Background BCG (Bacillus Calmette-Guerin), as reported by international guidelines, represents a first choice adjuvant immunotherapy for intracavitary treatment after transurethral resection of the bladder (TURB) for non muscle-invasive tumours, Ta-T1 and carcinoma in situ (CIS) at high risk of progression. When possible these patients should be treated for 6 weeks, followed by maintenance treatment for at least 1 year. If BCG is unavailable, the AIFA (Italian Medicines Agency), in several notes, although highlighting the risks and the possible disadvantages for patients, has recommended ensuring that all patients have induction treatment for 6 weeks, limiting the ongoing maintenance regime and starting a high surveillance of patients. Among the other therapeutic options to consider, it suggests to clinicians the use of radical cystectomy replacing the conservative treatment, with considerable discomforts for patients. The efficacy of different BCG strains (CONNAUGHTS, TICE, MOSCOW) seems to be comparable, according to EBM. Purpose To evaluate the clinical and pharmacoeconomic impact on the treatment of patients with bladder cancer as result of an international lack of BCG, which lasted for several months. Materials and methods A retrospective analysis was conducted on BCG patients treated during the drug shortage (Jan 2012-Sep 2013), evaluating their therapeutic courses in the following months (treatment suspension, final interruptions, reduced treatments, shift among different bacillus strains); furthermore the economic impact due to the drug shortage and to the following purchase abroad was evaluated. Results Of 68 patients who were treated in the period under study only 52, who completed the treatment, are considered evaluable. These patients have undergone: 29 complete induction treatments and 3 complete maintenance treatments; 22 complete treatments with shift among different drug strains (2 induction, 20 maintenance); 12 treatments with treatment interruption (5 induction, 7 maintenance with an average of 4 administered cycles). 16 patients are still in treatment (3 induction, 13 maintenance). Of 31 patients who completed the induction, under close monitoring, 19 (61.3%) resumed the treatment after an average interruption of 3.4 months after recurrence or progression. The average cost of a bladder instillation increased from 61.8 €, before the shortage (CONNOUGHT strain) to 152.3 € (+146%) (TICE, MOSCOW strains), after importing from abroad. Conclusions In high-risk patients, still considered suitable for conservative treatment or where that was not possible doing a radical cystectomy, induction and maintenance treatment should be the first choice, considering the high percentage of recurrence. Unfortunately the lack of drug has in many cases meant the interruption or temporary suspension of the treatment; this could cause long-lasting negative effects which will require further investigations to be confirmed. No conflict of interest.

Background Sodium dichloroacetate (Na-DCA), not a patented substance, which is used in the treatment of rare diseases with congenital defects of the pyruvate-dehydrogenase complex (PDHC), produces a marked reduction in acid-base imbalance and lactic acid levels toxic to the brain parenchyma. Purpose To evaluate the physical-chemical stability of sodium dichloroacetate in aqueous solution. Materials and Methods Six grammes of sodium dichloroacetate were dissolved in 60 ml of water for injections (WFI). The exact concentration of the solution obtained was calculated by extrapolation from a calibration curve, recording the absorbance value at the wavelength of 198 nm of suitable standard solutions (5–50 µg/ml) of sodium dichloroacetate dissolved in water for injections (WFI). The solution was divided between 3 dark glass containers. The first container was kept at room temperature (r.t.), the second one in a refrigerator at +4°C, the third one in a freezer at −20°C. The stability of the samples, kept at different temperatures, was checked at 31, 45, 54 and 60 days; for each sample, using appropriate dilution, absorbance values were recorded (λ = 198 nm) and through the sodium dichloroacetate calibration curve made daily, the concentrations of the substance being analysed were calculated. The results were expressed as percentages of sodium dichloroacetate in solution. Results Samples kept at +4°C were stable throughout the observation period. Samples kept at r.t. were stable until 30 days from preparation, while afterwards a slow and gradual decay could be observed. Samples kept at −20°C showed a progressive increase in concentration. Conclusions The observed increase in samples at −20°C can be explained by the formation of a secondary species with a higher extinction coefficient than sodium dichloroacetate. Data suggest that sodium dichloroacetate solutions should not be stored at −20°C or at r.t. for more than 30 days. No conflict of interest.

