Pages Experimental Methods. Imaging Methods. Back Matter Pages About this book Introduction As part of a collaboration between two different groups in chemistry and biochemistry, Thom Sharp presents here his thesis work on the development of new methods for cryoelectron microscopy. Throughout his PhD, Thom had to master a whole range of techniques including modelling, molecular biology and microscopy. Using these skills to tackle an outstanding problem, the pursuit of high-resolution structures of peptide-based materials, Thom highlights in this thesis his newly developed methods for analysing and processing this particular type of electron microscopy data.
This thesis gives the first molecular description of a de-novo designed peptide-based material. In addition, nanomaterials or nanoparticles have the advantage that they can be easily modified with different molecules and delivered to the tumor along blood vessels with immune systems [ 10 , 11 , 12 , 13 ]. In this chapter, we explain the benefits of optical imaging and the importance of nanomaterial-based imaging agents for effective cancer therapy.
Multimodality Cardiovascular Molecular Imaging, Part I
We will focus on research on using nanomaterials as optical imaging agents and their diverse applications Figure 1. Schematic illustration of a multifunctional nanocomposite. Reproduced from Ref. Various nanoparticle systems are being explored for their potential use in bioimaging for cancer diagnosis or treatment because of their unique properties, including their large surface-to-volume ratio, high biocompatibility, facile surface modification, and overall structural robustness.
In addition, they have unique optical, magnetic, and electron properties, which make them ideal candidates for signal generation and transduction in the development of sensing systems [ 5 , 6 , 7 , 8 , 12 , 14 , 15 , 16 , 17 , 18 , 19 , 20 ]. Moreover, some nano-sized materials exhibit unique physical properties, such as a proper size, surface charge, stability, shape, and hydrophilicity, which can aid their effective delivery to the desired site.
The delivery of nano-sized agents is affected by the enhanced permeability and retention EPR effect, which is a unique property of solid tumors that is related to their anatomical and pathological differences from normal tissues. This EPR effect leads to the passive accumulation of large molecules and small particles in tumor tissues due to the cut-off size of the leaky vasculature and retention with long circulation times, which is called passive targeting [ 21 , 22 , 23 , 24 , 25 , 26 ].
Main advantages of the PEGylated proteins. PEG is a shielding the protein surface from degradation agents by steric hindrance. Moreover, the increased size of the conjugate decreases the kidney clearance of the PEGylated protein. In addition, positively charged cationic nanoparticles can easily enhance endocytosis or phagocytosis for cell labeling via electrostatic interactions with the negatively charged cellular membrane.
Among bio-imaging techniques, well-tailored superparamagnetic nanocrystals are of great interest for cancer detection via magnetic resonance MR imaging due to their high sensitivity and specificity due to the nanoeffect. Fluorescence and optical imaging techniques are important tools for in vivo and cellular imaging, and they can provide vital information for cancer diagnosis and therapy in its early stages. In particular, for the fluorescence wavelength, near-infrared NIR light is preferred for tissue and in vivo imaging compared to visible light because of its minimal damage to the tissue, which allows deep tissue penetration, and low auto-fluorescence interference due to the reduced scattering of long wavelength photons [ 9 ].
Active targeting, is also called as ligand-mediated targeting, involves utilizing targeting moieties that are anchored on the surface of nanoparticles and form strong interactions with a particular cell surface marker e. A schematic illustration showing methods used for active targeting of nanoparticles. A Antibody-based targeting, B Aptamer-based targeting, and C Ligand-based targeting of nanoparticels.
Targeting moieties, such as antibodies, peptides Arg-Glyc-Asp RGD , nucleic acids aptamers , and polysaccharides hyaluronan, dextran , lead to enhanced selective delivery and uptake in the target cells, tissues, organs, or subcellular domains and minimize uptake by the RES system [ 34 , 35 , 36 , 37 , 38 , 39 , 40 , 41 , 42 , 43 , 44 , 45 , 46 , 47 , 48 , 49 , 50 , 51 , 52 , 53 ].
