotherapy were examined by ELISA and real-time PCR. RESULTS: Sensitized mice were identified with high specific IgE against allergenic cocktail when compared with healthy mice. Immunotherapy with the rHM induced the highest ratio of the IgG2a/IgG1 levels compared to allergenic cocktail or C. album pollen extract. The rHM was able to induce proliferative responses as well as the allergenic cocktail in cultured splenocytes. Immunotherapy with the rHM significantly improved secretion of IFN-γ and IL-10, while secretion of IL-13 rapidly diminished. Interestingly, mRNA expression of GATA3 was strongly decreased in rHM-treated mice whereas mRNA expression of T-bet and Foxp3 was significantly increased. CONCLUSION: Our results prove that immunotherapy with the rHM effectively controlled allergic responses by shifting from a Th2-like immune response to a Th1-dominated immune response.

Elevated expression of the AIM2 gene in response to Helicobacter pylori along with the decrease of NLRC4 inflammasome is associated with peptic ulcer development.

Helicobacter Pylori (H. Pylori) cause peptic ulcer disease (PUD), but the inflammasome's role in PUD is not well understood. Therefore this study has investigated inflammasome compartment expression and IL-1ß production in gastritis (G) and peptic ulcer disease. This study was based on gene expression of inflammasome compartments on stomach biopsies of 50 patients with PUD as cases and 50 individuals with gastritis as controls. The expression of NLRC4, ASC, IL-18, and serum IL-1ß decreased in the PUD group compared to the control group. AIM2 gene expression increased, and NLRP12 gene expression decreased in H. pylori-seropositive positive (HP+ ) individuals compared to H. pylori-seronegative (HP- ) individuals. The G-HP+ subjects had higher serum IL-1ß and AIM2 gene expression than G-HP- subjects but lower NLRP3 and NLRP12 gene expression. The PUD-HP+ had lower serum IL-1ß, but higher AIM2 and IL-18 expression than PUD-HP- . The PUD-HP- patients had decreased IL-18 expression than G-HP- group. The PUD-HP+ had lower serum IL-1ß and NLRC4 expression than G-HP+, while NLRP1 and NLRP3 were higher in expression in PUD-HP+ . The expression of caspase-1, NLRP3 and NAIP were correlated with IL-1ß and IL-18. In conclusion, a decrease in NLRC4, IL-18, ASC genes, and IL-1ß levels in PUD patients compared to gastritis may act in the development of PUD. H. pylori caused AIM2 induction and reduced NLRP12, indicating their contribution to bacterial responses. Decreased NLRC4 expression and IL-1ß protein, together with enhanced NLRP1, and NLRP3 expression, promotes H. pylori to develop peptic ulcers.

Tollip deficiency exaggerates airway type 2 inflammation in mice exposed to allergen and influenza A virus: role of the ATP/IL-33 signaling axis.

Toll-interacting protein (Tollip) is a negative regulator of the pro-inflammatory response to viruses, including influenza A virus (IAV). Genetic variation of Tollip has been associated with reduced airway epithelial Tollip expression and poor lung function in patients with asthma. Whether Tollip deficiency exaggerates type 2 inflammation (e.g., eosinophils) and viral infection in asthma remains unclear. We sought to address this critical, but unanswered question by using a Tollip deficient mouse asthma model with IAV infection. Further, we determined the underlying mechanisms by focusing on the role of the ATP/IL-33 signaling axis. Wild-type and Tollip KO mice were intranasally exposed to house dust mite (HDM) and IAV with or without inhibitors for IL-33 (i.e., soluble ST2, an IL-33 decoy receptor) and ATP signaling (i.e., an antagonist of the ATP receptor P2Y13). Tollip deficiency amplified airway type 2 inflammation (eosinophils, IL-5, IL-13 and mucins), and the release of ATP and IL-33. Blocking ATP receptor P2Y13 decreased IL-33 release during IAV infection in HDM-challenged Tollip KO mice. Furthermore, soluble ST2 attenuated airway eosinophilic inflammation in Tollip KO mice treated with HDM and IAV. HDM challenges decreased lung viral load in wild-type mice, but Tollip deficiency reduced the protective effects of HDM challenges on viral load. Our data suggests that during IAV infection, Tollip deficiency amplified type 2 inflammation and delayed viral clearance, in part by promoting ATP signaling and subsequent IL-33 release. Our findings may provide several therapeutic targets, including ATP and IL-33 signaling inhibition for attenuating excessive airway type 2 inflammation in human subjects with Tollip deficiency and IAV infection.

