DISCO

The aim of this study was to investigate p l a s m a m e m b r a n e fluidity in h u m a n keratinocytes using fluorescence anisotropy of 1,6-diphenyl-l,3,5-hexatriene (DPH) and its cationic derivative 1-[4-(trimethylamino)-phenyl]-6-phenyl-l,3,5-hexatriene (TMA-DPH). Keratinocytes from n o r m a l or psoriatic skin were isolated using trypsin-EDTA or dispase. In keratinocytes isolated from n o r m a l skin, T M A -D P H anisotropy values were higher than those observed using DPH; the difference must be related to the different localization of the two probes. In fact, D P H in whole cells localizes in p l a s m a as well as intracellular m e m b r a n e s , yielding an average value of fluidity, while the cationic derivative T M A -D P H resides in the p l a s m a m e m b r a n e of the whole cells for a sufficient time for anisotropy measurements. Moreover, it has to be considered that p l a s m a m e m b r a n e is more ordered than intracellular m e mbranes. The kinetics of incorporation of T M A -D P H was similar in keratinocytes isolated using trypsin-EDTA and those isolated using; dispase, however, the fluorescence anisotropy values were lower in keratinocytes isolated with dispase (0.260 + 0.01 vs 0.270 + 0.01, p = 0.029). This difference is p r o b a b l y related to modifications of lipid-protein interactions after trypsin treatment. Since no d a m a g e to plasma m e m b r a n e after incubation with dispase seems to have been reported, we decided to use this separation procedure to study p l a s m a m e m b r a n e fluidity in psoriasis, a h u m a n pathological condition characterized by excessive cell proliferation and incomplete differentiation. Lower anisotropy values (0.260 + 0.01 vs 0.270 + 0.01, p = 0.001), indicating an increase in fluidity, were observed O. Simonetti (1~) in keratinocytes isolated f r o m skin of psoriatic patients than in epidermal cells isolated from n o r m a l hum a n skin. We suggest that the m e a s u r e m e n t of fluorescence anisotropy in living cells is a convenient and useful tool to study m e m b r a n e fluidity in h u m a n keratinocytes isolated f r o m n o r m a l and diseased skin. Its application represents a technical advance because plasma m e m b r a n e fluidity can be measured using very limited a m o u n t s of tissue, as obtained from biopsies.

The minimum inhibitory concentrations (MICs), the minimal fungicidal concentrations (MFCs), the fungal biomass (FB) and hyphal viability employing the dye 3-4,5 dimethyl-2-thiazolyl-2,5-diphenyl-2H tetrazolium bromide (MTT) were used to compare the in vitro effects of fluconazole (FLU) with those of the Nterminal palmitoyl-lipidated peptide, Pal-Lys-Lys-NH 2 (PAL), and a tea tree oil component, g-Terpinene (TER), against several clinical isolates of Microsporum canis and Trichophyton rubrum. In general, FLU and PAL MICs were significantly lower than those observed with TER, while no differences in the three drugs were found in the MFCs. However, they were from two to 16-times higher than their respective MICs. FB of M. canis treated with either FLU or PAL, but not with TER, was significantly reduced over untreated controls. Only PAL and TER, in a medium-dependent fashion, but not FLU, reduced the FB of T. rubrum. Finally, PAL was found to be significantly more active than FLU at reducing the hyphal viability against both genera of dermatophytes. This study shows that PAL exerts an in vitro activity against dermatophytes at least similar to that observed with FLU and suggests that this compound might be a promising candidate in the treatment of infections due to dermatophytes.

Among the most common complications of both chronic wound and surgical sites are staphylococcal skin infections, which slow down the wound healing process due to various virulence factors, including the ability to produce biofilms. Furthermore, staphylococcal skin infections are often caused by methicillin-resistant Staphylococcus aureus (MRSA) and become a therapeutic challenge. The aim of this narrative review is to collect the latest evidence on old and new anti-staphylococcal therapies, assessing their anti-biofilm properties and their effect on skin wound healing. We considered antibiotics, quorum sensing inhibitors, antimicrobial peptides, topical dressings, and antimicrobial photo-dynamic therapy. According to our review of the literature, targeting of biofilm is an important therapeutic choice in acute and chronic infected skin wounds both to overcome antibiotic resistance and to achieve better wound healing.

