Anti-endotoxic Effects Research Articles

Does taurolidine have any effect on liver regeneration and oxidation in the experimental hepatectomy model?

Taurolidine is a bicyclic molecule produced by the natural amino acid taurine. Antibacterial, antiendotoxic and cytoprotective effects of taurolidine have been shown experimentally. Data on the effects of taurolidine on oxidative stress and hepatic regeneration are limited. The aim of the study was to evaluate the effect of taurolidine on hepatic regeneration and oxidative stress in rats undergoing partial hepatectomy. Forty adult, male Wistar Albino rats were randomly divided into four equal groups: sham (S) group (n= 10), post-sham opera- tion taurolidine administered (ST) group (n= 10), partial hepatectomy (H) group (n= 10) and post-partial hepatectomy taurolidine administered (HT) group (n= 10). 100 mg/kg/day taurolidine was administered for seven days. Blood and liver tissue samples were collected on postoperative day seven. Liver tissue malondialdehyde, glutathione and Cu-Zn superoxide dismutase activity (SOD) were measured to assess oxidative stress. Binuclear hepato- cyte and Ki-67 antigen levels were measured to evaluate hepatic regeneration. There was no difference between the groups for malondialdehyde, Cu-Zn superoxide dismutase and glutathione levels (p> 0.05). Binuclear nuclei levels were comparable between the H and HT groups (p= 0.06), while taurolidine decreased binuclear hepatocyte levels in the sham operated groups (p= 0.02). Taurolidine application decreased Ki-67 levels after partial hepatectomy (p= 0.001). Taurolidine may cause anti-regenerative effects after partial hepatectomy without causing oxidative damage.

Anti-pseudomonal and anti-endotoxic effects of surfactin-stabilized biogenic silver nanocubes ameliorated wound repair in streptozotocin-induced diabetic mice

Pseudomonas aeruginosa and its lipopolysaccharides play a key role in the pathogenesis of diabetic foot infection, for which, currently no effective therapeutic agents are available. Hence, newer forms of therapeutic agents are required for treating Pseudomonas aeruginosa infection. In this present study, nanocrystalline silver nanoparticles (AgNPs) were synthesized using culture filtrate of Brevibacillus brevis KN8(2) followed by an investigation of its in vivo anti-pseudomonal and anti-endotoxic properties. Biosynthesized AgNPs was predominantly cubical in shape with an average particle size of 15.40 nm as observed through field emission scanning electron microscopy (FESEM) and X-ray diffraction analysis. The LC-ESI-MS/MS analysis indicates the presence of surfactin in culture filtrate of B. brevis KN8(2). The MIC of surfactin-stabilized AgNPs against P. aeruginosa was 10 μg ml−1 and its wound repair activity was observed in P. aeruginosa-infected wounds of diabetic mice by measuring wound area closure, bacterial counts, mRNA expressions, and histopathology. Further, surfactin-stabilized AgNPs suppressed the transcription of LPS-triggered expression of the TNF-α in wounds that LPS-assisted extension of wound repair in diabetes mellitus conditions was circumvented quite well. Results gathered in this study established that surfactin-stabilized AgNPs could effectively offer to the novel treatment of Gram-negative bacilli infection in diabetic wounds.

Impact of tyloxapol to combat acute phase response in comparison with betamethasone and flunixin meglumine following an ovine experimentally induced endotoxemia model

