Organ-specific biological clocks: Ageotyping for personalized anti-aging medicine.

Aging is a complex multidimensional, progressive remodeling process affecting multiple organ systems. While many studies have focused on studying aging across multiple organs, assessment of the contribution of individual organs to overall aging processes is a cutting-edge issue. An organ's biological age might influence the aging of other organs, revealing a multiorgan aging network. Recent data demonstrated a similar yet asynchronous inter-organs and inter-individuals progression of aging, thereby providing a foundation to track sources of declining health in old age. The integration of multiple omics with common clinical parameters through artificial intelligence has allowed the building of organ-specific aging clocks, which can predict the development of specific age-related diseases at high resolution. The peculiar individual aging-trajectory, referred to as ageotype, might provide a novel tool for a personalized anti-aging, preventive medicine. Here, we review data relative to biological aging clocks and omics-based data, suggesting different organ-specific aging rates. Additional research on longitudinal data, including young subjects and analyzing sex-related differences, should be encouraged to apply ageotyping analysis for preventive purposes in clinical practice.

Nanomicellar eye drops: a review of recent advances.

INTRODUCTION: Research on nanotechnology in medicine has also involved the ocular field and nanomicelles are among the applications developed. This approach is used to increase both the water solubility of hydrophobic drugs and their penetration/permeation within/through the ocular tissues since nanomicelles are able to encapsulate insoluble drug into their core and their small size allows them to penetrate and/or diffuse through the aqueous pores of ocular tissues. AREAS COVERED: The present review reports the most significant and recent literature on the use of nanomicelles, made up of both surfactants and amphiphilic polymers, to overcome limitations imposed by the physiology of the eye in achieving a high bioavailability of drugs intended for the therapeutic areas of greatest commercial interest: dry eye, inflammation, and glaucoma. EXPERT OPINION: The results of the numerous studies in this field are encouraging and demonstrate that nanomicelles may be the answer to some of the challenges of ocular therapy. In the future, new molecules self-assembling into micelles will be able to meet the regulatory requirements for marketing authorization for their use in ophthalmic formulations.

A comparison between the effects of two liposome-encapsulated bevacizumab formulations on ocular neovascularization inhibition.

Bevacizumab (BVZ), an anti-VEGF antibody, has demonstrated reliable outcomes in the treatment of irritating ocular neovascularization. Frequent intravitreal injections are necessitated due to rapid clearance and short local accessibility. We recruited liposome as a highly prevailing drug delivery system to enhance drug availability. Two liposome formulations were characterized and their in vitro stability was analyzed. The toxicity of the formulations on some ocular cell lines was also evaluated. In addition, the anti-angiogenic effects of formulations were examined. Drug permeation was measured across ARPE-19 and HCE cell lines as in vitro cellular barrier models. Results revealed that NLP-DOPE-BVZ acquired high stability at 4 °C, 24 °C, and 37 °C for 45 days. It also showed more capacity to entrap BVZ in NLP-DOPE-BVZ (DEE% 69.1 ± 1.4 and DLE% 55.66 ± 1.15) as compared to NLP-BVZ (DEE% 43.57 ± 14.64, and DLE% 37.72 ± 12.01). Although both formulations inhibited the migration and proliferation of HUVECs, NLP-DOPE-BVZ was more effective at inhibiting angiogenesis. Furthermore, NLP-DOPE-BVZ better crossed our established barrier cellular models. Based on the findings, the inclusion of DOPE in NLPs has significantly enhanced the features of drug carriers. This makes them a potential candidate for treating ocular neovascularization and other related ailments.

Binary Polymeric Surfactant Mixtures for the Development of Novel Loteprednol Etabonate Nanomicellar Eyedrops.

