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Different immunomodulation strategies have been used to manage COVID‐19 due to the complex immune‐inflammatory processes involved in the pathogenesis of this infection. Curcumin with its powerful anti‐inflammatory and antiviral properties could serve as a possible COVID‐19 therapy. In this study, a randomized, double‐blinded, placebo‐controlled trial was performed to investigate the effectiveness and safety of nano‐curcumin oral soft gels as a complementary therapy in moderate–severe COVID‐19 patients. Hydroxychloroquine (HCQ) plus sofosbuvir was routinely administered to all 42 COVID‐19 patients, who were randomly assigned to receive 140 mg of nano‐curcumin or placebo for 14 days. CT scans of the chest were taken, and blood tests were run for all patients at time points of 0, 7, and 14 days. Our results indicated that C‐reactive protein (CRP) and erythrocyte sedimentation rate (ESR) levels significantly decreased from baseline in the nano‐curcumin‐treated group on day 7. Furthermore, blood levels of D‐dimer, CRP, serum ferritin, ESR, and inflammatory cytokines including IL‐6, IL‐8, and IL‐10 decreased more significantly in the nano‐curcumin‐treated group after 14 days. Additionally, the nano‐curcumin group showed significant improvements in chest CT scores, oxygen saturation levels, and hospitalization duration. Based on our data, oral administration of nano‐curcumin may be regarded as a promising adjunct treatment for COVID‐19 patients due to its ability to speed up chest clearance and recovery. [ABSTRACT FROM AUTHOR]

Phage display is well recognized as a promising high‐throughput screening tool for the discovery of novel cancer‐targeting peptides. Here, we screened a phage display library of 7‐mer random peptides through in vitro biopanning to isolate peptide ligands binding to SW480 human colon adenocarcinoma cells. Three rounds of negative and positive selection caused a remarkable enrichment of colon cancer cell‐binding phage clones with a significant enhancement of phage recovery efficiency (about 157‐fold). A number of phage clones were picked out from the eluted phages of last selection round and sequenced. According to the results of cell binding assay and phage cell‐based ELISA, one of the isolated peptides denoted as CCBP1 (with the sequence HAMRAQP) was indicated to have the highest binding efficiency, selectivity, and specificity toward colon cancer cells with no significant binding to control cells. Peptide competitive inhibition assay revealed that binding of the phage‐displayed CCBP1 is competitively inhibited by the same free peptide, suggesting that CCBP1 specific binding to the target cell is independent of the phage context. Taken together, our findings provide support for the notion that CCBP1 binds specifically to colon cancer cells and might be a potential lead candidate for targeted delivery of imaging agents or therapeutic genes/drugs to colon tumors. [ABSTRACT FROM AUTHOR]

Chimeric antigen receptor (CAR) therapy is a method directing T lymphocytes against antigens on the surface of tumors, increasing target cell elimination. Genetic engineering enhances the capability of immune cells to detect new antigens expressed on cell surfaces. CAR T cell therapy is a significant breakthrough for treating human malignancies; however, different side effects (e.g., cytokine release syndrome) restrict its application. Improving design and using various combined receptors enhance the performance of these cells. This review discusses limitations and risk factors associated with CAR T cell therapy. We also review some alternative approaches for developing the next generation of CAR T cells. [ABSTRACT FROM AUTHOR]

Different cellular and molecular mechanisms contribute to chondrocyte and osteocyte development. Although vital roles of the mothers against decapentaplegic homolog 4 (also called 'SMAD4') have been discussed in different cancers and stem cell‐related studies, there are a few reviews summarizing the roles of this protein in the skeletal development and bone homeostasis. In order to fill this gap, we discuss the critical roles of SMAD4 in the skeletal development. To this end, we review the different signalling pathways and also how SMAD4 defines stem cell features. We also elaborate how the epigenetic factors—ie DNA methylation, histone modifications and noncoding RNAs—make a contribution to the chondrocyte and osteocyte development. To better grasp the important roles of SMAD4 in the cartilage and bone development, we also review the genotype‐phenotype correlation in animal models. This review helps us to understand the importance of the SMAD4 in the chondrocyte and bone development and the potential applications for therapeutic goals. [ABSTRACT FROM AUTHOR]

