The P387 thrombospondin-4 variant promotes accumulation of macrophages in atherosclerotic lesions.

Thrombospondin-4 (TSP4) is a pro-angiogenic protein that has been implicated in tissue remodeling and local vascular inflammation. TSP4 and, in particular, its SNP variant, P387 TSP4, have been associated with cardiovascular disease. Macrophages are central to initiation and resolution of inflammation and development of atherosclerotic lesions, but the effects of the P387 TSP4 on macrophages remain essentially unknown. We examined the effects of the P387 TSP4 variant on macrophages in cell culture and in vivo in a murine model of atherosclerosis. Furthermore, the levels and distributions of the two TSP4 variants were assessed in human atherosclerotic arteries. In ApoE-/- /P387-TSP4 knock-in mice, lesions size measured by Oil Red O did not change, but the lesions accumulated more macrophages than lesions bearing A387 TSP4. The levels of inflammatory markers were increased in lesions of ApoE-/- /P387-TSP4 knock-in mice compared to ApoE-/- mice. Lesions in human arteries from individuals carrying the P387 variant had higher levels of TSP4 and higher macrophage accumulation. P387 TSP4 was more active in supporting adhesion of cultured human and mouse macrophages in experiments using recombinant TSP4 variants and in cells derived from P387-TSP4 knock-in mice. TSP4 supports the adhesion of macrophages and their accumulation in atherosclerotic lesions without changing the size of lesions. P387 TSP4 is more active in supporting these pro-inflammatory events in the vascular wall, which may contribute to the increased association of P387 TSP4 with cardiovascular disease.

Artificial Intelligence in Intracoronary Imaging.

PURPOSE OF REVIEW: This paper investigates present uses and future potential of artificial intelligence (AI) applied to intracoronary imaging technologies. RECENT FINDINGS: Advances in data analytics and digitized medical imaging have enabled clinical application of AI to improve patient outcomes and reduce costs through better diagnosis and enhanced workflow. Applications of AI to IVUS and IVOCT have produced improvements in image segmentation, plaque analysis, and stent evaluation. Machine learning algorithms are able to predict future coronary events through the use of imaging results, clinical evaluations, laboratory tests, and demographics. The application of AI to intracoronary imaging holds significant promise for improved understanding and treatment of coronary heart disease. Even in these early stages, AI has demonstrated the ability to improve the prediction of cardiac events. Large curated data sets and databases are needed to speed the development of AI and enable testing and comparison among algorithms.

Response of bone marrow derived connective tissue progenitor cell morphology and proliferation on geometrically modulated microtextured substrates.

Varying geometry and layout of microposts on a cell culture substrate provides an effective technique for applying mechanical stimuli to living cells. In the current study, the optimal geometry and arrangement of microposts on the polydimethylsiloxane (PDMS) surfaces to enhance cell growth behavior were investigated. Human bone marrow derived connective tissue progenitor cells were cultured on PDMS substrates comprising unpatterned smooth surfaces and cylindrical post microtextures that were 10 µm in diameter, 4 heights (5, 10, 20 and 40 µm) and 3 pitches (10, 20, and 40 µm). With the same 10 µm diameter, post heights ranging from 5 to 40 µm resulted in a more than 535 fold range of rigidity from 0.011 nNµm⁻¹ (40 µm height) up to 5.888 nNµm⁻¹(5 µm height). Even though shorter microposts result in higher effective stiffness, decreasing post heights below the optimal value, 5 µm height micropost in this study decreased cell growth behavior. The maximum number of cells was observed on the post microtextures with 20 µm height and 10 µm inter-space, which exhibited a 675 % increase relative to the smooth surfaces. The cells on all heights of post microtextures with 10 µm and 20 µm inter-spaces exhibited highly contoured morphology. Elucidating the cellular response to various external geometry cues enables us to better predict and control cellular behavior. In addition, knowledge of cell response to surface stimuli could lead to the incorporation of specific size post microtextures into surfaces of implants to achieve surface-textured scaffold materials for tissue engineering applications.

