Alpha-lipoic acid, but not di-hydrolipoic acid, activates Nrf2 response in primary human umbilical-vein endothelial cells and protects against TNF-α induced endothelium dysfunction.

The antioxidants role in cell response regulation attracted great interest in the last decades and it is undergoing to a profound reconsideration. The mere concept of "biological antioxidant" has been frequently misconceived or misused, possibly leading to the misinterpretation of some experimental observation. Organosulfur compounds in general and α-lipoic acid, a dithiol molecule, can be considered a typical example of the kind. Reduced α-lipoic acid, dehydrolipoic acid has been in fact originally considered a bona fide, reducing, electron donor molecule. A more recent approach, according to stoichiometric and thermodynamic evidences, lead to a reinterpretation of the biochemical role of "antioxidants". The electrophilic nature of oxidized nucleophilic molecules, including α-lipoic acid, renders more plausible a mechanism based on the ability to activate Nrf2/EpRE mediated hormetic response. In this study, we demonstrate that nmolar concentrations of oxidized α-lipoic acid, but not dehydrolipoic acid, protect human umbilical primary endothelial cells (HUVEC) from TNF-α induced dysfunction, inhibit NF-κB activation and block apoptosis following the activation of Nrf2 transcription factor. Our observations corroborate the concept that the major, if not the unique, mechanism by which α-lipoic acid can non-enzymatically exert its reducing activity is related to the electrophilic nature of the oxidized form.

Experimental exposure of blue mussels (Mytilus galloprovincialis) to high levels of benzo[a]pyrene and possible implications for human health.

Polycyclic aromatic hydrocarbons (PAHs) are lipophilic compounds able to accumulate in the food chain. Mussels showed to bioaccumulate contaminants, such as PAHs, so that recurrent consumption of such contaminated food represents a risk for human health. This study was aimed to elucidate if acute exposure of Mediterranean blue mussel (Mytilus galloprovincialis), a bivalve of great economic importance in several countries, to a PAH, benzo[a]pyrene (B[a]P), at doses able to induce cytochrome P450 1A (CYP1A) and pathological changes in mussel gills, can produce accumulation in soft tissue. We explored the cytotoxic effects (cell viability, DNA laddering, and glutathione levels) of in vitro exposure of human peripheral blood mononuclear cells (PBMCs) to organic extracts obtained from blue mussels previously exposed for 12 and 72h via water to B[a]P (0.5-1mg/L). In our experimental conditions, B[a]P induced CYP1A induction and morphological changes in mussel gills and a significant B[a]P accumulation in soft tissue. Conversely, exposing PBMCs to organic extracts obtained from contaminated mussels, resulted in a significant reduction of cell viability and cell glutathione content, and in an increase in DNA laddering. This confirms that consumption of mussels from B[a]P polluted waters might affect human health. Our data lead us to suggest that CYP1A activity in mussel gills may be useful (more than the amount of detected PAHs in the mussel edible tissue) as a marker in assessment of risk for health of consumers exposed to PAHs through ingestion of shellfish.

Exposure of sea bream (Sparus aurata) to toxic concentrations of benzo[a]pyrene: possible human health effect.

Polycyclic aromatic hydrocarbons (PAHs) can accumulate in the food chain, due to their lipophilic properties. Fish can accumulate contaminants including PAHs and frequent consumption of such contaminated fish can pose risk to human health. The aim of this study was to clarify if acute exposure of sea bream (Sparus aurata, a fish species of great economic importance in the Atlantic and Mediterranean areas) to a PAH, benzo[a]pyrene (B[a]P), at a dose that can induce CYP1A and pathological changes in fish gills, liver and muscle, can induce accumulation in muscle. We investigated the cytotoxic effects (as changes in cell viability, DNA laddering and glutathione content) of in vitro exposure of human peripheral blood mononuclear cells (PBMCs) to organic extracts obtained from muscle of sea breams previously exposed via water to B[a]P (2mg/l, for 12, 24 and 72 h). At this level of exposure, B[a]P caused morphological changes, inflammatory response and CYP1A induction not only in sea bream gills and liver but also in muscle; furthermore, in fish muscle we observed a substantial B[a]P accumulation, which may be associated with the increased CYP1A activity in liver and especially in muscle. However, when PBMCs were exposed to organic extracts obtained from sea bream muscle contaminated with B[a]P, a toxic, although modest effect was revealed, consisting in a significant decrease in cell glutathione levels without alterations in cell viability and DNA laddering. This suggests that consumption of sea breams from B[a]P contaminated waters might represent a risk for human health.