Innovative composites of biopolymers and nanomaterials have been exploited to fabricate wound dressings which show functional abilities to improve different stages of wound healing by a variety of mechanisms. In this study, a polymeric nanocomposite dressing is fabricated by electrospinning of a blend of sodium alginate (SA), poly vinyl alcohol (PVA) and graphene nanoplatelets (Gnp). The crosslinking of the nanofibers is done by thermal treatment followed by ionic bonding of the fibers. The crosslinked fibers are loaded by curcumin, a natural potent anti‐inflammatory compound, encapsulated in monomethoxy poly ethylene glycol‐oleate micelles/polymersomes (NCur). Results indicate that by incorporation of Gnp and NCur into the SA/PVA scaffold the tensile strength is not changed (~7 MPa) but the elongation to break and toughness of the scaffolds significantly increase from 11.25±2.6 and 50.56 to 35.5±5.1% and 125.9 Jm‐3, respectively. The scaffolds support the controlled release of curcumin for 24 h in vitro. Biocompatibility of the scaffolds has been confirmed by cell viability assay on mouse fibroblast cells. Overall, the findings demonstrate the potential applications of the spun fibers for wound dressing purposes. [ABSTRACT FROM AUTHOR]

Curcumin is an anti‐cancerous agent, but its low‐solubility limits its clinical use. The relationship between deregulation of miRNAs and their targets suggested that miRNAs can be interest targets of curcumin in treatment of different cancers. In this study, to overcome essential defects of the clinical usage of this golden drug, curcumin‐encapsulated polymersome nanoparticles (CPNs) have been developed, and the cytotoxicity effects were studied on MDA‐MB‐231 breast cancer cells. The expression level of miR‐182/125b and the expression pattern of some potential targets in apoptotic pathway, predicted by in silico approaches, were analyzed by RT‐qPCR in CPNs‐treated and untreated cells. Moreover, the amount of CASP9 and CASP8 proteins were determined by Western blotting. The effect of CPNs on cell migration were studied by scratch test and the level of EGFR, E‐cadherin, and beta‐catenin proteins were monitored in CPNs‐treated and untreated cells by western blotting. RT‐qPCR analysis identified the downregulation of miR‐125b and miR‐182 in CPNs‐treated cells and the upregulation of some predicted apoptotic target genes such as P53, CASP9 and BAX after 24 hours. Western blotting confirmed the effects of curcumin on the increase of cleaved CASP9 protein. Based on data from the current experiment, the migration of MDA‐MB‐231 cells was decreased after CPNs treatment. According to the results, CPNs, as suitable and compatible nanocarriers, can deliver curcumin into cancerous cells more effectively and can increase the therapeutic effects of curcumin on MDA‐MB‐231 cells partly by suppression of miR‐125b and miR‐182 as well as induction of apoptosis and inhibition of metastatic progression. [ABSTRACT FROM AUTHOR]

Curcumin as a safe traditional compound has various benefits such as anticancer activities. However, low solubility in water is a problem. Herein, curcumin encapsulated in polymersome nanoparticles (CPNs) have been developed, the physicochemical properties have been evaluated, and cytotoxicity effects on HT29 cells were evaluated by MTT assay and annexin V/PI staining. The expression of stemness markers including CD44, CD133, and CD24, as well as miRNAs (miR‐126, miR‐34a, miR‐21, miR‐155, miR‐221, and miR‐222) and some potential targets, was evaluated in CPNs‐treated and untreated cells. Physicochemical analysis confirmed the encapsulation of curcumin in polymersomes and showed a spherical shape, an appropriate mean size of 259.5±1.5 nm, the acceptable polydispersity index of ~ 0.465, and the zeta potential of (‐8.74±0.2), as well as long‐term storage of CPNs at 4°C. According to the result, CPNs with the IC50 of 14 μg/ml increased apoptosis and induced S arrest in treated cells. Flow cytometry analysis showed the decrease in cancer stemness markers. RT‐qPCR analysis identified the downregulation of miR‐21, miR‐155, and miR‐221/222, as well as upregulation of miR‐34a, miR‐126, and deregulation of some apoptotic targets such as P53, CASP9, CASP8, CASP3, BAX, and BCl‐2 in CPNs‐treated cells. As a result, CPNs can be a safe and effective complementary agent to diminish cancer stem cells and tumor recurrence in colorectal cancer therapy. [ABSTRACT FROM AUTHOR]