The treatment of several ocular inflammatory conditions affecting different areas of the ocular globe involves the administration of topical ophthalmic formulations containing corticosteroids. This research was aimed at evaluating the solubilising efficacy of 5.0% w/w of different binary mixtures of commercial amphiphilic polymeric surfactants with the purpose of obtaining nanomicellar solutions containing a high amount of loteprednol etabonate (LE). The selected LE-TPGS/HS nanomicelles, containing 0.253 mg/mL of the drug, had a small size (=13.57 nm) and uniform distribution (Polydispersity Index = 0.271), appeared completely transparent and perfectly filterable through 0.2 µm membrane filter, and remained stable up to 30 days at 4 °C. The critical micellar concentration (CMCTPGS/HS) was 0.0983 mM and the negative value of the interaction parameter between the polymeric-surfactant-building unit (ßTPGS/HS = -0.1322) confirmed the ability of the polymeric surfactants to interact, favouring the dissolution of LE into nanomicelles. The disappearance of the endothermic peak of LE in the DSC analysis confirmed the interactions of LE with the polymeric surfactants. LE-TPGS/HS produced in vitro LE which sustained diffusion for 44 h (more than 40% of encapsulated LE). Furthermore, the lack of a significant cytotoxic effect on a sensitive corneal epithelial cell line makes it a candidate for further biological studies.

Centenarian clocks: epigenetic clocks for validating claims of exceptional longevity.

Claims surrounding exceptional longevity are sometimes disputed or dismissed for lack of credible evidence. Here, we present three DNA methylation-based age estimators (epigenetic clocks) for verifying age claims of centenarians. The three centenarian clocks were developed based on n = 7039 blood and saliva samples from individuals older than 40, including n = 184 samples from centenarians, 122 samples from semi-supercentenarians (aged 105 +), and 25 samples from supercentenarians (aged 110 +). The oldest individual was 115 years old. Our most accurate centenarian clock resulted from applying a neural network model to a training set composed of individuals older than 40. An epigenome-wide association study of age in different age groups revealed that age effects in young individuals (age < 40) are correlated (r = 0.55) with age effects in old individuals (age > 90). We present a chromatin state analysis of age effects in centenarians. The centenarian clocks are expected to be useful for validating claims surrounding exceptional old age.

A Targeted Epigenetic Clock for the Prediction of Biological Age.

Epigenetic clocks were initially developed to track chronological age, but accumulating evidence indicates that they can also predict biological age. They are usually based on the analysis of DNA methylation by genome-wide methods, but targeted approaches, based on the assessment of a small number of CpG sites, are advisable in several settings. In this study, we developed a targeted epigenetic clock purposely optimized for the measurement of biological age. The clock includes six genomic regions mapping in ELOVL2, NHLRC1, AIM2, EDARADD, SIRT7 and TFAP2E genes, selected from a re-analysis of existing microarray data, whose DNA methylation is measured by EpiTYPER assay. In healthy subjects (n = 278), epigenetic age calculated using the targeted clock was highly correlated with chronological age (Spearman correlation = 0.89). Most importantly, and in agreement with previous results from genome-wide clocks, epigenetic age was significantly higher and lower than expected in models of increased (persons with Down syndrome, n = 62) and decreased (centenarians, n = 106; centenarians' offspring, n = 143; nutritional intervention in elderly, n = 233) biological age, respectively. These results support the potential of our targeted epigenetic clock as a new marker of biological age and open its evaluation in large cohorts to further promote the assessment of biological age in healthcare practice.

Sporopollenin Microcapsule: Sunscreen Delivery System with Photoprotective Properties.

In recent years, the demand for high-quality solar products that combine high efficacy with environmentally friendly characteristics has increased. Among the coral-safe sunscreens, ethylhexyl triazone (Uvinul® T150) is an effective organic UVB filter, photostable and practically insoluble in water, therefore difficult to be formulated in water-based products. Oil-free sunscreens are considered ideal for most skin types, as they are not comedogenic and do not leave the skin feeling greasy. Recent studies reported that pollen grains might represent innovative drug delivery systems for their ability to encapsulate and release active ingredients in a controlled manner. Before being used, the pollen grains must be treated to remove cellular material and biomolecules, which could cause allergic reactions in predisposed subjects; the obtained hollow structures possess uniform diameter and a rigid wall with openings that allow them to be filled with bioactive substances. In the present work, pollen from Lycopodium clavatum has been investigated both as a delivery system for ethylhexyl triazone and as an active ingredient by evaluating its photoprotective capacity. The goal is to obtain environmentally friendly solar aqueous formulations that take advantage of both sunscreen and sporopollenin microcapsules' UV protection with a relatively low cost, as these pollen grains are widely available.

