Jeremy C.M. Teo

Characterization of membrane materials and membrane coatings for bioreactor units of bioartificial kidneys.

The bioreactor unit of bioartificial kidneys contains porous membranes seeded with renal cells. For clinical applications, it is mandatory that human primary renal proximal tubule cells (HPTCs) form differentiated epithelia on the membranes. Here, we show that HPTCs do not grow and survive on a variety of polymeric membrane materials. This applies also to membranes consisting of polysulfone/polyvinylpyrrolidone (PSF/PVP), which have been used in the bioreactor unit of bioartificial kidneys after coating with an extracellular matrix (ECM). Our data reveal that coating with just an ECM does not sufficiently improve HPTC performance on non-HPTC-compatible membrane materials. On the other hand, we have characterized the effects of a variety of surface treatments and coatings, and found that double coating with 3,4-dihydroxy-l-phenylalanine and an ECM markedly improves HPTC performance and results in the formation of differentiated epithelia on PSF/PVP membranes. We have also synthesized alternative membrane materials, and characterized membranes consisting of polysulfone and Fullcure. We found that these membranes sustain proper HPTC performance without the need for surface treatments or coatings. Together, our data reveal that the materials that have been previously applied in bioartificial kidneys are not suitable for applications with HPTCs. This study elucidates the types of membrane materials and coatings that are favorable for the bioreactor unit of bioartificial kidneys.

Intravital Immunofluorescence for Visualizing the Microcirculatory and Immune Microenvironments in the Mouse Ear Dermis

Visualizing the dynamic behaviors of immune cells in living tissue has dramatically increased our understanding of how cells interact with their surroundings, contributing important insights into mechanisms of leukocyte trafficking, tumor cell invasion, and T cell education by dendritic cells, among others. Despite substantial advances with various intravital imaging techniques including two-photon microscopy and the generation of multitudes of reporter mice, there is a growing need to assess cell interactions in the context of specific extracellular matrix composition and microvascular functions, and as well, simpler and more widely accessible methods are needed to image cell behaviors in the context of living tissue physiology. Here we present an antibody-based method for intravital imaging of cell interactions with the blood, lymphatic, and the extracellular matrix compartments of the living dermis while simultaneously assessing capillary permeability and lymphatic drainage function. Using the exposed dorsal ear of the anesthetized mouse and a fluorescence stereomicroscope, such events can be imaged in the context of specific extracellular matrix proteins, or matrix-bound chemokine stores. We developed and optimized the method to minimize tissue damage to the ear, rapidly immunostain for multiple extracellular or cell surface receptors of interest, minimize immunotoxicity with pre-blocking Fcγ receptors and phototoxicity with extracellular antioxidants, and highlight the major dermal tissue structures with basement membrane markers. We demonstrate differential migration behaviors of bone marrow-derived dendritic cells, blood-circulating leukocytes, and dermal dendritic cells, with the latter entering sparse CCL21-positive areas of pre-collecting lymphatic vessels. This new method allows simultaneous imaging of cells and tissue structures, microvascular function, and extracellular microenvironment in multiple skin locations for 12 hours or more, with the flexibility of immunolabeling in addition to genetic-based fluorescent reporters.

Computational biomechanical modelling of the lumbar spine using marching-cubes surface smoothened finite element voxel meshing

There is a need for the development of finite element (FE) models based on medical datasets, such as magnetic resonance imaging and computerized tomography in computation biomechanics. Direct conversion of graphic voxels to FE elements is a commonly used method for the generation of FE models. However, conventional voxel-based methods tend to produce models with jagged surfaces. This is a consequence of the inherent characteristics of voxel elements; such a model is unable to capture the geometries of anatomical structures satisfactorily. We have developed a robust technique for the automatic generation of voxel-based patient-specific FE models. Our approach features a novel tetrahedronization scheme that incorporates marching-cubes surface smoothing together with a smooth-distortion factor (SDF). The models conform to the actual geometries of anatomical structures of a lumbar spine segment (L3). The resultant finite element analysis (FEA) at the surfaces is more accurate compared to the use of conventional voxel-based generated FE models. In general, models produced by our method were superior compared to that obtained using the commercial software ScanFE.