Recent concern for the possible effects of endocrine disrupting chemicals on humans and wildlife [1] has resulted in considerable interest in environmental contaminants, including pesticides, that affect aspects of reproduction and early development. Vinclozolin (VIN), a pesticide used to treat fungal infections in plant-derived foodstuffs, plants and grasses, causes abnormal male sexual development in rats [1]. As a result of its widespread use, the analysis of VIN on food samples has been incorporated into the US Food and Drug Administration Pesticide Residue Monitoring Program [2]. VIN can undergo chemical hydrolysis, photolysis or metabolic transformation by mammals and bacterial systems. Szeto et al. [2] isolated three hydrolysis products from aqueous buffers which were identified as 2-[[(3,5-dichlorophenyl)-carbamoyl]oxy]-2-methyl-3-butenoic acid (M1), 3'5'-dichloro-2-hydroxy-2-methylbut-3-enanilide (M2) and 3,5-dichloroaniline (M3). While specific metabolic pathways have been proposed for VIN, information on the in vivo disposition of VIN and its metabolites is limited, particularly in the target organs of mammalian species [3]. The two hydrolysis products of VIN, M1 and M2, are competitive antagonists of the androgen receptor (AR) [1]. AR antagonism by M1 and M2 can ultimately decrease androgen-dependent gene expression, resulting in adverse effects in developing, pubertal and adult male rats. Administration of VIN during the critical period of sexual differentiation resulted in sexual abnormalities expressed later in the adult male rat. Pregnant rats exposed to VIN at specific gestational stages had male offspring with a significant degree of morphological feminization and demasculinization [1]. The characterization of the role of VIN and its metabolites M1 and M2 in human cells, such as prostate cells, has received limited attention. This is due in part to analytical difficulties such as instability of VIN in aqueous media [4]. Prostate is a critical but neglected target in reproductive toxicology. Prostate function is critical for male fertility and its well-known oncological biomarker, the prostate-specific antigen (PSA), can be also used to monitor prostate epithelial human cells upon treatment with pharmaceutical drugs or natural bioactive compounds [5,6]. The objective of this study was to characterize the biological effects of VIN and its metabolites on prostate epithelium cells investigating i) their anti-androgenic ability to inhibit DHT-induced PSA secretion, and ii) their intracellular distribution, in presence or absence of sex steroid (DHT), To verify whether and to which extent VIN and its metabolites are able to enter the cell and to reach the nucleus, the target of their supposed transcriptional modulatory activity upon binding to sex steroid receptors. For this purpose we used the LNCaP human prostate cell line as a model system to investigate chemicals affecting prostate epithelium functionality by means of a tiered approach integrating two different toxicological endpoints, cell viability (MTS assay) and PSA secretion, using well established, widely used commercial assays. Data will be presented and discussed.

The bioaccessibility of nutrients and phytochemicals from almond skin has not been previously evaluated. We quantified the release of lipid, protein and polyphenols during simulated human digestion from natural (NS) and blanched (BS) skins, the latter being a by-product of the almond industry. Higher percentages of polyphenols were released from NS compared to BS during in vitro digestion. Most of the limited release of lipid and protein occurred during gastric digestion, with no significant differences between NS and BS. The total dietary fibre content was 45% for NS and 46% for BS, glucose and galacturonic acid being the major sugars present. No changes in dietary fibre composition and distribution of autofluorescent phenolics were observed in the cell walls of almond skin after simulated digestion. In the GI tract, the cell walls may therefore function as a useful source of fermentable fibre with beneficial implications for gut health.

Almond skins and blanch water are underutilized by-products of the almond processing industry. Nevertheless, they contain exploitable components that may contribute to the health benefits associated with almond consumption. We have compared natural almond skin powder (NS) prepared by a novel freeze-thawing method with blanched almond skin powder (BS). Microstructural studies were carried out, and we analyzed both types of almond skin for phenolic compounds (by HPLC), lipids (by solvent extraction), proteins (by micro-Kjeldahl), and fibre content (by the enzymatic-gravimetric AOAC method). Antioxidant activity (by measuring the reduction of the 2,2-diphenyl-1-picrylhydrazyl radical) was also monitored. We identified a combination of flavonols, flavan-3-ols, hydroxybenzoic acids and flavanones in NS, BS and in industrially obtained blanch water (BW). As expected, the total phenolic content was higher in NS compared to BW and BS, although the latter showed high antioxidant properties. Almond skins had high fibre content as well as significant amounts of lipid; both of these components may be relevant to fermentation in the large intestine. In addition, the processing of almond skins and blanch water clearly has economic potential for lowering the environmental impact of waste fill and pollution.

