Department of Pharmacology and Toxicology, University of Arizona, Tucson, Arizona 85721, USA.
Toxicol Sci. 2010 Jul;116(1):44-57. doi: 10.1093/toxsci/kfq106. Epub 2010 Apr 7.
Arsenic is a known human bladder carcinogen; however, the mechanisms underlying arsenical-induced bladder carcinogenesis are not understood. Previous research has demonstrated that exposure of a nontumorigenic human urothelial cell line, UROtsa, to 50 nM monomethylarsonous acid (MMA(III)) for 52 weeks resulted in malignant transformation. To focus research on the early mechanistic events leading to MMA(III)-induced malignancy, the goal of this research was to resolve the critical period in which continuous MMA(III) exposure (50 nM) induces the irreversible malignant transformation of UROtsa cells. An increased growth rate of UROtsa cells results after 12 weeks of MMA(III) exposure. Anchorage-independent growth occurred after 12 weeks with a continued increase in colony formation when 12-week exposed cells were cultured for an additional 12 or 24 weeks without MMA(III) exposure. UROtsa cells as early as 12 weeks MMA(III) exposure were tumorigenic in severe combined immunodeficiency mice with tumorigenicity increasing when 12-week exposed cells were cultured for an additional 12 or 24 weeks in the absence of MMA(III) exposure. To assess potential underlying mechanisms associated with the early changes that occur during MMA(III)-induced malignancy, DNA methylation was assessed in known target gene promoter regions. Although DNA methylation remains relatively unchanged after 12 weeks of exposure, aberrant DNA methylation begins to emerge after an additional 12 weeks in culture and continues to increase through 24 weeks in culture without MMA(III) exposure, coincident with the progression of a tumorigenic phenotype. Overall, these data demonstrate that 50 nM MMA(III) is capable of causing irreversible malignant transformation in UROtsa cells after 12 weeks of exposure. Having resolved an earlier timeline in which MMA(III)-induced malignant transformation occurs in UROtsa cells will allow for mechanistic studies focused on the critical biological changes taking place within these cells prior to 12 weeks of exposure, providing further evidence about potential mechanisms of MMA(III)-induced carcinogenesis.
砷是一种已知的人类膀胱癌致癌物；然而，砷诱导膀胱癌发生的机制尚不清楚。先前的研究表明，将非致瘤性人尿路上皮细胞系 UROtsa 暴露于 50 nM 一甲基胂酸（MMA(III)）中 52 周导致恶性转化。为了将研究重点放在导致 MMA(III)诱导恶性的早期机制事件上，本研究的目标是确定连续暴露 MMA(III)(50 nM)导致 UROtsa 细胞不可逆恶性转化的关键时期。暴露于 MMA(III) 12 周后，UROtsa 细胞的生长速度增加。12 周后出现无锚定依赖性生长，当 12 周暴露的细胞在无 MMA(III)暴露的情况下再培养 12 或 24 周时，集落形成继续增加。早在暴露于 MMA(III) 12 周时，UROtsa 细胞即可在严重联合免疫缺陷小鼠中致瘤，当 12 周暴露的细胞在无 MMA(III)暴露的情况下再培养 12 或 24 周时，致瘤性增加。为了评估与 MMA(III)诱导的恶性发生过程中早期变化相关的潜在机制，评估了已知靶基因启动子区域的 DNA 甲基化。尽管暴露 12 周后 DNA 甲基化相对不变，但在培养 12 周后，异常 DNA 甲基化开始出现，并在无 MMA(III)暴露的情况下继续培养 24 周时持续增加，与致瘤表型的进展一致。总的来说，这些数据表明，暴露于 50 nM MMA(III) 12 周后，UROtsa 细胞可发生不可逆恶性转化。解决了 UROtsa 细胞中 MMA(III)诱导恶性转化发生的早期时间线，将使机制研究能够集中于暴露前 12 周内这些细胞中发生的关键生物学变化，为 MMA(III)诱导致癌的潜在机制提供进一步证据。
