A Multimodal Spectroscopic Approach Combining Mid-infrared and Near-infrared for Discriminating Gram-positive and Gram-negative Bacteria.

The rapid and accurate identification of pathogenic bacteria is crucial for combating the growing threat of antibiotic resistance, nosocomial infections, and food safety concerns. This study presents a novel and comprehensive comparison of two vibrational spectroscopic techniques - attenuated total reflection Fourier transform infrared (ATR-FTIR) spectroscopy and a low-cost miniature near-infrared (NIR) spectrometer - for distinguishing Gram-positive and Gram-negative bacterial samples grown using the same stock media solution. This is the first report of NIR spectroscopy being applied to differentiate Gram-positive and Gram-negative bacteria, as well as the first direct comparison of ATR-FTIR and NIR for the combined multimodal analysis of clinical bacterial isolates.

Gold Nanoparticles: Tunable Characteristics and Potential for Nasal Drug Delivery.

A general procedure to prepare gold nanourchins (GNUs) via a seed-mediated method was followed using dopamine hydrochloride as a reducing agent and silver nitrate salt (AgNO) as a shape-directing agent. The novelty of this study comes from the successful incorporation of the prepared gold urchins as an aqueous suspension in a nasal pressurized metered dose inhaler (pMDI) formulation and the investigation of their potential for olfactory targeting for direct nose-to-brain drug delivery (NTBDD). The developed pMDI formulation was composed of 0.

A Cell-Based Nasal Model for Screening the Deposition, Biocompatibility, and Transport of Aerosolized PLGA Nanoparticles.

The olfactory region of the nasal cavity directly links the brain to the external environment, presenting a potential direct route to the central nervous system (CNS). However, targeting drugs to the olfactory region is challenging and relies on a combination of drug formulation, delivery device, and administration technique to navigate human nasal anatomy. In addition, in vitro and in vivo models utilized to evaluate the performance of nasal formulations do not accurately reflect deposition and uptake in the human nasal cavity.

An in vitro and ex vivo wound infection model to test topical and systemic treatment with antibiotics.

Aims: This study aimed to develop a wound infection model that could be used to test antibiotic-loaded electrospun matrices for the topical treatment of infected skin and compare the effectiveness of this treatment to systemically applied antibiotics.

Methods And Results: 3D-printed flow chambers were made in which Staphylococcus aureus biofilms were grown either on a polycarbonate membrane or explanted porcine skin. The biofilms were then treated either topically, by placing antibiotic-loaded electrospun matrices on top of the biofilms, or systemically by the addition of antibiotics in the growth medium that flowed underneath the membrane or skin.

Intestinal Transcytosis of a Protein Cargo and Nanoparticles Mediated by a Non-Toxic Form of Exotoxin A.

The low permeability of nanoparticles (NPs) across the intestinal epithelium remains a major challenge for their application of delivering macromolecular therapeutic agents via the oral route. Previous studies have demonstrated the epithelial transcytosis capacity of a non-toxic version of exotoxin A (ntPE). Here, we show that ntPE can be used to deliver the protein cargo green fluorescent protein (GFP) or human growth hormone (hGH), as genetic fusions, across intact rat jejunum in a model where the material is administered by direct intra-luminal injection (ILI) in vivo in a transcytosis process that required less than 15 min.

In Vitro Evaluation of Nasal Aerosol Depositions: An Insight for Direct Nose to Brain Drug Delivery.

The nasal cavity is an attractive route for both local and systemic drug delivery and holds great potential for access to the brain via the olfactory region, an area where the blood-brain barrier (BBB) is effectively absent. However, the olfactory region is located at the roof of the nasal cavity and only represents ~5-7% of the epithelial surface area, presenting significant challenges for the deposition of drug molecules for nose to brain drug delivery (NTBDD). Aerosolized particles have the potential to be directed to the olfactory region, but their specific deposition within this area is confounded by a complex combination of factors, which include the properties of the formulation, the delivery device and how it is used, and differences in inter-patient physiology.

Decellularized grass as a sustainable scaffold for skeletal muscle tissue engineering.

Scaffold materials suitable for the scale-up and subsequent commercialization of tissue engineered products should ideally be cost effective and accessible. For the in vitro culture of certain adherent cells, synthetic fabrication techniques are often employed to produce micro- or nano-patterned substrates to influence cell attachment, morphology, and alignment via the mechanism of contact guidance. Here we present a natural scaffold, in the form of decellularized amenity grass, which retains its natural striated topography and supports the attachment, proliferation, alignment and differentiation of murine C2C12 myoblasts, without the need for additional functionalization.