Systemic inflammatory responses to circulating lipopolysaccharide lead to high mortality rates in affected animals and thus effective treatments of this situation are crucial. However, despite different endotoxemia therapeutic regimens, the lack of an effective treatment still remains a clinical problem in sheep. Tyloxapol is a potential pro- and anti-inflammatory molecule to treat endotoxemia in farm animals. Hence, the present study was an attempt to clarify the antiendotoxic effects of tyloxapol in comparison with betamethasone and flunixin meglumine in experimentally induced endotoxemia in sheep. Thirty clinically healthy 1-year old Iranian fat-tailed ewes were randomly divided into 6 equal experimental (n=5) groups, comprising Negative and Positive control, Flunixin meglumine, Betamethasone, Tyloxapol 1 and Tyloxapol 2. Phenol extracted lipopolysaccharide from <em>Escherichia coli</em> serotype O55:B5 was infused at 2 μg/kg intravenously. Ninety min after endotoxemia induction, Flunixin meglumine (2.2 mg/kg), betamethasone (1 mg/kg) and tyloxapol (200 and 400 mg/kg) were injected to respective groups, over 60 min along with intravenous fluids. Blood samples were collected from all ewes prior and 1.5, 3, 4.5, 6, 24 and 48 hours after lipopolysaccharide injection and sera and plasmas were separated, subsequently. Haptoglobin, serum amyloid A, tumor necrosis factor-alpha, interferon-gamma, superoxide dismutase and glutathione peroxidase were measured in all samples. Serum concentrations of haptoglobin, serum amyloid A, tumor necrosis factor-alpha, interferongamma in Flunixin meglumine group were found to be lower than other experimental ones after hour 3. Serum levels of haptoglobin, serum amyloid A, tumor necrosis factor-alpha, interferon-gamma in Tyloxapol groups were significantly lower and higher than Betamethasone and Flunixin meglumine ones, respectively. Superoxide dismutase and glutathione peroxidase activities in Flunixin meglumine group were significantly higher than other experimental groups after administration of drugs. In conclusion, the efficacy of tyloxapol was significantly higher and lower than betamethasone and flunixin meglumine, respectively. Furthermore, tyloxapol doesn’t induce its effects by dose dependent manner and its anti- and pro-inflammatory effects at 200 and 400 mg/kg were statistically similar.

Attenuating the endotoxin induced acute phase response by pentoxifylline in comparison with dexamethasone and ketoprofen in sheep

Treating the endotoxemia in animals is necessary to prevent high mortality rates. Despite the suggestion of different endotoxemia therapeutic regimens, the lack of an effective treatment still remains a clinical problem in sheep. Hence, the present study was performed to clarify the antiendotoxic effects of pentoxifylline in comparison with dexamethasone and ketoprofen in experimentally induced endotoxemia in sheep. Thirty clinically healthy 1-year old Iranian fat-tailed ewes were randomly divided into 6 equal experimental (n=5) groups, comprising negative and positive control, ketoprofen (3mg/kg), dexamethasone (1mg/kg) and pentoxifylline (30 and 60mg/kg). Lipopolysaccharide from Escherichia coli serotype O55:B5 was infused at 2μg/kg intravenously. Ketoprofen, dexamethasone and pentoxifylline were injected to related groups, at 90min after endotoxemia induction over 60min along with intravenous fluids. Blood samples were collected from all ewes prior and 1.5, 3, 4.5, 6, 24 and 48h after lipopolysaccharide injection and sera and plasmas were separated, subsequently. Haptoglobin, serum amyloid A, tumor necrosis factor-alpha, interferon-gamma, superoxide dismutase and glutathione peroxidase were measured in all samples. Serum levels of haptoglobin, serum amyloid A, tumor necrosis factor-alpha, interferon-gamma in both pentoxifylline groups were significantly lower and higher than dexamethasone and ketoprofen ones, respectively. In conclusion, the efficacy of pentoxifylline was significantly higher and lower than dexamethasone and ketoprofen, respectively. Furthermore, its anti- and pro-inflammatory effects at 30 and 60mg/kg were statistically similar together.

The comparative efficacy of tyloxapol versus pentoxifylline against induced acute phase response in an ovine experimental endotoxemia model

