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Genetic association studies of age-related, chronic human diseases often suffer from a lack of power to detect modest effects. Here we propose an alternative approach of including healthy centenarians as a more homogeneous and extreme control group. As a proof of principle we focused on type 2 diabetes (T2D) and assessed allelic/genotypic associations of 31 SNPs associated with T2D, diabetes complications and metabolic diseases and SNPs of genes relevant for telomere stability and age-related diseases. We hypothesized that the frequencies of risk variants are inversely correlated with decreasing health and longevity. We performed association analyses comparing diabetic patients and non-diabetic controls followed by association analyses with extreme phenotypic groups (T2D patients with complications and centenarians). Results drew attention to rs7903146 (TCF7L2 gene) that showed a constant increase in the frequencies of risk genotype (TT) from centenarians to diabetic patients who developed macro-complications and the strongest genotypic association was detected when diabetic patients were compared to centenarians (p_value = 9.066*10−7). We conclude that robust and biologically relevant associations can be obtained when extreme phenotypes, even with a small sample size, are compared.

The effect of exposure to extremely low-frequency pulsed electromagnetic fields (EMFs) on DNA repair capability and on cell survival in human lymphocytes damaged in vitro with γ rays was studied by...

Aging is a complex process which is accompanied with the decline and the reshaping of different functions of the body. In particular the immune system is characterized, during ageing (immunosenescence) by a remodeling of innate immunity (well preserved, up-regulated) and clonotypical immunity (severely altered) and by the occurrence of a chronic inflammatory process (inflammaging) which are, at least in part, genetically controlled. In this scenario, it can be anticipated that a crucial role is played by age-related structural and functional alterations and modifications of proteasomes and immunoproteasomes, the last being a key component of antigen processing and MHC class I antigen presentation. A variety of experimental data are available, suggesting that proteasomes are affected by age, and that in centenarians they are relatively preserved. On the contrary, few data are available on immunoproteasomes, likely as a consequence of the poverty of suitable cellular models. Lymphoblastoid cell lines from EBV immortalized B cells from old donors is envisaged as a possible model for the study of immunoproteasomes in humans and their changes with age. Thus, basic questions such as those related to possible consequences, for immune responses in infectious diseases and cancer, of age-related alterations of antigen processing and presenting, change with age of self-antigen repertoire, and the genetic basis of immunoprotesome activity and its change with age, remain largely unanswered. © 2003 Elsevier Ireland Ltd. All rights reserved.

A significant broad substrate specificity, that crosses over the usual β-lactam derivatives, has been observed with an industrial glutaryl-7-aminocephalosporanic acid acylase (GA). This enzyme possesses significant enantioselective amidase and even esterase activity, with a stereopreference for the S-enantiomer. The easy separation of products from unreacted reagents, possessing different physical-chemical properties, is achieved by solvent extraction, avoiding chromatography or distillation during reaction work-up.

The production of cytokines during aging, except interleukin (IL)-2, has been neglected in humans. We measured the in vitro production of IL-6, tumor necrosis factor (TNF)-alpha, interferon (IFN)-gamma and IL-1 beta by peripheral mononuclear cells from selected healthy young (mean age 26.8 years) and aged (mean age 80.2 years) subjects. Significant increases of IL-6, TNF-alpha and IL-1 beta levels were found in mitogen-stimulated cultures from aged donors, occurring at 24 to 72 h after stimulation. No significant differences were observed for IFN-gamma production. Proliferative capability of cells stimulated with PHA was not impaired in aged subjects. Since the amounts of all cytokines studied were similar in unstimulated cultures from young and aged subjects, and also serum levels of TNF-alpha did not differ, these data indicate that the cellular machinery for the production of these cytokines is well preserved in aging, and also that cells from old people are able to up-regulate their production in response to appropriate stimuli. The increases in cytokine synthesis were not dependent on changes in the number of monocytes, nor were they related to the significant rise of CD45RO+, and the concomitant decrease of CD45RA+, occurring in both CD4+ and CD8+ lymphocytes from aged subjects. The increased production of pro-inflammatory cytokines by stimulated mononuclear cells of healthy aged subjects may be relevant to several aspects of age-associated pathological events, including atherosclerosis, osteoporosis, fibrosis and dementia.

