Current Pharmaceutical Design - Volume 14, Issue 26, 2008

Ageing is a post-maturational process that, due to a diminished homeostatic capacity and increased vulnerability, reduces responsiveness to environmental stimuli. Nowdays, individuals can live until 80-120 years but this increase in lifespan is not free of disabilities and diseases. Therefore, current attempts are aimed at finding out drug targets by which one may correct functional deficits in ageing and/or prevent age-associated diseases. Current knowledge on ageing and age-associated diseases and anti-ageing therapies will be reviewed in this issue of Current Pharmaceutical Design. Jirillo and associates [1] will discuss the state of the art of anti-ageing therapies with special reference to immunosenescence, Alzheimer's disease, cardiovascular diseases and diet and age-related diseases. Henson and associates [2] will examining the role of CTLA-4 and PD-1 in regulating immune response and their therapeutic potential in the elderly. Amadio and associates [3] will show that the RNA-binding protein ELAV/HuR can affect, post-transcriptionally, the fate of HSP70 mRNA following H2O2-mediated oxidative stress in SH-SY5Y human neuroblastoma cells. An impairment of this regulatory mechanism may lead to the defective cellular response to oxidative stress as evidenced in cerebral senescence. Porcellini and associates [4] will discuss on the role of α-1-anti-chymotrypsin (ACT) in Alzheimer's disease and the development of compounds able to interfere with ACT biological activity. Lanni and associates [5] will focus on p53 which has been involved in ageing and Alzheimer's disease. With ageing a conformational change of p53 occurs which impairs its ability to activate an apoptotic program when cells are exposed to noxious stimuli. Balistreri and associates [6] will demonstrate a correlation between the polymorphisms of TLR4 and CD14 genes and Alzheimer's disease with special reference to the +896A/G TLR4 single nucleotide polymorphism. Listì and associates [7] will report that -765G COX-2 and -1708A 5-LO single nucleotide polymorphisms are independent positive risk factors for developing myocardial infarction, thus allowing to determine which patients to treat with specific inhibitors of the eicosanoids or their enzymes. Iacoviello and associates [8] will show that high thyroid stimulating hormone levels in chronic heart failure patients are independently associated with a greater likelihood of heart failure progression, thus supporting prospective studies for establishing the most appropriate treatment of clinical hypothyroidism in such patients. Chiloiro and associates [9] will examine the relationship between moderate obesity and glucose metabolism, insulin sensitivity and suspected fatty liver in children. Results will clarify that moderately obese children exhibit a clear increase of insulin and insulin resistance which may lead to a future metabolic syndrome. Di Renzo and associates [10] will investigate the effect of body fat mass on the relationships between -174G/C IL-6 promoter gene polymorphism, IL-6 circulating level and insulin resistance. Data will show that fat mass percentage is a major determinant of increase in IL-6 production and insulin resistance. Trosko [11] will emphasize the need to provide elderly subjects with a correct dietary intake that might also affect stem cells, whose alterations can compromise the ageing of that organ. Mocchegiani and associates [12] will demonstrate that the association “zinc plus selenium” improves humoral immunity in old subjects after influenza vaccination. Furthermore, the association “zinc plus niacin (a precursor of NAD+)” in elderly is in progress. Magrone and associates [13-15] will investigate the ability of red wine polyphenols to promote the in vitro release from human healthy peripheral blood mononuclear cells of regulatory, proinflammatory and antiinflammatory cytokines as well as of immunoglobulins. The involvement of polyphenols in the activation of p38 and ERK1/2 as well as nitric oxide production will be discussed. Collectively, in vitro data will show that a moderate use of red wine in humans is able to elicit at cellular and molecular levels a protective response for the host mostly in age-related disorders.

A lasting dream of human beings is to reverse or at least postpone ageing. During the last years, an increasing number of scientific meetings, articles, and books have been devoted to anti-ageing therapies. This subject, full of misleading, simplistic, or wrong ideas, is very popular among the general public, whose imagery has been fascinated by all possible tools to delay ageing, getting immortality. Here, we discuss anti-ageing strategies aimed not to rejuvenate but to slow ageing and delay the onset of age-related diseases. These approaches should be able to substantially slow down the ageing process, extending our productive, youthful lives.

Inhibitory receptors of the CD28 family, CTLA-4 and PD-1 deliver negative signals that regulate the balance between T cell activation, tolerance, and immunopathology. Manipulation of these pathways has been utilized by pathogens and tumors to establish chronic infections or to promote tumor survival. In this review, we examine the role of CTLA-4 and PD-1 in regulating immune response and discuss their therapeutic potential during aging.

