Protective Potential of a Botanical-Based Supplement Ingredient against the Impact of Environmental Pollution on Cutaneous and Cardiopulmonary Systems: Preclinical Study.

Air pollution is a growing threat to human health. Airborne pollution effects on respiratory, cardiovascular and skin health are well-established. The main mechanisms of air-pollution-induced health effects involve oxidative stress and inflammation. The present study evaluates the potential of a polyphenol-enriched food supplement ingredient comprising Lippia citriodora, Olea europaea, Rosmarinus officinalis, and Sophora japonica extracts in mitigating the adverse effects of environmental pollution on skin and cardiopulmonary systems. Both in vitro and ex vivo studies were used to assess the blend's effects against pollution-induced damage. In these studies, the botanical blend was found to reduce lipid peroxidation, inflammation (by reducing IL-1α), and metabolic alterations (by regulating MT-1H, AhR, and Nrf2 expression) in human skin explants exposed to a mixture of pollutants. Similar results were also observed in keratinocytes exposed to urban dust. Moreover, the ingredient significantly reduced pollutant-induced ROS production in human endothelial cells and lung fibroblasts, while downregulating the expression of apoptotic genes (bcl-2 and bax) in lung fibroblasts. Additionally, the blend counteracted the effect of urban dust on the heart rate in zebrafish embryos. These results support the potential use of this supplement as an adjuvant method to reduce the impact of environmental pollution on the skin, lungs, and cardiovascular tissues.

Small Molecule Pytren-4QMn Metal Complex Slows down Huntington's Disease Progression in Male zQ175 Transgenic Mice.

Huntington's disease (HD) is an inherited neurodegenerative disorder considered a rare disease with a prevalence of 5.7 per 100,000 people. It is caused by an autosomal dominant mutation consisting of expansions of trinucleotide repeats that translate into poly-glutamine enlarged mutant huntingtin proteins (mHTT), which are particularly deleterious in brain tissues. Since there is no cure for this progressive fatal disease, searches for new therapeutic approaches are much needed. The small molecule pytren-4QMn (4QMn), a highly water-soluble mimic of the enzyme superoxide dismutase, has shown in vivo beneficial anti-inflammatory activity in mice and was able to remove mHTT deposits in a C. elegans model of HD. In this study, we assessed 4QMn therapeutic potential in zQ175 neo-deleted knock-in mice, a model of HD that closely mimics the heterozygosity, genetic injury, and progressive nature of the human disease. We provide evidence that 4QMn has good acute and chronic tolerability, and can cross the blood-brain barrier, and in male, but not female, zQ175 mice moderately ameliorate HD-altered gene expression, mHtt aggregation, and HD disease phenotype. Our data highlight the importance of considering sex-specific differences when testing new therapies using animal models and postulate 4QMn as a potential novel type of small water-soluble metal complex that could be worth further investigating for its therapeutic potential in HD, as well as in other polyglutamine diseases.

Mn(II) Quinoline Complex (4QMn) Restores Proteostasis and Reduces Toxicity in Experimental Models of Huntington's Disease.

Huntington's disease (HD) is an autosomal dominant neurodegenerative disorder, of the so-called minority diseases, due to its low prevalence. It is caused by an abnormally long track of glutamines (polyQs) in mutant huntingtin (mHtt), which makes the protein toxic and prone to aggregation. Many pathways of clearance of badly-folded proteins are disrupted in neurons of patients with HD. In this work, we show that one Mn(II) quinone complex (4QMn), designed to work as an artificial superoxide dismutase, is able to activate both the ubiquitin-proteasome system and the autophagy pathway in vitro and in vivo models of HD. Activation of these pathways degrades mHtt and other protein-containing polyQs, which restores proteostasis in these models. Hence, we propose 4QMn as a potential drug to develop a therapy to treat HD.

An antioxidant boehmite amino-nanozyme able to disaggregate Huntington's inclusion bodies.

A novel amino-nanozyme, based on boehmite nanoparticles (BNPs) functionalised with a tetra-azapyridinophane (L1), has been designed to undermine some of the key issues underlying Huntington disease. L1 forms Cu2+ complexes with a striking SOD activity, while when grafted to the BNPs displays mitoROS scavenging properties and ability to disaggregate mutant huntingtin deposits in cells.

