Dietary Calorie Restriction from Adulthood Through Old Age in Rats: Improved DNA Polymerase β and DNA Gap Repair Activity in Cortical Neurons.

It is well established now that dietary calorie restriction (CR) leads to extension of life span in many species, although the exact mechanism of this effect is still eluding. In the present study, we examined the effect of 40 % CR imposed during a prolonged period of life span (from 6 to 30 months) of rats on the activity of DNA polymerase β (pol β) in view of its role in short gap base excision DNA repair and template driven primer extension. DNA pol β activity is very low at this late age.

Topoisomerase IIβ regulates base excision repair capacity of neurons.

Topoisomerase IIβ (TopoIIβ), an enzyme involved in DNA rearrangements, is predominantly present in brain and its levels are shown to decrease with age. This study characterizes the function of TopoIIβ in regulating BER (base excision repair) activity. TopoIIβ deficient granule neurons (CGNT⁻) show greater sensitivity to N-ethyl N-nitroso urea (ENU)-mediated DNA damage.

Age-dependent decline of DNA base excision repair activity in rat cortical neurons.

Synthetic oligonucleotide duplexes containing a single uracil (U) or 8-oxoguanine (8-oxoG) were used as a model substrates to assess the base excision repair (BER) ability of neuronal extracts prepared from the cerebral cortex of young (7 days), adult (180 days) and old (720 days) rats. Our results demonstrate that BER activity in neurons markedly declines with age. The decline in BER could be attributed to decrease in the expression levels and activities of BER enzymes.

Age-related decline in DNA polymerase β activity in rat brain and tissues.

Fidelity of DNA polymerases is vital for maintaining genomic integrity. Deficient DNA repair leads to age related disorders or cancer. If the age at which the decline in activity of predominant DNA repair enzymes starts is identified, and the deficient proteins supplemented, then the manifestation of these diseases can be delayed promoting healthy aging.

Studies on the molecular correlates of genomic stability in rat brain cells following Amalakirasayana therapy.

Adult Wistar NIN (WNIN) rats (6 months old) of both sexes were orally fed Amalakirasayana at a dose of 4.5 g per kg body weight, five days in a week. The Amalakirasayana was prepared by Arya Vaidya Sala, Kottakkal, Kerala, India, which is considered as gold standard.

Study of DNA damage via the comet assay and base excision repair activities in rat brain neurons and astrocytes during aging.

Earlier we have used biochemical approach to assess the number of single (SSBs) and double (DSBs) strand breaks in brain cellular DNA. However, a quick method to obtain a reliable measure of DNA damage in cells was in need for population studies. Therefore, single cell gel electrophoresis technique (popularly known as "comet" assay) has been standardized using the Trevigen protocol.

Free radical induced oxidative damage to DNA: relation to brain aging and neurological disorders.

Free radicals are produced in cells by cellular metabolism and by exogenous agents. These species react with biomolecules in cells and one of the important targets is DNA. This kind of damage, often referred to as oxidative DNA damage, has consequences in various organs and particularly in brain, in view of its high metabolic activity and oxygen consumption.

On the inhibitory affect of some dementia drugs on DNA polymerase Beta activity.

Some drugs are routinely prescribed for dementia that sets in either due to normal ageing or due to neurodegenerative disorders. We have studied the effect of three of these drugs, Donepezil hydrochloride, Rivastigmine tartrate and Nootropyl, on the activity of DNA polymerases beta, a crucial enzyme in the base excision repair pathway, the most important mode of DNA repair in brain. All the three drugs inhibited DNA polymerase beta activity to varying degrees although the affects of Donepezil being the least and inconsistent.

Mechanisms of disease: DNA repair defects and neurological disease.

In this Review, familial and sporadic neurological disorders reported to have an etiological link with DNA repair defects are discussed, with special emphasis placed on the molecular link between the disease phenotype and the precise DNA repair defect. Of the 15 neurological disorders listed, some of which have symptoms of progeria, six--spinocerebellar ataxia with axonal neuropathy-1, Huntington's disease, Alzheimer's disease, Parkinson's disease, Down syndrome and amyotrophic lateral sclerosis--seem to result from increased oxidative stress, and the inability of the base excision repair pathway to handle the damage to DNA that this induces. Five of the conditions (xeroderma pigmentosum, Cockayne's syndrome, trichothiodystrophy, Down syndrome, and triple-A syndrome) display a defect in the nucleotide excision repair pathway, four (Huntington's disease, various spinocerebellar ataxias, Friedreich's ataxia and myotonic dystrophy types 1 and 2) exhibit an unusual expansion of repeat sequences in DNA, and four (ataxia-telangiectasia, ataxia-telangiectasia-like disorder, Nijmegen breakage syndrome and Alzheimer's disease) exhibit defects in genes involved in repairing double-strand breaks.

DNA double strand break repair in brain: reduced NHEJ activity in aging rat neurons.

Linearised pUC 19 DNA with cohesive, blunt and non-matching ends, generated by prior treatment with different restriction enzymes was presented as substrate to measure the NHEJ activity to repair DNA double strand breaks in extracts prepared from isolated neurons from neonatal, young adult and old rat cerebral cortex. Highest end joining activity was noticed with the substrates having cohesive 3' overhang (PstI) or 5' overhang (EcoRI) ends and this activity is significantly reduced with age. However, blunt and non-matching ends were very poorly repaired at all ages.

The presence of 3'-5' exonuclease activity in rat brain neurons and its role in template-driven extension of 3'-mismatched primers by DNA-polymerase beta in aging neurons.

A three-step reaction strategy has been developed to examine the mechanism of extension of a mismatched primer in an oligoduplex substrate by rat neuronal extracts and DNA polymerase beta. The results revealed that in the case of duplexes with a mismatch at 3'-end of primer, significant extension by DNA polymerase beta has taken place only after the removal of the mismatched base, thus indicating the presence of a proof reading 3'-5' exonuclease activity in neuronal extracts of all ages. A closer examination of the neuronal exonuclease activity revealed that bases are excised from the 3' end in a sequential and nonspecific manner, although initial excision of a mismatched base was slightly faster.

Dietary calorie restriction, DNA-repair and brain aging.

It is now well established, in many species, that dietary calorie restriction confers beneficial effects like slowing down many age dependent processes and extending the lifespan. There are indications that this phenomenon may be applicable even in non-human primates and humans. However the precise mechanism through which these effects are achieved is not known.

DNA-polymerase alpha, beta, delta and epsilon activities in isolated neuronal and astroglial cell fractions from developing and aging rat cerebral cortex.

The relative proportions of DNA-polymerases alpha, beta, delta and epsilon (pols alpha, beta, delta and epsilon ) activities in isolated neuronal and astroglial cell fractions from developing, adult and aging rat brain cerebral cortex, were examined. This was achieved through a protocol that takes advantage of the reported differential sensitivities of different DNA-polymerases towards certain inhibitors like butylphenyl and butylanilino nucleotide analogs, 2',3'-dideoxythymidine triphosphate (ddTTP), monoclonal antibody of human alpha polymerase and the use of two template primers as substrates. The results indicate that while DNA-polymerase beta (pol beta) is the predominant enzyme, significant levels of DNA-polymerases alpha and delta/epsilon (pols alpha and delta/epsilon ) are also present in both cell types at all the post-natal ages studied.