Leukemic cells can impact the bone marrow niche to create a tumor‐favorable microenvironment using their secreted factors. Little knowledge is available about immunosuppressive and tumor‐promoting properties of chronic myeloid leukemia derived exosomes in bone marrow stromal components. We report here that K562‐derived exosomes can affect the gene expression, cytokine secretion, nitric oxide (NO) production, and redox potential of bone marrow mesenchymal stem cells (BM‐MSCs) and macrophages. Human BM‐MSCs and mouse macrophages were treated with K562‐derived exosomes. Our results demonstrated that the expression of the genes involved in hematopoietic developmental pathways and immune responses, including C‐X‐C motif chemokine 12 (Cxcl12), Dickkopf‐related protein 1 (DKK1), wnt5a, interleukin 6 (IL‐6), transforming growth factor‐beta, and tumor necrosis factor‐alpha (TNF‐alpha), changed with respect to time and exosome concentration in BM‐MSCs. The TNF‐alpha level was higher in exosome‐treated BM‐MSCs compared with the control. Exosome treatment of BM‐MSCs led to an increased production of NO and a decreased production of reactive oxygen species (ROS) in a time‐ and concentration‐dependent manner. We have shown that K562‐derived exosomes induce overexpression of IL‐10 and TNF‐alpha and downregulation of iNOS transcript levels in macrophages. The enzyme‐linked immunosorbent assay results showed that TNF‐alpha and IL‐10 secretions increased in macrophages. Treatment of macrophages with purified exosomes led to reduced NO and ROS levels. These results suggest that K562‐derived exosomes may alter the local bone marrow niche toward a leukemia‐reinforcing microenvironment. They can modulate the inflammatory molecules (TNF‐alpha and NO) and the redox potential of BM‐MSCs and macrophages and direct the polarization of macrophages toward tumor‐associated macrophages. K562‐derived exosomes modulate the gene expression and cytokine secretion of bone marrow mesenchymal stem cells (BM‐MSC) and macrophages. These leukemic exosomes alter the redox potential and nitric oxide production capacity of BM‐MSCs and macrophages. In conclusion, K562‐derived exosomes may alter the local bone marrow niche toward an immunosuppressive and leukemia‐reinforcing microenvironment. [ABSTRACT FROM AUTHOR]

Viral vectors have a wide range of applications in biology, particularly in gene therapy. Based on their integration capacity, viral vectors are classified as either integrating or nonintegrating vectors. Although integrating vectors, such as lentivectors, have the ability to direct prolonged expression of exogenous genes, manipulation of the host genome is an inappropriate feature of these gene delivery tools. Non-integrating vectors, such as episomal replicating plasmids, can replicate and persist in host cells for long periods without any chromosomal interruption. These advantages made them good tools for gene induction purposes in gene therapy and basic studies. Due to the necessity of gene induction in stem cells for study of mammalian development and targeted differentiation, the use of integrating vectors for prolonged expression of genes of interest has been developed. Application of replicating plasmids can overcome some drawbacks associated with integrating vectors, although replication and maintenance of these plasmids can differ between cell types. Previously, it has been shown that such plasmids can be maintained in human embryonic stem cells for more than one month, but the rate of the plasmid replication during the host cell cycle has not been elucidated. In the present study, we showed that an EBV-based plasmid can replicate simultaneously with host in pluripotent and multipotent human and mouse stem cells and can be sustained for long time periods in dividing cells. [ABSTRACT FROM AUTHOR]

Aberrant expression and function of microRNAs (miRNAs) in leukemia have added a new layer of complexity to the understanding of development and progression of the disease state. However, their targeting of specific signaling pathways responsible for the maintenance and survival properties of leukemic stem cell (LSC) still remains to be further clarified. Hedgehog (Hh) signaling, a highly conserved developmental pathway, has been proven as a functional pathway for LSCs, and loss of this pathway impairs the development of BCR-ABL-induced chronic myeloid leukemia (CML) and depletes CML stem cells. Here, we revealed that upregulation of the Hh smoothened (Smo) signal transducer was associated with reduced expression of miR-326 in the CD34+ cells from a group of patients with CML at diagnosis. Additionally, overexpression of miR-326 led to downregulation of Smo, resulted in decreased cell proliferation and elevated rate of apoptosis in CML CD34+ cells. Interestingly, restoration of Smo expression levels reversed the effect of miR-326 and rescued K562 cells from the antiproliferative effects of this miRNA. Thus, Smo appears to be an essential target of miR-326 during the pathogenesis of CML. These findings lead us to suggest that downregulation of miR-326 may be a possible mechanism for unrestricted activation of Smo signal transducer of the oncogenic Hh pathway in CML; therefore, the restoration of miR-326 expression could be of benefit in eradicating CD34+ CML stem/progenitor cells that represent a potential source of relapse in patients suffering CML. [ABSTRACT FROM AUTHOR]

