Clinical evaluation of a novel subcutaneous lactate monitor.

Lactate levels are commonly used as an indirect measure to assess metabolic stress in clinical conditions like sepsis. Dynamic lactate measurements are recommended to assess and guide treatment in patients with shock and other critical care conditions. A minimally invasive, continuous lactate monitor has potential to improve clinical decisions and patient care. The purpose of the study was to evaluate continuous lactate measurements of a novel enzymatic Continuous Lactate Monitor (CLM) developed in our laboratory. Lactate levels were monitored during incremental cycling exercise challenges as a tool for hyperlactatemia. Six healthy individuals 18-45 y/o (4 males, 2 females) participated in the study. CLM devices were inserted subcutaneously in the postero-lateral trunk below the renal angle, one hour before the exercise challenge. Each exercise challenge consisted of a 3 to 12-min warm up period, followed by up to 7, 4-min incremental workload bouts separated by rest intervals. Continuous lactate measurements obtained from CLM were compared with commercial lactate analyzer (Abbott iSTAT) measurements of venous blood (plasma) drawn from the antecubital vein. Blood was drawn at up to 25 time points spanning the duration of before exercise, during exercise, and up to 120 min post exercise. Area under the curve (AUC), and delay time were calculated to compare the CLM readings with plasma lactate concentration. Average plasma lactate concentration increased from 1.02 to 16.21 mM. Ratio of AUC derived from CLM to plasma lactate was 1.025 (0.990-1.058). Average dynamic delay time of CLM to venous plasma lactate was 5.22 min (2.87-10.35). Insertion sites examined 48 h after CLM removal did not show signs of side effects and none required medical attention upon examination. The newly developed CLM has shown to be a promising tool to continuously measure lactate concentration in a minimally invasive fashion. Results indicate the CLM can provide needed trends in lactate over time. Such a device may be used in the future to improve treatment in clinical conditions such as sepsis.

Transcutaneous Flexible Sensor for In Vivo Photonic Detection of pH and Lactate.

Clinical research shows that frequent measurements of both pH and lactate can help guide therapy and improve patient outcome. However, current methods of sampling blood pH and lactate make it impractical to take readings frequently (due to the heightened risk of blood infection and anemia). As a solution, we have engineered a subcutaneous pH and lactate sensor (PALS) that can provide continuous, physiologically relevant measurements. To measure pH, a sheet containing a pH-sensitive fluorescent dye is placed over 400 and 465 nm light-emitting diodes (LEDs) and a filter-coated photodetector. The filter-coated photodetector collects an emitted signal from the dye for each LED excitation, and the ratio of the emitted signals is used to monitor pH. To measure lactate, two sensing sheets comprising an oxygen-sensitive phosphorescent dye are each mounted to a 625 nm LED. One sheet additionally comprises the enzyme lactate oxidase. The LEDs are sequentially modulated to excite the sensing sheets, and their phase shift at the LED drive frequency is used to monitor lactate. In vitro results indicate that PALS successfully records pH changes from 6.92 to 7.70, allowing for discrimination between acidosis and alkalosis, and can track lactate levels up to 9 mM. Both sensing strategies exhibit fast rise times (< 5 min) and stable measurements. Multianalyte in vitro models of physiological disorders show that the sensor measurements consistently quantify the expected pathophysiological trends without cross talk; in vivo rabbit testing further indicates usefulness in the clinical setting.

A bench-top model of middle ear effusion diagnosed with optical tympanometry.

OBJECTIVES: To assess the validity of a bench-top model of an optical tympanometry device to diagnose in vitro model of middle ear effusion (MEE). METHODS AND MATERIALS: We illuminated an in vitro model of ear canal and tympanic membrane with broadband light and relayed remitted light to a spectrometer system. We then used our proprietary algorithm to extract spectral features that, together with our logistic regression classifiers, led us to calculate a set of simplified indices related to different middle ear states. Our model included a glass vial covered with a porcine submucosa (representing the tympanic membrane) and filled with air, water, or milk solution (representing different MEE), and a set of cover-glass slips filled with either blood (representing erythema) or cerumen. By interchanging fluid types and cover-glass slips, we made measurements on combinations corresponding to normal healthy ear and purulent or serous MEE. RESULTS: Each simulated condition had a distinct spectral profile, which was then employed by our algorithm to discriminate clean and cerumen-covered purulent and serous MEE. Two logistic purulent and serous MEE classifiers correctly classified all in vitro middle ear states with 100% accuracy assessed by leave-one-out and k-fold cross validation. CONCLUSIONS: This proof-of-concept in vitro study addressed an unmet need by introducing a device that easily and accurately can assess middle ear effusion. Future in vivo studies aimed at collecting data from clinical settings are warranted to further elucidate the validity of the technology in diagnosing pediatric acute otitis media.

