Louise Smith

Infections arising from bacterial adhesion and colonization on medical device surfaces are a significant healthcare problem. Silver based antibacterial coatings have attracted a great deal of attention as a potential solution. This paper reports on the development of a silver nanoparticles based antibacterial surface that can be applied to any type of material surface. The silver nanoparticles were surface engineered with a monolayer of 2-mercaptosuccinic acid, which facilitates the immobilization of the nanoparticles to the solid surface, and also reduces the rate of oxidation of the nanoparticles, extending the lifetime of the coatings. The coatings had excellent antibacterial efficacy against three clinically significant pathogenic bacteria i.e. Staphylococcus epidermidis, Staphylococcus aureus and Pseudomonas aeruginosa. Studies with primary human fibroblast cells showed that the coatings had no cytotoxicity in vitro. Innate immune studies in cultures of primary macrophages demonstrated that the coatings do not significantly alter the level of expression of pro-inflammatory cytokines or the adhesion and viability of these cells. Collectively, these coatings have an optimal combination of properties that make them attractive for deposition on medical device surfaces such as wound dressings, catheters and implants.

ABSTRACT Poly (N-vinylpyrrolidinone) has many uses in biotechnology and medicine. Here we review our recent work on materials based on this polymer that have applications in drug release or as potential mitogens. The synthesis of crosslinked PNVP hydrogels and highly branched polymers is covered.

ABSTRACT Aim:  The purpose of this study was to assess the effectiveness of combining freshly excised oral mucosa tissue with different biological and synthetic scaffolds for production of graft material for one-stage soft-tissue reconstruction. Material and Methods:  Small oral mucosa biopsies were finely minced and transplanted onto a larger circular subcutaneous wound bed in rats. Test groups included explants seeded onto (1) an electrospun scaffold, (2) acellular dermis, and (3) directly on the muscle fascia. The soft-tissue implants were examined using histology and immunohistochemical analysis for cytokeratins (CKs) 5, 10, 13, and 19. Results:  Mucosal explants grew into fully developed epithelium with keratinisation. Surprisingly, instead of spreading over the surface of the membranes, the epithelial explants curled up and formed cyst-like structures in all test groups. The epithelial layer of the cysts originated from mucosal explants grown on the electrospun scaffold showed weak positive staining for CK5, CK10, and CK19 similar to native rat oral mucosa. Other groups showed negative staining for the CKs. Conclusions:  Subcutaneous transplantation of freshly excised oral mucosa explants resulted in the growth of the explants and formation of inclusion cyst-like structures. The phenotype of the cyst epithelium was affected by the underlying connective tissue substrate.

Optical coherence tomography (OCT) is an imaging technique based on the low coherence interferometry, in which signals are obtained based on the coherent addition of the back reflected light from the sample. Applying computational methods and automated algorithms towards the classification of OCT images allows a further step towards enhancing the clinical applications of OCT. One attempt towards classification could

Human keratinocytes are difficult to isolate and have a limited lifespan. Traditionally, immortalised keratinocyte cell lines are used in vitro due to their ability to bypass senescence and survive indefinitely. However these cells do not fully retain their ability to differentiate in vitro and they are unable to form a normal stratum corneum in organotypic culture. Here we aimed to generate a pool of phenotypically similar keratinocytes from human donors that could be used in monolayer culture, without a fibroblast feeder layer, and in 3D human skin equivalent models. Primary human neonatal epidermal keratinocytes (HEKn) were cultured in low calcium, (0.07mM) media, +/-10μM Y-27632 ROCK inhibitor (HEKn-CaY). mRNA and protein was extracted and expression of differentiation markers Keratin 14 (K14), Keratin 10 (K10) and Involucrin (Inv) assessed by qRT-PCR and Western blotting. The differentiation potential of the HEKn-CaY cultures was assessed by increasing calcium levels and removing the Y-27632 for 72hrs prior to assessment of K14, K10 and Inv. The ability of the HEKn-CaY, to form a stratified epithelium was assessed using a human skin equivalent (HSE) model in the absence of Y-27632. Increased proliferative capacity, expansion potential and lifespan of HEKn was observed with the combination of low calcium and 10μM ROCK inhibitor Y-27632. The removal of Y-27632 and the addition of high calcium to induce differentiation allowed the cells to behave as primary keratinocytes even after extended serial passaging. Prolonged lifespan HEK-CaYs were capable of forming an organised stratified epidermis in 3D HSE cultures, demonstrating their ability to fully stratify and retain their original, primary characteristics. In conclusion, the use of 0.07mM Calcium and 10μM Y-27632 in HEKn monocultures provides the opportunity to culture primary human keratinocytes without a cell feeder layer for extended periods of culture whilst retaining their ability to differentiate and form a stratified epithelium.

