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Valery V. Tuchin,1,2,3 Martin J. Leahy,4 Ruikang K. Wang5
1Saratov State Univ. (Russian Federation) 2Tomsk State Univ. (Russian Federation) 3Institute of Precision Mechanics and Control of the RAS (Russian Federation) 4National Univ. of Ireland, Galway (Ireland) 5Univ. of Washington (United States)
This PDF file contains the front matter associated with SPIE Proceedings Volume 11239 including the Title Page, Copyright information, Table of Contents, Introduction, and Conference Committee listing.
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In this work, using 930-nm optical coherence tomography (OCT) and reflection spectroscopy in a wide wavelength range from UV to NIR, we investigated and confirmed the concept of the possibility of fast and efficient optical clearing of tissue, in particular human skin in vivo when using various agents and enhancers of skin permeability. The combination of a number of physico-chemical factors enhancing permeability increases the OCT signal several times with more than double the penetration depth. For dark skin and in the UV wavelength range, where absorption is high, optical clearing is not so effective, but everything is equally useful.
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Here, we demonstrated that natural magnetic nanoparticles (nMNP) from magnetotactic bacteria can serve as photoacoustic high-contrast agents for single cell analysis. The main benefits of nMNP include biocompatibility and relatively low laser fluence for photoacoustic detection. These features together with safety of photoacoustics provide potential for clinical translation. Further decoration of nMNP with NIR-absorbing gold nanorods provided hybrid nanoparticles with strong NIR absorption and plasmonic effects for cancer cell theranostics. In studies in vivo, we showed that circulating tumor cells labeled with bioinspired hybrids generated transient ultrasharp photoacoustic resonances as the basis for new super-resolution photoacoustic flow cytometry in vivo.
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We apply time-domain diffuse correlation spectroscopy (TD DCS) to quantify dynamics in samples with mixed dynamics, containing both static and dynamic scatterers. We demonstrate that standard TD DCS processing is incapable to properly quantify dynamics at short source-detector distances due to the strong influence of the static component. To solve this problem, we introduce a novel model, which allows recovering the autocorrelation decay of the dynamic part properly. We then apply this novel approach in humans in vivo. We recovered the blood flow index of the leg muscle covered by the thin static turbid layer during the cuff occlusion challenge.
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Non-invasive imaging modalities, especially optical coherence tomography (OCT) are capable of providing high resolution structural and functional imaging capabilities for ophthalmic applications. Apart from functional imaging, OCT has also been used to extract functional dynamics within the microvasculature in response to changes in local environment. In mammals, various physiological processes, such as energy metabolisms, cardiovascular functions and circadian rhythms exhibit fluctuations in response to change in the local and external environment. These rhythmic oscillations within the body have been observed within the neurons, microvasculature, muscles and heart and play a vital role in modulating the biological processes and associated physiological response by an organism. Cornea is the transparent, avascular layer of the eye that controls the entry of light into the eye and also helps to refract the light onto the retina. Corneal injuries caused by various chemical, physical, and pathological stimuli damages the corneal epithelium, the stroma and the endothelium, thereby hindering its proper functioning. The maintenance of corneal transparency is vital for optimum vision, and this is ensured by the avascular cornea evenly spaced collagen fibrils of uniform diameter within the stroma, and also the level of hydration within the stromal layer. Recently, nano-sensitive OCT (nsOCT) technique has been proposed by our group that retains the high spatial frequency information in an OCT image, thereby enabling the detection of nanoscale structural alterations in in vivo imaging of tissues. In this paper, we describe nsOCT based approach to detect the dynamic/temporal structural changes within the cornea following an alkali injury using a high speed swept source OCT.
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We present an imaging-based quantitative approach for studying the localized pumping mechanism of the early tubular heart in live mouse embryos. The method relies on 4D (3D+time) imaging of cardiodynamics and hemodynamics of the embryonic heart using structural and Doppler optical coherence tomography (OCT). Our results from the mouse embryo at embryonic day 9.0 (E9.0) show an interesting relationship between the endocardial luminal areas and the localized volumetric blood flows, suggesting that a localized pressure gradient induced by the heart wall movement causes the variation of blood flows, including both the velocity magnitude and flow direction. Data provide new insights into the pumping mechanism of the mammalian tubular heart at the early developmental stage.
