Kevin E. Loewke

Non-invasive imaging of human embryos before embryonic genome activation predicts development to the blastocyst stage

We report studies of preimplantation human embryo development that correlate time-lapse image analysis and gene expression profiling. By examining a large set of zygotes from in vitro fertilization (IVF), we find that success in progression to the blastocyst stage can be predicted with >93% sensitivity and specificity by measuring three dynamic, noninvasive imaging parameters by day 2 after fertilization, before embryonic genome activation (EGA). These parameters can be reliably monitored by automated image analysis, confirming that successful development follows a set of carefully orchestrated and predictable events. Moreover, we show that imaging phenotypes reflect molecular programs of the embryo and of individual blastomeres. Single-cell gene expression analysis reveals that blastomeres develop cell autonomously, with some cells advancing to EGA and others arresting. These studies indicate that success and failure in human embryo development is largely determined before EGA. Our methods and algorithms may provide an approach for early diagnosis of embryo potential in assisted reproduction.

Dynamic blastomere behaviour reflects human embryo ploidy by the four-cell stage

Previous studies have demonstrated that aneuploidy in human embryos is surprisingly frequent with 50–80% of cleavage-stage human embryos carrying an abnormal chromosome number. Here we combine non-invasive time-lapse imaging with karyotypic reconstruction of all blastomeres in four-cell human embryos to address the hypothesis that blastomere behaviour may reflect ploidy during the first two cleavage divisions. We demonstrate that precise cell cycle parameter timing is observed in all euploid embryos to the four-cell stage, whereas only 30% of aneuploid embryos exhibit parameter values within normal timing windows. Further, we observe that the generation of human embryonic aneuploidy is complex with contribution from chromosome-containing fragments/micronuclei that frequently emerge and may persist or become reabsorbed during interphase. These findings suggest that cell cycle and fragmentation parameters of individual blastomeres are diagnostic of ploidy, amenable to automated tracking algorithms, and likely of clinical relevance in reducing transfer of embryos prone to miscarriage.

In vivo near-infrared dual-axis confocal microendoscopy in the human lower gastrointestinal tract

Near-infrared confocal microendoscopy is a promising technique for deep in vivo imaging of tissues and can generate high-resolution cross-sectional images at the micron-scale. We demonstrate the use of a dual-axis confocal (DAC) near-infrared fluorescence microendoscope with a 5.5-mm outer diameter for obtaining clinical images of human colorectal mucosa. High-speed two-dimensional en face scanning was achieved through a microelectromechanical systems (MEMS) scanner while a micromotor was used for adjusting the axial focus. In vivo images of human patients are collected at 5 frames/sec with a field of view of 362×212 μm(2) and a maximum imaging depth of 140 μm. During routine endoscopy, indocyanine green (ICG) was topically applied a nonspecific optical contrasting agent to regions of the human colon. The DAC microendoscope was then used to obtain microanatomic images of the mucosa by detecting near-infrared fluorescence from ICG. These results suggest that DAC microendoscopy may have utility for visualizing the anatomical and, perhaps, functional changes associated with colorectal pathology for the early detection of colorectal cancer.

In Vivo Micro-Image Mosaicing

Recent advances in optical imaging have led to the development of miniature microscopes that can be brought to the patient for visualizing tissue structures in vivo. These devices have the potential to revolutionize health care by replacing tissue biopsy with in vivo pathology. One of the primary limitations of these microscopes, however, is that the constrained field of view can make image interpretation and navigation difficult. In this paper, we show that image mosaicing can be a powerful tool for widening the field of view and creating image maps of microanatomical structures. First, we present an efficient algorithm for pairwise image mosaicing that can be implemented in real time. Then, we address two of the main challenges associated with image mosaicing in medical applications: cumulative image registration errors and scene deformation. To deal with cumulative errors, we present a global alignment algorithm that draws upon techniques commonly used in probabilistic robotics. To accommodate scene deformation, we present a local alignment algorithm that incorporates deformable surface models into the mosaicing framework. These algorithms are demonstrated on image sequences acquired in vivo with various imaging devices including a hand-held dual-axes confocal microscope, a miniature two-photon microscope, and a commercially available confocal microendoscope.

