Purification of PaTx-II from the Venom of the Australian King Brown Snake and Characterization of Its Antimicrobial and Wound Healing Activities.

Infections caused by multi-drug-resistant (MDR) bacteria are a global threat to human health. As venoms are the source of biochemically diverse bioactive proteins and peptides, we investigated the antimicrobial activity and murine skin infection model-based wound healing efficacy of a 13 kDa protein. The active component PaTx-II was isolated from the venom of Pseudechis australis (Australian King Brown or Mulga Snake). PaTx-II inhibited the growth of Gram-positive bacteria in vitro, with moderate potency (MICs of 25 µM) observed against S. aureus, E. aerogenes, and P. vulgaris. The antibiotic activity of PaTx-II was associated with the disruption of membrane integrity, pore formation, and lysis of bacterial cells, as evidenced by scanning and transmission microscopy. However, these effects were not observed with mammalian cells, and PaTx-II exhibited minimal cytotoxicity (CC50 > 1000 µM) toward skin/lung cells. Antimicrobial efficacy was then determined using a murine model of S. aureus skin infection. Topical application of PaTx-II (0.5 mg/kg) cleared S. aureus with concomitant increased vascularization and re-epithelialization, promoting wound healing. As small proteins and peptides can possess immunomodulatory effects to enhance microbial clearance, cytokines and collagen from the wound tissue samples were analyzed by immunoblots and immunoassays. The amounts of type I collagen in PaTx-II-treated sites were elevated compared to the vehicle controls, suggesting a potential role for collagen in facilitating the maturation of the dermal matrix during wound healing. Levels of the proinflammatory cytokines interleukin-1ß (IL-1ß), interleukin-6 (IL-6) and tumor necrosis factor-α (TNF-α), cyclooxygenase-2 (COX-2) and interleukin-10 (IL-10), factors known to promote neovascularization, were substantially reduced by PaTx-II treatment. Further studies that characterize the contributions towards efficacy imparted by in vitro antimicrobial and immunomodulatory activity with PaTx-II are warranted.

A-kinase anchor protein 12 (AKAP12) inhibits cell migration in breast cancer.

A-kinase anchor protein 12 (AKAP12) also known as Gravin and SSeCKS, is a novel potent scaffold protein for many key signaling factors, such as protein kinase C (PKC), PKA, cyclins as well as F-actin. AKAP12 expression is known to be suppressed in several human malignancies including breast, prostate, gastric and colon cancers. In this study, we evaluated the role of AKAP12 in the migration of breast cancer cells, an important cellular process in cancer progression. AKAP12 gene expression was analyzed in human breast cancer tissues using the Gene expression-based Outcome for Breast cancer Online (GOBO) database and TissueScan array, followed by relapse free survival (RFS) analysis with the Kaplan-Meier Plotter. AKAP12 protein was then analyzed in normal MCF10A breast cell line and six different breast cancer cell lines (AU565, Hs578T, MCF7, MDA-MB-231, T47D and ZR751). After which, siRNA-mediated knockdown of AKAP12 was carried out in MCF10A, MDA-MB-231 and Hs578T cells, followed by phenotypic assays. AKAP12 was observed to be reduced in breast cancer tissues as analyzed by GOBO and TissueScan array. Kaplan Meier survival analysis revealed that patients with AKAP12 gene expression had a higher RFS survival. There was also decreased AKAP12 protein expression in breast cancer cell lines compared to MCF10A normal epithelial breast cell line. Knockdown of AKAP12 in both MCF10A cells and Hs578T cells induced cell migration but did not alter cell proliferation. Moreover, siAKAP12 in aggressive MDA-MB-231 breast cancer cells led to an increase in cell migration. Immunofluorescence analysis of AKAP12 depleted MCF10A cells also revealed formation of thick stress fibers which could affect cell migration. Hence, the findings in this study suggest that AKAP12 is a potential metastasis suppressor in breast cancer.

Diagnosis and biomarkers for ocular tuberculosis: From the present into the future.

