Engineered Recombinant Protein Production Systems Originated from Escherichia coli

Selection of a suitable host strain is an important step in recombinant production of heterologous proteins. As the prokaryotic systems and especially Escherichia coli (E. coli) cells are the most attractive host in therapeutic protein production, various derivatives of this organism have been developed to overcome the limitations exist in recombinant protein production, such as inefficient expression and folding of target proteins with eukaryotic origin. This review summarized key strategies for E. coli engineering and introduced some new and applicable engineered E. coli strains that produce more complex proteins for therapeutic and research use in the future. HIGHLIGHTS• Selection of a suitable host is an important step in recombinant protein production.• Escherichia coli cells are the most attractive host in therapeutic protein production.• Engineered strains had developed to produce active protein with suitable conformation in industrial scale.• Classical mutagenesis & genome engineering are two important strain engineering strategies

Soluble Form Production of Recombinant Human Insulin-Like Growth Factor-1 by NusA Fusion Partner in E. coli

Insulin-Like Growth Factor-1 (IGF-1) is a small peptide with 70 amino acids and 7.6 kDa molecular weight that acts as the major mediator of growth hormone. According to the previous studies, recombinant production of human IGF-1 (rhIGF-1) in E. coli has resulted in an inactive form of protein (inclusion body). There are several strategies to transform inclusion body to a soluble form. Production in the form of fusion proteins as a suitable strategy, helps researcher in recombinant production of proteins in the soluble and active form. In current study, NusA fusion protein was used to produce IGF-1 soluble form, instead of insoluble protein. In previous study, rhIGF-1 was optimally expressed in inclusion body with 1.2 g/L concentration. rhIGF1 -NusA construct was cloned and expressed in E. coli, then, cell lysate was analyze by SDS-PAGE and densitometry techniques, to assay soluble and insoluble form of rhIGF-1. Results showed that rhIGF-1 concentration in soluble phase was 0.14 g/L, indicating that about 12% of total expression of rhIGF-1 was in the soluble form through NusA-fusion protein strategy. These results confirmed that some fusion proteins like NusA could improve the solubility of recombinant proteins expressed in heterogeneous bacterial hosts. HIGHLIGHTS Fusion proteins is a suitable strategy for recombinant production of proteins in e soluble form. NusA fusion tag improves the solubility of recombinant proteins expressed in bacterial hosts.  NusA fusion protein convert IGF-1 insoluble form to soluble form in E. coli

Direct Immobilization of Coagulation Factor VIII on Au/Fe3O4 Shell/Core Magnetic Nanoparticles for Analytical Application

Protein-coated nanoparticles have diverse applications in biomedical science. The protein hydrophobic domains or surface electrostatic charge conducts adsorption of proteins to different surfaces. This property can be customized to immobilize specific molecules on solid supports for experimental screenings or purification processes. To develop highly selective affinity ligands—such as aptamers—against specific protein targets, protein-coated magnetic particles have been successfully applied. This approach could be highly efficient in affinity ligand development against coagulation factor VIII.In this study, magnetic nanoparticles were prepared by co-precipitation method and, then, a gold coating was run on the MNPs’ surface. The gold coating could add some attractive specifications to the protein immobilized nanoparticles during the aptamer selection process, such as simultaneous affinity determination of aptameric oligonucleotides by fluorescence-based methods. The gold surface has been indicated as a specific feature for covalent binding to the sulphur functional groups of various molecules. In proteins, sulphur units of cysteine or methionine might be bound covalently to the gold surface. In addition, nonspecific and non-covalent attachment of proteins to the gold particles may be performed. Therefore, a series of samples containing different mass ratios of protein to gold magnetic nanoparticles (GMNPs) were evaluated to find the best conditions for coagulation factor VIII immobilization. The results showed that the best condition for high coating efficiency was 48 h incubation at 4 ºC of protein and GMNPs with a mass ratio of 0.5% in PBS 25mM, with pH=7 as binding buffer.Highlights:Magnetic nanoparticles are the most attractive nanostructures in biomedical and bio-analytical fields.The protein coating on MNPs has been found to have wide clinical and analytical applications.Coagulation factor VIII (FVIII) is a valuable therapeutic human protein in the market.Attachment of a large protein like F VIII to GMNPs is affected by various environmental factors