Brain aging is associated with a progressive imbalance between intracellular concentration of Reactive Oxygen Species (ROS) and cells ability to activate defensive genes. Heat Shock Protein 70 (HSP70) has been shown to act as a fundamental defensive mechanism for neurons exposed to an oxidant challenge, and its expression decreases during senescence. In the present report we show that the RNA-binding protein ELAV/HuR can affect, post-transcriptionally, the fate of HSP70 mRNA following H2O2-mediated oxidative stress in SH-SY5Y human neuroblastoma cells. As a consequence of H2O2 treatment (1mM for 30 minutes), HSP70 mRNA accumulates in the ribosomes associated to the cytoskeleton, where parallel Western blotting experiments reveal statistically significant increase for both HuR and HSP70 protein levels. We also confirm the capability of HuR to bind to HSP70 mRNA, and describe how the biological effect of this ELAV protein on the HSP70 mRNA could be due to a direct phosphorylation in serine/threonine residues of HuR itself by the early (10 minutes) H2O2-mediated activation of PKCα. Our findings shed light on the post-transcriptional regulation of HSP70 expression, suggesting the existence of a new molecular cascade -involving PKC/HuR/HSP70- that possibly represents an early event in the cellular response to H2O2-mediated oxidative stress in SH-SY5Y human neuroblastoma cells. The present results lead us to speculate that an impairment in this regulatory mechanism might directly contribute to the defective cellular response to oxidative stress, thus helping to dissect a potential tool useful to counteract some aspects associated to cerebral senescence.

α-1-antichymotrypsin (ACT), is an acute phase protein and a protease inhibitor produced by the liver and brain. ACT is involved in the pathogenesis of Alzheimer's disease (AD), since elevated ACT concentration was found in cerebrospinal fluid (CSF) and brain from AD. ACT has also been shown to influence amyloid deposition in vitro and in animal models of AD. In this investigation 830 healthy controls, 69 subjects with cognitive impairment and not dementia (CIND), 53 patients with severe clinical AD and 142 patients with mild AD were investigated. Plasma levels of ACT were measured with a new competitive immune enzyme linked immune-assay (ELISA). ACT levels were higher in AD patients than in CIND or controls. An age dependent increase of plasma ACT was present in both healthy elderly and CIND. Patients with mild clinical AD were followed up for two years and stratified according to the rate of clinical deterioration. CT plasma levels were elevated in AD patients that showed an accelerated rate of cognitive deterioration during the follow up; this increment being prominent in AD with the Apolipoprotein E (APOE) ε4 allele. Therefore, increased peripheral ACT levels in APOE 4 positive patients appear to predict an accelerated clinical progression. Plasma ACT might be used as a surrogate marker to monitor the conversion of pre-dementia stages to AD and the progression of the disease. The development of compounds able to interfere with the ACT biological activity (protease inhibition and/or promotion of amyloid deposition) might have therapeutic relevance for the disease.

In spite of the fact that the aging organism is the result of complex life-long gene/environment interactions, making peculiar the susceptibility to diseases and the response to drugs, pharmacogenetics studies are largely neglected in the aged. Altered response to drugs, cardiovascular and metabolic alterations, cancer and dementia are among the age associated ailments. The latter two are the major contributors to illness burden for the aged. Aging, dementia and cancer share a critical set of altered cellular functions in the response to DNA damage, genotoxic stress, and other insults. Aging in higher animals may be influenced by the balance of cell survival versus death, a decision often governed by checkpoint proteins in dividing cells. The paper is mainly focused on one of such proteins, p53 which has been recently shown to be involved in aging and Alzheimer's Disease (AD). Within this reference frame we studied p53 in aged controls and demented patients finding that with aging there is an increase of mutant like conformation state of p53 in peripheral blood cells, which is more pronounced in AD patients. As a result of such conformational change, p53 partially loses its activity and may become unable to properly activate an apoptotic program when cells are exposed to a noxious stimulus. Moreover we found that the tertiary structure of p53 and the sensitivity to p53-dependent apoptosis are affected by low concentrations of soluble beta amyloid, the peptide that accumulates in AD brain but also present in peripheral tissues. It is possible that p53 conformers may occur in the presence of misfolded molecules such as, but not limited to, beta amyloid. In particular at neuronal level the altered function of cell cycle proteins may affect synaptic plasticity rather than cell duplication.