The effective treatments of endotoxemia are necessary to prevent high mortality rates. Hence, the present study was performed to clarify the antiendotoxic effects of tyloxapol and pentoxifylline in experimentally induced endotoxemia in sheep. Thirty clinically healthy 1-year-old Iranian fat-tailed ewes were randomly divided into six equal experimental (n=5) groups, comprising Negative and Positive control, Tyloxapol 1, Tyloxapol 2, Pentoxifylline 1 and Pentoxifylline 2. Phenol extracted lipopolysaccharide from Escherichia coli serotype O55:B5 was infused at 2µg/kg intravenously. Tyloxapol (200 and 400mg/kg) and pentoxifylline (30 and 60mg/kg) were injected to Tyloxapol and Pentoxifylline groups, respectively, at 90min after endotoxemia induction over 60min along with intravenous fluids. Blood samples were collected from all ewes prior and 1.5, 3, 4.5, 6, 24 and 48h after lipopolysaccharide injection and sera and plasmas were separated, subsequently. Haptoglobin, serum amyloid A, tumor necrosis factor-alpha (TNF-α), interferon-gamma (IFN-γ), superoxide dismutase and glutathione peroxidase were measured in all samples. Serum concentrations of haptoglobin, serum amyloid A, TNF-α and IFN-γ in Tyloxapol 1 and 2 and Pentoxifylline 1 and 2 groups were significantly lower than Positive control one after hour 3. There were no significant differences among Tyloxapol and Pentoxifylline groups (P>0.05). Superoxide dismutase and glutathione peroxidase activities in Tyloxapol 1 and 2 and Pentoxifylline 1 and 2 groups were significantly lower than Positive control one after hour 3. There were no significant differences among Tyloxapol 1 and 2 and Pentoxifylline 1 and 2 groups (P>0.05). Tyloxapol and pentoxifylline act as the anti-inflammatory mediators by decreasing pro-inflammatory cytokines and hepatic APPs and modulating oxidative enzymes activity after endotoxemia induction in sheep. Furthermore, their efficacies at different doses were significantly similar together and both drugs don't induce their effects by dose dependent manner and the anti- and pro-inflammatory effects of them were statistically similar.

Anti-endotoxic and antibacterial effects of a dermal substitute coated with host defense peptides

Biomaterials used during surgery and wound treatment are of increasing importance in modern medical care. In the present study we set out to evaluate the addition of thrombin-derived host defense peptides to human acellular dermis (hAD, i.e. epiflex®). Antimicrobial activity of the functionalized hAD was demonstrated using radial diffusion and viable count assays against Gram-negative Escherichia coli, Pseudomonas aeruginosa and Gram-positive Staphylococcus aureus bacteria. Electron microscopy analyses showed that peptide-mediated bacterial killing led to reduced hAD degradation. Furthermore, peptide-functionalized hAD displayed endotoxin-binding activity in vitro, as evidenced by inhibition of NF-κB activation in human monocytic cells (THP-1 cells) and a reduction of pro-inflammatory cytokine production in whole blood in response to lipopolysaccharide stimulation. The dermal substitute retained its anti-endotoxic activity after washing, compatible with results showing that the hAD bound a significant amount of peptide. Furthermore, bacteria-induced contact activation was inhibited by peptide addition to the hAD. E. coli infected hAD, alone, or after treatment with the antiseptic substance polyhexamethylenebiguanide (PHMB), yielded NF-κB activation in THP-1 cells. The activation was abrogated by peptide addition. Thus, thrombin-derived HDPs should be of interest in the further development of new biomaterials with combined antimicrobial and anti-endotoxic functions for use in surgery and wound treatment.

Anti-inflammatory effects of four potential anti-endotoxaemic drugs assessed in vitro using equine whole blood assays.

Despite the severity and common occurrence of equine endotoxaemia, the available anti-endotoxic treatments do not effectively target key inflammatory mechanisms such as leucocyte activation and cytokine production. In this study, four compounds with potential anti-endotoxic effects, namely rolipram, azithromycin, ethyl pyruvate and metformin, were investigated in vitro using equine whole blood stimulated with bacterial lipopolysaccharide. TNF-α and IL-1β production were measured in plasma. Rolipram was the most potent inhibitor of cytokine production (IC50 0.84 and 4.68 μm for TNF-α and IL-1β, respectively) with almost complete inhibition of TNF-α, but inhibited IL-1β by only 39.46%. Azithromycin produced almost complete inhibition of both cytokines, but tended to be less potent than rolipram (IC50 10.66 and 17.4 μm for TNF-α and IL-1β, respectively). Metformin inhibited TNF-α production with similar potency to rolipram and azithromycin (IC50 3.35 μm) but showed significantly lower efficacy (45.93%; P < 0.05), and had no inhibitory effect on IL-1β. Ethyl pyruvate was the least potent (IC50 68.35 μm and >10 mm for TNF-α and IL-1β production, respectively). Further work is required to investigate whether these or related compounds may have potential use in the treatment of equine endotoxaemia in vivo.

