Staphylococci are a major health threat because of increasing resistance to antibiotics. An alternative to antibiotic treatment is preventing virulence by inhibition of bacterial cell-to-cell communication using the quorum-sensing inhibitor RNAIII-inhibiting peptide (RIP). In this work, we identified 2Ј,5-di-O-galloyl-Dhamamelose (hamamelitannin) as a nonpeptide analog of RIP by virtual screening of a RIP-based pharmacophore against a database of commercially available small-molecule compounds. Hamamelitannin is a natural product found in the bark of Hamamelis virginiana (witch hazel), and it has no effect on staphylococcal growth in vitro; but like RIP, it does inhibit the quorum-sensing regulator RNAIII. In a rat graft model, hamamelitannin prevented device-associated infections in vivo, including infections caused by methicillin-resistant Staphylococcus aureus and Staphylococcus epidermidis strains. These findings suggest that hamamelitannin may be used as a suppressor to staphylococcal infections.

We investigated the efficacy of LL-37, the C-terminal part of the only cathelicidin in humans identified to date (termed human cationic antimicrobial protein), in three experimental rat models of gram-negative sepsis. Adult male Wistar rats (i) were given an intraperitoneal injection of 1 mg Escherichia coli 0111:B4 LPS, (ii) were given 2 ؋ 10 10 CFU of Escherichia coli ATCC 25922, or (iii) had intra-abdominal sepsis induced via cecal ligation and puncture. For each model, all animals were randomized to receive intravenously isotonic sodium chloride solution, 1-mg/kg LL-37, 1-mg/kg polymyxin B, 20-mg/kg imipenem, or 60-mg/kg piperacillin. Lethality; growth of bacteria in blood, peritoneum, spleen, liver, and mesenteric lymph nodes; and endotoxin and tumor necrosis factor alpha (TNF-␣) concentrations in plasma were evaluated. All compounds reduced lethality compared to levels in controls. Endotoxin and TNF-␣ plasma levels were significantly higher in conventional antibiotic-treated rats than in LL-37-and polymyxin B-treated animals. All drugs tested significantly reduced bacterial growth compared to saline treatment. No statistically significant differences between LL-37 and polymyxin B were noted for antimicrobial and antiendotoxin activities. LL-37 and imipenem proved to be the most effective treatments in reducing all variables measured. Due to its multifunctional properties, LL-37 may become an important future consideration for the treatment of sepsis.

Resveratrol (Res) has been reported to possess cancer chemopreventive activity on the basis of its in vitro effects on tumor cells and in vivo experimental models of rodents transplanted with parental tumors or treated with carcinogens. We investigated the effects of Res on the development of mammary tumors appearing spontaneously in HER-2/neu transgenic mice at an early age. The mechanisms involved in the Res antitumor effect were evaluated by studying the immune effectiveness, tumor apoptosis and expression of mRNA and protein for HER-2/neu in tumoral mammary glands from Res-treated mice and in tumor cell lines. Res supplementation delayed the development of spontaneous mammary tumors (p < 0.001), reduced the mean number and size of mammary tumors (p < 0.0001) and diminished the number of lung metastases in HER-2/neu transgenic mice. The effects of Res were associated with downregulation of HER-2/neu gene expression and increased apoptosis both in tumoral mammary glands and in murine (N202) and human (SKBr3) tumor cell lines. Neither the basal, the IL-2-induced NK activity nor the lymphocyte number and proliferation was modified in Res-supplemented compared to control mice. Our results demonstrate that Res supplementation delays the development and reduces the metastasizing capacity of spontaneous mammary tumors in HER-2/neu transgenic mice. The antitumor effect of Res might be related to the downregulation of HER-2/neu expression and the induction of apoptosis in tumor cells. ' 2005 Wiley-Liss, Inc.Key words: resveratrol; HER-2/neu; mammary tumor; transgenic mouse; apoptosis Cancer chemoprevention, the prevention of cancer by chemical agents that reduce the risk of carcinogenesis, is one of the most direct ways to reduce morbidity and mortality. Res (trans-3,4 0 , 5-trihydroxystilbene) is a naturally occurring polyphenolic antioxidant compound present in grapes, mulberries, peanuts and red wine. It has been identified as an excellent candidate cancer chemopreventive, based on its safety and efficacy in experimental models of carcinogenesis. It has been found to inhibit diverse cellular events associated with tumor initiation, promotion and progression. 1 Most of the antitumor activity of Res has been discovered through in vitro studies in tumor cell lines. In these studies, Res was generally found to induce growth inhibition and apoptosis of a panel of human and murine tumor cell lines. [2][3][4][5][6][7] Fewer studies have been conducted on the antitumor effect exerted by in vivo supplementation with Res. Res was found to inhibit tumorigenesis in a mouse skin cancer model. 1 In other studies, it protected rats from ascites hepatoma 8 and colon carcinogenesis 9 and exerted inhibitory effects on tumor growth and lung metastasis in mice bearing highly metastatic Lewis lung carcinoma. 10 In the latter study, the effect of Res was related to its proapoptotic and antiangiogenic action on tumor cells, with no evidence of modulation of the immune response. 10 Res also inhibited the growth of experimentally implante...

