Morin inhibits Listeria monocytogenes virulence in vivo and in vitro by targeting listeriolysin O and inflammation.

BACKGROUND: Listeria monocytogenes (L. monocytogenes) is a global opportunistic intracellular pathogen that can cause many infections, including meningitis and abortion in humans and animals; thus, L. monocytogenes poses a great threat to public safety and the development of the aquaculture industry. The isolation rate of Listeria monocytogenes in fishery products has always been high. And the pore-forming toxin listeriolysin O (LLO) is one of the most important virulence factors of L. monocytogenes. LLO can promote cytosolic bacterial proliferation and help the pathogen evade attacks from the host immune system. In addition, L. monocytogenes infection can trigger a series of severe inflammatory reactions. RESULTS: Here, we further confirmed that morin lacking anti-Listeria activity could inhibit LLO oligomerization. We also found that morin can effectively alleviate the inflammation induced by Listeria in vivo and in vitro and exerted an obvious protective effect on infected cells and mice. CONCLUSIONS: Morin does not possess anti-Listeria activity, neither does it interfere with secretion of LLO. However, morin inhibits oligomerisation of LLO and morin does reduce the inflammation caused during Listeria infection.

Optimizing the Composition of Irrigation Fluid to Reduce the Potency of Staphylococcus aureus α-Toxin: Potential Role in the Treatment of Septic Arthritis.

OBJECTIVE: Septic arthritis is commonly caused by Staphylococcus aureus and is a medical emergency requiring antibiotics and joint irrigation. The bacteria produce α-toxin causing rapid cartilage cell (chondrocyte) death. Saline (0.9%NaCl) lavage is normally used to remove bacteria and toxins, however, its composition might be suboptimal to suppress the lethal effects of α-toxin. We utilized rabbit erythrocyte hemolysis as a sensitive, biologically relevant assay of α-toxin levels to determine if changes to osmolarity, temperature, pH, and divalent cation (Mg2+, Ca2+) concentration were protective. DESIGN: Erythrocytes were incubated in the various conditions and then exposed to α-toxin ("chronic" challenge) or incubated with α-toxin and then exposed to experimental conditions ("acute" challenge). RESULTS: Raising osmolarity from 300 mOsm (0.9%NaCl) to 400, 600, or 900 mOsm (sucrose addition) when applied chronically, significantly reduced hemolysis linearly. As an acute challenge, osmotic protection was significant and similar over 400 to 900 mOsm. Reducing temperature chronically from 37°C to 25°C and 4°C significantly reduced hemolysis, however, when applied as an acute challenge although significant, was less marked. Divalent cations (Mg2+, Ca2+ at 5mM) reduced hemolysis. Varying pH (6.5, 7.2, 8.0) applied chronically marginally reduced hemolysis. The optimized saline (0.9% NaCl; 900 mOsm with sucrose, 5 mM MgCl2 (37°C)) rapidly and significantly reduced hemolysis compared with saline and Hank's buffered saline solution applied either chronically or acutely. CONCLUSIONS: These results on the effect of S. aureus α-toxin on erythrocytes showed that optimizing saline could markedly reduce the potency of S. aureus α-toxin. Such modifications to saline could be of benefit during joint irrigation for septic arthritis.

Impact of Cigarette Smoke Condensate on Adhesion-Related Traits and Hemolysin Production of Oral Candida dubliniensis Isolates.