The Drosophila γ-glutamyl carboxylase (dγC) has substrate recognition properties similar to that of the vertebrate γ-carboxylase (γC), and its carboxylated product yield, in vitro, was shown to be more than that obtained with the human enzyme. However, whether the Drosophila enzyme is able to γ-carboxylate the human vitamin K-dependent (VKD) proteins, such as the human coagulation factor IX (hFIX), as synthesized in cultured Drosophila cells was not known. To examine this possibility, the Drosophila Schnider (S2) cell line was transfected with a metallothionein promoter-regulated hFIX-expressing plasmid. After induction with copper ion, expression efficiency of the active hFIX was analyzed by performing enzyme-linked immunosorbent assey (ELISA) and coagulation test on the culture supernatant of the transfected S2 cells during 72 h of postinduction. In comparison with Chinese hamster ovary cell line, S2 cells showed higher (∼12-fold) expression level of the hFIX. The γ-carboxylation of the Drosophila-derived hFIX was confirmed by evaluation of the expressed protein, after being precipitated with barium citrate. The biological activity of the S2 cell-derived hFIX indicated the capability of S2 cells to fulfill the required γ-carboxylation of the expressed hFIX. Coexpression of the human γ-glutamyl carboxylases (hγC) was also shown to improve both expression and γ-carboxylation of the hFIX. This is the first in vivo data to describe the ability of the dγC to recognize the human-based propeptide as substrate, which is an essential step for production of biologically active γ-carboxylated VKD proteins. © 2011 American Institute of Chemical Engineers Biotechnol. Prog., 2012 [ABSTRACT FROM AUTHOR]

Background The ability to direct efficiently and specifically carriers toward target cells and express the transgene of interest is a critical step in gene therapy trails. The display of targeting molecules on the surface of phage particles might represent a potent solution. In the present study, we evaluated a chemical coupling strategy for displaying human holotransferrin as a targeting molecule on the surface of phage lambda particles for specifically delivering green fluorescent protein (GFP) encoding gene into a human cell line. Methods Human holotransferrin was coupled on the phage lambda particles bearing a GFP-expression cassette by a chemical coupling strategy to formulate transferrin-targeted lambda-GFP (Tf-targeted-λ-GFP) gene nanocarrier. The carrier was then characterized by phage-enzyme-linked immunosorbent assay experiments and used for transfection of the human 293T cell line. Particle internalization into the cells was evaluated by immunocytochemical staining and transfection efficacy was studied using fluorescence-activated cell sorting (FACS) analysis. Results Characterization of the nanocarrier showed a rather high copy number (274 molecules) of transferrin molecules coupled per phage particle. Immunocytochemical staining revealed efficient internalization of the Tf-targeted-λ-GFP compared to wild lambda-GFP (λ-GFP) particles. FACS analysis showed 6.72% GFP positive cells for transfections mediated by Tf-targeted-λ-GFP, whereas the value was 0.61% for wild lambda-GFP particles. Conclusions Our findings highlight chemical coupling as an efficient and straightforward strategy for displaying a targeting molecule at high density on the phage surface, which, in turn, may improve the efficiency of phage-mediated gene transfer and expression. Copyright © 2011 John Wiley & Sons, Ltd. [ABSTRACT FROM AUTHOR]

Aim: Rheumatoid arthritis (RA) is the most common chronic inflammatory erosive joint disease with the worldwide distribution of approximately 0.5–1.0%. Etiology of RA is not exactly known but immunologic and genetic factors play an important role in the pathogenesis of the disease. Genetic factors such as human leukocyte antigens (HLA) are responsible for many autoimmune diseases; therefore we decided to look for a correlation between RA and the presence of HLA-DQβ1 alleles as possible genetic markers. Methods: Genomic DNA from the whole blood samples of 25 patients with RA and 86 normal individuals as control group were extracted by salting out method. The genomic DNA was amplified by polymerase chain reaction-sequence specific primer (PCR-SSP) technique. HLA-typing was done by this method after optimizing the PCR reaction for each allele. In this procedure seven serological subclasses of HLA-DQβ1 can be detected. Results: Comparing the results between the patients and controls show a significant increase in the frequency of HLA-DQ8 (*0302, *0305) alleles in RA patients. The P-values were 0.007 and the relative risk for these alleles was evaluated higher than 1. Conclusions: The results suggest that DQ8 is the dominant HLA-DQβ1 allele that is associated with susceptibility to RA in north-eastern Iran. [ABSTRACT FROM AUTHOR]

A novel amino acid supplementation strategy was developed for enhancing the production of IL-2 (interleukin-2; as a model protein) by recombinant Escherichia coli BL21 (pET21a-hil2) in fed-batch high-cell-density cultures. The amino acids most needed and their amounts were determined using a stoichiometric model, and full factorial design experiments were conducted to determine the effects of single amino acids and amino acid mixtures on production. One of the most effective amino acid mixtures was found to be leucine, aspartic acid and glycine. This amino acid mixture was utilized for the production of IL-2 in batch and fed-batch fermentations. The amount of IL-2 produced increased from 403 to 722 mg/l and from 5.15 × 10³ to 8.08 × 10³ mg/l in batch and fed-batch cultures respectively. The results also revealed that the above amino acid mixture specifically increases IL-2 concentration in the cells.