Tuberculosis is an airborne disease caused by Mycobacterium tuberculosis (Mtb) and can manifest both pulmonary and extrapulmonary disease, including ocular tuberculosis (OTB). Accurate diagnosis and swift optimal treatment initiation for OTB is faced by many challenges combined with the lack of standardized treatment regimens this results in uncertain OTB outcomes. The purpose of this study is to summarize existing diagnostic approaches and recently discovered biomarkers that may contribute to establishing OTB diagnosis, choice of anti-tubercular therapy (ATT) regimen, and treatment monitoring. The keywords ocular tuberculosis, tuberculosis, Mycobacterium, biomarkers, molecular diagnosis, multi-omics, proteomics, genomics, transcriptomics, metabolomics, T-lymphocytes profiling were searched on PubMed and MEDLINE databases. Articles and books published with at least one of the keywords were included and screened for relevance. There was no time limit for study inclusion. More emphasis was placed on recent publications that contributed new information about the pathogenesis, diagnosis, or treatment of OTB. We excluded abstracts and articles that were not written in the English language. References cited within the identified articles were used to further supplement the search. We found 10 studies evaluating the sensitivity and specificity of interferon-gamma release assay (IGRA), and 6 studies evaluating that of tuberculin skin test (TST) in OTB patients. IGRA (Sp = 71-100%, Se = 36-100%) achieves overall better sensitivity and specificity than TST (Sp = 51.1-85.7%; Se = 70.9-98.5%). For nuclear acid amplification tests (NAAT), we found 7 studies on uniplex polymerase chain reaction (PCR) with different Mtb targets, 7 studies on DNA-based multiplex PCR, 1 study on mRNA-based multiplex PCR, 4 studies on loop-mediated isothermal amplification (LAMP) assay with different Mtb targets, 3 studies on GeneXpert assay, 1 study on GeneXpert Ultra assay and 1 study for MTBDRplus assay for OTB. Specificity is overall improved but sensitivity is highly variable for NAATs (excluding uniplex PCR, Sp = 50-100%; Se = 10.5-98%) as compared to IGRA. We also found 3 transcriptomic studies, 6 proteomic studies, 2 studies on stimulation assays, 1 study on intraocular protein analysis and 1 study on T-lymphocyte profiling in OTB patients. All except 1 study evaluated novel, previously undiscovered biomarkers. Only 1 study has been externally validated by a large independent cohort. Future theranostic marker discovery by a multi-omics approach is essential to deepen pathophysiological understanding of OTB. Combined these might result in swift, optimal and personalized treatment regimens to modulate the heterogeneous mechanisms of OTB. Eventually, these studies could improve the current cumbersome diagnosis and management of OTB.

Excessive neutrophils and neutrophil extracellular traps contribute to acute lung injury of influenza pneumonitis.

Complications of acute respiratory distress syndrome (ARDS) are common among critically ill patients infected with highly pathogenic influenza viruses. Macrophages and neutrophils constitute the majority of cells recruited into infected lungs, and are associated with immunopathology in influenza pneumonia. We examined pathological manifestations in models of macrophage- or neutrophil-depleted mice challenged with sublethal doses of influenza A virus H1N1 strain PR8. Infected mice depleted of macrophages displayed excessive neutrophilic infiltration, alveolar damage, and increased viral load, later progressing into ARDS-like pathological signs with diffuse alveolar damage, pulmonary edema, hemorrhage, and hypoxemia. In contrast, neutrophil-depleted animals showed mild pathology in lungs. The brochoalveolar lavage fluid of infected macrophage-depleted mice exhibited elevated protein content, T1-α, thrombomodulin, matrix metalloproteinase-9, and myeloperoxidase activities indicating augmented alveolar-capillary damage, compared to neutrophil-depleted animals. We provide evidence for the formation of neutrophil extracellular traps (NETs), entangled with alveoli in areas of tissue injury, suggesting their potential link with lung damage. When co-incubated with infected alveolar epithelial cells in vitro, neutrophils from infected lungs strongly induced NETs generation, and augmented endothelial damage. NETs induction was abrogated by anti-myeloperoxidase antibody and an inhibitor of superoxide dismutase, thus implying that NETs generation is induced by redox enzymes in influenza pneumonia. These findings support the pathogenic effects of excessive neutrophils in acute lung injury of influenza pneumonia by instigating alveolar-capillary damage.

