Current Aging Science - Volume 6, Issue 1, 2013

We have investigated the effect of melatonin administration on the cytometric and endocrine functions of the ovary during aging. Young cyclic (3 months old), middle-aged pre-acyclic (13 months old), and old acyclic (22 months old) female Wistar rats were used for two months, in both control and melatonin-treated groups. Cell cycle by flow cytometry: the percentage of ovarian cells in the G0-G1 phase was the highest in both control and melatonin-treated rats. However, melatonin treatment significantly reduced (P< 0.05) the percentage of cells in the G0-G1 phase compared to control rats. This reduction of cells in the G0-G1 phase is derived to the S phase in cyclic and acyclic rats. The possibility that a tumoral process leads to a proliferative effect observed in young and acyclic melatonin-treated rats was ruled out because no significant differences were found for p53 and Ki67 expression levels between control and melatonin groups. Density of oocytes: the oocyte number per ovary unit volume was not affected by melatonin treatment in the three age ranges studied. Melatonin treatment in middle-aged (pre-acyclic) rats resulted in significantly higher (P< 0.05) ovarian volume; higher oocyte volume, without significant differences, and oocytes in circular form were significantly (P< 0.05) higher than in control rats. Melatonin treatment during pre-acyclic age range could resynchronize the estrous cycle periodicity. Melatonin treatment was able to maintain the same levels of estradiol in the pre-acyclic age groups studied as those observed in the young cyclic rats. The present results indicate that melatonin administration to middle-aged female rats produces beneficial effects that extend the reproductive function of the ovary.

A very small tripeptide amide L-pyroglutamyl-L-histidyl-L-prolineamide (L-PHP, Thyrotropin-Releasing Hormone, TRH), was first identified in the brain hypothalamus area. Further studies found that L-PHP was expressed in pancreas. The biological role of pancreatic L-PHP is still not clear. Growing evidence indicates that L-PHP expression in the pancreas may play a pivotal role for pancreatic development in the early prenatal period. However, the role of L-PHP in adult pancreas still needs to be explored. L-PHP activation of pancreatic β cell Ca2+ flow and stimulation of β-cell insulin synthesis and release suggest that L-PHP involved in glucose metabolism may directly act on the β cell separate from any effects via the central nervous system (CNS). Knockout L-PHP animal models have shown that loss of L-PHP expression causes hyperglycemia, which cannot be reversed by administration of thyroid hormone, suggesting that the absence of L-PHP itself is the cause. L-PHP receptor type-1 has been identified in pancreas which provides a possibility for L-PHP autocrine and paracrine regulation in pancreatic function. During pancreatic damage in adult pancreas, L-PHP may protect beta cell from apoptosis and initiate its regeneration through signal pathways of growth hormone in β cells. L-PHP has recently been discovered to affect a broad array of gene expression in the pancreas including growth factor genes. Signal pathways linked between L-PHP and EGF receptor phosphorylation suggest that L-PHP may be an important factor for adult β- cell regeneration, which could involve adult stem cell differentiation. These effects suggest that L-PHP may benefit pancreatic β cells and diabetic therapy in clinic.

