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In this study, age-related changes in the monoamine oxidases ( MAO) were studied in the optic nerve ( ON) of both young and aged male rats. The aim of the study was to assess the role of MAO in age-related changes in the rat ON and explain the mechanisms of neuroprotection mediated by MAO-B-specific inhibitors. Fifteen three month old and fifteen 26 month old Sprague-Dawley rats were used. The animals were killed by terminal anaesthesia. Staining of MAO, quantitative analysis of images, biochemical assays and statistical analysis of data were carried out. Samples of the ON were washed in water, fixed in Bowen fluid, dehydrated and embedded in Entellan. Histological sections were stained for MAO-enzymatic activities. The specificity of the reaction was evaluated by incubating control sections in a medium either without substrate or without dye. The quantitative analysis of images was carried out at the same magnification and the same lighting using a Zeiss photomicroscope. The histochemical findings were compared with the biochemical results. After enzymatic staining, MAO could be demonstrated in the ON fibres of both young and aged animals; however, MAO were increased in the nerve fibres of the elderly rats. These morphological findings were confirmed biochemically. The possibility that age-related changes in MAO levels may be attributed to impaired energy production mechanisms and/or represent the consequence of reduced energy needs is discussed. [ABSTRACT FROM AUTHOR]

Di-(2-ethylhexyl)-phthalate (DEHP), employed in polyvinyl chloride fabrication and released by endotracheal tubes, is known to cause alterations to several mammalian tissues, markedly in immature animals. The high incidence and severity of bronchopulmonary dysplasia and retinopathy in preterm babies submitted to endotracheal intubation prompted us to investigate the effects of DEHP in lung and retina perinatal development. We previously demonstrated that in rats delivered and breast-fed by DEHP-treated mothers lung alveolarisation is severely impaired. In the present research, the maturation of retinal vessels was studied in (a) flat-mounted retinas obtained after intracardiac injection of FITC-conjugated dextran, (b) flat-mounted retinas incubated with FITC-conjugated Bandeira simplicifolia isolectin B4, marker of vascular endothelial cells, and (c) eyecup sections incubated with biotinylated IB4 and revealed by ABC. DEHP-induced vascular alterations mainly affected the superficial plexus, where the radial vessels showed non-perfused as well as remarkably dilated and branched segments, capillary net appeared coarsely arranged and locally absent; periarteriolar capillary-free regions were still found in 14-day-old animals. This extensive vascular remodelling and generally the high responsiveness to DEHP shown by the immature rat retina confirm previous hypothesis that the phthalate released by PVC medical devices remarkably affects perinatal development of several tissues in different body districts. [ABSTRACT FROM AUTHOR]

Many studies have shown that the presence of 1,25-dihydroxyvitamin D3 in the eye is able to modulate inflammatory responses. In fact, it has been demonstrated that topical administration of vitamin D3 inhibits Langerhans cells migration from the central cornea, corneal neovascularization, and production of cytokines (i.e., interleukin-1-6-8) in experimental animals. Moreover, both in vitro and in vivo studies have demonstrated that vitamin D is a potent inhibitor of retinal neovascularization. It has been shown that calcitriol, the biologically active form of vitamin D, inhibits angiogenesis both in cultured endothelial cells and in retinas from guinea pigs with retinoblastoma or oxygen-induced ischemic retinopathy. In addition, it seems that this compound is able to prevent the progression from early to neovascular age-related macular degeneration (AMD) and, at the same time, to down-regulate the characteristic inflammatory cascade at the retinal pigment epithelium–choroid interface due to its anti-inflammatory and immunomodulatory capabilities. Furthermore, 1,25-dihydroxyvitamin D3 and its analogue, 2-methylene-19-nor-1,25-dihydroxyvitamin D3, are able to modulate intraocular pressure (IOP) through gene expression. Several studies have suggested a role in glaucoma and diabetic retinopathy therapies for vitamin D3. In conclusion, this review summarizes our current knowledge on the potential use of vitamin D3 in the protection and treatment of ocular diseases in ophthalmology. [ABSTRACT FROM AUTHOR]