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The cochlea of the inner ear transduces sound energy into electrical signals that are essential for audition. This transduction is processed in nano-scale vibration of the cochlear sensory epithelia. In mammals, the epithelia contain various cells and structures: inner hair cells, outer hair cells (OHCs), Deiters’ cells, basilar membrane, reticular lamina, etc. The sound elicits vibration in all these constituents. Among them, only OHCs cell body actively and periodically changes in length in association with the vibration. The unique mechanical activity of OHCs modifies the sound elicited vibration in the epithelia with a feedback mechanism. Although the modification is considered to critically contribute to the high sensitivity and sharp tuning in hearing through sensory IHCs, the real motion of OHCs remains uncertain. Vibrometrical studies of cochlear mechanics has revealed important vibration of the cell bodies involving the epithelia. However, difference in vibration pattern of the apical and basal ends of the cell has remain uncertain due to low spatial resolution of the system and low reflectivity of the cells. We performed a spectral domain OCT (SD-OCT) vibrometry by using the modified commercial SD-OCT system. Because the broad spectral bandwidth and strong power of the light source improve a performance of OCT systems in both of imaging and vibrometry, we introduced a supercontinuum light source into the commercial system. Our system achieved cellular-level tomographic imaging and subnano-scale vibration measurement in the transparent epithelia with the recording time of 100 ms in in vivo animal.
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Pulsatile signals from the cutaneous blood flow could be informative for evaluating the health condition of an individual. One of the popular optical measuring devices, photoplethysmogram (PPG) is often used to detect the pulse signal from skin. However, the origin of the PPG signal still remains controversial. Benefiting from the non-invasive, label-free, 3D imaging tool, optical coherence tomography (OCT) is able to capture the intrinsic tissue signals at different penetration depth in high spatial and temporal resolution. Periodic pulse signal was observed by taking advantage of the optical microangiography (OMAG) algorithm which is sensitive to the motion of blood flow. The pulsatile pattern from the capillary and arteriole was successfully differentiated and their morphology showed distinct property at different local blood pressure. The pulse signal from the arteriole is more consistent and has similar waveform as the PPG signals. The result indicated that the PPG signal could be deceive by the mixing signal from the capillary bed and arterioles since it measures the total blood volume change in the plexuses. This study may shed some new light on understanding the mechanical property of how blood travel through different types of vasculature networks and elucidate its potential application in disease assessments.
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In the United States, 20% of pregnant women are estimated to smoke, thus affecting 800,000 babies annually. Maternal nicotine exposure is known to have several detrimental effects on the developing fetus including intrauterine growth restriction, perinatal mortality and morbidity, placental abruption, and other childhood disorders. In humans, studies evaluating the association between maternal cigarette smoking during pregnancy and behavioral development in offsprings have shown negative influences of nicotine on brain development. Although several studies have documented lower birth weights, morphological and behavioral changes, not much has been done evaluating the acute changes in brain vasculature after prenatal exposure to nicotine. This work uses correlation mapping optical coherence angiography (cm-OCA), a functional extension of optical coherence tomography, to evaluate changes in murine fetal brain vasculature, in utero, minutes after maternal nicotine exposure. A rapid and significant decrease in vasculature was observed compared to the sham group.
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Our team has recently shown the SNR and depth-sensitivity advantages of using 1064 nm light for diffuse correlation spectroscopy as well as the challenges of commercially available single-photon detectors at this wavelength. We will review two strategies for custom readout integrated circuit designs that simultaneously target lower pixel dead times and lower afterpulsing probabilities. Both designs use macropixels comprising many detectors, each having a programmable hold-off time. We will compare simulated autocorrelations for our detector models and compare predicted performance against commercial InGaAs/InP detectors.
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Breast cancer is a serious threat to women worldwide due to its high incidence and mortality. The early detection is very crucial for the treatment of breast cancer. Currently, for breast cancer patients, mammography and stereotactic needle biopsy followed by time-consuming pathological observations are the primary diagnostic approaches. In our previous study, it was found that the characteristic features of breast carcinoma tissues often include fibrous structures induced by inflammatory reactions, which can be quantitatively evaluated by polarimetric techniques. In this study, we further measure the transmission Mueller matrix microscopic images of 30 breast ductal tissue samples at different progression stages. We calculate the Mueller matrix derived parameters, which can provide the quantitative information on the location, density and distribution behavior of the fibrous structures in the tissues. To evaluate the distribution behavior of fibrous structures more quantitatively and precisely, we also analyze the parameters δ and θ using the gray level cooccurrence matrix (GLCM) analyzing method. The results demonstrate that, the GLCM features Contrast, Energy, Correlation and Homogeneity of δ and θ can be used to describe different textures of fibers distributions among the healthy, ductal carcinoma in situ (DCIS) and invasive ductal carcinoma (IDC) tissue samples, whereas the parameters of unpolarized light intensity images show no prominent differences. The Mueller matrix derived parameters combined with image analyzing methods can be used for label-free detecting and quantitative staging of breast carcinoma tissues, which can be helpful for clinical diagnosis.
