Age-correlated DNA damage in human muscle tissue

Abstract

This investigation represents the largest study so far published on human DNA damage and aging. The subject of this investigation is damage, determined as DNA alterations which give rise to complete molecular breaks in the course of treatment of purified DNA solutions with single-strand-specific nucleases. The DNA is derived from milligram samples of human muscle of individuals mostly undergoing surgical treatment. Care has been taken to bring the muscle samples, once shut off from blood circulation to liquid nitrogen temperatures within few seconds. The DNA is prepared by a procedure keeping breaks by handling and by DNAase attack as low as possible, however pushing DNA purity, especially with respect to protein as high as possible. Highly purified DNA treated in this way has some sites which are susceptible to single-strand (ss) specific DNAase splitting (ss-events). Three different deoxyribonucleases have been used: Nuclease S 1, Nuclease BAL31 and Pea Endo-Nuclease. They give very similar results, i.e. splitting of the DNA so as to yield DNA pieces of given distribution. The lengths of these double-strand (ds) pieces have been determined from their electron microscopical pictures, either by following the image contours with a magnetostrictive stylo of the projected photo on a pad, by following the contours with a mileage ruler, or by integrating the silver grains on the photo. The molecular weight averages of the ds DNA threads between two ss-events for each individual have been determined from 20 to 200 molecules.