Aims:To evaluate the antimicrobial properties of flavonoid-rich fractions derived from natural and blanched almond skins, the latter being a by-product from the almond processing industry.Methods and Results:Almond skin extracts were tested against Gram-negative bacteria (Escherichia coli, Pseudomonas aeruginosa, Salmonella enterica, Serratia marcescens), Gram-positive bacteria (Listeria monocytogenes, Enterococcus hirae, Staphylococcus aureus, Enterococcus durans) and the yeast Candida albicans. Almond skin fractions were found to have antimicrobial activity against L. monocytogenes and Staph. aureus in the range 250-500 mu g ml-1, natural skins showing antimicrobial potential against the Gram-negative Salm. enterica. The interactions between three almond skin flavonoids were also evaluated with isobolograms.Conclusions:Pairwise combinations of protocatechuic acid, naringenin and epicatechin showed both synergistic and indifferent interactions against Salm. enterica and Staph. aureus. Antagonism was observed against L. monocytogenes with all combinations tested. Further studies need to be performed to understand the mechanisms responsible for these interactions.Significance and Impact of the Study:Almond skins are a potential source of natural antimicrobials.

It is known that oxidative stress is involved in the pathogenesis of multiple sclerosis (MS), in particular, protein carbonylation. The aim of this study was to evaluate whether cerebrospinal fluid (CSF) carbonylated proteins could be used as a marker of cerebral oxidative activity in MS patients. The serum concentrations of malondialdehyde/hydroxynonenal, thiol groups and protein carbonyl groups were evaluated, together with the CSF levels of thiol groups and protein carbonyl groups in 33 MS patients and 19 patients with other inflammatory neurological diseases. Findings suggest that CSF carbonylated proteins might be used as a marker of cerebral oxidative activity in MS patients.

Herpes simplex virus type 1 (HSV-1) and herpes simplex virus type 2 (HSV-2) are common human pathogens that in particular cases can also cause severe problems especially in immunodeficient patients. The present paper reports the antiviral and immunomodulatory properties of a methanolic extract of C. spinosa buds (CAP), rich in flavonoids, including several quercetin and kaempferol glycosides. In particular we have investigated whether the in vitro exposure of human peripheral blood mononuclear cells (PBMCs) to CAP might inhibit the replication of HSV-2 and modulate the induction kinetics of IL-12, TNF-alpha IFN-gamma. Our findings have shown that CAP treatment interferes with HSV-2 replication in PBMCs inhibiting the extracellular virus release upregulating their production of IL-12, IFN-gamma and TNF-alpha. One could speculate that CAP may contribute in improving immune surveillance of PBMCs toward virus infection by up-regulating expression of peculiar proinflammatory cytokines; it should thus be successfully employed for treatment of HSV-2 infections in immunocompromised hosts.

Several studies have shown that UV radiation on the skin results in the formation of reactive oxygen species (ROS) that interact with proteins, lipids and DNA, thus altering cellular functions. The epidermis is composed mainly of keratinocytes, rich in ROS detoxifying enzymes and in low-molecular-mass antioxidant molecules. However, the increased generation of ROS can overwhelm the natural defences against oxidative stress. Therefore treatment of the skin with products containing plant-derived antioxidant ingredients may be a useful strategy for the prevention of UV-mediated cutaneous damage. In the present study we have investigated the in vitro capability of a Jacquez grapes wine extract (containing a significant level of proanthocyanidins, together with lower amounts of anthocyanins and hydroxycinnamic acids; JW-E), to protect skin against UVB-induced oxidative damage by using a three-dimensional tissue culture model of human epidermis. The endpoints of our experiments were cell viability, release of interleukin-1alpha and prostaglandin E(2) (well-known mediators of cutaneous inflammatory processes), accumulation in the epidermis of malondialdehyde/4-hydroxynonenal and protein carbonyl groups (derived by the oxidative damage respectively of lipids and proteins) and tissue redox balance (expressed by the levels of reduced glutathione, oxidized glutathione, glutathione peroxidase and glutathione reductase). Taken together, our findings demonstrate that the JW-E is an efficient botanical mixture able to prevent skin oxidative damage induced by UV-B exposure and may thus be a potential promising candidate as a skin photoprotective agent.