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Background/Aims: Head and neck squamous cell carcinoma (HNSCC) ranks sixth worldwide for tumor-related mortality. A subpopulation of tumor cells, termed cancer stem cells (CSCs), has the ability to support cancer growth. Therefore, profiling CSC-enriched populations could be a reliable tool to study cancer biology. Methods: We performed phenotypic characterization of 7 HNSCC cell lines and evaluated the presence of CSCs. CSCs from Hep-2 cell line and HNSCC primary cultures were enriched through sphere formation and sphere-forming cells have been characterized both in vitro and in vivo. In addition, we investigated the expression levels of Nicotinamide N-methyltransferase (NNMT), an enzyme overexpressed in several malignancies. Results: CSC markers were markedly expressed in Hep-2 cell line, which was found to be highly tumorigenic. CSC-enriched populations displayed increased expression of CSC markers and a strong capability to form tumors in vivo. We also found an overexpression of CSC markers in tumor formed by CSC-enriched populations. Interestingly, NNMT levels were significantly higher in CSC-enriched populations compared with parental cells. Conclusion: Our study provides an useful procedure for CSC identification and enrichment in HNSCC. Moreover, results obtained seem to suggest that CSCs may represent a promising target for an anticancer therapy.

Staphylococcus aureus is a prevalent cause of bacterial infections associated with indwelling medical devices. RNA III inhibiting peptide (RIP) is known to inhibit S. aureus pathogenesis by disrupting quorum-sensing mechanisms. RIP was tested in the present study for its ability to inhibit S. aureus biofilm formation in a rat Dacron graft model. The activity of RIP was synergistic with those of antibiotics for the complete prevention of drug-resistant S. aureus infections.Staphylococci are a common cause of nosocomial infections related to bacterial biofilm formation on implanted devices (17). Central venous catheters, urinary catheters, prosthetic heart valves, orthopedic implants, and dialysis catheters are among the most common targets of infection (21, 28). Staphylococcus aureus and Staphylococcus epidermidis are often the predominant species found on biofilms after the removal of devices (20,28). Infections related to bacterial biofilm formation may result in longer hospitalizations, a need for surgery, and even death (6). In hemodialysis patients, S. aureus is the leading cause of bacteremia, with an attributable mortality rate of 5 to 19%, often due to endocarditis (20). Peritoneal catheter-related infections (exit site and tunnel) are also often caused by S. aureus isolates and, as in hemodialysis patients, can be difficult to eradicate.Infections characterized by bacterial biofilm formation can be described as follows. The biofilm structure is highly resistant to antibiotic treatment. Several explanations for the mechanism of biofilm resistance have been given, such as the failure of antimicrobial agents to penetrate the biofilm; the different metabolic states of the cells in the biofilm, which results in reduced susceptibilities to antimicrobial agents; and the expression of different genes by bacteria on a surface compared with those expressed by the planktonic counterparts (7, 10). One of the most intriguing explanations, however, is the possibility that antimicrobial resistance in the biofilm is acquired as a multicellular strategy, in which a "multicellular organism" collectively withstands antimicrobial treatments that would kill a lone cell (11,12,28). A novel way to prevent biofilm formation is to interfere with the bacterial cell-cell communication that leads to the virulence phenotype (4, 11, 27). The organization of the biofilm into complex structures is regulated by the exchange of chemical signals between cells in a process known as quorum sensing. Until now, two quorum-sensing mechanisms have been described in S. aureus. The first one is composed of the autoinducer RNA III-activating peptide (RAP) and its target protein, TRAP (3, 4). The second is composed of the peptide AIP and its receptor, AgrC (18,19). When the amount of RAP reaches a threshold concentration, it induces the phosphorylation of TRAP, which, through an unknown mechanism, leads to increased cell adhesion and to the activation of the regulatory system agr, which encodes AIP and AgrC. AIP downregulates TRAP phosphorylation, ...

Sepsis remains a major cause of morbidity and mortality in hospitalized patients, despite intense efforts to improve survival. The primary lead for septic shock results from activation of host effector cells by endotoxin, the lipopolysaccharide (LPS) associated with cell membranes of gram-negative bacteria. For these reasons, the quest for compounds with antiendotoxin properties is actively pursued. We investigated the efficacy of the amphibian skin antimicrobial peptide temporin L in binding Escherichia coli LPS in vitro and counteracting its effects in vivo. Temporin L strongly bound to purified E. coli LPS and lipid A in vitro, as proven by fluorescent displacement assay, and readily penetrated into E. coli LPS monolayers. Furthermore, the killing activity of temporin L against E. coli was progressively inhibited by increasing concentrations of LPS added to the medium, further confirming the peptide's affinity for endotoxin. Antimicrobial assays showed that temporin L interacted synergistically with the clinically used ␤-lactam antibiotics piperacillin and imipenem. Therefore, we characterized the activity of temporin L when combined with imipenem and piperacillin in the prevention of lethality in two rat models of septic shock, measuring bacterial growth in blood and intra-abdominal fluid, endotoxin and tumor necrosis factor alpha (TNF-␣) concentrations in plasma, and lethality. With respect to controls and single-drug treatments, the simultaneous administration of temporin L and ␤-lactams produced the highest antimicrobial activities and the strongest reduction in plasma endotoxin and TNF-␣ levels, resulting in the highest survival rates.

The results of the in vivo study demonstrated that APDT with RLP068/Cl may be useful in the management of chronic infected wounds, accelerating the repair process through a significant bacterial inhibition.