Mutations of mitochondrial (mt) DNA play a role in neurodegeneration, normal aging, premature aging of the skin (photoaging), and tumors. We and others could demonstrate that mtDNA mutations can be induced in skin cells in vitro and in normal human skin in vivo by repetitive, sublethal ultraviolet (UV)-A-irradiation. These mutations are mediated by singlet oxygen and persist in human skin as long-term biomarkers of UV exposure. Although mtDNA exclusively encodes for the respiratory chain, involvement of the energy metabolism in mtDNA mutagenesis and a protective role of the energy precursor creatine have thus far not been shown. We assessed the amount of a marker mutation of mtDNA, the so-called common deletion, by real-time PCR. Induction of the common deletion was paralleled by a measurable decrease of oxygen consumption, mitochondrial membrane potential, and ATP content, as well as an increase of matrix metalloproteinase-1. Mitochondrial mutagenesis as well as functional consequences could be normalized by increasing intracellular creatine levels. These data indicate that increase of the energy precursor creatine protects from functionally relevant, aging-associated mutations of mitochondrial DNA.

In photoaged skin, wrinkles result from an increased degradation and a decreased de novo synthesis of collagen fibers. At the molecular level, photoaged skin is characterized by increased amounts of large-scale deletions of the mitochondrial (mt) genome such as the 4,977 bp common deletion. The common deletion can be generated in dermal fibroblasts through repetitive ultraviolet (UV) A irradiation, and this was found to be associated with an increased expression of the collagen-degrading enzyme matrix metalloproteinase-1 (MMP-1). These studies did not clarify whether increased MMP-1 expression was caused by a disturbance of mtDNA integrity or whether it occurred independently. We have therefore generated a phenocopy of cells bearing large-scale deletions of mtDNA by gradually depleting the mtDNA from unirradiated human skin fibroblasts. Gradual depletion of mtDNA caused a gene expression profile, which was reminiscent of that observed in photoaged skin. Accordingly, in these cells an increased expression of MMP-1 without a concomitant change in tissue inhibitor metalloproteinase-1 as well as a decreased expression of collagen type 1 alpha-1, that is, a gene involved in collagen de novo synthesis, was observed. This altered gene expression resulted from intracellular, mitochondria-derived oxidative stress. Our results support the concept that disruption of mt integrity, for example, by UV-induced mtDNA mutagenesis, is of pathogenetic relevance for photoaging of human skin.

Xeroderma pigmentosum (XP) is a genetic disorder characterised by hypo-/hyperpigmentation, increased sensitivity to ultraviolet (UV)-radiation and an up to 2000-fold increased skin cancer risk. Cells from XP-patients are defective in nucleotide excision repair (NER) responsible for repair of UV-induced DNA damage. This defect accounts for their mutator phenotype but does not predict their increased skin cancer risk. Therefore, we carried out array analysis to measure the expression of more than 1000 genes after UVB-irradiation in XP cells from three complementation groups with different clinical severity (XP-A, XP-D, XP-F) as well as from patients with normal DNA repair but increased skin cancer risk (Ն2 basal or squamous cell carcinoma at age Ͻ40yrs). Of 144 genes investigated, 20 showed differential expression with p Ͻ 0.05 after irradiation of cells with 100 mJ/cm 2 of UVB. A subset of six genes showed a direct association of expression levels with clinical severity of XP in genes affecting carcinogenesis relevant pathways. Genes identifi ed in XP cells could be confi rmed in cells from patients with no known DNA repair defects but increased skin cancer risk. Thus, it is possible to identify a small gene subset associated with clinical severity of XP patients also applicable to individuals with no known DNA repair defects.