Search

Moderate heating of collagenous tissues such as cartilage and cornea by infrared laser irradiation can produce biologically nondestructive structural rearrangements and relaxation of internal stresses resulting in the tissue reshaping. The reshaping results and eventual changes in optical and biological properties of the tissue strongly depend on the laser-irradiation regime. Here, a speckle-contrast technique based on monochromatic illumination of the tissue in combination with strain mapping by means of optical coherence elastography (OCE) is applied to reveal the interplay between the temperature and thermal stress fields producing tissue modifications. The speckle-based technique ensured en face visualization of cross correlation and contrast of speckle images, with evolving proportions between contributions of temperature increase and thermal-stresses determined by temperature gradients. The speckle-technique findings are corroborated by quantitative OCE-based depth-resolved imaging of irradiation-induced strain-evolution. The revealed relationships can be used for real-time control of the reshaping procedures (e.g., for laser shaping of cartilaginous implants in otolaryngology and maxillofacial surgery) and optimization of the laser-irradiation regimes to ensure the desired reshaping using lower and biologically safer temperatures. The figure of waterfall OCE-image demonstrates how the strain-rate maximum arising in the heating-beam center gradually splits and drifts towards the zones of maximal thermal stresses located at the temperature-profile slopes.

In recent pre-clinical studies, we have demonstrated that analysis of speckle statistics in low-scattering, low SNR regions may have applications for accurate lymphatic vessels (and nerves) mapping in optical coherence tomography (OCT) of biotissues. Specifically, we utilized a Rayleigh fitting goodness-of-fit threshold parameter to isolate low scattering biological structures, in particular to distinguish them from noise. The threshold as well as the size of region of interest (ROI) for such speckle statistics evaluation were both chosen empirically. Here we present a model simulation study aimed at optimizing and automatically determining these empirically chosen important parameters. We explore the Rayleigh fit and speckle contrast metrics parameter space to enable accurate label-free OCT mapping of weakly scattering structures such as lymphatic vessels.

Development of noninvasive diagnostic techniques in clinical practice, especially sufficiently fast methods suitable for intraoperative use, remains a topical and challenging problem in modern medicine. Compressional Optical Coherence Elastography (OCE) that has a high potential to be intraoperatively applied. This study reports preliminary results on application of compressional OCE for identification of tumor margins and even differentiation of breast cancer types with low- and high- histological grades, i.e., a step towards creation of a kind of OCE-based optical biopsy. The reported variant of OCE utilized B-scans with scales ~2-4 mm comparable with typical scales of histological samples. The approach is based on the ability of OCE to quantitatively visualize distribution of the Young modulus (stiffness) in studied tissue samples, which then is used to perform "elasto-spectroscopy" generically resembling the mass-spectroscopy procedures. The procedures of the OCE examination do not require any special preparation of tissue samples and thus can be applied intraoperatively on time intervals ~ a few minutes with immediately available results. In the reported study, the samples after OCE examinations were subjected to histological studies and classification made by an experienced histopathologist. Comparison between the OCE results and histological data made it possible not only to distinguish between normal tissue and tumor (for determining resection boundary), but also to formulate OCE-based criteria allowing for differentiation between carcinomas of low grades and fairly good treatment prognosis and high-grade invasive tumors with poor prognosis. Similarly, identification of not only invasive breast tumors, but also benign breast lesions, intermediate between norm and malignant tumors can be made.

We present an optical coherence tomography (OCT) lymphangiography visualization approach based on analysis of speckle statistics. For in-vivo experimentation, normal and tumor tissues are examined in mouse dorsal skin window chamber model. In order to evaluate the speckle statistics, OCT datasets are acquired in 3 spatial and 1 temporal dimensions to be divided then into smaller volumes of interests. In temporal dimension, repeated same-location scanning is performed for simultaneous blood vessel detection through speckle variance processing. Speckle statistics in each volume of interest are tested for similarity to known distributions corresponding either to noise or to tissue. We show that lymphatic vessels could be detected using a specific parameter range of speckle statistics as a filter, to separate the surrounding tissues and blood vessels. The proposed approach does not require numerous post-processing steps that are often used in lymphatic detection methods that are based on low signal amplitude regions (e.g., OCT signal attenuation compensation, inversion, amplitude thresholding etc.). Instead, we use a fast 2-step filtering procedure to reveal lymphatic vessels in imaged tissues.

We present a realization of real-time OCT-based strain mapping by estimating interframe phase-variation gradient using the developed "vector" method. This technique allows for mapping both fairly fast and large, as well as rather small strains, slowly-varying on intervals ~tens of minutes. Optimization of interframe interval for improving signal-to-noise ratio is discussed and experimentally demonstrated. Ultimate stability of strain estimation with the designed OCT setup is experimentally estimated using stable phantoms. Examples of spatially resolved maps of slowly-varying strains are demonstrated. The developed methods can be used in emerging techniques of laser-assisted modification of collagenous tissues (e.g., for such perspective application as fabrication of cartilaginous implants).

A computationally efficient and fairly realistic model of OCT-scan formation in spectral-domain optical coherence tomography is described. The model is based on the approximation of discrete scatterers and ballistic character of scattering, these approximations being widely used in literature. An important feature of the model is its ability to easily account for arbitrary scatterer motions and computationally efficiently generate large sequences of OCT scans for gradually varying configurations of scatterers. This makes the proposed simulation platform very convenient for studies related to the development of angiographic processing of OCT scans for visualization of microcirculation of blood, as well as for studies of decorrelation of speckle patterns in OCT scans due to random (Brownian type) motions of scatterers. Examples demonstrating utilization of the proposed model for generation OCT scans imitating perfused vessels in biological tissues, as well as evolution of speckles in OCT scans due to random translational and rotational motions of localized (but not-point-like) scatterers are given. To the best of our knowledge, such numerical simulations of large series of OCT scans in the presence of various types of motion of scatterers have not been demonstrated before.