Silk fibroin/gelatin microcarriers as scaffolds for bone tissue engineering.

Microcarrier cell scaffolds have potential as injectable cell delivery vehicles or as building blocks for tissue engineering. The use of small cell carriers allows for a 'bottom up' approach to tissue assembly when moulding microparticles into larger structures, which can facilitate the introduction of hierarchy by layering different matrices and cell types, while evenly distributing cells through the structure. In this work, silk fibroin (SF), purified from Bombyx mori cocoons, was blended with gelatin (G) to produce materials composed of varying ratios of the two components (SF: G 25:75, 50:50, and 75:25).

The effect of ozone gas sterilization on the properties and cell compatibility of electrospun polycaprolactone scaffolds.

The growing area of tissue engineering has the potential to alleviate the shortage of tissues and organs for transplantation, and electrospun biomaterial scaffolds are extremely promising devices for translating engineered tissues into a clinical setting. However, to be utilized in this capacity, these medical devices need to be sterile. Traditional methods of sterilization are not always suitable for biomaterials, especially as many commonly used biomedical polymers are sensitive to chemical-, thermal- or radiation-induced damage.

Platelet lysate gel and endothelial progenitors stimulate microvascular network formation in vitro: tissue engineering implications.

Revascularisation is a key step for tissue regeneration and complete organ engineering. We describe the generation of human platelet lysate gel (hPLG), an extracellular matrix preparation from human platelets able to support the proliferation of endothelial colony forming cells (ECFCs) in 2D cultures and the formation of a complete microvascular network in vitro in 3D cultures. Existing extracellular matrix preparations require addition of high concentrations of recombinant growth factors and allow only limited formation of capillary-like structures.

Ozone Gas as a Benign Sterilization Treatment for PLGA Nanofiber Scaffolds.

The use of electrospun nanofibers for tissue engineering and regenerative medicine applications is a growing trend as they provide improved support for cell proliferation and survival due, in part, to their morphology mimicking that of the extracellular matrix. Sterilization is a critical step in the fabrication process of implantable biomaterial scaffolds for clinical use, but many of the existing methods used to date can negatively affect scaffold properties and performance. Poly(lactic-co-glycolic acid) (PLGA) has been widely used as a biodegradable polymer for 3D scaffolds and can be significantly affected by current sterilization techniques.

Electrospun Zein/PCL Fibrous Matrices Release Tetracycline in a Controlled Manner, Killing Staphylococcus aureus Both in Biofilms and Ex Vivo on Pig Skin, and are Compatible with Human Skin Cells.

Purpose: To investigate the destruction of clinically-relevant bacteria within biofilms via the sustained release of the antibiotic tetracycline from zein-based electrospun polymeric fibrous matrices and to demonstrate the compatibility of such wound dressing matrices with human skin cells.

Methods: Zein/PCL triple layered fibrous dressings with entrapped tetracycline were electrospun. The successful entrapment of tetracycline in these dressings was validated.

An investigation into the stability of commercial versus MG63-derived hepatocyte growth factor under flow cultivation conditions.

The scale-up of tissue engineering cell culture must ensure that conditions are maintained while also being cost effective. Here we analyse the stability of hepatocyte growth factor (HGF) to investigate whether concentrations change under dynamic conditions, and compare commercial recombinant human HGF as an additive in 'standard medium', to HGF secreted by the osteosarcoma cell line MG63 as a 'preconditioned medium'. After 3 h under flow conditions, HGF in the standard medium degraded to 40% of its original concentration but HGF in the preconditioned medium remained at 100%.

A hydrazide-anchored dendron scaffold for chemoselective ligation strategies.

Chemoselective ligation, including "click" chemistry, has found wide utility in general synthetic strategies and the specific modification of polymers and biomolecules. This has resulted in a number of applications of such approaches, particularly in the biomedical area, including diagnostic imaging and drug delivery. However, tools to chemoselectively decorate target molecules with multiple copies of a particular drug, ligand or label are lacking.

Zein/polycaprolactone electrospun matrices for localised controlled delivery of tetracycline.

We report the controlled release of the antibiotic tetracycline (Tet) from triple-layered (3L) electrospun matrices consisting of zein or a zein/PCL blend, where the drug was loaded into the central layer with the two outer layers acting as diffusion barriers. These fibrous matrices successfully encapsulated Tet and efficiently inhibited the growth of a clinical isolate, the methicillin-resistant Staphylococcus aureus strain MRSA252, as demonstrated in a modified Kirby-Bauer disc assay over 5 days. Whilst untreated zein fibres are unstable in an aqueous environment, rapidly shrinking due to plasticisation and film formation, blending zein with PCL stabilised the electrospun matrices and prevented them from shrinking.