Viper metalloproteinase (Agkistrodon halys pallas) with antimicrobial activity against multi-drug resistant human pathogens.

Metalloproteinases are abundant enzymes in crotalidae and viperidae snake venoms. Snake venom metalloproteinases (SVMPs) comprise a family of zinc-dependent enzymes, which display many different biological activities. A 23.1 kDa protein was isolated from Agkistrodon halys (pallas, Chinese viper) snake venom. The toxin is a single chain polypeptide with a molecular weight of 23146.61 and an N-terminal sequence (MIQVLLVTICLAVFPYQGSSIILES) relatively similar to that of other metalloprotein-like proteases isolated from the snake venoms of the Viperidae family. The antibacterial effect of Agkistrodon halys metalloproteinase (AHM) on Burkholderia pseudomallei (strains TES and KHW), Escherichia coli, Enterobacter aerogenes, Proteus vulgaris, Proteus mirabilis, Pseudomonas aeruginosa (Gram-negative bacteria) and Staphylococcus aureus (Gram-positive bacterium) was studied at a concentration 120 microM. Interestingly, we found that the metalloproteinase exhibited antibacterial properties and was more active against S. aureus, P. vulgaris, P. mirabilis and multi-drug resistant B. pseudomallei (strain KHW) bacteria. AHM variants with high bacteriostatic activity (MIC 1.875-60 microM) also tended to be less cytotoxic against U-937 human monocytic cells up to 1 mM concentrations. These results suggest that this metalloprotein exerts its antimicrobial effect by altering membrane packing and inhibiting mechanosensitive targets.

Structure, self-assembly, and dual role of a beta-defensin-like peptide from the Chinese soft-shelled turtle eggshell matrix.

Biomineral matrix formation and molecular recognition are two important processes associated with eggshell biomineralization. To understand these two processes, a major intracrystalline peptide, pelovaterin, was isolated from turtle (Pelodiscus sinensis) eggshell and its tertiary and quaternary structures were established. The global fold of pelovaterin is similar to that of human beta-defensins but has a large hydrophobic core and a short hydrophilic N-terminal segment, which is not preserved in defensins. Pelovaterin exhibits strong antimicrobial activity against two pathogenic gram-negative bacteria, Pseudomonas aeruginosa and Proteus vulgaris, and stabilizes a thin film of metastable vaterite. We show that pelovaterin self-aggregates in the form of micellar nanospheres and the aggregation in solution is entropy-driven. It is suggested that the micellar aggregation of pelovaterin is responsible for the induction and stabilization of the metastable phase by altering the interfacial energy. The results demonstrate the adaptability of an extracellular matrix protein to perform multiple tasks: polymorph discrimination and protection of the contents of the egg against bacterial invasion.

Ethnobotanical survey of folk plants for the treatment of snakebites in Southern part of Tamilnadu, India.

Ethnobotanical surveys were conducted in four different indigenous groups in Southern parts of Tamilnadu, India, using a questionnaire. The herbal practitioners in the study area were interviewed, and information on medicinal plants was collected from the traditional healers called "Vaidyars". This survey covers 72 medicinal plants belonging to 53 families that are used for the treatment of snakebite in a traditional way. Traditional approach was evaluated scientifically with some selected plant extracts (7.2 mg/kg bw) and partially purified fractions (2.4 mg/kg bw) were orally administered to mice experimentally envenomed with rattlesnake venom s.c. injection (2.5-15 microg/kg bw). Tested fractions (Aristolochia indica, Hemidesmus indicus, Gloriosa superba, Strychnos nux-vomica, Eclipta prostrata, and Andrographis paniculata) showed potent neutralizing effect against the venom. Compared to the extracts, administration of purified fractions was more effective in increasing the body weight. Control mice injected with the venom alone showed weight loss and severe toxicity at 15 microg/kg bw. The purified fractions (2.4 mg/kg bw) produced significant protection against venom induced changes in serum SOD and LPx levels. The isolated fractions effectively inhibited the toxic effect of snake venoms in vitro than in vivo. The above observations confirmed the protective activity of plants-Aristolochia indica, Hemidesmus indicus, Gloriosa superba, Strychnos nux-vomica, Eclipta prostrata, and Andrographis paniculata against the lethal action of snake venom and need further investigation.