Flavonoid profile, antioxidant and antiglycation properties of Retama sphaerocarpa fruits extracts.

Retama sphaerocarpa occurs in the Mediterranean area of North-east Africa and in the Iberian Peninsula, and grows on a variety of soil types and climatic conditions. Used in Algerian folk medicine, it is a valuable species for revegetation and soil restoration. The aim of this study is to evaluate flavonoid composition and antioxidant and antiglycation properties of methanolic and aqueous extracts from R. sphaerocarpa fruits. HPLC-PDA/ESI-MS was used to identify/quantify flavonoid content. Antioxidant activity was evaluated by Folin-Ciocalteu, ORAC, FRAP, TEAC, and DPPH assays, and antiglycation capability by glucose/fructose-BSA assay. Results showed that fruits contain isoflavones (daidzein and genistein derivatives) and flavonols (apigenin, isorhamnetin, kaempferol and quercetin derivatives), and extracts (especially the methanolic one, richer in flavonoids) possess good in vitro antioxidant and antiglycation properties. These findings evidence that R. sphaerocarpa fruits are a source of valuable phytochemicals, with potential applications in the field of phytopharmaceuticals and in food industry.

Antimicrobial susceptibility and beta-lactamase production of anaerobic and aerobic bacteria isolated from pus specimens from orofacial infections.

Most suppurative orofacial infections are polymicrobial. Information regarding the antimicrobial susceptibility of the microorganisms involved can be useful in the choice of an effective antibiotic therapy. In this study we determined the antimicrobial susceptibility of a total 235 anaerobic and aerobic bacteria recently isolated from pus specimens of orofacial infections. All the viridans streptococci were susceptible to penicillin, cefotaxime, cefoxitin, imipenem and levofloxacin. Imipenem and levofloxacin were active against 100% of the anaerobic Gram-positive organisms isolated. Among the anaerobic Gram-negative rods beta-lactamase production was detected in all species except Campylobacter rectus. Amoxicillin-clavulanate, cefoxitin, imipenem and metronidazole were active against all the isolates of anaerobic Gram-negative species. Isolates resistant to erythromycin were found in all the species tested, however, resistance to clindamycin was only detected in Porphyromonas gingivalis and Bacteroides ureolyticus. Isolates resistant to levofloxacin were detected in P. gingivalis and Prevotella sp.

In vitro protective effect of a Jacquez grapes wine extract on UVB-induced skin damage.

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.

Immunomodulatory properties of cefaclor: in vivo effect on cytokine release and lymphoproliferative response in rats.

The proper and coordinated response of the host immune system to bacterial infections is known to play a central role in the eradication of an infection. Therefore, the impact of antibiotics on both innate and acquired host immunity may be involved in the therapeutic outcome. The aim of this study was to evaluate the effects of the widely used cephalosporin cefaclor on some parameters of the immune system in ex vivo conditions. The results demonstrated that short-term (3 to 6 days) treatment with this antibiotic induced pleiotropic modification of rat spleen cells upon ex vivo stimulation with the polyclonal mitogen PHA, entailing increased lymphoproliferative responses, augmented IFN-gamma, IL-2 and IL-10 synthesis and decreased production of IL-4 and IL-6 in comparison to spleen cells from control rats. The mononuclear spleen cells of healthy rats released larger amounts of IFN-gamma and IL-2 in culture supernatants in response to polyclonal mitogenic stimulation with PHA compared to the spleens of the control rats receiving vehicle only. Simultaneously, the treatment with cefaclor augmented PHA-induced lymphoproliferative responses and reduced the synthesis of IL-4 and IL-6. These data depict a type 1 cytokine inducing and immunostimulatory pharmacological profile that, by activating the innate and acquired immune system, would be synergistic with cefaclor antibacterial activity.

Antibacterial activity of propolis and its active principles alone and in combination with macrolides, beta-lactams and fluoroquinolones against microorganisms responsible for respiratory infections.

Propolis is produced by bees and is reported to have several pharmaceutical properties. Its antibacterial activity against strains causing upper respiratory tract infections is particularly important: propolis might be used as a therapeutic agent to prevent the bacterial infections that sometimes overlap viral infections. In this study the in vitro activity of both an alcoholic solution and a hydroglyceric extract of propolis, as well as its active principles, was tested against bacteria responsible for respiratory infections (Streptococcus pneumoniae, Haemophilus influenzae, Haemophilus parainfluenzae, Moraxella catarrhalis and Streptococcus pyogenes). We also evaluated the in vitro activity of a combination of propolis and its active principles and some beta-lactams, macrolides and fluoroquinolones. Our results, though not demonstrating a clearly synergistic activity between antibiotics and propolis and its constituents, show the possibility of using natural preparations, due to their antimicrobial and anti-inflammatory properties, to enhance antibacterial therapy.