According to our conception, the aging process is caused by cell proliferation restriction-induced accumulation of various macromolecular defects (mainly DNA damage) in cells of a mature organism or in a cell population. In the case of cell cultures, the proliferation restriction is related to so-called contact inhibition and to the Hayflick’s limit, while in the case of multicellular organisms, it is related to the appearance, in the process of differentiation, of organs and tissues consisting of postmitotic and very slowly dividing cells. It is assumed that the proliferation of intact cells prevents accumulation of various errors in a cell population. However, the continuous propagation of all the cells in a multicellular organism is absolutely incompatible with its normal functioning. Thus, the program of development, when it generates postmitotic or slowly dividing cells, automatically leads also to the onset of the aging process (mortality increase with age). Therefore, any additional special program for aging simply becomes unnecessary. This, however, doesn’t reject, for some organisms, the reasonability of programmed death, which makes possible the elimination of harmful, from the species point of view, individuals. It is also very important to emphasize that increase or decrease of an organism’s lifespan under the effects of various external factors is not always necessarily related to modification of the aging process, though the experimental results in the field are usually interpreted in just this way. I called the experimental-gerontological models similar to the Hayflick’s model "correlative", since they are based on some correlations only and not related necessarily to the gist of the aging phenomenon. So, for the Hayflick’s model, it is the relationship between population doubling level and donor age, between population doubling potential and species lifespan, between some cell changes in vivo and in vitro, and so forth. If the rationale of the "Hayflick phenomenon" is used, we can’t explain why we age. Nevertheless, many authors virtually put a sign of equality between aging in vitro and aging in vivo, which generates conclusions that are of quite doubtful accuracy. A classic illustration of this is the telomere concept of aging. Originally, the principle of shortening end-segments of DNA (telomeres) during each cell division was formulated at the beginning of seventies by the Russian scientist Aleksey Olovnikov and used by him to explain the limited "proliferative" lifespan in vitro of normal cells. Subsequently, the existence of this phenomenon was confirmed by the results of many research reports, the culmination of which was a publication in which the authors demonstrated the possibility of increasing the proliferative potential of normal cells by introducing the enzyme telomerase to them, thus restoring the lost telomere segments. At the moment it looks like the telomere shortening contributes to aging in vitro only, but not to aging in vivo because an organism never realizes the full proliferative potential of its cells. Besides, the most "responsive to aging" are the organs and tissues consisting of postmitotic cells, for which the concept of proliferative potential loses any meaning in practical terms. We developed another "correlative" model–a model for testing of geroprotectors and geropromoters – the "cell kinetics model." It is based on the well-known correlation between the "age" of cultured cells (age of their donor) and their saturation density. The model allowed us to perform preliminary testing of a lot of different compounds and factors that are interesting from a gerontological point of view, but it revealed no information about the real mechanisms of aging. However, the second model we use in our studies – the "stationary phase aging" model – obviously, is a "gist" model. It is based on the assumption that in the cells of stationary cultures various intracellular changes similar to those of an aging organism can be observed. The proliferation restriction in the case is provided, as a rule, just by contact inhibition. Many experimental results confirming this assumption were obtained. "Age-related" changes that are well known from organismal studies were shown to really occur in our experimental stationary cell culture model. Besides, such experiments can be carried out on nearly any type of cells from various biological species. Thus, the evolutionary approach to analysis of the data is provided. Moreover, the changes in the stationary cell cultures become detectable very soon – as a rule, in 2 to 3 weeks after beginning the experiment. All this allows us to suppose that the "stationary phase aging" model should provide a very effective approach to testing of different substances and their cocktails on their activities in terms of accelerating or retarding aging – of course, if their effect is realized on the cell level only.

Advanced age is associated with an increased incidence of immune and degenerative disorders, mediated by metabolic changes, dysregulation of proinflammatory signals, and apoptosis. Concurrently, there is a progressive decline in self-recognition. Investigations on biologic functions of transferrin (Tf) other than iron transport showed that Tf has a profound cytoprotective (anti-apoptotic) effect on lympho-hematopoietic cells and the thymus, and interferes with stressinduced signals. Tf protects hepatocytes against Fas-induced cell death by reducing BID cleavage, inhibiting caspase-3 and -9 activation and up-regulating survival signals such as Bcl-xL. The involvement in the regulation of alloreactivity and apoptosis suggests that Tf participates in the maintenance of “self-identity” mechanisms, which are tightly linked to the capacity of the immune system to recognize and react against any noxious agent. Some of the disorders associated with aging are thought to be related to thymic involution, reflecting alterations in the interplay of neural, endocrine and immune factors. We established a murine model of thymic involution induced by stereotactically placed electrolytic lesions in the anterior hypothalamic area. The events observed in this model mimic those observed during senescence including thymus involution, i.e. enhanced glucocorticoid reaction to distress, and obesity. The described properties of Tf can be exploited to modify immune responses and provide cytoprotection against pro-apoptotic and cytotoxic signals when neuroimmunomodulatory mechanisms are impaired, as is the case with aging.

Hyperhomocysteinemia is a risk factor for a number of neurodegenerative and cardiovascular diseases. We have shown that homocysteine induces excitotoxic effects in cells expressing glutamate receptors of the NMDA class. These receptors were found not only in neurons but also in immune-competent cells, neutrophils, red blood cells, cardiomyocytes, and osteoblasts. Activation of these cells by homocysteine results in an increase in cytoplasmic calcium ions, accumulation of reactive oxygen species, and activation of MAP kinase. An overload of immune-competent cells activates both necrotic and apoptotic cell death, whereas the neuropeptide carnosine (an antioxidant and immune modulator) protects cells against both processes. In a model of prenatal hyperhomocysteinemia in rats, we have found that carnosine protects animals against homocysteine toxicity with no change of the blood homocysteine levels. The efficiency of carnosine has also been demonstrated in clinical trials of chronic brain ischemia and Parkinson’s disease.

