Uptake of surface-functionalized thermo-responsive polymeric nanocarriers in corticospinal tract motor neurons.

Nanocarrier drug delivery systems are attractive options for targeted delivery of survival- and regeneration-enhancing therapeutics to neurons damaged by degenerative or traumatic central nervous system (CNS) lesions. Functional groups on nanocarrier surfaces allow derivatization with molecules to target specific cells but may affect cellular interactions and nanocarrier uptake. We synthesized differently sized -COOH and -NH2 surface functionalized polymeric nanocarriers (SFNCs) by emulsion copolymerization and assessed uptake by different cell types in mixed cortical cultures. Following 60-min incubation with SFNCs, mean intensity measurements of fluorescently labeled SFNCs indicated that corticospinal tract motor neurons (CSMNs) took up more COOH- or NH2- functionalized SFNCs with similar sizes (150 nm), compared to glia. However, larger diameter (750 nm) SFNCs were taken up at higher concentrations compared to smaller COOH-derivatized SFNCs (150 nm). These data suggest that larger SFNCs may provide an advantage for enhanced uptake by targeted neurons.

Enhanced proliferation of PC12 neural cells on untreated, nanotextured glass coverslips.

Traumatic injury to the central nervous system is a significant health problem. There is no effective treatment available partly because of the complexity of the system. Implementation of multifunctional micro- and nano-device based combinatorial therapeutics can provide biocompatible and tunable approaches to perform on-demand release of specific drugs. This can help the damaged cells to improve neuronal survival, regeneration of axons, and their reconnection to appropriate targets. Nano-topological features induced rapid cell growth is especially important towards the design of effective platforms to facilitate damaged neural circuit reconstruction. In this study, for the first time, feasibility of neuron-like PC12 cell growth on untreated and easy to prepare nanotextured surfaces has been carried out. The PC12 neuron-like cells were cultured on micro reactive ion etched  nanotextured glass coverslips. The effect of nanotextured topology as physical cue for the growth of PC12 cells was observed exclusively, eliminating the possible influence(s) of the enhanced concentration of coated materials on the surface. The cell density was observed to increase by almost 200% on nanotextured coverslips compared to plain coverslips. The morphology study indicated that PC12 cell attachment and growth on the nanotextured substrates did not launch any apoptotic machinery of the cell. Less than 5% cells deformed and depicted condensed nuclei with apoptotic bodies on nanotextured surfaces which is typical for the normal cell handling and culture. Enhanced PC12 cell proliferation by such novel and easy to prepare substrates is not only attractive for neurite outgrowth and guidance, but may be used to increase the affinity of similar cancerous cells (ex: B35 neuroblastoma) and rapid proliferation thereafter-towards the development of combinatorial theranostics to diagnose and treat aggressive cancers like neuroblastoma.

Moderate level exposure to magnetic nanodots encased in tunable poly(ethylene glycol) analouge biopolymer shell do not deleteriously affect neurite outgrowth.

Recently, huge interest has been generated in investigating the possible therapeutic use of tunable magnetic nanostructures to overcome the existing challenges to treat central nervous system damage related conditions. However, several issues (e.g., biocompatibility or remote controlled actuation for multi-modal therapeutics) limit the use of conventional magnetic nanoparticles for biomedical applications. To address many of these shortcomings, we have synthesized a monodisperse nanoscale system consisting highly water dispersible magnetic nanodots encased in a remotely tunable polyethylene glycol analouge biopolymer shell. The monodisperse nature of the nanospheres, their response to external magnetic field and volumetric transition near physiological temperatures are very attractive, especially for drug delivery systems where triggered release is necessary. To further analyze the potential for combinatorial therapeutics for central nervous system damage related conditions, we have explored the efficiency of the uptake of nanospheres into pheochromocytoma cell line 12 (PC12) cells and assessed several additional measures of neurite outgrowth. We find that nanospheres were readily incorporated into the cytosolic compartment within 3 hours and did not alter the morphology of cellular processes compared to cells not exposed to nanospheres. Quantification of neurite outgrowth did not reveal any significant differences in neurite initiation or elongation between cells treated with moderate level nanomagnet exposure compared to control cultures under similar conditions. Thus, this study reports an attractive nano-scale system with great potential to deliver therapeutics to precise locations within the nervous system for axonal outgrowth and guidance.

