Regulation of immune responses to infection through interaction between stem cell-derived exosomes and toll-like receptors mediated by microRNA cargoes.

Infectious diseases are among the factors that account for a significant proportion of disease-related deaths worldwide. The primary treatment approach to combat microbial infections is the use of antibiotics. However, the widespread use of these drugs over the past two decades has led to the emergence of resistant microbial species, making the control of microbial infections a serious challenge. One of the most important solutions in the field of combating infectious diseases is the regulation of the host's defense system. Toll-like receptors (TLRs) play a crucial role in the first primary defense against pathogens by identifying harmful endogenous molecules released from dying cells and damaged tissues as well as invading microbial agents. Therefore, they play an important role in communicating and regulating innate and adaptive immunity. Of course, excessive activation of TLRs can lead to disruption of immune homeostasis and increase the risk of inflammatory reactions. Targeting TLR signaling pathways has emerged as a new therapeutic approach for infectious diseases based on host-directed therapy (HDT). In recent years, stem cell-derived exosomes have received significant attention as factors regulating the immune system. The regulation effects of exosomes on the immune system are based on the HDT strategy, which is due to their cargoes. In general, the mechanism of action of stem cell-derived exosomes in HDT is by regulating and modulating immunity, promoting tissue regeneration, and reducing host toxicity. One of their most important cargoes is microRNAs, which have been shown to play a significant role in regulating immunity through TLRs. This review investigates the therapeutic properties of stem cell-derived exosomes in combating infections through the interaction between exosomal microRNAs and Toll-like receptors.

Activated mesenchymal stem cells increase drug susceptibility of methicillin-resistant Staphylococcus aureus and Pseudomonas aeruginosa.

Methicillin-resistant Staphylococcus aureus (MRSA) and Pseudomonas aeruginosa are major causes of hospital-acquired infections and sepsis. Due to increasing antibiotic resistance, new treatments are needed. Mesenchymal stem cells (MSCs) have antimicrobial effects, which can be enhanced by preconditioning with antibiotics. This study investigated using antibiotics to strengthen MSCs against MRSA and P. aeruginosa. MSCs were preconditioned with linezolid, vancomycin, meropenem, or cephalosporin. Optimal antibiotic concentrations were determined by assessing MSC survival. Antimicrobial effects were measured by minimum inhibitory concentration (MIC), minimum bactericidal concentration (MBC), and antimicrobial peptide (AMP) gene expression. Optimal antibiotic concentrations for preconditioning MSCs without reducing viability were 1 µg/mL for linezolid, meropenem, and cephalosporin and 2 µg/mL for vancomycin. In MIC assays, MSCs preconditioned with linezolid, vancomycin, meropenem, or cephalosporin inhibited MRSA or P. aeruginosa growth at lower concentrations than non-preconditioned MSCs (p ≤ 0.001). In MBC assays, preconditioned MSCs showed enhanced bacterial clearance compared to non-preconditioned MSCs, especially when linezolid and vancomycin were used against MRSA (p ≤ 0.05). Preconditioned MSCs showed increased expression of genes encoding the antimicrobial peptide genes hepcidin and LL-37 compared to non-preconditioned MSCs. The highest hepcidin expression was seen with linezolid and vancomycin preconditioning (p ≤ 0.001). The highest LL-37 expression was with linezolid preconditioning (p ≤ 0.001). MSCs' preconditioning with linezolid, vancomycin, meropenem, or cephalosporin at optimal concentrations enhances their antimicrobial effects against MRSA and P. aeruginosa without compromising viability. This suggests preconditioned MSCs could be an effective adjuvant treatment for antibiotic-resistant infections. The mechanism may involve upregulation of AMP genes.

The effects of functional foods mixture on inflammatory cytokines and biochemical findings in hospitalized patients with COVID-19: a randomized double-blind controlled trial.

