Gene Therapy for Regenerative Medicine.

The development of biological methods over the past decade has stimulated great interest in the possibility to regenerate human tissues. Advances in stem cell research, gene therapy, and tissue engineering have accelerated the technology in tissue and organ regeneration. However, despite significant progress in this area, there are still several technical issues that must be addressed, especially in the clinical use of gene therapy. The aims of gene therapy include utilising cells to produce a suitable protein, silencing over-producing proteins, and genetically modifying and repairing cell functions that may affect disease conditions. While most current gene therapy clinical trials are based on cell- and viral-mediated approaches, non-viral gene transfection agents are emerging as potentially safe and effective in the treatment of a wide variety of genetic and acquired diseases. Gene therapy based on viral vectors may induce pathogenicity and immunogenicity. Therefore, significant efforts are being invested in non-viral vectors to enhance their efficiency to a level comparable to the viral vector. Non-viral technologies consist of plasmid-based expression systems containing a gene encoding, a therapeutic protein, and synthetic gene delivery systems. One possible approach to enhance non-viral vector ability or to be an alternative to viral vectors would be to use tissue engineering technology for regenerative medicine therapy. This review provides a critical view of gene therapy with a major focus on the development of regenerative medicine technologies to control the in vivo location and function of administered genes.

Developing 3D Technology for Drug Discovery.

Developing 3D living systems will open many doors and lead to significant improvements in biological tools, drug discovery process, lead identification as well as therapeutic approaches. The miniaturization of this approach allows one to perform many more experiments than previously possible more simply. 3D in vitro technology aims to develop a set of tools that are simple, inexpensive, portable, and robust that could be commercialized and used in various fields of biomedical sciences, such as drug discovery, diagnostic tools, therapeutic approaches, and regenerative medicine.

Safety Evaluation of Nanotechnology Products.

Nanomaterials are now being used in a wide variety of biomedical applications. Medical and health-related issues, however, have raised major concerns, in view of the potential risks of these materials against tissue, cells, and/or organs and these are still poorly understood. These particles are able to interact with the body in countless ways, and they can cause unexpected and hazardous toxicities, especially at cellular levels. Therefore, undertaking in vitro and in vivo experiments is vital to establish their toxicity with natural tissues. In this review, we discuss the underlying mechanisms of nanotoxicity and provide an overview on in vitro characterizations and cytotoxicity assays, as well as in vivo studies that emphasize blood circulation and the in vivo fate of nanomaterials. Our focus is on understanding the role that the physicochemical properties of nanomaterials play in determining their toxicity.

Retraction Note: In Vitro Transfection of Plasmid DNA by Amine Derivatives of Gelatin Accompanied with Ultrasound Irradiation.

The Editor-in-Chief has retracted this article [1] at the request of the corresponding author. Figure 1B appears to be identical to Figure 1D, despite being under different experimental conditions.

Cyclopropenium Nanoparticles and Gene Transfection in Cells.

Non-viral vectors for the transfection of genetic material are at the frontier of medical science. In this article, we introduce for the first time, cyclopropenium-containing nanoparticles as a cationic carrier for gene transfection, as an alternative to the common quaternary ammonium transfection agents. Cyclopropenium-based cationic nanoparticles were prepared by crosslinking poly(ethylene imine) (PEI) with tetrachlorocyclopropene. These nanoparticles were electrostatically complexed with plasmid DNA into nanoparticles (~50 nm). Their cellular uptake into F929 mouse fibroblast cells, and their eventual expression in vitro have been described. Transfection is enhanced relative to PEI with minimal toxicity. These cyclopropenium nanoparticles possess efficient gene transfection capabilities with minimal cytotoxicity, which makes them novel and promising candidates for gene therapy.

Bone defect healing is induced by collagen sponge/polyglycolic acid.

We have evaluated the capability of a collagen/poly glycolic acid (PGA) scaffold in regeneration of a calvarial bone defects in rabbits. 4 bone critical size defects (CSD) were created in the calvarial bone of each rabbit. The following 4 treatment modalities were tested (1) a collagen/PGA scaffold (0.52% w/w); (2) the collagen/PGA scaffold (0.52% w/w) seeded with adipose-derived mesenchymal stem cells (AD-MSCs, 1 × 106 cells per each defect); (3) AD-MSCs (1 × 106 cells) no scaffold material, and (4) blank control. The rabbits were then divided into 3 random groups (of 5) and the treatment outcomes were evaluated at 4, 8 and 12 weeks. New bone formation was histologically assessed. Experimental groups were analyzed by CT scan and real-time PCR. Histological analysis of bone defects treated with collagen/PGA alone exhibited significant fibrous connective tissue formation at the 12 weeks of treatments (P ≤ 0.05). There was no significant difference between collagen/PGA alone and collagen/PGA + AD-MSCs groups. The results were confirmed by CT scan data showing healing percentages of 34.20% for the collage/PGA group alone as compared to the control group and no difference with collagen/PGA containing AD-MSCs (1 × 106 cells). RT-PCR analysis also indicated no significant differences between collagen/PGA and collagen/PGA + AD-MSC groups, although both scaffold containing groups significantly express ALP and SIO rather than groups without scaffolds. Although there was no significant difference between the scaffolds containing cells with non-cellular scaffolds, our results indicated that the Collagen/PGA scaffold itself had a significant effect on wound healing as compared to the control group. Therefore, the collagen/PGA scaffold seems to be a promising candidate for research in bone regeneration.