Alzheimer's disease (AD) is a heterogeneous and progressive neurodegenerative disease which in Western society mainly accounts for clinical dementia. Inflammation plays a key role in AD and dissecting the genetics of inflammation may provide an answer to the possible treatment. Hence, the better understanding of different molecular and cellular inflammatory mechanisms is crucial for complete knowledge of AD pathophysiology, and for its prevention and drug therapy. Accordingly, in the present study we evaluated whether the pro-inflammatory polymorphisms of lipopolysaccaride- receptors, +896A/G Toll-Like Receptor (TLR4) and -260C/T CD14, are risk factors for AD. The study included both 626 AD patients (427 women and 199 men; age range: 53-98 years; mean age: 74.88 ± 8.44) from Northern Italy and age and gender matched controls. Our results demonstrate that the +896A/G TLR4 single nucleotide polymorphism (SNP) is associated with AD, whereas no association has been observed with -260C/T CD14 SNP. Furthermore, no differences have been observed evaluating the combined presence of +896A+TLR4/-260T+CD14 “high responder”(proinflammatory- profile). However, our results showing the involvement of TLR4 in AD pathophysiology, strengthen the suggestion that systemic inflammation plays a key role in AD. Carriers of high responder SNP, affected by mild cognitive impairment might, be the ideal target for a preventive treatment with biologics as monoclonal antibodies directed against the pro-inflammatory cytokines to decrease the level of systemic inflammation involved in AD pathophysiology.

Inflammation and genetics play an important role in the pathogenesis of coronary heart disease (CHD). However, despite the increasing appreciation of the role of genetics in CHD and myocardial infarction (MI) pathogenesis, pharmacogenomic approaches to uncover drug target have not been extensively explored. Cyclo-oxygenases (COXs) and 5-lipoxygenase (5-LO) are the key enzymes in the conversion of arachidonic acid to prostaglandins (PG) and leukotrienes (LT) and are implicated in a wide variety of inflammatory disorders, including atherosclerosis. In fact, PGE2 activates Matrix Metallo-proteinases whereas LTB4 is a chemoactractant for monocytes and activates gene expression in inflammatory cells. We have tested the hypothesis that anti-inflammatory variants of these genes confer genetic resistance to MI and conversely favour longevity. So, we analyzed MI patients, age-related controls and centenarians. The proinflammatory alleles of COX-2 and 5-LO were overrepresented in MI and under-represented in centenarians whereas agerelated controls displayed intermediate values. MI is a multifactorial disease, hence MI might be the result of a cumulative effect which contributes with different timing to achieve a threshold where the chance to develop the diseases is very high. In particular, differences in inflammatory status can contribute to the chance of developing a risk phenotype. However, these studies might contribute to the determination of a risk profile which may allow both the early identification of individuals susceptible to disease and the possible discovery of potential targets for drug.

Background. It has been suggested that low thyroid hormones levels may be associated with increased mortality in patients with cardiovascular disease. Aim. To evaluate the prognostic role of thyroid function deficiency in patients with chronic heart failure (CHF). Methods. We evaluated 338 consecutive outpatients with stable CHF receiving conventional therapy, all of whom underwent a physical examination, electrocardiography and echocardiography. Blood samples were drawn to assess renal function, and Na+, hemoglobin, NT-proBNPs, fT3, fT4 and TSH levels. Patients with hyperthyroidism were excluded. Results. During the follow-up (15±8 months), heart failure progression was observed in 79 patients (including 18 who died of heart failure after hospitalisation and six who underwent transplantation). Univariate regression analysis showed that TSH (p<0.0001), fT3 (p<0.0001), fT4 (p=0.016) and fT3/fT4 (p<0.0001) were associated with heart failure progression but multivariate analysis showed that only TSH considered as a continuous variable (p = 0.001) as well as subclinical hypothyroidism (TSH &gt5.5 mUI/l; p=0.014) remained significantly associated with the events. Conclusions. In CHF patients TSH levels even slightly above normal range are independently associated with a greater likelihood of heart failure progression. This supports the need for prospective studies aimed at clarifying the most appropriate therapeutic approach to sub-clinical hypothyroidism in such patients.