Metallothioneins (MTs) are small, cysteine-rich proteins which have been implicated in various forms of stress providing cytoprotective action against oxidative injury, DNA damage and apoptosis. Owing to their high affinity for physiological metals, such as zinc and copper MTs are also critical components of regulatory proteins involved in cell growth and multiplication, as well as in the maintenance of immune homeostasis. To elucidate the role of MTs in the pathomechanisms of autoimmune CNS disorders we estimated the expression of MT I+II proteins and the content of free Zn ions in the brain, spinal cord and in the liver early in the course of chronic relapsing experimental autoimmune encephalomyelitis (CR-EAE) pathogenesis, i.e. before the onset of any clinical symptoms. Disease was induced in the genetically susceptible Dark Agouti (DA) rats by subcutaneous injection of bovine brain homogenate in CFA. Control animals were treated with CFA alone. The data, obtained by immuno-histochemistry and in situ fluorescent labeling of free zinc ions, have shown that in the presymptomatic phase of CR-EAE (on the seventh postimmunization day) MTs I+II were markedly upregulated in the cells that form blood-brain and blood-cerebrospinal fluid barriers, as well as in the cerebellar parenchyma and hippocampal dentate gyri. Furthermore, we found that the liver also becomes a site of extensive MTs I+II synthesis shortly after immunization. Simultaneously, tissue content of free zinc ions increased at the sites of MTs induction, reflecting their antioxidative activity. The data, described in this paper point to regulatory and neuroprotective role of MTs in the pathogenesis of CR-EAE.

Parkinsons’disease (PD), a common neurodegenerative disorder, is characterized by progressive loss of dopaminergic (DAergic) neurons in the subtantia nigra pars compacta (SNpc) and gliosis. The cause and mechanisms underlying the demise of nigrostriatal DAergic neurons are not completely clarified, but interactions between genes and environmental factors are recognized to play a critical role in modulating the vulnerability to PD. Current evidence points to reactive glia as a pivotal factor in PD, but whether astroglia activation may protect or exacerbate DAergic neuron loss is presently the subject of much debate. Astrocytes and microglia are the key players in neuroinflammatory responses, by secreting an array of pro- and anti-inflammatory cytokines, anti-oxidant and neurotrophic factors. Here, the contribution of astrocytes and their ability to influence DAergic neurodegeneration, neuroprotection and neurorepair will be discussed. In particular, the dynamic interplay between astrocyte-derived factors and neurogenic signals in MPTP-induced plasticity of nigrostriatal DAergic neurons will be summarized together with recent findings showing that reactive astrocytes may contribute to promote DAergic neurogenesis from midbrain adult neural stem/precursor cells (NPCs). Within a host of astrocyte- derived factors, we unveiled Wingless-type MMTV integration site (Wnt)/β-catenin signalling was unveiled, as a strong candidate in MPTP-induced DAergic neuroplasticty/neurorepair. Understanding the intrinsic plasticity of nigrostriatal DAergic neurons and decifering the signals facilitating the crosstalk between astrocytes and midbrain neuroprogenitors may have implications for the role of stem cells technology in PD and for identifying potential therapeutic targets to promote endogenous neurorepair.

Physiologic autoantibodies, that is, those with an active physiologic role, are an important part of the normal human immune system and are essential in maintaining homeostasis. Evidence suggests that the body uses autoantibodies to prevent disease and to self-treat diseases once started. This suggests a potential therapeutic role for autoantibodies, or, even better, a way to use them to prevent disease. Their capacity to remove aged, damaged cells is well established. Immunoglobulin (Ig) G autoantibodies bind to senescent cell antigen (SCA), which is an altered band 3 anion exchanger protein found mainly on aged cells. Once bound, IgG triggers the removal of the senescent cells by macrophages. Band 3 is altered primarily by oxidation, which in turn generates SCA. These studies demonstrated that oxidation can generate neoantigens that the immune system will recognize. Band 3 isoforms are ubiquitous: they have been found in all mammalian cells and species so far examined. The innate immune response to band 3 membrane proteins, and their regulation of cellular lifespan and therapeutic potential will be presented. Examples of other potential innate and physiologic autoantibodies include neuroprotective antibodies to amyloidgenic toxic peptides and antibodies to oxidized LDL (OxLDL), which modify the natural progression of atherosclerosis.

This study demonstrates the effects of Quinton’s isotonic and hypertonic solution on mononuclear cells of peripheral blood. This involved assessing cell viability, morphology, number and size of aggregated cells; possible effects on cellular proliferation; effects on cellular proliferation in different lymphocyte populations; effect on hemoglobin released into the medium.

