Saba H. Syed

Hemodynamic measurements from individual blood cells in early mammalian embryos with Doppler swept source OCT

The most common and lethal birth defects affect the cardiovascular (CV) system. The mouse is a superior model for identifying and understanding mammalian CV birth defects, but there is a great need for tools that can detect early and subtle deficiencies in cardiac function in mouse embryos. We combined swept source optical coherence tomography (SS-OCT) with live mouse embryo culture protocols to generate structural two-dimensional and three-dimensional imaging and hemodynamic measurements in a live 8.5 day embryo just a few hours after the beginning of a heartbeat. Our data show that individual circulating blood cells can be visualized with structural SS-OCT, and using Doppler SS-OCT the velocity of single moving blood cells were measured during different phases of the heartbeat cycle. These results demonstrate that Doppler SS-OCT is an extremely useful tool for structural and hemodynamic analysis at the earliest stages of mammalian blood circulation.

Optical coherence tomography for high-resolution imaging of mouse development in utero

Although the mouse is a superior model to study mammalian embryonic development, high-resolution live dynamic visualization of mouse embryos remain a technical challenge. We present optical coherence tomography as a novel methodology for live imaging of mouse embryos through the uterine wall thereby allowing for time lapse analysis of developmental processes and direct phenotypic analysis of developing embryos. We assessed the capability of the proposed methodology to visualize structures of the living embryo from embryonic stages 12.5 to 18.5 days postcoitus. Repetitive in utero embryonic imaging is demonstrated. Our work opens the door for a wide range of live, in utero embryonic studies to screen for mutations and understand the effects of pharmacological and toxicological agents leading to birth defects.

Optical coherence tomography for live phenotypic analysis of embryonic ocular structures in mouse models

Mouse models of ocular diseases provide a powerful resource for exploration of molecular regulation of eye development and pre-clinical studies. Availability of a live high-resolution imaging method for mouse embryonic eyes would significantly enhance longitudinal analyses and high-throughput morphological screening. We demonstrate that optical coherence tomography (OCT) can be used for live embryonic ocular imaging throughout gestation. At all studied stages, the whole eye is within the imaging distance of the system and there is a good optical contrast between the structures. We also performed OCT eye imaging in the embryonic retinoblastoma mouse model Pax6-SV40 T-antigen, which spontaneously forms lens and retinal lesions, and demonstrate that OCT allows us to clearly differentiate between the mutant and wild type phenotypes. These results demonstrate that OCTin utero imaging is a potentially useful tool to study embryonic ocular diseases in mouse models.

Quantification of molecular diffusion in arterial tissues with optical coherence tomography and fluorescence microscopy

Alternations in vascular permeability for different molecules, drugs, and contrast agents might be a significant early marker of development of various diseases such as atherosclerosis. However, up to date experimental studies of molecular diffusion across vascular wall have been limited. Recently, we demonstrated that the Optical Coherence Tomography (OCT) technique could be applied for noninvasive and nondestructive quantification of molecular diffusion in different biological tissues. However, the viability of the OCT-based assessment of molecular diffusion should be validated with established methods. This study focused on comparing molecular diffusion rates in vascular tissues measured with OCT and standard fluorescent microscopy. Noninvasive quantification of tetramethylrhodamine (fluorescent dye) permeability in porcine vascular tissues was performed using a fiber-based OCT system. Concurrently, standard histological examination of dye diffusion was performed and quantified with fluorescent microscopy. The permeability of tetramethylrhodamine was found to be (2.08 ± 0.31) × 10−5 cm/s with the fluorescent technique (n = 8), and (2.45 ± 0.46) × 10−5 cm/s with the OCT (n = 3). Good correlation between permeability rates measured by OCT and histology was demonstrated, suggesting that the OCT-based method could be used for accurate, nondestructive assessment of molecular diffusion in multilayered tissues.

Optical coherence tomography guided microinjections in live mouse embryos: high-resolution targeted manipulation for mouse embryonic research.

Abstract. The ability to conduct highly localized delivery of contrast agents, viral vectors, therapeutic or pharmacological agents, and signaling molecules or dyes to live mammalian embryos is greatly desired to enable a variety of studies in the field of developmental biology, such as investigating the molecular regulation of cardiovascular morphogenesis. To meet such a demand, we introduce, for the first time, the concept of employing optical coherence tomography (OCT)-guide microinjections in live mouse embryos, which provides precisely targeted manipulation with spatial resolution at the micrometer scale. The feasibility demonstration is performed with experimental studies on cultured live mouse embryos at E8.5 and E9.5. Additionally, we investigate the OCT-guided microinjection of gold–silica nanoshells to the yolk sac vasculature of live cultured mouse embryos at the stage when the heart just starts to beat, as a potential approach for dynamic assessment of cardiovascular form and function before the onset of blood cell circulation. Also, the capability of OCT to quantitatively monitor and measure injection volume is presented. Our results indicate that OCT-guided microinjection could be a useful tool for mouse embryonic research.

Time dependent changes in aortic tissue during cold storage in physiological solution.