We examined the relationship between moderate obesity and glucose metabolism, insulin sensitivity and suspected fatty liver in children. We measured body mass index (BMI), z-score BMI, caliper skinfold thickness, waist and hip circumference in 94 participants (mean age 9.7 ±2.2 years). Fasting blood glucose, insulin, HOMA score, lipid profile and transaminases (ALT, AST) were measured. Fatty liver and skinfold thickness were evaluated by means of ultrasound. The z-score BMI was 2.01 ±0.39 (mean ± SD), and the duration of obesity was 4.3±3.03 years. A positive correlation was found between caliper and US skinfold thickness for tricipital (r= 0.33; p= 0.003) and sovrailiac skinfold (r= 0.34; p=0.003). Fatty liver was diagnosed in 64% of children and it was positively related to anthropometric measurements. The three sub-groups-group 0 (normal US liver and normal transaminases); group 1 (US fatty liver and normal transaminases); group 2 (US fatty liver and elevated transaminases)-showed a difference concerning z-score BMI, insulin and HOMA parameters (Tukey test: z score BMI group 1 vs group 0 and 2 vs group 0; serum insulin: group 2 vs group 1 and group 2 vs group 0; HOMA IR: group 2 vs group 1 and group 2 vs group 0). Moderately obese children with steatosis exhibited a clear increase of insulin and insulin resistance which represents indices of a future metabolic syndrome. In addition, it is important to perform a liver ultrasound since transaminases seems to be not adequate for the diagnosis of fatty liver.

Insulin resistance and obesity are intimately related to a chronic low grade systemic inflammation. Interleukin- 6 (IL-6) may influence the pathogenesis of obesity-related diseases. The aim of this study is to investigate the effect of body's fat mass on the relationships between -174G/C IL-6 promoter gene polymorphism, IL-6 circulating level and insulin resistance. A population of 150 Caucasian women was studied, subdivided according to their body composition in non-obese (NW), Normal Weight Obese (NWO) and preobese-obese (OB). The NWO subjects were found in an intermediate position between the NW and OB subjects in terms of body weight, fat mass percentage (FM%), abdominal FAT%, hs-CRP and plasma triglyceride level. Fasting plasma IL-6 concentration was positively correlated with the homeostasis model assessment for insulin resistance (HOMA-IR) in all subjects analyzed (P=0.0014). In NWO and OB women a significantly increased IL-6 mean value was observed compared with NW subjects. In G/G population, the IL-6 plasma level of NWO and OB was significantly higher with respect to NW. No significant differences of IL-6 concentrations were observed in the three groups carrying G/C genotype. NWO and OB women homozygous for the allele C have significantly lower value of IL-6 with respect to NW subjects. IL-6 concentration was positively correlated with FM% in G/G (R2=0.397, P<0.001) and was negatively correlated in C/C (R2=0.459, P=0.002). No significant correlation was observed in G/C genotype (R2=0.041, P=0.173). In conclusion our study confirms that, at least in Italian Caucasian females, the FM% is a major determinant of an increase in IL-6 production and insulin resistance. -174 G/C IL-6 promoter polymorphism represents a marker which could help to identify, time in advance, “vulnerable” individuals at risk of age and obesity related diseases.

An integrative synthesis of concepts and an explosion of experimental and epidemiological findings allow new insights as to how the interactions of genetic, environmental, dietary, cultural (social, psychological, economic) factors can influence the aging and diseases of aging processes. Although the net effect of the best dietary maintenance of homeostatic control of cell proliferation, cell differentiation and apoptosis, systems breakdown of the human being and death will inevitably be the ultimate end result. Reduction of the quantity of the stem cell pool in any tissue will affect the “aging” of that organ. This, in turn, will affect the homeostatic maintenance of the organ systems of the human. Clearly, not all organs of the body age uniformly. The quality of the stem cells in any organ, depending on circumstances, can contribute to various disease pathogeneses. In the case where the quality of the stem cells is altered in utero or early postnatal development by some mutagenic mechanism that could lead to the initiation step of carcinogenesis, then the individual can, to some degree, control the fate of those prenatally and early postnatally-derived initiated stem cells by choosing those environmentally and dietary factors that either enhance or prevent the clonal expansion of these initiated stem cells during the promotion phase of carcinogenesis. This might explain the Barker hypothesis which suggests that prenatal and early postnatal exposures to toxic agents can lead to diseases later in life.