Vision based 3-D shape sensing of flexible manipulators

Rigid robotic manipulators employ traditional sensors such as encoders or potentiometers to measure joint angles and determine end-effector position. Manipulators that are flexible, however, introduce motions that are much more difficult to measure. This is especially true for continuum manipulators that articulate by means of material compliance. In this paper, we present a vision based system for quantifying the 3-D shape of a flexible manipulator in real-time. The sensor system is validated for accuracy with known point measurements and for precision by estimating a known 3-D shape. We present two applications of the validated system relating to the open-loop control of a tendon driven continuum manipulator. In the first application, we present a new continuum manipulator model and use the sensor to quantify 3-D performance. In the second application, we use the shape sensor system for model parameter estimation in the absence of tendon tension information.

3-D Near-Infrared Fluorescence Imaging Using an MEMS-Based Miniature Dual-Axis Confocal Microscope

We demonstrate a fast miniature microelectro-mechanical-system-based near-infrared fluorescence dual-axis confocal microscope in a 10-mm-diameter package for 3-D imaging in both ex vivo and in vivo samples. The miniature microscope, while in contact with the targeted tissue, can reveal subsurface structure or anatomy as deep as 300 mum. The lateral and axial resolutions are 5 and 7 mum, respectively. Real-time en face mosaicing image of in vivo human skin is demonstrated to enlarge the overall FOV to be over 3 mm at acquisition frame rate of 5 frames/s.

An Optical Fiber Proximity Sensor for Haptic Exploration

This paper presents the design of an optical fiber proximity sensor for haptic exploration with a robotic finger. The sensor uses emitter and receiver optical fiber pairs to measure the intensity of light reflected off surrounding objects in a 2-D workspace. We present the design and construction a 32-point sensor array mounted within a 36 mm diameter finger and describe software techniques to process data acquired by an inexpensive Web cam. We experimentally characterize the sensor performance and demonstrate applications for haptic exploration such as pre-contact velocity reduction and non-contact contour following based on object curvature.

Force control of a permanent magnet for minimally-invasive procedures

Magnetic actuation can be used for minimally invasive control of medical devices such as robotic catheters. However, current systems that use large permanent magnets are limited in their ability to modulate the magnetic force. In this paper we present a proof-of-concept system for closed-loop force control of a permanent magnet using shielding materials. Our system consists of a device that actuates pieces of high-permeability metal to redirect magnetic lines of flux. This is used to regulate the attractive force exerted by a large controlling magnet on a smaller moving magnet. We compare the performance of our system to FEA simulations and present experimental results for constant-force control at forces and distances that are medically relevant.

Deformable Image Mosaicing for Optical Biopsy

Traditional image mosaicing usually relies on rigid image transformations. In many medical applications, however, tissue deformation during image acquisition or 3D parallax effects may require nonrigid transformations in the mosaicing process. This paper presents a new method that integrates deformable surface models into the image mosaicing algorithms. Our approach has two main contributions. First, we present a global alignment algorithm to efficiently deal with accumulated image registration errors. Second, we introduce a local alignment algorithm to accommodate local scene deformations. These two problems are integrated into a single optimization problem that simultaneously recovers the motion of the camera as well as the structure of the scene. Our approach is demonstrated on simulations, images from a hand-held digital camera, and microscopic images acquired with a micro-endoscope.

The Role of Time-Lapse Microscopy in Stem Cell Research and Therapy

Stem cell therapy holds enormous potential for treating a wide range of genetic and sporadic degenerative disorders. However, one of the major hurdles facing stem cell therapy is the ability to assess cell fate or outcome prior to transplantation. Recent studies have shown that time-lapse microscopy may be a useful tool to assess cell fate via observation of dynamic behavior at the single-cell and population levels. The ideal embodiment of time-lapse microscopy would be a high-throughput, noninvasive device that can identify stem cells that form nontumorigenic differentiated progeny capable of integration into mature tissues. Such technologies are on the horizon and hold promise for clinical and therapeutic applications.