BACKGROUND: The coronavirus disease 2019 (COVID-19) pandemic has been shown to affect nutritional recommendations. Some functional foods have been demonstrated to be useful in the treatment of people with COVID-19. However, little is known about the impact of combining functional foods on disease control. This study aimed to investigate the effects of functional foods mixture on serum levels of inflammatory cytokines and biochemical findings in patients with COVID-19. METHODS: A randomized double-blind controlled trial was conducted in Baqiyatallah Al-Azam hospital in Tehran, Iran. Sixty patients were randomly assigned to receive either a soup containing functional foods (n = 30) or a usual soup (control group) (n = 30). Participants' sociodemographic information was gathered using a general questionnaire. Blood levels of inflammatory markers and biochemical findings were assessed using standard protocols. RESULTS: The results showed that soup containing functional foods was more effective in controlling serum levels of D-dimer, blood urea nitrogen, and creatinine than the control group (P < 0.05). Also, more significant improvement was found in the intervention group vs control group in terms of interleukin (IL)-1ß, IL-6, IL-17, IL-10, and tumor necrose factor-α (P < 0.05). In contrast, the control intervention more efficiently controlled potassium levels and reduced quantitative C-reactive protein than the intervention group (P < 0.05). CONCLUSIONS: This study indicates a soup containing functional foods could alleviate biomarkers of inflammation in patients with COVID-19. However, its effectiveness on biochemical findings remained inconclusive which warranted further research. TRIAL REGISTRATION: IRCT, IRCT20180201038585N11. Registered 23 August 2021, https://www.irct.ir/trial/57338.

Targeting glioblastoma multiforme using a novel fusion protein comprising interleukin-13 and staphylococcal enterotoxin B in vitro.

Targeting cell surface receptors with immunotoxins provides a novel, unique and highly potent treatment against cancers. A high expression of interleukin-13 (IL13) receptor α2 (IL13Rα2) has been reported in different types of cancers including glioblastoma multiforme (GBM). In this paper, to target IL13Rα2 on GBM cells, a fusion protein was generated comprising human IL13 and staphylococcal enterotoxin B (SEB), termed IL13-linker-SEB. The fusion protein was cloned into pET28a(+) and expressed in Escherichia coli strain BL21 (DE3); U251 (IL13Rα2-positive) and T98G (IL13Rα2-negative) GBM cell lines were employed and the functional activity of IL13-linker-SEB was evaluated by cell ELISA, cytotoxicity (MTT and LDH), apoptosis (flow cytometry and caspase-3 activity), adhesion, scratch and RT-PCR tests. SEB and chemotherapeutic drugs were employed to be compared to IL13-linker-SEB function. The IL13-linker-SEB exhibited higher binding affinity and cytotoxicity compared to SEB on U251 cells, although both recombinant proteins had shown similar behavior regarding T98G cells. Furthermore, the highest induction of apoptosis was observed in U251 cells treated with IL13-linker-SEB which was confirmed by Bax/Bcl-2 ratio. The expression of MMP2, MMP9 and VEGFR2 in U251 cells experienced a significant reduction after treatment with IL13-linker-SEB compared to SEB and T98G treated cells. The data showed that IL13-linker-SEB can be considered as a novel potential agent for GBM treatment; however, further research is needed to investigate the efficacy.

A comparative study of the arazyme-based fusion proteins with various ligands for more effective targeting cancer therapy: an in-silico analysis.

Background and purpose: Recently, the use of immunotoxins for targeted cancer therapy has been proposed, to find new anticancer drugs with high efficacy on tumor cells with minimal side effects on normal cells. we designed and compared several arazyme (AraA)-based fusion proteins with different ligands to choose the best-targeted therapy for interleukin 13 receptor alpha 2 (IL13Rα2)-overexpressed cancer cells. For this purpose, IL13Rα2 was selected as a receptor and IL13 and IL13.E13K were evaluated as native and mutant ligands, respectively. In addition, Pep-1 and A2b11 were chosen as the peptide ligands for targeted cancer therapy. Experimental approach: Several bioinformatics servers were used for designing constructs and optimization. The structures of the chimeric proteins were predicted and verified by I-TASSER, Q-Mean, ProSA, Ramachandran plot, and Verify3D program. Physicochemical properties, toxicity, and antigenicity were predicted by ProtParam, ToxinPred, and VaxiJen. HawkDock, LigPlot+, and GROMACS software were used for docking and molecular dynamics simulation of the ligand-receptor interaction. Findings/Results: The in silico results showed AraA-A2b11 has higher values of confidence score and Q-mean score was obtained for high-resolution crystal structures. All chimeric proteins were stable, non-toxic, and non-antigenic. AraA-(A(EAAAK)4ALEA(EAAAK)4A)2-IL13 retained its natural structure and based on ligand-receptor docking and molecular dynamic analysis, the binding ability of AraA-(A(EAAAK)4ALEA(EAAAK)4A)2-IL13 to IL13Rα2 was sufficiently strong. Conclusion and implications: Based on the bioinformatics result AraA-(A(EAAAK)4ALEA(EAAAK)4A)2-IL13 was a stable fusion protein with two separate domains and high affinity with the IL13Rα2 receptor. Therefore, AraA-(A(EAAAK)4ALEA(EAAAK)4A)2-IL13 fusion protein could be a new potent candidate for target cancer therapy.