BACKGROUND: Cigarette smoke is associated with higher oral Candida carriage and possible predisposition and increased susceptibility to oral candidal infection. Candida dubliniensis is associated with oral candidosis. Candidal adherence to buccal epithelial cells (BEC) and denture acrylic surfaces (DAS), germ tube (GT) formation, cell surface hydrophobicity (CSH) and hemolysin production are pathogenic traits of Candida. OBJECTIVES: The impact of exposure to cigarette smoke on the aforementioned pathogenic attributes of oral C. dubliniensis has not been studied. Hence, the impact of cigarette smoke condensate (CSC) on adhesion to BEC and DAS, GT formation, CSH and hemolysin production of 20 oral C. dubliniensis isolates after exposure to CSC for 24, 48 and 72 h was ascertained. METHODS: After preparation of the CSC, using an in-house smoking device, the Candida isolates were exposed to the CSC for 24, 48 and 72 h, by a previously described in vitro method. Thereafter, the adhesion to BEC and DAS, GT formation, CSH and hemolysin production of C. dubliniensis isolates was investigated by hitherto described in vitro assays. RESULTS: Exposure to CSC significantly increased the ability of C. dubliniensis oral isolates to adhere to BEC, DAS, GT formation, CSH and produce hemolysin following 24-h, 48-h and 72-h exposure periods to CSC (P < 0.001 for all attributes tested). CONCLUSIONS: Exposure of oral C. dubliniensis isolates to CSC may significantly promote in vitro adhesion traits and hemolysin production of these isolates, thereby augmenting its pathogenicity in vitro in the presence of cigarette smoke.

In vitro Post-Antifungal Effect of Posaconazole and Its Impact on Adhesion-Related Traits and Hemolysin Production of Oral Candida dubliniensis Isolates.

OBJECTIVE: Candidal adherence to denture acrylic surfaces (DAS) and oral buccal epithelial cells (BEC), formation of candidal germ tubes (GT), candidal cell surface hydrophobicity (CSH), and hemolysin production are important pathogenic traits of Candida. The antifungal drug-induced post-antifungal effect (PAFE) also impacts the virulence of Candida. Candida dubliniensis isolates are associated with the causation of oral candidiasis which could be managed with posaconazole. Thus far there is no evidence on posaconazole-induced PAFE and its impact on adhesion-related attributes and production of hemolysin by C. dubliniensis isolates. Hence, the PAFE, adhesion to DAS and BEC, formation of GT, CSH, and hemolysin production of 20 oral C. dubliniensis isolates after brief exposure to posaconazole was ascertained. MATERIALS AND METHODS: The PAFE, adherence to DAS and BEC, formation of GT, candidal CSH, and hemolysin production were investigated by hitherto described in vitro assays. RESULTS: The mean PAFE (h) induced by posaconazole on C. dubliniensis isolates was 1.66. Exposure to posaconazole suppressed the ability of C. dubliniensis to adhere to DAS, BEC, formation of candidal GT, candidal CSH and to produce hemolysin by a reduction of 44, 33, 34, 36, and 15% (p < 0.005 to p < 0.001), respectively. CONCLUSION: Exposure of C. dubliniensis isolates to posaconazole for a brief period induced an antimycotic impact by subduing its growth in addition to suppressing pathogenic adherence-associated attributes, as well as production of hemolysin.

Antioxidant Properties and Antibacterial Effects of Eucalyptus camaldulensis Ethanolic Leaf Extract on Biofilm Formation, Motility, Hemolysin Production, and Cell Membrane of the Foodborne Pathogen Listeria monocytogenes.

Consumer concerns toward chemical preservatives have resulted in increased search for healthy green alternative. In this study, the antioxidant activity and antibacterial effects of Eucalyptus camaldulensis ethanolic leaf extract against Listeria monocytogenes, a serious foodborne pathogen, was evaluated. Total phenolic and flavonoid contents of the extract were 11.10 mg garlic acid equivalent/mg extract and 15.05 mg quercetin equivalent/mg extract, respectively. The minimum inhibitory concentration (MIC) and minimum bactericidal concentration of the extract was 64-128 µg/mL and 256-512 µg/mL, respectively. Time-kill assay revealed growth inhibitory effects after 4-h treatment of the bacteria with the extract. A reduction of ≈2-3 log colony-forming units per milliliter was observed against the tested food and environmental isolates after challenging the pathogens with the extract at MIC for 6 h. Sub-MICs of the extract significantly inhibited motility and listeriolysin O production up to 80%, with 60% inhibition of biofilm formation (p < 0.05). Antioxidant assay revealed free radical scavenging activity with 50% inhibitory concentration (IC50) of 57.07 µg/mL for 2,2-diphenyl-1-picrylhydrazyl and 29.01 µg/mL for ABTS [2,2'-azino-bis(3-ethylbenzothiazoline-6-sulfonic acid)] assay. Ferric reducing antioxidant power assay further showed a total antioxidant power equivalent to 92.93 µM ascorbic acid equivalent/mg extract. As the extract exhibited profound antilisterial activity and good radical scavenging ability, it might serve as a potential alternative source of biopreservative agent against L. monocytogenes.