Antimicrobial Effects of Helix D-derived Peptides of Human Antithrombin III

Antithrombin III (ATIII) is a key antiproteinase involved in blood coagulation. Previous investigations have shown that ATIII is degraded by Staphylococcus aureus V8 protease, leading to release of heparin binding fragments derived from its D helix. As heparin binding and antimicrobial activity of peptides frequently overlap, we here set out to explore possible antibacterial effects of intact and degraded ATIII. In contrast to intact ATIII, the results showed that extensive degradation of the molecule yielded fragments with antimicrobial activity. Correspondingly, the heparin-binding, helix D-derived, peptide FFFAKLNCRLYRKANKSSKLV (FFF21) of human ATIII, was found to be antimicrobial against particularly the Gram-negative bacteria Escherichia coli and Pseudomonas aeruginosa. Fluorescence microscopy and electron microscopy studies demonstrated that FFF21 binds to and permeabilizes bacterial membranes. Analogously, FFF21 was found to induce membrane leakage of model anionic liposomes. In vivo, FFF21 significantly reduced P. aeruginosa infection in mice. Additionally, FFF21 displayed anti-endotoxic effects in vitro. Taken together, our results suggest novel roles for ATIII-derived peptide fragments in host defense.

Galleria mellonella native and analogue peptides Gm1 and ΔGm1. II) Anti-bacterial and anti-endotoxic effects

Antimicrobial peptides (AMPs) are important components of the innate immune system of animals, plants, fungi and bacteria and are recently under discussion as promising alternatives to conventional antibiotics. We have investigated two cecropin-like synthetic peptides, Gm1, which corresponds to the natural overall uncharged Galleria mellonella native peptide and ΔGm1, a modified overall positively charged Gm1 variant. We have analysed these peptides for their potential to inhibit the endotoxin-induced secretion of tumour necrosis factor-α (TNF-α) from human mononuclear cells. Furthermore, in a conventional microbiological assay, the ability of these peptides to inhibit the growth of the rough mutant bacteria Salmonella enterica Minnesota R60 and the polymyxin B-resistant Proteus mirabilis R45 was investigated and atomic force microscopy (AFM) measurements were performed to characterize the morphology of the bacteria treated by the two peptides. We have also studied their cytotoxic properties in a haemolysis assay to clarify potential toxic effects.Our data revealed for both peptides minor anti-inflammatory (anti-endotoxin) activity, but demonstrated antimicrobial activity with differences depending on the endotoxin composition of the respective bacteria. In accordance with the antimicrobial assay, AFM data revealed a stronger morphology change of the R45 bacteria than for the R60. Furthermore, Gm1 had a stronger effect on the bacteria than ΔGm1, leading to a different morphology regarding indentations and coalescing of bacterial structures. The findings verify the biophysical measurements with the peptides on model systems. Both peptides lack any haemolytic activity up to an amount of 100μg/ml, making them suitable as new anti-infective agents.

Importance of lipopolysaccharide aggregate disruption for the anti-endotoxic effects of heparin cofactor II peptides

Lipid membrane and lipopolysaccharide (LPS) interactions were investigated for a series of amphiphilic and cationic peptides derived from human heparin cofactor II (HCII), using dual polarization interferometry, ellipsometry, circular dichroism (CD), cryoTEM, and z-potential measurements. Antimicrobial effects of these peptides were compared to their ability to disorder bacterial lipid membranes, while their capacity to block endotoxic effects of LPS was correlated to the binding of these peptides to LPS and its lipid A moiety, and to charge, secondary structure, and morphology of peptide/LPS complexes. While the peptide KYE28 (KYEITTIHNLFRKLTHRLFRRNFGYTLR) displayed potent antimicrobial and anti-endotoxic effects, its truncated variants KYE21 (KYEITTIHNLFRKLTHRLFRR) and NLF20 (NLFRKLTHRLFRRNFGYTLR) provide some clues on structure–activity relations, since KYE21 retains both the antimicrobial and anti-endotoxic effects of KYE28 (although both attenuated), while NLF20 retains the antimicrobial but only a fraction of the anti-endotoxic effect, hence locating the anti-endotoxic effects of KYE28 to its N-terminus. The antimicrobial effect, on the other hand, is primarily located at the C-terminus of KYE28. While displaying quite different endotoxic effects, these peptides bind to a similar extent to both LPS and lipid A, and also induce comparable LPS scavenging on model eukaryotic membranes. In contrast, fragmentation and densification of LPS aggregates, in turn dependent on the secondary structure in the peptide/LPS aggregates, correlate to the anti-endotoxic effect of these peptides, thus identifying peptide-induced packing transitions in LPS aggregates as key for anti-endotoxic functionality. This aspect therefore needs to be taken into account in the development of novel anti-endotoxic peptide therapeutics.

