Preparation of Cerium Oxide Nanoparticles and Their Cytotoxicity Evaluation In vitro and In vivo

Background: Nanotechnology plays a significant role in medicine, especially in diagnosis and treatment as a carrier to drugs and vaccinology. Several studies were conducted using nanoparticles as an adjuvant. The main aim of this study was in vivo and in vitro toxicity evaluation of synthesized Cerium Nanoparticles (CeNPs).Methods: In the present study, cerium nanoparticles were prepared using the wet chemical method. The formation of cerium nanoparticles was confirmed by scanning electron microscopy, transmission electron microscopes, x-ray diffraction analysis, dynamic light scattering. In vivo and in vitro toxicity of synthesized nanoparticles was evaluated in three different amounts of cerium nanoparticles (30 µg, 50 µg, & 100 µg) in mice and human fibroblast cell lines, respectively.Results: Cerium nanoparticles were successfully synthesized, and the identity was confirmed by x-ray diffraction analysis. The shape and size of nanoparticles were spherical and <100 nm, respectively. The prepared nanoparticles had a charge of -26.6 mV and a hydrodynamic radius of 446 nm. MTT assay indicated that none of the concentration of cerium was toxic, and in vivo toxicity also clarified the safety of cerium nanoparticles in mice; no significant un-normal behavioral and physical symptoms were observed in mice after CeNP administrationConclusion: Cerium nanoparticles have special properties, especially low toxicity, unique capabilities in stimulating the immune system. Cerium nanoparticles can be considered an effective and safe candidate in vaccines

Development of a bivalent protein-based vaccine candidate against invasive pneumococcal diseases based on novel pneumococcal surface protein A in combination with pneumococcal histidine triad protein D

Extensive efforts have been made toward improving effective strategies for pneumococcal vaccination, focusing on evaluating the potential of multivalent protein-based vaccines and overcoming the limitations of pneumococcal polysaccharide-based vaccines. In this study, we investigated the protective potential of mice co-immunization with the pneumococcal PhtD and novel rPspA proteins against pneumococcal sepsis infection. The formulations of each antigen alone or in combination were administered intraperitoneally with alum adjuvant into BALB/c mice three times at 14-day intervals. The production of antigen-specific IgG, IgG1 and IgG2a subclasses, and IL-4 and IFN-γ cytokines, were analyzed. Two in vitro complement- and opsonophagocytic-mediated killing activities of raised antibodies on day 42 were also assessed. Finally, the protection against an intraperitoneal challenge with 106 CFU/mouse of multi-drug resistance of Streptococcus pneumoniae ATCC49619 was investigated. Our findings showed a significant increase in the anti-PhtD and anti-rPspA sera IgG levels in the immunized group with the PhtD+rPspA formulation compared to each alone. Moreover, the results demonstrated a synergistic effect with a 6.7- and 1.3- fold increase in anti-PhtD and anti-rPspA IgG1, as well as a 5.59- and 1.08- fold increase in anti-PhtD and anti-rPspA IgG2a, respectively. Co-administration of rPspA+PhtD elicited a mixture of Th-2 and Th-1 immune responses, more towards Th-2. In addition, the highest complement-mediated killing activity was observed in the sera of the immunized group with PhtD+rPspA at 1/16 dilution, and the opsonophagocytic activity was increased from 74% to 86.3%. Finally, the survival rates showed that mice receiving the rPspA+PhtD formulation survived significantly longer (100%) than those receiving protein alone or PBS and exhibited the strongest clearance with a 2 log10 decrease in bacterial load in the blood 24h after challenge compared to the control group. In conclusion, the rPspA+PhtD formulation can be considered a promising bivalent serotype-independent vaccine candidate for protection against invasive pneumococcal infection in the future

Synthesis and characterization of a novel chemically designed (Globo)3–DTPA–KLH antigen

In recent years, many experiments have been conducted for the production and evaluation of anticancer glycoconjugated vaccines in developed countries and many achievements have been accomplished with Globo H derivatives. In the current experiment, a new chemically designed triplicate version of (Globo H)3–diethylenetriamine pentaacetic acid (DTPA)–KLH antigen was synthesized and characterized. Immunization with (Globo H)3-DTPA-KLH, a hexasaccharide that is a member of a family of antigenic carbohydrates that are highly expressed in various types of cancers conjugated with DTPA and KLH protein, induced a high level of antibody titer along with an elevated level of IL-4 in mice. Treatment of tumors with the collected sera from immunized mice decreased the tumor size in nude mice as well. None of the immunized mice illustrated any sign of tumor growth after injection of MCF-7 cells compared to the control animals. These findings, based on the newly presented structure of the Globo H antigen, lend exciting and promising evidence for clinical advancement in the development of a therapeutic vaccine in the future

In-silico design and evaluation of an epitope-based serotype-independent promising vaccine candidate for highly cross-reactive regions of pneumococcal surface protein A