Use of Medaka Fish as Vertebrate Model to Study the Effect of Cocoa Polyphenols in the Resistance to Oxidative Stress and Life Span Extension.

Oxidative stress (OS) can induce cell apoptosis and thus plays an important role in aging. Antioxidant foods protect tissues from OS and contribute to a healthier lifestyle. In this study, we described the used of medaka embryos (Oryzias latipes) to study the putative antioxidant capacity of dietary cocoa extract in vertebrates. A polyphenol-enriched cocoa extract regulated the expression of several genes implicated in OS, thereby protecting fish embryos from induced OS. The cocoa extract activated superoxide dismutase enzyme activity in embryos and adult fish tissues, suggesting a common mechanism for protection during embryonic development and adulthood. Furthermore, long-term feeding of the cocoa extract increased fish life span. Our study demonstrates that the polyphenol-enriched cocoa extract decreases OS and extends life span in medaka fish, validating the use of medaka embryos as an economical platform to screen the antioxidant capacity of food compounds.

Analysis of the Ush2a gene in medaka fish (Oryzias latipes).

Patients suffering from Usher syndrome (USH) exhibit sensorineural hearing loss, retinitis pigmentosa (RP) and, in some cases, vestibular dysfunction. USH is the most common genetic disorder affecting hearing and vision and is included in a group of hereditary pathologies associated with defects in ciliary function known as ciliopathies. This syndrome is clinically classified into three types: USH1, USH2 and USH3. USH2 accounts for well over one-half of all Usher cases and mutations in the USH2A gene are responsible for the majority of USH2 cases, but also for atypical Usher syndrome and recessive non-syndromic RP. Because medaka fish (Oryzias latypes) is an attractive model organism for genetic-based studies in biomedical research, we investigated the expression and function of the USH2A ortholog in this teleost species. Ol-Ush2a encodes a protein of 5.445 aa codons, containing the same motif arrangement as the human USH2A. Ol-Ush2a is expressed during early stages of medaka fish development and persists into adulthood. Temporal Ol-Ush2a expression analysis using whole mount in situ hybridization (WMISH) on embryos at different embryonic stages showed restricted expression to otoliths and retina, suggesting that Ol-Ush2a might play a conserved role in the development and/or maintenance of retinal photoreceptors and cochlear hair cells. Knockdown of Ol-Ush2a in medaka fish caused embryonic developmental defects (small eyes and heads, otolith malformations and shortened bodies with curved tails) resulting in late embryo lethality. These embryonic defects, observed in our study and in other ciliary disorders, are associated with defective cell movement specifically implicated in left-right (LR) axis determination and planar cell polarity (PCP).

Metabolomic changes in the rat retina after optic nerve crush.

PURPOSE: To identify metabolic pathways and metabolites affected by optic nerve crush that can act as predictors of the disease or therapeutic targets. METHODS: The left optic nerve of adult rats was intraorbitally crushed and retinas were dissected 24 hours or 14 days after the lesion (n = 10 per group). Metabolic profiling analysis was carried out by Metabolon, Inc. A total of 195 metabolites were unambiguously detected. Data were normalized and the regulated metabolites were identified after comparing the different conditions. Metabolite concentration changes were analyzed using single and multivariate statistical analysis to detect discriminatory metabolites. Functional clustering and meta-analysis of the regulated metabolites was run through the Metacore platform. RESULTS: Comparison of 24 hours versus control, 14 days versus control samples, and 24 hours versus 14 days identified 9, 19, and 32 regulated metabolites, respectively. Single and multivariate analysis identified a total of 27 and 36 metabolites to discriminate between control and 14 days and between 24 hours and 14 days, respectively. Enrichment analysis showed alterations in the amino acid, carbohydrate, and lipid metabolism, which were further linked to translation, oxidative stress, energy (glucose and tricarboxylic acid cycle), and apoptosis through ceramide pathways. CONCLUSIONS: Our analysis differentiates a set of metabolites that clearly discriminate control and early-injury samples from late-injury samples. These metabolites could have potential use as diagnostic molecules.