Identification of natural peptides as a new class of antimalarial drugs by in silico approaches.

Malaria is one of the most widespread and serious parasitic diseases worldwide. Currently available antimalarial drugs have side effects, and many strains of Plasmodia have developed resistance to such drugs. The present review examines the use of annexins and of natural peptides from snake venom as a new class of anti-malarial agents, with the key property of reducing inflammation. Severe cases of malaria manifest elevated serum levels of liver enzymes, inflammation, fibrin deposition, apoptosis, and reduction in peripheral CD8+ T cells. The annexin-A1/5 proteins trigger inflammation via increased expression of diverse cytokines (tumor necrosis factor alpha, interleukin-1 beta, interleukin-10), however, by shielding microbial phospholipids they prevent injury via damage-associated molecular patterns (DAMPs). Here, we also review an in silico-based bioengineering approach that may allow for a better design, synthesis and characterization of novel peptides from snake venom as a more effective approach to treatment due to their improved antimalarial activity.

Anti-tumor promoting potential of luteolin against 7,12-dimethylbenz(a)anthracene-induced mammary tumors in rats.

In this study, we evaluated the anti-tumor potential of luteolin (30mg/kg, p.o.), combined with cyclophosphamide (10mg/kg, i.p.) (LU+CYC) orally administered for 20 days; and CYC individually for 10 days against 7,12-dimethylbenz(a)anthracene (DMBA)-induced mammary carcinogenesis in Wistar rats. Combination treatment (LU+CYC) inhibited the incidence rate of tumors and decreased tumor volume significantly without changing the total body weight of the animals. Long-term treatment did not show any apparent toxicity in rats. The CYC-treated group showed potential reduction of tumor volume (74%), severe toxicity, and loss of body weight. In order to elucidate the anticancer mechanism of luteolin, antioxidant activities such as superoxide dismutase (SOD), catalase (CAT), and glutathione peroxidase (GPx) generation in the liver, kidney and breast, as well as protein profiles, were also examined. Biochemical analysis of the combination-treated group showed significant (P<0.01; P<0.05) inhibition of lipid peroxide (LPx) formation (oxygen-free radicals), the level and the activity of SOD, CAT and GPx were found to be very high than the LU and CYC individually treated rats at a 30mg/kg dose. 2D gel electrophoresis analysis revealed that (56kDa) high molecular weight protein was detected in tumors of rats receiving combination treatment than the cancer controls. The biological significance of that protein involved for the dysfunction of cancer cells and induces apoptosis. Histopathological changes also confirmed the formation of tumor tubules and neovascularization after the treatment. Overall, these results suggest that the combination treatment provided antioxidant defense with strong chemopreventive activity against the genesis of DMBA-induced mammary tumors.

Purification of antibacterial agents from Tragia involucrata--a popular tribal medicine for wound healing.