Abstract Background The pathogenicity of pneumococcus with high morbidity, mortality, and multi-drug resistance patterns has been increasing. The limited coverage of the licensed polysaccharide-based vaccines and the replacement of the non-vaccine serotypes are the main reasons for producing a successful serotype-independent vaccine. Pneumococcal surface protein A (PspA) is an extremely important virulence factor and an interesting candidate for conserved protein-based pneumococcal vaccine classified into two prominent families containing five clades. PspA family-elicited immunity is clade-dependent, and the level of the PspA cross-reactivity is restricted to the same family. Methods To cover and overcome the clade-dependent immunity of the PspAs in this study, we designed and tested a PspA1-5c+p vaccine candidate composed of the highest immunodominant coverage of B- and T-cell epitope truncated domain of each clade focusing on two cross-reactive B and C regions of the PspAs. The antigenicity, toxicity, physicochemical properties, 3D structure prediction, stability and flexibility of the designed protein using molecular dynamic (MD) simulation, molecular docking of the construct withHLADRB1*(01:01) and human lactoferrin N-lop, and immune simulation were assessed using immunoinformatics tools. In the experimental section, after intraperitoneal immunization of the mice with Alum adjuvanted recombinant PspA1-5c+p, we evaluated the immune response, cross-reactivity, and functionality of the Anti-PspA1-5c+p antibody using ELISA, Opsonophagocytic killing activity, and serum bactericidal assay. Results For the first time, this work suggested a novel PspA-based vaccine candidate using immunoinformatics tools. The designed PspA1-5c+p protein is predicted to be highly antigenic, non-toxic, soluble, stable with low flexibility in MD simulation, and able to stimulate both humoral and cellular immune responses. The designed protein also could interact strongly with HLADRB1*(01:01) and human lactoferrin N-lop in the docking study. Our immunoinformatics predictions were validated using experimental data. Results showed that the anti-PspA1-5c+p IgG not only had a high titer with strong and same cross-reactivity coverage against all pneumococcal serotypes used but also had high and effective bioactivity for pneumococcal clearance using complement system and phagocytic cells. Conclusion Our findings elucidated the potential application of the PspA1-5c+p vaccine candidate as a serotype-independent pneumococcal vaccine with a strong cross-reactivity feature. Further in-vitro and in-vivo investigations against other PspA clades should be performed to confirm the full protection of the PspA1-5c+p vaccine candidate

Additional file 1 of Indirect optimization of staphylokinase expression level in dicistronic auto-inducible system

Additional file 1: Table S1. The fluorescent intensity measured in each run for indirect optimization of SAK expression

<span style="font-size:15.0pt;font-family: "Times New Roman","serif";mso-fareast-font-family:"Times New Roman";mso-bidi-font-family: Mangal;mso-ansi-language:EN-GB;mso-fareast-language:EN-US;mso-bidi-language: HI" lang="EN-GB">Identification of new <i style="mso-bidi-font-style:normal">Streptomyces griseus</i> strains with potential antimicrobial activity isolated from Caspian Sea</span>

2277-2280<span style="font-size:9.0pt;font-family: " times="" new="" roman","serif";mso-fareast-font-family:"times="" roman";mso-bidi-font-family:="" mangal;mso-ansi-language:en-gb;mso-fareast-language:en-us;mso-bidi-language:="" hi"="" lang="EN-GB">With the emergence of new antibiotic-resistant bacteria, the global announcement for new antimicrobial agents has initiated. Many scientists have been focused on identification and characterization of novel marine species which produce more potent antimicrobial agents. In this study, 162 strains were isolated from Caspian Sea and analyzed based on antimicrobial activity. Bacteria were grown in wide period of time, 24 h to 20 days, on Muller Hinton Agar medium. Among 162 isolates, 4 strains showed antimicrobial activity to Bacillus subtilis, Escherichia coli, Pseudomonas aeruginosa and Staphylococcus aureus as reference strains. But the maximum effect was observed on Bacillus subtilis and Staphylococcus aureus (Gram positive bacteria). The positive strains were subjected to 16S rDNA PCR sequencing and the results were BLASTed against the NCBI database to evaluate genetic distribution and bacterial classification. The strains were submitted to NCBI as new Streptomyces griseus strains.</span

The correlation of long non-coding RNAs IFNG-AS1 and ZEB2-AS1 with IFN-γ and ZEB-2 expression in PBMCs and clinical features of patients with coronary artery disease

