Abstract
In this study, we aimed to study the role of growth factor receptor-bound protein 2 (Grb2) in palmitic acid-induced steatosis and other “fatty liver” symptoms in vitro. HepG2 cells, with or without stably suppressed Grb2 expression, were incubated with palmitic acid for 24 h to induce typical clinical “fatty liver” features, including steatosis, impaired glucose metabolism, oxidative stress, and apoptosis. MTT and Oil Red O assays were applied to test cell viability and fat deposition, respectively. Glucose uptake assay was used to evaluate the glucose utilization of cells. Quantitative polymerase chain reaction and Western blot were used to measure expressional changes of key markers of insulin signaling, lipid/glucose metabolism, oxidative stress, and apoptosis. After 24-h palmitic acid induction, increased fat accumulation, reduced glucose uptake, impaired insulin signaling, enhanced oxidative stress, and increased apoptosis were observed in HepG2 cells. Suppression of Grb2 in HepG2 significantly reduced fat accumulation, improved glucose metabolism, ameliorated oxidative stress, and restored the activity of insulin receptor substrate-1/Akt and MEK/ERK pathways. In addition, Grb2 deficiency attenuated hepatic apoptosis shown by reduced activation of caspase-3 and fluorescent staining. Modulation of Bcl-2 and Bak1 also contributed to reduced apoptosis. In conclusion, suppression of Grb2 expression in HepG2 cells improved hepatic steatosis, glucose metabolism, oxidative stress, and apoptosis induced by palmitic acid incubation partly though modulating the insulin signaling pathway.
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Editor: T. Okamoto
Xiangxiang Shan and Yufeng Miao contributed equally to this work.
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ESM Fig. 1
Deficiency of Grb2 did not influence the fat accumulation and cell viability of untreated HepG2 cells. Representative pictures for Oil Red O staining of HepG2 cells of (A) control and (B) Grb2 suppression group. Quantification of Oil Red O signals was concluded in C. Cell viability was measured by MTT assay (D). Error bars represent standard error of the mean of samples within a group (n = 5). Magnification ×400. ns Not significant (JPEG 33 kb)
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Shan, X., Miao, Y., Fan, R. et al. Suppression of Grb2 expression improved hepatic steatosis, oxidative stress, and apoptosis induced by palmitic acid in vitro partly through insulin signaling alteration. In Vitro Cell.Dev.Biol.-Animal 49, 576–582 (2013). https://doi.org/10.1007/s11626-013-9646-9
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DOI: https://doi.org/10.1007/s11626-013-9646-9