Abstract
Spinal cord injury (SCI) is a traumatic injury to the spinal cord which is characterized by a cascade of pathophysiological changes that include disruption of the blood spinal cord barrier (BSCB) and generation of reactive oxygen species (ROS) that ultimately leads to apoptosis cell death. The current therapy for treatment of SCI includes the delivery of high doses of neuroprotective agents which have not been proven to be effective. The challenge in therapy is to deliver low and effective doses of therapeutics to ameliorate the injury condition. Thus, this study focuses on developing nanoparticles (NP) loaded with naturally occurring antioxidant α–tocopherol (α–TP) for delivery to the injured site in SCI. Calcium alginate NP (CA–NP) and poly lactic co–glycolic acid (PLGA) NP were prepared by ionotropic gelation and solvent evaporation technique to obtain a particle size (PS) of 21.9 ± 11.19 and 152.4 ± 10.6 nm, respectively and were characterized. An ultra–high pressure liquid chromatography (UPLC) based method with a fluorescence detector was developed and validated for the detection and quantification of α-TP. The CA–NP and
PLGA were found to have an entrapment efficiency of 4.00 ± 1.63% and 76.61 ± 11.45%, respectively. In vitro studies on human astrocyte–spinal cord (HA–sp) cells were conducted to determine the effect of NP on the cell lines. Blank CA–NP were found to decrease cell viability as their concentrations were increased, indicating that the NP themselves, had a deleterious effect on HA–sp cells. PLGA NP were found to be less toxic compared the pure drug solutions. The anti–oxidative effect of pure drug solution was compared to α–TP loaded PLGA NP against oxidative stress induced by 400 μM H2O2. It was observed that α–TP PLGA NP showed two-fold higher protection as compared to the pure drug solutions. Thus, it was concluded that CA–NP are unsuitable for the treatment against oxidative stress in SCI. Additionally, pure α–TP solution and α–TP loaded PLGA NP showed protection gainst oxidative stress conditions with a dose of 100 μM. However, α–TP loaded PLGA NP showed significantly (p < 0.05) higher protection as compared to pure drug solution. The in vitro data suggested that in vivo studies need to be investigated and compared.