Brain Development in Prolidase Deficient Mice

Prolidase deficiency (PD) is a rare autosomal recessive disorder caused by mutations in the prolidase gene, the PEPD, causing the reduction or the loss of the prolidase enzyme activity. PD patients present a variable onset, and severe skin ulcers mainly characterize the pathology. However, PD has a broad spectrum of phenotypes including mental impairment and developmental delay of variable degrees. Prolidase is a member of the matrix metalloproteinase (MMP) family. MMPs together with their inhibitors (tissue inhibitor of metalloproteinases, TIMPs) regulate the extracellular matrix maturation and remodelling life-long. Among them, prolidase is able to cleave dipeptides when prolin or hydroxyprolin residues are located at the C-terminal end. According to prolidase activity, its function has an impact on the metabolism of many biologically important molecules, particularly during the biosynthesis and degradation of collagen and procollagen. Therefore, prolidase indirectly has a role in the ECM remodelling. In particular, the ECM adjustments are essential in the brain, especially during the critical period of development: from passive structural property, to a direct influence on cell proliferation, migration, axonal guidance, synaptogenesis, homeostatic plasticity, learning and memory processes, and angiogenesis. In particular, the basement membrane beside the pial meninx (pBM) is a specialized structure of ECM whose integrity and proper assembly is essential for a correct cortical development and the collagen IV plays an essential role in pBM stability. Ruptures, even localized, in the pBM are accompanied by changes in the morphology of radial glia cells, subsequent cortical dysplasia, overmigration of neurons, decrease in the proliferation and migration of granule cell precursors, and reduction in Purkinje neuron dendrites. Recently, a mutant mouse with reduced prolidase activity has been identified with a spontaneous 4 bp deletion in the exon 14 of Pepd gene. The mutant mouse was named dark-like (dal) because of its darkened-coat color in homozygosis. The dal/dal phenotype includes small body size, reproductive degeneration, vacuolated cells at the cortical medullary junction of the adrenal gland, mild hydrocephalus, dark urine and altered bone phenotype. The prolidase activity was strongly reduced in cerebrum and cerebellum in dal mice. Moreover, they develop hypertrophic cardiomyopathy, but neither skin lesions nor recurrent infections were reported (in contrast to the reported human cases). The aim of this thesis was the study the brain development of dal/+ and dal/dal mice. Since no information were available, the analysis started with a morphological evaluation of the cerebellum, neocortex and hippocampus, through histological stainings. Then, immunohistochemistry reactions and western blotting analysis helped the anomalies characterizations. In particular, the attention has been mainly focused on the cerebellum, since it is the structure in which the ontogenetic events occurred also postnatally. The neocortex and hippocampal results were not described in details. Our results suggested that the absence of a full functional prolidase enzyme in the dal/dal mice results in a damage of the integrity of the pBM with an altered collagen, laminin and reelin profile. Such damage, could affect as a cascade of developmental events the proper lamination process of the cerebellum, leading to a cortical dysplasia together with the presence of degenerating and ectopic cells, defects in cerebellar lobulation and in the excitation/inhibition pattern of the cerebellar circuit

Antioxidant Efficacy of a Standardized Red Orange (<i>Citrus sinensis</i> (L.) Osbeck) Extract in Elderly Subjects: A Randomized, Double Blind, Controlled Study

The world population is rapidly aging. This should cause us to reflect on the need to develop a new nutritional approach to mitigate the accumulation of reactive oxygen species (ROS)-induced damage. A randomized, double blind, controlled study was carried out on 60 elderly male and female subjects. Product efficacy was measured before and after 2 and 8 weeks of product intake. The reduced (GSH) and oxidized (GSSG) glutathione concentrations in the erythrocytes and the reactive oxygen metabolites (d-ROMs) hematic concentration were measured to assess the antioxidant efficacy. The tumor necrosis factor-alpha (TNF-α) levels in the serum were measured to assess the anti-inflammatory effectiveness. The wellbeing was assessed by Short Form Health Survey (SF-36) questionnaire (male) and by Menopause Rating Scale (MRS) (female). Blood, urine analysis and electrocardiography (ECG) were carried out to assess the product’s safety. The results showed that GSH/GSSG ratio increased by 22.4% and 89.0% after 2 and 8 weeks of product intake. Serum TNF-α levels decreased by 2.5% after 8 weeks of product intake. The SF-36 QoL and the MRS questionnaire outputs indicate, preliminarily, a positive effect of the extract intake in ameliorating the wellbeing of both male and female subjects. The product was well-tolerated. Our findings suggest that the test product has antioxidant and anti-inflammatory efficacy and has a positive effect on the wellbeing of elderly female and male subjects