Method measuring oxygen tension and transport within subcutaneous devices.

Cellular therapies hold promise to replace the implantation of whole organs in the treatment of disease. For most cell types, in vivo viability depends on oxygen delivery to avoid the toxic effects of hypoxia. A promising approach is the in situ vascularization of implantable devices which can mediate hypoxia and improve both the lifetime and utility of implanted cells and tissues. Although mathematical models and bulk measurements of oxygenation in surrounding tissue have been used to estimate oxygenation within devices, such estimates are insufficient in determining if supplied oxygen is sufficient for the entire thickness of the implanted cells and tissues. We have developed a technique in which oxygen-sensitive microparticles (OSMs) are incorporated into the volume of subcutaneously implantable devices. Oxygen partial pressure within these devices can be measured directly in vivo by an optical probe placed on the skin surface. As validation, OSMs have been incorporated into alginate beads, commonly used as immunoisolation devices to encapsulate pancreatic islet cells. Alginate beads were implanted into the subcutaneous space of Sprague­Dawley rats. Oxygen transport through beads was characterized from dynamic OSM signals in response to changes in inhaled oxygen. Changes in oxygen dynamics over days demonstrate the utility of our technology.

Lens-free computational imaging of capillary morphogenesis within three-dimensional substrates.

Endothelial cells cultured in three-dimensional (3-D) extracellular matrices spontaneously form microvessels in response to soluble and matrix-bound factors. Such cultures are common for the study of angiogenesis and may find widespread use in drug discovery. Vascular networks are imaged over weeks to measure the distribution of vessel morphogenic parameters. Measurements require micron-scale spatial resolution, which for light microscopy comes at the cost of limited field-of-view (FOV) and shallow depth-of-focus (DOF). Small FOVs and DOFs necessitate lateral and axial mechanical scanning, thus limiting imaging throughput. We present a lens-free holographic on-chip microscopy technique to rapidly image microvessels within a Petri dish over a large volume without any mechanical scanning. This on-chip method uses partially coherent illumination and a CMOS sensor to record in-line holographic images of the sample. For digital reconstruction of the measured holograms, we implement a multiheight phase recovery method to obtain phase images of capillary morphogenesis over a large FOV (24 mm2) with ≈ 1.5 µm spatial resolution. On average, measured capillary length in our method was within approximately 2% of lengths measured using a 10 × microscope objective. These results suggest lens-free on-chip imaging is a useful toolset for high-throughput monitoring and quantitative analysis of microvascular 3-D networks.

Quantification of local matrix deformations and mechanical properties during capillary morphogenesis in 3D.

Reciprocal mechanical interactions between cells and the extracellular matrix (ECM) are thought to play important instructive roles in branching morphogenesis. However, most studies to date have failed to characterize these interactions on a length scale relevant to cells, especially in three-dimensional (3D) matrices. Here we utilized two complementary methods, spatio-temporal image correlation spectroscopy (STICS) and laser optical tweezers-based active microrheology (AMR), to quantify endothelial cell (EC)-mediated deformations of individual ECM elements and the local ECM mechanical properties, respectively, during the process of capillary morphogenesis in a 3D cell culture model. In experiments in which the ECM density was systematically varied, STICS revealed that the rate at which ECs deformed individual ECM fibers on the microscale positively correlated with capillary sprouting on the macroscale. ECs expressing constitutively active V14-RhoA displaced individual matrix fibers at significantly faster rates and displayed enhanced capillary sprouting relative to wild-type cells, while those expressing dominant-negative N19-RhoA behaved in an opposite fashion. In parallel, AMR revealed a local stiffening of the ECM proximal to the tips of sprouting ECs. By quantifying the dynamic physical properties of the cell-ECM interface in both space and time, we identified a correlation linking ECM deformation rates and local ECM stiffening at the microscale with capillary morphogenesis at the macroscale.