Mitomycin-C (MMC), a potent antibiotic-antineoplastic drug, has recently proven to be useful as additional treatment in ocular surgery (pterygium, glaucoma filtering, photorefractive laser keratectomy, etc.) when inhibiting the epithelial wound healing response and reducing surgical scarring is beneficial to surgical success. However, the therapeutic benefits of MMC are frequently offset by ocular toxicity and undesirable side effects (retinal toxicity, corneal edema, corneal perforation, cataract, secondary glaucoma, iritis, scleral calcification, pain, etc.). The purpose of this study was to evaluate if a liposomal preparation containing MMC was capable of reducing the corneal healing rate and drug toxicity of a corneal lesion in a rabbit model. To do this, a liposomal formulation containing 0.2 mg/ml of MMC was prepared and tested against aqueous solutions and viscous formulations based on tamarind seeds polysaccharide (TSP), which were used as references. In vitro release of MMC from the vehicles through a dialysis membrane was characterized by a fast diffusion of MMC from the aqueous solution, while a more gradual release from the TSP and the liposomal formulations was observed. To evaluate the intraocular penetration of MMC, the drug amount in the aqueous and vitreous humor of the animals was determined 60 min after treatment. MMC toxicity was analyzed by monitoring the proliferation and viability of a rabbit corneal epithelial cell line (RCE) in the different formulations under study. These results showed reduced cytotoxicity for MMC in TSP viscous vehicles; however, only the liposomal MMC vehicle appeared to produce both lower toxicity for a rabbit corneal epithelial cell line (RCE) culture and a substantial reduction of the corneal healing rate in vivo.

The effect of the exposure of mitogen-stimulated lymphocytes from subjects infected by human immunodeficiency virus to extremely low frequency pulsed electromagnetic fields (PEMFs) was studied, by evaluating the incorporation of tritiated thymidine, the expression of IL-2 receptor, and the amount of activated T lymphocytes. Four groups of subjects were considered patients with acquired immunodeficiency syndrome (AIDS), asymptomatic seropositive subjects, seronegative drug users, and young healthy controls. PEMFs increased cell proliferation only in the group of healthy controls, as measured at the 72nd hour of culture, but an increase in the number of activated T lymphocytes was observed by cytofluorimetric analysis after 18 hrs of PEMF exposure in cultures from AIDS patients.

In continuation of a previous study dealing with the promoting effect of niaouli essential oil (NIA) on transdermal permeation of estrogens, the effect of vehicle composition on permeation of estradiol (ES) ex vivo across hairless mouse skin was investigated using a diffusion cell technique. Three types of gel vehicles possessing suitable esthetic and spreading characteristics, formulated with Carbopol1342 and containing NIA at two concentrations (1.0 and 10.0%) were tested. Promoter-free vehicles and two commercial alcohol-based gels (Gelestra and Sandrena) were used as reference. The solubility of ES in the solvent phase of the gels and the rheological properties of all formulations were also evaluated. The mean transdermal fluxes of ES at steady state from Gelestra and Sandrena were not significantly different, although Gelestra showed a shorter lag time and a greater amount of ES permeated after 30 h compared with Sandrena. In the gels under test, the presence of 1.0% NIA significantly increased the ES transdermal flux over the reference vehicles only when in association with ethanol and water. The present results may warrant further investigations on other drugs and skin models, aimed at optimizing the promoting activity of NIA, and the synergistic effect of solvents.

A wide difference in the susceptibility to undergo in vitro apoptosis exists among individuals, and this fact has potential implications in predicting the in vivo response to apoptotic agents, such as those employed in chemotherapy. In this report, we addressed the question whether the natural variability at p53 locus (the proline-arginine substitution at codon 72) affects the capacity of peripheral-blood mononuclear cells from healthy subjects to undergo in vitro apoptosis in response to the cytotoxic drug cytosine arabinoside. We found that cells from subjects carrying the arginine/arginine genotype undergo in vitro apoptosis at a higher extent in comparison to those from arginine/proline subjects. This finding suggests that naturally occurring genetic variability at p53 gene explains part of the inter-individual difference in the in vitro susceptibility to a chemotherapeutic drug, thus resulting as an eligible predictor marker of in vivo response to chemotherapy and its adverse effects.