A 100-V Withstanding Analog-Front-End for High-Resolution Intravascular Ultrasound Imaging.

Intravascular Ultrasound ultrasonic imaging (IVUS) can microscopically image blood vessels and reveal tissue layers from within the blood vessel lumen. It has high tissue penetration ability for lesion classification and can image through blood. Compared to optical techniques, however, IVUS has lower resolution arising from low acoustic bandwidths which cannot resolve sharp edges. The presented 100-V withstanding Analog-Front-End (AFE) was developed to enable a high resolution, low cost IVUS system using a high-bandwidth focused polymer transducer with 40-MHz center frequency. The fabricated AFE interfaced with the transducer with minimal insertion loss, could withstand and duplex 100-V high voltage pulses and echo signal, and had a total signal chain gain of 9.8 dB. The AFE achieved a signal-to-noise ratio (SNR) of 20.1 dB including the insertion loss of the high-impedance transducer. AFE SNR was limited by input impedance required for high-voltage pulse clamping circuitry, but was sufficient for IVUS echo reception.Clinical Relevance- This work has the potential to enable much higher resolution, and potentially cheaper, IVUS imaging in blood vessels by integrating low-cost acoustic transducers with interface amplifiers directly on the catheter.

Microfluidic chip for graduated magnetic separation of circulating tumor cells by their epithelial cell adhesion molecule expression and magnetic nanoparticle binding.

The enumeration of circulating tumor cells (CTCs) in the peripheral bloodstream of metastatic cancer patients has contributed to improvements in prognosis and therapeutics. There have been numerous approaches to capture and counting of CTCs. However, CTCs have potential information beyond simple enumeration and hold promise as a liquid biopsy for cancer and a pathway for personalized cancer therapy by detecting the subset of CTCs having the highest metastatic potential. There is evidence that epithelial cell adhesion molecule (EpCAM) expression level distinguishes these highly metastatic CTCs. The few previous approaches to selective CTC capture according to EpCAM expression level are reviewed. A new two-stage microfluidic device for separation, enrichment and release of CTCs into subpopulations sorted by EpCAM expression level is presented here. It relies upon immunospecific magnetic nanoparticle labeling of CTCs followed by their field- and flow-based separation in the first stage and capture as discrete subpopulations in the second stage. To fine tune the separation, the magnetic field profile across the first stage microfluidic channel may be modified by bonding small Vanadium Permendur strips to its outer walls. Mathematical modeling of magnetic fields and fluid flows supports the soundness of the design.

Three-dimensional in vitro follicle growth: overview of culture models, biomaterials, design parameters and future directions.

In vitro ovarian follicle culture is a new frontier in assisted reproductive technology with tremendous potential, especially for fertility preservation. Folliculogenesis within the ovary is a complex process requiring interaction between somatic cell components and the oocyte. Conventional two-dimensional culture on tissue culture substrata impedes spherical growth and preservation of the spatial arrangements between oocyte and surrounding granulosa cells. Granulosa cell attachment and migration can leave the oocyte naked and unable to complete the maturation process. Recognition of the importance of spatial arrangements between cells has spurred research in to three-dimensional culture system. Such systems may be vital when dealing with human primordial follicles that may require as long as three months in culture. In the present work we review pertinent aspects of in vitro follicle maturation, with an emphasis on tissue-engineering solutions for maintaining the follicular unit during the culture interval. We focus primarily on presenting the various 3-dimensional culture systems that have been applied for in vitro maturation of follicle:oocyte complexes. We also try to present an overview of outcomes with various biomaterials and animal models and also the limitations of the existing systems.

A microfluidic bioreactor with integrated transepithelial electrical resistance (TEER) measurement electrodes for evaluation of renal epithelial cells.

