Investigating Internalization of Reporter-Protein-Functionalized Polyhedrin Particles by Brain Immune Cells.

Achieving sustained drug delivery to the central nervous system (CNS) is a major challenge for neurological injury and disease, and various delivery vehicles are being developed to achieve this. Self-assembling polyhedrin crystals (POlyhedrin Delivery System; PODS) are being exploited for the delivery of therapeutic protein cargo, with demonstrated efficacy in vivo. However, to establish the utility of PODS for neural applications, their handling by neural immune cells (microglia) must be documented, as these cells process and degrade many biomaterials, often preventing therapeutic efficacy. Here, primary mouse cortical microglia were cultured with a GFP-functionalized PODS for 24 h. Cell counts, cell morphology and Iba1 expression were all unaltered in treated cultures, indicating a lack of acute toxicity or microglial activation. Microglia exhibited internalisation of the PODS, with both cytosolic and perinuclear localisation. No evidence of adverse effects on cellular morphology was observed. Overall, 20-40% of microglia exhibited uptake of the PODS, but extracellular/non-internalised PODS were routinely present after 24 h, suggesting that extracellular drug delivery may persist for at least 24 h.

Enhancing the regenerative potential of stem cell-laden, clinical-grade implants through laminin engineering.

Protected delivery of neural stem cells (NSCs; a major transplant population) within bioscaffolds has the potential to improve regenerative outcomes in sites of spinal cord injury. Emergent research has indicated clinical grade bioscaffolds (e.g. those used as surgical sealants) may be repurposed for this strategy, bypassing the long approval processes and difficulties in scale-up faced by laboratory grade materials. While promising, clinical scaffolds are often not inherently regenerative. Extracellular molecule biofunctionalisation of scaffolds can enhance regenerative features such as encapsulated cell survival/distribution, cell differentiation into desired cell types and nerve fibre growth. However, this strategy is yet to be tested for clinical grade scaffolds. Here, we show for the first time that Hemopatch™, a widely used, clinically approved surgical matrix, supports NSC growth. Further, functionalisation of Hemopatch™ with laminin promoted homogenous distribution of NSCs and their daughter cells within the matrix, a key regenerative criterion for transplant cells.

Magnetic nanoparticle mediated transfection of neural stem cell suspension cultures is enhanced by applied oscillating magnetic fields.

Safe genetic modification of neural stem cell (NSC) transplant populations is a key goal for regenerative neurology. We describe a technically simple and safe method to increase transfection in NSCs propagated in the neurosphere (suspension culture) model, using magnetic nanoparticles deployed with applied oscillating magnetic fields ('magnetofection technology'). We show that transfection efficiency was enhanced over two-fold by oscillating magnetic fields (frequency=4 Hz). The protocols had no effect on cell viability, cell number, stem cell marker expression and differentiation profiles of 'magnetofected' cultures, highlighting the safety of the technique. As far as we are aware, this is the first successful application of magnetofection technology to suspension cultures of neural cells. The procedures described offer a means to augment the therapeutic potential of NSCs propagated as neurospheres - a culture model of high clinical translational relevance - by safe genetic manipulation, with further potential for incorporation into 'magneto-multifection' (repeat transfection) protocols. FROM THE CLINICAL EDITOR: This team of investigators describe a simple and safe method to increase transfection in neural stem cells using magnetic nanoparticles deployed with oscillating magnetic fields, demonstrating a greater than two-fold transfection efficiency increase by applying low frequency magnetic oscillation.