Differentially Expressed mRNA Targets of Differentially Expressed miRNAs Predict Changes in the TP53 Axis and Carcinogenesis-Related Pathways in Human Keratinocytes Chronically Exposed to Arsenic

Abstract

Arsenic is a widely distributed toxic natural element. Chronic arsenic ingestion causes several cancers, especially skin cancer. Arsenic-induced cancer mechanisms are not well defined, but several studies indicate that mutation is not the driving force and that microRNA expression changes play a role. Chronic low arsenite exposure malignantly transforms immortalized human keratinocytes (HaCaT), serving as a model for arsenic-induced skin carcinogenesis. Early changes in miRNA expression in HaCaT cells chronically exposed to arsenite will reveal early steps in transformation. HaCaT cells were maintained with 0/100 nM NaAsO2 for 3 and 7 weeks. Total RNA was purified. miRNA and mRNA expression was assayed using Affymetrix microarrays. Targets of differentially expressed miRNAs were collected from TargetScan 6.2, intersected with differentially expressed mRNAs using Partek Genomic Suite software, and mapped to their pathways using MetaCore software. MDM2, HMGB1 and TP53 mRNA, and protein levels were assayed by RT-qPCR and Western blot. Numerous miRNAs and mRNAs involved in carcinogenesis pathways in other systems were differentially expressed at 3 and 7 weeks. A TP53 regulatory network including MDM2 and HMGB1 was predicted by the miRNA and mRNA networks. Total TP53 and TP53-S15-phosphorylation were induced. However, TP53-K382-hypoacetylation suggested that the induced TP53 is inactive in arsenic exposed cells. Our data provide strong evidence that early changes in miRNAs and target mRNAs may contribute to arsenic-induced carcinogenesis.

Dynamic alteration in miRNA and mRNA expression profiles at different stages of chronic arsenic exposure-induced carcinogenesis in a human cell culture model of skin cancer

“…In order to isolate total RNA (including mRNA and small RNA) from cells harvested at 7, 19 and 28-wk time points, mirVana™ RNA isolation kit was employed as described previously (Al-Eryani et al 2018b ) following the manufacturer’s recommendations. The quality of the isolated RNA was determined using the Agilent RNA 6000 Pico Kit, Eukaryote, version 2.6 and the Agilent 2100 Bioanalyzer instrument (Agilent Technologies, Inc., Santa Clara, CA, USA).…”

Sublethal exposure of small few-layer graphene promotes metabolic alterations in human skin cells

“…Although genotoxicity assays are still required, the results of this work show the relevance of refining our roadmap for interpreting data for a better understanding of the interactions between different GRMs and human barriers, with particular attention paid to metabolism and mitochondria even when an evident phenotype is not noticeable. These studies are also important because some GRMs have already been proposed for use in contact with skin, for example in textiles 29 or wearable electronics 30 , 31 . Special care must be taken to study the degradation and release of these materials during service life.…”

Arsenic-Induced Carcinogenesis: The Impact of miRNA Dysregulation

“…Several modes of action have been suggested in arsenic carcinogenesis including DNA repair inhibition, comutagenicity, oxidative stress, cell proliferation, aneuploidy, interaction with zinc finger proteins and epigenetic alterations including changes in DNA methylation, histone modification, and miRNA expression (Bailey and Fry, 2014;Reichard and Puga, 2010;States, 2015;Zhou et al, 2014). Recent in vitro studies support an association of exposure to arsenic with altered miRNA expression and alternative mRNA splicing (Al-Eryani et al, 2018;Riedmann et al, 2015).…”