Technology of RNA Interference in Advanced Medicine.

BACKGROUND: RNA interference (RNAi) and related pathways involving small interfering RNAs (siRNAs), microRNAs (miRNAs), and PIWI-interacting RNAs (piRNAs) regulate processes such as antiviral defense, genome surveillance, heterochromatin formation, and gene expression in animals, plants, and fungi. Studies on RNAi have revealed a two-step mechanism: (i) Degradation of dsRNA into small interfering RNAs (siRNAs), 21 to 25 nucleotides long, by an RNase III-like activity. (ii) The siRNAs join an RNase complex, RISC (RNA-induced silencing complex), which acts on the mRNA and degrades it. OBJECTIVE: Molecular structures of Dicer, Argonaute proteins, and RNA-bound complexes have offered insights into the underlying mechanisms of RNA-silencing pathways. METHODS: Sequence specific gene silencing using small interfering RNA (siRNA) is now being evaluated as a novel therapeutic strategy. RESULTS: Recently, promising data have been obtained from clinical trials for the treatment of respiratory syncytial virus and age-related macular degeneration. The exact mechanism of the RNAi pathways is still unclear. CONCLUSION: Our review summarizes the RNAi pathways and the known functions of siRNAs, miRNAs, and piRNAs in lower and higher organisms (mostly focusing on mammals) and discusses the potential applications of RNAi.

Tissue engineering strategies for the induction of angiogenesis using biomaterials.

Angiogenesis is touted as a fundamental procedure in the regeneration and restoration of different tissues. The induction of de novo blood vessels seems to be vital to yield a successful cell transplantation rate loaded on various scaffolds. Scaffolds are natural or artificial substances that are considered as one of the means for delivering, aligning, maintaining cell connection in a favor of angiogenesis. In addition to the potential role of distinct scaffold type on vascularization, the application of some strategies such as genetic manipulation, and conjugation of pro-angiogenic factors could intensify angiogenesis potential. In the current review, we focused on the status of numerous scaffolds applicable in the field of vascular biology. Also, different strategies and priming approaches useful for the induction of pro-angiogenic signaling pathways were highlighted.

Biomolecule-Responsive Hydrogels in Medicine.

Recent advances and applications of biomolecule-responsive hydrogels, namely, glucose-responsive hydrogels, protein-responsive hydrogels, and nucleic-acid-responsive hydrogels are highlighted. However, achieving the ultimate purpose of using biomolecule-responsive hydrogels in preclinical and clinical areas is still at the very early stage and calls for more novel designing concepts and advance ideas. On the way toward the real/clinical application of biomolecule-responsive hydrogels, plenty of factors should be extensively studied and examined under both in vitro and in vivo conditions. For example, biocompatibility, biointegration, and toxicity of biomolecule-responsive hydrogels should be carefully evaluated. From the living body's point of view, biocompatibility is seriously depended on the interactions at the tissue/polymer interface. These interactions are influenced by physical nature, chemical structure, surface properties, and degradation of the materials. In addition, the developments of advanced hydrogels with tunable biological and mechanical properties which cause no/low side effects are of great importance.

Long bone mesenchymal stem cells (Lb-MSCs): clinically reliable cells for osteo-diseases.

Mesenchymal stem cells (MSCs) have been designated as the most reliable cells in clinics to treat osteo-diseases because of their versatile nature. MSCs, isolated from long bone (Lb-MSCs) are rarely reported and named as RIA-MSCs because of the reamer-irrigator-aspirator (RIA) device. The potential of these cells in the treatment of non-union bone fractures made them the ideal candidates to be studied for clinical practices. In this work, effect of cryopreservation on the proliferation and differentiation capabilities of long bone MSCs (Lb-MSCs) has been studied. For this purpose, Lb-MSCs were isolated via RIA device and characterized using flow cytometry and differentiation assays. Cells were cryopreserved for 3, 6 and 12 months and thereafter were characterized using differentiation assays and genetic markers specific for osteogenic, chondrogenic, and adipogenic potential quantitatively by qRT-PCR. Lb-MSCs were found expressing MSC characteristic markers defining their identity. The population doubling time (PDT) was about 2.5 ± 0.5 days and colonies appeared after 7-10 days. Differentiation potential and gene expression of 3, 6 and 12 months cryopreserved Lb-MSCs were unaltered. The results show that cryopreservation did not have an effect on the differentiation potential of human Lb-MSCs. Therefore, our work offers Lb-MSCs as clinically cells for treating osteo-diseases.

Transferrin-conjugated magnetic dextran-spermine nanoparticles for targeted drug transport across blood-brain barrier.