Infections arising from bacterial adhesion and colonization on medical device surfaces are a significant healthcare problem. Silver based antibacterial coatings have attracted a great deal of attention as a potential solution. This paper reports on the development of a silver nanoparticles based antibacterial surface that can be applied to any type of material surface. The silver nanoparticles were surface engineered with a monolayer of 2-mercaptosuccinic acid, which facilitates the immobilization of the nanoparticles to the solid surface, and also reduces the rate of oxidation of the nanoparticles, extending the lifetime of the coatings. The coatings had excellent antibacterial efficacy against three clinically significant pathogenic bacteria i.e. Staphylococcus epidermidis, Staphylococcus aureus and Pseudomonas aeruginosa. Studies with primary human fibroblast cells showed that the coatings had no cytotoxicity in vitro. Innate immune studies in cultures of primary macrophages demonstrated that the coatings do not significantly alter the level of expression of pro-inflammatory cytokines or the adhesion and viability of these cells. Collectively, these coatings have an optimal combination of properties that make them attractive for deposition on medical device surfaces such as wound dressings, catheters and implants.

Background: This review focuses on looking at recent developments in the non-invasive imaging of skin, in particular at how such imaging may be used at present or in the future to detect cutaneous melanoma.

Methods: A MEDLINE search was performed for papers using imaging techniques to evaluate cutaneous melanoma, including melanoma metastasis.

Results: Nine different techniques were found: dermoscopy, confocal laser scanning microscopy (including multiphoton microscopy), optical coherence tomography, high frequency ultrasound, positron emission tomography, magnetic resonance imaging, and Fourier, Raman, and photoacoustic spectroscopies. This review contrasts the effectiveness of these techniques when seeking to image melanomas in skin.

Conclusions: Despite the variety of techniques available for detecting melanoma, there remains a critical need for a high-resolution technique to answer the question of whether tumours have invaded through the basement membrane.

There is an increasing need for a robust, simple to use, non-invasive imaging technology to follow tissue-engineered constructs as they develop. Our aim was to evaluate the use of swept-source optical coherence tomography (SS-OCT) to image tissue-engineered skin as it developed over several weeks. Tissue-engineered skin was produced using both de-epithelialized acellular dermis (DED) and amorphous collagen gels. In both cases the epidermis could be readily distinguished from the neodermis, based on a comparison with standard destructive histology of samples. Constructs produced with DED showed more epidermal/dermal maturation than those produced using collagen. The development of tissue-engineered skin based on DED was accurately monitored with SS-OCT over 3 weeks and confirmed with conventional histology.

Poly(N-vinylpyrrolidinone) (PNVP) has been used in various biomedical applications for many years. This study explores two PNVP hydrogels for their biocompatibility with skin cells and their ability to support the growth of skin cells in direct and indirect contact with the cells. Two crosslinked PNVP's were investigated, one crosslinked with ethylene glycol dimethacrylate (EGDMA) and the other crosslinked with diethylene glycol bisallylcarbonate (DEGBAC). The different crosslinkers lead to hydrogels with different mechanical and slightly different biological properties. While neither hydrogel proved to be a suitable substrate for culturing cells (based on fibroblasts and a range of other cells), indirect contact with both showed them to be biocompatible and even stimulatory to fibroblasts. The P(NVP-co-DEGBAC) hydrogel stimulated fibroblast viability more reliably than the P(NVP-co-EGDMA) hydrogel when in indirect contact with cells. This effect was shown to be independent of the presence of foetal calf serum in the culture media, and could not be explained by any hydrogel breakdown products during the course of these experiments. Rather the phenomenon was observed to be the result of a dynamic interaction between the hydrogels and the cells.