Photoprotective and Antiaging Effects of a Standardized Red Orange (<i>Citrus sinensis</i> (L.) Osbeck) Extract in Asian and Caucasian Subjects: A Randomized, Double-Blind, Controlled Study

The increase in solar ultraviolet radiation (UVR) that reaches the Earth’s surface should make us reflect on the need to develop new approaches in protecting the skin from UVR exposure. The present study aims to evaluate the photoprotective and antiaging efficacy of a red orange extract (100 mg/day) in both Asian and Caucasian subjects. A randomized, double-blind, controlled study was carried out in 110 Asian and Caucasian subjects. Product efficacy was measured as follows: (1) the photoprotective effect was measured by the minimal erythema dose (MED) assessment; (2) the efficacy in decreasing the UVA+B-induced skin redness was measured by colorimetry; (3) the antioxidant efficacy was measured by the ferric-reducing antioxidant power (FRAP) and the malondialdehyde (MDA) assay; and (4) skin moisturization, skin elasticity, skin radiance, the intensity of melanin staining, transepidermal water loss (TEWL), and wrinkles were measured to assess the antiaging efficacy. The intake of the product for 56 days was effective in improving the skin reaction to UV exposure; in increasing the skin antioxidant capacity as well as in decreasing UVA-induced lipid peroxidation; in increasing the skin moisturization, skin elasticity, and skin radiance; and in decreasing TEWL, the intensity of melanin staining inside dark spots, and wrinkle depth. Our results suggest that the test product is effective in counteracting both the harmful effects of UVR exposure and aging signs

Cerebellum neurotransmission during postnatal development: [Pt(O,O'-acac)(γ-acac)(DMS)] vs cisplatin and neurotoxicity

Abstract Several chemotherapeutic drugs are known to cause neurotoxicity. Platinum-based agents in use or in clinical trials display neurotoxic potential accompanied by neurological complications; recent studies have identified a large number of behavioural issues in paediatric oncology patients. To understand the toxicity of platinum drugs at the molecular and cellular levels, this study compares the possible cytotoxic effects of an older platinum compound, cisplatin and a new platinum compound, [Pt(O,O′-acac)(γ-acac)(DMS)], on the \{CNS\} of postnatally developing rats, which is much more vulnerable to injury than the \{CNS\} of adult rats. Since several drugs interact with neurotransmitters during neuronal maturation, we performed immunostainings with antibodies raised against markers of glutamate and GABA, the major neurotransmitters in the cerebellum. After a single injection of cisplatin at postnatal day 10 (PD10), the labelling of Purkinje cells with the neurotransmitter markers evidenced alterations between \{PD11\} and PD30, i.e. atrophy of the dendrite tree, changes in the distribution of synaptic contacts of parallel and climbing fibres, delay in the elimination of transient synapses on cell soma and severely impaired pinceau formation at the axon hillock. After treatment with [Pt(O,O′-acac)(γ-acac)(DMS)], the sole relevant change concerned the timing of climbing fibres elimination; the transient synapses disappearance on the Purkinje cell soma was delayed in some cells; instead, the growth of Purkinje cell dendrite tree was normal as was the formation of inhibitory synaptic contacts on these neurons. These findings add new evidence not only on the lower neurotoxicity of [Pt(O,O′-acac)(γ-acac)(DMS)] vs cisplatin but also on the involvement of neurotransmitters and relative synaptic connections in the maturation of central nerve tissue

Metabolic profiling, in vitro bioaccessibility and in vivo bioavailability of a commercial bioactive Epilobium angustifolium L. extract

Abstract The global diffusion of benign prostatic hyperplasia (BPH) demands the search for safe and effective treatment alternatives to the drugs commonly used, which exert both side and adverse effects. Among plant-based products, the extracts of Epilobium angustifolium L. (EAEs) could improve BPH symptoms thanks to the presence of ellagitannins and their anti-inflammatory metabolites, urolithins. This study focused its attention on a commercial EAE, standardized to contain ≥ 15 % oenothein B, to determine a) the metabolic profile and the chemical degradation induced by digestion, b) in vivo bioavailability after acute and prolonged treatments of CD1 mice, and c) in vitro antioxidant activity. Utilizing RP-HPLC-PDA-ESI-MSn analysis, 20 different compounds were identified. Polyphenols suffered from degradation after both orogastric and duodenal digestion processes, suggesting that gastro-resistant coating agents are required to preserve the bioactive components occurring in the EAE phytocomplex from orogastric digestion. In vivo data underlined the presence of urolithins only after the prolonged treatment, confirming that the gut fermentation process requires at least 24 h to produce urolithins. Finally, an increase of Superoxide Dismutase-1 (SOD-1), which represents one of the fundamental endogenous antioxidant defenses, was determined in an EAE pretreated LNCap cell model system, confirming EAE antioxidant activity

