A novel nanoliposome model platform mimicking SARS-CoV-2 as a bioreceptor to dissect the amperometric response in biosensor applications.

In this study, we proposed to investigate the response of an electrochemical-based immunosensor via nanoliposomes carrying the SARS-CoV-2 Spike-S1 protein. In this regard, we prepared RNA encapsulated nanoliposome functionalized with a specific SARS-CoV-2 Spike-S1 protein as a SARS-CoV-2 model. Then, this new nanoliposome mimicking SARS-CoV-2 was used as the bio-recognizing agent of an immunosensor developed to detect the SARS-CoV-2 within the scope of the study. The working electrode of the immunosensor was coated with chitosan polymer, decorated with SARS-CoV-2 Spike antibody, to achieve antibody-antigen matching on the electrode surface. SARS-CoV-2 mimicking nanoliposomes at various concentrations was used to achieve an amperometric response and the analytical parameters of the sensor were calculated from the relationship between the immunosensor's current values depending on the number of these matches with regard to varying antigen concentrations. Linear measurement range, LOD and measurement sensitivity were calculated as 53 pM-8 nM, 3.79 pM and 55.47 µA nM-1 cm-2, respectively. The standard deviation of the same measurements in the developed immunosensor was 0.33 %.

Collagen-infilled 3D printed scaffolds loaded with miR-148b-transfected bone marrow stem cells improve calvarial bone regeneration in rats.

Differentiation of progenitors in a controlled environment improves the repair of critical-sized calvarial bone defects; however, integrating micro RNA (miRNA) therapy with 3D printed scaffolds still remains a challenge for craniofacial reconstruction. In this study, we aimed to engineer three-dimensional (3D) printed hybrid scaffolds as a new ex situ miR-148b expressing delivery system for osteogenic induction of rat bone marrow stem cells (rBMSCs) in vitro, and also in vivo in critical-sized rat calvarial defects. miR-148b-transfected rBMSCs underwent early differentiation in collagen-infilled 3D printed hybrid scaffolds, expressing significant levels of osteogenic markers compared to non-transfected rBMSCs, as confirmed by gene expression and immunohistochemical staining. Furthermore, after eight weeks of implantation, micro-computed tomography, histology and immunohistochemical staining results indicated that scaffolds loaded with miR-148b-transfected rBMSCs improved bone regeneration considerably compared to the scaffolds loaded with non-transfected rBMSCs and facilitated near-complete repair of critical-sized calvarial defects. In conclusion, our results demonstrate that collagen-infilled 3D printed scaffolds serve as an effective system for miRNA transfected progenitor cells, which has a promising potential for stimulating osteogenesis and calvarial bone repair.

In vitro chondrogenesis by BMP6 gene therapy.

In this study, the promotion of in vitro chondrogenesis was investigated by using chitosan scaffolds and rat bone marrow-derived mesenchymal stem cells (rBMSCs) which are transfected by BMP6 (bone morphogenetic protein-6) encoding gene. For this purpose, plasmid DNA (pShuttle-rBMP6), the expression vector consisting of the coding sequence of the BMP6 was obtained, and then, it was entrapped in chitosan scaffolds to obtain a gene-activated matrix (GAM). The chitosan scaffolds performed the controlled and sustained release of plasmid DNA, thus they continuously provided the modification of rBMSCs to induce chondrogenic differentiation. In addition, the cells were transfected by lipid-based agent (Lipofectamine) and then, these modified cells were inoculated into the chitosan scaffolds. Furthermore, a group of chitosan scaffolds with nontransfected rBMSCs with recombinant BMP6 free in culture medium was used as control. Comparative results showed that, mitochondrial activities of modified rBMSCs by Lipofectamine and chitosan GAM were significantly higher than those of nontransfected rBMSCs. The observations from scanning electron microscopy analysis confirmed that BMP6 gene-modified rBMSCs differentiated to the chondrogenic phenotype. Highest amount of glycosaminoglycan contents of rBMSCs on GAM concluded that BMP6 gene-activated chitosan scaffold has a potential in the application of cartilage regeneration.