Photo-Magnetic Irradiation-Mediated Multimodal Therapy of Neuroblastoma Cells Using a Cluster of Multifunctional Nanostructures.

The use of high intensity chemo-radiotherapies has demonstrated only modest improvement in the treatment of high-risk neuroblastomas. Moreover, undesirable drug specific and radiation therapy-incurred side effects enhance the risk of developing into a second cancer at a later stage. In this study, a safer and alternative multimodal therapeutic strategy involving simultaneous optical and oscillating (AC) magnetic field stimulation of a multifunctional nanocarrier system has successfully been implemented to guide neuroblastoma cell destruction. This novel technique permitted the use of low-intensity photo-magnetic irradiation and reduced the required nanoparticle dose level. The combination of released cisplatin from the nanodrug reservoirs and photo-magnetic coupled hyperthermia mediated cytotoxicity led to the complete ablation of the B35 neuroblastoma cells in culture. Our study suggests that smart nanostructure-based photo-magnetic hybrid irradiation is a viable approach to remotely guide neuroblastoma cell destruction, which may be adopted in clinical management post modification to treat aggressive cancers.

Role of engineered nanocarriers for axon regeneration and guidance: current status and future trends.

There are approximately 1.5 million people who experience traumatic injuries to the brain and 265,000 who experience traumatic injuries to the spinal cord each year in the United States. Currently, there are few effective treatments for central nervous system (CNS) injuries because the CNS is refractory to axonal regeneration and relatively inaccessible to many pharmacological treatments. Smart, remotely tunable, multifunctional micro- and nanocarriers hold promise for delivering treatments to the CNS and targeting specific neurons to enhance axon regeneration and synaptogenesis. Furthermore, assessing the efficacy of treatments could be enhanced by biocompatible nanovectors designed for imaging in vivo. Recent developments in nanoengineering offer promising alternatives for designing biocompatible micro- and nanovectors, including magnetic nanostructures, carbon nanotubes, and quantum dot-based systems for controlled release of therapeutic and diagnostic agents to targeted CNS cells. This review highlights recent achievements in the development of smart nanostructures to overcome the existing challenges for treating CNS injuries.

Alternating Magnetic Field Controlled, Multifunctional Nano-Reservoirs: Intracellular Uptake and Improved Biocompatibility.

Biocompatible magnetic nanoparticles hold great therapeutic potential, but conventional particles can be toxic. Here, we report the synthesis and alternating magnetic field dependent actuation of a remotely controllable, multifunctional nano-scale system and its marked biocompatibility with mammalian cells. Monodisperse, magnetic nanospheres based on thermo-sensitive polymer network poly(ethylene glycol) ethyl ether methacrylate-co-poly(ethylene glycol) methyl ether methacrylate were synthesized using free radical polymerization. Synthesized nanospheres have oscillating magnetic field induced thermo-reversible behavior; exhibiting desirable characteristics comparable to the widely used poly-N-isopropylacrylamide-based systems in shrinkage plus a broader volumetric transition range. Remote heating and model drug release were characterized for different field strengths. Nanospheres containing nanoparticles up to an iron concentration of 6 mM were readily taken up by neuron-like PC12 pheochromocytoma cells and had reduced toxicity compared to other surface modified magnetic nanocarriers. Furthermore, nanosphere exposure did not inhibit the extension of cellular processes (neurite outgrowth) even at high iron concentrations (6 mM), indicating minimal negative effects in cellular systems. Excellent intracellular uptake and enhanced biocompatibility coupled with the lack of deleterious effects on neurite outgrowth and prior Food and Drug Administration (FDA) approval of PEG-based carriers suggest increased therapeutic potential of this system for manipulating axon regeneration following nervous system injury.