Freeze-dried multiscale porous nanofibrous three dimensional scaffolds for bone regenerations

Introduction: Simulating hydrophobic-hydrophilic composite face with hierarchical porous and fibrous architectures of bone extracellular matrix (ECM) is a key aspect in bone tissue engineering. This study focused on the fabrication of new three-dimensional (3D) scaffolds containing polytetrafluoroethylene (PTFE), and polyvinyl alcohol (PVA), with and without graphene oxide (GO) nanoparticles using the chemical cross-linking and freeze-drying methods for bone tissue application. The effects of GO on physicochemical features and osteoinduction properties of the scaffolds were evaluated through an in vitro study. Methods: After synthesizing the GO nanoparticles, two types of 3D scaffolds, PTFE/PVA (PP) and PTFE/PVA/GO (PPG), were developed by cross-linking and freeze-drying methods. The physicochemical features of scaffolds were assessed and the interaction of the 3D scaffold types with human adipose mesenchymal stem cells (hADSCs) including attachment, proliferation, and differentiation to osteogenic like cells were investigated. Results: GO nanoparticles were successfully synthesized with no agglomeration. The blending of PTFE as a hydrophobic polymer with PVA polymer and GO nanoparticles (hydrophilic compartments) were successful. Two types of 3D scaffolds had nano topographical structures, good porosities, hydrophilic surfaces, thermal stabilities, good stiffness, as well as supporting the cell attachments, proliferation, and osteogenic differentiation. Notably, GO incorporating scaffolds provided a better milieu for cell behaviors. Conclusion: Novel multiscale porous nanofibrous 3D scaffolds made from PTFE/ PVA polymers with and without GO nanoparticles could be an ideal candidate for bone tissue engineering as a 3D template

HYSSOP and POLIUM could help to prevent COVID-19 in high-risk population: The results of a parallel randomized placebo-controlled field trial

243-253This study was conducted to evaluate the effect of HYSSOP (composed of Hyssopus officinalis L., Echium amoenum Fisch & C. A. Mey and Glycyrrhiza glabra L.) and POLIUM (contained Teucrium polium L., Cuscuta epithymum Murr and Cichorium intybus L.) combined distilled herbal medicines compared to placebo in the prevention of COVID-19. This is a double-blind parallel placebo-controlled field trial conducted on 751 asymptomatic individuals whose one of the family members recently had a positive RT-PCR test for COVID-19. They were divided into three groups including POLIUM, HYSSOP and placebo using random blocks with a 1:1:1 allocation ratio. Participants received daily 5 cc (under 12 years) or 10 cc (over 12 years) of allocated oral medications for 20 days. The primary outcome was the frequency of positive RT-PCR test among participants who became symptomatic. The mean age of participants was 36.6. Nineteen participants get infected by COVID-19 during the intervention; fifteen of them belonged to the placebo and four to the POLIUM group. Fisher's exact test indicated significant differences between HYSSOP and placebo (p<0.001) as well as POLIUM and placebo (p=0.009) groups in terms of COVID-19 confirmed by PCR tests. Cox regression model adjusted for confounders illustrated that the hazard of getting infection by COVID-19 in POLIUM and HYSSOP groups decreased by 66% (OR (95% CI): 0.34 (0.12 to 0.94); p=0.038) and 93% (OR (95% CI): 0.07 (0.01to 0.56); p=0.012) respectively, compared to placebo .Oral administration of HYSSOP and POLIUM with the other supportive health care could decrease the risk of getting COVID-19

Nanobodies As Novel Agents for Targeting Angiogenesis in Solid Cancers

Solid cancers are dependent on angiogenesis for sustenance. The FDA approval of Bevacizumab in 2004 inspired many scientists to develop more inhibitors of angiogenesis. Although several monoclonal antibodies (mAbs) are being administered to successfully combat various pathologies, the complexity and large size of mAbs seem to narrow the therapeutic applications. To improve the performance of cancer therapeutics, including those blocking tumor angiogenesis, attractive strategies such as miniaturization of the antibodies have been introduced. Nanobodies (Nbs), small single-domain antigen-binding antibody fragments, are becoming promising therapeutic and diagnostic proteins in oncology due to their favorable unique structural and functional properties. This review focuses on the potential and state of the art of Nbs to inhibit the angiogenic process for therapy and the use of labeled Nbs for non-invasive in vivo imaging of the tumors