Lipopolysaccharide Interactions of C-Terminal Peptides from Human Thrombin

Interactions with bacterial lipopolysaccharide (LPS), both in aqueous solution and in lipid membranes, were investigated for a series of amphiphilic peptides derived from the C-terminal region of human thrombin, using ellipsometry, dual polarization interferometry, fluorescence spectroscopy, circular dichroism (CD), dynamic light scattering, and z-potential measurements. The ability of these peptides to block endotoxic effects caused by LPS, monitored through NO production in macrophages, was compared to peptide binding to LPS and its endotoxic component lipid A, and to size, charge, and secondary structure of peptide/LPS complexes. While the antiendotoxic peptide GKY25 (GKYGFYTHVFRLKKWIQKVIDQFGE) displayed significant binding to both LPS and lipid A, so did two control peptides with either selected D-amino acid substitutions or with maintained composition but scrambled sequence, both displaying strongly attenuated antiendotoxic effects. Hence, the extent of LPS or lipid A binding is not the sole discriminant for the antiendotoxic effect of these peptides. In contrast, helix formation in peptide/LPS complexes correlates to the antiendotoxic effect of these peptides and is potentially linked to this functionality. Preferential binding to LPS over lipid membrane was furthermore demonstrated for these peptides and preferential binding to the lipid A moiety within LPS inferred.

Innate Immune Cocktail Partially Protects Broilers Against Cellulitis and Septicemia

Antimicrobial/host defense peptides (A/HDP) are natural compounds that are found in leucocyte cells and on the skin and bodily fluids of birds, reptiles, and mammals. Not only do they possess antibacterial, antiviral, and antiparasitic characteristics but they also stimulate the host immune system to combat infectious diseases and may play a role in the promotion of wound repair. Gamma-amino butyric acid (GABA) is an amino acid-based inhibitory neurotransmitter in the brain that has also been shown to promote wound healing on skin. The objective of this study was to establish a therapeutic cocktail that protects birds against Escherichia coli-related disease and lesions in broilers. We injected a cocktail of six A/HDPs with or without GABA into 3-wk-old broilers by a subcutaneous or intramuscular route followed 24 hr later by challenge with a field isolate of serogroup O2 E. coli. Birds were examined for 5-6 days post-E. coli challenge and clinical, pathologic, and bacteriologic assessments were conducted. Birds that were subcutaneously injected with an A/HDP plus GABA cocktail had significantly higher survival rates and lower levels of bacteremia (P < 0.05), but a similar percentage of the surviving birds had large cellulitis lesions compared to the surviving phosphate-buffered saline-injected control birds. When this cocktail was administered intramuscularly, there was a trend towards protection against E. coli-related death, although the results were not statistically significant and there was no reduction in bacteremia. A significant number of birds had a reduced bacterial load on cellulitis lesions but no reduction in lesion size, which suggests that when the cocktail was administered intramuscularly it failed to protect against cellulitis. These results suggest that the route of administration of the cocktail influences disease outcome. Gene expression analysis was performed to investigate whether the cocktail induced immunomodulatory functions in avian cells that complemented their antimicrobial and anti-endotoxic effects. A/HDP plus GABA mediated temporal induction of pro-inflammatory cytokines but no induction of any of the chemokines under investigation. This cocktail shows potential to protect against E. coli-related death, which is a major economic burden to the poultry industry.