Haptics in computer-mediated simulation: training in vertebroplasty surgery

Surgical simulators and computer games share the enabling technologies in the human-machine interface. With appropriate design and development, the computer-game-like medical training simulator could be used in surgical training. The authors describe a PC-based system for the simulation of the vertebroplasty procedure. In vertebroplasty, the surgeon or radiologist relies on sight and feel to properly insert the bone needle through various tissue types and densities and monitor the injection and reflux of the polymethylmethacrylate (PMMA), or cement, into the vertebra. This article focuses on the provision of a near-realistic haptic feel in bone needle insertion and manual PMMA injection. This involves an efficient biomechanical modeling of bone needle insertion and PMMA flow in bone for haptic rendering, as well as reliable delivery of forces via haptic devices. The authors show that with virtual reality gaming technologies, the surgical simulator can become a virtual trainer for a potentially risky spinal interventional procedure.

A component-oriented software toolkit for patient-specific finite element model generation

A component-oriented software system, i.BioMech (image-based biomechanical modeling) is proposed for generation of patient-specific finite element model. It applies a systematic software engineering approach to patient/subject-specific meshing and assignment of material properties. The prototype program is based on the component object model (COM), which enables ease of combination of existing mesh generation algorithms and material property assignment schemes, and incorporation of new ones. It also facilitates utilization by other programming languages or platforms. Data input comprises a series of medical images captured from the patient. The output is a patient-specific finite element model for computational analysis using commercially available finite element software. The prototype software system provides a framework to compare the different finite element mesh generation methods as well as schemes for material property assignment. Our focus is on patient/subject-specific modeling of the human vertebrae.

Preliminary study on biomechanics of vertebroplasty: a computational fluid dynamics and solid mechanics combined approach.

Study Design. Algorithm development for the automatic finite element modeling of patient vertebra. Objective. To present a technique for automatic generation of patient specific computational fluid dynamics (CFD) models for intraosseous PMMA cement flow simulation. The secondary objective is to demonstrate the possibility of using resultant PMMA cement distribution for post-PVP stress-strain analyses. Summary of Background Data. There are no noninvasive methods for the visualization of PMMA cement flow. In addition, optimum volume and distribution of PMMA cement are still not known. Computational models that allow patient specific intraosseous PMMA cement flow visualization as well as postvertebroplasty mechanical evaluation would be advantageous. Methods. We developed an algorithm coded into a custom platform that inputs patient CT datasets. Hounsfield unit values were used to assign permeability values as well as modulus to the finite element model before analyses. Several user inputs are required, and these reflect the decisions made by physicians that practice vertebroplasty. As a case study, we isolated a single L1 vertebra from patient CT dataset and used our platform for model generation. Simulated vertebroplasty was performed for different PMMA cement volumes and at different placements to study the effects of varying distribution. Results. Increased needle injection pressure was observed as the volume of PMMA increases and as the distribution of PMMA is in close proximity to the cortical walls. Stiffness of augmented vertebral body also increases with increased volume of PMMA administered. Varying distributions, for the same volume, of PMMA cement did not alter stiffness drastically. Conclusion. Our custom platform and technique for modeling vertebral bodies may contribute significantly to the science of vertebroplasty. Intraosseous PMMA cement flow can be visualized before vertebroplasty, and needle position altered for optimization. Also, parametric computational studies on the postvertebroplasty biomechanical effects of vertebroplasty are now enhanced with such a modeling capability.