Functional Characterization and Synthetic Application of Is2-SDR, a Novel Thermostable and Promiscuous Ketoreductase from a Hot Spring Metagenome.

In a metagenome mining-based search of novel thermostable hydroxysteroid dehydrogenases (HSDHs), enzymes that are able to selectively oxidize/reduce steroidal compounds, a novel short-chain dehydrogenase/reductase (SDR), named Is2-SDR, was recently discovered. This enzyme, found in an Icelandic hot spring metagenome, shared a high sequence similarity with HSDHs, but, unexpectedly, showed no activity in the oxidation of the tested steroid substrates, e.g., cholic acid. Despite that, Is2-SDR proved to be a very active and versatile ketoreductase, being able to regio- and stereoselectively reduce a diversified panel of carbonylic substrates, including bulky ketones, α- and ß-ketoesters, and α-diketones of pharmaceutical relevance. Further investigations showed that Is2-SDR was indeed active in the regio- and stereoselective reduction of oxidized steroid derivatives, and this outcome was rationalized by docking analysis in the active site model. Moreover, Is2-SDR showed remarkable thermostability, with an apparent melting temperature (TM) around 75 °C, as determined by circular dichroism analysis, and no significant decrease in catalytic activity, even after 5 h at 80 °C. A broad tolerance to both water-miscible and water-immiscible organic solvents was demonstrated as well, thus, confirming the potential of this new biocatalyst for its synthetic application.

Functionalized Hyaluronic Acid for "In Situ" Matrix Metalloproteinase Inhibition: A Bioactive Material to Treat the Dry Eye Sydrome.

Hyaluronic acid (HA) is a naturally occurring polysaccharide with many molecular functions, including maintaining the structure and physiology of the tissues, tissue remodeling, and inflammation. HA is found naturally in physiological tear fluid, possesses excellent mucus-layer-adhesive properties, and is successfully employed in the treatment of dry eye syndrome (DES). However, HA has as major drawback: its rapid in vivo degradation by hyaluronidase. We report on a unique material, namely, HA-3, obtained by the functionalization of HA with the metalloproteinase inhibitor 3 (MMPI). This material is characterized by an increased resistance to hyaluronidase degradation, associated with MMP inhibition properties. The ability of HA-3 to prevent dehydration of human corneal epithelial cells in vitro and in vivo may accelerate the development of more efficient DES treatment and broaden the application of HA in human diseases.

MAGL inhibitor NanoMicellar formulation (MAGL-NanoMicellar) for the development of an antiglaucoma eye drop.

The ocular endocannabinoid system (ECS) including enzymes and CB1/CB2 receptors determines various substantial effects, such as anti-inflammatory activity and reduction of the intraocular pressure (IOP). The modulation of 2-arachidonoylglycerol (2-AG) levels obtained via MAGL inhibition is considered as a promising pharmacological strategy to activate the ECS. Within the scope of this study, the effect of a selective monoacylglycerol lipase (MAGL) inhibitor (MAGL17b) was investigated by measuring the IOP reduction in normotensive rabbits after performing a solubilisation process of the molecule with non-ionic surfactants, to produce suitable eye drops containing the highest possible concentration of the drug. Furthermore, the study involved the evaluation of cytotoxicity and of in vitro/ex vivo corneal permeation of MAG17b of selected formulations based on polyoxyl(35)castor oil (C-EL) and polyethylene glycol (80) sorbitan monolaurate (TW80). The solubilisation of 0.5 mM MAGL17b with 3 % w/w TW80 (TW80/3-17b), through the formation of NanoMicellar structures (diameter of 12.3 nm), determined a significant permeation of MAGL17b, both through excised rabbits corneas and reconstituted corneal epithelium, with a limited corneal epithelial cells death. The blockade of MAGL activity induced a IOP reduction up to 4 mmHg in albino and pigmented rabbits after topical instillation, thus confirming the potential efficacy of the MAGL inhibition approach in the treatment of ocular pathologies.