Membrane and lipopolysaccharide interactions of C-terminal peptides from S1 peptidases

The mechanisms underlying antimicrobial and anti-endotoxic effects were investigated for a series of structurally related peptides derived from the C-terminal region of S1 peptidases. For this purpose, results on bacterial killing were compared to those on peptide-induced liposome leakage, and to ellipsometry and dual polarization interferometry results on peptide binding to, and disordering of, supported lipid bilayers. Furthermore, the ability of these peptides to block endotoxic effects caused by bacterial lipopolysaccharide (LPS), monitored through NO production in macrophages, was compared to the binding of these peptides to LPS, and to secondary structure formation in the peptide/LPS complex. Bacteria killing, occurring through peptide-induced membrane lysis, was found to correlate with liposome rupture, and with the extent of peptide binding to the lipid membrane, no adsorption threshold for peptide insertion being observed. Membrane and LPS binding was found to depend on peptide net charge, illustrated by LPS binding increasing with increasing peptide charge, and peptides with net negative charge being unable to lyse membranes, kill bacteria, and block LPS-induced endotoxic effect. These effects were, however, also influenced by peptide hydrophobicity. LPS binding was furthermore demonstrated to be necessary, but not sufficient, for anti-endotoxic effect of these peptides. Circular dichroism spectroscopy showed that pronounced helix formation occurs in peptide/LPS complexes for all peptides displaying anti-endotoxic effect, hence potentially linked to this functionality. Similarly, ordered secondary structure formation was correlated to membrane binding, lysis, and antimicrobial activity of these peptides. Finally, preferential binding of these peptides to LPS over the lipid membrane was demonstrated.

Using non-steroidal anti-inflammatory drugs around calving: Maximizing comfort, productivity and fertility

Non-steroidal anti-inflammatory drugs (NSAIDs) have analgesic, anti-inflammatory, anti-endotoxic and anti-pyretic effects in cattle. As such, they could be expected to have significant effects in cows and calves in the post-calving period. This review evaluates the published data on the use of NSAIDs in the dam and its calf after dystocia, the impact of NSAIDs on uterine involution, the restoration of ovarian function and prevention and treatment of the metritis complex, and the benefits of using NSAIDs in the recumbent cow. Overall, the published data are very limited, despite frequent use of NSAIDs by veterinarians in the post-calving cow, and the small number of published studies focus on blanket treatment of calving cows rather than targeted treatment after dystocia. Blanket treatment had no economic benefit; indeed, some studies reported adverse effects, such as pyrexia and increased risk of metritis. There is even less information on the value of treating calves with NSAIDs after dystocia, despite significant tissue damage which may benefit from NSAID use. Appreciably more studies have evaluated the influence of NSAIDs on uterine and ovarian function, but clinical relevance is limited. In cows with a normal puerperium, prolonged treatment with NSAIDs may slow the restoration of normal function, but most reported studies are small and use NSAIDs more frequently and for longer periods than is common in general practice. The evidence of a clinical benefit in cows with puerperal disease is limited and equivocal, and the evidence base for the use of NSAIDs in the treatment of recumbent cows is also small, even though an expert panel concluded that NSAIDs were a key aspect of veterinary treatment of downer cows. The lack of evidence identified by this review supports the contention that NSAIDs are likely to be under-used and sub-optimally prescribed in the post calving period. Further research on the use of NSAIDs in the post-calving cow and calf is required.

C-terminal Peptides of Tissue Factor Pathway Inhibitor Are Novel Host Defense Molecules

Tissue factor pathway inhibitor (TFPI) inhibits tissue factor-induced coagulation, but may, via its C terminus, also modulate cell surface, heparin, and lipopolysaccharide interactions as well as participate in growth inhibition. Here we show that C-terminal TFPI peptide sequences are antimicrobial against the gram-negative bacteria Escherichia coli and Pseudomonas aeruginosa, gram-positive Bacillus subtilis and Staphylococcus aureus, as well as the fungi Candida albicans and Candida parapsilosis. Fluorescence studies of peptide-treated bacteria, paired with analysis of peptide effects on liposomes, showed that the peptides exerted membrane-breaking effects similar to those seen for the "classic" human antimicrobial peptide LL-37. The killing of E. coli, but not P. aeruginosa, by the C-terminal peptide GGLIKTKRKRKKQRVKIAYEEIFVKNM (GGL27), was enhanced in human plasma and largely abolished in heat-inactivated plasma, a phenomenon linked to generation of antimicrobial C3a and activation of the classic pathway of complement activation. Furthermore, GGL27 displayed anti-endotoxic effects in vitro and in vivo in a mouse model of LPS shock. Importantly, TFPI was found to be expressed in the basal layers of normal epidermis, and was markedly up-regulated in acute skin wounds as well as wound edges of chronic leg ulcers. Furthermore, C-terminal fragments of TFPI were associated with bacteria present in human chronic leg ulcers. These findings suggest a new role for TFPI in cutaneous defense against infections.