Imaging metabolic syndrome
Active tumor targeting is more efficient and specific than passive targeting, and can facilitate early cancer detection. In particular, active tumor-targeted imaging can quantify the target expression through molecular imaging, so it is an indispensable tool in diagnosis and disease management.
For example, for the selective detection of tumors expressing a high level of epidermal growth factor receptors EGFR , anti-EGFR antibody-modified nanoparticles are widely used as imaging agents for MR, CT, and optical imaging.
CD44 is a cell surface glycoprotein that is overexpressed in breast cancer and gastric cancer stem cells and is associated with cancer growth, migration, invasion, and angiogenesis. Hyaluronan HA , which is an immune—neutral polysaccharide, forms a specific interaction with CD Angiogenesis appears to be one of the most crucial steps in tumor translation to the metastatic form, in which it is capable of spreading to other parts of the body by degrading the basement membrane and forming a new vascular structure. Targeted-molecular imaging of vascular or angiogenesis can provide accurate anatomic details for effective cancer management.
Aptamers Apt are short nucleic acid molecules that can bind to target antigens with high affinity and specificity. The tumor microenvironment plays a critical role in tumor initiation, progression, metastasis, and resistance to therapy [ 54 , 55 , 56 , 57 , 58 , 59 , 60 , 61 ]. The microenvironment differs from that of normal tissues because of the dynamic network within normal tissues, including blood and lymphatic vessels, extracellular matrix proteins, and both enzyme and immune components. These unique characteristics lead to a matrix remodeling e.
The changes in the physiological characteristics of tumor microenvironments are consistent, regardless of the type of cancer, so it is possible to use these as a universal indicator for cancer detection. Vesicular pH plays a pivotal role in cell metabolism processes, such as proliferation and apoptosis. They fabricated polyaniline-based nanoprobes that exhibited convertible transition states according to the proton concentration as an in situ indicator of the vesicular transport pH [ 58 ]. A Schematic illustration of organic nanoindicator based on polyaniline nanoparticle for the detection of endolysosomal compartments.
Synthesis steps of nanoindicator based on polyaniline in mesosilica template when using heterometal nanoparticle IsNP as oxidant. While migrating from endosomes to lysosomes, transition state of polyaniline transferred to emeraldine salt state due to the increment of proton concentration. The tumor pH is usually more acidic than that of normal tissues due to increased aerobic glycolysis, which is called the Warburg effect tumor have a pH of 6.
This can promote tumor metastasis by generating an invasive environment for tearing down the extracellular matrix and for tissue remodeling. Core—shell MnO Mn 3 O 4 urchin-shaped nanoparticles are synthesized via an anisotropic etching process. The manganese ions released from the MnO phase in the low-pH sites within tumor cells lead to an enhanced T1 contrast image for the entire tumor mass. In addition, specific stromal cell-derived proteases, such as matrix metalloproteases MMP , matrix cysteine cathepsins, and serine proteases, are overexpressed in primary tumors.
These proteases induce the epithelial-to-mesenchymal transition EMT and promote invasion and metastasis by degrading the extracellular membrane. Molecular imaging of the activity of proteases has the potential to determine tumor malignancy, guide the development of diagnostic tools, and evaluate the efficacy of treatment Figure 5 A [ 60 ].
MMPs are the most prominent family of proteases associated with tumorigenesis. Their expression and activity are highly enhanced in many types of human cancer and are strongly implicated in advanced cancer states. Tumor microenvironment-targeted molecular imaging has the potential to provide clinically significant progress. Emerging evidence suggests that microRNAs can also function as a diagnostic biomarker for human cancers because they can act as tumor suppressor genes or oncogenes.
Imaging the intracellular distribution of specific miRNAs should provide insight into the mechanisms of metastasis and invasion. A Schematic illustration of the dual imaging process of anchored proteinase-targetable optomagentic nanoprobes with activatable fluorogenic peptides MNC-ActFP. B Schematic illustration of miRa beacon delivery system for targeted intracellular recognition of miRa based on Hyaluronic acid HA -coated nanocontainers that encapsulate the miRa beacons bHNCs.