Selective Supercritical CO2 Extraction and Biocatalytic Valorization of Cucurbita pepo L. Industrial Residuals.

The valorization of biomass residuals constitutes a key aspect of circular economy and thus a major challenge for the scientific community. Among industrial wastes, plant residuals could represent an attractive source of bioactive compounds. In this context, a residue from the industrial extraction of Cucurbita pepo L. seeds, whose oil is commercialized for the treatment of genito-urinary tract pathologies, has been selected. Supercritical CO2 technology has been employed as a highly selective "green" methodology allowing the recovery of compounds without chemical degradation and limited operational costs. Free fatty acids have been collected in mild conditions while an enrichment in sterols has been selectively obtained from sc-CO2 extracts by appropriate modulation of process parameters (supercritical fluid pressure and temperature), hence demonstrating the feasibility of the technique to target added-value compounds in a selective way. Obtained fatty acids were thus converted into the corresponding ethanol carboxamide derivatives by lipase-mediated biocatalyzed reactions, while the hydroxylated derivatives of unsaturated fatty acids were obtained by stereoselective hydration reaction under reductive conditions in the presence of a selected FADH2-dependent oleate hydratase.

Ciclopirox Hydroxypropyl Chitosan (CPX-HPCH) Nail Lacquer and Breathable Cosmetic Nail Polish: In Vitro Evaluation of Drug Transungual Permeation Following the Combined Application.

BACKGROUND: Onychomycosis produces nail chromatic alterations that lead patients to mask them with cosmetic enamels. Objectives: Evaluate drug transungual permeation and antimycotic activity against selected strains after application of CPX-HPCH nail lacquer (NL) on the nail pre-covered with breathable cosmetic polish. METHODS: CPX transungual permeation after applying CPX-HPCH NL once or twice a day on bovine hoof membranes pre-covered with a breathable cosmetic nail polish was compared to that obtained applying CPX-HPCH NL directly on the membrane. The relevant experimental permeates underwent an in vitro susceptibility test. RESULTS: After CPX-HPCH NL application once a day, the drug transungual flux in the presence of cosmetic product tended to decrease while maintaining the antifungal activity. Two daily applications of CPX-HPCH NL on the membrane pre-covered with cosmetic polish exhibited the same permeation profile as daily application of the medicated lacquer directly on the nail as well as the same microbiological activity. CONCLUSIONS: The breathable cosmetic nail polish can be applied on the nail affected by onychomycosis in association with CPX-HPCH NL to mask the imperfections. The application of CPX-HPCH NL twice a day appears to be a good solution to obtain the same results as for a daily application without the presence of the cosmetic layer.

Doxorubicin-induced senescence in normal fibroblasts promotes in vitro tumour cell growth and invasiveness: The role of Quercetin in modulating these processes.

Ageing is a complex biological phenomenon representing the major risk factor for developing age-related diseases, such as cardiovascular pathologies, neurodegenerative diseases, and cancer. Geroscience, the new vision of gerontology, identifies cellular senescence as an interconnected biological process that characterises ageing and age-related diseases. Therefore, many strategies have been employed in the last years to reduce the harmful effects of senescence, and among these, the most intriguing ones use nutraceutical compounds. Here we show that a pre-treatment with Quercetin, a bioactive flavonoid present in many fruits and vegetables, increasing cellular antioxidant defence, can alleviate Doxorubicin (Doxo)-induced cellular senescence in human normal WI-38 fibroblasts. Furthermore, our work demonstrates that Quercetin pre-treatment, reducing the number of senescent cells and the production of the senescence-associated secretory phenotype (SASP) factors, can decrease the pro-tumour effects of conditioned medium from Doxo-induced senescent fibroblasts on osteosarcoma cells. Overall, our findings are consistent with the hypothesis that targeting senescent cells can be an emerging strategy for cancer treatment, especially in elderly patients, in which senescent cells are already abundant in several tissues and organs.

DNA Methylation Analysis of Ribosomal DNA in Adults With Down Syndrome.