Staphylococcal enterotoxin B as DNA vaccine against breast cancer in a murine model.

Recently, many efforts have been made to treat cancer using recombinant bacterial toxins and this strategy has been used in clinical trials of various cancers. Therapeutic DNA cancer vaccines are now considered as a promising strategy to activate the immune system against cancer. Cancer vaccines could induce specific and long-lasting immune responses against tumors. This study aimed to evaluate the antitumor potency of the SEB DNA vaccine as a new antitumor candidate against breast tumors in vivo. To determine the effect of the SEB construct on inhibiting tumor cell growth in vivo, the synthetic SEB gene, subsequent codon optimization, and embedding the cleavage sites were sub-cloned to an expression vector. Then, SEB construct, SEB, and PBS were injected into the mice. After being vaccinated, 4T1 cancer cells were injected subcutaneously into the right flank of mice. Then, the cytokine levels of IL-4 and IFN-γ were estimated by the ELISA method to evaluate the antitumor activity. The spleen lymphocyte proliferation, tumor size, and survival time were assessed. The concentration of IFN-γ in the SEB-Vac group showed a significant increase compared to other groups. The production of IL-4 in the group that received the DNA vaccine did not change significantly compared to the control group. The lymphocyte proliferation increased significantly in the mice group that received SEB construct than PBS control group (p < 0.001). While there was a meaningful decrease in tumor size (p < 0.001), a significant increase in tumor tissue necrosis (p < 0.01) and also in survival time of the animal model receiving the recombinant construct was observed. The designed SEB gene construct can be a new model vaccine for breast cancer because it effectively induces necrosis and produces specific immune responses. This structure does not hurt normal cells and is a safer treatment than chemotherapy and radiation therapy. Its slow and long-term release gently stimulates the immune system and cellular memory. It could be applied as a new model for inducing apoptosis and antitumor immunity to treat cancer.

Evaluation of the Role of Probiotics As a New Strategy to Eliminate Microbial Toxins: a Review.

Probiotics are living microorganisms that have favorable effects on human and animal health. The most usual types of microorganisms recruited as probiotics are lactic acid bacteria (LAB) and bifidobacteria. To date, numerous utilizations of probiotics have been reported. In this paper, it is suggested that probiotic bacteria can be recruited to remove and degrade different types of toxins such as mycotoxins and algal toxins that damage host tissues and the immune system causing local and systemic infections. These microorganisms can remove toxins by disrupting, changing the permeability of the plasma membrane, producing metabolites, inhibiting the protein translation, hindering the binding to GTP binding proteins to GM1 receptors, or by preventing the interaction between toxins and adhesions. Here, we intend to review the mechanisms that probiotic bacteria use to eliminate and degrade microbial toxins.

A comparative study of laboratory findings in PCR-positive and PCR-negative COVID-19 hospitalized patients.

INTRODUCTION: Given the many misconceptions in terms of both diagnosis and treatment, SARS-CoV-2 continues to infect and victimize. Notwithstanding molecular testing is the gold standard method of in vitro diagnostic, the often long-waiting time, as well as false-negative results are daunting challenges facing us. In this study, we aimed to report the diagnostic value of laboratory findings in COVID-19 patients, with an extensive focus on the differences between PCR-positive and PCR-negative cases. PATIENTS AND METHODS: We did a retrospective single-centre study on a large cohort of 1546 COVID-19 patients in Tehran, Iran. Based on clinical symptoms, chest CTs were performed for all the patients. Also, molecular testing of swab specimens was also performed for 1450 cases. RESULTS: All the data on laboratory results were retrospectively extracted from medical records. Of the 1546 patients, 1040 (67.5%) were male and 506 (32.5%) were female with the mean age of 55.67. On admission, 31.4% of the whole study population displayed lymphopenia and 38.9% showed neutrophilia. Decreased hemoglobin and mild thrombocytopenia were also found in 40% and 18.6% of cases, respectively. Elevated lactate dehydrogenase in nearly 75% of COVID-19 cases was the most common alteration amongst biochemical parameters which together with increased ESR and CRP could serve as diagnostic markers in SARS-CoV-2 infection. Of the 1450 patients with a PCR result, 439 (28.3%) were PCR-negative and 1011 (65.3%) were PCR-positive. Notably, lymphopenia and increased AST were higher in the PCR-positive group than their negative counterparts. Albeit being in the normal range, a significant decrease in the number of monocytes was also evident in the PCR-positive cases. CONCLUSIONS: As far we are aware, this is the first time that we reported a comprehensive exploration of laboratory characteristics of a large cohort of hospitalized COVID-19 patients from Iran, hoping that these data will cast more light on the diagnostic significance of these parameters.

