Norwegian Radiation Protection Authority

Results

Some single strand breaks are induced in the cells by blue light irradiation alone, and the amount of DNA-damage induced by light is increased in TMG-1 cells pre-incubated with SnPP or ZnPP (Table 1). DNA-damage is induced much more efficiently by ZnPP than by SnPP. The explanation for this observation may be either that ZnPP produces singlet oxygen more efficiently or that more ZnPP is taken up by the cells. The uptake of porphyrins was measured in cell cultures containing approximately the same number of cells and the same concentration of cellular protein. Seventeen times more ZnPP was bound to the cells compared to SnPP when both had been present in the cell culture medium at a concentration of 32 mM for 1 h (data not shown). The high uptake of ZnPP (10 mM incubated for 1 h) is illustrated in Fig. 1. It can be observed that the ZnPP fluorescence is located in distinct spots in the peri-nuclear region of the cells and that the fluorescence intensity is different in different cells. In order to observe fluorescence at all from SnPP the cells had to be incubated with 100 mM for 3 h. The fluorescence from SnPP observed in the fluorescence microscope was diffusely distributed in the cells.

None of the Cr-derivatives produced detectable increases in the frequency of single strand breaks when they were tested under the same conditions as SnPP and ZnPP in Table 1. The same conclusion could be drawn from experiments where the cells were not preincubated with porphyrins, but were exposed to light in the presence of 8 mM of the Cr-derivatives in PBS (data not shown).

[Up]

The DNA-damage increases with the concentration of SnPP and ZnPP (Table 1) as well as with the incubation time in the presence of porphyrins, probably due to a higher uptake of porphyrins (data not shown).

Since different photosensitizers may increase the photooxidation of bilirubin (19), and since bilirubin is a potent scavenger of oxygen radicals (13,14,15), it was of interest to study the photosensitised damage to cells in the presence of metalloporphyrins and bilirubin together.

To assay the phototoxicity of the metalloporphyrins alone and bilirubin alone and in combination, the 308-cells (mouse epidermis) were incubated in growth medium in the presence of 5 m M metalloporphyrin for 1 h. The medium was removed and PBS containing BSA with a concentration sufficient to give an albumin concentration of 10 mM was added. In some of the samples 10 mM bilirubin was dissolved in this solution. Cells that had not been preincubated with metalloporphyrins were used as a control of the effect of light alone and bilirubin alone. Figure 2 shows that light alone does not induce any cell inactivation at the two doses shown. Addition of bilirubin induces a small degree of cell inactivation in light exposed samples (significant at the highest light dose, p<0.05). Bilirubin did not induce any dark toxicity under the conditions used. Both metalloporphyrins showed a dose dependent increase in cell inactivation. Upon addition of bilirubin to the metalloporphyrin labelled cells, the cell in- activation increased compared to the effect of metalloporphyrins alone. At the highest light dose the bilirubin-induced increase in inactivation was significant for both SnPP and ZnPP (p<0.05).

All the porphyrins tested caused photooxidation of tryptophan. The rate constants for the chromium derivatives (Tab. II) were smaller than for the other derivatives, but significantly different from 0 (p<<0.05).

[Up]

[Previous] [Next] [Index]