Zimei Rong

Influence of nanopatterns on endothelial cell adhesion: Enhanced cell retention under shear stress.

In this study, nanopatterned crosslinked films of collagen Type I were seeded with human microvascular endothelial cells and tested for their suitability for vascular tissue engineering. Since the films will be rolled into tubes with concentric layers of collagen, nutrient transfer through the collagen films is quite crucial. Molecular diffusivity through the collagen films, cell viability, cell proliferation and cell retention following shear stress were studied. Cells were seeded onto linearly nanogrooved films (groove widths of 332.5, 500 and 650nm), with the grooves aligned in the direction of flow. The nanopatterns did not affect cell proliferation or initial cell alignment; however, they significantly affected cell retention under fluid flow. While cell retention on unpatterned films was 35+/-10%, it was 75+/-4% on 332.5nm patterned films and even higher, 91+/-5%, on 650nm patterned films. The films were found to have diffusion coefficients of ca. 10(-6)cm(2)s(-1) for O(2) and 4-acetaminophenol, which is comparable to that observed in natural tissues. This constitutes another positive asset of these films for consideration as a scaffold material for vascular tissue engineering.

Oxygen diffusion through collagen scaffolds at defined densities: implications for cell survival in tissue models.

For the success of any biomaterial for tissue engineering, its mechanical properties and ability to support nutrient diffusion will be critical. Collagen scaffolds are ideal candidates, due to their ability to immerse cells in a biomimetic nanofibrous matrix. We have established O2 diffusion coefficients through native, dense collagen scaffolds at two tissue‐like densities, with and without photo‐chemical crosslinking, by adapting an optical fibre‐based system for real‐time core O2 monitoring deep within collagen constructs. Using a Ficks law model, we then derived O2 diffusion coefficients; 4.5 × 10−6 cm2/s for 11% density collagen scaffolds; 1.7 × 10−6 cm2/s for 34% collagen scaffolds; 3.4 × 10−6 cm2/s for photochemically crosslinked collagen scaffolds at 11%. Both O2 diffusion coefficients of the 11% collagen fall within the range of native intestinal submucosa. The high diffusion coefficients of these collagen scaffolds, as well as their material properties, render them viable tissue‐engineering matrices for tissue replacement. Copyright

Mathematical modelling of fibre-enhanced perfusion inside a tissue-engineering bioreactor

We develop a simple mathematical model for forced flow of culture medium through a porous scaffold in a tissue-engineering bioreactor. Porous-walled hollow fibres penetrate the scaffold and act as additional sources of culture medium. The model, based on Darcys law, is used to examine the nutrient and shear-stress distributions throughout the scaffold. We consider several configurations of fibres and inlet and outlet pipes. Compared with a numerical solution of the full Navier-Stokes equations within the complex scaffold geometry, the modelling approach is cheap, and does not require knowledge of the detailed microstructure of the particular scaffold being used. The potential of this approach is demonstrated through quantification of the effect the additional flow from the fibres has on the nutrient and shear-stress distribution.

Fabrication of Biomaterials via Controlled Protein Bubble Generation and Manipulation

In this work, we utilize a recently developed microbubbling process to generate controlled protein (bovine serum albumin, BSA) coated bubbles and then manipulate these to fabricate a variety of structures suitable for several generic biomedical applications, tissue engineering, and biosensor coatings. Using BSA solutions with varying concentrations (20, 25, and 30 wt %) and cross-linking (terephthaloyl chloride) mechanisms, structures were fabricated including porous thin films with variable pore sizes and thickness (partially cross-linked coupled to bubble breakdown), scaffolds with variable pore morphologies (fully cross-linked), and coated bubbles (no cross-linking), which can be used as stand-alone delivery devices and contrast agents. The movement of typical biosensor chemicals (catechol and hydrogen peroxide) across appropriate film structures was studied. The potential of formed scaffold structures for tissue engineering applications was demonstrated using mouse cell lines (L929). In addition to low cost, providing uniform structure generation and high output, the size of the bubbles can easily be controlled by adjusting simplistic processing parameters. The combination of robust processing and chemical modification to uniform macromolecule bubbles can be utilized as a competing, yet novel, tool with current technologies and processes in advancing the biomaterials and biomedical engineering remits.

Needle Enzyme Electrode for Lactate Measurement In Vivo

Electrochemical lactate needle enzyme electrodes were fabricated based on lactate oxidase with a conventional hydrogen peroxide detection regimen with a linear range up to 7 mM, response time ~ 3 min, and sensitivity ~ 1 nA/mM. A negatively charged inner (sulphonated polyether ether sulphone-polyether sulphone) membrane was applied for ensuring selectivity by limiting oxidazible anion diffusion to the Pt working electrode; polyurethane outer membrane layers were dip coated over the enzyme layer to limit substrate diffusion to the enzyme layer to achieve: (1) stir independence and (2) a low oxygen requirement. Lactate was monitored subcutaneously in rats during controlled haemorrhage and hypovolaemic shock. Tissue lactate showed agreement with blood lactate before haemorrhage and for limited haemorrhage (up to 2 ml blood withdrawal from 16 ml total blood volume) but with blood loss above 3 ml the tissue lactate rise was less pronounced than in blood. Loss of intercompartmental equilibrium due to diffusion limitation is suggested as a factor in causing this difference. An experimental in vitro model was developed which employed the needle lactate electrode within a cylindrical collagen gel to monitor inward diffusion of lactate as a basis for determining lactate diffusion coefficient. The high precision measurement gave a diffusion coefficient consistent with report values 3.54 times 10-6 cm2/s. The simplified experimental approach could allow lactate transport studies across tissue analogues.