Fishing pluripotency mechanisms in vivo.

To understand the molecular mechanisms that regulate the biology of embryonic stem cells (ESCs) it is necessary to study how they behave in vivo in their natural environment. It is particularly important to study the roles and interactions of the different proteins involved in pluripotency and to use this knowledge for therapeutic purposes. The recent description of key pluripotency factors like Oct4 and Nanog in non-mammalian species has introduced other animal models, such as chicken, Xenopus, zebrafish and medaka, to the study of pluripotency in vivo. These animal models complement the mouse model and have provided new insights into the evolution of Oct4 and Nanog and their different functions during embryonic development. Furthermore, other pluripotency factors previously identified in teleost fish such as Klf4, STAT3, Sox2, telomerase and Tcf3 can now be studied in the context of a functional pluripotency network. The many experimental advantages of fish will fuel rapid analysis of the roles of pluripotency factors in fish embryonic development and the identification of new molecules and mechanisms governing pluripotency.

Aqueous humor neutrophil gelatinase-associated lipocalin levels in patients with idiopathic acute anterior uveitis.

PURPOSE: The purpose of this study was to evaluate the levels of neutrophil gelatinase-associated lipocalin (NGAL) in the aqueous humor in eyes with idiopathic acute anterior uveitis (AAU). METHODS: A comparative control study. Aqueous humor was collected from 20 eyes of 20 patients with idiopathic AAU. The control group included 20 aqueous samples from 20 patients about to undergo cataract surgery and without any other ocular or systemic diseases. The level of NGAL was determined with a commercially available ELISA kit. RESULTS: The concentration of NGAL in aqueous humor was markedly higher in patients with idiopathic AAU than in control subjects (Mann-Whitney U test, p<0.001). The level of NGAL was 139,197.38+/-183,426.36 (mean+/-SD) pg/ml in eyes with AAU and 3,169.96+/-1,595.78 pg/ml in the eyes of the control group. CONCLUSIONS: The aqueous humor NGAL level is increased in eyes with idiopathic AAU. These results imply that NGAL is associated with the regulation of inflammation in patients with AAU and could be used as a biomarker of ocular inflammation and immunomodulatory treatment response.

Nanog regulates primordial germ cell migration through Cxcr4b.

Gonadal development in vertebrates depends on the early determination of primordial germ cells (PGCs) and their correct migration to the sites where the gonads develop. Several genes have been implicated in PGC specification and migration in vertebrates. Additionally, some of the genes associated with pluripotency, such as Oct4 and Nanog, are expressed in PGCs and gonads, suggesting a role for these genes in maintaining pluripotency of the germ lineage, which may be considered the only cell type that perpetually maintains stemness properties. Here, we report that medaka Nanog (Ol-Nanog) is expressed in the developing PGCs. Depletion of Ol-Nanog protein causes aberrant migration of PGCs and inhibits expression of Cxcr4b in PGCs, where it normally serves as the receptor of Sdf1a to guide PGC migration. Moreover, chromatin immunoprecipitation analysis demonstrates that Ol-Nanog protein binds to the promoter region of Cxcr4b, suggesting a direct regulation of Cxcr4b by Ol-Nanog. Simultaneous overexpression of Cxcr4b mRNA and depletion of Ol-Nanog protein in PGCs rescues the migration defective phenotype induced by a loss of Ol-Nanog, whereas overexpression of Sdf1a, the ligand for Cxcr4b, does not restore proper PGC migration. These results indicate that Ol-Nanog mediates PGC migration by regulating Cxcr4b expression.

Wnt signaling has different temporal roles during retinal development.

Differentiation of neural retinal precursor (NRP) cells in vertebrates follows an established order of cell-fate determination associated with exit from the cell cycle. Wnt signaling regulates cell cycle in colon carcinoma cells and has been implicated in different aspects of retinal development in various species. To better understand the biological roles of Wnt in the developing retina, we have used a transgenic and pharmacological approach to manipulate the Wnt signaling pathway during retinal development in medaka embryos. With the use of both approaches, we observed that during the early phase of retinal development Wnt signaling regulated cell cycle progression, proliferation, apoptosis, and differentiation of NRP cells. However, during later phases of retinal development, proliferation and apoptosis were not affected by manipulation of Wnt signaling. Instead, Wnt regulated Vsx1 expression, but not the expression of other retinal cell markers tested. Thus, the response of NRP cells to Wnt signaling is stage-dependent.

