Category: Preclinical Development
Purpose: Several lines of evidence indicate that sildenafil (SF), a phosphodiesterase 5 inhibitor used for treatment of erectile dysfunction, crosses the blood brain barrier and could have beneficial therapeutic effects in the treatment of stroke, subarachnoid hemorrhage, dementia, and neurodegenerative disorders. In addition, SF may improve delivery of chemotherapeutic agents to brain tumors. Such effects could be enhanced by targeted nanoparticles (NPs) known to improve drug delivery to the brain. However, nanoencapsulation and surface modification of NPs for targeting purposes may alter the toxicity profile of encapsulated drugs. Thus, the objective of the current study was to develop new Tween 80 (T80)-targeted SF-loaded chitosan NPs (T80-SF-NPs) for potential brain delivery and to assess the effects of nanoencapsulation and T80 coating on functional and structural hepatic features in male rats.
Methods: Sildenafil chitosan NPs (SF-NPs) and T80-SF-NPs) were prepared by ion gelation and characterized for particle size, polydispersity index (PdI), zeta potential (ZP), entrapment efficiency (EE %) and morphology by TEM imaging. The effects of SF-NPs and T80-SF-NPs in comparison with SF solution at 1.5 mg/kg dose for 21 consecutive days on the liver of male rats was assessed biochemically and histopathologically using saline as control. Biochemical testing included changes in the activity of phase I drug metabolizing enzymes, namely Cytochrome c reductase (CcR), Aryl hydrocarbon hydroxylase (AAH) and 7-Ethoxycoumarin-O-deethylase (EOD) and the protein expression of different cytochrome P isozymes by Western immunoblotting analysis as well as the activity of antioxidant enzymes including glutathione S transferase (GST), glutathione reductase (GSR), glutathione peroxidase (GTP), catalase (CAT) and superoxide dismutase (SOD). Histopathological changes in the livers of rats were examined using H&E staining.
Results: SF-NPs and T80-SF-NPs spherical chitosan nanospheres showing in vitro characteristics (mean size, PdI and ZP) suitable for brain delivery were developed by modulating experimental variables. Biochemical assessments indicated heterogeneous effects of SF-NPs and T80-SF-NPs in comparison with SF solution on the activity of metabolic enzymes and protein expression of CYP2E1, CYP34A and CYT b5. Data for antioxidant enzymes indicated that SF did not change SOD, reduced GST and GR but increased GPx activities (p< 0.05). While chitosan nanoencapsulation did not change CAT level relative to SF, it significantly attenuated the SF-induced decrease in GST and GR levels and augmented the increase in GPx and SOD activity. T80 coating did not significantly modify the effect of SF-NPs on GST, GR and GPx activities. However, it brought SOD activity back to normal and significantly increased CAT activity. Histopathological changes in liver sections (Fig. 1) indicated obvious SF-induced necrobiotic changes, fibrosis, hemolysis and inflammation. These were reduced by chitosan NPs, yet ballooning degeneration of hepatocytes was clear. Interestingly, T80 coating considerably improved liver architecture and cells with prevention of inflammation and hemolysis.
Conclusion: This study provides the first preclinical evidence of the effect of chitosan nano-encapsulation and T80 surface modification of NPs on the SF-induced hepatotoxicity. In general, T80-coated chitosan NPs greatly reduced the adverse effects of SF on the liver, enhancing the protective effect provided by chitosan nanoencapsulation.