Banerjee Mayukh, Lykoudi Angeliki, Hwang Jae Y, Pan Jianmin, Rai Shesh N, Park Juw W, States J Christopher
Department of Pharmacology and Toxicology, University of Louisville, 505 S. Hancock Street, Louisville, KY 40202, USA; Center for Integrative Environmental Health Sciences, University of Louisville, 505 S. Hancock Street, Louisville, KY 40202, USA.
Department of Pharmacology and Toxicology, University of Louisville, 505 S. Hancock Street, Louisville, KY 40202, USA.
Toxicol Appl Pharmacol. 2025 Feb;495:117209. doi: 10.1016/j.taap.2024.117209. Epub 2024 Dec 22.
Dysregulated miRNA expression contributes to development of arsenic-induced cutaneous squamous cell carcinoma (cSCC). hsa-miR-186 (miR-186) is overexpressed in arsenical cSCC tissues as well as in preclinical cell line model of arsenical cSCC. Simultaneous miR-186 overexpression and chronic inorganic trivalent arsenite (iAs; 100 nM) exposure transformed human HaCaT cell line preferentially over miR-186 overexpression or iAs exposure alone. Both iAs and miR-186 regulate the expression of wide range of mRNA targets. However, how their interaction impacts the transcriptome-wide mRNA expression landscape ushering in cancer is unknown. We performed longitudinal RNA-seq analysis in passage-matched HaCaT cell clones (±miR-186 overexpression) with simultaneous chronic iAs exposure (0/100 nM) at 12 and 29 weeks. We determined the impact of each factor and their interaction towards differential gene expression and pathway dysregulation employing two different statistical approaches (t-statistic and 2-factor ANOVA). We show that a core set of pathways are dysregulated deterministically irrespective of the statistical approach chosen, possibly representing necessary changes for transformation. The data suggest that each clonal line could take a unique route to dysregulate this core set of pathways necessary for transformation, highlighting the possible role of stochasticity in cancer development. Evidence is presented to sift the strengths and weaknesses of each statistical methodology in providing biological understanding of events that play crucial roles in carcinogenesis in large datasets with multiple contributing variables.
微小RNA（miRNA）表达失调促成了砷诱导的皮肤鳞状细胞癌（cSCC）的发生。hsa-miR-186（miR-186）在砷性cSCC组织以及砷性cSCC的临床前细胞系模型中均有过表达。miR-186过表达与慢性无机三价亚砷酸盐（iAs；100 nM）同时暴露，相较于单独的miR-186过表达或iAs暴露，更优先地转化了人HaCaT细胞系。iAs和miR-186都调控多种mRNA靶标的表达。然而，它们的相互作用如何影响导致癌症的全转录组mRNA表达格局尚不清楚。我们对传代匹配的HaCaT细胞克隆（±miR-186过表达）进行了纵向RNA测序分析，并在第12周和第29周同时进行慢性iAs暴露（0/100 nM）。我们采用两种不同的统计方法（t统计量和双因素方差分析）确定了每个因素及其相互作用对差异基因表达和通路失调的影响。我们表明，无论选择何种统计方法，一组核心通路都会确定性地失调，这可能代表了转化所需的必要变化。数据表明，每个克隆系可能会采取独特的途径来失调转化所需的这组核心通路，突出了随机性在癌症发展中的可能作用。本文提供的证据筛选了每种统计方法在为具有多个影响变量的大型数据集中对致癌作用起关键作用的事件提供生物学理解方面的优缺点。