It has been demonstrated that the balance between proteases and protease-inhibitors system plays a key role in maintaining cellular and tissue homeostasis. Indeed, its alteration has been involved in many ocular and systemic diseases. In particular, research has focused on keratoconus, corneal wounds and ulcers, keratitis, endophthalmitis, age-related macular degeneration, Sorsby fundus dystrophy, loss of nerve cells and photoreceptors during optic neuritis both in vivo and in vitro models. Protease-inhibitors have been extensively studied, rather than proteases, because they may represent a therapeutic approach for some ocular diseases. The protease-inhibitors mainly involved in the onset of the above-mentioned ocular pathologies are: α2-macroglobulin, α1-proteinase inhibitor (α1-PI), metalloproteinase inhibitor (TIMP), maspin, SERPINA3K, SERPINB13, secretory leukocyte protease inhibitor (SLPI), and calpeptin. This review is focused on the several characteristics of dysregulation of this system and, particularly, on a possible role of proteases and protease-inhibitors in molecular remodeling that may lead to some ocular diseases. Recently, researchers have even hypothesized a possible therapeutic effect of the protease-inhibitors in the treatment of injured eye in animal models. [ABSTRACT FROM AUTHOR]

Resveratrol (RV) differentially affects UV-induced death/pro-survival pathways in normal and tumor cells. On these grounds, RV-containing topical products have been developed to prevent UV-associated tumorigenesis/damage to human skin. In this study, we evaluated mechanisms of combined effects of RV and low-dose solar simulated UVA+UVB or 6-formylindo[3,2-b]carbazole (FICZ), a product of tryptophan photo-oxidation known to mediate UV effects, on the inflammatory, metabolic and proliferative responses of cultured normal human epidermal keratinocytes (HEK). Applied alone, RV, UV and FICZ induced time- and dose-dependent activation of aryl hydrocarbon receptor (AhR) pathway followed by over-expression of Cyp1A1 (metabolic response), UV and RV induced IL-8 expression (inflammatory response), while RV enhanced also HEK proliferation revealed by MTT assay and 3H-thymidine incorporation. In the combined treatment, RV synergized with both UV and FICZ, leading to further activation of AhR machine, Cyp1A1 transcription and IL-8 expression, the latter partly AhR-dependent as assessed by AhR silencing. RV enhanced UV-induced NFkappaB activation and nuclear translocation of epidermal growth factor receptor. By contrast, proliferative effect of RV was abolished in the presence of UV, whereas synergic anti-proliferative action of RV+UV was observed in the Nrf2-silenced HEK. Our data suggest cooperative effects of RV-specific and UV-/FICZ-activated transcription factors leading to deregulated inflammatory, metabolic and proliferative responses of HEK. [ABSTRACT FROM AUTHOR]

Background: Classic infantile onset of Pompe disease (c-IOPD) leads to hypotonia and hypertrophic cardiomyopathy within the first days to weeks of life and, without treatment, patients die of cardiorespiratory failure in their first 1–2 years of life. Enzymatic replacement therapy (ERT) with alglucosidase alfa is the only available treatment, but adverse immune reactions can reduce ERT's effectiveness and safety. It is therefore very important to identify strategies to prevent and manage these complications. Several articles have been written on this disease over the last 10 years, but no univocal indications have been established. Methods: Our study presents a review of the current literature on management of immune responses to ERT in c-IOPD as considered by an Italian study group of pediatric metabolists and immunologists in light of our shared patient experience. Results: We summarize the protocols for the management of adverse reactions to ERT, analyzing their advantages and disadvantages, and provide expert recommendations for their optimal management, to the best of current knowledge. However, further studies are needed to improve actual management protocols, which still have several limitations. [ABSTRACT FROM AUTHOR]