We have developed a bilayer microfluidic system with integrated transepithelial electrical resistance (TEER) measurement electrodes to evaluate kidney epithelial cells under physiologically relevant fluid flow conditions. The bioreactor consists of apical and basolateral fluidic chambers connected via a transparent microporous membrane. The top chamber contains microfluidic channels to perfuse the apical surface of the cells. The bottom chamber acts as a reservoir for transport across the cell layer and provides support for the membrane. TEER electrodes were integrated into the device to monitor cell growth and evaluate cell-cell tight junction integrity. Immunofluorescence staining was performed within the microchannels for ZO-1 tight junction protein and acetylated α-tubulin (primary cilia) using human renal epithelial cells (HREC) and MDCK cells. HREC were stained for cytoskeletal F-actin and exhibited disassembly of cytosolic F-actin stress fibers when exposed to shear stress. TEER was monitored over time under normal culture conditions and after disruption of the tight junctions using low Ca(2+) medium. The transport rate of a fluorescently labeled tracer molecule (FITC-inulin) was measured before and after Ca(2+) switch and a decrease in TEER corresponded with a large increase in paracellular inulin transport. This bioreactor design provides an instrumented platform with physiologically meaningful flow conditions to study various epithelial cell transport processes.

Permeability - Selectivity Analysis for Ultrafiltration: Effect of Pore Geometry.

The effects of pore size on the performance of ultrafiltration membranes are fairly well understood, but there is currently no information on the impact of pore geometry on the trade-off between the selectivity and permeability for membranes with pore size below 100 nm. Experimental data are presented for both commercial ultrafiltration membranes and for novel silicon membranes having slit-shaped nanopores of uniform size fabricated by photolithography using a sacrificial oxide technique. Data are compared with theoretical calculations based on available hydrodynamic models for solute and solvent transport through membranes composed of a parallel array of either cylindrical or slit-shaped pores. The results clearly demonstrate that membranes with slit-shaped pores have higher performance, i.e., greater selectivity at a given value of the permeability, than membranes with cylindrical pores. Theoretical calculations indicate that this improved performance becomes much less pronounced as the breadth of the pore size distribution increases. These results provide new insights into the effects of pore geometry on the performance of ultrafiltration membranes.

Design and analysis of MEMS based PVDF ultrasonic transducers for vascular imaging.

Polyvinilidene fluoride (PVDF) single-element transducers for high-frequency (>30 MHz) ultrasound imaging applications have been developed using MEMS (Micro-electro-Mechanical Systems) compatible techniques. Performance of these transducers has been investigated by analyzing the sources and effects of on-chip parasitic capacitances on the insertion-loss of the transducers. Modeling and experimental studies showed that on-chip parasitic capacitances degraded the performance of the transducers and an improved method of fabrication was suggested and new devices were built. New devices developed with minimal parasitic effects were shown to improve the performance significantly. A 1-mm aperture PVDF device developed with minimal parasitic effects has resulted in a reduction of insertion loss of 21 dB compared with devices fabricated using a previous method.

Towards safe operation of an active retinal prosthesis during functional MRI and diffusion tensor imaging.

OBJECTIVE: To determine if the Argus II retinal prosthesis can operate during functional MRI (fMRI) and diffusion tensor imaging (DTI) acquisitions and if currents induced in the prosthesis by imaging are at safe levels. MATERIALS AND METHODS: One Argus II retinal prosthesis was modified to enable current measurements during imaging. Active electronics were modified to enable operation during scans. Induced current was measured during diagnostic scans, which were previously shown to be safe for implanted patients, and during fMRI and DTI scans. All measurements were performed using an ASTM phantom to ensure reproducible placement. RESULTS: The prosthesis was able to maintain communication with the external RF coil during the fMRI and DTI scans except briefly during pre-scans. Current levels induced during fMRI and DTI scans were consistently below those measured during diagnostic scans. CONCLUSIONS: fMRI and DTI may be safely performed while the Argus II retinal prosthesis is operating.

Development of multistage magnetic deposition microscopy.