BACKGROUND Stored vascular tissues are employed in biomedical research for studies in imaging, in biomechanics, and/or in assessing vessel diseases. In the present study, the stability of aortic tissue in phosphate buffer saline (PBS) at 4°C was monitored over a course of 10 days as determined by the rate of glucose permeation measured by optical coherence tomography (OCT) and validated by histology. METHODS AND RESULTS The initial mean permeability through fresh porcine aorta was (2.32 ± 0.46)× 10(-5)cm/s (n=5); after maintaining the tissue at 4°C for 10 days, the mean rate was (7.37 ± 0.41)× 10(-5)cm/s (n=4), an increase of nearly 300%. A z-test verified that a significant change in the permeability rate (p<0.05) had occurred after 4 days of 4°C storage. Histology was used to quantify changes in tissue pore area. The increase in average pore area paralleled the increase in permeability rate over 10 days. CONCLUSIONS These results suggest that (1) the structural integrity of aortic tissue at 4°C is retained for at least the first three days after resection and (2) OCT is a powerful technology well suited for evaluating tissue structural integrity over time. GENERAL SIGNIFICANCE Functional OCT imaging provides for a noninvasive and quantitative technique in determining the structural integrity of aortic tissue stored at 4°C. This modality may be used for assessing the efficacy of other preservation techniques.

Permeation of human plasma lipoproteins in human carotid endarterectomy tissues: measurement by optical coherence tomography.

Atherosclerosis is an inflammatory process occurring in arterial tissue, involving the subintimal accumulation of LDL. Measurement of the rate at which LDL and other lipoproteins, such as HDL and VLDL, enter and exit the tissue can provide insight into the mechanisms involved in the development of atherosclerotic lesions. Permeation of VLDL, LDL, HDL, and glucose was measured for both normal and atherosclerotic human carotid endarterectomy tissues (CEA) at 20°C and 37°C using optical coherence tomography (OCT). The rates for LDL permeation through normal CEA tissue were (3.16 ± 0.37) × 10−5 cm/s at 20°C and (4.77 ± 0.48) × 10−5 cm/s at 37°C, significantly greater (P < 0.05) than the rates for atherosclerotic CEA tissue at these temperatures [(1.97 ± 0.34) × 10−5 cm/s at 20°C and (2.01 ± 0.23) × 10−5 cm/s at 37°C]. This study effectively used OCT to measure the rates at which naturally occurring lipoproteins enter both normal and diseased carotid intimal tissue.

Real-Time Imaging of Circulating Individual Blood Cells in Mammalian Embryos with Doppler SSOCT

Congenital cardiovascular (CV) defects are present in approximately 1% of live births. Moreover, among deaths due to birth defects, cardiovascular failures are the most likely. Therefore, characterization of mammalian embryonic cardio dynamics and understanding of the hemodynamic changes that occur during embryonic development is crucial for improving diagnostics, prevention, and treatment of cardiovascular defects and diseases. In this work, we combined Swept Source Optical Coherence Tomography (SSOCT) with live mouse embryo culture protocols to generate structural 2-D and 3-D imaging and hemodynamic measurements in live 8.5 day embryos. Our data show that individual circulating blood cells can be visualized with structural SSOCT and the velocity of single moving blood cells were measured during different phases of heartbeat cycle with Doppler SSOCT. These results demonstrate that this imaging modality could be an extremely useful tool for structural and hemodynamic analysis at the earliest stages of mammalian blood circulation.

OCT guided microinjections for mouse embryonic research

Optical coherence tomography (OCT) is gaining popularity as live imaging tool for embryonic research in animal models. Recently we have demonstrated that OCT can be used for live imaging of cultured early mouse embryos (E7.5-E10) as well as later stage mouse embryos in utero (E12.5 to the end of gestation). Targeted delivery of signaling molecules, drugs, and cells is a powerful approach to study normal and abnormal development, and image guidance is highly important for such manipulations. Here we demonstrate that OCT can be used to guide microinjections of gold nanoshell suspensions in live mouse embryos. This approach can potentially be used for variety of applications such as guided injections of contrast agents, signaling molecules, pharmacological agents, cell transplantation and extraction, as well as other image-guided micromanipulations. Our studies also reveal novel potential for gold nanoshells in embryonic research.

Mouse embryo manipulations with OCT guidance

Optical coherence tomography (OCT) is a three-dimensional, non-invasive optical imaging technique that relies on low-coherence interferometry. OCT has the capability of imaging 2 – 3 mm into tissue, which enables imaging of deeper structures within the embryo with a relatively high spatial resolution (2 - 15μm). Within the past decade, OCT has been increasingly used as a live imaging tool for embryonic cardiovascular research in several animal models. Research in our lab has recently shown that OCT can be used in combination with embryo culture for the visualization of early mammalian cardiovascular development (E7.5 – E10.0). Here, we demonstrate that OCT can be used for the guided microinjection of gold-silica nanoshell suspension into the cardiovascular system in live embryos without deleterious effect. This approach shows a promising application for the OCT guided delivery of contrast agents, viral vectors, therapeutic or pharmacological agents, signaling molecules or dyes to specific organ systems or tissues in live embryos and demonstrates a great potential for gold-silica nanoshells as a contrast agent in embryonic studies.