The effect of extremely-low-frequency electromagnetic fields on in-vitro mitogen-induced lymphocyte proliferation was studied in two groups of healthy subjects (23 and 85 years, mean ages). An increase of cell proliferation was observed to be much higher in the group of aged subjects. The effects depended on the mitogen dose and involved interleukin-2 production and/or utilization. The DNA repair capability and cell survival in the exposed cultures after gamma irradiation was also evaluated, and no genotoxic effect was found. >

Thymectomy (Tx) is a common therapeutic option to treat myasthenia gravis (MG), but its effects on the immune system are still obscure in humans.We sought to evaluate long-term immunologic effects of therapeutic Tx in patients with MG.T- and B-cell subsets and T-cell repertoire were analyzed in 35 patients with MG, 16 with previous Tx (at least 8 years before), 6 with recent (1 year) Tx, and 13 without Tx, as well as in 32 healthy subjects used as normal control subjects. Serum immunoglobulins and a variety of autoantibodies were also measured. A subsequent 3-year clinical follow-up was performed to verify the possible appearance of systemic autoimmune diseases.The long-term thymectomized (Txd) patients had mild T-cell lymphopenia and an expansion of some Vbeta families among circulating CD4+ and CD8+ T cells. They displayed a normal number of total B and CD5+ B-circulating lymphocytes, but they also displayed a polyclonal increase in serum IgM and IgG associated with the presence of high levels of a variety of organ- and nonorgan-specific autoantibodies, including anti-dsDNA and anticardiolipin, without clinical evidence of autoimmune disease. These serologic abnormalities were not detectable in both non-Txd and recently Txd patients. After 3 years, 2 long-term Txd patients had systemic lupus erythematosus and an undifferentiated connective tissue disease.The association between MG and laboratory findings of systemic autoimmune disease may be in part related to Tx rather than to MG. Tx may represent a risk for the development of systemic autoimmune disorders over years in patients with MG.

A common polymorphism at codon 72 in p53 gene leads to an arginine to proline aminoacidic substitution which affects in an age-dependent manner the susceptibility of cells to undergo apoptosis after oxidative stress. Here we report that dermal fibroblasts from Proline allele carriers (Pro+) display a higher expression of p21WAF1 gene, in both basal conditions and after treatment with doxorubicin or camptothecin. This phenomenon is accompanied by a lower susceptibility of Pro+ cells to undergo apoptosis, a lower capability to over cross G1-S transition and an increased propensity to express markers of cell senescence, with respect to fibroblasts from Arginine homozygotes (Pro-). All these phenomena are particularly evident in cells from centenarians. We conclude that the functional difference between the two p53 codon 72 alleles exerts a broad impact on the capability of cell from aged people to respond to stressors such as cytotoxic drugs.

The exposure of human peripheral blood mononuclear cells to extremely low frequency pulsed electromagnetic fields (PEMFs) increased both the spontaneous and the PHA- and TPA-induced production of interleukin-1 (IL-1) and IL-6. Our results suggest that cells of the monocytic lineage, which are good producers of both IL-1 and IL-6, can be important cellular targets for PEMFs. Taking into account that these cytokines are among the most pleiotropic ones, these data can help us understand the previous reported effects of PEMFs on the proliferation of human lymphocytes and the effects exerted by such fields on cartilage and bone cells, whose physiological activity is highly dependent on IL-1 and IL-6.