Diagnostic Accuracy and Validity of Serological and Molecular Tests for Hepatitis B and C.

INTRODUCTION: Hepatitis B and C viruses are one of the leading causes of health problems in the world and early diagnosis and treatment of them are very important. Thereby, this study aimed to evaluate the validity and reliability of usable diagnostic tests for the detection of hepatitis B and C viruses in the clinical setting and to compare them with each other. MATERIALS AND METHODS: In this review article, we have searched major online databases, including PubMed and EMBASE. 42 retrieved articles were published between January 2000 and January 2020, which are summarized in this review. RESULTS: Immunoassay approaches are general techniques for the identification of pathogenic agents, among which ELISA is the gold standard for the detection of HBsAg. While serological techniques are not conclusive, molecular assays are really important because of the high sensitivity to detect chronic hepatitis B without HBeAg, in which viral loads are very low. Biosensors have more elevated selectivity and sensitivity and faster responses compared to other methods. CONCLUSION: This study suggests that all of the molecular, serological, and biotechnological assays have advantages and disadvantages for diagnosing hepatitis B and C viruses which are dependent on the condition, so we should choose one of them in regards to the time, cost, and laboratory equipment along with the clinical symptoms.

TGFαL3-SEB fusion protein as an anticancer against ovarian cancer.

TGFαL3-SEB is a new synthetic fusion protein produced by the combination of the third loop of transforming growth factor with staphylococcal enterotoxin type B. In the current study, the anti-tumor effects of TGFαL3-SEB were evaluated against SKOV3 cells, which highly expressed the epidermal growth factor receptor (EGFR). Our findings showed that incubation of SKOV3 cells with 75, 100 and 150 µg/ml of TGFαL3-SEB significantly reduces the proliferation rate in a concentration-dependent manner (P < 0.05) and its IC50 value was 110 µg/ml. Caspase-3 activity was increased from 100% for control cells to 109, 144, and 169% for 75, 100 and 150 µg/ml of TGFαL3-SEB treatment, respectively. Caspase-9 activity and bax/bcl-2 ratio were also confirmed the apoptosis induction ability of TGFαL3-SEB (P < 0.001). Flow cytometry examination also showed that apoptosis was induced and the number of apoptotic cells was increased from 8.2% in un-treated cells to 20.9, 50, and 90% in response to 75, 100 and 150 µg/ml of TGFαL3-SEB fusion protein in a concentration-dependent manner (P < 0.05). The mRNA expression level of VEGF was also reduced to 0.89, 0.69, and 0.60, respectively in response to 75, 100 and 150 µg/ml of TGFαL3-SEB fusion protein exposure, respectively (P < 0.5). In summary, the findings of our study uncovered that TGFαL3-SEB fusion protein induced apoptosis and reduced angiogenesis in SKOV3 ovarian cancer cells in a concentration-dependent manner. This protein has the potential to act against EGFR expressing malignant cells to serve as a pro-apoptotic and angiogenesis blocker agents; however, further studies are needed to confirm its ability.

Mesenchymal stem cells preconditioned by staphylococcal enterotoxin B enhance survival and bacterial clearance in murine sepsis model.