The present study was designed to verify the antidiarrhoeal effects of a lyophilized extract of wine from Jacquez grapes (Ord. Rhamnales; Fam. Vitaceae; Sp. Vitis aestivalis M.-cinerea E. x Vitis vinifera L.), studying its influence on castor oil-induced diarrhoea and enteropooling, and on gastrointestinal transit (measured by a charcoal marker) in mice. The pre-treatment of the animals with the JWE (Jacquez wine extract) produced a significant inhibition against castor oil induced-diarrhoea and intestinal fluid accumulation; furthermore the extract significantly decreased the propulsive movement of the charcoal meal. These findings suggest a potential beneficial use of the JWE in the treatment of diarrhoeal diseases.

A simple synthetic methodology, based on the inverse electron demand hetero Diels-Alder reaction of electron-poor dienic o-thioquinones with electron-rich styrenes used as dienophiles, allowed the preparation of several polyhydroxylated 4-thiaflavans. Such compounds, as a function of the nature and position of the substituents on the aromatic rings, as well as of the oxidation state of the sulfur atom, are able to behave in vitro as efficient antioxidants mimicking the action of catechol containing flavonoids or/and tocopherols. The possibility of joining together the potentialities of two relevant families of natural polyphenolic antioxidants appears particularly appealing since an efficient protection against free radicals and other reactive oxygen species (ROS) depends in vivo upon the synergic action of different antioxidant derivatives.

Administration of granulocyte colony-stimulating factor (G-CSF) is widely used for harvesting an adequate number of CD34+ stem cells by leukapheresis in normal donors. G-CSF is the most established agent for the mobilization of stem cells in current clinical practice, because it has been proven to be superior to any other agent tested to date in terms of not only mobilization capacity, but also of tolerance. However, although regulatory and accrediting agencies have provided guidelines to protect donors, the short- and long-term side effects of G-CSF need to be further studied. In this study, we evaluated the time course of plasma myeloperoxidase (MPO) levels measured in a group of donors given recombinant human G-CSF (rHuG-CSF) at different intervals: (i) before starting rHuG-CSF administration, (ii) on day 5 of rHuG-CSF administration, (iii) on the same day soon after the end of the first leukapheresis procedure and (iv) 1 week after rHuG-CSF withdrawal. Plasma MPO levels significantly increased in the donors after 5 days of rHuG-CSF treatment, returning to the baseline values within 7 days following rHuG-CSF withdrawal. These findings may contribute to a better understanding of G-CSF safety profile in stem cell donors.

Granulocyte-colony stimulating factor (G-CSF), the most established agent for the mobilization of stem cells in current clinical practice, could induce a condition of oxidative stress. Herein plasmatic levels of protein carbonyl groups (a biomarker of oxidative stress) were measured in a group of donors given recombinant human G-CSF (rHuG-CSF) at different times: (1) before starting rHuG-CSF administration, (2) on day 5 of rHuG-CSF administration, (3) on the same day immediately after the end of the first leukapheresis procedure and (4) one week after rHuG-CSF withdrawal. Plasma levels of protein carbonyl groups enhanced significantly in donors after 5 days of rHuG-CSF treatment and appeared further slightly increased following leukapheresis procedure; 7 days following rHuG-CSF withdrawal, they showed a tendency to normalization. These findings may contribute to better understand the oxidative reactions following rHuG-CSF treatment and leukapheresis.

A dysregulation of the redox homoeostasis has been reported in various neoplastic disorders. Malondialdehyde/4-hydroxy-2,3-nonenal (MDA/HNE) and protein carbonyl groups represent in vivo indexes of lipid peroxidation and protein oxidation, respectively, suitable to investigate radical-mediated physio-pathological conditions. We evaluated MDA/HNE and protein carbonyl groups in sera of untreated Hodgkin's lymphoma (HL) patients in advanced disease stages, in order to quantify the oxidative stress. HL patients displayed significantly higher levels of both MDA/HNE and protein carbonyl groups as compared with healthy controls. This is the first evidence that a strong increase in HL is one of the most common haematological malignancies, representing approximately 30% of all lymphomas in the circulating protein carbonyl content in HL. These findings may contribute to a better definition of the redox homoeostasis dysregulation in HL.