It has been suggested that arterial stiffness is one of the most important risk factors for the development of a cardiac infarction or stroke. As cardiovascular disease remains the number one killer of individuals before the age of 75, the early detection of cardiovascular disease and its prevention remains paramount in order to sustain a healthy longevity. This article looks at the latest noninvasive technology that can measure arterial stiffness quickly and easily and also highlights a small open trial in which supplements were used to determine their efficacy in helping to reverse/improve arterial stiffness.

First, the latest scientific and clinical reports will be evaluated to separate the wheat from the chaff, that is, good data versus merely anecdotal evidence. Thus, the famous (infamous) Stromboli Cocktail will be brought up to date. Second, longevity statistics will be reviewed: Why do the most scientifically advanced countries have such low (comparatively) life expectancies? Scientific knowledge expands exponentially each decade, whereas there have been no significant advances in our knowledge, government, economics, politics, anti-corruption, and so forth since the dawn of history. What can we expect in the future? Will the human species outlive the cockroach? Can we expect to get closer to that theoretical asymptote of 120 years of human life? Will this ceiling ever be lifted? Finally, we offer two vital challenges to scientists of today.

Thyrotropin-releasing hormone (TRH) aroused our interest when we were engaged in related experiments, so we decided to study its effects on organs, tissues, and aging-related metabolic and hormonal markers when administered in acute or chronic (oral) doses at various time points in its cyclic circadian pattern. We also wanted to determine what effects, if any, it had on aging processes in two essential systems, namely gonadal-reproductive and kidney-urinary. Our results show positive changes as a result of short-term acute and long-term chronic oral administration of TRH to old mice that included rapid correction to more juvenile levels of most typical aging-related hormonal and metabolic measurements. Remarkably, testes function was maintained by means of a 4-month oral treatment with TRH in aging mice. As we suspected upon seeing a significant increase in testes weight, TRH resulted in maintenance or even reconstitution of testes structure and function when administered in the drinking water. This was demonstrated by the active formation and proliferation of mature spermatogonia and the intensive spermatogenesis in the follicles. The same TRH treatment led to protection for the kidneys from amyloid and hyalin infiltration of tubuli and glomeruli, which typically occurs in aging mice. In fact, we observed massive deposits of amyloid and hyalin material infiltrating the shrunken glomeruli and negatively affecting filtration capacity of the untreated mice, whereas this was barely present in the TRH-treated mice. Advanced hyalin degeneration could also be observed in the tubular vessels of the untreated control mice. These experiments with TRH supplementation show clear aging-delaying and apparently even aging-reversing effects of the neuropeptide, whether it was administered parenterally or orally. TRH, like melatonin, is an anti-aging agent with a broad spectrum of activities that, because of their actions, suggest that TRH has a fundamental role in the regulation of metabolic and hormonal functions.

The restoration of the thymic functions and the thymic re-growth may be achieved in old mice by some endocrinological (melatonin) or nutritional interventions (arginine or zinc), suggesting that the thymic involution in old age is a phenomenon secondary to age-related alterations occurring in neuroendocrine–thymus interactions. The targets for the thymic restoration may be hormone receptors and cytokines, strictly related to the presence of two nutritional factors, such as arginine and zinc, which are in turn essential for the efficiency of neuroendocrine–immune network both in ontogeny and ageing. The effect of melatonin is largely due to the presence of its specific receptors on cell membrane of thymocytes and Thymic Epithelial Cells (TECs). TECs synthesize thymulin peptide that is required for T-cell differentiation and maturation within the thymus gland. In this context, the role of zinc is pivotal because it is involved, through “zinc finger motifs”, in the gene expression of melatonin receptors, in cell proliferation, apoptosis and thymulin reactivation. Zinc is also required for the biological action of arginine, via Nitric Oxide pathway. Therefore, the beneficial effect of melatonin or arginine on neuroendocrine-thymus interaction in ageing can also occur via a better zinc pool redistribution within the body where the capability of the zinc-binding proteins Metallothioneins (MT) in zinc release has a key role. These findings suggest that zinc, via MT buffering, can be a single mediator in modulating neuroendocrine-thymus interaction in ageing.