Application of many vital hydrophilic medicines have been restricted by blood-brain barrier (BBB) for treatment of brain diseases. In this study, a targeted drug delivery system based on dextran-spermine biopolymer was developed for drug transport across BBB. Drug loaded magnetic dextran-spermine nanoparticles (DS-NPs) were prepared via ionic gelation followed by transferrin (Tf) conjugation as targeting moiety. The characteristics of Tf conjugated nanoparticles (TDS-NPs) were analyzed by different methods and their cytotoxicity effects on U87MG cells were tested. The superparamagnetic characteristic of TDS-NPs was verified by vibration simple magnetometer. Capecitabine loaded TDS-NPs exhibited pH-sensitive release behavior with enhanced cytotoxicity against U87MG cells, compared to DS-NPs and free capecitabine. Prussian-blue staining and TEM-imaging showed the significant cellular uptake of TDS-NPs. Furthermore, a remarkable increase of Fe concentrations in brain was observed following their biodistribution and histological studies in vivo, after 1 and 7 days of post-injection. Enhanced drug transport across BBB and pH-triggered cellular uptake of TDS-NPs indicated that these theranostic nanocarriers are promising candidate for the brain malignance treatment. © 2017 Wiley Periodicals, Inc. J Biomed Mater Res Part A: 105A: 2851-2864, 2017.

Prospects of siRNA applications in regenerative medicine.

Small interfering RNA (siRNA) has established its reputation in the field of tissue engineering owing to its ability to silence the proteins that inhibit tissue regeneration. siRNA is capable of regulating cellular behavior during tissue regeneration processes. The concept of using siRNA technology in regenerative medicine derived from its ability to inhibit the expression of target genes involved in defective tissues and the possibility to induce the expression of tissue-inductive factors that improve the tissue regeneration process. To date, siRNA has been used as a suppressive biomolecule in different tissues, such as nervous tissue, bone, cartilage, heart, kidney, and liver. Moreover, various delivery systems have been applied in order to deliver siRNA to the target tissues. This review will provide an in-depth discussion on the development of siRNA and their delivery systems and mechanisms of action in different tissues.

In-Vitro Assessment of Magnetic Dextran-Spermine Nanoparticles for Capecitabine Delivery to Cancerous Cells.

Cationic polymeric nanoparticles have great potential for developing drug delivery systems with limited side effects for tumor medication. The goal of this research is investigation of cationic dextran-spermine polymer (DS) efficacy for improvement of hydrophilic drug delivery to negatively charged cancerous cells. Capecitabine (as a hydrophilic antineoplastic drug) was loaded into the magnetic dextran-spermine nanoparticles (DS-NPs) via ionic gelation. Design of experiments was applied to specify how the significant factors affect size, surface charge and capecitabine entrapment efficiency of the DS-NPs. Physicochemical properties, in-vitro release profile and cellular studies of the optimized DS-NPs were evaluated. The experimental results indicated that DS-NPs with favorable properties can be achieved at an optimized condition of 2 mg/mL DS and 0.75 mg/mL tri-polyphosphate (TPP) concentrations, TPP addition rate of 35 mL/min, pH 3 of DS solution and super paramagnetic iron oxide nanoparticles (SPION)/DS mass ratio of 0.5. The entrapment efficiency of capecitabine was 26.1% at optimum condition and drug release at neutral pH after 24 h and acidic pH within 3 h was 56 and 98%, respectively. The cytotoxicity assessment exhibited that capecitabine loaded DS-NPs was more toxic than corresponding free drug as control. Significant cellular uptake of capecitabine loaded DS-NPs by U87MG glioblastoma cells were proved by Prussian blue staining and TEM, qualitatively. DS-NPs are suitable candidates for delivery of the hydrophilic drugs in cancer treatment and due to positive charge of the dextran-spermine, the uptake of the hydrophilic drugs by the cancerous cells was improved.

PGA-incorporated collagen: Toward a biodegradable composite scaffold for bone-tissue engineering.

Nowadays composite scaffolds based on synthetic and natural biomaterials have got attention to increase healing of non-union bone fractures. To this end, different aspects of collagen sponge incorporated with poly(glycolic acid) (PGA) fiber were investigated in this study. Collagen solution (6.33 mg/mL) with PGA fibers (collagen/fiber ratio [w/w]: 4.22, 2.11, 1.06, 0.52) was freeze-dried, followed by dehydrothermal cross-linking to obtain collagen sponge incorporating PGA fibers. Properties of scaffold for cell viability, proliferation, and differentiation of mesenchymal stem cells (MSCs) were evaluated. Scanning electron microscopy showed that collagen sponge exhibited an interconnected pore structure with an average pore size of 190 µm, irrespective of PGA fiber incorporation. The collagen-PGA sponge was superior to the original collagen sponge in terms of the initial attachment, proliferation rate, and osteogenic differentiation of the bone marrow-MSCs (BM-MSC). The shrinkage of sponges during cell culture was significantly suppressed by fiber incorporation. Incorporation of PGA fiber is a simple and promising way to reinforce collagen sponge without impairing biocompatibility. © 2016 Wiley Periodicals, Inc. J Biomed Mater Res Part A: 104A: 2020-2028, 2016.