Why Do We Age? What Do Dolly'sTelomeres Tell Us?
Two Theories Linking DNA Damage and Aging: Free Radical/Oxidation vs.Telomere Shortening

Ruwaida Dhala-Vakil, BSc, MSc

There are many factors involved in human aging, and significantprogress has been made in this field over the last few decades.Recent evidence from cloned calves suggests that scientists may notmerely be able to reverse the cellular damage accumulating withage--they may, in fact, be able to prolong cell life. The cells fromthese cloned animals lived longer in culture and had longer telomeresthan their normal counterparts. If this extension of the cellularlife span can be translated into longer life for the entire organism,the calves may live fifty percent longer than normal.¹Additionally, studies on antioxidants show that transgenic Drosophila(the fruit fly), which overexpress antioxidant genes, live 34% longerthan controls.² This article will focus on both the freeradical/oxidation theory of aging, and the role of telome-rase inaging.

Free Radical/Oxidation Theory of Aging
In 1956, Denham Harman suggested that there is an age-relatedaccumulation of reactive oxygen species (ROS) which causes damage tocellular components. The damage is targeted to the proteins and DNAin the nucleus and mitochondria, as well as to the proteins andlipids in the cell membrane, and the proteins of the cytoplasm.Mitochondrial DNA is located near the inner mitochondrial membrane,close to the sites where free radicals form.tochondrialDNA more susceptible to damage due to ROS. Additionally,mitochondrial DNA lacks advanced DNA repair mechanisms and thereforeis unable to counter-act ROS-induced damage. As a result, it is moreaffected by ROS damage than is nuclear DNA. With increasing age,there is an increase, in mammalian cells, of accumulated geneticdamage, that can lead to a decrease in mitochondrial activity in thecells of organs such as the heart and kidney.

Oxidative damage is also one of the factors involved in thepathogenesis of age-related disorders such as Alzheimer'sdisease.³ Neurons appear to be particularly susceptible tofree radicals. Post-mortem examination of the brains of Alzheimer'sdisease patients show many signs of free radical attacks such asdamaged mitochondrial and nuclear DNA, lipid peroxidation and proteinoxidation. Oxidative damage is also one of the factors involved inthe pathogenesis of age-related disorders such as cancer andarteriosclerosis.³

ROS are generated by metabolic processes. Two to three percent ofthe oxygen that is taken up is reduced by the addition of electronsand converted into ROS. These ROS include the superoxide anions,hydrogen peroxide and the hydroxyl radical.² These freeradicals are atoms or molecules that have at least one unpairedelectron, rendering them chemically unstable and highly reactive withother molecules in the body. As a result, the free radicals andreactive oxidants cause extensive damage to both DNA and tissues. Atthe cellular level, DNA damage has a negative impact on the processesof transcription and replication, and causes cell cycle arrest,apoptosis and the accumulation of genomic mutations (genomicinstability). Although this damage can be repaired by DNA repairmechanisms, some damage accumulates due to the inability of therepair mechanisms to correct the defects as quickly as theyarise.

Cells respond to free radical oxidative damage by neutralizingoxygen radicals through antioxidant enzymes, and through othercellular defense mechanisms such as DNA repair. The DNA repairmechanisms involved in the removal of oxidative DNA lesions or DNAbase modifications include base excision repair and nucleotideexcision repair. Base excision repair involves the removal of smallbase modifications, and nucleotide excision repair involves removalof bulky helix distorting additions. At some point the accumulationof free radical damage overwhelms the DNA repair mechanisms. Thisaffects cellular functions such as growth and energy production and,in some cases, leads to cell death.⁴

Role of Telomerase in Aging
Telomeres are repeating DNA sequences located at the ends ofeukaryotic chromosomes. Telomeres and telomerase help maintainchromosome integrity by preventing the loss, during cell division, ofgenes that are required for cell function. Telomerase is aribonucleoprotein enzyme that extends the telomeres by addinghexameric nucleotide repeats to the ends of chromosomes. Without theactivity of telomerase, telomeres continue to shrink during cellulardivision, thereby losing genes that are important for cellfunction.⁷ Once the telomere reaches a critical length,the cell stops growing. This phenomenon is termed replicative orcellular senescence.²

The shortening of telomeres results from DNA replication and aprocess known as the 'end replication problem'. The mechanism ofreplication for the leading and lagging strands of the DNA doublehelix is different. The lagging strand is synthesized as a series offragments, each of which requires a new RNA primer to initiatesynthesis. Since there is no DNA beyond the end of the chromosome toserve as the template, the gap between the final lagging strand andthe end of the chromosome cannot be filled in. Thus, the 5' end ofthe lagging strand will lose some nucleotides each time a cellreplicates.⁵ This results in the progressive shortening oftelomeres upon cellular division.

In1961, Leonard Hayflick discovered that cells grown in culture woulddivide a limited number of times before they lost the ability todivide and died.⁸ This was suggestive of a link betweencellular senescence and cell death. The mechanism responsible forcell death was unknown. In 1986, Howard Cooke noticed that thetelomeres of reproductive cells were longer than the telomeres ofsomatic cells. Since telomeres shorten each time a cell divides, hespeculated that somatic cells do not make telomerase. This wouldexplain the shortening of telomeres in somatic cells as they divide.After a certain number of cell divisions the telomeres would reach acritical length, preventing the cell from further replication,resulting in cellular senescence.⁷ Evidence from in vitroculture and in vivo data proves that telomeric DNA is lost from humancells over the course of many cell divisions.^9,10,11

Recent evidence from a study in Science points to a direct linkbetween telomerase and cellular aging. In this study the researchersintroduced a genetic component that activated telomerase into twotelomerase-negative, normal, human cell types--retinal pigmentepithelial cells and foreskin fibroblasts. These cells remainedhealthy and continued to divide without senescing, exceeding normalcellular life span by at least 20 replications.¹² Bycontrast, data from a mouse study showed that the mice, in which theessential RNA subunit of telomerase had been deleted, had cells thatcould undergo malignant transformation. It is possible that in micethe mechanisms underlying senescence and tumor regulation differ fromthose in humans.¹³ Therefore it is possible that slowingdown the rate of telomere shortening in humans could slow down theaging process.

Evidence to support the role of telomeres in cancer comes from invitro studies of immortal human cell lines. These cell lines havelong telomeres and active telomerase.^15,16 Removing humantelomerase activity using antisense RNA causes the cancer cell to dieafter only 23-26 divisions.¹⁷ Since the likelihood ofgetting most cancers increases with age, slowing down the rate oftelomere shortening to try and delay the aging process may increasethe risk of cancer.¹² Therefore, in attempting to reversethe aging process we may increase the risk of cancer.

Conclusion
Although aging is a complicated process, scientists are now beginningto tease apart some of the variables involved. It was discovered thatthe chromosomes of Dolly, the cloned sheep, had shorter telomeresthan the chromosomes of her siblings. This suggested that Dolly mightbe aging more rapidly than her siblings. However, further researchfrom the cloning of cattle showed that the telomeres of these animalswere longer or the same length as those of their normal siblings. Inaddition, these animals were cloned from cells that had undergone asmany as 50 doublings, whereas Dolly was cloned from relativelyyounger cells.¹ This demonstrated that cloning may allowfor a reversal of the aging process.

Clearly the role of telomeres in aging requires further study.Since there is a clear link between the presence of telomerase andthe possibility of malignant transformation, efforts to reverse theprocess of aging must take into account the increased risk of cancer.Additional evidence also points to the role of ROS in aging. Aging isa complex process that has many variables. Understanding thesevariables may eventually make possible therapeutic interventions,which could slow down the aging process.

References

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