Bilirubin‐induced cell death during continuous and intermittent phototherapy and in the dark

Abstract

Aim: To compare continuous and intermittent light exposure in the presence of bilirubin with respect to cellular damage. Furthermore, it was of interest to characterize the nature of cellular toxicity of bilirubin in the dark. Method: A murine lymphoma cell line, L5178Y‐R (LY‐R), was exposed to solutions of bilirubin (160 μM) supplemented with human serum albumin (200 μM) and irradiated with phototherapy light (Philips 20W/52) at a constant total dose of approximately 500 kJ/m2. The irradiation was given either as intermittent or continuous treatment with light of variable irradiance. The three lower irradiance levels were clinically relevant. Cells treated with bilirubin were also kept in the dark for various periods of time. Cell toxicity was determined by measuring apoptosis and necrosis. Apoptosis was measured by terminal deoxynucleotide transferase and propidium iodide staining assay, while trypan blue assay was used for detection of necrosis. Results: There was no difference (n=6, p>0.05) between continuous and intermittent irradiation in the induction of early and late apoptotic cell death. Necrosis was more pronounced after intermittent treatment. Bilirubin dark toxicity was observed and classified as both apoptotic and necrotic. Conclusion: Continuous and intermittent light exposure caused the same degree of apoptotic cell death, while the cells underwent more necrotic death after intermittent exposure. Bilirubin was cytotoxic in the dark by both cell death mechanisms.

Formation of photoproducts and cytotoxicity of bilirubin irradiated with turquoise and blue phototherapy light

“…The multiplication of cells exposed to 24 h of green irradiation was comparable to the multiplication of cells exposed to 12 h blue irradiation. Based on data from previous studies [13,16], it may be concluded that the cellular effects are induced by a phototoxic effect of BR. Hydrogen peroxide and other peroxides can be formed in irradiated solutions of BR and cause the observed cytotoxicity [5,6].…”

“…Photosensitizing effects of bilirubin (BR) on cells in culture have been shown previously [5,[12][13][14][15]. Studies on mouse lymphoma cells, L5178Y-R (LY-R), have shown that BR in the dark and in combination with blue light can induce apoptosis in this cell line [15,16].…”

Formation of photoproducts and cytotoxicity of bilirubin irradiated with turquoise and blue phototherapy light

“…Photosensitizing effects of bilirubin (BR) on cells in culture have been shown previously 512–15. Studies on mouse lymphoma cells, L5178Y‐R (LY‐R), have shown that BR in the dark and in combination with blue light can induce apoptosis in this cell line 15, 16.…”

Phototherapy: Old questions, new answers

“…Why does phototherapy fail in some patients with hereditary spherocytosis and hemolytic jaundice? In this month's issue, Bruzell Roll et al present three studies that shed some light on these areas 3–5.…”

“…Others have argued that it is only the total dose (irradiance multiplied by irradiation time) that matters. In an in vitro study using murine lymphoma cells to compare the effects of continuous versus intermittent phototherapy on cytotoxicity, cells were treated with bilirubin and human serum albumin and exposed to blue light at four different irradiance levels from 3 to 18 h to achieve the same total irradiation dose 3. Increased apoptosis and necrosis were noted with longer exposure periods, irrespective of irradiance, and was possibly due to increased photo‐oxidation and cell damage.…”