Killing bacteria within biofilms by sustained release of tetracycline from triple-layered electrospun micro/nanofibre matrices of polycaprolactone and poly(ethylene-co-vinyl acetate).

We report the controlled release of the antibiotic tetracycline (tet) HCl from a triple-layered electrospun matrix consisting of a central layer of poly(ethylene-co-vinyl acetate (PEVA) sandwiched between outer layers of poly-ε-caprolactone (PCL). These micro/nanofibre layers with tet successfully encapsulated (essentially quantitatively at 3 and 5 % w/w) in each layer, efficiently inhibited the growth of a panel of bacteria, including clinical isolates, as shown by a modified Kirby-Bauer disc assay. Furthermore, they demonstrated high biological activity in increasingly complex models of biofilm formation (models that are moving closer to the situation in a wound) by stopping biofilm formation, by killing preformed biofilms and killing mature, dense biofilm colonies of Staphylococcus aureus MRSA252.

Oxidized alginate hydrogels as niche environments for corneal epithelial cells.

Chemical and biochemical modification of hydrogels is one strategy to create physiological constructs that maintain cell function. The aim of this study was to apply oxidised alginate hydrogels as a basis for development of a biomimetic niche for limbal epithelial stem cells that may be applied to treating corneal dysfunction. The stem phenotype of bovine limbal epithelial cells (LEC) and the viability of corneal epithelial cells (CEC) were examined in oxidised alginate gels containing collagen IV over a 3-day culture period.

Multicellular aggregation of maltol-modified cells triggered by Fe(3+) ions.

The synthesis of a maltol-derived hydrazide is described which, once attached to a cell surface, induces rapid multicellular aggregation selectively in the presence of Fe(3+) ions. Heterocellular aggregates are also reported.

Electrospun matrices for localised controlled drug delivery: release of tetracycline hydrochloride from layers of polycaprolactone and poly(ethylene-co-vinyl acetate).

We report the controlled release of tetracycline (Tet) HCl from a three-layered electrospun matrix for the first time. Five formulations of electrospun poly-ε-caprolactone (PCL) and poly(ethylene-co-vinyl acetate) (PEVA) have been designed, prepared as micro/nanofibre layers, and assayed for the controlled release of the clinically useful antibiotic Tet HCl with potential applications in wound healing and especially in complicated skin and skin-structure infections. Tet HCl was also chosen as a model drug possessing a good ultraviolet (UV) chromophore and capable of fluorescence together with limited stability.

Simple pyrazoline and pyrazole "turn on" fluorescent sensors selective for Cd2+ and Zn2+ in MeCN.

An efficient two-step synthesis of pyrazoline ligand is described which is an effective "turn on" fluorescent sensor for Cd(2+) in MeCN. Oxidation to the corresponding pyrazole ligand creates a "turn on" fluorescent sensor now selective for Zn(2+) and able to distinguish it from Cd(2+).

Accelerated formation of multicellular 3-D structures by cell-to-cell cross-linking.

The three-dimensional (3-D) arrangement of cells within tissues is integral to their development and function. Advances in stem cell science and regenerative medicine have stimulated interest in the replication of this architecture in vitro. We have developed a versatile method for controlling short-term cell-cell and cell-matrix interactions via a facile cell surface engineering process that enables the rapid formation of specific 3-D interactions for a range of cell types.

A spectrophotometric assay for fatty acid amide hydrolase suitable for high-throughput screening.

Signalling via the endocannabinoids anandamide and 2-arachidonylglycerol appears to be terminated largely through the action of the enzyme fatty acid amide hydrolase (FAAH). In this report, we describe a simple spectrophotometric assay to detect FAAH activity in vitro using the ability of the enzyme to hydrolyze oleamide and measuring the resultant production of ammonia with a NADH/NAD+-coupled enzyme reaction. This dual-enzyme assay was used to determine Km and Vmax values of 104 microM and 5.

Chemical modification of mammalian cell surfaces.

The mammalian cell surface is a highly heterogeneous chemical environment with proteins, carbohydrates, lipids and composite molecules controlling vital cell functions. Chemical modification of this environment is a challenge due to the complexity of the surface chemistry and the fragility of the cell. Here, we review recent attempts to perform targeted, non-genetically controlled, changes to cell surface chemistry.