How To Evaluate The Toxicology Of Nanoparticles
Nano-drug delivery systems are constantly exposed to complex physiological environments in vivo. The interaction between nano-materials, proteins and cells, their effects and possible toxicity is the key to evaluate and understand the compatibility and toxicity of nano-materials. Cell-based toxicity evaluation is the main method for the in vitro toxicity evaluation of nanometer drug delivery systems, including cell uptake and processing of nanomaterials, effects on cell signals, interference with membranes, effects on cell electron transfer beams, cytochemical factors and reactive oxygen species (ROS) production, intercellular interaction and intercellular transport, gene regulation, obvious toxic reactions, potential toxicity, and cell necrosis or apoptosis.
Toxicology In Vitro Research Method Of Nano Drug Delivery System
A variety of cells can be used for the cytotoxic biological evaluation of nano-drug delivery systems. For targeting nanoparticles, target cells need to be selected for evaluation. Cell detection mainly includes 5 indicators: cell viability, oxidative stress, cell stress response, cell morphology and particle uptake of cells. Among them, cell morphology and particle uptake of cells are mainly checked by microscope (TEM and BSE) combined with fluorescence technology. The other three indicators are checked as follows.
- Cell Viability
Cell viability testing is the study of the ability of nano drug delivery systems to kill living cells. Cell viability assays are mostly based on dyes, using the different of uptake, discharge, and conversion of dyes or enzyme-converted dyes of live cells and dead cells to quantify cell viability by colorimetry or fluorescence. Dyes generally include: neutral red, trypan blue, and lactic acid, dehydrogenase, LIVE / DEAD system, formazan (MTT, MTS, WST), Alma Blue, Coomassie Blue, ATP fluorescein luminescence, adenylate conversion kinase release, mitochondrial membrane potential, and sulfur spraying.
- Oxidative Stress
Many nanoparticles produce reactive oxygen species (ROS), and the oxidative stress (OS) mechanism caused by reactive oxygen species is the main reason for the toxicity of nanoparticles. Oxidative stress is that when the body is subjected to various harmful stimuli, the body’s highly active molecules such as reactive oxygen free radicals ROS and reactive nitrogen free radicals produce too much, the degree of oxidation exceeds the removal of oxides, and the oxidative system and the antioxidant system are out of balance, leading to tissue damage. Oxidative stress is a negative effect. It is an important factor leading to aging and diseases. It can damage proteins, lipids and DNA, and is associated with many diseases. The oxidative stress evaluation method of the nanometer drug delivery system is to directly measure the ROS production of cells, and the methods include a detection method based on fluorescein compounds and an electron paramagnetic resonance (EPR).
- Cell Stress Response
Nanoparticles may bring non-fatal damage to cells and changes in cell behavior, which are mainly manifested in the regulation of protein and gene expression, regulation of phagocytic capacity, and inflammatory response. Gene and protein expression changes can be measured by polymerase chain reaction (PCR), western blotting, and total protein analysis (such as BCA and Bradford). For example, quantitative real-time PCR (qPCR) can be used to measure CDKN1A (cell cycle arrest gene), GADD45β (DNA damage dependent), IL-6 (inflammatory response), NFKBIA (inflammatory response), EGFP (reporter/gene), and NF-rB (including Fibrosis/inflammatory growth factor promoter/enhancer regions, cytokines and adherent cells) gene expression levels. These assays can explain the effects of toxicity on gene expression and the mechanism of toxicity.