Answer To: סמינריון בוגר- הנחיות למורים ולתלמידים:FROM THE UNIVERCITY Graduate seminar - instructions for...
Dr Insiyah R. answered on Dec 19 2022
Introduction 2
Method 2
Results 2
Discussion 3
Limitations 4
Conclusion 4
Reference 6
Introduction
The process of genetically altering the cells of a patient in order to enhance their health is known as gene therapy. An innovative approach to treating both inherited and acquired disorders involves inserting therapeutic gene information and its associated regulatory mechanisms into the nucleus. This allows for the correction of mutation-induced loss of function or the expression of the defective gene product at physiological levels (Foldvari et a,2016). It is well-known that almost all human illnesses are brought on by a lack of one gene or a mutated combination of genes. There are two approaches to carrying out CNS gene transfer in living creatures: in vivo, where the gene-vector conjugate is immediately injected further into the animal; and ex vivo, in which the desired gene is first transfected into neuronal cell culture before being reintroduced into the animal (Ramamoorth & Narvekar,2015). The scientific community has speculated for more than 50 years that treatments based on the transmission of genetic elements would be a desirable choice to combat human illnesses. Theoretically, this approach—so-called gene therapy—would provide the opportunity to achieve a long-lasting and curative therapeutic effect. This method is often used in clinical trials, and some have only recently been given the status of authorised medications in the United States and Europe (Al Qtaish et al,2022). To provide a more accurate and translational milieu for an in vivo model, further gene delivery studies using nano-duplexes at a 5/1 lipid/DNA ratio in primary CNS cells from cerebral and retinal sources have been conducted. The immunocytochemistry detected the GFP signal in both primary cell cultures, demonstrating the ability of nano-duplexes to transfer genetic material to CNS cells successfully. Additionally, it is likely that NDs, rather than other carbon-based nanomaterials, would have a better correlation with biological systems, making them a promising contender for biomedical applications (Al Qtaish et al,2022). The discovery that NDs play a growing function in niosomes for gene delivery applications is significant since CNS illnesses represent a field where the creation of novel treatment approaches represents a critical need. The non-viral techniques for in vivo gene transfer in the CNS will be the main emphasis of this review. This assignment will explain the non-viral gene therapy in the CNS in detail.
Method
In order to achieve the precise tomographic tilt series alignment needed, a 1 mg/mL sample was diluted to 0.5 mg/mL using bovine serum albumin (BSA) gold nanoparticles (10 nm). Following vortex shaking, 3 L of the sample was placed to the Cu/ Rh R2/2 Quantifoil grid and vitrified at 22 °C with 95% humidity using a ThermoFisher Scientific Vitrobot Mark IV (Al Qtaish et al,2022). Utilising ThermoFisher Scientific's Tomography programme (step 3°, "65° at 28.000," with a pixel size of 1.44 nm/pix), tilted series were acquired to an actual figure dose of 130 e- 2 utilising the dose-symmetric tilt method. Utilising IMOD, the tilt series synchronisation was carried out, evaluated, and rendered in three dimensions using TOMO3D and SIRT reconstruction. Rebuilt volumes for USFC Chimera (Al Qtaish et al,2022).
Results
Compared to nioplexes, nanodiaplexes had greater levels of endocytosis via clathrins (p 0.05), higher lysosomal colocalisation (p 0.05), and endosomal escape characteristics. However, endocytosis mediated by caveolae was the most effective mechanism in the case of nanodiaplexes (Al Qtaish et al,2022). Nanodiaplexes successfully transfected retinal and neuronal cells, according to investigations in CNS primary cells...