Phenotype and proliferative ability of peripheral blood lymphocytes from 15 noninstitutionalized children affected with Down Syndrome (DS), in apparently good health, were studied and compared with those of 16 healthy control children of the same age. A complex derangement of all the major peripheral blood cell subsets, i.e., B cells, T cells, and natural killer (NK) cells, was present in DS children. A significant decrease of the absolute number of circulating lymphocytes, a marked and significant decrease of B lymphocyte absolute number and percentage, and dramatic modifications of the T-cell subsets were observed. The absolute number of CD4+ cells was significantly decreased, whereas CD8+ cells increased significantly in percentage but not in absolute number. A derangement of cells bearing markers associated with NK activity, such as CD57, CD16, and CD56, was observed. Among the most important alterations, the presence of a high number of CD57+, CD16- cells, of CD57+, CD8+ lymphocytes, and of CD3+, CD56+ lymphocytes was seen. Many of these alterations are similar to those characteristic of chromosomally normal subjects of advanced age. The hypothesis that the reduced thymic endocrine activity and the zinc deficiency characteristic of DS are responsible for the derangement of T and NK subsets is discussed.

The ene-reductase-mediated reduction of the carbon-carbon double bond of some alkyl 2-substituted butenedioates was investigated. The stereochemical outcome of the reaction was found to be influenced by steric effects. Ethyl and butyl citraconates were converted into the corresponding alkyl (R)-2-methylsuccinates with excellent enantioselectivity, whereas ethyl and butyl mesaconates were completely unreactive. Methyl 2-substituted fumarates were reduced to enantiomerically enriched methyl (S)-2-substituted succinates, whereas the (Z)-stereoisomers were left unreacted by ene-reductases. Labelling experiments were performed to investigate the mechanism of these bioreductions and explain their stereochemical outcome.

Free radicals are formed in the body as a consequence of aerobic metabolism. Cells have developed a variety of antioxidant systems, that include classical antioxidant enzymes (superoxide dismutase, glutathione peroxidase, and catalase) as well as nonenzymatic oxy-radicals scavengers (vitamin E, urea, s-carotene and some more recently described substances such as carnosine) (1). However, a certain fraction of active oxygen species escapes the cellular defence and may cause transient or permanent damage to cellular components. According to one of the most interesting theory of aging is the “free radical theory of aging”, proposed by D. Harman (2) more than thirty years ago, where oxidative damage has been suggested as a major cause of aging. One of the prediction of this theory is an age-related decrease of the efficiency of antioxidant defence mechanisms.

Ciclopirox is a topical antimycotic agent belonging to the chemical class of hydroxypyridones and not related to azoles or any other class of antifungal agents. Its antimicrobial profile includes nearly all of the clinically relevant dermatophytes, yeasts and moulds, and is therefore broader than that of most other antimycotics. It is also active against certain frequently azole-resistant Candida species and against some bacteria. The mechanism of action of ciclopirox is different from that of other topical antifungal drugs, which generally act through ergosterol inhibition. The high affinity of ciclopirox for trivalent metal cations, resulting in inhibition of the metal-dependent enzymes that are responsible for the degradation of peroxides within the fungal cell, appears to be the major determinant of its antimicrobial activity. This unique and multilevel mechanism of action provides a very low potential for the development of resistance in pathogenic fungi, with cases of resistance rarely reported. Ciclopirox also displays mild anti-inflammatory effects in biochemical and pharmacological models; effects also shown in small clinical studies. Scavenging of reactive oxygen species released from inflammatory cells is a likely contributor to these anti-inflammatory effects. Ciclopirox, and its olamine salt, is available in multiple topical formulations, suitable for administration onto the skin and nails and into the vagina. The pharmaceutical forms most widely investigated are 1% ciclopirox olamine cream and 8% ciclopirox acid nail lacquer, but lotion, spray, shampoo, pessary, solution, gel and douche formulations have also been used. Ciclopirox penetrates into the deep layers of the skin, mucosal membranes and nail keratin, reaching concentrations exceeding the minimal fungicidal concentrations for most medically important fungi. A large number of clinical trials were and are still being performed with ciclopirox, starting in the early 1980s. Ciclopirox was first developed for fungal skin infections and vaginal candidiasis, and is currently well established in these indications. More recently, the drug has been clinically investigated in seborrhoeic dermatitis and onychomycosis, showing good efficacy and excellent tolerability. Emphasis in this review is given to a ciclopirox medicated nail lacquer, which is based on an original technology and has superior properties in terms of its affinity to keratin and nail permeation. It has been found to have superior efficacy and safety to another commercially available formulation in the treatment of onychomycosis. The safety features of ciclopirox are well known. The topical drug is devoid of systemic adverse reactions. Mild local reactions characterized by a burning sensation of the skin, irritation, redness, pain or pruritus, generally in less than 5% of treated patients, can be observed following skin and vaginal application. With nail application, the most common adverse event is the appearance of mild erythema in 5% of the treated population. As a general conclusion, although less effective than some oral antimycotic agents in various indications, ciclopirox compares very well in terms of the benefit/risk ratio due to its excellent tolerability and complete absence of serious adverse effects.

