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Mechanical forces are key to the structure, dynamics, and interactions of living systems. In the last two decades, Brillouin Microscopy (BM) has emerged as a non-invasive optical tool for the mechanical characterisation of biomatter at GHz frequencies and on a microscale. Viscous and elastic properties of biosamples in this spatio-temporal regime are effectively an uncharted territory that is important for the potential impact on function and physiology.
Since its inception, BM has been applied to address a myriad of biological and medical questions and has shown key capabilities for cell mechanobiology and tissue histopathology. Our team has developed and applied BM to study tissue mechanics and revealed the ability of BM to map the acoustic anisotropy of extracellular matrix proteins in isolated fibres and tissue biopsies. For these studies, we have introduced the correlative Brillouin–Raman method as a chemical-specific mechanical probe of biosamples.
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Brillouin spectrometers use etalons to analyze Brillouin spectra. Line-scan Brillouin microscopes improve the image acquisition time ~100-fold than what was previously achievable by coupling line illumination with etalon spectrometers to multiplex the spectral measurement a row of pixels at a time. Multiplexing represents a way to improve Brillouin imaging speeds, but etalon-based spectrometers cannot multiplex a full image. Here, we investigate the potential of a new spectrometer based on atomic hyperfine transitions, enabling simultaneous analysis of a full field of view. We show that the spectrometer can fully transmit an image without distortions, thus proving the potential for 2D multiplexing.
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Brillouin spectroscopy has emerged as a great modality to non-invasively target mechanical properties in material and biological samples, although it requires high-performance spectrometers and long acquisition times to extract the Brillouin peaks with high SNR and precision. Stimulated Brillouin scattering (SBS) has the potential to improve speed and resolution, achieving a resonant amplification of the scattered signal through the interaction of two counterpropagating laser beams. However, the overall performances of current SBS spectrometers result just comparable to spontaneous Brillouin, and this may indicate that the system is operating with suboptimal acquisition parameters. Here, we will investigate this hypothesis introducing the localization theory in the estimation of the peak position in SBS spectroscopy and demonstrating a ten times improvement in acquisition speed, retaining SNR and precision, by simply designing an SBS spectrometer with proper acquisition parameters.
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Brillouin spectroscopy and microscopy is an emerging tool of biological imaging and biomedical spectroscopy, which allows assessing local viscoelastic properties of biological materials.One of the greatest untapped potentials of Brillouin microscopy is its compatibility with almost all known optical imaging modalities for multimodal optical imaging. In the past, we successfully demonstrated that Brillouin spectroscopy can be successfully combined with Raman spectroscopy for simultaneous imaging of local chemical and physical properties of cells and biomaterials. In this report, we describe our ongoing efforts to extend the multimodal capabilities of Brillouin microscopy for better understanding mechanical properties of biological systems and their relationship with the local structure, chemistry and metabolic activity.
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Neural tube closure, or neurulation, has been studied across a range of vertebrates as it is the basis of embryonic development. Closure failure can lead to severe congenital malformations. Current technologies require fixed specimens and physical contact to extract a modulus. Here we investigate the mechanical changes of the neural plate from formation to closure within intact live chick embryos using time-lapse Brillouin imaging and ex-ovo culture. We observed an increase in the Brillouin modulus of the neural plate as the embryo develops in ex-ovo culture. By quantifying the timing and the extent of the forces that drive neural tube closure, we can more accurately identify when and why neural tube defects occur.
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Altered tissue stiffness has raised as both the cause and the consequence of breast tumorigenesis. Imaging the heterogeneous mechanical properties of tissue microenvironment provides crucial insights into micro-mechanical drivers of carcinogenesis. Here, we introduce laser Speckle rHEologicAl micRoscopy (SHEAR), which enables mapping the frequency-dependent shear viscoelastic modulus, G*(x,y,ω), with a spatial resolution of < 50 mm, over multiple cm2, in solid tumors within minutes. SHEAR mapping in a cohort of excised breast specimens demonstrates that |G*(x,y)| closely agrees with tumor histopathology, its gradient, |∇|G*(x,y,ω)||, is increased at the tumor invasive front, and both metrics are associated with tumor prognosis.
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Reconstructive skin surgeries drive the clinical need for non-contact objective measurements of skin elasticity. Here we demonstrate that all three of skin’s elastic constants (in-plane and out-of-plane shear moduli and an additional modulus defining skin’s tensile anisotropy) and the orientation of collagen fibers in dermis can be determined from Rayleigh wave anisotropy in-plane with acoustic micro-tapping (AuT) OCE. A nearly-incompressible transverse isotropic (NITI) model was used to reconstruct skin’s moduli from OCE measurements in human forearm in vivo for five healthy volunteers. Co-registered polarization-sensitive (PS-) OCT shows that optical and mechanical axes are co-aligned at measured sites.
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Diabetic retinopathy, a slow progressive complication of diabetes, is the leading cause of vision impairment and blindness among working-age adults. In the presence of hyperglycemia, ocular tissues become stiffer in response to the increased non-enzymatic cross-linking of collagen fibrils. Ocular rigidity may serve as a cumulative response indicator of hyperglycemia. We have implemented an in vivo approach to estimate ocular rigidity using dynamic optical coherence tomography, which allowed us to investigate the diabetic retinopathy-associated biomechanical changes in a clinical setting.
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Methods to quantify airway wall compliance are needed for diagnostics, stratification, and monitoring in upper airway disorders and inhalation injury. Endoscopic OCT tracks micron-scale airway deformation during respiration, and in conjunction with in situ pressure monitoring, maps local and cross-sectional compliances. Airway phantoms are employed to validate the accuracy of the methods, and experiments in ex vivo and in vivo pig airways are compared to CINE CT. Findings include the importance of centroid tracking, best practices for endoscopic procedures, hysteresis in airway pressure-volume curves, changes in elastic properties during burn injury, and modeling methods to extract elasticity and viscoelasticity.