Magnetic deposition microscropy (MDM) combines magnetic deposition and optical analysis of magnetically tagged cells into a single platform. Our multistage MDM uses enclosed microfabricated channels and a magnet assembly comprising four zones in series. The enclosed channels alleviate the problem plaguing previous versions of MDM: scouring of the cell deposition layer by the air-liquid interface as the channel is drained. The four-zone magnet assembly was designed to maximize capture efficiency, and experiments yielded total capture efficiencies of >99% of fluorescent- and magnetically-labeled Jurkat cells at reasonable throughputs (10(3) cells/min). A digital image processing protocol was developed to measure the average pixel intensities of the deposited cells in different zones, indicative of the marker expression. Preliminary findings indicate that the multistage MDM may be suitable for depositing cells and particles in successive zones according to their magnetic properties (e.g., magnetic susceptibilities or magnetophoretic mobilities). The overall goal is to allow the screening of multiple disease conditions in a single platform.

High-Performance Silicon Nanopore Hemofiltration Membranes.

Silicon micromachining provides the precise control of nanoscale features that can be fundamentally enabling for miniaturized, implantable medical devices. Concerns have been raised regarding blood biocompatibility of silicon-based materials and their application to hemodialysis and hemofiltration. A high-performance ultrathin hemofiltration membrane with monodisperse slit-shaped pores was fabricated using a sacrificial oxide technique and then surface-modified with poly(ethylene glycol) (PEG). Fluid and macromolecular transport matched model predictions well. Protein adsorption, fouling, and thrombosis were significantly inhibited by the PEG. The membrane retained hydraulic permeability and molecular selectivity during a 90 hour hemofiltration experiment with anticoagulated bovine whole blood. This is the first report of successful prolonged hemofiltration with a silicon nanopore membrane. The results demonstrate feasibility of renal replacement devices based on these membranes and materials.

A preliminary biomechanical evaluation in a simulated spinal fusion model. Laboratory investigation.

OBJECT: A preliminary in vitro biomechanical study was conducted to determine if the pressure at a bone graft-mortise interface and the load transmitted along a ventral cervical plate could be used as parameters to assess fusion status. METHODS: An interbody bone graft and a ventral plate were placed at the C3-4 motion segment in six fresh cadaveric goat spines. Polymethylmethacrylate (PMMA) was used to simulate early bone fusion at the bone graft site. The loads along the plate and the simultaneous pressures induced at the graft-endplate interfaces were monitored during simulated stages of bone healing. Each specimen was nondestructively tested in compression loading while the pressures and loads at the graft site were recorded continuously. Each specimen was tested under five conditions (Disc, Graft, Plate, PMMA, and Removal). RESULTS: The pressure at the interface of the bone graft and vertebral endplate did not change significantly with the addition of the ventral plate. The interface pressure and segmental stiffness did increase following PMMA augmentation of the bone graft (simulating an intermediate phase of bone fusion). The load transmitted along the ventral plate in compression increased after the addition of the bone graft, but decreased after PMMA augmentation. Thus, there was an increase in pressure at the graft-endplate interface and a decrease in load transferred along the ventral plate after the simulation of bone fusion. Upon removal of the ventral plate, the simulated fusion bore most of the axial load, thus explaining a further increase in graft site pressure. CONCLUSIONS: These observations support the notions of load sharing and the redistribution of loads occurring during and after bone graft incorporation. In the clinical setting, these parameters may be useful in the assessment of fusion after spine surgery. Although feasibility has been demonstrated in this preliminary study, further research is needed.

Catheter-Mounted CMOS Front-Ends for Broadband Intravascular Ultrasonic Imaging.

