ABSTRACT
Chronic wounds have become a growing concern as they can have a profound impact on individuals, potentially resulting in mortality. It is crucial to prevent and manage bacterial infections, particularly drug-resistant ones. Antimicrobial peptides, such as LL-37, can firmly eliminate pathogens. Additionally, the process of angiogenesis, facilitated by growth factors like VEGF, is essential for tissue repair and wound healing. To enhance the stability and bioavailability of therapeutic agents, targeted delivery strategies utilizing Chitosan-based carriers have been employed. Electrospun biopolymers in advanced wound dressings have revolutionized wound care by providing a more effective and efficient solution for promoting tissue regeneration and speeding up the healing process. The present investigation utilized Chitosan nanoparticles to encapsulate the recombinant LL37 peptide and VEGF. An in-depth investigation was carried out to analyze the biophysical and morphological traits of the LL37-CSNPs and VEGF-CSNPs. The first support layer consisted of PCL electrospun nanofiber, followed by the electrospinning of PVA/CsLL37, PVA/CsVEGF, and PVA/CsLL37/CsVEGF onto the PCL layer. An in vitro examination assessed the fabricated nanofibers' morphological, mechanical, and biological characteristics. The antimicrobial effects were tested on methicillin-resistant Staphylococcus aureus (MRSA). The in vivo experiments assessed the antibacterial and wound-healing capabilities of the nanofibers. The findings validated the continuous release of LL37 and VEGF. The composite material PCL/PVA/CsLL37/CsVEGF demonstrated potent bactericidal and antioxidant characteristics. The cytotoxic assay demonstrated the biocompatibility of the fabricated nano mats and their potential to accelerate fibroblast cell proliferation. The efficacy of PVA/CsLL37/CsVEGF in promoting wound healing was confirmed through an in vivo wound healing assay. Furthermore, the histological analysis provided evidence of faster epidermal formation and improved antibacterial activity in wounds covered with PVA/CsLL37/CsVEGF. Adding LL37 and VEGF to the composite material improves the immune response and promotes blood vessel formation, accelerating wound healing and decreasing inflammation.
ABSTRACT
BACKGROUND: Vascular endothelial growth factor-A (VEGF-A), an endothelial cell-specific mitogen produced by various cell types, plays important roles in cell differentiation and proliferation. In this study we investigated the effect of recombinant VEGF-A on differentiation of mesenchymal stem cells (MSCs) to endothelial cells (ECs). METHODS: VEGF-A was expressed in E. coli BL21 (DE3) and BL21 pLysS competent cells with the pET32a expression vector. Recombinant VEGF-A protein expression was verified by SDS-PAGE and western blotting. Mesenchymal stem cell differentiation to ECs in the presence of VEGF-A was evaluated by flow cytometry and fluorescence microscopy. RESULTS: Recombinant VEGF-A was produced in E. coli BL21 (DE3) cells at 0.8 mg/mL concentration. Expression of CD31 and CD 144 was significantly greater, while expression of CD90, CD73, and CD44 was significantly less, in MSCs treated with our recombinant VEGF-A than in those treated with the commercial protein (p < 0.05). CONCLUSION: Recombinant VEGF-A expressed in a prokaryotic system can induce MSCs differentiation to ECs and can be used in research and likely therapeutic applications.
ABSTRACT
OBJECTIVES: Vascular endothelial growth factor (VEGF) is one of the most effective proteins in angiogenesis, mesenchymal stem cells (MSCs) differentiation and wound healing. These abilities are therapeutic potential of VEGF in diabetic retinopathy, nephropathy and other tissue damage circumstances. In this study, recombinant VEGF was produced in Escherichia coli (E. coli) system and then biological activity of this protein was evaluated in animal wound healing. MATERIALS AND METHODS: E. coli BL21 (DE3) competent cells were transformed with pET32a-VEGF clone and induced by isopropyl-ß-D-thio-galactoside (IPTG). The recombinant protein was purified by affinity chromatography. Recombinant VEGF-A-based ointment (VEGF/Vaseline 0.8 mg/100 w/w) was used for external wound (25×15mm thickness) healing in animal model. In vivo activity of ointment was evaluated by clinical evidences and cytological microscopic assessment. RESULTS: The recombinant protein with molecular weight of 45 kilodaltons (kDa) and concentration of 0.8 mg/ml was produced. Immunoblotting data showed that the antigenic region of VEGF can be expressed in E. coli and the recombinant protein has similar epitopes with close antigenic properties to the natural form. Macroscopic findings and microscopic data showed that the recombinant VEGF-A ointment was effective on excisional wound healing. CONCLUSION: Recombinant VEGF-A produced by pET32a in E. coli, possesses acceptable structure and has wound healing capability.
ABSTRACT
OBJECTIVE: To evaluate the viral antibodies in new Iranian multiple sclerosis (MS) patients. METHODS: In a cross-sectional study, sera from 61 MS patients and 60 healthy individuals were collected from January 2009 to March 2010 in the Immunology Department of Arak University of Medical Sciences, Arak, Iran, and examined for the presence of the anti-Epstein-Barr virus (EBV), human herpes virus 6 (HHV-6), measles, mumps, and para-influenza viruses IgG and IgM using an enzyme-linked immunosorbent assay or immunofluorescence. RESULTS: There were significant differences between the MS patients and the healthy individuals (controls) in the seroprevalence of anti-HHV-6 IgM (odds ratio [OR]=4.3, 95% confidence interval [CI]=2-9.3, p=0.001); anti-HHV-6 IgG (OR=2, 95% CI=1-4, p=0.04); anti-measles IgM (OR=3.2, 95% CI=1.5-6.9, p=0.002); and the anti-mumps IgM (OR=4.1, 95% CI=1.9-8.8, p=0.0001) and IgG (OR=9.5, 95% CI=3-29.6, p=0.0001). Almost all MS patients and the control individuals were negative to EBV and parainfluenza IgM. CONCLUSION: These results confirm an association between the incidence of MS and the antibodies to HHV-6 and the measles and mumps viruses, and show induction of a primary immune response (IgM), or virus reactivation, in MS patients. These viruses may have an important role in development of MS as an initial trigger in this geographical area.
