Alkaline sucrose sedimentation analysis as an indicator of repair capability of xeroderma pigmentosum fibroblasts for 4-nitroquinoline-1-oxide damage

Abstract

4-Nitroquinoline-1-oxide (4NQO) damage to DNA and its repair in normal and xeroderma pigmentosum (XP) fibroblast stains were followed by sedimentation in an alkaline sucrose gradient. Two forms of analysis were employed. In the brief lysis technique, the cells were exposed to alkali for 30 min before centrifuging. In the long lysis technique, the cells were exposed to alkali overnight before centrifuging. The fibroblast cultures were derived from 6 normal individuals, 2 XP variants, 2 from complementation group C and 1 each from complementation groups A, B and E. by means of the brief lysis technique all of the fibroblasts were found to repair the 4NQO induced damage. By means of the long lysis technique the fibroblasts from normal individuals, from XP variants and from XP group E were found to repair the 4NQO induced damage. The fibroblasts from groups A, B and C showed no repair of the lesion. This latter result, on the repair capability of 4NQO damaged XP cells, correlates with the repair capability of XP cells shown by other methods. It suggest that if sedimentation analysis in alkaline sucrose is to be used to demonstrate the induction of chemically induced damage to DNA and its repair, both brief and long lysis periods should be employed.