Protective activity of an anthocyanin-rich extract from bilberries and blackcurrants on acute acetaminophen-induced hepatotoxicity in rats.

Acetaminophen (N-acetyl-p-aminophenol, APAP) overdosage can produce fatal centrilobular hepatic necrosis in humans. The present study attempted to investigate the protective effect of an anthocyanin-rich extract from bilberries and blackcurrants (AE) against APAP-induced acute hepatic damage in rats. Treatment with AE normalised blood activities of glutamate oxaloacetate and glutamate pyruvate transaminase and prevented APAP-induced plasmatic and tissutal alterations in biomarkers of oxidative stress, probably due to various bioproperties of the components of the extract.

Antibiotic susceptibility and serotype distribution in Streptococcus pneumoniae circulating in Italy: results of the SEMPRE surveillance study (2000-2002).

During 2000-2002, 20 clinical microbiology centres collected 1623 Streptococcus pneumoniae isolates. Susceptibility to penicillin, amoxicillin, amoxicillin/clavulanic acid, cefaclor, cefuroxime, cefotaxime, clarithromycin, ciprofloxacin, levofloxacin, rifampicin and teicoplanin was determined locally by the Etest and/or by the microdilution method by three co-ordinating centres. Total resistance to penicillin increased from 15.2% to 16.1% and macrolide resistance increased from 37.9% to 43.7%. Overall, the most effective drugs (>99% susceptible strains) were amoxicillin, amoxicillin/clavulanic acid, levofloxacin and rifampicin. The most frequent serotypes were: 23F (15.8%), 3 (10.8%) 14 (9.1%), 19F (9.1%), 6B (7.2%), 19A (6.9%) and 6A (4.8%). In conclusion, penicillin and macrolide resistance is increasing in Italy, whilst fluoroquinolone currently remains active. The most common serotypes circulating are included in the heptavalent conjugate vaccine, with the exception of type 3.

The Sentinel Project: an update on the prevalence of antimicrobial resistance in community-acquired respiratory Streptococcus pneumoniae and Haemophilus spp. in Italy.

A total of 460 Streptococcus pneumoniae and Haemophilus spp. collected from respiratory infections during 2000 was tested for their susceptibility to 15 selected antibiotics. Overall, penicillin resistance among pneumococci was 10.5%, while lack of susceptibility to macrolides, co-trimoxazole, tetracycline and chloramphenicol reached 35.2%, 26.2%, 22.6% and 6.0%, respectively. Amoxicillin/clavulanic acid and levofloxacin were the most potent compounds (100% and 99.9% susceptible strains, respectively). Among isolates of Haemophilus influenzae and Haemophilus parainfluenzae, beta-lactamase production (12.5% and 10%, respectively), and co-trimoxazole (19.9% and 40.0%) and clarithromycin (11.2% and 40.0%) resistance were the prevalent threats. This study confirms the trend observed in Italy since 1992: macrolide resistance among respiratory microorganisms is increasing, while several drugs including amoxicillin/clavulanic acid, third generation injectable cephalosporins and fluoroquinolones remain active on the great majority of these pathogens.

Palmitate-induced endothelial dysfunction is attenuated by cyanidin-3-O-glucoside through modulation of Nrf2/Bach1 and NF-κB pathways.

Free fatty acids (FFA), commonly elevated in diabetes and obesity, have been shown to impair endothelial functions and cause oxidative stress, inflammation, and insulin resistance. Anthocyanins represent one of the most important and interesting classes of flavonoids and seem to play a role in preventing cardiovascular diseases. Herein, we investigated the in vitro protective effects of cyanidin-3-O-glucoside (C3G) on cell signaling pathways in human umbilical vein endothelial cells (HUVECs) exposed to palmitic acid (PA), the most prevalent saturated FFA in circulation. Our data reported a significant augmentation of free radicals and oxidative stress in HUVECs exposed to PA for 3h, while C3G pretreatment improved intracellular redox status altered by FFA. Moreover, C3G significantly inhibited NF-κB proinflammatory pathway and adhesion molecules induced by PA, and these effects were attributed to the activation of Nrf2/EpRE pathway. In fact, C3G induced Nrf2 nuclear localization and activation of cellular antioxidant and cytoprotective genes at baseline and after PA exposure in endothelial cells. Our data confirm the hypothesis that natural Nrf2 inducers, such as C3G, might be a potential therapeutic strategy to protect vascular system against various stressors preventing several pathological conditions.

