Richard G. A. Faragher

Telomerase prevents the accelerated cell ageing of Werner syndrome fibroblasts.

Werner syndrome (WS) is a rare disorder inherited in an autosomal recessive manner and characterized by accelerated ageing. WS fibroblasts display an accelerated rate of senescence in vitro, which has been linked to this progeroid phenotype. The senescence of normal human fibroblasts is triggered by telomere shortening, whereas the premature senescence of WS fibroblasts has been assumed to reflect the accumulation of DNA damage. Here we show that forced expression of telomerase in WS confers extended cellular lifespan and probable immortality.

Ocular biomaterials and implants

The maintenance of vision is a key determinant of healthy ageing. This has been facilitated over recent decades by the development of a wide range of implants and biomedical devices to correct the functional deficiencies of disease, age and ocular trauma. This brief overview provides an insight into the structure of this unique organ, the major physiological functions of the component tissues and the present state of the art with respect to modern ocular implants. The review focuses primarily on the existing limitations of existing ocular biomaterials used in the fabrication of contact lenses, intraocular lenses, glaucoma filtration implants, keratoprostheses, intracorneal implants, scleral buckles and viscoelastic replacement agents. The challenge of improving ocular compatibility and ensuring the longevity of indwelling ocular devices is addressed along with the need to improve the physicochemical and mechanical properties of existing ocular biomaterials.

How might replicative senescence contribute to human ageing

Cell senescence is the limited ability of primary human cells to divide when cultured in vitro. This eventual cessation of division is accompanied by a specific set of changes in cell physiology, morphology, and gene expression. Such changes in phenotype have the potential to contribute to human ageing and age‐related diseases. Until now, senescence has largely been studied as an in vitro phenomenon, but recent data have for the first time directly demonstrated the presence of senescent cells in aged human tissues. Although a direct causal link between the ageing of whole organisms and the senescence of cells in culture remains elusive, a large body of data is consistent with cell senescence contributing to a variety of pathological changes seen in the aged. This review considers the in vitro phenotype of cellular senescence and speculates on the various possible routes whereby the presence of senescent cells in old bodies may affect different tissue systems. BioEssays 20:985–991, 1998.

Cell divisions and mammalian aging: integrative biology insights from genes that regulate longevity

Despite recent progress in the identification of genes that regulate longevity, aging remains a mysterious process. One influential hypothesis is the idea that the potential for cell division and replacement are important factors in aging. In this work, we review and discuss this perspective in the context of interventions in mammals that appear to accelerate or retard aging. Rather than focus on molecular mechanisms, we interpret results from an integrative biology perspective of how gene products affect cellular functions, which in turn impact on tissues and organisms. We review evidence suggesting that mutations that give rise to features resembling premature aging tend to be associated with cellular phenotypes such as increased apoptosis or premature replicative senescence. In contrast, many interventions in mice that extend lifespan and might delay aging, including caloric restriction, tend to either hinder apoptosis or result in smaller animals and thus may be the product of fewer cell divisions. Therefore, it appears plausible that changes in the number of times that cells, and particularly stem cells, divide during an organisms lifespan influence longevity and aging. We discuss possible mechanisms related to this hypothesis and propose experimental paradigms. BioEssays 30:567–578, 2008.

Aging and the cornea.

Aging, the persistent decline in age specific fitness of an organism as a result of internal physiological deterioration, is a common process among multicellular organisms.1 In humans, aging is usually monitored in relation to time, which renders it difficult to differentiate between time dependent biological changes and damage from environmental insults. There are essentially three types of aging at work in any adult tissue; the aging of long lived proteins, the aging of dividing cells, and the aging of non-dividing cells.2 Dividing cells may be derived from renewing populations in which the rate of cell loss and division is great. An example is the corneal epithelium in which complete turnover occurs within 5–7 days after terminal differentiation.34Conditional renewal populations, which normally have an extremely low proliferation rate, can also produce dividing cells in response to extrinsic stimuli. Stromal keratocytes are a prime example of a conditional renewing population.5 Corneal endothelial cells retain the capacity to undergo mitosis and conditional renewal in humans although they very seldom do so.6-9 Non-dividing cells are those from static cell populations (exemplified by cerebral neurons) which never divide during adult life.3 Corneal aging produces both structural and functional changes. These changes in turn can affect the ability of the organ to refract light, to repair itself, and to protect itself and the internal structures of the eye.10 A variety of corneal aging changes have been reported. However, as it is difficult to distinguish age specific deterioration from degenerations modified by environmental and genetic factors, we think it is helpful to consider these alterations within the broader framework of the aging process. The study over the past 30 years of isolated cells in culture as a model system for aging changes has greatly advanced our understanding of …

Microarray analysis of senescent vascular smooth muscle cells: A link to atherosclerosis and vascular calcification

Little is known about the senescent phenotype of human vascular smooth muscle cells (VSMCs) and the potential involvement of senescent VSMCs in age-related vascular disease, such as atherosclerosis. As such, VSMCs were grown and characterised in vitro to generate senescent VSMCs needed for microarray analysis (Affymetrix). Comparative analysis of the transcriptome profiles of early (14 CPD) and late (39-42 CPD) passage VSMCs found a total of 327 probesets called as differentially expressed: 149 are up-regulated in senescence and 178 repressed (p-value<0.5%, minimum effect size of at least 2-fold differential regulation, explore data at http://www.madras.cf.ac.uk/vsmc). Data mining shows a differential regulation of genes at senescence associated with the development of atherosclerosis and vascular calcification. These included genes with roles in inflammation (IL1beta, IL8, ICAM1, TNFAP3, ESM1 and CCL2), tissue remodelling (VEGF, VEGFbeta, ADM and MMP14) and vascular calcification (MGP, BMP2, SPP1, OPG and DCN). The microarray data for IL1beta, IL8 and MGP were validated by either, ELISA, Western blot analysis or RT-PCR. These data thus provide the first evidence for a role of VSMC senescence in the development of vascular calcification and provides further support for the involvement of senescent VSMCs in the progression of atherosclerosis.