Identification and Validation of an Antivirulence Agent Targeting HlyU-Regulated Virulence in Vibrio vulnificus.

Antimicrobial resistance (AMR) in pathogens is the result of indiscriminate use of antibiotics and consequent metabolic/genetic modulation to evolve survival strategies and clonal-selection in AMR strains. As an alternative to antibiotic treatment, antivirulence strategies are being developed, not only to combat bacterial pathogenesis, but also to avoid emerging antibiotic resistance. Vibrio vulnificus is a foodborne pathogen that causes gastroenteritis, necrotizing wound infections, and sepsis with a high rate of mortality. Here, we developed an inhibitor-screening reporter platform to target HlyU, a master transcriptional regulator of virulence factors in V. vulnificus by assessing rtxA1 transcription under its control. The inhibitor-screening platform includes wild type and ΔhlyU mutant strains of V. vulnificus harboring the reporter construct P rtxA1::luxCDABE for desired luminescence signal detection and control background luminescence, respectively. Using the inhibitor-screening platform, we identified a small molecule, fursultiamine hydrochloride (FTH), that inhibits the transcription of the highly invasive repeat-in-toxin (rtxA1) and hemolysin (vvhA) along with other HlyU regulated virulence genes. FTH has no cytotoxic effects on either host cells or pathogen at the tested concentrations. FTH rescues host cells from the necrotic cell-death induced by RtxA1 and decreases the hemolytic activity under in vitro conditions. The most important point is that FTH treatment does not induce the antivirulence resistance. Current study validated the antivirulence strategy targeting the HlyU virulence transcription factor and toxin-network of V. vulnificus and demonstrated that FTH, exhibits a potential to inhibit the pathogenesis of deadly, opportunistic human pathogen, V. vulnificus without inducing AMR.

Eriodictyol protects against Staphylococcus aureus-induced lung cell injury by inhibiting alpha-hemolysin expression.

Staphylococcus aureus (S. aureus) is a common pathogenic bacterium that causes various diseases in both humans and animals. With the increased prevalence of methicillin-resistant S. aureus, the therapeutic effects of commonly used antibiotics are limited against S. aureus infection. Novel treatment strategies and new antibiotics are needed urgently to address this concern. Many studies have shown that virulence factors secreted from S. aureus play vital roles in their pathogenic processes. Alpha-hemolysin (Hla), an important exotoxin in S. aureus, is one such virulence factor that increases sensitivity of multiple host cells to S. aureus resulting in various diseases. Eriodictyol is a flavonoid compound that exists in many fruits and vegetables. In this study, eriodictyol was demonstrated to inhibit the expression of Hla by hemolysis assays, western blotting, and RT-qPCR at the sub-minimal inhibitory concentration. In live/dead and cytotoxicity assays, the results showed that eriodictyol protected A549 cells against Hla-induced injury in a dose-dependent manner. The minimal inhibitory concentration of eriodictyol against S. aureus was 512 µg/mL. Eriodictyol can downregulate S. aureus Hla at both the expressional and transcriptional levels without affecting S. aureus growth. In addition, cell assays had proved that eriodictyol could protect A549 cells against Hla damage. Eriodictyol could therefore have the potential to treat S. aureus infection targeting Hla.

Lysionotin attenuates Staphylococcus aureus pathogenicity by inhibiting α-toxin expression.

α-Toxin, one of the best known pore-forming proteins produced by Staphylococcus aureus (S. aureus), is a critical virulence factor in multiple infections. The necessity of α-toxin for S. aureus pathogenicity suggests that this toxin is an important target for the development of a potential treatment strategy. In this study, we showed that lysionotin, a natural compound, can inhibit the hemolytic activity of culture supernatants by S. aureus by reducing α-toxin expression. Using real-time PCR analysis, we showed that transcription of hla (the gene encoding α-toxin) and agr (the locus regulating hla) was significantly inhibited by lysionotin. Lactate dehydrogenase and live/dead assays indicated that lysionotin effectively protected human alveolar epithelial cells against S. aureus, and in vivo studies also demonstrated that lysionotin can protect mice from pneumonia caused by S. aureus. These findings suggest that lysionotin is an efficient inhibitor of α-toxin expression and shows significant protection against S. aureus in vitro and in vivo. This study supports a potential strategy for the treatment of S. aureus infection by inhibiting the expression of virulence factors and indicates that lysionotin may be a potential treatment for S. aureus pneumonia.