Sepsis, a health-threatening progressive infectious disease, is the major cause of morbidity and mortality worldwide. Cell therapy using mesenchymal stromal cells (MSCs) is an innovative strategy with excessive therapeutic potential in the treatment of sepsis. Staphylococcal enterotoxin B (SEB) preconditioning aims to prolong the interval of survival of transplanted MSCs which induces the production of cytoprotective agents, anti-apoptotic and anti-inflammatory factors. The MSCs were preconditioned with an optimum dose of SEB (470 µmol/L). The expression levels of apoptosis genes and antibacterial activity of MSC and SEB-MSC and their conditioned medium (CM), as well as cell survival, were studied in vitro in an oxidative stress and serum deprivation condition. Following treatment of the septic mice with MSCs and SEB-MSCs, pro/anti-inflammatory cytokines, hematological factors, bacterial clearance and animal survival were assessed. The apoptotic and pro-inflammatory cytokine's genes expression was down-regulated while antibacterial peptides and anti-inflammatory cytokines were up-regulated in SEB-MSC-treated mice. The animal survival rates were improved; bacterial clearance was enhanced in the peritoneal fluids, blood and organs; aspartate aminotransferase (AST) and alanine aminotransferase (ALT) were reduced in blood, compared with saline and MSCs alone. This research concludes that transplantation of SEB-MSCs presents improved therapeutic effects on a live bacterial model of sepsis.

Antimicrobial effects of mesenchymal stem cells primed by modified LPS on bacterial clearance in sepsis.

BACKGROUND AND OBJECTIVES: Mesenchymal stem cells (MSCs)-based regenerative therapy is now considered as an alternative approach to revive infectious diseases, including sepsis. Nevertheless, the efficiency of MSC application is limited by the poor survival rate of engrafted MSCs. Hence, preconditioning was established as a strategy to increase the cells' efficiency. METHODS: MSCs were preconditioned with 1 µg/ml of three different lipopolysaccharides (LPSs) of Pseudomonas (Pse-LPS), Acinetobacter (Ac-LPS), and Acinetobacter inactivated lipid A by PagL (Ac-LPS-PagL). Then, preconditioned MSCs were exposed to oxidative stress and serum deprivation followed by evaluation of the antibacterial activity, survival, and apoptosis of MSCs. Then, the murine sepsis model treated with 100 µl phosphate-buffered saline (control group, sepsis group), 100 µl of 1 × 10 6 wild MSCs (MSC group), and three remained groups received 100 µl of 1 × 10 6 LPS-preconditioned MSCs (Pse-LPS-MSCs group: LPS purified from Pseudomonas, or Ac-LPS-MSCs group: LPS purified from Acinetobacter, and Ac-PagL-LPS-MSCs group: detoxified LPS Pagl). RESULTS: After 4 days, LPS-preconditioned MSC transplantation modulated the immune response and reduced inflammation in septic mice. Apoptosis of Pse-LPS/Ac-LPS-preconditioned-MSCs was obviously reduced in vitro, and the survival rate of engrafted mice was evidently elevated in Pse-LPS-MSCs and Ac-LPS-MSCs groups compared with other three groups. CONCLUSION: LPS preconditioning provides an innovative strategy for evolving functional and biological properties of MSCs and ameliorates the survival rate of the mouse model of sepsis after MSC transplantation, protects cells from apoptosis and organ damages, and evaluates therapeutic properties, including immunemodulatory.

Staggered Target SELEX, a novel approach to isolate non-cross-reactive aptamer for detection of SEA by apta-qPCR.

BACKGROUND AND OBJECTIVE: Aptamers or chemical antibodies are oligonucleotides (DNA or RNA) that are able to bind to various targets with high specificity and affinity such as toxins which are isolated by an in vitro method known as SELEX. To date, there are many SELEX procedures for the isolation of novel aptamers against proteins. However not all modified SELEX are suitable for similar protein based on sequence homology such as staphylococcal enterotoxins. Staphylococcal enterotoxin type A (SEA) is the most prevalent toxin involved in staphylococcal food poisoning (SFP) worldwide. SEA is homologous to Staphylococcal enterotoxin type D (SED) and Staphylococcal enterotoxin type E (SEE) about 50% and 83%, respectively. Here, we have developed Staggered Target SELEX (ST-SELEX) as a novel designed SELEX procedure to acquire specific non-cross-reactive aptamers against SEA as a model protein. METHODS: In this study, isolated ssDNA aptamers by ST-SELEX were used for detection of SEA via apta-Real time PCR (apta-qPCR). After in silico analysis of SEA protein with SEE and finding the specific region on the surface of protein, ST-SELEX was carried out in two steps (classical SELEX and Second SELEX). Finally, after isolating high specific aptamers, the apta-qPCR was used for the detection of the SEA. In this technique, poly-clonal antibody against SEA was immobilized on protein G sepharose beads (Ab-PGs). Then, the SEA protein was captured by poly clonal antibody as the target that immobilized on sepharose beads. The isolated aptamers were bound on the surface of SEA protein that captured by Ab-PGs. Finally, the heat-released aptamers were amplified by qPCR. RESULT: Our investigation showed that the aptamers were generated in vitro by a ten-round selection process based on ST-SELEX procedure with dissociation constant (KD) value 7.44± 0.6 nM and limit of detection (LOD) of 146.67 fM. DISCUSSION AND CONCLUSION: The advantage of ST-SELEX compared to other SELEX methods was to select a specific non cross-reactive aptamer against two or more proteins with high sequence homology. These aptamers can be used in sensitive detection methods such as apta-qPCR.

