Date of Award
Master of Science (MS)
Webb , Ken
Vyavahare , Naren
Traumatic brain injury (TBI) is a leading cause of death and disability worldwide. There is currently no effective therapeutic for the treatment of TBI. Primary injury, from the initial injury, causes contusions and hemorrhaging at the site of impact. Diffuse damage is caused throughout the brain from the impact; this includes axonal injury, hypoxic brain damage, brain swelling and vascular injury. Brain damage continues as the secondary injury; which is characterized by hypoxia, hypotension, amino acid excitotoxicity and ionic imbalance. All of these conditions cause additional cell death and damage. Inflammation, brought about by reduced cyclic AMP levels is also seen post-injury. After injury, the glial scar and myelin produce neurite growth inhibitory molecules. There are several different types of myelin associated inhibitors expressed by oligodendrocytes; these interact with multiple types of neuron surface receptors triggering the RhoA cascade, which inhibits actin polymerization and neurite outgrowth. Chondroitin sulfate proteoglycans (CSPG) expressed on astrocytes also inhibits growth through the same RhoA pathway. Several strategies have elected to knockdown the RhoA gene or other genes involved in the growth inhibition pathway. The objective of this work is to develop novel neuron-specific nanotherapeutics for combinatorial therapy of drug and small interfering RNA (siRNA) targeting both extrinsic and intrinsic barriers to neuroplasticity. This neuron-specific polymeric micelle nanotherapeutics will be designed as follows: First, a cell-type specific targeting moiety such as an antibody can be conjugated to the polymeric micelle nanoparticle surface to deliver nanotherapeutics specifically to neurons. Second, RhoA siRNA, can be targeted to common intracellular signal transduction pathways for inhibitory molecules such as myelin and CSPGs. Third, a hydrophobic drug, a phosphodiesterase 4 inhibitor (rolipram) will be incorporated in the PgP micelle to increase intrinsic neuronal growth capacity by preventing injury-induced reductions in cAMP levels. To achieve this goal, we synthesized amphiphilic block copolymers, poly (lactide-co-glycolide)-graft-polyethylenimine (PLGA-g-PEI: PgP) using PLGA as a hydrophobic core forming block and PEI as a hydrophilic shell forming block and characterized the physico-chemical properties of the PgP micelle as a delivery carrier for combinatorial therapy of nucleic acid and drug. We demonstrated that the PgP micelle is a promising nucleic acid delivery carrier using phMGFP plasmid as a reporter gene in C6 (glioma) cells and primary chick forebrain neurons (CFN) cells in 10% serum containing media in vitro. We also studied incorporating rolipram in the PgP micelle and successfully conjugated an antibody (mouse IgG) on the surface of PgP. Currently, we are evaluating PgP as a siRNA delivery carrier to primary CFN cells and preparing PgP-mNgR1 using NgR1 monoclonal antibody and evaluating the feasibility of PgP-mNgR1 as a neuron-specific nucleic acid carrier for targeting neuron cells in a rat cortical neuron /astrocyte co-culture system. In the future, we will study rolipram-loaded PgP-Ab as a nucleic acid/drug carrier using RhoA siRNA in hypoxic conditions as a TBI model in vitro and a rat traumatic brain injury model in vivo.
Nice, Justin, "SYNTHESIS AND CHARACTERIZATION OF POLYMERIC MICELLE DELIVERY SYSTEM AS A DRUG AND" (2014). All Theses. 1930.