The majority of chronic diseases, most notably those accompanying aging, result from progressive deterioration of central neuroimmunoendocrine control, often referred to as immunological surveillance. This is as true of cancer as it is of the development of cardiovascular, autoimmune, and neurodegenerative disease, in all of these immunological surveillance break downs, leading to an unraveling of the neuroimmunoendocrine process that inhibits proliferation of preneoplastic and neoplastic cells already existing in the body. The onset of cancer is anticipated by changes in the hormonalimmune coordination resulting in chronic quantitative alterations in the synthesis and release of hormones and the loss of the natural synchronicity of that release, which occurs according to circadian rhythms in the healthy organism, principally under the control of the pineal network. Periodic circadian hormonal release is the source of immune system regulation, thus altering hormone rhythms impairs the immune system’s ability to maintain control over emerging tumor cells, not necessarily to eliminate them, but to inhibit proliferation. Malignancy, then, is the result of suppression of or interference with the regular release of hormones that maintain strict regulation of the thymo-lymphatic immune system’s maturation and activity. This understanding means that we can act to prevent cancer by means of efficiently monitoring and maintenance of physiological hormonal values. For the cyclic synthesis of malignancies that are metastasized, a means of xenogeneic bone marrow transplantation is proposed as an alternative therapeutic approach.

There is virtually no correlation between what are generally accepted to be the symptoms of deficient androgen in men and levels of androgens as measured in the laboratory. Now that androgen deficiency is being shown to play a part in conditions as diverse as metabolic syndrome, diabetes, and coronary heart disease, a hypothesis is needed to explain this apparent discrepancy between measured androgen levels and our understanding of the symptoms of androgen deficiency. When the possible mechanisms for androgen actions are considered, one explanation emerges that androgen may act much like insulin in persons with type 2 diabetes mellitus: the degree of androgen resistance may be variable depending on the organs or systems considered. Therefore, the symptoms can result from altered or damaged synthesis of androgen synthesis or regulation, elevated androgen binding, a reduction in tissue response, or decreased as a result of polymorphism and aging. Genomic transcription and translation may also be affected. As with diabetes, in adult male androgen deficiency, it is suggested that the definition of androgen deficiency should be based on individual physiology, with the requirements of the individual at a particular stage of life setting the baseline against which any deficiency of androgens or androgen metabolites, either absolute or relative, is determined. This approach will affect the terminology, etiology, diagnosis, and treatment of androgen deficiency.

The pineal hormone melatonin (MLT) has potent anti-breast cancer activity, its actions are heavily mediated via the MT1 receptor and subsequent modulation of downstream signaling pathways including cAMP/PKA, Erk/MAPK, p38, and Ca2+/calmodulin. Also, via the MT1 pathway, MLT can repress the transcriptional activity of some mitogenic nuclear receptors including ERα, GR, and RORα, while potentiating the activity of other receptors (RARα and RXRα) involved in differentiation, anti-proliferation, and apoptosis. A review of the literature supports the view that MLT, via its MT1 receptor, can suppress all phases of breast cancer including initiation, promotion, and progression. During the fifth and sixth decades of life, the production of MLT diminishes, concurrently with an increase in the incidence of breast cancer. Inasmuch as MLT has been demonstrated to have anti-cancer activity, we hypothesized that there may be a causal link between the reduction in MLT production in the pineal gland and the incidence of breast cancer which increases with age. We designed this study to establish whether a truly inverse relationship exists between tissue-isolated mammary tumor growth in young (2 months), adult (12 months), and old (20 months) female Buffalo rats and the decrease in both MLT and the MT1 receptor with age, such that a causal link could be found. Serum MLT levels were measured in both the light and dark phases. A significant 29% decrease in serum MLT levels, measured at the nocturnal peak, was found in the adult and senescent rats (75% decrease) in comparison to that in young rats. In young rats, the nocturnal pineal gland MLT content exceeded daytime levels by 19-fold compared to a sevenfold increase in old mice. Also, the MT1 receptor was found to be significantly lower in the nighttime and early morning in the senescent rat uterus as compared to uteri from young and adult rats. Analysis of the rate of growth in transplanted, tissue-isolated, mammary tumors induced by N-nitroso-n-methyl-urea (NMU) showed a significant increase in the senescent rats, but not in the young or adult rats Additionally, diminished response to the inhibitory action on tumor growth of exogenous MLT was noted in senescent rats such that tumor growth was suppressed by only 33% compared to 48% and 66% in adult and young rats, respectively. The diminution of the response of tumors to exogenous MLT was found to correlate with reduced MT1 receptor expression in senescent compared to young and adult rats. These data suggest that the observed age-associated enhanced growth of tumors is related to the much reduced levels of MLT and its receptor in aged animals which reduce the sensitivity of tumors to inhibition by exogenous MLT.