Chapter 5: Summary and Future prospects
2.5 Silk based scaffolds in Tissue Engineering
2.6.2 Nanotools for Neuro-imaging
Visualizing the mind at work and acquiring knowledge about its functioning has always been the ultimate aim of neurologists. However, the anatomical and physiological barriers always remained a major challenge. Nanotechnology has had a profound impact on improving the diagnosis of neurological disorders by enhancing the specificity and clarity of neuro-imaging techniques.
2.6.2.1 Magnetic nanoparticles in Neuro-imaging
Liposomes and iron-oxide nanoparticles are the most used nanomaterials in MRI because of their ability to cross the BBB and tunable magnetic properties respectively. Like most other biomedical innovations, the use of super-paramagnetic nanoparticles as contrast agents for neuro-MRI was also inspired from natural physiology itself. It was back in the late 80s and early 90s that the role of iron imbalance in neurodegenerative diseases was observed. In the body, iron remains in nano-form as ferritin comprising of a 1nm thick protein coating called apoferritin and a 25nm iron oxide core (Theil et al, 1987). A separate investigation revealed that Parkinson’s patients exhibited an increased ferritin level corresponding to enhanced MR signal in the substantia nigra region (Gorell et al, 1995). Although ultra small iron oxide particles (USPIO) were already used in MR studies of liver, kidney and respiratory diseases, the correlation of ferritin levels with Parkinson’s lead to exponential increase in research activity involving application of USPIO in neuro- MRI. These USPIOs possessed long circulating life and were found to accumulate at the margins of brain tumor leading to improved delineation of tumors in MR images (Enochs et al, 1999). However, their limited ability to cross the BBB as well as
toxicity concerns was a major impediment towards the use of USPIOs in therapeutic applications and intracellular imaging in the brain. Once again, the solution lay with nature herself. Taking cue from viruses like HIV which can easily cross the BBB, a Harvard group conjugated USPIOs with HIV derived Tat peptide (responsible for cellular uptake of HIV). They further coated the nanoparticles with dextran thereby making them more biocompatible (Josephson et al, 1999). The high degree of cellular uptake by lymphocytes encouraged them to develop USPIO based multifunctional systems. They further conjugated the dextran coated USPIO-Tat complex with near-infrared fluorescent probes (NIRF) like Cy5.5 rendering a multimodal aspect to the system. Such a system could be used in preoperative MRI to delineate tumor margins as well as in intra-operative imaging of brain tumors (Kircher et al, 2003). The enhanced cellular uptake of surface functionalized iron- oxide nanoparticles encouraged studies using USPIOs as vehicles for drug delivery to brain tumors (Chertok et al, 2008). Further, glioma specific USPIOs were also designed by conjugation with Chlorotoxin, a peptide having high affinity and specificity for matrix-metalloproteinase (MMP-2) expressing tumors of neuro- ectodermal origin (Sun et al, 2008). Conjugating such target specific iron-oxide nanoparticles with NIRF probe Cy5.5 enabled MRI as well as intra-operative pathology detection at cellular level (Veiseh et al, 2005). A Chinese team designed and reported similar glioma specific super-paramagnetic iron oxide nanoparticles (SPIONs) by conjugating with Transferrin which can bind with the Transferrin receptors over-expressed in glioma cells. MR imaging even 2 days post administration of SPIONs revealed significant contrast enhancement on T2-weighed images of glioma as well as Prussian blue staining of tumor indicated enhanced retention of nanoparticles inside the tumor cell cytoplasm (Jiang et al, 2012).
Enhanced permeability and retention (EPR) of such magnetic nanoparticles were obtained by temporary disruption of BBB by focused ultrasound prior to injecting the nanoparticles. The technique was found to be useful in delivering macromolecular chemotherapeutic agents inside the CNS (Liu et al, 2010). The advent of several commercial varieties of SPIOs like Feridex (Ferimoxides by AMAG Pharma), Resovist (Ferucarbotran by Bayer Healthcare), Combidex (Ferumoxtran-10 by AMAG Pharma) and Clariscan (PEGylated SPIO suspended in glucose) led to their comparison with conventional gadolinium based MRI contrast enhancers.
Gadolinium based agents suffer low circulation time and fast diffusion from cells with
low target tissue specificity and certain side effects (Jiang et al, 2012). Studies comparing commercial SPION - based contrast enhancers and gadolinium reported identical efficiency of both as MRI contrast enhancers. Notably, the SPIONs were found to accumulate along the periphery of tumor and hence were primarily effective in determining the tumor margins (Varallyay et al, 2002; Manninger et al, 2005). A separate study by the same group with Ferumoxtran-10 (Combidex), a dextran coated SPION, showed TI and T2 MR signals even in very low magnetic field of up to 0.15 Tesla, which did not enhance with gadolinium. Further, Ferumoxtran derived T1 signal enhancement was persistent for longer duration (2-5 days) which could be advantageous for postoperative imaging (Neuwelt et al, 2004). Although many SPION based commercial contrast enhancers were approved in the past, most were banned or got discontinued at a later stage. Hence, presently, gadolinium chelate based complexes remain the preferred MRI contrast enhancers. Recent reports of application of SPIOs in monitoring cerebral blood volume (CBV) have been encouraging. Due to their long half-life and enhanced sensitivity, USPIO nanoparticles can play an important role in estimating CBV through CBV weighted functional MRI (fMRI). The CBV being a critical indicator of tissue viability and vascular activity, fMRI involving USPIOs have been reported to provide enhanced spatial specificity and better monitoring of neurological changes induced upon functional activity and pharmacological interventions (Kim et al, 2013).
2.6.2.2 Liposomes in Neuro-imaging
Liposomes are microscopic artificial phospholipids which have been recognized as promising vehicles for delivery of therapeutic and diagnostic agents to various tissues in the body. Moreover, the ability of such colloidal particles to cross the BBB and accumulate in the CNS along with their easily tunable surface properties makes liposomes an attractive vehicle for neuro-diagnostic applications. It was back in the early 1990s, that a group of radiologists from Connecticut Health Centre, USA experimented with lipid coated microbubbles (air filled bubbles which were efficient reflectors of sound) as contrast agents for neurosonography. They observed that the lipid coated microbubbles were stable for over 6 months in vitro with an in vivo half- life of 20 hours (Simon et al, 1990). Upon direct injection of these microbubbles as contrast agents into brain parenchyma it was possible to detect growing lesions in
Fig 2.15: Lipoproteins as contrast agents. A) Schematic depiction of lipoprotein structure.
B) The ways in which lipoproteins may be modified to act as contrast agents. (Adapted with