A Randomized, Double-Blind, Placebo-Controlled Trial: Efficacy of <i>Opuntia ficus</i>-<i>indica</i> Prebiotic Supplementation in Subjects with Gut Dysbiosis

Gut dysbiosis refers to an imbalance in gut microbiota composition and function. Opuntia ficus-indica extract has been shown to modulate gut microbiota by improving SCFA production in vivo and gastrointestinal discomfort (GD) in humans. The aim of this study was to demonstrate the efficacy of OdiliaTM on gastrointestinal health by changing the microbial diversity of species involved in inflammation, immunity, oxidation, and the brain–gut–muscle axis. A randomized, double-blind clinical trial was conducted in 80 adults with gut dysbiosis. The intervention consisted of a 300 mg daily intake of OdiliaTM (n = 40) or maltodextrin as a placebo (n = 40), administered for 8 weeks. Intervention effect was evaluated using 16S metagenomics and GIQLI/GSAS scores at baseline, at 4 and 8 weeks. Eight weeks of OdiliaTM supplementation positively modulates gut microbiota composition with a significant reduction in the Firmicutes to Bacteroidetes ratio (p = 0.0012). Relative abundances of beneficial bacteria (Bacteroides and Clostridium_XIVa) were significantly increased (p p TM may represent an effective and well-tolerated treatment in subjects with gut dysbiosis

[Pt(O,O'-acac)(γ-acac)(DMS)] versus cisplatin: apoptotic effects in B50 neuroblastoma cells.

Cisplatin is one of the most active chemotherapeutic agents used in the treatment of childhood and adult malignancies. Cisplatin induces cell death through different pathways. Despite its effectiveness, the continued clinical use of cisplatin is limited by onset of severe side effects (nephrotoxicity, ototoxicity and neurotoxicity) and drug resistance. Therefore, one of the main experimental oncology purpose is related to the search for new platinum-based drugs to create different types of adducts or more specific and effective subcellular targets. Thus, [Pt(O,O'-acac)(γ-acac)(DMS)], which reacts preferentially with protein thiols or thioether, was synthesized. In our research, different approaches were used to compare cisplatin and [Pt(O,O'-acac)(γ-acac)(DMS)] effects in B50 rat neuroblastoma cells. Our results, using immunocytochemical, cytometric and morphological techniques, showed that these compounds exert a cytostatic action and activate apoptosis with different pathways. Long-term effects demonstrated that [Pt(O,O'-acac)(γ-acac)(DMS)] exerts cytotoxic effects in neuronal B50 cell line not inducing drug resistance. Analysis was performed both to compare the ability of these platinum compounds to induce cell death and to investigate the intracellular mechanisms at the basis of their cytotoxicity

[Pt(O,O'-acac)(γ-acac)(DMS)] versus cisplatin: apoptotic effects in B50 neuroblastoma cells

Cisplatin is one of the most active chemotherapeutic agents used in the treatment of childhood and adult malignancies. Cisplatin induces cell death through different pathways. Despite its effectiveness, the continued clinical use of cisplatin is limited by onset of severe side effects (nephrotoxicity, ototoxicity and neurotoxicity) and drug resistance. Therefore, one of the main experimental oncology purpose is related to the search for new platinum-based drugs to create different types of adducts or more specific and effective subcellular targets. Thus, [Pt(O,O'-acac)(γ-acac)(DMS)], which reacts preferentially with protein thiols or thioether, was synthesized. In our research, different approaches were used to compare cisplatin and [Pt(O,O'-acac)(γ-acac)(DMS)] effects in B50 rat neuroblastoma cells. Our results, using immunocytochemical, cytometric and morphological techniques, showed that these compounds exert a cytostatic action and activate apoptosis with different pathways. Long-term effects demonstrated that [Pt(O,O'-acac)(γ-acac)(DMS)] exerts cytotoxic effects in neuronal B50 cell line not inducing drug resistance. Analysis was performed both to compare the ability of these platinum compounds to induce cell death and to investigate the intracellular mechanisms at the basis of their cytotoxicity