Combining Three In Vitro Assays For
Detecting Early Signs Of UVB Cytotoxicity
In Cultured Human Skin Fibroblasts
Author(s)
C. Khalil
Abstract
The aim of this study was to determine the most sensitive approach for detecting
the early signs of UVB-induced cellular damage using human skin fibroblasts.
UVB-induced cell damage was assessed immediately and 24 h post irradiation
using 3 in vitro colorimetric assays: neutral red (NR); 5-(3-carboxy-methoxyphenyl)-
2-(4.5-dimethylthiazolyl)-3-(4-sulphonyl) MTS; and (iii) LDH
enzyme release. A good correlation was observed immediately post exposure
between the MTS and NR in measuring damage levels, but was lost 24 h post
exposure. This loss of correlation was the result of delayed expression of
lysosomal damage and led to investigating cell membrane damage using LDH
cell leakage assay. LDH levels observed immediately post UVB irradiation
indicated significant LDH release at exposure doses of 2.2 and 2.8 J/cm 2 , while
LDH release reported 24 h post exposure was recorded for doses as low as 0.70
J/cm 2 . The data reported in this paper indicated that cell viability and damage
were significantly affected in a dose dependent manner as a result of exposure
doses. The assays used displayed different sensitivities in detecting damage with
the earliest signs of cellular UVB damage best-measured 24 h following
exposure using the LDH assay. Furthermore UVB contributed to denaturing the
cellular LDH released during irradiation. Therefore the use of LDH cytotoxicity
based assays with UVB exposure must be considered with extreme care.
Keywords: cells, viability, MTS, neutral red, UVB, LDH, cytotoxicity.
Keywords
cells, viability, MTS, neutral red, UVB, LDH, cytotoxicity.
Related Book
Other papers in this volume
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