Antibiofilm activity of Vetiveria zizanioides root extract against methicillin-resistant Staphylococcus aureus.

Methicillin-resistant Staphylococcus aureus (MRSA) is a leading human pathogen responsible for causing chronic clinical manifestation worldwide. In addition to antibiotic resistance genes viz. mecA and vanA, biofilm formation plays a prominent role in the pathogenicity of S. aureus by enhancing its resistance to existing antibiotics. Considering the role of folk medicinal plants in the betterment of human health from the waves of multidrug resistant bacterial infections, the present study was intended to explore the effect of Vetiveria zizanioides root on the biofilm formation of MRSA and its clinical counterparts. V. zizanioides root extract (VREX) showed a concentration-dependent reduction in biofilm formation without hampering the cellular viability of the tested strains. Micrographs of scanning electron microscopy (SEM) and confocal laser scanning microscopy (CLSM) portrayed the devastating impact of VREX on biofilm formation. In addition to antibiofilm activity, VREX suppresses the production of biofilm related phenotypes such as exopolysaccharide, slime and α-hemolysin toxin. Furthermore, variation in FT-IR spectra evidenced the difference in cellular factors of untreated and VREX treated samples. Result of mature biofilm disruption assay and down regulation of genes like fnbA, fnbB, clfA suggested that VREX targets these adhesin genes responsible for initial adherence. GC-MS analysis revealed the presence of sesquiterpenes as a major constituent in VREX. Thus, the data of present study strengthen the ethnobotanical value of V. zizanioides and concludes that VREX contain bioactive molecules that have beneficial effect over the biofilm formation of MRSA and its clinical isolates.

Crataeva nurvala nanoparticles inhibit virulence factors and biofilm formation in clinical isolates of Pseudomonas aeruginosa.

Green synthesized nanoparticles have gained great attention due to their non-toxic and non-hazardous nature. In the present study, bark extract of the medicinal plant in Ayurveda Crataeva nurvala (Buch-Ham) (CN) was chosen for the biosynthesis of silver nanoparticles (AgNPs). These NPs were characterized by Ultra violet visible spectroscopy, Fourier Transform Infra Red, Atomic Force Microscopy, and Transmission Electron Microscopy (TEM). The average particle size of green synthesized CN-AgNPs was 15.2 ± 1.01 nm. Gas chromatography- mass spectrometry analysis of methanolic bark extract involved in the formation of CN-AgNPs revealed lupeol as a major active component. In this study, CN-AgNPs (15 µg ml-1 ) efficiently suppressed the production of quorum sensing mediated virulence factors viz. pyocyanin, protease, hemolysin, and biofilm formation in Pseudomonas aeruginosa. The pyocyanin production was strongly inhibited (74.64%) followed by proteolysis (47.3%) and hemolysin production (47.7%). However, the biofilm forming ability was maximally reduced up to 79.70%. Moreover, the Confocal Laser Scanning Microscopic Analysis showed that CN-AgNPs inhibit colonization of P. aeruginosa on to the surface. Furthermore, TEM analysis revealed internalization of CN-AgNPs inside the bacterial cell. It is concluded that green synthesized AgNPs have great potential to inhibit virulence factors and biofilm forming ability of drug-resistant clinical isolates of P. aeruginosa.

Impact of short-term exposure of antifungal agents on hemolysin activity of oral Candida dubliniensis isolates from Kuwait and Sri Lanka.