Aim of this study was to investigate the activation of the IFNγ pathway in the affected liver and in the blood of patients with secondary hemophagocytic lymphohistiocytosis (sHLH). To this purpose, the mRNA expression levels of IFNG and IFNγ-inducible genes as well as Tyrosine (701)-phosphorylated signal transducer and activator of transcription 1 (STAT1) protein levels were evaluated in the liver and in peripheral blood mononuclear cells (PBMCs) of three patients with sHLH with predominant liver involvement. The mRNA expression levels of IFNG and IFNγ-inducible genes were markedly higher in patient livers compared to control livers and to one disease control liver. Conversely, slight differences in the expression levels of Type I IFN-inducible genes and other classical inflammatory cytokine genes were found. Further supporting the activation of the IFNγ pathway, higher protein levels of phosphorylated and total STAT1 were detected in patient livers compared to control livers. When the expression of the same genes analysed in liver tissues was evaluated in PBMCs collected from 2 out of 3 patients before the liver biopsy, we found that mRNA levels of IFNγ-inducible genes were markedly increased. Accordingly, high circulating levels of IFNγ-inducible CXCL9 were observed in patients. Altogether, these data demonstrate the selective and marked up-regulation of the IFNγ pathway in the liver tissue and blood of patients with active sHLH. Finally, we show that measurement of circulating CXCL9 levels and evaluation of IFNγ–inducible gene expression levels in PBMCs may represent a new valid tool to better identify patients with suspected HLH with predominant liver involvement. [ABSTRACT FROM AUTHOR]

Background: Lysosphingolipids, the N-deacylated forms of sphingolipids, have been identified as potential biomarkers of several sphingolipidoses, such as Gaucher, Fabry, Krabbe and Niemann-Pick diseases and in GM1 and GM2 gangliosidoses. To date, different methods have been developed to measure various lysosphingolipids (LysoSLs) in plasma. Here, we present a novel liquid chromatography tandem mass spectrometry (LC-MS/MS) assay for a simultaneous quantification of LysoSLs (HexSph, LysoGb3, LysoGM1, LysoGM2, LysoSM and LysoSM509) in dried blood spot (DBS). This LC-MS/MS method was used to compare the levels of LysoSLs in DBS and plasma in both affected patients and healthy controls. Methods: Lysosphingolipids were extracted from a 3.2 mm diameter DBS with a mixture of methanol:acetonitrile:water (80:15:5, v/v) containing internal stable isotope standards. Chromatographic separation was performed using a C18 column with a gradient of water and acetonitrile both with 0.1% formic acid in a total run time of 4 min. The compounds were detected in the positive ion mode electrospray ionization (ESI)-MS/MS by multiple reaction monitoring (MRM). Results: The method was validated on DBS to demonstrate specificity, linearity, lowest limit of quantification, accuracy and precision. The reference ranges were determined in pediatric and adult populations. The elevated levels of LysoSLs were identified in Gaucher disease (HexSph), Fabry disease (LysoGb3), prosaposin deficiency (HexSph and LysoGb3) and Niemann-Pick disease types A/B and C (LysoSM and LysoSM509). The correlation in the levels between DBS and plasma was excellent for LysoGb3 and HexSph but poor for LysoSM and LysoSM509. Conclusions: Despite the fact that plasma LysoSLs determination remains the gold standard, our LC-MS/MS method allows a rapid and reliable quantification of lysosphingolipids in DBS. The method is a useful tool for the diagnosis of different sphingolipidoses except for Niemann-Pick type C. [ABSTRACT FROM AUTHOR]