Intravascular ultrasonic (IVUS) imaging catheters currently use ceramic piezoelectric transducers to form radial images of blood vessel walls. Further improvements in image quality may be enabled through Capacitive and Piezoelectric Micromachined Ultrasonic Transducers (CMUTs and PMUTs). Polymer PMUTs offer many benefits in imaging quality, however, the low acoustic sensitivity and high electrical impedance of polymer PMUTs prevents unbuffered use with 50-Ω IVUS cables. Here, we present the design and optimization of an integrated CMOS front-end specifically designed to be integrated on a 0.8-mm imaging catheter. A series-duplexer topology with active limiter was selected for simplicity of integration, and compatibility with conventional high-voltage IVUS pulser-receiver systems. Noise optimization of the front-end revealed that an impedance-matching optimum exists, which balances the system parasitic capacitances relative to the transducer complex impedance. Optimized design equations compared favorably to simulation results in predicting front-end bandwidth and transducer-referred SNR. Large-signal transient simulation showed that the front-end can withstand 100-V pulses while recovering in 240 ns for echo reception. The integrated front-end occupied 0.74 × 1.8 mm die area (including large solder-bump bond pads), with an estimated transducer-referred noise floor of 6.5 n V R M S / H z over a 100-MHz imaging bandwidth.

Evaluation of silicon nanoporous membranes and ECM-based microenvironments on neurosecretory cells.

Understanding the interactions between microfabricated synthetic interfaces and cultured cells expressing a neuronal phenotype are critical for advancing research in the field of neural engineering such as neural recording and stimulation and neural microdevice interactions with the human brain. Here we explore the integration of these two components for therapeutic applications of neural prostheses. Microfabricated silicon nanoporous membranes were investigated for their effects on survival, proliferation, and differentiation of the well-known PC12 clonal line. Specifically, cell morphology, examined through fluorescence staining, were comparable in many respects on both silicon membrane and widely-used polystyrene culture surfaces. The attachment and differentiation of PC12 cells cultured on collagen and laminin-modified membranes and standard tissue culture surfaces were similar. Lastly, the differentiation response and tyrosine hydroxylase activity of PC12 cells embedded in a type I collagen matrix on experimental membrane substrates while exposed to NGF were significant and indistinguishable from tissue-culture polystyrene (TC-PS) surfaces. Results from this research suggest that microfabricated silicon nanoporous membranes may be useful, biocompatible permselective structures for neuroprosthetic applications and that collagen may be a useful immobilizing matrix for PC12 cells loaded in implantable macroencapsulation devices designed for the treatment of neurodegenerative disorders.

Electrochemical detection and characterization of proteins.

On-line detection of serum proteins is of clinical relevance, in detecting leaks and biofouling in hemofiltration equipment, biofilm growth on prosthetic devices, or hemolysis within a prosthetic or therapeutic device. Electrochemical impedance spectroscopy (EIS) and cyclic voltammetry (CV) were employed to detect and analyze micromolar concentrations of four globular proteins of clinical importance. CV testing showed that identification and quantification of each of these proteins was possible through analysis of current changes at specific potentials. Preliminary CV studies into the contamination of Bovine Serum Albumin with a microgram amount of one of the other three proteins illustrated that direct detection of the contaminant protein was possible. The analysis of the EIS data demonstrated that with increase in relative concentration of proteins, the amount of electroactive proteins adsorption at the interface increases, leading to increase in surface charge density and capacitance, especially for lower molecular weight proteins. The impedance data was used to determine the values of Gibbs adsorption energy, adsorption coefficients for the four proteins, and develop an equivalent circuit model for the protein-containing solutions.

Orthogonal-coil RF probe for implantable passive sensors.

A versatile orthogonal-coil radio frequency (RF) probe suitable for detecting the resonant frequency of miniature implantable passive sensors has been designed and tested. The probe sensitivity has been tested using printed-circuit spiral inductors of various sizes (3-15 mm) in series with discrete surface-mount capacitors designed to resonate over a range of frequencies (50-200 MHz). Close agreement between theoretical calculations and experimental results has been obtained. An equation is derived for transmit/receive (T/R) isolation that agrees with experimental measurements over the frequency range 1-500 MHz. The probe includes an additional coil to compensate for the effect of eddy currents in the human body on the probe. T/R isolation of at least 90 dB over the frequency range 1-100 MHz can be achieved when the probe is placed in close proximity to the human body.