OBJECTIVE: Ability to produce hemolysin by Candida species is an important determinant of its pathogenicity. Candida dubliniensis is implicated in the causation of oral candidosis, which can be treated with polyene, echinocandin, and azole groups of antifungal agents as well as chlorhexidine. After oral application, however, the concentrations of these agents tend to decrease quickly to subtherapeutic levels due to the peculiarity of the oral environment. In this study, we have evaluated the effect of short-term exposure of sublethal concentrations of these drugs on hemolysin production by oral C. dubliniensis isolates obtained from two different geographical locale. MATERIALS AND METHODS: Twenty C. dubliniensis oral isolates obtained from Kuwait and Sri Lanka were exposed to sublethal concentrations of nystatin, amphotericin B, caspofungin, ketoconazole, fluconazole, and chlorhexidine for 1 h. Thereafter, the drugs were removed by dilution and the hemolysin production determined by a previously described plate assay. RESULTS: Hemolysin production of these isolates was significantly suppressed with a percentage reduction of 17.09, 16.45, 17.09, 11.39, 8.23 and 12.03 following exposure to nystatin, amphotericin B, caspofungin, ketoconazole, fluconazole, and chlorhexidine, respectively. CONCLUSION: Brief exposure to sublethal concentrations of drugs with antifungal properties appears to reduce the pathogenic potential of C. dubliniensis isolates by suppressing hemolysin production.

Impact of brief exposure to antifungal agents on the post-antifungal effect and hemolysin activity of oral Candida albicans.

Post-antifungal effect (PAFE) of Candida and its production of hemolysin are determinants of candidal pathogenicity. Candida albicans is the foremost aetiological agent of oral candidosis, which can be treated with polyene, azole, and echinocandin antifungals. However, once administered, the intraoral concentrations of these drugs tend to be subtherapeutic and transient due to the diluent effect of saliva and cleansing effect of the oral musculature. Hence, intra-orally, Candidamay undergo a brief exposure to antifungal drugs.Objective Therefore, the PAFE and hemolysin production of oral C. albicans isolates following brief exposure to sublethal concentrations of the foregoing antifungals were evaluated.Material and Methods A total of 50 C. albicans oral isolates obtained from smokers, diabetics, asthmatics using steroid inhalers, partial denture wearers and healthy individuals were exposed to sublethal concentrations of nystatin, amphotericin B, caspofungin, ketoconazole and fluconazole for 60 min. Thereafter, the drugs were removed and the PAFE and hemolysin production were determined by previously described turbidometric and plate assays, respectively.Results Nystatin, amphotericin B, caspofungin and ketoconazole induced mean PAFE (hours) of 2.2, 2.18, 2.2 and 0.62, respectively. Fluconazole failed to produce a PAFE. Hemolysin production of these isolates was suppressed with a percentage reduction of 12.27, 13.47, 13.33, 8.53 and 4.93 following exposure to nystatin, amphotericin B, caspofungin, ketoconazole and fluconazole, respectively.Conclusions Brief exposure to sublethal concentrations of antifungal drugs appears to exert an antifungal effect by interfering with the growth as well as hemolysin production of C. albicans.

Impact of brief exposure to antifungal agents on the post-antifungal effect and hemolysin activity of oral Candida albicans

AbstractPost-antifungal effect (PAFE) of Candida and its production of hemolysin are determinants of candidal pathogenicity. Candida albicans is the foremost aetiological agent of oral candidosis, which can be treated with polyene, azole, and echinocandin antifungals. However, once administered, the intraoral concentrations of these drugs tend to be subtherapeutic and transient due to the diluent effect of saliva and cleansing effect of the oral musculature. Hence, intra-orally, Candidamay undergo a brief exposure to antifungal drugs.Objective Therefore, the PAFE and hemolysin production of oral C. albicans isolates following brief exposure to sublethal concentrations of the foregoing antifungals were evaluated.Material and Methods A total of 50 C. albicans oral isolates obtained from smokers, diabetics, asthmatics using steroid inhalers, partial denture wearers and healthy individuals were exposed to sublethal concentrations of nystatin, amphotericin B, caspofungin, ketoconazole and fluconazole for 60 min. Thereafter, the drugs were removed and the PAFE and hemolysin production were determined by previously described turbidometric and plate assays, respectively.Results Nystatin, amphotericin B, caspofungin and ketoconazole induced mean PAFE (hours) of 2.2, 2.18, 2.2 and 0.62, respectively. Fluconazole failed to produce a PAFE. Hemolysin production of these isolates was suppressed with a percentage reduction of 12.27, 13.47, 13.33, 8.53 and 4.93 following exposure to nystatin, amphotericin B, caspofungin, ketoconazole and fluconazole, respectively.Conclusions Brief exposure to sublethal concentrations of antifungal drugs appears to exert an antifungal effect by interfering with the growth as well as hemolysin production of C. albicans.