In Situ Fabrication of Cross-Linked Protein Membranes by Using Microfluidics

We report a novel technique for preparing cross‐linked protein membranes within microchannels by using an interfacial cross‐linking reaction. Glass microchannels with a Y input were assembled by using a simple adhesive bonding technique to achieve dual, parallel laminar flows. Membrane formation utilised an interfacial reaction at the liquid–liquid interface, which involved bovine serum albumin (aqueous solution with a flow rate of 300 μL min−1) and terephthaloyl chloride (xylene solution with a flow rate of 700 μL min−1), to form thin (∼25 μm) cross‐linked films along the length of the channel under the continuous pressure‐driven laminar flow. Such microfabricated membranes could extend the separation potential of any microfluidic structure to provide a stable barrier layer. Furthermore, degradation of the membrane was possible by using an alkali sodium dodecyl sulfate solution, which led to the complete disappearance of the membrane. These membranes could facilitate additional modification to allow for different permeability properties by controlled degradation. The one‐step in situ membrane‐fabrication methodology reported here generated precisely localised membranes and avoided the complexities of subcomponent assembly, which require complicated alignment of small, preformed membranes. This methodology could become the basis for sophisticated microseparation systems, biosensors and several “lab‐on‐a‐chip” devices.

Study of albumin and fibrinogen membranes formed by interfacial crosslinking using microfluidic flow

Microfluidics enables scale reduction in sample volume with obvious benefits for reagent conservation. In contrast to conventional macro-scale flow, microfluidics also offers unprecedented control over flow dynamics. In particular, laminar flow is readily achieved, allowing for new analytical and synthetic strategies. Here, two parallel flows of buffer and xylene were used to create a stable liquid-liquid interface within linear micro-channels. These, respectively, carried protein (albumin or fibrinogen) and an acyl chloride to effect protein crosslinking. This created robust, micro-membranes at the interface that bisected the fluid channel. Membrane formation was self-limiting, with fibrinogen membranes showing greater solute permeability than albumin, based on dye transport (Ponceau S, Meldola Blue). The crosslinker isophthaloyl dichloride led to thinner, less permeable membranes than terephthaloyl chloride. Larger surface area membranes formed at a static liquid-liquid interface served as a more physically accessible model and allowed precise electrochemical determination of acetaminophen, catechol and peroxide diffusion coefficients, which confirmed the greater fibrinogen permeability. Scanning electron microscopy (SEM) of the membranes also indicated a higher population of discrete nanopores at the fibrinogen surface. A crosslinking pH had a strong effect on overall permeability. Adhesion of B50 neuronal cells was demonstrated, and it is proposed that the membranes could facilitate cell growth through bidirectional nutrient supply in a micrbioreactor format.

Displacement matrix elements of Deng-Fan oscillators

Analytic expressions for matrix elements of integral powers of the displacement of the coordinate from equilibrium for the Deng-Fan oscillator have been derived. A comparison is made with the corresponding matrix elements for the Morse oscillator. These matrix elements are useful in high overtone spectroscopy and in models for DNA melting.

Calculation of displacement matrix elements for Morse oscillators

Displacement matrix elements of the Morse oscillators are widely used in physical and chemical computation. Many papers have been published about the analytical expressions of the displacement matrix elements. People still use an integration method rather than the analytical expressions to calculate the displacement matrix elements. Analytical expressions of the diagonal and off diagonal displacement matrix elements with intermediate variables are derived, where v, m and n are non-negative integers and n • 6. Calculation with these expressions is superior to that with numerical integration in precision, calculation speed and convenience.

O 2 Diffusion through Collagen Scaffolds at Defined Densities: Implications for Cell Survival and Angiogenic Signalling in Tissue Models

The success of any biomaterial for tissue engineering is dominated by its mechanical properties and ability to support nutrient diffusion. Collagen scaffolds are ideal candidates due to their ability to immerse cells in a biomimetic nano-fibrous matrix. We have established O2 diffusion coefficients through native, dense collagen scaffolds at two tissue-like densities, with and without photo-chemical crosslinking, by adapting an optical fibre-based system for real-time core O2 monitoring deep within collagen constructs. The high diffusion coefficients of these collagen scaffolds, as well as their material properties, render them viable tissue engineering matrices for tissue replacement. Due to this O2 diffusion through cell-seeded collagen type I scaffolds, natural gradients of O2 form and cells in different locations are subject to varying levels of O2. These gradients were controlled by varying cell density, as it was found that cell consumption of O2 played a greater role compared to material diffusion in formation of such O2 gradients. Potent angiogenic signaling molecules were upregulated at both the gene and protein level, particularly within the core of 3D scaffolds, where O2 was low, but remained within physiological hypoxia. By incorporating phosphate-based dissolving glass fibres into collagen constructs, as they are produced, it was possible to introduce channels throughout the construct in a gradual manner. Where channeled architecture was introduced to the 3D constructs, thus delivery of sustained O2 to all cells even within the core, this upregulation of angiogenic factors was abolished. We can now engineer collagen type I scaffolds with varying density, varying degrees of crosslinking and various architectural features to control delivery of O2 to all cells embedded within the construct.