Continuous flow magnetic cell fractionation based on antigen expression level.

Cell separation is important in medical and biological research and plays an increasingly important role in clinical therapy and diagnostics, such as rare cancer cell detection in blood. The immunomagnetic labeling of cells with antibodies conjugated to magnetic nanospheres gives rise to a proportional relationship between the number of magnetic nanospheres attached to the cell and the cell surface marker number. This enables the potential fractionation of cell populations by magnetophoretic mobility (MM). We exploit this feature with our apparatus, the Dipole Magnet Flow Fractionator (DMFF), which consists of an isodynamic magnetic field, an orthogonally-oriented thin ribbon of cell suspension in continuous sheath flow, and ten outlet flows. From a sample containing a 1:1 mixture of immunomagnetically labeled (label+) and unlabeled (label-) cells, we achieved an increase in enrichment of the label+ cell fraction with increasing outlet numbers in the direction of the magnetic field gradient (up to 10-fold). The total recovery of the ten outlet fractions was 90.0+/-7.7%. The mean MM of label+ cells increased with increasing outlet number by up to a factor of 2.3. The postulated proportionality between the number of attached magnetic beads and the number of cell surface markers was validated by comparison of MM measured by cell tracking velocimetry (CTV) with cell florescence intensity measured by flow cytometry.

Differentiated growth of human renal tubule cells on thin-film and nanostructured materials.

Over 300,000 Americans are dependent on hemodialysis as treatment for renal failure, and kidney transplantation is limited by scarcity of donor organs. This shortage has prompted research into tissue engineering of renal replacement therapy. Existing bioartificial kidneys are large and their use labor intensive, but they have shown improved survival compared to conventional therapy in preclinical studies and an US Food and Drug Administration-approved phase 2 clinical trial. This hybrid technology will require miniaturization of hemofilters, cell culture substrates, sensors, and integration of control electronics. Using the same harvesting and isolation techniques used in preparing bioartificial kidneys for clinical use, we characterized human renal tubule cell growth on a variety of silicon and related thin-film material substrates commonly used in the construction of microelectromechanical systems (MEMS), as well as novel silicon nanopore membranes (SNMs). Human cortical tubular epithelial cells (HCTC) were seeded onto samples of single-crystal silicon, polycrystalline silicon, silicon dioxide, silicon nitride, SU-8 photoresist, SNMs, and polyester tissue culture inserts, and grown to confluence. The cells formed confluent monolayers with tight junctions and central cilia. Transepithelial resistances were similar between SNMs and polyester membranes. The differentiated growth of human tubular epithelial cells on MEMS materials strongly suggests that miniaturization of the existing bioartificial kidney will be feasible, paving the way for widespread application of this novel technology.

Inductive passive sensor for intraparenchymal and intraventricular monitoring of intracranial pressure.

Accurate measurement of intracranial hypertension is crucial for the management of elevated intracranial pressure (ICP). Catheter-based intraventricular ICP measurement is regarded as the gold standard for accurate ICP monitoring. However, this method is invasive, time-limited, and associated with complications. In this paper, we propose an implantable passive sensor that could be used for continuous intraparenchymal and intraventricular ICP monitoring. Moreover, the sensor can be placed simultaneously along with a cerebrospinal fluid shunt system in order to monitor its function. The sensor consists of a flexible coil which is connected to a miniature pressure sensor via an 8-cm long, ultra-thin coaxial cable. An external orthogonal-coil RF probe communicates with the sensor to detect pressure variation. The performance of the sensor was evaluated in an in vitro model for intraparenchymal and intraventricular ICP monitoring. The findings from this study demonstrate proof-of-concept of intraparenchymal and intraventricular ICP measurement using inductive passive pressure sensors.