Anti-inflammatory activity of Punica granatum L. (Pomegranate) rind extracts applied topically to ex vivo skin

Coadministered pomegranate rind extract (PRE) and zinc (II) produces a potent virucidal activity against Herpes simplex virus (HSV); however, HSV infections are also associated with localised inflammation and pain. Here, the objective was to determine the anti-inflammatory activity and relative depth penetration of PRE, total pomegranate tannins (TPT) and zinc (II) in skin, ex vivo. PRE, TPT and ZnSO4 were dosed onto freshly excised ex vivo porcine skin mounted in Franz diffusion cells and analysed for COX-2, as a marker for modulation of the arachidonic acid inflammation pathway, by Western blotting and immunohistochemistry. Tape stripping was carried out to construct relative depth profiles. Topical application of PRE to ex vivo skin downregulated expression of COX-2, which was significant after just 6 h, and maintained for up to 24 h. This was achieved with intact stratum corneum, proving that punicalagin penetrated skin, further supported by the depth profiling data. When PRE and ZnSO4 were applied together, statistically equal downregulation of COX-2 was observed when compared to the application of PRE alone; no effect followed the application of ZnSO4 alone. TPT downregulated COX-2 less than PRE, indicating that tannins alone may not be entirely responsible for the anti-inflammatory activity of PRE. Punicalagin was found throughout the skin, in particular the lower regions, indicating appendageal delivery as a significant route to the viable epidermis. Topical application of TPT and PRE had significant anti-inflammatory effects in ex vivo skin, confirming that PRE penetrates the skin and modulates COX-2 regulation in the viable epidermis. Pomegranates have potential as a novel approach in ameliorating the inflammation and pain associated with a range of skin conditions, including cold sores and herpetic stromal keratitis

Bacillus subtilis vegetative isolate surviving chlorine dioxide exposure: an elusive mechanism of resistance

Aims Oxidizing agents such as chlorine dioxide are widely used microbicides, including for disinfection of medical equipment. We isolated a Bacillus subtilis isolate from a washer-disinfector whose vegetative form demonstrated unique resistance to chlorine dioxide (0·03%) and hydrogen peroxide (7·5%). The aim of this study was to understand the mechanisms of resistance expressed by this isolate. Methods and Results A range of resistance mechanisms were investigated in the B. subtilis isolate and a reference B. subtilis strain (ATCC 6051) to include bacterial cell aggregation, the presence of profuse exopolysaccharide (EPS), and the expression of detoxification enzymes. The basis of resistance of the isolate to high concentrations of oxidizing agents was not linked to the presence of endospores. Although, the presence of EPS, aggregation and expression of detoxification enzymes may play a role in bacterial survival to low concentrations of chlorine dioxide, it is unlikely that the mechanisms helped tested to survive the bactericidal effect of higher oxidizer concentrations. Conclusions Overall, the mechanisms conferring resistance to chlorine dioxide and hydrogen peroxide remains elusive. Based on recent advances in the mode of action of oxidizing agents and notably hydrogen peroxide, we postulate that additional efficient intracellular mechanisms may be involved to explain significant resistance to in-use concentrations of commonly used high-level disinfectants

Potentiated virucidal activity of pomegranate rind extract (PRE) and punicalagin against Herpes simplex virus (HSV) when co-administered with zinc (II) ions, and antiviral activity of PRE against HSV and aciclovir-resistant HSV

