Springer protocols feed by Imaging/Radiology 
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This is an RSS file. You can use it to subscribe to this data in your favourite RSS reader or to display this data on your own website or blog.Imaging of Electrotransferred siRNA
SiRNA delivery to the cytoplasm can be obtained through the application of calibrated electric field pulses to a mixture of cells and oligonucleotides. To investigate the uptake pathway, time lapse confocal fluorescence microscopy provides a direct visualization of the transfer. SiRNA is electrophoretically drifted directly to the cytoplasm during the pulse. No post pulse transfer is observed. The uploaded siRNA then freely diffuse in the cytoplasm with no access to the nuclei. (Source: Springer protocols feed by Imaging/Radiology)
Source: Springer protocols feed by Imaging/Radiology - October 11, 2015 Category: Radiology Source Type: news
Indium-Labeling of siRNA for Small Animal SPECT Imaging
Ever since the discovery of RNA interference (RNAi), therapeutic delivery of siRNA has attracted a lot of interest. However, due to the nature and structure of siRNA, a carrier is needed for any mode of systemic treatment. Furthermore, specific imaging techniques are required to trace where the deposition of the siRNA occurs throughout the body after treatment. Tracking in vivo siRNA biodistribution allows understanding and interpreting therapeutics effects and side effects. A great advantage of noninvasive imaging techniques such as SPECT imaging is that several time points can be assessed in the same subject. Thus, the t...
Source: Springer protocols feed by Imaging/Radiology - October 11, 2015 Category: Radiology Source Type: news
Analyses of Tumor Burden In Vivo and Metastasis Ex Vivo Using Luciferase-Expressing Cancer Cells in an Orthotopic Mouse Model of Neuroblastoma
Cancer xenograft mouse models are useful for examining and understanding tumor growth and cancer progression in vivo. With the development of bioluminescent imaging, these parameters can now be monitored noninvasively with relative ease. Herein we describe imaging of luciferase-expressing cancer cells to quantitatively measure tumor burden in vivo and metastases ex vivo. Specifically, we detail the methodology to examine the effect of shRNA-mediated knockdown of a target gene on the growth and spread of neuroblastoma tumors in immune-deficient mice. (Source: Springer protocols feed by Imaging/Radiology)
Source: Springer protocols feed by Imaging/Radiology - October 11, 2015 Category: Radiology Source Type: news
Targeted Delivery with Imaging Assessment of siRNA Expressing Nanocassettes into Cancer
Molecular therapy using small interfering RNA (siRNA) shows great promise in the development of novel therapeutics for cancer. Although various approaches have been developed for in vivo delivery of siRNAs into tumors, stability of siRNA in blood circulation, and low efficiency of siRNA delivery into tumor cells are the major obstacles for further translation into cancer therapeutics. In this protocol, we describe methods of the production of shRNA expressing DNA nanocassettes by PCR amplification of double-stranded DNA fragments containing a U6 promoter and a shRNA gene. Those DNA nanocassettes can be conjugated to the po...
Source: Springer protocols feed by Imaging/Radiology - October 11, 2015 Category: Radiology Source Type: news
Magnetic Resonance Spectroscopy of siRNA-Based Cancer Therapy
Small interfering RNA (siRNA) is routinely used as a biological tool to silence specific genes, and is under active investigation in cancer treatment strategies. Noninvasive magnetic resonance spectroscopy (MRS) provides the ability to assess the functional effects of siRNA-mediated gene silencing in cultured cancer cells, and following nanoparticle-based delivery in tumors in vivo. Here we describe the use of siRNA to downregulate choline kinase, a critical enzyme in choline phospholipid metabolism of cancer cells and tumors, and the use of 1H MRS of cells and 1H magnetic resonance spectroscopic imaging (MRSI) of tumors t...
Source: Springer protocols feed by Imaging/Radiology - October 11, 2015 Category: Radiology Source Type: news
In Vivo Magnetic Resonance Imaging of Small Interfering RNA Nanodelivery to Pancreatic Islets
Pancreatic islet transplantation is a promising therapeutic approach for type 1 diabetes. (Source: Springer protocols feed by Imaging/Radiology)
Source: Springer protocols feed by Imaging/Radiology - October 11, 2015 Category: Radiology Source Type: news
Imaging Functional Nucleic Acid Delivery to Skin
Monogenic skin diseases arise from well-defined single gene mutations, and in some cases a single point mutation. As the target cells are superficial, these diseases are ideally suited for treatment by nucleic acid-based therapies as well as monitoring through a variety of noninvasive imaging technologies. Despite the accessibility of the skin, there remain formidable barriers for functional delivery of nucleic acids to the target cells within the dermis and epidermis. These barriers include the stratum corneum and the layered structure of the skin, as well as more locally, the cellular, endosomal and nuclear membranes. A ...
Source: Springer protocols feed by Imaging/Radiology - October 11, 2015 Category: Radiology Source Type: news
MicroRNA Imaging in Combination with Diagnostic Ultrasound and Bubble Liposomes for MicroRNA Delivery
MicroRNA (miRNA) is expected to play an important role in the diagnosis and therapy of various diseases. In miRNA therapy, the development of delivery tools to the target site is considered to be essential. By using a delivery tool possessing imaging ability, miRNA colocalized with the carrier could be visualized after administration. We prepared polyethylene glycol (PEG)-modified liposomes containing echo-contrast gas, “Bubble liposomes” (BLs), and confirmed that BLs containing cationic lipid were capable of loading miRNA. Furthermore, we also achieved the imaging and delivery of systemically injected miRNA to...