Psoriasis has been associated with an abnormal plasma lipid metabolism, and changes of erythrocyte membrane lipid composition and fluidity have been shown in adult patients. To investigate whether the alterations of plasma lipids appear also in pediatric patients, we have studied plasma lipids and lipoproteins in 15 prepubertal children affected by mild-to-moderate psoriasis with respect to healthy controls. The patients showed higher levels of plasma total cholesterol (4.44 + 0.78 versus 4.03 f 0.58 mmol/L), a significant increase of cholesterol associated with HDL (1.39 +. 0.26 versus 1.13 +. 0.28 mmol/ L, p = 0.02), and a significant decrease of the ratio LDL cholesterol to HDL cholesterol (1.73 + 0.6 versus 2.46 2 0.8, p = 0.02). By using fluorescence polarization of 1,6diphenyl-1,3,5-hexatriene, we have shown a significant increase in fluidity in erythrocyte membrane of psoriatic children that was associated with a slight, but not significant, decrease in the cholesterol to protein ratio (422 + 127 versus 503 f 117 nmol/mg). No significant changes of phospholipid fatty acid composition have been shown, in disagreement with previous studies in adult patients. Our results support the relation between childhood psoriasis and plasma lipid changes, which are likely related to the slight compositional changes in erythrocytes. However, the observed abnormalities are expressed differently in children than in adults. (Pediatr Res 33: 506-509, 1993) Abbreviations DPH, 1,6-diphenyl-1,3,5-hexatriene HDL-C, cholesterol associated with high-density lipoprotein LDL-C, cholesterol associated with low-density lipoprotein Pf, fluorescence polarization TC, total cholesterol TG, triglyceride VLDL-C, cholesterol associated with very low-density lipoprotein Psoriasis is a chronic inflammatory skin disease characterized by an accelerated turnover of epidermal cells and an incomplete differentiation in lesional epidermidis. The etiology of psoriasis is unknown, but genetic, metabolic, and immunologic mechanisms have been proposed (1). Various disorders of the plasma lipid and lipoprotein pattern that include an increase in TG in VLDL and a decrease in HDL-C have been shown in adult

Aims There is evidence that plasma coenzyme Q 10 (CoQ 10 ) levels decrease in patients with advanced chronic heart failure (CHF). However, it is not known whether oral CoQ 10 supplementation may improve cardiocirculatory efficiency and endothelial function in patients with CHF.

This paper reports on the synthesis and properties of a new UV-absorber (OC-NO) based on the most popular UV filter worldwide, ethylhexyl methoxycinnamate (OMC) in which the methoxy group has been replaced with a pyrrolidine nitroxide bearing antioxidant activity. This sunscreen active has therefore both UVabsorbing and antioxidant properties which could ideally address both the UV-B and UV-A skin photodamage. For broad-spectrum coverage, the combinations of OC-NO with two commonly used UV-A absorbers (BMDBM and DHHB) were also studied. The results obtained reveal that OC-NO: (a) is as photostable as OMC after UV-A exposure; (b) acts as free radical scavenger as demonstrated by EPR and chemical studies; (c) reduces UV-A and UV-A+BMDBM induced lipid peroxidation in liposomes and cells, measured as reduced TBARS levels and increased C11-BODIPY red fluorescence, respectively; (d) has comparable antioxidant activity to that of vitamin E and BHT commonly used in skin care formulations; (e) is non-cytotoxic to human skin fibroblasts as assessed with the MTT assay when exposed to increasing doses of UV-A; and (f) OC-NO+ DHHB is a promising, photostable broad spectrum UV-filter combination that concomitantly reduces UVinduced free radical damage. These results suggest that nitroxide/antioxidant-based UV-absorbers may pave the way for the utilization of 'multi-active' ingredients in sunscreens thereby reducing the number of ingredients in these formulations.

Aims This randomized controlled study was designed to determine whether oral coenzyme Q 10 (CoQ 10 ) supplementation (100 mg tid) was able to improve extracellular superoxide dismutase (ecSOD) activity and endothelium-dependent (ED) vasodilation in patients with coronary artery disease (CAD). ecSOD, a major antioxidant enzyme system of the vessel wall, is reduced in patients with CAD. Moreover, there is a strong correlation between endothelium-bound ecSOD and the ED dilation of conduit arteries. CoQ 10 has been recently shown to improve the ED relaxation in diabetic patients. Methods and results Thirty-eight CAD patients (33 M/5 F, mean age 55 + 4 years, ejection fraction 57.5 + 8%) were randomized into two groups. One group (n ¼ 19) received CoQ 10 orally at doses of 300 mg/day for 1 month, whereas the other group received a placebo. On entry and after 1 month, all patients underwent brachial artery ED assessment, cardiopulmonary exercise test, and the measurement of endothelium-bound ecSOD activity. A total of 33 patients completed the study. ecSOD, ED relaxation, as well as peak VO 2 and O 2 pulse increases in the CoQ 10 -treated group were statistically greater vs. the variations in the placebo group. In particular, improvements elicited by CoQ 10 supplementation were remarkable in subjects presenting low initial endothelium-bound ecSOD and thus more prone to oxidative stress. Conclusion Improvements in the ED relaxation and endothelium-bound ecSOD activity might be related to CoQ 10 capability of enhancing endothelial functionality by counteracting nitric oxide oxidation. The enhancement of peak VO 2 and of O 2 pulse is likely due to the bioenergetic effect of CoQ 10 ; on the other end, the improved VO 2 could also depend on the observed enhanced peripheral endothelial function.

