The temporins are a family of small, linear antibiotic peptides with intriguing biological properties. We investigated the antibacterial, haemolytic and cytotoxic activities of temporin L (FVQWFSKFLGRIL-NH2), isolated from the skin of the European red frog Rana temporaria. The peptide displayed the highest activity of temporins studied to date, against both human erythrocytes and bacterial and fungal strains. At variance with other known temporins, which are mainly active against Gram-positive bacteria, temporin L was also active against Gram-negative strains such as Pseudomonas aeruginosa A.T.C.C. 15692 and Escherichia coli D21 at concentrations comparable with those that are microbiocidal to Gram-positive bacteria. In addition, temporin L was cytotoxic to three different human tumour cell lines (Hut-78, K-562 and U-937), causing a necrosis-like cell death, although sensitivity to the peptide varied markedly with the specific cell line tested. A study of the interaction of temporin L with liposomes of different lipid compositions revealed that the peptide causes perturbation of bilayer integrity of both neutral and negatively charged membranes, as revealed by the release of a vesicle-encapsulated fluorescent marker, and that the action of the peptide is modulated to some extent by membrane lipid composition. In particular, the presence of negatively charged lipids in the model bilayer inhibits the lytic power of temporin L. We also show that the release of fluorescent markers caused by temporin L is size-dependent and that the peptide does not have a detergent-like effect on the membrane, suggesting that perturbation of bilayer organization takes place on a local scale, i.e. through the formation of pore-like openings.

Temporins, antimicrobial peptides of 10±13 residues, were isolated from secretions of Rana temporaria [Simmaco, M., Mignogna, G., Canofeni, S., Miele, R., Mangoni, M.L. & Barra, D. (1996) Eur. J. Biochem. 242, 788±792]. These molecules are specific to this amphibian species, which is also able to secrete on its skin other antimicrobial peptides similar to those found in different Rana species. The effect of temporins A, B and D (13 residues, net charge +2), and H (10 residues, net charge +1 and +2, respectively) against both artificial membranes of differing lipid composition and bacteria has been investigated in order to gain insight into their mechanisms of action. The results indicate that: the lytic activity of temporins is not greatly affected by the membrane composition; temporins A and B allow the leakage of large-size molecules from the bacterial cells; temporin H renders both the outer and inner membrane of bacteria permeable to hydrophobic substances of low molecular mass; and temporin D, although devoid of antibacterial activity, has a cytotoxic effect on erythrocytes. The results allow important conclusions to be drawn about the minimal structural requirements for lytic efficiency and specificity of temporins.

Antimicrobial peptides are produced by all organisms in response to microbial invasion and are considered as promising candidates for future antibiotics. There is a wealth of evidence that many of them interact and increase the permeability of bacterial membranes as part of their killing mechanism. However, it is not clear whether this is the lethal step. To address this issue, we studied the interaction of the antimicrobial peptide temporin L with Escherichia coli by using fluorescence, confocal and electron microscopy. The peptide previously isolated from skin secretions of the frog Rana temporaria has the sequence FVQWFSKFLGRIL-NH2. With regard to fluorescence microscopy, we applied, for the first time, a triple-staining method based on the fluorochromes 5-cyano-2,3-ditolyl tetrazolium chloride, 4',6-diamidino-2-phenylindole and FITC. This technique enabled us to identify, in the same sample, both living and total cells, as well as bacteria with altered membrane permeability. These results reveal that temporin L increases the permeability of the bacterial inner membrane in a dose-dependent manner without destroying the cell's integrity. At low peptide concentrations, the inner membrane becomes permeable to small molecules but does not allow the killing of bacteria. However, at high peptide concentrations, larger molecules, but not DNA, leak out, which results in cell death. Very interestingly, in contrast with many antimicrobial peptides, temporin L does not lyse E. coli cells but rather forms ghost-like bacteria, as observed by scanning and transmission electron microscopy. Besides shedding light on the mode of action of temporin L and possibly that of other antimicrobial peptides, the present study demonstrates the advantage of using the triple-fluorescence approach combined with microscopical techniques to explore the mechanism of membrane-active peptides in general.

The innate immunity of multicellular organisms relies in large part on the action of antimicrobial peptides (AMPs) to resist microbial invasion. Crafted by evolution into an extremely diversified array of sequences and folds, AMPs do share a common amphiphilic 3-D arrangement. This feature is directly linked with a common mechanism of action that predominantly (although not exclusively) develops upon interaction of peptides with cell membranes of target cells. This minireview reports on current understanding of the modes of interaction of AMPs with biological and model membranes, especially focusing on recent insights into the folding and oligomerization requirements of peptides to bind and insert into lipid membranes and exert their antibiotic effects. Given the potential of AMPs to be developed into a new class of anti-infective agents, emphasis is placed on how the information on peptide-membrane interactions could direct the design and selection of improved biomimetic synthetic peptides with antibiotic properties.

Resveratrol, a naturally occurring polyphenol, shows pleiotropic health beneficial effects, including anti-oxidant, anti-inflammatory, anti-aging, cardioprotective and neuroprotective activities. Due to the several protective effects and since this compound is widely distributed in the plant kingdom, resveratrol can be envisaged as a chemo-preventive/curative agent introduced almost daily with the diet. Currently, a number of preclinical findings suggest resveratrol as a promising nature's weapon for cancer prevention and treatment. A remarkable progress in elucidating the molecular mechanisms underlying anti-cancer properties of resveratrol has been achieved in the last years. Concerning the resveratrol mechanism of action as a protective (vs. normal cells and tissues) and toxic (vs. cancer cells) compound, many studies focus on its antioxidant capacity as well as on its ability to trigger and favor the apoptotic cascade in malignant cells. However, a generalized mechanism of action able to explain this dual effect of resveratrol has not yet been clearly established. In addition to these important functions, resveratrol is reported to exhibit several other biological/biochemical protective effects on heart, circulation, brain and age-related diseases which are summarized in this Review.