Diagnosis of Chenopodium album allergy with a cocktail of recombinant allergens as a tool for component-resolved diagnosis.

Chenopodium album pollen is one of the main sources of pollen allergy in desert and semi-desert areas and contains three identified allergens, so the aim of this study is comparison of the diagnostic potential of C. album recombinant allergens in an allergenic cocktail and C. album pollen extract. Diagnostic potential of the allergenic cocktail was investigated in 32 individuals using skin prick test and obtained results were compared with the acquired results from C. album pollen extract. Specific IgE reactivity against the pollen extract and allergenic cocktail was determined by ELISA and western blotting tests. Inhibition assays were performed for the allergenic cocktail characterization. The exact sensitization profile of all patients was identified which showed that 72, 81 and 46% of allergic patients had IgE reactivity to rChe a 1, rChe a 2 and rChe a 3, respectively. Almost all of C. album allergic patients (30/32) had specific IgE against the allergenic cocktail. In addition, there was a high correlation between IgE levels against the allergenic cocktail and IgE levels against the pollen extract. The allergenic cocktail was able to completely inhibit IgE binding to natural Che a 1, Che a 2 and Che a 3 in C. album extract. In addition, positive skin test reactions were seen in allergic patients that tested by the allergenic cocktail. The reliable results obtained from this study confirmed that the allergenic cocktail with high diagnostic potential could be replaced with natural C. album allergen extracts in skin prick test and serologic tests.

Applying high throughput and comprehensive immunoinformatics approaches to design a trivalent subunit vaccine for induction of immune response against emerging human coronaviruses SARS-CoV, MERS-CoV and SARS-CoV-2.

Coronaviruses (CoVs) cause diseases such as severe acute respiratory syndrome (SARS), Middle East respiratory syndrome (MERS), and coronavirus disease 2019 (COVID-19). Therefore, this study was conducted to combat major CoVs via a trivalent subunit vaccine, which was engineered by implementing sequences of spike (S) protein, nucleocapsid (N), envelope (E), membrane (M) protein, non-structural protein (nsp) 3, and nsp8 antigens. The CTL, HTL, MHC I, and IFN-γ epitopes were predicted via CTLPRED, IEDB, and IFN epitope servers, respectively. Also, to stimulate strong helper T lymphocytes (HTLs) responses, Pan HLA DR-binding epitope (PADRE) was used. Also, for boosting the immune response, ß-defensin 2 was added to the construct as an adjuvant. Furthermore, TAT was applied to the vaccine to facilitate the intracellular delivery. Finally, TAT, adjuvant, PADRE, and selected epitopes were appropriately assembled. Based on the predicted epitopes, a trivalent multi-epitope vaccine with a molecular weight of 74.8 kDa was constructed. Further analyses predicted the molecule to be a strong antigen, and a non-allergenic and soluble protein. Secondary and tertiary structures were predicted. Additionally, analyses validated the stability of the proposed vaccine. Molecular docking and molecular dynamics simulation (MDS) showed binding affinity and stability of the vaccine-TLR3 complex was favorable. The predicted epitopes demonstrated a strong potential to stimulate T and B-cell mediated immune responses. Furthermore, codon optimization and in silico cloning guaranteed increased expression. In summary, investigations demonstrated that this next-generation approach might provide a new horizon for the development of a highly immunogenic vaccine against SARS-CoV, MERS-CoV, and SARS-CoV-2.Communicated by Ramaswamy H. Sarma.

