Arsenic is a universal environmental toxicant associated mostly with skin related diseases in people exposed to low doses over a long term. Low dose arsenic trioxide (ATO) with long exposure will lead to chronic exposure. Experiments were performed to provide new knowledge on the incompletely understood mechanisms of action of chronic low dose inorganic arsenic in keratinocytes. Cytotoxicity patterns of ATO on long-term cultures of HaCaT cells on collagen IV was studied over a time course of 14 days. DNA damage was also assessed. The percentages of viable cells after exposure were measured on Day 2, Day 5, Day 8, and Day 14. Statistical and visual analytics approaches were used for data analysis. In the result, a biphasic toxicity response was observed at a 5 μg/ml dose with cell viability peaking on Day 8 in both chronic and acute exposures. Furthermore, a low dose of 1 μg/ml ATO enhanced HaCaT keratinocyte proliferation but also caused DNA damage. Global gene expression study using microarray technique demonstrated differential expressions of genes in HaCaT cell exposed to 0.5 μg/ml dose of ATO up to 22 passages. Four of the up-regulated and 1 down-regulated genes were selected and confirmed with qRT-PCR technique. These include; Aldo-Keto Reductase family 1, member C3 (AKR1C3), Insulin Growth Factor-Like family member 1 (IGFL1), Interleukin 1 Receptor, type 2 (IL1R2) and Tumour Necrosis Factor [ligand] Super-Family, member 18 (TNFSF18), and down-regulated Regulator of G-protein Signalling 2 (RGS2). The decline in growth inhibiting gene (RGS2) and increase in AKR1C3 may be the contributory path to chronic inflammation leading to metaplasia. This pathway is proposed to be a mechanism leading to carcinogenesis in skin keratinocytes. The observed over expression of IGFL1 may be a means of triggering carcinogenesis in HaCaT keratinocytes. In conclusion, it was established that at very low doses, arsenic is genotoxic and induces aberrations in gene expression though it may appear to enhance cell proliferation. The expression of two genes encoding membrane proteins IL1R2 and TNFSF18 may serve as possible biomarkers of skin keratinocytes intoxication due to arsenic exposure. This research provides insights into previously unknown gene markers that may explain the mechanisms of arsenic-induced dermal disorders including skin cancer / Environmental Sciences / D. Phil. (Environmental science)

Arsenic is a universal environmental toxicant associated mostly with skin related diseases in people exposed to low doses over a long term. Low dose arsenic trioxide (ATO) with long exposure will lead to chronic exposure. Experiments were performed to provide new knowledge on the incompletely understood mechanisms of action of chronic low dose inorganic arsenic in keratinocytes. Cytotoxicity patterns of ATO on long-term cultures of HaCaT cells on collagen IV was studied over a time course of 14 days. DNA damage was also assessed. The percentages of viable cells after exposure were measured on Day 2, Day 5, Day 8, and Day 14. Statistical and visual analytics approaches were used for data analysis. In the result, a biphasic toxicity response was observed at a 5 μg/ml dose with cell viability peaking on Day 8 in both chronic and acute exposures. Furthermore, a low dose of 1 μg/ml ATO enhanced HaCaT keratinocyte proliferation but also caused DNA damage. Global gene expression study using microarray technique demonstrated differential expressions of genes in HaCaT cell exposed to 0.5 μg/ml dose of ATO up to 22 passages. Four of the up-regulated and 1 down-regulated genes were selected and confirmed with qRT-PCR technique. These include; Aldo-Keto Reductase family 1, member C3 (AKR1C3), Insulin Growth Factor-Like family member 1 (IGFL1), Interleukin 1 Receptor, type 2 (IL1R2) and Tumour Necrosis Factor [ligand] Super-Family, member 18 (TNFSF18), and down-regulated Regulator of G-protein Signalling 2 (RGS2). The decline in growth inhibiting gene (RGS2) and increase in AKR1C3 may be the contributory path to chronic inflammation leading to metaplasia. This pathway is proposed to be a mechanism leading to carcinogenesis in skin keratinocytes. The observed over expression of IGFL1 may be a means of triggering carcinogenesis in HaCaT keratinocytes. In conclusion, it was established that at very low doses, arsenic is genotoxic and induces aberrations in gene expression though it may appear to enhance cell proliferation. The expression of two genes encoding membrane proteins IL1R2 and TNFSF18 may serve as possible biomarkers of skin keratinocytes intoxication due to arsenic exposure. This research provides insights into previously unknown gene markers that may explain the mechanisms of arsenic-induced dermal disorders including skin cancer / Environmental Sciences / D. Phil. (Environmental science)