Stilbenes reduce Staphylococcus aureus hemolysis, biofilm formation, and virulence.

Stilbenoids have a broad range of beneficial health effects. On the other hand, the emergence of antibiotic-resistant Staphylococcus aureus presents a worldwide problem that requires new antibiotics or nonantibiotic strategies. S. aureus produces α-hemolysin (a pore-forming cytotoxin) that has been implicated in the pathogenesis of sepsis and pneumonia. Furthermore, the biofilms formed by S. aureus constitute a mechanism of antimicrobial resistance. In this study, we investigated the hemolytic and antibiofilm activities of 10 stilbene-related compounds against S. aureus. trans-Stilbene and resveratrol at 10 µg/mL were found to markedly inhibit human blood hemolysis by S. aureus, and trans-stilbene also inhibited S. aureus biofilm formation without affecting its bacterial growth. Furthermore, trans-stilbene and resveratrol attenuated S. aureus virulence in vivo in the nematode Caenorhabditis elegans, which is normally killed by S. aureus. Transcriptional analysis showed that trans-stilbene repressed the α-hemolysin hla gene and the intercellular adhesion locus (icaA and icaD) in S. aureus, and this finding was in line with observed reductions in virulence and biofilm formation. In addition, vitisin B, a stilbenoid tetramer, at 1 µg/mL was observed to significantly inhibit human blood hemolysis by S. aureus.

Integrating artificial with natural cells to translate chemical messages that direct E. coli behaviour.

Previous efforts to control cellular behaviour have largely relied upon various forms of genetic engineering. Once the genetic content of a living cell is modified, the behaviour of that cell typically changes as well. However, other methods of cellular control are possible. All cells sense and respond to their environment. Therefore, artificial, non-living cellular mimics could be engineered to activate or repress already existing natural sensory pathways of living cells through chemical communication. Here we describe the construction of such a system. The artificial cells expand the senses of Escherichia coli by translating a chemical message that E. coli cannot sense on its own to a molecule that activates a natural cellular response. This methodology could open new opportunities in engineering cellular behaviour without exploiting genetically modified organisms.

[Effect of plant extracts on cytotoxic activity of Vibrio cholerae hemolysin].

AIM: Study of plant extracts that have the ability to neutralize cytotoxic activity of hemolysin. MATERIALS AND METHODS: Preparations of purified and recombinant V. cholerae eltor hemolysin as well as supernatants of V. cholerae strains were used. Determination ofcytotoxic activity of hemolysin and neutralizing activity of plant extracts were carried out by using cell cultures CHO-K1 and CaCo2. RESULTS: Out of 9 water extracts only 3 - extracts of Rhei rhizome, Limonium gmelinii and Quercus robur neutralized hemolysin in cell culture CHO-K1 and CaCo2, whereas the other extracts--Humulus lupulus, Ocimum basilicum, Chelidonium majus, Juglans regia, Achillea milefolium and Hypericum perforatum did not have anti-cytotoxic effect. Neutralizing properties of extracts are exhibited during their co-incubation with hemolysin preparations and supernatants of V. cholerae strains already within 10 minutes. CONCLUSION: Plant extracts that have anti-cytotoxic activity against hemolysin are perspective for development oftherapeutic-prophylaxis preparations.

Flavone reduces the production of virulence factors, staphyloxanthin and α-hemolysin, in Staphylococcus aureus.