Topical Botulinum Toxin: A Non-invasive Way for Treatment of Muscle Disorders.

Botulinum neurotoxin type A (BoNT/A) is a toxin that inhibits the release of stimulatory neurotransmitter (acetylcholine) at the neuromuscular synapses. In recent years, many patients with muscle contraction disorders have greatly benefited from the therapeutic ability of this biological drug. On the other hand, the injection of this bio drug is accompanied by some side effects such as irritation, bruising, inflammation, pain, bleeding at the site of injection. Recently, a tendency has been observed among scientists to create new techniques to offer conventional injectable drugs the ability of transdermal delivery. Such promising drugs can be applied in various forms from gel, cream, and ointments to ready-to-use pads. This would eliminate a need for high drug doses to release the drug gradually at the site of application while at the same time, lower the side effects. Here, we discuss the possibility of noninvasive administration of BoNT/A in order to reduce the side effects of drug injection.

Multi-Drug Resistant Clinical Pseudomonas aeruginosas Inhibited by Ferula gummosa Boiss.

BACKGROUND: Multi-drug resistance among Pseudomonas aeruginosa (P. aeruginosa) clinical isolates is increasing and becoming a serious problem for public health authorities worldwide. OBJECTIVE: The aim of the current study is to introduce a potent antibacterial compound against the resistant P. aeruginosa. METHODS: In this study, we evaluated the antibacterial effects of extracts and essential oils of Ferula gummosa Boiss (F. gummosa) on 33 P. aeruginosa clinical isolates by microdilution method and assessed the association of antimicrobial activity with the extended spectrum ß-lactamase (ESBL) producing, biofilm forming and aliginate production of the strains. In addition, the presence of some genes involved in these properties, including blaGES- 1, blaRER-1, blaCTX-M, blaVEB-1, blaOXA-1, blaOXA-4, blaOXA-10, ppyR, pslA, pelA, algU, algL, algD, fliC and oxaA was determined using PCR. RESULTS: We revealed that all of our extracts and essential oils had significant antibacterial effects (p<0.001), but the aqueous extracts showed a relatively lower antibacterial activity compared with the methanolic ones. Furthermore, the minimum inhibitory concentration required for the ESBL producing strains was significantly higher than the non-ESBL producing ones (p<0.001). Loss of some genes such as blaPER-1, blaGES-1, blaOXA-1 and blaOXA-4 caused sensitivity to F. gummosa derivatives (p<0.05). CONCLUSION: The findings of this study indicate that the antibacterial effects of the extract and essential oils of F. gummosa may be a potential novel treatment against drug-resistant P. aeruginosa clinical isolates.

An integrative in silico approach to the structure of Omp33-36 in Acinetobacter baumannii.

Omp33-36 in A. baumannii, a bacterium causing serious nosocomial infections, is a virulence factor associated with the pathogen metabolic fitness as well as its adherence and invasion to human epithelial cells. This protein is also involved in interaction of the bacteria with host cells by binding to fibronectin. Moreover, Omp33-36 renders cytotoxicity to A. baumannii in addition to inducing apoptosis and modulation of autophagy. In the present study, an integrated strategy is launched to pierce into the 3D structure of Omp33-36 protein. The signal peptide within the sequence was determined, then, topology as well as secondary and tertiary structures of the protein were predicted. The mature protein assigned as a 14-stranded barrel in which residues 1-19 is removed as signal peptide. The obtained 3D models were evaluated in terms of quality; and then, served as queries to find similar protein structures. The hits were analyzed regarding topology among which 14-stranded were considered. The most qualified model was refined and then its sequence aligned to its counterpart similar structure protein (CymA from Klebsiella oxytoca). The determined structure of Omp33-36 could justify its porin function and carbapenem-resistance associated with the loss of this protein.