Background There is a clinical need for new therapeutic products against Herpes simplex virus (HSV). The pomegranate, fruit of the tree Punica granatum L, has since ancient times been linked to activity against infection. This work probed the activity of pomegranate rind extract (PRE) and co-administered zinc (II) ions. Materials and methods PRE was used in conjunction with zinc (II) salts to challenge HSV-1 and aciclovir-resistant HSV in terms of virucidal plaque assay reduction and antiviral activities in epithelial Vero host cells. Cytotoxicity was determined by the MTS assay using a commercial kit. Results Zinc sulphate, zinc citrate, zinc stearate and zinc gluconate demonstrated similar potentiated virucidal activity with PRE against HSV-1 by up to 4-fold. A generally parabolic relationship was observed when HSV-1 was challenged with PRE and varying concentrations of ZnSO4, with a maximum potentiation factor of 5.5. Punicalagin had 8-fold greater virucidal activity than an equivalent mass of PRE. However, antiviral data showed that punicalagin had significantly lower antiviral activity compared to the activity of PRE (EC50 = 0.56 μg mL-1) a value comparable to aciclovir (EC50 = 0.18 μg mL-1); however, PRE also demonstrated potency against aciclovir-resistant HSV (EC50 = 0.02 μg mL-1), whereas aciclovir showed no activity. Antiviral action of PRE was not influenced by ZnSO4. No cytotoxicity was detected with any test solution. Conclusions The potentiated virucidal activity of PRE by coadministered zinc (II) has potential as a multi-action novel topical therapeutic agent against HSV infections, such as coldsores

A Partially Purified Acinetobacter baumannii Phage Preparation Exhibits no Cytotoxicity in 3T3 Mouse Fibroblast Cells

A surge in the level and scale of antibiotic resistance has prompted renewed interest in the application of bacteriophages to treat bacterial infections. However, concerns still exist over their efficacy and safety. The cytotoxic effect of a single Acinetobacter baumannii phage was evaluated in mouse fibroblast 3T3 cells using four different assays: trypan blue; staining with Hoechst and propidium iodide; lactate dehydrogenase release; and the MTS assay. The addition of phage concentrations up to 2 x 109 pfu/mL showed little to no impact on the viability of 3T3 cells after 24h exposure using the different assays. This study demonstrates that phage BS46 is non-cytotoxic to 3T3 cells using four different assays and that appropriate quality assurance protocols for phage therapeutics are required

In vitro permeation and biological activity of punicalagin and zinc (II) across skin and mucous membranes prone to Herpes simplex virus infection

Coadministration of pomegranate rind extract (PRE) and zinc (II) ions has recently been reported as a potential new topical treatment for Herpes simplex virus (HSV) infections. In the current work we examined the in vitro topical delivery of punicalagin (major phytochemical of PRE) and zinc from hydrogels across epithelial membranes that can become infected with HSV. Porcine epidermal, buccal and vaginal mucous membranes were excised and mounted in Franz diffusion cells and dosed with a simple hydrogel containing PRE and zinc sulphate (ZnSO4). The permeation of punicalagin and zinc were determined by HPLC and ICPMS respectively; punicalagin was also determined in the basal layers by reverse tape stripping. Receptor phases from the epidermal membrane experiment were also used to challenge HSV-1 in Vero host cells, and ex vivo porcine skin was used to probe COX-2 modulation. Punicalagin and zinc permeated each of the three test membranes, with significantly greater amounts of both delivered across the epidermal membrane. The amounts of punicalagin permeating the buccal and vaginal membranes were similar, although the amount of zinc permeating the vaginal membrane was comparatively very large – it is known that zinc interacts with vaginal mucosa. The punicalagin recovered by reverse tape stripping of the epidermal, buccal and vaginal membranes gave 0.47 ± 0.016, 0.45 ± 0.052 and 0.51 ± 0.048 nM cm− 2 respectively, and were statistically the same (p < 0.05). A 2.5 log reduction was achieved against HSV-1 using diffusion cell receptor phase, and COX-2 expression was reduced by 64% in ex vivo skin after 6 h. Overall, a hydrogel containing 1.25 mg mL− 1 PRE and 0.25 M ZnSO4 was able to topically deliver both the major bioactive compound within PRE and Zn (II) across all membranes and into the site specific region of Herpes simplex vesicular clusters, while maintaining potentiated virucidal and anti-inflammatory properties. This novel therapeutic system therefore has potential for the topical treatment of HSV infections

