The effect of silica nanoparticle exposure on cultured human keratocytes

Abstract

PurposeSilica nanoparticles (SiNPs) are closely related to our daily life including drug delivery, cosmetics and fine dust. However, the influence of SiNPs on human corneal keratocyte cells has not yet been widely studied. In this study, we investigated the effect of SiNPs on cultured human keratocytes.MethodsHuman keratocytes were cultured in DMEM/F12(1:1) medium containing 10% FBS and Antibiotic‐Antimycotic. SiNPs exposure was performed by adding 50, 100 and 150 nm of non‐porous SiNPs into culture media with different concentrations (10, 20, 50, 100 μg/ml) for 24 and 48 h. Keratocytes viability was measured using CCK‐8 reagent after 24 and 48 h exposure of SiNPs. Release of Lactate Dehydrogenase (LDH) was measured by LDH cytotoxicity detection kit. The measurement of intracellular reactive oxygen species (ROS) generation was performed using Fluorometric Intracellular Ros Kit. Cellular autophagy activity (LC3B and Beclin 1) and mTOR pathway activation (p‐mTOR and mTOR) were detected by Western Blot.ResultsIn human keratocytes, significant cellular cytotoxicity and membrane damage were not detected after exposure to three different sizes of SiNPs for 24 and 48 h. Intracellular ROS generation was slightly increased at high concentration (100 μg/ml) of three sizes of SiNPs. And Cellular autophagy was significantly activated in concentration‐dependent manner after exposure to SiNPs for 24 h with increase of western blot for LC3A/B. The upstream of autophagy signaling, the mTOR pathway, was slightly activated after exposure to three sizes of SiNPs.ConclusionsSiNPs (50, 100, 150 nm) induced no significant cytotoxicity in cultured human keratocytes.

Chlorpyrifos activates cell pyroptosis and increases susceptibility on oxidative stress‐induced toxicity by miR‐181/SIRT1/PGC‐1α/Nrf2 signaling pathway in human neuroblastoma SH‐SY5Y cells

“…Formation of ROS in cells was detected using the Fluorometric Intracellular ROS kit (Sigma-Aldrich; Merck KGaA) according to the manufacturer's instruction. 28 Briefly, cells were cultured in serum-free RPMI-1640 at 37 C and were incubated with 10 μM 2 0 ,7 0 -dichlorofluorescin diacetate for 20 min. The florescent signal was then measured by flow cytometry (BD Biosciences, San Jose, California).…”