Staphylococcus aureus is a leading cause of nosocomial infections due to its resistance to diverse antibiotics. This bacterium produces a large number of extracellular virulence factors that are closely associated with specific diseases. In this study, diverse plant flavonoids were investigated to identify a novel anti-virulence compound against two S. aureus strains. Flavone, a backbone compound of flavonoids, at subinhibitory concentration (50 µg/mL), markedly reduced the production of staphyloxanthin and α-hemolysin. This staphyloxanthin reduction rendered the S. aureus cells 100 times more vulnerable to hydrogen peroxide in the presence of flavone. In addition, flavone significantly decreased the hemolysis of human red blood by S. aureus, and the transcriptional level of α-hemolysin gene hla and a global regulator gene sae in S. aureus cells. This finding supported the usefulness of flavone as a potential antivirulence agent against antibiotic-resistant S. aureus.

Polysaccharide extracted from Rheum tanguticum prevents irradiation-induced immune damage in mice.

AIM: To investigate the protective effect of purified fraction 1 polysaccharide extracted from Rheum tanguticum RTP1 on irradiation-induced immune damage in mice. METHODS: Kunming mice were randomly divided into five groups: normal group (NC), irradiation control group (IC), RTP1 low dose (200 mg/kg), middle dose (400 mg/kg) and high dose (800 mg/kg) groups. RTP1 was administered by the gastric route for 14 d, mice in the NC and IC groups being given by 0.9% sodium chloride solution in the same way. The mice in all groups except NC group were irradiated with 2.0 Gy6°Co γ-ray on the fourteenth day. Immune indives of non-specific immune function, cellular immunity and humoral immunity were assessed at the 24th hour after radiation. RESULTS: Compared with the IC group, the spleen index, thymus index, rate of carbon clearance, phagocytic function of macrophages, lymphocyte proliferation, hemolysin value of blood serum and NK activity were increased markedly (P < 0.05 or P < 0.05). CONCLUSION: RTP1 has an obvious protective effects on damage in γ-ray radiated mice.

Assessment of biomedical and pharmacological activities of sea anemones Stichodactyla mertensii and Stichodactyla gigantea from Gulf of Mannar Biosphere Reserve, southeast coast of India

Cnidarians comprise an old and diverse animal phylum, and possess a wide variety of biologically active substances. Sea anemones contain a diversity of interesting biologically active compounds including some potent toxins. In the present work, the sea anemones Stichodactyla mertensii and Stichodactyla gigantea, collected from the Mandapam coast, are characterized biomedically and pharmacologically. The crude protein was obtained by using methanol and aqueous extracts. The respective protein contents of S. mertensii and S. gigantea were found to be 2.10 µg/mL and 1.87 µg/mL. The methanol and aqueous extracts of S. mertensii and S. gigantea yielded six and nine bands by SDS-PAGE on 12 percent gel. In the hemolytic assay, both extracts exhibited hemolytic effect on chicken, goat, cow and human erythrocytes ('A', 'B' and 'O'). The neurotoxic effects of these crude extracts were determined in vivo using the sea shore crab Ocypode macrocera and mortality was observed. The mouse bioassay for lethality was performed on male albino mice. The crude extract of S. mertensii showed higher lethality (58 seconds at 1 mL-dose) than that of S. gigantea (2 minutes and 10 seconds at 0.75 mL-dose). The analgesic activity test was also carried out on albino mice by Eddy's hot plate and tail-flick methods. The extracts showed moderate analgesic effect by both hot-plate and tail-flick methods. These characteristics emphasize the need for the isolation and molecular characterization of new active toxins in S. mertensii and S. gigantea.(AU)

5- and 4'-Hydroxylated flavonoids affect voltage gating of single alpha-hemolysin pore.

Molecular mechanisms of the influence of flavonoids on the voltage gating of a single alpha-hemolysin channel in planar lipid membranes are studied. It is shown that the addition of flavonoids hydroxylated in position 5 of the A-ring and in position 4' of the B-ring into bilayer bathing solution shifts the voltage dependence of channel switching from high- to low-conductance states to voltages nearer zero. It is concluded that the effect is likely to be attributed to a specific interaction of at least three flavonoid molecules with the voltage sensor of an alpha-hemolysin pore. Possible flavonoid binding sites and identification of amino acid residues included into the voltage sensor domain of the alpha-hemolysin channel are discussed.