Ling Cao

Date of Award

9-7-2012

Document Type

Dissertation

Degree Name

Doctor of Philosophy (PhD)

Department

Biology

First Advisor

Seema Somji

Abstract

Arsenic and cadmium (Cd2+) are environmental carcinogens that have been implicated in the development of bladder cancer. However, there are no good in vitro model systems to study the development of bladder cancer caused by these carcinogens. Previous studies from this laboratory have shown that arsenite (As3+) and Cd2+ exposure can cause direct malignant transformation of an immortalized human urothelial cell line, the UROtsa. In the present study the repeatability of the transformation process was examined by isolating and characterizing 6 additional Cd2+- transformed lines and 5 additional As3+- transformed cell lines. All the transformed cell lines formed tumors when injected into nude mice and these tumors had histologies similar to the transitional carcinoma of the bladder. The tumors had distinct areas of squamous differentiation which is an indicator of poor prognosis in bladder cancer. Some of the transformed cell lines overexpressed the intermediate filament proteins keratin 6 and keratin 16. However all the tumor heterotransplants had increased expression of keratin 6 and 16 and the expression was localized to the areas of the tumor that had squamous differentiation suggesting that keratin 6/16 may be a marker for cells that have the potential to undergo squamous differentiation. Treatment of the parent UROtsa cells with arsenic and Cd2+ did not induce the expression of keratin 6 suggesting that an in vivo environment may be necessary to induce the expression of these proteins. The effect of various growth factors on the expression of keratin 6/16 was also studied. UROtsa parent cells and the transformed cell lines were treated with epidermal growth factor (EGF) and or insulin for 24 h and the expression levels of keratin 6/16 was determined. The results indicate that EGF and insulin can induce the expression of keratin 6/16 in the parent UROtsa cell line but do not have an effect on the transformed cell lines irrespective of the basal levels of keratin 6/16 expression suggesting that a constitutive mechanism may be involved in the expression of keratin 6/16 in the transformed cell lines. The signal transduction pathway involved in the up-regulation of keratin 6 in the normal and transformed cell lines was also determined. The data obtained suggests that the extracellular signal-regulated kinase, ERK 1 and 2 (ERK1/2) pathway was involved in the induction of keratin 6. Inhibition of this pathway resulted in a decrease in the expression of keratin 6 in the parent and transformed cell line, further implicating this pathway in the up-regulation of keratin 6.

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