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CHARACTERIZATION OF HUMAN URINARY BLADDER CELL LINE UROTSA TRANSDUCED WITH RAT ASLLL-METHYLTRANSFERASE
Citation:
Drobna, Z., S. B. Waters, Walton, Felecia S, V. Devesa, M Styblo, AND D J. Thomas. CHARACTERIZATION OF HUMAN URINARY BLADDER CELL LINE UROTSA TRANSDUCED WITH RAT ASLLL-METHYLTRANSFERASE. Presented at Society of Toxicology, Baltimore, MD, March 21-25, 2004.
Description:
In humans, the biomethylation of arsenic (As) is catalyzed by an As(III)-methyltransferase (Cyt19) and yields pentavalent and trivalent methylated arsenicals. Cyt19 activity and expression levels vary among tissues. For example, Cyt19 mRNA is not detected in UROtsa cells, a human uroepithelial cell line that cannot methylate As. Thus, UROtsa cells are an ideal null background to assess the consequences of As methylation and production of methylated arsenicals. A retroviral gene delivery system (pLEGFP-N1) was used to transduce UROtsa cells with a rat Cyt19 (rCyt19) construct. After neomycin selection (G418), UROtsa/rCyt19 cells were checked for the transduction efficiency. Both Touch-down RT-PCR analysis for Cyt19mRNA and Western Blot analysis of UROtsa/rCyt19 cell lysate (using preimmune rabbit serum) gave positive signals in transduced cells. Cyt19 activity in these cells was tested by examining methylation of radiolabeled arsenite. After 65 hours, UROtsa/rCyt19 cells converted between 20 to 25% of radioarsenic to mono- and dimethylated arsenicals (MAs and DMAs). DMAs was the predominant metabolite in cell lysate and medium. At steady state, UROtsa/rCyt19 cells retained less radioarsenic than did similarly-exposed UROtsa cells. UROtsa/rCyt19 cells were more resistant to the acute cytotoxicity of iAs(III) and MAs(III) than was the parental cell line. However, transduced cells were more sensitive than parental cells to the acute cytotoxicity of DMAs(III). Establishment of cell lines differing in capacity to methylate As provides opportunities to examine relationships between production of methylated arsenicals and the metalloid's action as a toxin and carcinogen. (This abstract does not reflect US EPA policy.)