Current Issue : October - December Volume : 2020 Issue Number : 4 Articles : 5 Articles
Drug delivery to the brain is highly hindered by the presence of the bloodâ??brain barrier\n(BBB), which prevents the entry of many potential drugs/biomolecules into the brain. One of the\ncurrent strategies to achieve gene therapy for neurodegenerative diseases involves direct injection of\na viral vector into the brain. There are various disadvantages of viral vectors, including limitations of\ncargo size and safety concerns. Nanomolecules, such as dendrimers, serve as an excellent alternative\nto viral delivery. In this study, as proof-of-concept, we used a surface-modified dendrimer complex\nand delivered large plasmids to cells in vitro and in vivo in healthy rats via intracranial injection.\nThe dendrimers were biodegradable by chemicals found within cells and toxicity assays revealed\nthat the modified dendrimers were much less toxic than unmodified amine-surface dendrimers.\nAs mentioned in our previous publication, these dendrimers with appropriately modified surfaces are\nsafe, can deliver large plasmids to the brain, and can overcome the cargo size limitations associated\nwith viral vectors. The biocompatibility of this dendritic nanomolecule and the ability to finely tune\nits surface chemistry provides a gene delivery system that could facilitate future in vivo cellular\nreprograming and other gene therapies....
Recent advancements in stem cell therapy have led to an increased interest within\nthe auditory community in exploring the potential of mesenchymal stem cells (MSCs) in the treatment\nof inner ear disorders. However, the biocompatibility of MSCs with the inner ear, especially when\ndelivered non-surgically and in the immunocompetent cochlea, is not completely understood. In this\nstudy, we determined the effect of intratympanic administration of rodent bone marrow MSCs\n(BM-MSCs) on the inner ear in an immunocompetent rat model. The administration of MSCs did not\nlead to the generation of any oxidative stress in the rat inner ear. There was no significant production\nof proinflammatory cytokines, tumor necrosis factor (TNF), interleukin (IL)-1, IL-6 and IL-12, due\nto BM-MSCs administration into the rat cochlea. BM-MSCs do not activate caspase 3 pathway, which\nplays a central role in sensory cell damage. Additionally, transferase dUTP nick end labeling (TUNEL)\nstaining determined that there was no significant cell death associated with the administration of\nBM-MSCs. The results of the present study suggest that trans-tympanic administration of BM-MSCs\ndoes not result in oxidative stress or inflammatory response in the immunocompetent rat cochlea....
Site-directed RNA editing (SDRE) technologies have great potential for treating genetic\ndiseases caused by point mutations. Our group and other researchers have developed SDRE methods\nutilizing adenosine deaminases acting on RNA (ADARs) and guide RNAs recruiting ADARs to target\nRNAs bearing point mutations. In general, effcient SDRE relies on introducing numerous guide\nRNAs relative to target genes. However, achieving a large ratio is not possible for gene therapy\napplications. In order to achieve a realistic ratio, we herein developed a system that can introduce\nan equal number of genes and guide RNAs into cultured cells using a fusion protein comprising\nan ADAR fragment and a plasmid vector containing one copy of each gene on a single construct.\nWe transfected the single construct into HEK293T cells and achieved relatively high effciency (up to\n42%). The results demonstrate that effcient SDRE is possible when the copy number is similar for all\nthree factors (target gene, guide RNA, and ADAR enzyme). This method is expected to be capable of\nhighly ecient gene repair in vivo, making it applicable for gene therapy....
Fibrin scaffold fits as a provisional platform promoting cell migration and proliferation,\nangiogenesis, connective tissue formation and growth factors stimulation. We evaluated a unique\nheterologous fibrin biopolymer as scaffold to mesenchymal stem cells (MSCs) to treat a critical-size\nbone defect. Femurs of 27 rats were treated with fibrin biopolymer (FBP); FBP + MSCs; and FBP\n+ MSC differentiated in bone lineage (MSC-D). Bone repair was evaluated 03, 21 and 42 days later\nby radiographic, histological and scanning electron microscopy (SEM) imaging. The FBP + MSC-D\nassociation was the most effective treatment, since newly formed Bone was more abundant and early\nmatured in just 21 days. We concluded that FBP is an excellent scaffold for MSCs and also use of\ndifferentiated cells should be encouraged in regenerative therapy researches. The FBP ability to\nmaintain viable MSCs at Bone defect site has modified inflammatory environment and accelerating\ntheir regeneration....
p53 gene (TP53) replacement therapy has shown promising results in cancer gene therapy.\nHowever, it has been hampered, mostly because of the gene delivery vector of choice. CRISPR-Cas9\ntechnology (clustered regularly interspaced short palindromic repeats/CRISPR-associated protein 9)\ncan knock out the mutated TP53 (mutTP53), but due to its large size, many viral vectors are not\nsuitable or require implemented strategies that lower the therapeutic effciency. Here, we introduced a\nbacteriophage or phage-based vector with the ability to target cancer cells and aimed to investigate the\nfeasibility of using this vector to deliver CRISPR-Cas9 transgene in human lung adenocarcinoma cells.\nFirst, we produced a tumour-targeted bacteriophage carrying a CRISPR-Cas9 transgene cassette.\nNext, we investigated any negative impact on vector titers via quantitative polymerase chain\nreaction (qPCR) and colony-forming agar plate. Last, we combined Western blot analysis and\nimmunofluorescence staining to prove cell transduction in vitro. We showed that the tumour-targeted\nbacteriophage can package a large-size vector genome, ~10 kb, containing the CRISPR-Cas9 sequence\nwithout any negative impact on the active or total number of bacteriophage particles. Then,wedetected\nexpression of the Cas9 in human lung adenocarcinoma cells in a targeted and effcient manner. Finally,\nwe proved loss of p53 protein expression when a p53 gRNA was incorporated into the CRISPR-Cas9\nphage DNA construct. These proof-of-concept findings support the use of engineered bacteriophage\nfor TP53 replacement therapy in lung cancer....
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