In this study we assessed whether widely accepted risk factors for atherosclerotic vascular diseases such as lipoprotein(a) [Lp(a)], a cholesterol-rich lipoprotein under strict genetical control, and other lipid parameters change with age. The variations of blood levels and the pathophysiological role of Lp(a) in old people, and particularly in the oldest old, are unknown. Accordingly, we measured Lp(a) levels as well as total, LDL, and HDL cholesterol (CT), and triglycerides (TG) in sera from 75 healthy centenarians, 114 randomly selected subjects under 65 years, 73 randomly selected elderly people, and 30 healthy selected elderly people. The results showed that Lp(a) serum levels did not vary by age group, including centenarians. Remarkably, one-quarter of the centenarians had high Lp(a) serum levels even though they never suffered from atherosclerosis-related diseases. At variance with young and aged people, centenarians with high Lp(a) serum levels also had high plasma concentrations of the proinflammatory cytokine IL-6, suggesting that genetic control of the Lp(a) serum level may attenuate with age and that environmental factors such as chronic subclinical inflammatory processes may play a role. We also showed that most centenarians are paradoxically characterized by low HDL-CT and relatively high TG levels, which together are considered to be strong risk factors for coronary heart disease. On the whole, these data support the hypothesis that a continuous and complex reshaping of lipid metabolism occurs in physiological aging, likely contributing to successful aging.

Stopped flow experiments gave evidence of the formation of a biliverdin-superoxide complex and/or a biliverdin radical anion by reaction of aqueous O2- with biliverdin. Such transient species are likely intermediates both in the bleaching of biliverdin, during exposure to the aerobic xanthine oxidase reaction, and in the reduction of ferricytochrome c under the same conditions.

Since 2018, we are living in a world where there are more people over age 65 than there are children under five. Predictions indicate, if this trend continues, by the year 2050, the number of people over 65 will be double the number of people under five (1). Consequently, an understanding of the optimal physiological, endocrinological, and anthropometric conditions associated with better health during aging is to be considered a priority topic. In parallel with the increasing aging of the population, there is a parallel increase of overweight and obese individuals among older adults (2). Normal aging involves important changes to body composition, including decreased muscle mass and increased fat mass (3). Basal metabolism, for the majority of the elderly, is the main daily energetic expenditure and its decrease with age provides one explanation for the tendency to gain weight, with age. In addition to this physiological statement, lifestyle changes in aged people and the associated reduction in physical activity level favors weight increase with age. Total body fat peaks at about 65–70 years, while in advanced old age it decreases. Aging, indeed, modifies adipose tissue accumulation and redistribution resulting in accumulation of abdominal fat. These age-related changes alter many physiological functions including inflammation and contribute to age-related diseases such as cardiovascular events, diabetes mellitus, hypertension, stroke, and several types of cancer (4). However, to what extent, the age-related adipose tissue remodeling impacts the health status in elderly is incompletely understood. To highlight and clarify the main age-related changes in adipose tissue and discuss its implications on health status with particular regard to age-related diseases, we dedicated a Research Topic to the alteration of lipid storage, the redistribution and the types of fat, the production of different mediators contributing to a pro-inflammatory status in aging.