Background Hunter Syndrome is an X-linked lysosomal storage disorder due to the deficit of iduronate 2- sulfatase, an enzyme catalysing the degradation of the glycosaminoglycans (GAG) dermatanand heparan-sulfate. Treatment of the disease is mainly performed by Enzyme Replacement Therapy (ERT) with idursulfase, in use since 2006. Clinical efficacy of ERT has been monitored mainly by the Hunter Outcome Survey (HOS) while very few independent studies have been so far conducted. The present study is a 3.5-years independent follow-up of 27 Hunter patients starting ERT between 1.6 and 27 years of age, with the primary aim to evaluate efficacy of the therapy started at an early age (<12 years). Methods In this study, we evaluated: urinary GAG content, hepato/splenomegaly, heart valvulopathies, otorinolaryngological symptoms, joint range of motion, growth, distance covered in the 6 minute-walk test, neurological involvement. For data analysis, the 27 patients were divided into three groups according to the age at start of ERT: ⩽5 years, >5 and ⩽ 12 years and > 12 years. Patients were analysed both as 3 separate groups and also as one group; in addition, the 20 patients who started ERT up to 12 years of age were analysed as one group. Finally, patients presenting a "severe" phenotype were compared with "attenuated" ones. Results Data analysis revealed a statistically significant reduction of the urinary GAG in patients ⩽5 years and ⩽ 12 years and of the hepatomegaly in the group aged >5 and ⩽ 12 years. Although other clinical signs improved in some of the patients monitored, statistical analysis of their variation did not reveal any significant changes following enzyme administration. The evaluation of ERT efficacy in relation to the severity of the disease evidenced slightly higher improvements as for hepatomegaly, splenomegaly, otological disorders and adenotonsillar hypertrophy in severe vs attenuated patients. Conclusions Although the present protocol of idursulfase administration may result efficacious in delaying the MPS II somatic disease progression at some extent, in this study we observed that several signs and symptoms did not improve during the therapy. Therefore, a strict monitoring of the efficacy obtained in the patients under ERT is becoming mandatory for clinical, ethical and economic reasons. [ABSTRACT FROM AUTHOR]

Background: Hunter Syndrome is an X-linked lysosomal storage disorder due to the deficit of iduronate 2-sulfatase, an enzyme catalysing the degradation of the glycosaminoglycans (GAG) dermatan- and heparan-sulfate. Treatment of the disease is mainly performed by Enzyme Replacement Therapy (ERT) with idursulfase, in use since 2006. Clinical efficacy of ERT has been monitored mainly by the Hunter Outcome Survey (HOS) while very few independent studies have been so far conducted. The present study is a 3.5-years independent follow-up of 27 Hunter patients, starting ERT between 1.6 and 27 years of age, with the primary aim to evaluate efficacy of the therapy started at an early age (<12 years).Methods: In this study, we evaluated: urinary GAG content, hepato/splenomegaly, heart valvulopathies, otorinolaryngological symptoms, joint range of motion, growth, distance covered in the 6-minute walk test, neurological involvement. For data analysis, the 27 patients were divided into three groups according to the age at start of ERT: ≤5 years, >5 and ≤ 12 years and > 12 years. Patients were analysed both as 3 separate groups and also as one group; in addition, the 20 patients who started ERT up to 12 years of age were analysed as one group. Finally, patients presenting a "severe" phenotype were compared with "attenuated" ones.Results: Data analysis revealed a statistically significant reduction of the urinary GAG in patients ≤5 years and ≤ 12 years and of the hepatomegaly in the group aged >5 and ≤ 12 years. Although other clinical signs improved in some of the patients monitored, statistical analysis of their variation did not reveal any significant changes following enzyme administration. The evaluation of ERT efficacy in relation to the severity of the disease evidenced slightly higher improvements as for hepatomegaly, splenomegaly, otological disorders and adenotonsillar hypertrophy in severe vs attenuated patients.Conclusions: Although the present protocol of idursulfase administration may result efficacious in delaying the MPS II somatic disease progression at some extent, in this study we observed that several signs and symptoms did not improve during the therapy. Therefore, a strict monitoring of the efficacy obtained in the patients under ERT is becoming mandatory for clinical, ethical and economic reasons. [ABSTRACT FROM AUTHOR]