Bilirubin- and light induced cell death in a murine lymphoma cell line.

Cells from the mouse lymphoma cell line L5178Y-R were exposed to blue light from phototherapy lamps in the presence of solutions of 160 microM bilirubin supplemented with serum albumin. HPLC analysis showed that the bilirubin solution was photooxidised as a function of increasing light dose. The cells were stained with trypan blue to score necrosis, and apoptosis was assayed by the terminal deoxynucleotide transferase assay (TdT) or by studying the nuclear structure in cells stained with propidium iodide. A rapidly developing apoptosis was observed after light doses killing 60-80% of the cells as judged from the trypan blue exclusion test. The fraction of apoptotic cells was smaller than the fraction of necrotic cells. Exposure of the cells to fractions of light at a high dose rate was compared to the effect of the same total dose at a lower dose rate given as a single fraction. No large differences were found, however, there was a tendency of a higher degree of necrosis as well as apoptosis in the cells receiving the light in fractions at a high dose rate.

Cells, bilirubin and light: formation of bilirubin photoproducts and cellular damage at defined wavelengths.

Cultured cells from one human and one murine cell line were treated with bilirubin and irradiated with visible light of different wavelengths, either from phototherapy lamps or from a Xenon/Mercury lamp equipped with a monochromator. Bilirubin bound to human serum albumin was also irradiated with light. After irradiation, the bilirubin and its photoisomers were extracted and analysed with High Pressure Liquid Chromatography. The formation of single strand breaks in the DNA of treated cells was studied using a fluorescence marker. Cytotoxicity in the mouse skin cell line was measured by loss of the ability to form visible colonies in vitro. Green light exposure favours the production of lumirubin, while blue light causes more DNA damage and cytotoxicity. Green light may be more efficient and safer than shorter wavelength exposure when treating jaundiced newborns with phototherapy.

Bilirubin bound to cells does not form photoisomers.

Cultured cells from one human and one murine cell line were incubated with bilirubin by different methods that allowed bilirubin to be bound to cells. The cells were irradiated with visible light of different wavelengths. Bilirubin bound to human serum albumin was also irradiated with light. After irradiation, bilirubin and its photoisomers were extracted and analyzed by HPLC. No photoisomers were found in samples of irradiated cells, while the types and amounts of photoisomers that were expected from the literature were found in samples of irradiated bilirubin/albumin mixtures. We conclude that the formation of therapeutically active photoisomers during phototherapy most probably does not take place in skin cells, but most likely in bilirubin bound to albumin in the vessels or in the interstitial space.

Impairment in drivers due to cannabis in combination with other drugs.

Blood samples from 425 suspected drugged drivers who were clinically impaired and negative for alcohol were analysed. Fifty-six percent of the samples were positive for tetrahydrocannabinol (THC). Tetrahydrocannabinol-positive blood samples were analysed for amphetamines, barbiturates, benzodiazepines, cocaine metabolites and opiates. Eighty-two percent of the samples were found to be positive for one or more drugs in addition to THC, and the concentrations of these drugs were often high. Thus, THC in combination with other drugs seems to be a much more frequent reason for impairment than THC alone among Norwegian drugged drivers.

Single-strand breaks in the DNA of human cells exposed to visible light from phototherapy lamps in the presence and absence of bilirubin.

Clinical evidence indicates that phototherapy of hyperbilirubinaemia in newborn infants is a safe and efficient form of therapy. The short-term side effects are not serious and seem to be well controlled. There are few long-term follow-up studies of phototherapy-treated infants. Therefore one cannot completely exclude the possibility that side effects can be found in future studies. With this background we undertook the present study of possible genotoxic effects of phototherapy. Human cells of the established glioblastoma cell line TMG-1 were used. The cells were exposed to visible light in the presence of different concentrations of bilirubin or in the absence of bilirubin. DNA was unwound in alkaline solution and the induction of strand breaks was assayed by a method taking advantage of the fluorescence from the dye Hoechst 33258. Blue light induced single-strand breaks in the DNA of cells in culture in the absence of bilirubin. During irradiation of bilirubin solutions with blue and green phototherapy light, long-lived toxic photoproducts were formed under in vitro conditions. At high and clinically relevant bilirubin concentrations, the effects of blue and green light were relatively similar. At low concentrations, there was a smaller effect of green light as expected from the absorption spectrum of bilirubin. It remains to be seen whether the genotoxic effect observed in the present studies can occur in vivo.