Acrylamide exerts its cytotoxicity in NIH/3T3 fibroblast cells by apoptosis

Abstract

Objective:

Acrylamide is a chemical utilized in various industries, and many studies have demonstrated its toxicity. The NIH/3T3 mouse embryonic cell line is the standard cell line of fibroblasts, which have a pivotal role with their versatile functions in the body. However, only two studies have attempted to investigate the effect of acrylamide on these crucial cells. To fill this knowledge gap, we aimed to determine the effects of acrylamide on NIH/3T3 cells.

Method:

First, we performed the 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl-2H-tetrazolium bromide assay and calculated the IC₅₀ dose of acrylamide. Then, we treated cells with the IC₅₀ dose of acrylamide for 24 h and determined whether the dominant death mode of NIH/3T3 cells was apoptosis or necrosis by annexin V and caspase 3/7 assays. Finally, we performed confocal microscopy and transmission electron microscope (TEM) analysis for observing the morphological alterations.

Results:

MTT assay results showed that acrylamide treatment reduced the viability of NIH/3T3 cells dose-dependently and that the IC₅₀ of acrylamide was 6.73 mM. Based on annexin V and caspase 3/7 assays, the dominant death mode of NIH/3T3 cells was determined to be apoptosis. Also, caspase 3/7 activities of the acrylamide-treated NIH/3T3 cells were three times greater than those of the untreated NIH/3T3 cells. Furthermore, we observed membrane blebbing, nuclear chromatin clumping, and cytoplasmic vacuolization in TEM analysis and apparent apoptotic bodies, nuclear fragmentations, and condensations in confocal microscopy.

Conclusions:

In conclusion, our results suggested that the IC₅₀ of acrylamide against NIH/3T3 cells for 24 h was 6.73 mM and that acrylamide exerted its cytotoxic and anti-proliferative effects on these cells mainly via apoptosis.