Reactive oxygen species seem to play an important role in vascular homeostasis. In conditions of high oxidative stress, such as chronic heart failure and multiple coronary risk factors, the rate of inactivation of nitric oxide to peroxynitrite by superoxide anions may be reduced by CoQ10, which can also protect against nitrosative damage. CoQ10 may also influence vascular function indirectly via inhibition of oxidative damage to LDL. Patients with lower levels of extracellular superoxide dismutase (ecSOD) demonstrate greater improvements than patients with normal ec-SOD levels, suggesting that the higher the oxidative stress the greater the improvement in the endothelium-dependent relaxation after the administration of a compound with antioxidant properties like CoQ10. Future studies are needed to inquire whether these effects may translate into benefits in clinical practice.

a b s t r a c t Extracellular superoxide dismutase (SOD3) is the primary enzymatic antioxidant defence of the vascular wall. The physiopathological role of SOD3 has been examined in vascular-related diseases, atherosclerosis, hypertension, diabetes, ischaemia-reperfusion injury, lung disease, various inflammatory conditions, and neurological diseases. An important single nucleotide polymorphism (SNP), nt.760 G > C of the SOD3 gene (rs#1799895) leads to the amino acid substitution Arg(213)Gly (R213G) in the center of the heparinbinding domain and consequently to a lowered affinity for the endothelium. This mutation, which occurs with a relatively high frequency in the population (4% of Swedish, 3% of Australian and 6% of Japanese people), is associated with decreased tissue antioxidant defences and increased risk of ischaemic heart disease. The identification of patients carrying this mutation is therefore of great interest in order to highlight lowered antioxidant defences at a vascular level which could lead to increased susceptibility toward coronary artery disease and atherogenesis. Here we describe a method to detect the 760 G > C single nucleotide polymorphism based on Real Time PCR strategy using locked nucleic acid (LNA) probes. This technique, a modification of classic TaqMan probes SNP genotyping, amplifies and detects the mutation in a single reaction tube. Moreover, the implementation of LNA probes remarkably increases the specificity of the reaction. The proposed method enables unambigous and rapid discrimination of wild type and mutant genotype both in plasmid and genomic DNA samples. In light of the role of SOD3 polymorphism, the genotyping of 760 G > C mutant has important clinical implications. The proposed assay combines rapidity, high specificity, can be easily automated and overall reduces labor and cost of analyses. Moreover, identification of patients with lowered vascular antioxidant defences could address pharmacogenomical approaches to the therapy of cardiovascular diseases.

UV-C radiation is able to impair cellular functions by directly damaging DNA, and by inducing an increased formation of reactive oxygen species that leads to a condition of oxidative stress. In this study we evaluated different responses to UV insult of two leukemia cell lines, HL-60 and Raji, and the relationship with their CoQ10 content. DNA damage was monitored by means of the alkaline single cell gel electrophoresis (Comet assay); intracellular levels of ROS, mitochondrial depolarization and cell viability was measured by flow cytometry. Raji cells appeared more resistant to the UV insult; moreover, they did not show any increase in ROS content and the extent of mitochondrial depolarisation was much lower than in HL 60 cell line. Raji cells also contained significantly higher levels of CoQ10 and their ability to incorporate and to reduce exogenous CoQ10 added to the culture medium was remarkably elevated compared with HL 60.

Background: There is evidence that plasma CoQ10 levels decrease in patients with advanced chronic heart failure (CHF).Objective: To investigate whether oral CoQ10 supplementation could improve cardiocirculatory efficiency in patients with CHF.Methods: We studied 21 patients in NYHA class II and III (18M, 3W, mean age 59 ±9 years) with stable CHF secondary to ischemic heart disease (ejection fraction 37 ± 7%), using a double-blind, placebo-controlled cross-over design. Patients were assigned to oral CoQ10 (100 mg tid) and to placebo for 4 weeks, respectively.Results: CoQ10 supplementation resulted in a threefold increase in plasma CoQ10 level (P<0.0001 vs placebo). Systolic wall thickening score index (SWTI) was improved both at rest and peak dobutamine stress echo after CoQ10 supplementation (+12.1 and 15.6%, respectively, P<0.05 vs placebo). Left ventricular ejection fraction improved significantly also at peak dobutamine (15% from study entry P<0.0001) in relation to a decrease in LV end-systolic volume index (from 57 ± 7 mL/m2 to 45 mL/m2, P<0.001). Improvement in the contractile response was more evident among initially akinetic (+33%) and hypokinetic (+25%) segments than dyskinetic ones (+6%). Improvement in SWTI was correlated with changes in plasma CoQ10 levels (r=−0.52, P<0.005). Peak VO2 was also improved after CoQ10 as compared with placebo (+13%, <0.005). No side effects were reported with CoQ10.Conclusions: Oral CoQ10 improves LV contractility in CHF without any side effects. This improvement is associated with an enhanced functional capacity.