Evaluation of the Factors Associated with AIDS Prevention Performance among Male Barbers Based on the Health Belief Model in Fasa

Background and Objectives: The number of patients inflicted with AIDS is on a growing trend in Iran, and one of the routes of disease transmission is the infection of sharps that are used for different people without disinfection. This study was designed to evaluate the performance of male barbers regarding AIDS prevention based on the Health Belief Model in Fasa, Iran. Materials and Methods: This cross-sectional study was conducted in 2014 on 220 male barbers working in Fasa, Iran. The participants were chosen through random multi-stage sampling. The data were collected using a researcher-made questionnaire including demographic data, knowledge, and constructs of the Health Belief Model using self-report. To analyze the data, linear regression was run in SPSS, version 18. Results: Our findings showed that among the participants, knowledge was good, perceived sensitivity was moderate, perceived severity was low, perceived benefits were high, perceived barriers were low, and practice was at a good level. Knowledge, perceived sensitivity, and perceived benefits constructs were predictors of barbers' performance in preventing AIDS. In general, 31.5% of variance predicted performance variables. Conclusions: Regarding the results of this study, by appropriate planning for raising awareness, perceived sensitivity, and perceived benefits regarding AIDS, barbers’ performance can be enhances, which is essential for preventing and controlling AIDS

An Update on Phytochemicals in Molecular Target Therapy of Cancer: Potential Inhibitory Effect on Telomerase Activity

Telomerase is a ribonucleoprotein enzyme, which has a significant role in synthesizing DNA telomeric in eukaryotes. Telomere maintenance can cause to immortalization and malignant transformation of human cells and thereby telomerase activity must be scrutinized as an important factor in most tumor cells. The proliferation of cancer cells or apoptosis induction can be suppressed by telomerase inhibition using different therapeutic agents without any side effects upon normal cells. Natural substances, with anti-tumor effects, such as those derived from plants can be suitable candidates due to their capabilities in preventing some side effects and resistance of tumors with respect to most chemotherapeutic drugs. In this regards, many studies have shown that natural phytochemicals have inhibitory effects on telomerase activity through affecting its subunits and components. Therefore, the aim of this paper is to review the recent studies on these kinds of phytochemicals in terms of property and mechanism. Moreover, strategies for improving the therapeutic efficacy of plant-derived substances such as combination therapy and nanoformulation based approaches are included

The Comparison between the Osteogenic Differentiation Potential of Clay-Polyacrylonitrile Nanocomposite Scaffold and Graphene-Polyacrylonitrile Scaffold in Human Mesenchymal Stem Cells

Nowadays, bone repair by means of stem cells potential is considered as a new approach in regenerative medicine. Adipose-derived mesenchymal stem cells (AD-MSCs) have been investigated as a plentiful cell source with the ability of osteogenic differentiation which can play an important role in bone tissue engineering applications. Discovering proper elements in combination of scaffolds structure to stimulate osteogenesis in adipose-derived stem cells is one of the major concerns in this issue. Porous polymeric scaffolds such as polyacrylonitrile (PAN) and susceptible nanoparticles have attracted a lot of attention recently due to biodegradability and differentiation potential respectively. In the present study, clay-PAN nanocomposite (CPN) and graphene-PAN scaffold have been electrospuned separately and evaluated from the point of the osteogenic potential in AD-MSCs. The objective of this study was to determine the effect of clay and graphene nanoparticles with PAN nanofibers on the fate of viability and osteogenesis of AD-MSCs. First, isolated mesenchymal cells were characterized by flow cytometry. After cell culture on the surface of scaffolds MTT assay, scanning electron microscope (SEM) and DAPI staining were done. The scaffolds were characterized and osteogenic differentiation potential of AD-MSCs has been investigated. The results have indicated that alkaline phosphatase (ALP) activity, calcium content and collagen expression of cells which cultured on clay-PAN nanofibers were higher than cells which cultured on graphene-PAN scaffold. Taken together, these results suggest that porous nanofiber clay-PAN scaffold can enhance the osteogenic differentiation of AD-MSCs, and can be used as a new biodegradable scaffold for bone tissue engineering applications