Internalization-associated proteins among Streptococcus pyogenes isolated from asymptomatic carriers and children with pharyngitis.

Sixty-two strains of Streptococcus pyogenes isolated from 30 asymptomatic school children and 32 children with pharyngitis were characterized to analyze the involvement of 2 fibronectin-binding proteins (F/SfbI and PrtF2/PfbpI) in S. pyogenes colonizing asymptomatic carriers and to determine the possible association between these proteins and the genes associated with macrolide resistance. In this study, we demonstrated that the proportion of S. pyogenes strains carrying the pfbpI gene was significantly higher among asymptomatic carriers (80%) than among children with pharyngitis (53%; P<.05). With regard to the proportion of prtF1-positive strains, no significant differences were found between the 2 groups (70% vs. 69%, for asymptomatic carriers and children with pharyngitis, respectively). Another important finding is the significant association between macrolide resistance and protein F/SfbI (P<.001) in both groups. These results suggest that the presence of the pfbpI gene can be linked to the ability of S. pyogenes to persist in the throat of asymptomatic carriers.

Enhanced percentage of Leu M3+DR+ and Leu M3+CD25+ cells in newly diagnosed IDDM patients.

The percentage of Leu M3+DR+ and of Leu M3+CD25+ cells was determined by means of immunofluorescence analysis in a group of patients with insulin dependent diabetes mellitus (IDDM). Our results show that an increased percentage of these cells may occur in the early stage of the disease. These data provide evidence for a "phenotypical" activation of Leu M3+ cells at the onset of the disease and warrant future studies to evaluate the potential role of these cells in the pathogenesis of IDDM.

Antimicrobial activity of quinupristin/dalfopristin, a new injectable streptogramin with a wide Gram-positive spectrum.

Quinupristin/dalfopristin (Synercid) is a new injectable streptogramin antibiotic proposed for the treatment of severe antimicrobial infections, that has been shown to be active against Gram-positive, multi-resistant cocci. We compared the in vitro activity of quinupristin/dalfopristin with that of amoxycillin, ampicillin, penicillin, cefixime, ceftriaxone, clindamycin, erythromycin, imipenem, meropenem, oxacillin, piperacillin/tazobactam, teicoplanin and vancomycin. The susceptibility of 37 Staphylococcus aureus (14 MS, 23 MR), 26 Staphylococcus epidermidis (16 MS, 10 MR), 20 Streptococcus pneumoniae, 33 Group A Streptococcus pyogenes, 15 Streptococcus agalactiae, 10 Enterococcus faecalis (1 vancomycin-resistant), 15 Enterococcus faecium (9 van A) was evaluated. Quinupristin/dalfopristin was active against all Gram-positive species tested, including met-R S. aureus (MIC < or = 2 mg/l), met-R S. epidermidis (MIC < or = 2 mg/l), S. pneumoniae (MIC < or = 1 mg/l), ery-R and ery-S streptococci (MIC < or = 1 mg/l). The strains of E. faecalis were generally less susceptible. Time-kill studies confirmed that quinupristin/dalfopristin at 4 x MIC concentration showed a complete bactericidal effect (3 log reduction) in about 4 6 h against all strains tested. A post-antibiotic effect (PAE) of 3.9-5.2 h was observed at 4 x MIC concentration of quinupristin/dalfopristin against staphylococci. A prolonged PAE was obtained for S. pneumoniae (8 h), S. pyogenes (9 h) and S. agalactiae (7 h), while the shortest PAE was seen for E. faecalis and E. faecium (about 4 h).

Molecular mechanisms of resistance in Pseudomonas aeruginosa to fluoroquinolones.

Pseudomonas aeruginosa is important in the field of infectious disease especially with respect to its role in nosocomial infections. Infections with P. aeruginosa may be a problem as the organism has intrinsic resistance to several antibiotics and a capability in acquiring resistance during antibiotic therapy. Fluoroquinolones are sometimes used during antibiotic therapy of P. aeruginosa infections even though resistance to fluoroquinolones may develop. Six strains of P. aeruginosa were studied in an attempt to elucidate the mechanisms of resistance to fluoroquinolones. These included the electrophoresis patterns of the outer membrane proteins (OMPs), random amplified polymorphic DNA (RAPD) and polymerase chain reaction (PCR) analyses. A method is described that improved the clarity of the OMP gels. Resistance in these P. aeruginosa strains could depend not only on DNA-gyrase modifications but also on membranes alterations and on the presence (qualitative and quantitative) of the efflux pump formed by three subunits.