Herceptin-decorated salinomycin-loaded nanoparticles for breast tumor targeting.

The use of conventional chemotherapeutic drugs was emerged as challenging for breast cancer therapy, because breast cancer stem cells cannot be destroyed due to their great nature of drug resistance. In this study, a novel nanoparticulate system of Herceptin (HER)-immobilized salinomycin (SAL)-encapsulated poly(lactic-co-glycolic acid) (PLGA) (HER-SAL-PLGA) nanoparticles were constructed and investigated for breast cancer targeting. SAL-encapsulated PLGA nanoparticles were characterized for their particle size, morphology, structural and thermal properties, and drug-encapsulation efficiency. HER-SAL-PLGA nanoparticles were characterized via particle size, surface chemistry, and herceptin-immobilization efficiency. In vitro release studies were performed for both nontargeting and targeting SAL-PLGA nanoparticles, which demonstrated a controlled release of SAL from nanoparticles. Cellular uptake of the HER-SAL-PLGA nanoparticles was assessed by fluorescence and optical microscopy and flow cytometry, which showed that the HER-SAL-PLGA nanoparticles were successfully uptaken by MCF7 cells. In conclusion, this novel drug-delivery system, HER-SAL-PLGA, was suggested as a promising targeting system for breast cancer therapy.

Chitosan scaffolds with BMP-6 loaded alginate microspheres for periodontal tissue engineering.

The aim of this study is to develop an effective growth factor releasing scaffold-microsphere system for promoting periodontal tissue engineering. Bone morphogenetic protein-6 (BMP-6)-loaded alginate microspheres in narrow size distribution were produced by optimising electrospraying conditions. The addition of these microspheres to chitosan gels produced a novel scaffold in which not only the pore sizes and interconnectivity were preserved, but also a controlled release vehicle was generated. Loading capacity was adjusted as 50 ng or 100 ng BMP-6 for each scaffold and the controlled release behaviour of BMP-6 from chitosan scaffolds was observed during seven days. Cell culture studies were carried out with rat mesenchymal stem cells derived from bone marrow in three groups; chitosan scaffolds, chitosan scaffolds containing BMP-6-loaded alginate microspheres and chitosan scaffolds with free BMP-6 in culture medium. Results showed that controlled delivery of BMP-6 from alginate microspheres has a significant effect on osteogenic differentiation.

A novel dermal substitute based on biofunctionalized electrospun PCL nanofibrous matrix.

In this study, nanofibrous matrices of polycaprolactone (PCL) and PCL/collagen with immobilized epidermal growth factor (EGF) were successfully fabricated by electrospinning for the purpose of damaged skin regeneration. Nanofiber diameters were found to be 284 ± 48 nm for PCL and 330 ± 104 nm for PCL/collagen matrices. The porosities were calculated as 85% for PCL and 90% for PCL/collagen matrices. The covalent immobilization of EGF onto the nanofibrous matrices was verified by the increase of surface atomic nitrogen ratio from 1.0 to 2.4% for PCL and from 3.7 to 4.7% for PCL/collagen. Moreover, EGF immobilization efficiencies of PCL and PCL/collagen matrices were determined as 98.5 and 99.2%, respectively. Human dermal keratinocytes (HS2) were cultivated on both neat and EGF immobilized PCL and PCL/collagen matrices to investigate the effects of matrix chemical composition and presence of EGF on cell proliferation and differentiation. EGF immobilized PCL/collagen matrices exerted early cell spreading and rapid proliferation. Statistically high expression levels of loricrin in HS2 cells cultivated on EGF immobilized PCL/collagen matrices were (p < 0.001) regarding superior differentiation ability of these cells compared to HS2 cells cultured on neat PCL and PCL/collagen matrices. In conclusion, this novel EGF immobilized PCL/collagen nanofibrous matrix could potentially be considered as an alternative dermal substitutes and wound healing material for skin tissue engineering applications.