Control of ribosome biogenesis is a critical aspect of the regulation of cell metabolism. As ribosomal genes (rDNA) are organized in repeated clusters on chromosomes 13, 14, 15, 21, and 22, trisomy of chromosome 21 confers an excess of rDNA copies to persons with Down syndrome (DS). Previous studies showed an alteration of ribosome biogenesis in children with DS, but the epigenetic regulation of rDNA genes has not been investigated in adults with DS so far. In this study, we used a targeted deep-sequencing approach to measure DNA methylation (DNAm) of rDNA units in whole blood from 69 adults with DS and 95 euploid controls. We further evaluated the expression of the precursor of ribosomal RNAs (RNA45S) in peripheral blood mononuclear cells (PBMCs) from the same subjects. We found that the rDNA promoter tends to be hypermethylated in DS concerning the control group. The analysis of epihaplotypes (the combination of methylated and unmethylated CpG sites along the same DNA molecule) showed a significantly lower intra-individual diversity in the DS group, which at the same time was characterized by a higher interindividual variability. Finally, we showed that RNA45S expression is lower in adults with DS. Collectively, our results suggest a rearrangement of the epigenetic profile of rDNA in DS, possibly to compensate for the extranumerary rDNA copies. Future studies should assess whether the regulation of ribosome biogenesis can contribute to the pathogenesis of DS and explain the clinical heterogeneity characteristic of the syndrome.

Hydrogels as Corneal Stroma Substitutes for In Vitro Evaluation of Drug Ocular Permeation.

Hydrogels are complex hydrophilic structures, consisting of crosslinked homopolymers or copolymers insoluble in water. Due to their controllable bio-physicochemical properties mimicking the morphology of the native extracellular matrix, they are a key part of a lot of research fields, including medicine, pharmaceutics, and tissue engineering. This paper was focused on the preparation and characterization of hydrogels from different blends of polyvinyl alcohol (PVA) with microcrystalline cellulose (MCC) and gelatin (GEL) at various ratios, and from gelatin and chitosan alone to understand their feasibility of utilizing as corneal stroma substitutes in permeability tests for drug candidate molecules in early stages of their development. The characterization was carried out by differential scanning calorimetry, electron microscopy (SEM), water content, mass loss, water permeability, wettability, and tensile stress-strain tests. After the physicochemical characterization, PVA/MCC blend and chitosan proved to be the most promising constructs, showing negligible mass loss after immersion in aqueous medium for two weeks and low hydrodynamic permeability. They were then employed in drug molecules permeation studies and these data were compared to that obtained through excised tissues. The results obtained showed that PVA/MCC hydrogels have similar mechanical and permeability properties to corneal stroma.

Nanostructured Drug Delivery Systems for Targeting 5-α-Reductase Inhibitors to the Hair Follicle.

Androgenetic alopecia is a multifactorial condition characterized by noticeable hair loss, affecting both men and women and representing a debilitating and chronic disorder that considerably affects the quality of life. Available topical treatments based on minoxidil or finasteride require repeated applications and are associated with a certain number of adverse effects. The challenges associated with current treatments pave the way for the research of new therapeutic strategies, more precise and selective, and capable of providing long-term results. In this context, the present review examines the new proposed formulation strategies to deliver 5-α-reductase inhibitors in order to obtain a targeted drug delivery, for improving drug retention at the site of action in the hair follicle, contemporaneously reducing drug systemic absorption, which is the cause of important adverse effects. In particular, the research will be focused on the several aspects that influence the performance of nanostructured drug delivery systems in creating a depot in the hair follicles, such as particle size, surface charge, excipients, and combined application with external stimuli (infrared radiation, mechanical massage, ultrasounds application).

Stereoselective Biocatalyzed Reductions of Ginger Active Components Recovered from Industrial Wastes.

Ginger is among the most widespread and widely consumed traditional medicinal plants around the world. Its beneficial effects, which comprise e. g. anticancer and anti-inflammatory activities as well as gastrointestinal regulatory effects, are generally attributed to a family of non-volatile compounds characterized by an arylalkyl long-chained alcohol, diol, or ketone moiety. In this work, ginger active components have been successfully recovered from industrial waste biomass of fermented ginger. Moreover, their recovery has been combined with the first systematic study of the stereoselective reduction of gingerol-like compounds by isolated alcohol dehydrogenases (ADHs), obtaining the enantioenriched sec-alcohol derivatives via a sustainable biocatalytic path in up to >99 % conversions and >99 % enantiomeric/diastereomeric excesses.