Telomere-based proliferative lifespan barriers in Werner-syndrome fibroblasts involve both p53-dependent and p53-independent mechanisms

Werner-syndrome fibroblasts have a reduced in vitro life span before entering replicative senescence. Although this has been thought to be causal in the accelerated ageing of this disease, controversy remains as to whether Werner syndrome is showing the acceleration of a normal cellular ageing mechanism or the occurrence of a novel Werner-syndrome-specific process. Here, we analyse the signalling pathways responsible for senescence in Werner-syndrome fibroblasts. Cultured Werner-syndrome (AG05229) fibroblasts senesced after ∼20 population doublings with most of the cells having a 2N content of DNA. This was associated with hypophosphorylated pRb and high levels of p16Ink4a and p21Waf1. Senescent AG05229 cells re-entered the cell cycle following microinjection of a p53-neutralizing antibody. Similarly, production of the human papilloma virus 16 E6 oncoprotein in presenescent AG05229 cells resulted in senescence being bypassed and extended cellular life span. Werner-syndrome fibroblasts expressing E6 did not proliferate indefinitely but reached a second proliferative lifespan barrier, termed Mint, that could be bypassed by forced production of telomerase in post-M1 E6-producing cells. The conclusions from these studies are that: (1) replicative senescence in Werner-syndrome fibroblasts is a telomere-induced p53-dependent event; and (2) the intermediate lifespan barrier Mint is also a telomere-induced event, although it appears to be independent of p53. Werner-syndrome fibroblasts resemble normal human fibroblasts for both these proliferative lifespan barriers, with the strong similarity between the signalling pathway linking telomeres to cell-cycle arrest in Werner-syndrome and normal fibroblasts providing further support for the defect in Werner syndrome causing the acceleration of a normal ageing mechanism.

Insights into CNS ageing from animal models of senescence

In recent years, novel model systems have made significant contributions to our understanding of the processes that control the ageing of whole organisms. However, there are limited data to show that the mechanisms that gerontologists have identified as having a role in organismal ageing contribute significantly to the ageing of the central nervous system. Two recent discoveries illustrate this particularly well. The first is the consistent failure of researchers to demonstrate a simple relationship between organismal ageing and oxidative stress — a mechanism often assumed to have a primary role in brain ageing. The second is the demonstration that senescent cells play a causal part in organismal ageing but remain essentially unstudied in a CNS context. We argue that the animal models now available (including rodents, flies, molluscs and worms), if properly applied, will allow a paradigm shift in our current understanding of the normal processes of brain ageing.

From old organisms to new molecules: integrative biology and therapeutic targets in accelerated human ageing

Abstract.Understanding the basic biology of human ageing is a key milestone in attempting to ameliorate the deleterious consequences of old age. This is an urgent research priority given the global demographic shift towards an ageing population. Although some molecular pathways that have been proposed to contribute to ageing have been discovered using classical biochemistry and genetics, the complex, polygenic and stochastic nature of ageing is such that the process as a whole is not immediately amenable to biochemical analysis. Thus, attempts have been made to elucidate the causes of monogenic progeroid disorders that recapitulate some, if not all, features of normal ageing in the hope that this may contribute to our understanding of normal human ageing. Two canonical progeroid disorders are Werner’s syndrome and Hutchinson-Gilford progeroid syndrome (also known as progeria). Because such disorders are essentially phenocopies of ageing, rather than ageing itself, advances made in understanding their pathogenesis must always be contextualised within theories proposed to help explain how the normal process operates. One such possible ageing mechanism is described by the cell senescence hypothesis of ageing. Here, we discuss this hypothesis and demonstrate that it provides a plausible explanation for many of the ageing phenotypes seen in Werner’s syndrome and Hutchinson-Gilford progeriod syndrome. The recent exciting advances made in potential therapies for these two syndromes are also reviewed.

Telomerase and the cellular lifespan: implications of the aging process.

ABSTRACT The aging process has multiple causes. However, there is now substantial evidence consistent with the hypothesis that (i) all normal mammalian somatic cells have a finite capacity to replicate and (ii) that gradual cell turnover throughout the lifespan of a mammal eventually exhausts this finite capacity. This results in a gradual accumulation of senescent (irreversibly post-mitotic) cells with increasing age. These cells display a radically different phenotype to their growing counterparts, which has the potential to compromise tissue function. Perhaps the best evidence for this is seen in Werner’s syndrome, a rare genetic disease, in which patients display most of the features of accelerated aging, together with a profoundly compromised replicative lifespan in certain tissue lineages. Several classes of human cells are now known to count divisions by monitoring the progressive attrition of chromosomal ends (telomeres), leading to the activation of a p53-p21waf - dependent G1 checkpoint. Ectopic expression of telomerase has been shown to prevent senescence in several cell types and offers the potential for interventions in the aging process based on tissue engineering, gene therapy or homeografts. However, this telomere-driven senescence mechanism seems to be absent from rodents, which use telomere-independent means (perhaps based upon p14arf) to count divisions. Similar senescence pathways are now being reported in humans, and this, coupled with the demonstration of tissue-specific telomeric loss rates, has the potential to render strategies based on the use of telomerase dependent on the characteristics of the target tissue. Werner’s syndrome may provide strong clues regarding the potential limitations and prospects of such future treatments.