Aims This randomized controlled study was designed to determine whether oral coenzyme Q 10 (CoQ 10 ) supplementation (100 mg tid) was able to improve extracellular superoxide dismutase (ecSOD) activity and endothelium-dependent (ED) vasodilation in patients with coronary artery disease (CAD). ecSOD, a major antioxidant enzyme system of the vessel wall, is reduced in patients with CAD. Moreover, there is a strong correlation between endothelium-bound ecSOD and the ED dilation of conduit arteries. CoQ 10 has been recently shown to improve the ED relaxation in diabetic patients. Methods and results Thirty-eight CAD patients (33 M/5 F, mean age 55 + 4 years, ejection fraction 57.5 + 8%) were randomized into two groups. One group (n ¼ 19) received CoQ 10 orally at doses of 300 mg/day for 1 month, whereas the other group received a placebo. On entry and after 1 month, all patients underwent brachial artery ED assessment, cardiopulmonary exercise test, and the measurement of endothelium-bound ecSOD activity. A total of 33 patients completed the study. ecSOD, ED relaxation, as well as peak VO 2 and O 2 pulse increases in the CoQ 10 -treated group were statistically greater vs. the variations in the placebo group. In particular, improvements elicited by CoQ 10 supplementation were remarkable in subjects presenting low initial endothelium-bound ecSOD and thus more prone to oxidative stress. Conclusion Improvements in the ED relaxation and endothelium-bound ecSOD activity might be related to CoQ 10 capability of enhancing endothelial functionality by counteracting nitric oxide oxidation. The enhancement of peak VO 2 and of O 2 pulse is likely due to the bioenergetic effect of CoQ 10 ; on the other end, the improved VO 2 could also depend on the observed enhanced peripheral endothelial function.

Objective: To evaluate the effectiveness of coenzyme Q 10 treatment in improving semen quality in men with idiopathic infertility. Design: Placebo-controlled, double-blind randomized trial. Setting: Patient(s): Sixty infertile patients (27-39 years of age) with the following baseline sperm selection criteria: concentration >20 Â 10 6 /mL, sperm forward motility <50%, and normal sperm morphology >30%; 55 patients completed the study. Intervention(s): Patients underwent double-blind therapy with coenzyme Q 10 , 200 mg/day, or placebo; the study design was 1 month of run-in, 6 months of therapy or placebo, and 3 months of follow-up. Main Outcome Measure(s): Variations in semen parameters used for patient selection and variations of coenzyme Q 10 and ubiquinol concentrations in seminal plasma and spermatozoa. Result(s): Coenzyme Q 10 and ubiquinol increased significantly in both seminal plasma and sperm cells after treatment, as well as spermatozoa motility. A weak linear dependence among the relative variations, baseline and after treatment, of seminal plasma or intracellular coenzyme Q 10 and ubiquinol levels and kinetic parameters was found in the treated group. Patients with a lower baseline value of motility and levels of coenzyme Q 10 had a statistically significant higher probability to be responders to the treatment. Conclusion(s): The exogenous administration of coenzyme Q 10 increases the level of the same and ubiquinol in semen and is effective in improving sperm kinetic features in patients affected by idiopathic asthenozoospermia.

Down syndrome (DS) is a chromosomal abnormality (trisomy 21) associated with mental retardation and Alzheimer like dementia, characteristic change of the individual's phenotype and premature ageing. Oxidative stress is known to play a major role in this pathology since a gene dose effect leads to elevated ratio of superoxide dismutase to catalase/glutathione peroxidase compared to controls in all age categories suggesting that oxidative imbalance contributes to the clinical manifestation of DS. Hyperuricemia is another feature of DS that has an interesting relationship with oxidative stress since uric acid represents an important free radical scavenger. However its formation is connected to the conversion of Xanthine dehydrogenase (XDH) to Xanthine oxidase (XO) which leads to concomitant production of free radicals. Here we report that plasma samples from DS patients in pediatric age, despite an increased total antioxidant capacity, largely due to elevated Uric acid content (UA), present significantly elevated markers of oxidative damage such as increased allantoin levels. Moreover DS plasma samples do not differ from healthy control ones in terms of Coenzyme Q10 and susceptibility to peroxidative stimuli. On the contrary, lymphocyte and platelet CoQ10 content was significantly lower in DS patients, a fact that might underlie oxidative imbalance at a cellular level.