Current imaging techniques play an important role in enabling the early detection of several diseases, including cancer, due to their ability to locate tumors and assess the tumor activity. However, these techniques are insufficient to provide reliable and accurate information at the disease site, due to their low sensitivity or limits in their spatial resolution Table 1. Computed tomography CT is useful for tumor staging but offers poor soft tissue contrast, with resulting poor sensitivity and specificity in screening. Magnetic resonance imaging MRI offers excellent contrast without ionizing radiation but has temporal and financial needs that are likely inconsistent with high-throughput screening.
Positron emission tomography PET , which has very high sensitivity, can investigate various molecular and biochemical properties but is more suitable for monitoring the response to therapy than for detecting early lesions due to its limited spatial resolution. Therefore, multimodal imaging, i. Recently, various types of hybrid nanoparticles have been used for multimodal imaging by combining the strengths of individual components into single nano-structured systems. Multimodal imaging probes enable both magnetic and optical imaging to provide great benefits for in vivo disease diagnosis and the in situ monitoring of living cells.
Uniformly sized tantalum oxide nanoparticles were synthesized using a microemulsion method and were modified using various silane derivatives, such as polyethylene glycol PEG and fluorescent dye molecules, through simple in situ sol-gel reaction. These nanoparticles exhibited remarkable performances in in vivo simultaneous fluorescence imaging as well as X-ray CT angiography and bimodal image-guided lymph node mapping [ 72 ].
Additionally, targeted multimodal imaging systems by modifying targeting moieties to increase the selective accumulation at the disease site has shown promising results. In this case, several factors should be considered, including the appropriate choice of a targeting moiety and its conjugation method.
A Illustration of simultaneously self-assembled fluorescent magnetic nanoprobes FMNPs as multimodal biomedical imaging probes. B MR images of tumor-bearing mice after injection of the FMNPs i: xenograft tumor model and ii: orthotopic bladder tumor model and C fluorescence images of their excised tumor slides, respectively. Recently, nanoparticles have provided significant progress in cancer theranostics due to their unique physicochemical properties, in which both diagnosis and therapeutic functions can be achieved simultaneously.
Theranostics aims to provide image-guided cancer therapy by integrating imaging and therapy, which are particularly interesting fields in Nanomedicine [ 12 ]. Theranostic nanoparticles comprise at least three components: i the biological payload, ii the carrier, and iii surface modifiers Figure 1 [ 74 , 75 , 76 , 77 , 78 , 79 , 80 , 81 , 82 , 83 , 84 , 85 , 86 , 87 , 88 , 89 , 90 , 91 , 92 , 93 ].
Biological payloads include imaging agents and therapeutic agents. Therefore, they can allow the simultaneous delivery of therapeutic agents to the tumor site and real-time tracking of their biodistribution in vivo. Optical imaging has advantages in theranostics because it allows the non-invasive monitoring of the progression of diseases and therapy [ 84 ].
The DOX-loaded ZnO-QD-chitosan-folate carrier exhibited highly stable QDs because of its hydrophilicity and the cationic charge of chitosan as well as a rapid drug release profile with a controlled release. They confirmed that the PTX-Cy5. Smart theranostic nanosystems that respond to environmental changes e. These nanogels demonstrated potential as excellent drug carriers, providing a high drug loading capacity for TMZ as a model anticancer drug and offering the possibility of pH-regulated drug delivery [ 93 ].
Biomolecular Imaging at High Spatial and Temporal Resolution In Vitro and In Vivo | SpringerLink
Theragnostic chitosan-based nanoparticles CNPs for cancer imaging and chemotherapy. A Conceptual description of theragnostic nanoparticles labeled with Cy5. The intensity maps on the fluorescence images display normalized photon counts and D confocal microscopy images of tumor sections from mice treated with Cy5. Green, red, and blue represent FL from miR, Cy5. In this system, the PLI acts to deliver miRNA to the site of action via the buffering effect of the imidazole residues under endosomal pH.
This systemic delivery of miRa using our NVs shows the most favorable delivery efficiency, a significant suppression of CD44 expression, and increased apoptosis in the in vivo models [ 94 ]. In optical imaging, fluorophores, such as fluorescent dyes, bioluminescent proteins and fluorescent proteins, are widely used to monitor molecular events.