Background Aberrant expression of long non-coding RNAs (lncRNAs) can contribute to the pathogenesis of coronary artery disease (CAD). In this study, we aimed to evaluate the expression of lncRNA interferon gamma-antisense 1 (IFNG-AS1), zinc finger E-box binding homeobox 2 antisense RNA 1 (ZEB2-AS1), and their direct target genes (IFN-gamma and ZEB2, respectively) in peripheral blood mononuclear cell (PBMC) from CAD and healthy individuals. Methods and results We recruited 40 CAD patients and 40 healthy individuals. After doing some bioinformatics analyses, the expressions of IFNG-AS1/ ZEB2-AS1 lncRNAs and IFN-gamma/ ZEB2 in PBMCs were measured using quantitative real-time PCR. The possible correlation between the putative lncRNAs and disease severity was also assessed. Receiver operating characteristic (ROC) curve analysis was used to evaluate the predictive role of lncRNAs as diagnostic biomarkers in CAD patients. The expressions of IFNG-AS1 lncRNA as well as IFN-gamma and ZEB2 genes were significantly reduced in CAD patients compared to healthy subjects. In contrast, the expression of ZEB2-AS1 was up-regulated in these patients. Linear regression analysis unveiled that there is a positive correlation between the expression of IFNG-AS1 and IFN-gamma, also similarly, ZEB2-AS1 and ZEB2 in PBMCs of subjects. Moreover, the expression of IFNG-AS1 and ZEB2-AS1 correlated with the Gensini score. The area under the ROC curves ranged from 0.633-0.742 for ZEB2-AS1/ZEB2 and IFNG-AS1/IFN-gamma, respectively. Conclusions Our results indicated that the dysregulation of IFNG-AS1/IFN-gamma and ZEB2-AS1/ZEB2 in PBMCs of CAD patients may be involved in CAD pathogenesis

The effect of manipulating glucuronic acid biosynthetic pathway in Bacillus subtilis strain on hyaluronic acid production

Abstract Hyaluronic acid (HA), composed of glucuronic acid (GlcUA) and N-acetyl glucoseamine (GlcNAc), is a versatile biopolymer with high commercial value and innumerous physiological roles and pharmaceutical applications. The hasA gene has main role in HA biosynthesis by Streptococcus strain as a natural producer. The hasB and hasC genes are also mediate GlcUA precursor biosynthesis. In the present study, S. equisimilis hasA gene; B. subtilis tuaD and gtaB genes for GlcUA precursors enhancement, and vgb gene coding bacterial hemoglobin as an oxygen provider were used to construct the B. subtilis strain for HA production. RBSHA (hasA), RBSHA2 (hasA/tuaD/gtaB), and RBSHA3 (hasA/tuaD/gtaB/vgb) strains were developed and confirmed through genotype and phenotype analysis. After HA production and purification, FTIR spectroscopy confirmed the produced HA structures. HA assay showed the highest HA titer for RBSHA3 (2.1 ± 0.18 mg/ml) and then RBSHA2 (1.9 ± 0.03 mg/ml), and RBSHA (0.6 ± 0.14 mg/ml). Statistical analysis indicated there is no significant difference in HA titer between RBSHA2 and RBSHA3 strains (p-value > 0.05), however, these strains produced HA approximately 4-fold higher than that of RBSHA strain. Agarose gel electrophoresis showed the same molecular weight (< 30 kDa) of produced HA by strains. Dynamic light scattering (DLS) revealed all HA polymers had a relatively low polydispersity index (PDI < 0.5). These findings demonstrate the successful GlcUA biosynthetic pathway engineering strategy in improving HA yield by recombinant B. subtilis, metabolically-robust, and industrially potential strain

Molecular Design, Expression and Evaluation of PASylated Human Recombinant Erythropoietin with Enhanced Functional Properties

Erythropoietin (EPO) is the principal hormone which, has somewhat short half-life involved in the differentiation and regulation of circulating red blood cells. The present study was carried out to evaluate the capability of a polyethylene glycol mimetic technology as a biological alternative to improve pharmaceutical properties of human recombinant EPO. In silico models of EPO fused to 200 amino acids of proline, alanine, and serine (PAS) were initially generated and assessed by molecular dynamic (MD) simulation. The fluctuations of the modeled structure reached a plateau after 6000 ps of MD simulation. The Phi and psi analysis showed \u3e99.2% of residues were located in the allowed regions. An expression vector consisting of EPO cDNA tagged to PAS coding sequences was synthesized and expressed in CHO-K1 Cells. The produced PASylated molecule was purified and characterized by standard analytical methods. The molecular weight of fusion protein was expanded to 70 kDa using sodium dodecyl sulfate polyacrylamide gel electrophoresis method. Analytical size exclusion chromatography revealed an approximately sevenfold increase in apparent size of produced protein. Although the in vitro potency of the fusion protein was significantly reduced (1.26 ± 0.05 vs. 0.24 ± 0.03 ng/ml) but, the in vivo activity was considerably increased up to 1.58 × 105 IU/ml in normocythemic mice assay. Pharmacokinetic animal studies revealed strongly 15.6-fold plasma half-life extension for the PASylated EPO (83.16 ± 13.28 h) in comparison to epoetin α (8.5 ± 2.4 h) and darbepoetin α (25.3 ± 2.2h)