Mohamad G. Ghosn

Live imaging of blood flow in mammalian embryos using Doppler swept-source optical coherence tomography

Studying hemodynamic changes during early mammalian embryonic development is critical for further advances in prevention, diagnostics, and treatment of congenital cardiovascular (CV) birth defects and diseases. Doppler optical coherence tomography (OCT) has been shown to provide sensitive measurements of blood flow in avian and amphibian embryos. We combined Doppler swept-source optical coherence tomography (DSS-OCT) and live mouse embryo culture to analyze blood flow dynamics in early embryos. SS-OCT structural imaging was used for the reconstruction of embryo morphology and the orientation of blood vessels, which is required for calculating flow velocity from the Doppler measurements. Spatially and temporally resolved blood flow profiles are presented for the dorsal aorta and a yolk sac vessel in a 9.5-day embryo. We demonstrate that DSS-OCT can be successfully used for structural analysis and spatially and temporally resolved hemodynamic measurements in developing early mammalian embryos.

Depth-resolved monitoring of glucose diffusion in tissues by using optical coherence tomography

We demonstrate the capability of the optical coherence tomography (OCT) technique for depth-resolved monitoring and quantifying of glucose diffusion in fibrous tissues (sclera). The depth-resolved and average permeability coefficients of glucose were calculated. We found that the glucose diffusion rate is not uniform throughout the tissue and is increased from approximately 2.39+/-0.73 x 10(-6) cm/s at the epithelial side to 8.63+/-0.27 x 10(-6) cm/s close to the endothelial side of the sclera. Results demonstrated that the OCT technique is capable of depth-resolved monitoring and quantification of glucose diffusion in sclera with a resolution of approximately 40 mum.

Optical Clearing for OCT Image Enhancement and In-Depth Monitoring of Molecular Diffusion

In this paper, we overview the basic principles, recent results, advantages, limitations, and future of the optical clearing method in application to many fields of biology and medicine. We also discuss the possibility of noninvasive assessment of molecular diffusion in tissues using the optical coherence tomography technique. Issues of safety and toxicity of application of different endogenous and exogenous molecules to tissues are outlined.

Differential permeability rate and percent clearing of glucose in different regions in rabbit sclera

Imaging of biological tissues with optical coherence tomography (OCT) poses a great interest for its capability to noninvasively outline subsurface microstructures within tissues. However, a major limitation for many optical imaging techniques is inadequate depth penetration of light in turbid media, which is bounded to just a few millimeters. There have been several attempts to improve light penetration depth in biological tissues, including application of different tissue optical clearing methods. In this study, an aqueous solution of glucose (40%) is added to rabbit sclera in vitro, where depth-resolved permeability coefficients and optical clearing are calculated with OCT. The permeability rate in regions in the upper 80- to 100-microm region is found to be different from that of regions in the deeper 100-microm region: (6.01+/-0.37)x10(-6) cmsec and (2.84+/-0.68)x10(-5) cmsec, respectively. A difference in percent clearing is also noted. Optical clearing of the upper region is about 10% and increased to 17 to 22% in the one beneath. These results demonstrate the capability of OCT-based methods to not only measure the diffusion rate and optical clearing of a tissue, but also its ability of functional differentiation between layers of epithelial tissues.

Monitoring of glucose permeability in monkey skin in vivo using Optical Coherence Tomography.

Topical trans-dermal delivery of drugs has proven to be a promising route for treatment of many dermatological diseases. The aim of this study is to monitor and quantify the permeability rate of glucose solutions in rhesus monkey skin noninvasively in vivo as a primate model for drug diffusion. A time-domain Optical Coherence Tomography (OCT) system was used to image the diffusion of glucose in the skin of anesthetized monkeys for which the permeability rate was calculated. From 5 experiments on 4 different monkeys, the permeability for glucose-20% was found to be (4.41 +/- 0.28) 10(-6) cm/sec. The results suggest that OCT might be utilized for the noninvasive study of molecular diffusion in the multilayered biological tissues in vivo.

Permeability of hyperosmotic agent in normal and atherosclerotic vascular tissues

Noninvasive cardiovascular imaging could lead to the early detection and timely treatment of complex atherosclerotic lesions responsible for major cardiovascular events. Recent investigations have suggested that optical coherence tomography (OCT) is an ideal diagnostic tool due to the high resolution this technology achieves in discriminating the different features of atherosclerotic lesions based on structural imaging. We explore the capability of OCT for functional imaging of normal and atherosclerotic aortic tissues based on time- and depth-resolved quantification of the permeability of biomolecules through these tissues. The permeability coefficient of 20% aqueous solution of glucose was found to be (6.80+/-0.18)x10(-6) cms in normal aortas and (2.69+/-0.42)x10(-5) cms in aortas with atherosclerotic disease. The results suggest that this new OCT functional imaging method-the assessment of the permeability coefficients of various physiologically neutral biomolecules in vascular tissues-could assist in early diagnosing and detecting the different components of atherosclerotic lesions.