Tragia involucrata has been widely used in traditional systems of medicine for a variety of diseases. In the present study, in vitro antibacterial properties of nine different compounds including vinyl hexylether, shellsol, 2,4-dimethyl hexane, 2-methylnonane and 2,6-dimethyl heptane were isolated from the leaf of Tragia involucrata studied against Escherichia coli, Proteus vulgaris and Staphylococcus aureus using the disc-diffusion method at 50 microg/ml concentrations. The compound vinyl hexylether showed a broad spectrum of activity. The highest activity was found in shellsol (50 microg/ml) against Proteusvulgaris and Staphylococcus aureus. Minimum inhibitory concentrations were determined for the effective compounds (MICs 2.5-40 microg/ml), shellsol and vinyl hexylether showed inhibitory action at the lowest dilution (10 microg/ml) than 2-methylnanone. Shellsol inhibited the growth of Staphylococcus aureus very effectively than the other compounds. These compounds showed bactericidal effects against all the tested bacteria (MBC, 12.25 microg/ml). However, the compound shellsol showed effective killing of wound causing bacteria (Staphylococcus aureus). So, the study was focused on the constituent to evaluate wound healing in rat model. Rats that received 50 microg/kg, b.w. of shellsol showed complete healing after 24 days. Histological examination revealed an increase in the fibroblast, neovascularization, granulation and thickness of scar tissue after the treatment of shellsol as compared to control. The topical application of shellsol did not cause any toxic response on rat skin. Thus, the antibacterial properties of the constituents give some scientific basis to its usage in traditional medicine.

A brief update on potential molecular mechanisms underlying antimicrobial and wound-healing potency of snake venom molecules.

Infectious diseases remain a significant cause of morbidity and mortality worldwide. A wide range of diverse, novel classes of natural antibiotics have been isolated from different snake species in the recent past. Snake venoms contain diverse groups of proteins with potent antibacterial activity against a wide range of human pathogens. Some snake venom molecules are pharmacologically attractive, as they possess promising broad-spectrum antibacterial activities. Furthermore, snake venom proteins (SVPs)/peptides also bind to integrins with high affinity, thereby inhibiting cell adhesion and accelerating wound healing in animal models. Thus, SVPs are a potential alternative to chemical antibiotics. The mode of action for many antibacterial peptides involves pore formation and disruption of the plasma membrane. This activity often includes modulation of nuclear factor kappa B (NF-κB) activation during skin wound healing. The NF-κB pathway negatively regulates the transforming growth factor (TGF)-ß1/Smad pathway by inducing the expression of Smad7 and eventually reducing in vivo collagen production at the wound sites. In this context, SVPs that regulate the NF-κB signaling pathway may serve as potential targets for drug development.

ANXA1 inhibits miRNA-196a in a negative feedback loop through NF-kB and c-Myc to reduce breast cancer proliferation.

MiRNAs are endogenous ~22 nt RNAs which play critical regulatory roles in a wide range of biological and pathological processes, which can act as oncogenes or tumor suppressor genes depending on their target genes. We have recently shown that ANXA1 inhibits the expression of miRNAs including miR196a. Here, we show that miR196a was highly expressed in ER+ MCF-7 breast cancer cells when compared to normal mammary gland cells, with expression levels negatively correlating to ANXA1. ANXA1 inhibits the biogenesis of oncogenic miR-196a by suppressing primary-miR196a indirectly through the stimulation of c-myc and NFkB expression and activity in breast cancer cells. In a negative feedback loop, miR-196a directly inhibits ANXA1 and enhances breast cancer cell proliferation in vitro. Finally, miR196a promotes breast tumor growth in vivo. This study reports a novel regulatory circuit between ANXA1, NF-kB, c-myc and miR-196a which regulates breast cancer cell proliferation and tumor growth.

Ageing and the telomere connection: An intimate relationship with inflammation.

Telomeres are the heterochromatic repeat regions at the ends of eukaryotic chromosomes, whose length is considered to be a determinant of biological ageing. Normal ageing itself is associated with telomere shortening. Here, critically short telomeres trigger senescence and eventually cell death. This shortening rate may be further increased by inflammation and oxidative stress and thus affect the ageing process. Apart from shortened or dysfunctional telomeres, cells undergoing senescence are also associated with hyperactivity of the transcription factor NF-κB and overexpression of inflammatory cytokines such as TNF-α, IL-6, and IFN-γ in circulating macrophages. Interestingly, telomerase, a reverse transcriptase that elongates telomeres, is involved in modulating NF-κB activity. Furthermore, inflammation and oxidative stress are implicated as pre-disease mechanisms for chronic diseases of ageing such as neurodegenerative diseases, cardiovascular disease, and cancer. To date, inflammation and telomere shortening have mostly been studied individually in terms of ageing and the associated disease phenotype. However, the interdependent nature of the two demands a more synergistic approach in understanding the ageing process itself and for developing new therapeutic approaches. In this review, we aim to summarize the intricate association between the various inflammatory molecules and telomeres that together contribute to the ageing process and related diseases.

