Potential of asiaticoside encapsulated alginate chitosan nanoparticles in the management of antiproliferative activity in C6 glioma cells

Abstract

Background

Asiaticoside is a pentacyclic triterpenoid saponin constituent of the medicinal herb Centella asiatica, known for its neuroprotective, anticancer and other biological effects. The present study aims to develop and characterise asiaticoside encapsulated alginate chitosan nanoparticles (ACNPs) and evaluate their antiproliferative potential on C6 glioma cells.

Methods

ACNPs were prepared by the ionic gelation polyelectrolyte complexation technique and characterised by dynamic light scattering, scanning electron microscopy and transmission electron microscopy. Cytotoxicity and antiproliferative effect of ACNPs were studied on C6 glioma cells using MTT and EdU-assays. Apoptosis/necrosis in C6 glioma cells was determined by annexin V and acridine orange/ethidium bromide (AO/EtBr) assays. Intracellular ROS generation in C6 glioma cells exposed with ACNP was determined by DCFDA assay using flow cytometry. Cell cycle analysis in C6 glioma cells treated with ACNP was performed by flow cytometry.

Results

Synthesised ACNPs showed spherical shape with 200 nm particle size, good thermal stability, and an acceptable polydispersity index. ASI from synthesized ACNPs demonstrated 90% encapsulation efficiency (EE) and 10% drug release within 24 h. Upon ACNP treatment mBA cells exhibited good cell viability and intact cell morphology. However, C6 cells displayed dose-dependent cytotoxicity when treated with ACNP. Annexin V and acridine orange/ethidium bromide (AO/EtBr) assays revealed late apoptosis and necrosis (p < 0.05) in C6 glioma cells treated with ACNP. Likewise, ACNPs significantly augmented reactive oxygen species generation (p < 0.05) in C6 glioma cells. ACNP treatment also resulted in cell cycle arrest at the G1 phase with chromatin condensation in C6 glioma cells.

Conclusions

These findings suggest that ACNPs act as an antiproliferative agent against C6 glioma cells via an increased intracellular ROS pathway that promotes apoptosis/necrosis. This study throws light on more in-depth mechanistic aspects of managing brain disorders using C. asiatica and its phytoconstituents.