The anti‐endotoxic effects of the KSL‐W decapeptide on Escherichia coli O55:B5 and various oral lipopolysaccharides

Host responses following the recognition of bacterial lipopolysaccharide can range from acute inflammation to septic shock. The aim of this study was to evaluate the ability of the KSL-W decapeptide to bind to and block the endotoxic effects of lipopolysaccharide. An enzyme-linked immunosorbent assay-based binding assay using fluorescently labeled KSL-W to detect adsorbed Escherichia coli O55:B5 lipopolysaccharide was employed. A commercially available recombinant Factor C lipopolysaccharide detection assay, hemagglutination of rabbit erythrocytes as well as E-selectin expression in human umbilical vein endothelial cells were used to assess the anti-endotoxic effects after KSL-W exposure to E. coli lipopolysaccharide as well as to oral lipopolysaccharide samples. Lipopolysaccharide-binding assays using E. coli O55:B5 lipopolysaccharide revealed both a higher maximal binding range (532-713 microM) and a half-maximum binding concentration (70-185 microM) for the KSL-W peptide when compared with its analog control. Significant inhibition of E-selectin expression in human umbilical vein endothelial cells (p < 0.0001) as well as hemagglutination of rabbit erythrocytes occurred after the interaction of KSL-W with E. coli lipopolysaccharide. Recombinant Factor C enzyme detection inhibition revealed dose-dependent inhibition values ranging from 1.0-51.8 microM. which were dependent upon the type of lipopolysaccharide sample tested. These results demonstrate that for the concentrations tested, the KSL-W decapeptide was nontoxic to mammalian cells and could bind to and block the host recognition and response towards enteric, as well as oral, lipopolysaccharide samples.

Effect of Peritoneal Lavage with Taurolidine on Primary Colonic Anastomosis in a Rat Model of Secondary Peritonitis

The reported antibacterial, antiendotoxic, and antiadhesive effects of taurolidine prompted us to study the benefits of intraperitoneal lavage with taurolidine on primary colonic anastomosis in a rat model of secondary peritonitis. We induced peritonitis in 40 rats by injecting Escherichia coli isolate intraperitoneally. We performed colonic resection and primary anastomosis 5 h later, after lavage with either taurolidine or saline solution. After the rats were killed, on postoperative day (POD) 3 (n = 10) or 7 (n = 10), we measured the bursting pressures and hydroxyproline levels, then examined the resected specimens histologically. Bursting pressures and tissue hydroxyproline levels were significantly higher in the taurolidine group than in the control group on PODs 3 and 7 (P < 0.05). Histopathological examination revealed significantly higher fibroblastic activity in the taurolidine group. The higher bursting pressures and tissue hydroxyproline levels in the rats given taurolidine showed the positive effect of taurolidine on anastomotic strength in secondary peritonitis. Taurolidine is a novel antibiotic with both antibacterial and antiendotoxic effects. Intraperitoneal lavage with taurolidine solution may reduce the risks associated with performing primary colonic anastomosis in patients with secondary peritonitis.