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The ability of diffuse correlation spectroscopy (DCS) to measure tissue perfusion paves the way for monitoring cerebral blood flow non-invasively. However, during measurements on human forehead, the measured blood flow index (BFi) is susceptible to contamination due to the blood flow in extracerebral tissue. Time domain DCS addresses this problem by selecting photons based on their travel time to obtain BFi at various depths. We have determined the gate start time(s) and width(s) that can lead to optimal sensitivity of BFi to brain blood flow during actual measurements on human subjects using commercially available hardware with accurate noise modelling.
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We present a new approach to Diffuse Correlation Spectroscopy (DCS) which overcomes the limited light throughput of single mode photon counting techniques, and operates with continuous wave illumination without disturbance from ambient light. Heterodyne holographic detection allows parallel measurement of the power spectrum of a fluctuating electric field across thousands of modes, from which we may directly compute flow parameters using a novel Fourier domain DCS model. Our detection and modelling strategy are rigorously validated by modulating the Brownian and flow components of an optical tissue phantom, demonstrating absolute measurements of the Brownian diffusion coefficient in excellent agreement with conventional methods. We demonstrate the feasability of in vivo measurement through the recovery of pulsatile flow rates measured in the human forearm.
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Desmoplastic stiffening of breast carcinoma is the bulk scale manifestation of the extra-cellular matrix (ECM) micromechanical remodeling during neoplastic transformation. We have developed a novel optical imaging technology, termed laser speckle micro-rheology (LSM) for evaluating the micro-mechanical properties of tissue and investigating the implications of the mechano-pathological phenomena to the disease progression. Here, we present a new enhancement to the LSM processing scheme to extract the optical properties from speckle patterns and correct for residual speckle modulations to quantify the shear viscoelastic modulus of breast tissues. By accounting for optical variations, we observed an increased significance in the shear moduli differences between benign lesions and tumors and also between tumors of different histological diagnosis.
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We report holistic quantification of cutaneous microcirculation with spatial frequency domain imaging (SFDI) and coherent hemodynamics spectroscopy (CHS) based on a dynamic microcirculation PIPE model. A simple device was developed to induce periodic variations in cutaneous blood volume and flow velocity. Both baseline and dynamic features of cutaneous microcirculation for healthy subjects were completely quantified. The CHS findings were further related to SSMD-SFDI imaging of the same healthy subjects under reactive hyperemia protocol. The results demonstrate spatial frequency domain imaging and coherent hemodynamics spectroscopy based on the dynamic microcirculation PIPE model provides a valuable tool for functional studies with hemodynamic-based techniques.
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The goal of this research was in testing magnetic resonance imaging (MRI) pulse sequences for monitoring local changes of proton relaxation times after the local application of skin optical clearing (OC) compositions in vivo. We used xenograft mouse models of cancer, i.e. nu/nu mice bearing subcutaneous tumors expressing endogenous TagRFP red fluorescent protein marker and tested the changes in fluorescence intensity and lifetime (FL) of the subcutaneous tumor foci after OC application (70% glycerol, 5% DMSO, 25% water) onto the skin. By using time-correlated single photon counting within 20-30 min after the OC we observed: 1) 30-40% increase in the overall photon numbers output; 2) 50 ps increase in the median FL of TagRFP. We subsequently performed tracking of MR signal intensity changes within selected regions of interest (ROI) located close to the skin surface before, during and after OC. The analysis of 1T MR T2-weighted (T2w) fast spin-echo images showed significant quantitative differences between Gaussian noise-normalized MRI signal intensities (Mann-Whitney test, p<0.05). Our results suggest that the application of OC may cause: 1) a transient change of the peripheral tumoral microenvironment and as a consequence, FL increase and shortening of mean proton relaxation times within the voxels of subcutaneous tumor (i.e. T2w hypointensity increase); 2) potential microviscosity change due to the permeability for the OC components resulting in shortening of tissue water proton relaxation times. The results suggest that T2w 1T MRI was useful for semi-quantitative monitoring of MR signal intensity longitudinal changes in the subcutaneous space during and after OC thereby enabling registration of optical and MR signal fluctuations in the same voxels of live tissue.
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Dynamic FFOCT allows us to see the sub-cellular motion of biological samples. We are able to follow the evolution in the same plane of biological samples for hours thanks to an autofocus procedure. RPE cells are involved in the integrity of retina and vision. We performed a linear scratch assay in RPE cell culture with a surgical scalpel blade, inducing border cell migration at 20.8 µm/h to close the scratch. We also recorded motility of microvilli, thanks to rapid GPU computing. Quantitative live imaging of RPE cell culture with DFFOCT is useful in development of disease models of retinal degeneration.