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Recent advances in dynamic OCE have resulted in tools that can generate/track sub-mm wavelength mechanical waves in tissue. However, reconstructing material elasticity from measured wavefields needs an appropriate model accounting for tissue anisotropy, structure and geometry. We assume that tissues consisting of collagen fibers can be locally described with a model of a nearly incompressible transverse isotropic (NITI) medium using three elastic parameters to describe shear and tensile behavior. Examples of NITI media are discussed and the problem of inversion of moduli from bulk shear, Rayleigh and guided waves is considered.
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Described in this study is an advanced co-axial acoustic stimulation technique with pre-compensation for acoustic frequency content and reconstruction of signals in the Fourier domain for use in Optical Coherence Tomography (OCT) Vibrography. The feasibility of the technique was demonstrated via the measurement of the first mechanical, flexural resonance modes of two contact lenses with varied elastic moduli and an ex-vivo porcine cornea, each with a maintained constant intraocular pressure. The measurement of these resonance modes was achieved through use of a Swept Source OCT system, operated in phase sensitive mode, to detect the nanometer scale displacements of these modes.
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Ocular Biomechanical Properties: Joint Session with Conferences 11941 and 11962
Irregularities of ocular pulsatility are associated with some of the most severe eye diseases. Here we present fundus elastography (FUEL) based on optical coherence tomography (OCT) for the quantitative assessment and depth-resolved mapping of pulsatile dynamics in the murine retina and choroid. Our FUEL approach is based on an analysis of the complex OCT signal dynamics across repeated frame acquisitions. We demonstrate in vivo FUEL imaging in the retinas of wildtype mice and mouse models of retinal diseases and reveal subtle structural deformations related to ocular pulsation. Our data in mouse eyes hold promise for a powerful retinal elastography technique that may enable a new paradigm of OCT based measurements and image contrast.
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The previous researches have demonstrated biomechanical elasticity of optic nerve head is associated with multiple ophthalmic diseases. In this work, we have demonstrated a method to quantify in-vivo elastography of ONH, by using a confocal lamb wave acoustic radiation force optical coherence elastography (ARF-OCE). The ARF-OCE system is based on a phase resolved SD-OCT system combined with an acoustic transducer. Experiments were performed on New Zealand White rabbit eyes in vivo after anesthesia. We have obtained 3D reconstructed OCT images of ONH and the time-resolved elastic map of peripheral retina under various Intraocular pressure.
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Cornea, the window of the eye, is critical for vision. The precise understanding of corneal biomechanical properties remains an unmet clinical need in early diagnosis and customized treatments for corneal blindness.
Our research group has developed a novel non-contact, non-invasive AuT-OCE method to quantify corneal elasticity. Because of the anisotropic corneal property due to its collagen structure, a nearly incompressible transversely isotropic (NITI) model was developed to characterize its elasticity. It has been shown that in-plane tensile and out-of-plane shear properties are defined by different moduli, E and G, respectively.
Our research offers new opportunities to develop a personalized biomechanical model based on quantitative maps of corneal mechanical moduli to quantify both biomechanical properties and predict final corneal shape, leading to potential early diagnosis and more precise surgical planning.
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The nucleus is the largest organelle in the cell. When deformed with techniques like AFM or micropipette aspiration, the nucleus appears to be highly elastic and much stiffer than the cytoplasm. Whether the nucleus behaves like a stiff elastic object when shaped by cellular forces and on physiological time scales, such as during migration through confining channels, is not clear. Here I will discuss our efforts to understand nuclear mechanics in cell migration. I will present live cell imaging experiments that reveal surprising nuclear mechanical behaviors such as drop-like deformation. I will show how the nucleus is likely shaped by viscous coupling between the nucleus and the cytoplasm rather than static cytoskeletal stresses.
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Altered biomechanical properties are an important precursor for degenerative tissue pathologies. In the eye, diagnostic tools are demanded to be non-invasive, quickly-performed and of high resolution. To meet this need, a novel technique is presented based on an under-pressure chamber applying a homogenous mechanical load on the ocular shell similar to the intraocular pressure, and simultaneous phase-sensitive recording of axial displacements and strain by optical coherence tomography. Results are presented that visualize the instantaneous mechanical effect of patterned corneal cross-linking in ex vivo rat eyes, and the creep response of cornea and crystalline lens in an in vivo subject.
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Quantification of the corneas´ biomechanical properties helps to diagnose corneal abnormalities early, which is key in keratoconus (KC) management and treatment. We recently introduced a multi-meridian air-puff ssOCT system capable of acquiring corneal deformation images during air-puff excitation on two meridians. Two healthy and three KC patients were measured with the system. The results were used to quantify deformation asymmetries and as input data for Finite Element (FE) modeling, which was used to estimate corneal biomechanical properties by means of an inverse analysis. Deformation asymmetry parameters and the estimated tangent modulus for healthy and KC corneas are presented and compared.
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AμT-OCE was used to quantify changes in both the in-plane Young’s (E) and out-of-plane shear (G) moduli in human cornea following riboflavin/UVA crosslinking in a non-contact, non-destructive manner. Since OCT methods are broadly accepted in Ophthalmology, it suggests fast translation of AμT-OCE into clinical practice if results are confirmed in vivo. In addition, AμT-OCE can change diagnostic criteria of ectatic corneal diseases, leading to early diagnosis, reduced complications, customized surgical treatment, and new opportunities to develop personalized biomechanical models of the eye.
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