Medaka Oct4 is expressed during early embryo development, and in primordial germ cells and adult gonads.

Oct4 is a crucial transcription factor for controlling pluripotency in embryonic stem cells and the epiblast of mouse embryos. We have characterized the expression pattern of medaka (Oryzias latipes) Ol-Oct4 during embryonic development and in the adult gonads. Genomic analysis showed that Ol-Oct4 is the ortholog of zebrafish spg/pou2. However, their expression patterns are not the same, suggesting that Oct4 may play different roles in zebrafish and medaka. Using specific antibodies for the Ol-Oct4 protein, we showed that Ol-Oct4 is also expressed in primordial germ cells, in the spermatogonia (male germ stem cells), and during different stages of oocyte development. These results suggest that Ol-Oct4 plays a post-embryonic role in the maturing gonads and gametes. The Ol-Oct4 mRNA and protein expression patterns are similar to those of mammalian Oct4 and introduce medaka fish as a valid model for the functional and evolutionary study of pluripotency genes in vivo.

Nanog regulates proliferation during early fish development.

Nanog is involved in controlling pluripotency and differentiation of stem cells in vitro. However, its function in vivo has been studied only in mouse embryos and various reports suggest that Nanog may not be required for the regulation of differentiation. To better understand endogenous Nanog function, more animal models should be introduced to complement the murine model. Here, we have identified the homolog of the mammalian Nanog gene in teleost fish and describe the endogenous expression of Ol-Nanog mRNA and protein during medaka (Oryzias latipes) embryonic development and in the adult gonads. Using medaka fish as a vertebrate model to study Nanog function, we demonstrate that Ol-Nanog is necessary for S-phase transition and proliferation in the developing embryo. Moreover, inhibition or overexpression of Ol-Nanog does not affect gene expression of various pluripotency and differentiation markers, suggesting that this transcription factor may not play a direct role in embryonic germ layer differentiation. STEM CELLS 2009;27:2081-2091.

Loss-of-function mutations in the human GLI2 gene are associated with pituitary anomalies and holoprosencephaly-like features.

Diminished Sonic Hedgehog (Shh) signaling is associated with the most common forebrain defect in humans, holoprosencephaly (HPE), which includes cyclopia, a phenotype also seen in mice and other vertebrates with defective Shh signaling. The secreted protein Shh acts as a crucial factor that patterns the ventral forebrain and is required for the division of the primordial eye field and brain into two discrete halves. Gli2 is one of three vertebrate transcription factors implicated as obligatory mediators of Shh signal transduction. Here, we show that loss-of-function mutations in the human GLI2 gene are associated with a distinctive phenotype (within the HPE spectrum) whose primary features include defective anterior pituitary formation and pan-hypopituitarism, with or without overt forebrain cleavage abnormalities, and HPE-like midfacial hypoplasia. We also demonstrate that these mutations lack GLI2 activity. We report on a functional association between GLI2 and human disease and highlight the role of GLI2 in human head development.

Pathways and consequences: Hedgehog signaling in human disease.

Signaling pathways that play a fundamental role during development are turning out to underlie many disease states when misregulated. Here, we review some of the recent findings in the Hedgehog (Hh) pathway and the role it plays in different human diseases. We present a summary of the diseases that result from the inactivation or inappropriate activation of the Hh pathway. The human phenotypes generally fit the findings in model organisms and help to identify some potential targets for therapy.

Growth, hedgehog and the price of GAS.

Embryonic development in a given species is orchestrated by genes regulating growth and differentiation in a stereotyped and conserved manner, resulting in embryos of consistent size and shape. Several signaling pathways, including that of Sonic Hedgehog (SHH), have been implicated in these processes. Recent experiments with Gas1 indicate that it may act as a growth-inducing gene, challenging its previous function as a gene specifically involved in growth arrest. Moreover, GAS1, a GPI-linked membrane protein, can bind SHH, suggesting an interacting link between growth and patterning through SHH and GAS1.