ASSESSMENT OF TISSUE OPTICAL CLEARING AS A FUNCTION OF GLUCOSE CONCENTRATION USING OPTICAL COHERENCE TOMOGRAPHY.

One of the major challenges in imaging biological tissues using optical techniques, such as optical coherence tomography (OCT), is the lack of light penetration due to highly turbid structures within the tissue. Optical clearing techniques enable the biological samples to be more optically homogeneous, allowing for deeper penetration of light into the tissue. This study investigates the effect of optical clearing utilizing various concentrations of glucose solution (10%, 30%, and 50%) on porcine skin. A gold-plated mirror was imaged beneath the tissue and percentage clearing was determined by monitoring the change in reflected light intensity from the mirror over time. The ratio of percentage clearing per tissue thickness for 10%, 30% and 50% glucose was determined to be 4.7 ± 1.6% mm(-1) (n = 6), 10.6 ± 2.0% mm(-1) (n = 7) and 21.8 ± 2.2% mm(-1) (n = 5), respectively. It was concluded that while higher glucose concentration has the highest optical clearing effect, a suitable concentration should be chosen for the purpose of clearing, considering the osmotic stress on the tissue sample.

High-Fat Feeding-Induced Hyperinsulinemia Increases Cardiac Glucose Uptake and Mitochondrial Function Despite Peripheral Insulin Resistance

In obesity, reduced cardiac glucose uptake and mitochondrial abnormalities are putative causes of cardiac dysfunction. However, high-fat diet (HFD) does not consistently induce cardiac insulin resistance and mitochondrial damage, and recent studies suggest HFD may be cardioprotective. To determine cardiac responses to HFD, we investigated cardiac function, glucose uptake, and mitochondrial respiration in young (3-month-old) and middle-aged (MA) (12-month-old) male Ldlr(-/-) mice fed chow or 3 months HFD to induce obesity, systemic insulin resistance, and hyperinsulinemia. In MA Ldlr(-/-) mice, HFD induced accelerated atherosclerosis and nonalcoholic steatohepatitis, common complications of human obesity. Surprisingly, HFD-fed mice demonstrated increased cardiac glucose uptake, which was most prominent in MA mice, in the absence of cardiac contractile dysfunction or hypertrophy. Moreover, hearts of HFD-fed mice had enhanced mitochondrial oxidation of palmitoyl carnitine, glutamate, and succinate and greater basal insulin signaling compared with those of chow-fed mice, suggesting cardiac insulin sensitivity was maintained, despite systemic insulin resistance. Streptozotocin-induced ablation of insulin production markedly reduced cardiac glucose uptake and mitochondrial dysfunction in HFD-fed, but not in chow-fed, mice. Insulin injection reversed these effects, suggesting that insulin may protect cardiac mitochondria during HFD. These results have implications for cardiac metabolism and preservation of mitochondrial function in obesity.

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.

Effect of temperature on permeation of low-density lipoprotein particles through human carotid artery tissues.

Quantification of the diffusion of small molecules and large lipid transporting lipoproteins across arterial tissues could be useful in elucidating the mechanism(s) of atherosclerosis. Optical coherence tomography (OCT) was used to determine the effect of temperature on the rate of diffusion of glucose and low-density lipoproteins (LDL) in human carotid endarterectomy tissue in vitro. The permeability rate for glucose was calculated to be (3.51 +/- 0.27) x 10(-5) cm/s (n = 13) at 20 degrees C, and (3.70 +/- 0.44) x 10(-5) cm/s (n = 5) at 37 degrees C; for LDL the rate was (2.42 +/- 0.33) x 10(-5) cm/s (n = 5) at 20 degrees C and (4.77 +/- 0.48) x 10(-5) cm/s (n = 7) at 37 degrees C, where n is the number of samples. These results demonstrate that temperature does not significantly influence the permeation of small molecules (e.g. glucose), however, raising the temperature does significantly increase the permeation of LDL. These results provide new information about the capacity of an atherogenic lipoprotein to traverse the intimal layer of the artery. These results also demonstrate the potential of OCT for elucidating the dynamics of lipoprotein perfusion across the arterial wall.