Epigenetic dysregulation in cadmium urothelial carcinogenesis

The urothelium, the epithelial lining of the bladder, is exposed to urinary-excreted carcinogens from environmental, occupational and dietary sources. These carcinogens include heavy metals such as cadmium. As cadmium is a weak mutagen, this suggests that genetic mechanisms are not responsible for cadmium-induced carcinogenesis. Non-genotoxic carcinogenesis is relatively poorly understood, however recent advances show that epigenetic dysregulation of gene expression may play an important role. The aim of my research was to investigate epigenetic dysregulation as the candidate mechanism underlying cadmium carcinogenesis of human urothelial cells. Normal human urothelial (NHU) cells were cultured as finite cell lines following isolation from surgical specimens. When established in vitro, NHU cells have a highly proliferative phenotype and can be induced to differentiate using two published methods: either through PPARγ-mediated differentiation or serum-mediated differentiation. Exposure of NHU cells to cadmium inhibited expression of the tumour suppressor genes, p16, APC and RASSF1A. As dysplasia is axiomatic of carcinoma in situ, the precursor to muscle invasive urothelial carcinoma, the potential of NHU cells to differentiate in the presence of cadmium was investigated. Following exposure to cadmium, there was a failure to upregulate archetypical differentiation-associated genes, including uroplakin 1A and 2, and cytokeratins 13 and 20. Trichostatin A, a histone deacetylase inhibitor was able to reverse some of these changes. Mass spectrometry and immunoblotting were utilised to investigate post-translational histone modification changes caused by cadmium exposure. Data showed that there was a change in histone modification marks present in NHU cell cultures exposed to cadmium that failed to upregulate differentiation markers. An increase in repressive histone marks such as methylation at H3K9 was found alongside a decrease in active marks such as acetylation at H3K18 and H3K23. This study presents evidence that cadmium exposure changes the epigenome of NHU cells and leads to compromised urothelial differentiation and downregulation of tumour suppressor genes.