Evaluation of the anti‐endotoxic effects of polymyxin‐E (colistin) in dogs with naturally occurred endotoxic shock

Endotoxin is a potent stimulator of the inflammatory response and is believed to initiate the pathology in gram-negative sepsis. Agents are being searched for that bind and neutralize or block the effects of endotoxin. The aim was to study the anti-endotoxic effects of polymyxin-E (colistin) in endotoxaemic dogs. The study included a total of 30 endotoxaemic dogs, which were divided into two groups (control = 15; test = 15) of both sexes, different breeds and ages. Hetastarch colloid solution (Expahes,10 mL/kg, i.v.) with lactated Ringer's solution (20 mL/kg, i.v., Q12 h) was given to all dogs. While ampicillin was administered (Alfasilin, 10 mg/kg, i.m., Q12 h) as an antibacterial to the control group, colistin (12,500 IU/kg, i.m., Q12 h) + ampicillin were administered to the test group. The clinical examination (body temperature, pulse and respiration rates, capillary filling times, peripheral pulse qualities, dehydration degrees), hematological and biochemical examinations (WBC, RBC, HGB, HCT, thrombocyte, serum urea, creatinine and TNF-alpha) were performed both before the treatment, and 2, 4, 12 and 24 h after the treatment. In comparison with the control group, it was observed that test group had shorter capillary filling time at 24 h (P < 0.001). Moreover, the degree of dehydration in test group, was significantly improved at 24 h (P < 0.01). While the differences in peripheral pulse qualities significantly differed between 0 and 2 h in controls, at 2, 4, 24 h after treatment it was found to be significantly increased when compared with 0 h in the test group. Serum TNF-alpha concentrations were statistically decreased in the test group between 0 h and other times (P < 0.01). When compared with controls, serum TNF-alpha concentrations were lower at 2, 4, 12 and 24 h in test group (P < 0.05). Results of the study indicated that polymyxin-E (colistin) has an anti-endotoxic effect and is safe for the dogs with endotoxemia at the dosage used in this study.

Anti-endotoxic effects of syringic acid ofRadix Isatidis

The anti-endotoxic effect of syringic acid (SA) isolated from Radix Isatidis (Banlangen, BLG) was studied. SA was extracted and isolated from BLG and diluted into 1% solution. The content of SA-pretreated endotoxin (ET) was quantitatively determined using Limulus test. The ability of fever induction of ET pretreated with SA was measured using endotoxin-induced fever test in rabbits. The LPS-induced death in mice pretreated with and without SA was compared. Results showed that after pretreatment with SA, 83.16% of ET was destroyed, the ET-induced fever in rabbits relieved markedly and the LPS-induced death rate in mice dropped from 68% to 20%. It was concluded that SA isolated from BLG had anti-endotoxic effects.

Antibacterial proteins of granulocytes differ in interaction with endotoxin. Comparison of bactericidal/permeability-increasing protein, p15s, and defensins.

Bactericidal/permeability-increasing protein (BPI), antibacterial 15-kDa protein isoforms (p15s), and defensins (neutrophil peptides or NPs) are granule-associated antibacterial proteins of polymorphonuclear leukocytes (PMN) that have both direct and synergistic growth inhibitory activity against Gram-negative bacteria. In this study, we have compared in vitro the abilities of these antibacterial proteins, alone and in combination, to inhibit the endotoxic activity of isolated LPS and whole bacteria. All three proteins blocked endotoxin activity in: 1) the Limulus amoebocyte lysate assay, 2) priming of PMN for enhanced arachidonate release, and 3) stimulating leukocyte oxidase activity in 1% blood. However, the proteins differ markedly in both relative potency (BPI >> p15s = NP1) in the presence of the plasma LPS-binding protein and in the range of LPS chemotypes that can be inhibited. BPI potently neutralizes LPS of any chemotype, but p15s and defensins are less active against long-chain (S-type) LPS. In whole blood ex vivo, the p15s and NP1 are approximately 1000-fold less potent than BPI, but at subinhibitory doses act in synergy with BPI to inhibit the TNF-inducing activity of a serum-resistant encapsulated strain of Escherichia coli (K1/r). The anti-endotoxic effects of p15 and NP1 against E. coli K1/r in whole blood appear secondary to growth arrest, because, in marked contrast to BPI, they are not evident against nonviable bacteria (pretreated with antibiotic) nor isolated LPS. Thus, BPI stands out for its ability to inhibit isolated or bacterial LPS under physiologic conditions. However, p15s and defensins may also contribute to suppression of endotoxic signaling by Gram-negative bacteria via synergistic (with BPI) growth inhibition upon extracellular release of these proteins from PMN during inflammation.