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We used laser Doppler holography, a full-field imaging technique, to measure blood flow in the retina and choroid with an unmatched temporal resolution. We here report on the measurement of the flow waveforms in retinal arteries and veins after spatio-temporal filtering of the power Doppler. We demonstrate a full field mapping of the local resistivity index, and the possibility to perform unambiguous identification of retinal arteries and veins on the basis of their systolodiastolic variations. This work demonstrates the potential held by laser Doppler holography to study ocular hemodynamics in healthy and diseased eyes.
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How to write a biophotonics app that reaches more than 500 million people
1. Focus on what you know. In our case that was tissue optics and microcirculation imaging.
2. Watch out for new and especially consumer technologies which could be harnessed in your field. By 2010 smartphones had a good light source, a good camera and sufficient processing power to be a useful biophotonics instrument.
3. Think of a bio problem which concerns many more than 500 million people and which could be solved using this new technology. The WHO had just declared that resting heart rate was a leading indicator of cardiovascular health.
4. Train someone in the necessary skill(s). In our case that was Java which would allow us to program an Android based smartphone. We already knew a lot about tissue optics and blood flow.
5. Give it away for free. We decided to publish the idea and have provided well documented simple versions of the code on GitHub, along with an educational version on Google Play.
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Biophotonics of Embryo Dynamics: Monitoring, Imaging, and Functional Control
Optical coherence tomography (OCT) is a promising research tool for non-invasive imaging of biological tissues. OCT can be used to capture images of the beating Drosophila heart in vivo with micrometer resolution and video-rate imaging speed. Due to its non-invasiveness nature, OCT enables longitudinal imaging of developing flies through the entire life cycle of the insect. Combined with non-invasive optogenetic pacing, this forms a powerful research platform to study developmental biology.
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We recently established a set of unique in vivo methods for investigation of early embryonic development and reproduction in mice. Without the use of any contrast agents, these methods allow for live dynamic volumetric imaging of mouse reproductive tract with micro-scale spatial resolution using optical coherence tomography (OCT), in vivo depth-resolved mapping of the cilia location and cilia beat frequency (CBF) in the Fallopian tube (oviduct), and tracking individual sperm and their motility in vivo in the oviduct. Our observations revealed intriguing findings, which question current views on role of cilia in gamete and embryo transfer during preimplantation pregnancy.
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Biomechanical signaling is required during cardiac development. Perturbation of heart biomechanics results in a malformed heart, the primary phenotype found in congenital heart disease. Cardiac progenitor cells are responsible for acquiring all cardiac cell fates, expanding, and organizing into a functional heart. How biomechanical signals govern cardiac progenitor cell behavior to achieve cardiac morphogenesis is unknown. In this work, we will implement optogenetic cardiac pacing as a means to control cardiac biomechanics and determine how altered mechanics affects cardiac progenitor cells.
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Optical clearing (OC) of biological tissues is a promising technology for a wide spread use in medical practice to increase the screening depth, spatial resolution and contrast of the resulting images/spectra. Nevertheless, despite the significant OC effect, some biocompatible optical clearing agents (OCAs) can adversely affect biological tissues, causing local hemostasis, morphological changes, d ehydration, and in some cases even tissue necrosis. The aim of this study was to study the effect of Omnipaque 300 and fructose solutions of various concentrations and exposure times on the intact skin using confocal Raman microspectroscopy. It was shown that the application of each of these OCAs on intact skin for 5 min also leads to an appreciable OC effect. The increase in OC was achieved using a mixture of Omnipaque 300 with DMSO; it was shown that the optical properties of the skin can be controlled at a depth of about 80 μm.
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Assessment of disease using OCT is an actively investigated problem, owing to many unresolved challenges in early disease detection, diagnosis and treatment response monitoring. The spatial scale to which the information can be obtained from the scattered light is limited by the diffraction limit (~λ/2; λ = wavelength of light is typically in the micron level) and the axial resolution of OCT systems is limited by the inverse of spectral bandwidth. Yet, onset or progression of disease /precancer is typically associated with subtle alterations in the tissue dielectric and its ultra-structural morphology. On the other hand, biological tissue is known to have ultra-structural multifractality. For both the fundamental study of biological processes and early diagnosis of pathological processes, information on the nanoscale in the tissue sub-micron structural morphology is crucial. Therefore, we have developed a novel spectroscopic and label-free 3D OCT system with nanoscale sensitivity in combination of multifractal analysis for extraction and quantification of tissue ultra-structural multifractal parameters. This present approach demonstrated its capability to measure nano-sensitive tissue ultra-structural multifractality. In an initial study, we found that nano-sensitive sub-micron structural multifractality changes in transition from healthy to tumor in pathologically characterized fresh tissue samples. This novel method for extraction of nanosensitive tissue multifractality promises to develop a non-invasive diagnosis tool for early cancer detection.
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