Tissue-specific deletion patterns of the mitochondrial genome with advancing age

doi:10.1016/j.exger.2006.03.010

Experimental Gerontology

Volume 41, Issue 5, May 2006, Pages 518-524

https://doi.org/10.1016/j.exger.2006.03.010 Get rights and content

Abstract

Aging is a multifactorial process and a lot of theories have been put forward to explain the deterioration of organ function with advancing age. The free radical hypothesis developed by Harman is amongst the most prominent today and has been focused on mitochondrial aging in the last decades. Applying a long PCR approach we screened human skeletal muscle, heart, caudate nucleus and cerebellum of 50 individuals for large-scale deletions of mitochondrial DNA (mtDNA). The most important observation of our study was the detection of age dependent tissue specific deletion patterns of mtDNA. The pattern of the same tissue of different individuals was more similar than the pattern of different tissues of the same individuals. Whereas deletions were barely detectable in cerebellar tissue, in caudate nucleus a specific banding pattern with deletions of 4–8 kb was already observed around the age of thirty. However, the increase of these large-scale deletions in number and variety over lifetime was more pronounced in skeletal muscle or heart. Our data support the notion that different tissues accumulate mtDNA damage in a specific manner. Although functional consequences of mitochondrial deletions are clearly supported by experimental data on the single-cell level in model organisms and mammals, their role regarding impaired function of organs with advancing age in humans remains unresolved.

Introduction

Aging is a multifactorial process, which leads to a decreased production of ATP from energy-rich compounds in different organs and is expressed as loss of cognitive ability and memory, cardiovascular function or muscular strength. The decreased production of ATP should be reflected on the level of the mitochondria, because mitochondria are the powerplants of the cell, where in the respiratory chain various metabolic substrates are finally oxidized and converted into ATP for energy supply. But up to 5% of mitochondrially consumed oxgen is converted to reactive oxygen species or free radicals generated as byproducts of oxidative phosphorylation (Chance et al., 1979, Richter, 1995). These free radicals can induce oxidative damage to many proteins, membrane lipids and especially nucleic acids (Harman, 1956). Mitochondrial DNA (mtDNA) is at a much higher risk for mutations, because it exhibits less efficient repair mechanisms and a close vicinity to the respiratory chain where free radicals are released (Ballard and Dean, 2001).

The mitochondrial genome is a double stranded ring with 16,569 bp in length that codes for 13 subunits of the electron transport chain (ETC), 22 tRNAs, and 2 rRNAs (Graeber et al., 1998). MtDNA is transcribed as a whole, with one primary transcript of the heavy and the light strand, starting at the origins of replication O_H and O_L. Oxidative damage to single base pairs inflicted by free radicals can lead to deletions or other rearrangements of mtDNA, because the level of 8-OHdG has been correlated with large-scale mitochondrial DNA deletions (Lezza et al., 1999, Albers and Beal, 2000). At first it has been proposed, that deletions can arise from slippage mispairing of direct repeats during replication; either directly (Shoffner et al., 1989, Elson et al., 2001), or indirectly through large-scale duplications, subsequent intramolecular recombination and loss of the duplicated regions (Wallace, 1997). Furthermore, occurrence of deletions has been linked to mutations of polymerase gamma, adenine nucleotide translocator and the helicase twinkle (Zeviani et al., 2003). More recently experimental evidence has been presented indicating that multiple mtDNA deletions may be promoted by double strand breaks (Prado et al., 2003, Srivastava and Moraes, 2005).

Since, the late 1980s studies about rearrangements of mitochondrial DNA have been reported and opened a new field of research. Interestingly, these deletions increase in an age dependent manner and are supposed to impair the ATP production of the cell, because coding sequences of the respiratory chain are removed. The earliest reports focused on a few single rearrangements, especially the 4977 bp deletion which has been studied in a lot of different tissues (Cortopassi et al., 1992). Upon screening for other large scale deletions of mitochondrial DNA in skeletal muscle or human heart of old age subjects, only a few or no molecules of normal length were detectable (Melov et al., 1995, Hayakawa et al., 1996, Kovalenko et al., 1997). Unfortunately, most studies regarding these rearrangements of mtDNA include only one tissue and only a few cases. It is very important to study different tissues of the same individuals, because between organ systems the susceptibility to mutations and the functional consequences can differ markedly (Kajander et al., 2000). The aim of our study was to compare the banding pattern of mitochondrial deletions in human caudate nucleus, cerebellum, skeletal muscle, and heart between 50 individuals.

Section snippets

Samples

Ten to thirty milligrams tissue from caudate nucleus, cerebellum, posterior wall of the left chamber of the heart, and iliopsoas muscle of 50 individuals was collected at autopsy. To exclude mitochondrial or degenerative diseases a complete microscopic examination of each tissue sample was performed. Only individuals who died of acute or peracute causes (stabbing, suffocation, drowning) were included in the study. Care was taken not to exceed an interval of more than 48 h between death and

Results

All the tissues reveal a panel of different mitochondrial deletions, which increase in number and variety over lifetime. For exclusion of PCR artefacts, called jackpots, each PCR was performed for three times. Jackpots result, when mutations arise during the early circles of the PCR due to a lack of processivity of the polymerase leading to stuttering at sequences of low complexity like homopolymer tracts (Hutchison et al., 2005). As indicated in Fig. 1 the banding pattern of the three

Discussion

Screening for large scale deletions in four different tissues of 50 individuals exhibit an age-dependent increase of somatically acquired mtDNA deletions in the four tissues. In cerebellar tissue this increase was less pronounced. Additional signals were detectable only in some single individuals, especially in the group older than 80 years. The low amount of deletions detectable in cerebellar tissue may be due to a higher expression of superoxide dismutase and catalase RNA as has been

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Tissue-specific deletion patterns of the mitochondrial genome with advancing age

Abstract

Introduction

Section snippets

Samples

Results

Discussion

Curr. Opin. Genet. Dev.

Biochem. Biophys. Res. Commun.

Biochem. Biophys. Res. Commun.

Am. J. Hum. Genet.

Mech. Ageing. Dev.

Biochem. Biophys. Res. Commun.

Mutat. Res.

Biochem. Biophys. Res. Comm.

Free Radic. Biol. Med.

Exp. Gerontol.

Int. J. Biochem. Cell Biol.

Curr. Opin. Genet. Dev.

Mitochondrial dysfunction and oxidative stress in aging and neurodegenerative disease

J. Neural. Transm.

Mitochondrial abnormalities are more frequent in muscles undergoing sarcopenia

J. Appl. Physiol.

Hydroperoxide metabolism in mammalian organs

Physiol. Rev.

Mitochondrial DNA deletions in human brain: regional variability and increase with advanced age

Nat. Genet.

A pattern of accumulation of a somatic deletion of mitochondrial DNA in aging human tissues

Proc. Natl Acad. Sci.