New frontiers for anti-biofilm drug development.

Pathogenic microbial biofilm, a consortium of microbial cells protected by a self-produced polymer matrix, is considered a worldwide challenge due to the inherent antibiotic resistance conferred by its lifestyle. Living, as it does, in a community of microbial organisms in a clinical situation, makes it responsible for severe and dangerous cases of infection. Combating this organisation of cells usually requires high antibiotic doses for a prolonged time, and these approaches often fail, contributing to infection persistence. In addition to therapeutic limitations, biofilms can be a source of infections when they grow in medical devices. The challenge imposed by biofilms has mobilised researchers in the entire world to prospect or develop alternatives to control biofilms. In this context, this review summarises the new frontiers that could be used in clinical circumstances in order to prevent or eliminate pathogenic biofilms.

Targeting transcription factor STAT3 for cancer prevention and therapy.

Signal Transducers and Activators of Transcription (STATs) comprise an important class of transcription factors that have been implicated in a wide variety of essential cellular functions related to proliferation, survival, and angiogenesis. Among various STAT members, STAT3 is frequently overexpressed in tumor cells as well as tissue samples, and regulates the expression of numerous oncogenic genes controlling the growth and metastasis of tumor cells. The current review briefly discusses the importance of STAT3 as a potential target for cancer therapy and also provides novel insights into various classes of existing pharmacological inhibitors of this transcription factor that can be potentially developed as anti-cancer drugs.

DAMPs and influenza virus infection in ageing.

Influenza A virus (IAV) is a serious global health problem worldwide due to frequent and severe outbreaks. IAV causes significant morbidity and mortality in the elderly population, due to the ineffectiveness of the vaccine and the alteration of T cell immunity with ageing. The cellular and molecular link between ageing and virus infection is unclear and it is possible that damage associated molecular patterns (DAMPs) may play a role in the raised severity and susceptibility of virus infections in the elderly. DAMPs which are released from damaged cells following activation, injury or cell death can activate the immune response through the stimulation of the inflammasome through several types of receptors found on the plasma membrane, inside endosomes after endocytosis as well as in the cytosol. In this review, the detriment in the immune system during ageing and the links between influenza virus infection and ageing will be discussed. In addition, the role of DAMPs such as HMGB1 and S100/Annexin in ageing, and the enhanced morbidity and mortality to severe influenza infection in ageing will be highlighted.

Viperatoxin-II: A novel viper venom protein as an effective bactericidal agent.

Infections caused by methicillin-resistant Staphylococcus aureus (MRSA) have become a rising threat to public health. There is an urgent need for development of promising new therapeutic agents against drug resistant bacteria like S. aureus. This report discusses purification and characterization of proteins from Indian Russell's viper snake venom. Novel 15-kDa proteins called "Viperatoxin" (VipTx-I and VipTx-II) were extracted from the whole venom and evaluated using in vitro antimicrobial experiments. The N-terminal amino acid sequence of "Viperatoxin" showed high sequence homology to daboiatoxin isolated from the same venom and also matched phospholipase A2 (PLA2) enzymes isolated from other snake venoms. In an in vitro plate assay, VipTx-II but not VipTx-I showed strong antimicrobial effects against S. aureus and Burkholderia pseudomallei (KHW & TES), Proteus vulgaris and P. mirabilis. The VipTx-II was further tested by a broth-dilution assay at 100-3.1 µg/ml concentrations. The most potent bactericidal effect was found at the lowest dilutions (MICs of 6.25 µg/ml) against B. pseudomallei, S. aureus and P. vulgaris (MICs of 12.25 µg/ml). Electron microscopic investigation revealed that the protein-induced bactericidal potency was closely associated with pore formation and membrane damage, even at the lowest concentrations (<20 µg/ml). The toxin caused a low level of cytotoxic effects as observed in human (THP-1) cells at higher concentrations. Molecular weight determinations of VipTx-II by sodium dodecyl sulfate-polyacrylamide gel electrophoresis showed one major, along with a few minor bands. The results indicate that VipTx-II plays a significant role in bactericidal and membrane damaging effects in vitro. Non-cytotoxic properties on human cells highlight it as a promising candidate for further evaluation of antimicrobial potential in vivo.