Association of rs3027178 polymorphism in the circadian clock gene PER1 with susceptibility to Alzheimer's disease and longevity in an Italian population.

Many physiological processes in the human body follow a 24-h circadian rhythm controlled by the circadian clock system. Light, sensed by retina, is the predominant "zeitgeber" able to synchronize the circadian rhythms to the light-dark cycles. Circadian rhythm dysfunction and sleep disorders have been associated with aging and neurodegenerative diseases including mild cognitive impairment (MCI) and Alzheimer's disease (AD). In the present study, we aimed at investigating the genetic variability of clock genes in AD patients compared to healthy controls from Italy. We also included a group of Italian centenarians, considered as super-controls in association studies given their extreme phenotype of successful aging. We analyzed the exon sequences of eighty-four genes related to circadian rhythms, and the most significant variants identified in this first discovery phase were further assessed in a larger independent cohort of AD patients by matrix assisted laser desorption/ionization-time of flight mass spectrometry. The results identified a significant association between the rs3027178 polymorphism in the PER1 circadian gene with AD, the G allele being protective for AD. Interestingly, rs3027178 showed similar genotypic frequencies among AD patients and centenarians. These results collectively underline the relevance of circadian dysfunction in the predisposition to AD and contribute to the discussion on the role of the relationship between the genetics of age-related diseases and of longevity.

Stable inversion clines in a grasshopper species group despite complex geographical history.

Chromosomal inversions are known to play roles in adaptation and differentiation in many species. They involve clusters of correlated genes (i.e., loci in linkage disequilibrium, LD) possibly associated with environmental variables. The grasshopper "species complex" Trimerotropis pallidipennis comprises several genetic lineages distributed from North to South America in arid and semi-arid high-altitude environments. The southernmost lineage, Trimerotropis sp., segregates for four to seven putative inversions that display clinal variation, possibly through adaptation to temperate environments. We analysed chromosomal, mitochondrial and genome-wide single nucleotide polymorphism data in 19 Trimerotropis sp. populations mainly distributed along two altitudinal gradients (MS and Ju). Populations across Argentina comprise two main chromosomally and genetically differentiated lineages: one distributed across the southernmost border of the "Andes Centrales," adding evidence for a differentiation hotspot in this area; and the other widely distributed in Argentina. Within the latter, network analytical approaches to LD found three clusters of correlated loci (LD-clusters), with inversion karyotypes explaining >79% of the genetic variation. Outlier loci associated with environmental variables mapped to two of these LD-clusters. Furthermore, despite the complex geographical history indicated by population genetic analyses, the clines in inversion karyotypes have remained stable for more than 20 generations, implicating their role in adaptation and differentiation within this lineage. We hypothesize that these clines could be the consequence of a coupling between extrinsic postzygotic barriers and spatially varying selection along environmental gradients resulting in a hybrid zone. These results provide a framework for future investigations about candidate genes implicated in rapid adaptation to new environments.

An inflammatory aging clock (iAge) based on deep learning tracks multimorbidity, immunosenescence, frailty and cardiovascular aging.

While many diseases of aging have been linked to the immunological system, immune metrics capable of identifying the most at-risk individuals are lacking. From the blood immunome of 1,001 individuals aged 8-96 years, we developed a deep-learning method based on patterns of systemic age-related inflammation. The resulting inflammatory clock of aging (iAge) tracked with multimorbidity, immunosenescence, frailty and cardiovascular aging, and is also associated with exceptional longevity in centenarians. The strongest contributor to iAge was the chemokine CXCL9, which was involved in cardiac aging, adverse cardiac remodeling and poor vascular function. Furthermore, aging endothelial cells in human and mice show loss of function, cellular senescence and hallmark phenotypes of arterial stiffness, all of which are reversed by silencing CXCL9. In conclusion, we identify a key role of CXCL9 in age-related chronic inflammation and derive a metric for multimorbidity that can be utilized for the early detection of age-related clinical phenotypes.