Ageing and moisture uptake in polymethyl methacrylate (PMMA) bone cements

Bone cements are extensively employed in orthopaedics for joint arthroplasty, however implant failure in the form of aseptic loosening is known to occur after long-term use. The exact mechanism causing this is not well understood, however it is thought to arise from a combination of fatigue and chemical degradation resulting from the hostile in vivo environment. In this study, two commercial bone cements were aged in an isotonic fluid at physiological temperatures and changes in moisture uptake, microstructure and mechanical and fatigue properties were studied. Initial penetration of water into the cement followed Fickian diffusion and was thought to be caused by vacancies created by leaching monomer. An increase in weight of approximately 2% was experienced after 30 days ageing and was accompanied by hydrolysis of poly(methyl methacrylate) (PMMA) in the outermost layers of the cement. This molecular change and the plasticising effect of water resulted in reduced mechanical and fatigue properties over time. Cement ageing is therefore thought to be a key contributor in the long-term failure of cemented joint replacements. The results from this study have highlighted the need to develop cements capable of withstanding long-term degradation and for more accurate test methods, which fully account for physiological ageing

Standardisation of a mental skills scale in South Africa and the United Kingdom

The contemporary science of sport and exercise psychology requires the standardisation of mental skills questionnaires to facilitate accurate assessment of and intervention for individuals and groups in various health and sport related contexts. The study presents international research findings regarding the standardisation of a Mental Skills Scale with a sample of university students (N=420) from South Africa (n=211) and the United Kingdom (n=209) respectively. Although further international and national standardisation in both English and other languages is recommended, factor and reliability analyses indicated satisfactory validity and reliability of the current English version of the scale

Reduction in exopolysaccharide viscosity as an aid to bacteriophage penetration through Pseudomonas aeruginosa biofilms

To cause an infection, bacteriophages must penetrate the alginate exopolysaccharide of Pseudomonas aeruginosa to reach the bacterial surface. Despite a lack of intrinsic motility, phage were shown to diffuse through alginate gels at alginate concentrations up to 8% (wt/vol) and to bring about a 2-log reduction in the cell numbers in 20-day-old biofilms of P. aeruginosa. The inability of alginate to act as a more effective diffusional barrier suggests that phage may cause a reduction in the viscosity of the exopolysaccharide. Samples (n = 5) of commercial alginate and purified cystic fibrosis (CF) alginate were incubated with 2 × 108 purified phage per ml for 24 h at 37°C. After incubation the samples and controls were subjected to rheological analysis with a Carrimed controlled stress rheometer. The viscosities of phage-treated samples were reduced by up to 40% compared to those of controls incubated in the absence of phage. The experiment was repeated by using phage concentrations of 1010 and 1012 phage per ml and samples taken for analysis at intervals up to 4 h. The results indicated that there was a time- and concentration-dependent reduction in viscosity of up to 40% compared to the viscosities of the controls. Commercial and purified CF alginate samples, both phage treated and untreated, were subjected to gel filtration chromatography by using Sephacryl High Resolution S-400 medium in order to obtain evidence of degradation. The results demonstrated that alginate treated with phage had a lower molecular weight than untreated alginate. The data suggest that bacteriophage migration through P. aeruginosa biofilms may be facilitated by a reduction in alginate viscosity brought about by enzymic degradation and that the source of the enzyme may be the bacterial host itself

Mechanism of sporicidal activity for the synergistic combination of peracetic acid and hydrogen peroxide

There is still great interest in controlling bacterial endospores. The use of chemical disinfectants and, notably, oxidizing agents to sterilize medical devices is increasing. With this in mind, hydrogen peroxide (H2O2) and peracetic acid (PAA) have been used in combination, but until now there has been no explanation for the observed increase in sporicidal activity. This study provides information on the mechanism of synergistic interaction of PAA and H2O2 against bacterial spores. We performed investigations of the efficacies of different combinations, including pretreatments with the two oxidizers, against wild-type spores and a range of spore mutants deficient in the spore coat or small acid-soluble spore proteins. The concentrations of the two biocides were also measured in the reaction vessels, enabling the assessment of any shift from H2O2 to PAA formation. This study confirmed the synergistic activity of the combination of H2O2 and PAA. However, we observed that the sporicidal activity of the combination is largely due to PAA and not H2O2. Furthermore, we observed that the synergistic combination was based on H2O2 compromising the spore coat, which was the main spore resistance factor, likely allowing better penetration of PAA and resulting in the increased sporicidal activity