Permeability study of vertebral cancellous bone using micro-computational fluid dynamics

Understanding of cancellous bone permeability is lacking despite its importance in designing tissue engineering scaffolds for bone regeneration and orthopaedic surgery that relies on infiltration of bone cement into porous cancellous bone. We employed micro-computational fluid dynamics to investigate permeability for 37 cancellous bone specimens, eliminating stringent technical requirements of bench-top testing. Microarchitectural parameters were also determined for the specimens and correlated, using uni-variate and multi-variate regression analyses, against permeability. We determined that bone surface density, trabecular pattern factor, structure model index and trabecular number are other possible predictors of permeability (with R values of 0.47, 0.44, 0.40 and 0.33), in addition to the commonly used porosity parameter (R value of 0.38). Pooling these parameters and performing multi-variate linear regression analysis improved yield the R-value of 0.50, indicating that porosity alone is a poor predictor of cancellous bone permeability and, therefore, other parameters should be included for a better and improved linear model.

A Fibrin-Based Tissue-Engineered Renal Proximal Tubule for Bioartificial Kidney Devices: Development, Characterization and In Vitro Transport Study

A bioartificial renal proximal tubule is successfully engineered as a first step towards a bioartificial kidney for improved renal substitution therapy. To engineer the tubule, a tunable hollow fiber membrane with an exterior skin layer that provides immunoprotection for the cells from extracapillary blood flow and a coarse inner surface that facilitates a hydrogel coating for cell attachment was embedded in a “lab-on-a-chip” model for the small-scale exploratory testing under flow conditions. Fibrin was coated onto the inner surface of the hollow fiber, and human renal proximal tubule epithelial cells were then seeded. Using this model, we successfully cultured a confluent monolayer, as ascertained by immunofluorescence staining for ZO-1 tight junctions and other proximal tubule markers, scanning electron microscopy, and FITC-inulin recovery studies. Furthermore, the inulin studies, combined with the creatinine and glucose transport profiles, suggested that the confluent monolayer exhibits functional transport capabilities. The novel approaches here may eventually improve current renal substitution technology for renal failure patients.

Surface characteristics of acrylic modified polysulfone membranes improves renal proximal tubule cell adhesion and spreading

Current polyvinylpyrrolidone-modified polysulfone (PVP-PSU) membranes in haemodialysers do not facilitate the attachment and proliferation of renal proximal tubule cells (RPTCs). For bioartificial kidney (BAK) development expensive extracellular matrices are employed to ensure the PVP-PSU membranes can serve as a substrate for RPTCs. In this study we modified PSU using an acrylic monomer (am-PSU) and polymerization using ultraviolet irradiation. We demonstrated that on adjusting the PSU or acrylic content of the membranes the wettability and surface chemistry were altered, and this affected the amount of fibronectin (Fn) that was adsorbed onto the membranes. Using an integrin blocking assay we ascertained that Fn is an important extracellular matrix component that mediates RPTC attachment. The amount of Fn adsorbed also led to different bioresponses of RPTCs, which were evaluated using attachment and proliferation assays and qualitative quantification of vinculin, focal adhesion kinase, zonula occludens and Na(+)/K(+) ATPase. Our optimized membrane, am-PSU1 (21.4% C-O groups, 19.1% PVP-PSU; contact angle 71.5-80.80, PVP-PSU: 52.4-67.50), supports a confluent monolayer of RPTCs and prevents creatinine and inulin diffusion from the apical to the basal side, meeting the requirements for application in BAKs. However, further in vivo evaluation to assess the full functionality of RPTCs on am-PSU1 is required.

Modeling of the Human Orbit from MR Images

We previously described a parametric eyeball modeling system for real-time simulation of eye surgery (MICCAI 01). However, in the simulation of ophthalmologic surgery, the model of the eyeball alone is not sufficient. The orbital structures are as important as the eyeball. In this paper, we describe the approach to model the orbital structures from patient specific MRI data set and integrate the orbital model with the parametric eyeball model. The orbital tissues including the eyeball, muscles, and orbital fat are segmented from MRI data. An interactive image-based geometrical modeling tool is developed to generate a finite element model of the orbit. Preliminary results include biomechanical models of three human subjects, one of which is a young patient with a benign tumor in the right orbit. The biomedical model can provide quantitative information that is important in diagnosis. It can also be used to accurately analyze the result of intervention, which is an important component of the simulator for training and treatment planning. Our analysis includes a deformation study on an eyeball subjected to simulated tumor growth using the finite element method.