Immunogenicity of chimeric MUC1-HER2 vaccine against breast cancer in mice.

OBJECTIVES: Breast cancer is one of the most common cancers in the world and is on the increase. MUC1 and HER2 as tumor-associated antigens (TAAs) are abnormally expressed to some extent in 75-80% of breast cancers. In our present research, a novel chimeric MUC1-HER2 (HM) protein was designed and used to study whether an immune response can be generated against these TAAs. In vitro analysis of the HER2-MUC1 construct confirmed the co-expression of MUC1 and HER2. MATERIALS AND METHODS: BALB/c mice were immunized with this novel chimeric protein. The humoral immune response was assessed by enzyme-linked immunosorbent assay (ELISA). Then, BALB/c mice were injected subcutaneously 2×105 4T1-MUC1-HER2 tumor cells. Subsequently, tumor size and tumor necrosis measurements, MTT, cytokines assay and survival test were performed. RESULTS: The results implied a critical role of HER2 and MUC1 antibodies in vaccination against breast cancer. This engineered protein can be a good vaccine to stop breast cancer. CONCLUSION: The results implied a critical role of HER2 and MUC1 antibodies in vaccination against breast cancer. This engineered protein can be a good vaccine to stop breast cancer.

High genetic diversity among Mycobacterium tuberculosis strains in Tehran, Iran.

INTRODUCTION: Tuberculosis (TB) still remains an important public health problem in Iran. The genotyping of Mycobacterium tuberculosis isolates is expected to lead to a better understanding of M. tuberculosis transmission in Tehran, the most populated city of Iran. MATERIALS AND METHODS: A total of 2300 clinical specimens were obtained from TB suspected patients who were referred to a TB center in Tehran from Jan 2014 to Dec 2016. Identification was performed using both conventional and molecular methods. The presence of resistance to rifampicin was examined by the GeneXpert MTB/RIF. The standard 15-locus mycobacterial interspersed repetitive units-variable number of tandem repeats (MIRU-VNTR) typing method was applied to genotype of clinical isolates. RESULTS: Of 2300 specimens, 80 isolates were identified as M. tuberculosis by using biochemical and molecular tests. Of 80 M. tuberculosis isolates, 76 (95%) had unique genotypic profiles and 4 (5%) shared a profile with one or more other strains. Based on single loci variation (SLV) 4 clonal complexes were observed. NEW-1 was found to be the most predominant lineage (22.5%) followed by West African (1.25%), Central Asian (CAS)/Delhi (1.25%), Bovis (1.25%), H37Rv (1.25%) and multiple matches (1.25%). Loci MIRU10, MIRU26, MTUB21 and QUB26 were found as highly discriminative. No mutation was detected in the hotspot region of rifampicin by using GeneXpert MTB/RIF. CONCLUSIONS: Our study findings show that there was considerable genotypic diversity among M. tuberculosis isolates in Tehran. The 15-locus MIRU-VNTR showed high HGDI and could be used as a first-line genotyping method for epidemiological studies.

Production of recombinant flagellin to develop ELISA-based detection of Salmonella Enteritidis

ABSTRACT Food-borne diseases, caused by the pathogenic bacteria, are highly prevalent in the world. Salmonella is one of the most important bacterial genera responsible for this. Salmonella Enteritidis (SE) is one of the non-typhoid Salmonellae that can be transmitted to human from poultry products, water, and contaminated food. In recent years, new and rapid detection methods such as enzyme-linked immunosorbent assay (ELISA) and polymerase chain reaction (PCR) have been developed. In this study, recombinant FliC (rFliC) was produced to be used as an antigen. The immunization was conducted in mice with the purified recombinant FliC (rFliC). The mice were subcutaneously immunized with rFliC and elicited significant rFliC specific serum IgG antibodies. An indirect ELISA system was established for the detection of Salmonella Enteritidis. Our results confirmed that the recombinant flagellin can be one of the excellent indicators for the detection of Salmonella Enteritidis.