Novel phospholipase A2 inhibitors from python serum are potent peptide antibiotics.

Antimicrobial peptides (AMPs) play a vital role in defense against resistant bacteria. In this study, eight different AMPs synthesized from Python reticulatus serum protein were tested for bactericidal activity against various Gram-positive and Gram-negative bacteria (Staphylococcus aureus, Burkholderia pseudomallei (KHW and TES strains), and Proteus vulgaris) using a disc-diffusion method (20 µg/disc). Among the tested peptides, phospholipase A2 inhibitory peptide (PIP)-18[59-76], ß-Asp65-PIP[59-67], D-Ala66-PNT.II, and D60,65E-PIP[59-67] displayed the most potent bactericidal activity against all tested pathogens in a dose-dependent manner (100-6.8 µg/ml), with a remarkable activity noted against S. aureus at 6.8 µg/ml dose within 6 h of incubation. Determination of minimum inhibitory concentrations (MICs) by a micro-broth dilution method at 100-3.125 µg/ml revealed that PIP-18[59-76], ß-Asp65-PIP[59-67] and D-Ala66-PNT.II peptides exerted a potent inhibitory effect against S. aureus and B. pseudomallei (KHW) (MICs 3.125 µg/ml), while a much less inhibitory potency (MICs 12.5 µg/ml) was noted for ß-Asp65-PIP[59-67] and D-Ala66-PNT.II peptides against B. pseudomallei (TES). Higher doses of peptides had no effect on the other two strains (i.e., Klebsiella pneumoniae and Streptococcus pneumoniae). Overall, PIP-18[59-76] possessed higher antimicrobial activity than that of chloramphenicol (CHL), ceftazidime (CF) and streptomycin (ST) (30 µg/disc). When the two most active peptides, PIP-18[59-76] and ß-Asp65-PIP[59-67], were applied topically at a 150 mg/kg dose for testing wound healing activity in a mouse model of S. aureus infection, the former accelerates faster wound healing than the latter peptide at 14 days post-treatment. The western blot data suggest that the topical application of peptides (PIP-18[59-67] and ß-Asp65-PIP[59-67]) modulates NF-kB mediated wound repair in mice with relatively little haemolytic (100-1.56 µg/ml) and cytotoxic (1000-3.125 µg/ml) effects evident on human cells in vitro.

Clostridium and bacillus binary enterotoxins: bad for the bowels, and eukaryotic being.

Some pathogenic spore-forming bacilli employ a binary protein mechanism for intoxicating the intestinal tracts of insects, animals, and humans. These Gram-positive bacteria and their toxins include Clostridium botulinum (C2 toxin), Clostridium difficile (C. difficile toxin or CDT), Clostridium perfringens (ι-toxin and binary enterotoxin, or BEC), Clostridium spiroforme (C. spiroforme toxin or CST), as well as Bacillus cereus (vegetative insecticidal protein or VIP). These gut-acting proteins form an AB complex composed of ADP-ribosyl transferase (A) and cell-binding (B) components that intoxicate cells via receptor-mediated endocytosis and endosomal trafficking. Once inside the cytosol, the A components inhibit normal cell functions by mono-ADP-ribosylation of globular actin, which induces cytoskeletal disarray and death. Important aspects of each bacterium and binary enterotoxin will be highlighted in this review, with particular focus upon the disease process involving the biochemistry and modes of action for each toxin.