Source: Springer protocols feed by Imaging/Radiology - October 11, 2015 Category: Radiology Source Type: news
Positive Bioluminescence Imaging of MicroRNA Expression in Small Animal Models Using an Engineered Genetic-Switch Expression System, RILES
MicroRNAs (miRNAs) are a class of small, noncoding RNAs which regulate gene expression by directing their target mRNA for degradation or translational repression. Since their discovery in the early 1990s, miRNAs have emerged as key components in the posttranscriptional regulation of gene networks, shaping many biological processes from development, morphogenesis, differentiation, proliferation and apoptosis. Although understanding of the molecular basis of miRNA biology is improving, methods to monitor the dynamic and the spatiotemporal aspects of miRNA expression under physiopathological conditions are required. However, ...
Source: Springer protocols feed by Imaging/Radiology - October 11, 2015 Category: Radiology Source Type: news
Microvesicles: Isolation, Characterization for In Vitro and In Vivo Procedures
Microvesicles and exosomes are released to the extracellular milieu and are detectable in body fluids. They act as unique vehicles for cargo transfer to other cells/tissues. They contain a set of membrane receptors, intracellular proteins and nucleic acids. Here, we describe basic techniques for their isolation and characterization from cell culture media or body fluids. We also describe critical techniques for characterizing their cargo (miRNAs) and their protein content. Finally, we present labeling methods for their use in in vitro procedures and for their in vivo delivery and subsequent analysis of their cargo transfer...
Source: Springer protocols feed by Imaging/Radiology - October 11, 2015 Category: Radiology Source Type: news
Controlling RNA Expression in Cancer Using Iron Oxide Nanoparticles Detectable by MRI and In Vivo Optical Imaging
Herein, we describe a protocol for the preparation of iron oxide nanoparticle-based contrast agents and drug delivery vehicles for noninvasive cancer imaging and therapy. In the first part of the chapter we describe the details of the contrast agent synthesis, functionalization, and characterization. In the second part we describe the methods for tumor imaging using the synthesized particles with noninvasive T2-weighted magnetic resonance imaging (MRI) and in vivo near infrared optical imaging. (Source: Springer protocols feed by Imaging/Radiology)
Source: Springer protocols feed by Imaging/Radiology - October 11, 2015 Category: Radiology Source Type: news
Hypoxia-Responsive Copolymer for siRNA Delivery
A wide variety of nanomedicine has been designed for cancer therapy. Herein, we describe the synthesis and evaluation of a hypoxia-responsive copolymer for siRNA delivery (Perche et al., Angew Chem Int Ed Engl 53:3362–3366, 2014). The synthesis is achieved using established coupling chemistry and accessible purification procedures. A polyelectrolyte-lipid conjugate (polyethyleneimine 1.8 kDa-dioleyl-phosphatidylinositol, PEI-PE) and polyethylene glycol 2000 (PEG) were assembled via the hypoxia-sensitive azobenzene (Azo) unit to obtain the PEG-Azo-PEI-DOPE copolymer. This copolymer can condense siRNA and shows hypoxia...
Source: Springer protocols feed by Imaging/Radiology - October 11, 2015 Category: Radiology Source Type: news
Molecular Beacon-Based MicroRNA Imaging During Neurogenesis
The fluorescence monitoring system for examining endogenous microRNA (miRNA) activity in cellular level provides crucial information on not only understanding a critical role of miRNA involving a variety of biological processes, but also evaluating miRNA expression patterns in a noninvasive manner. In this protocol, we report the details of a new procedure for a molecular beacon-based miRNA monitoring system, which includes the illustration scheme for miRNA detection strategy, exogenous miRNA detection, and measurement of endogenous miRNA expression level during neurogenesis. The fluorescence signal of miR-124a beacon quen...
Source: Springer protocols feed by Imaging/Radiology - October 11, 2015 Category: Radiology Source Type: news
Sensing miRNA: Signal Amplification by Cognate RISC for Intracellular Detection of miRNA in Live Cells
The ability to detect miRNA expression in live cells would leave these cells available for further manipulation or culture. Here, we describe the design of a miRNA sensor oligonucleotide whose sequence mimics the target mRNA. The sensor has a fluorescent label on one end of the oligo and a quencher on the other. When inside the cell, the sensor is recognized by its cognate miRNA-RISC and gets cleaved, setting the fluorophore free from its quencher. This results in fluorescence “turn on.” Since cleavage by the RISC complex is an enzymatic process, the described approach has a very high level of sensitivity (nM)....
Source: Springer protocols feed by Imaging/Radiology - October 11, 2015 Category: Radiology Source Type: news
Optogenetics: Optical Control of a Photoactivatable Rac in Living Cells
Recent developments in optogenetics have extended optical control of signaling to intracellular proteins, including Rac, a small G protein in the Rho family. A blue light-sensing LOV (light, oxygen, or voltage) domain derived from Avena sativa (oat) phototropin was fused to the N-terminus of a constitutively active mutant of Rac, via an α-helix (Jα) that is conserved among plant phototropins. The fused LOV domain occluded binding of downstream effectors to Rac in the dark. Exposure to blue light caused a conformational change of the LOV domain and unwinding of the Jα helix, relieving steric inhibition. Th...
Source: Springer protocols feed by Imaging/Radiology - November 13, 2014 Category: Radiology Source Type: news