In silico design of a TLR4-mediating multiepitope chimeric vaccine against amyotrophic lateral sclerosis via advanced immunoinformatics.

Amyotrophic lateral sclerosis (ALS) is the most prevalent motor neuron disorder worldwide. In ALS, progressing disease can result from misfolding and aggregation of superoxide dismutase-1 (SOD1) or TAR DNA-binding protein 43 kDa (TDP43). An efficient immunotherapy for ALS should spare intact SOD1 while eliminating its dysfunctional variant. We utilized advanced immunoinformatics to suggest a potential vaccine candidate against ALS by proposing a model of dynamic TLR4 mediation and induction of a specific Th2-biased shift against mutant SOD1, TDP43, and TRAF6, a protein that specifically interacts with dysfunctional SOD1. SOD1, TDP43, and TRAF6 were retrieved in FASTA. Immune Epitopes Database and CTLpred suggested T/B-cell epitopes from disease-specific regions of selected antigens. A TLR4-mediating adjuvant, RS01, was used. Sequences were assembled via suitable linkers. Tertiary structure of the protein was calculated. Refined protein structure and physicochemical features of the 3D structure were verified in silico. Differential immune induction was assessed via C-ImmSim. GROningen MAchine for Chemical Simulation was used to assess evolution of the docked vaccine-TLR4 complex in blood. Our protein showed high structural quality and was nonallergenic and immune inducing. Also, the vaccine-TLR4 complex stability was verified by RMSD, RMSF, gyration, and visual analyses of the molecular dynamic trajectory. Contact residues in the vaccine-TLR4 complex showed favorable binding energies. Immune stimulation analyses of the proposed candidate demonstrated a sustained memory cell response and a strong adaptive immune reaction. We proposed a potential vaccine candidate against ALS and verified its physicochemical and immune inducing features. Future studies should assess this vaccine in animal studies.

Anti-Cancer Effect of Bromelain and Its Combination with Cisplatin on HN5 Cell Line (Squamous Cell Carcinoma).

Statement of the Problem: Squamous cell carcinoma (SCC) comprises over 90% of oral malignancies. Cisplatin, as a selective chemotherapy agent to treat SCC, has many side effects despite its high effectiveness. There are some studies on the effects of bromelain derived from pineapple stems on different malignancies. Purpose: The aim of this study was to investigate the effect of bromelain alone and in combination with Cisplatin on oral squamous cell carcinoma (OSCC) and fibroblast cell lines. Materials and Method: In this interventional study, the HN5 cell line of OSCC and fibroblast cell line were treated with different concentrations of bromelain alone and in combination with cisplatin. Cell viability test was performed after 24, 48 and 72 hours using MTT (3-)4,5-dimethylthiazol-2-yl(-2,5 diphenyl tetrazolium bromide) assay. In the final stage, the drug-treated cells underwent flow cytometry to assess apoptosis patterns. Data were analyzed using SPSS 17, ANOVA (for general comparison of groups) and LSD post hoc tests (for comparison two groups). p< 0.05 was considered statistically significant. Results: The findings suggested that although bromelain showed toxic effects on HN5 cancer cells, its combination with Cisplatin resulted in little improvement in its effectiveness. Bromelain alone and in combination with Cisplatin presented cytotoxic effects against fibroblasts, which depended on the dosage and time exposure (p< 0.05). The flow cytometry results did not support the superior effect of the combination of two medications over Cisplatin alone (p> 0.05). Conclusion: According to the findings, although adding bromelain to Cisplatin reduced toxicity on normal tissues, the combination of these two drugs did not increase the anticancer effect of Cisplatin. Thus, bromelain in combination with Cisplatin is not recommended as an adjuvant drug for OSCC.