Ageing is an inevitable biological process with gradual and spontaneous biochemical and physiological changes and increased susceptibility to diseases. Some nutritional factors (zinc, niacin, selenium) may remodel these changes leading to a possible escaping of diseases, with the consequence of healthy ageing, because they are involved in improving immune functions, metabolic homeostasis and antioxidant defence. Experiments performed “in vitro” (human lymphocytes exposed to endotoxins) and “in vivo” (old mice or young mice with low zinc dietary intake) show that zinc is important for immune efficiency (both innate and adaptive), metabolic homeostasis (energy utilization and hormone turnover) and antioxidant activity (SOD enzyme). Niacin is a precursor of NAD+, the substrate for the activity of DNA repair enzyme PARP-1 and, consequently, may contribute to maintaining genomic stability. Selenium provokes zinc release by Metallothioneins (MT), via reduction of glutathione peroxidase. This fact is crucial in ageing because high MT may be unable to release zinc with subsequent low intracellular free zinc ion availability for immune efficiency, metabolic harmony and antioxidant activity. Taking into account the existence of zinc transporters (ZnT and ZIP family) for cellular zinc efflux and influx, respectively, the association between zinc transporters and MT is crucial in maintaining satisfactory intracellular zinc homeostasis in ageing. Improved immune performance, metabolic homeostasis, antioxidant defence occur in elderly after physiological zinc supplementation, which also induces prolonged survival in old, nude and neonatal thymectomized mice. The association “zinc plus selenium” improves humoral immunity in old subjects after influenza vaccination. The association “zinc plus niacin” in elderly is actually in progress.

Many studies have been conducted on the effects of red wine polyphenols on certain diseases, primarily, coronary heart disease (CHD) and, in this respect, evidence has been demonstrated that intake of red wine is associated with a reduction of CHD symptomatology. In this framework, the purpose of this review is to illustrate the effects of polyphenols on immune cells from human healthy peripheral blood. Data will show that polyphenols are able to stimulate both innate and adaptive immune responses. In particular, the release of cytokines such as interleukin (IL)-12, interferon (IFN)-γ, and IL-10 as well as immunoglobulins may be important for host protection in different immune related disorders. Another important aspect pointed out in this review is the release of nitric oxide (NO) from peripheral blood mononuclear cells (PBMC), stimulated by red wine polyphenols despite the fact that the majority of studies have reported NO production only by endothelial cells. Release of NO from PBMC may play an important role in cardiovascular disease, because it is known that this molecule acts as an inhibitor of platelet aggregation. On the other hand, NO exerts a protective role against infectious organisms. Finally, some molecular cytoplasmatic pathways elicited by polyphenols able to regulate certain immune responses will also be discussed. In particular, it seems that p38, a molecule belonging to the MAPK family, is involved in the release of IFN-γ and, therefore, in NO production. All these data confirm the beneficial effects of polyphenols in some chronic diseases.

Polyphenols contained in red wine possess a broad array of properties which seem to be beneficial to human and animal health. We have investigated the ability of red wine polyphenols to promote the in vitro release of both proinflammatory and antiinflammatory cytokines from human healthy mononuclear cells, as well as of immunoglobulins from B cells. Following red wine (Negroamaro) pretreatment of lymphomonocytes, results will show a production of regulatory [Interleukin( IL)-12], proinflammatory (IL-1β and IL-6), and anti-inflammatory (IL-10) cytokines, as well as of IgA and IgG. The fine balance between inflammation and antiinflammation, as well as the role of humoral immune response either systemic or mucosal will be discussed as a consequence of red wine intake. Finally, since ageing is characterized by a decline of many immune functions, our results suggest that moderate use of red wine may be beneficial in age-related disorders where the host immune response is very often not effective against a variety of antigens.

Red wine represents a source of polyphenols which exhibit a number of biological effects on various systems. In this respect, there is evidence that red wine polyphenols constitute one of the ingredients of the Mediterranean diet which is associated to a reduced risk of coronary heart disease according to current literature. Here, we have evaluated in vitro the molecular mechanisms elicited by polyphenols from red wine (Negroamaro) on human healthy mononuclear cells. In particular, we have investigated the involvement of polyphenols in the activation of p38 and ERK1/2 molecules belonging to the MAPK kinase family and on the expression of IκBα and p65/NFκB. Results will demonstrate that in cells both the expression of p38 and ERK1/2 augments in the presence of red wine polyphenols, but their expression drops in the presence of polyphenols plus lipopolysaccharides (LPS). This indicates that in Gram-negative infections polyphenols may attenuate triggering of inflammatory mediators as a response to LPS stimulation. Finally, the regulatory role of polyphenols on IκBα and p65/NFκB expression is discussed, pointing out that red wine might favor anti-atherogenic mechanisms in the course of cardiovascular disease.