Oxidation of lipids is one of the basic processes causing rancidity in food products. Since application of natural antioxidants may be one of the technically simplest ways of reducing fat oxidation, we studied the effect of heating on antioxidant effectiveness and the chemical composition of basil, cinnamon, clove, nutmeg, oregano and thyme essential oils. When maintained at room temperature, all the oils tested appeared endowed with good radical-scavenger properties in the DPPH assay (effectiveness order: clove≫cinnamon>nutmeg>basil⩾oregano≫thyme). When heated up to 180 °C, nutmeg oil (but not the other essential oils under study) showed a significantly higher free radical-scavenger activity and evident changes in its chemical composition. Furthermore, the ability of these essential oils to protect α-tocopherol, contained in virgin olive oil, against thermal oxidative degradation was investigated. All the essential oils tested appeared able to prevent α-tocopherol loss following oil heating at 180 °C for 10 min (efficiency order: clove>thyme⩾cinnamon>basil≫oregano>nutmeg). In conclusion, the essential oils under study exhibited good antioxidant properties and might be efficiently used to control lipid oxidation during food processing.

We studied the serum levels of vitamins A, E, zinc and copper in two hundred and twenty-five subjects of both sexes. They were divided into two groups: 87 healthy subjects who served as controls and 138 patients with neoplastic disease. The patients were subdivided according to the absence (n = 79) or the presence of metastatic disease (n =59). In 59 patients with cancer, who were in therapy with scavenger drugs of free radical such as calcium antagonists and the antagonists of receptors H2, we also studied the possible effect of the same therapy on the serum levels of vitamins, on the concentrations of the microelements and on membrane lipid peroxidation. We found that membrane lipid peroxidation, evaluated from the time of in vitro formation in the blood of so-called "Heinz bodies," decreased in all patients treated with scavenger drugs. In these patients the permeability of the erythrocyte membrane was similar to the controls and the serum levels of the vitamins were equal to the levels in patients who did not receive these therapies. Zinc concentration increased while copper remained unchanged. We also studied the levels of vitamins in some organs. The results are discussed considering the role of free radicals. We underline the importance of vitamins A and E in the protection from membranous peroxidation and from free radicals and the need to consider cancer as a systemic morbid event, apart from the contingent actual location.

Familial chronic nail candidiasis (FCNC.MIM 607644) is a rare disorder characterized by early onset infections caused by different species of Candida and restricted to the nails; this disorder is genetically associated with low serum concentration of intercellular adhesion molecule 1 (ICAM-1). Herein we report the evidence of high circulating levels of malondialdehyde (MDA) and 4-hydroxy-2,3-nonenal (HNE) in seven patients of a five-generation Italian family affected by FCNC.MIM 607644. The present data evidence, in these patients, an increase in circulating MDA and HNE levels. Only some merely speculative hypotheses may be suggested to explain the mechanisms subserving the oxidative stress condition observed in these genetically ICAM-1 deficient patients; however, one has to point out that a chronic oxidative stress condition could contribute to the development of concurrent pathological alterations in which an overproduction of free radicals may play a central role.

Familiar chronic nail candidiasis (FCNC) is a rare disorder characterized by early-onset infections caused by different species of Candida, restricted to the nail of the hands and feet, and associated with a low serum concentration of intercellular adhesion molecule 1. Host defense mechanisms against candidiasis require the cooperation of many immune cells through several candidacidal mechanisms, including oxygen-dependent killing mechanisms, mediated by a superoxide anion radical myeloperoxidase-H2O2- halide system, and reactive nitrogen intermediates. We analyzed protein carbonyl groups (considered a useful marker of oxidative stress) in the serum of patients belonging to a five-generation Italian family with an isolated form of FCNC. Serum protein carbonyl groups in FCNC patients were significantly lower than those measured in healthy donors. Also, if this hypothesis is merely speculative, we could suggest that the decreased circulating level of protein carbonyl groups in these patients is not a marker of a lower oxidative stress condition, but might be linked to a lower protease activity.

Benign prostate hypertrophy (BPH) is the most common benign tumor in men due to obstruction of the urethra and, finally, uremia. Malondialdehyde (MDA) is a product derived from peroxidation of polyunsaturated fatty acids and related esters. Evaluation of MDA in serum represents a non-invasive biomarker of oxidative stress. Prostate-specific antigen (PSA) is a sensitive marker for prostatic hypertrophy and cancer. We analyzed MDA serum levels to evaluate the oxidative stress in BPH. To this end, 22 BPH patients and 22 healthy donors were enrolled. Data show an increase of MDA level in BPH patients and a positive correlation between PSA and MDA levels. In conclusion, we describe a previously unknown relationship between PSA and MDA as an index of inflammation and oxidative stress in BPH.