Minimal inhibitory concentrations and time-kill determination of moxifloxacin against aerobic and anaerobic isolates.

Moxifloxacin is a new oral 8-methoxy-quinolone with a wide spectrum of activity against Gram-negative and anaerobic bacteria, atypical micro-organisms and multi-resistant Gram-positive bacteria. This study was designed to assess the in vitro activity of moxifloxacin against Gram-positive bacteria with different resistance patterns, anaerobes and atypical micro-organisms such as Chlamydia and Mycoplasma. Moxifloxacin had good activity against Streptococcus pneumoniae with all strains inhibited by < or =0.12 mg/l. The minimal inhibitory concentrations (MICs) of moxifloxacin for Streptococcus pyogenes and Streptococcus agalactiae ranged from 0.03 to 0.5 mg/l while those of ciprofloxacin were about two- to four-fold higher (MICs=0.12-1 mg/l). Moxifloxacin was poorly active against enterococci but its activity against Clostridium and Bacteroides spp. was in the same range as that of metronidazole and superior to that of clindamycin. Moxifloxacin was substantially more active than both ciprofloxacin and sparfloxacin against Chlamydia.

The Italian Epidemiological Survey 1997-1999. Antimicrobial susceptibility data of Haemophilus influenzae, Haemophilus parainfluenzae and Moraxella catarrhalis in Italy.

The Italian Epidemiological Survey began a surveillance study with the aim of monitoring the antimicrobial resistance of respiratory pathogens. From 1997 to 1999, 2028 strains of Haemophilus influenzae and 523 strains of Haemophilus parainfluenzae were collected from 59 Clinical Microbiology Laboratories distributed throughout Italy. In 1998, the study was extended to include Moraxella catarrhalis and a total of 360 isolates were collected. There was a significant increase in the beta-lactamase production both for H. influenzae (from 5% in 1997 to 16% in 1999) and for H. parainfluenzae (from 5% in 1997 to 22% in 1999). Beta-lactamase production in M. catarrhalis was 84% in 1998 and 87% in 1999. Beta-lactamase production affected the susceptibility to unprotected penicillins (87% in H. influenzae, 85% in H. parainfluenzae and 34% in M. catarrhalis), and in part the susceptibility to cefaclor (about 98%). Amoxycillin/clavulanate, cefixime, ceftriaxone and ciprofloxacin were active against all strains of H. influenzae, H. parainfluenzae and M. catarrhalis.

Antibacterial activity of moxifloxacin against periodontal anaerobic pathogens involved in systemic infections.

The in vitro activity of moxifloxacin was compared with that of penicillin G, amoxycillin/clavulanate, cefoxitin, erythromycin, clindamycin and metronidazole against 158 isolates associated with periodontal infections. MIC(50)/MIC(90) values of moxifloxacin were respectively 0.06/0.5 mg/l for Porphyromonas gingivalis (35), for Prevotella spp. (28) and Actinomyces spp. (35), 0.12/0.25 mg/l for Fusobacterium nucleatum (20) and 0.06/0.12 mg/l for Peptostreptococcus spp. (30). The minimum inhibitory concentration (MIC) range of moxifloxacin for Bacteroides forsythus (6) and Campylobacter rectus (4) was 0.06-0.12 mg/l. The minimum bactericidal concentrations were equal to or 2-4 times the MIC values. Moxifloxacin produced a bactericidal effect at 8 h. Our results show that moxifloxacin has good antibacterial activity against periodontal pathogens comparable with that of cefoxitin and amoxycillin/clavulanate, and better than that of clindamycin, metronidazole and penicillin.

Berberis aetnensis C. Presl. extracts: antimicrobial properties and interaction with ciprofloxacin.

Previous research showed that berberine-containing Berberis species synthesise the substances 5'-methoxyhydnocarpin-D (5'-MHC-D) and pheophorbide a, which have no antimicrobial activity but inhibit the expression of multidrug resistant efflux pumps (MDRs) in Staphylococcus aureus and potentiate the action of berberine. The MDR pumps extrude synthetic and natural antimicrobials from bacterial cells. We searched for these compounds in Berberis aetnensis C. Presl. (Berberidaceae), an endemic plant of the volcano Mount Etna. This work confirms the presence of pheophorbide a and permits us to hypothesise the presence of 5'-MHC-D in leaf extracts. In fact, the activity of ciprofloxacin was improved when two chromatographic fractions isolated from leaf extracts were added. These results are indicative of the presence of MDR pump inhibitors. Moreover, crude extracts were tested on several micro-organisms and showed antimicrobial activity mainly against Gram-positive bacteria and yeasts.