In this paper, we present a computationally highly efficient full-wave model of optical coherence tomography (OCT)-scan formation by focused beams in spectral-domain OCT. Similarly to some previous models, it is based on the summation of fields scattered by discrete sub-resolution scatterers, and enables one to account for the axial and lateral inhomogeneity of the illuminating Gaussian beam. The main feature ensuring the high computational efficiency of the described model is that, rather than numerical integration of the scattered signal over the receiving aperture, we apply an analytical description of both the illuminating-beam focusing and the collection of the scattered signals over the receiving aperture. Elimination of numerical integration over the receiving aperture increases the overall computation speed by a factor of ∼ 10 3 . This is of key importance in terms of the practical feasibility of simulations of 3D OCT data volumes for large amounts (∼ 10 5 − 10 7 ) of scatterers, corresponding to realistic densities of cells in biological tissues. We demonstrate the model’s possibilities by simulating the digital refocusing of strongly focused OCT beams in the presence moving scatterers, thereby presenting a novel principle of contrast-agent-free visualization of scatterer flows, with velocities typical of blood microcirculation.

Emerging methods of anti-tumor therapies require new approaches to tumor response evaluation, especially enabling label-free diagnostics and in vivo utilization. Here, to assess the tumor early reaction and predict its long-term response, for the first time we apply in combination the recently developed OCT extensions - optical coherence angiography (OCA) and compressional optical coherence elastography (OCE), thus enabling complementary functional/microstructural tumor characterization. We study two vascular-targeted therapies of different types, (1) anti-angiogenic chemotherapy (ChT) and (2) photodynamic therapy (PDT), aimed to indirectly kill tumor cells through blood supply injury. Despite different mechanisms of anti-angiogenic action for ChT and PDT, in both cases OCA demonstrated high sensitivity to blood perfusion cessation. The new method of OCE-based morphological segmentation revealed very similar histological structure alterations. The OCE results showed high correlation with conventional histology in evaluating percentages of necrotic and viable tumor zones. Such possibilities make OCE an attractive tool enabling previously inaccessible in vivo monitoring of individual tumor response to therapies without taking multiple biopsies.

In this report we discuss some practical obstacles/pitfalls arising in realization of quantitative compressional OCE, as well as discuss possible ways of their resolution. More specifically we consider (i) complications in quantification of the Young modulus of tissues related to influence of partial adhesion between the OCT probe and pre-calibrated reference layers - "compliant sensors", (ii) distorting influence of surface curvature/corrugation on strain distribution in the tissue bulk, (iii) ways of enhancement of effective SNR in OCT-based strain mapping without periodic averaging, and (iv) potentially significant influence of nonlinearity of the elastic response of biological tissues on quantification of their stiffness.

Strain mapping based on analysis of complexvalued signal of optical coherence tomography (OCT) is applied to monitor slow deformations of biological tissues (e.g., cartilage or cornea) subjected to thermo-mechanical reshaping by moderate heating with an IR laser. Insufficiently relaxed internal stresses may slowly distort the required tissue shape on a scale from minutes to hours with strain rate ~10−4 s−1. We show the ability of the developed OCT method to monitor strains with uncertainly ~10−5 on ~102 s intervals sufficient for shape-stability verification in intra-operational preparation of cartilaginous implants.

Approach to OCT-lymphangiography (OCT-LA) and interstitial-fluid-filled dislocations mapping that complement speckle variance (SV) OCT angiography is presented. OCT-LA can be extracted from the same OCT raw datasets that were acquired for SV OCT angiography. Lymphatic vessels and interstitial dislocations are filled with transparent fluids (e.g. lymph) and can be separated from the blood vessels and tissue. In vivo application of the OCT-LA approach is demonstrated on NRG mouse with grown BX-PC3 tumor (human pancreatic cancer cells).

Introduction of innovative noninvasive diagnostic techniques in clinical practice remains an actual topic of modern medicine. In the study, we assessed feasibility of multimodal OCT (MM OCT) that combines crosspolarization imaging and elastographic stiffness mapping to assess spatial structural organization and heterogeneity of breast cancer in the tumor center in comparison with normal mammary gland tissue. The research was carried out using human breast tissue mastectomy surgical samples including different histological types of breast cancer: ductal carcinoma in situ, invasive lobular and ductal carcinoma, fibroadenoma and normal tissue. The histological subtypes of breast cancer showed different structural and stiffness features of tumor tissue. In the case of invasive ductal carcinoma, the cross-polarization OCT image shows a more heterogeneous high-level OCT signal and higher stiffness in comparison with lobular breast cancer or fibroadenoma. These results indicate that assessing microstructures and elasticity changes yields complementary information about the microstructural features of breast cancer, which is important for selection of treatment tactics.

A new laser method for increasing uveoscleral outflow path for normalization of intraocular pressure in glaucomatous eyes is presented. Nonuniform laser heating affects the porous system of biological tissues. Formation of new pores in the paralimbal region of the eye can accelerate the flow of the intraocular fluid through the eye sclera and, thus, facilitate normalization of the intraocular pressure. A positive effect of laser impact is achieved, as a rule, in a narrow range of laser radiation parameters, which makes it difficult to choose the intensity and time parameters of laser irradiation due to such factors as nonstationary temperature fields, thermotensions and pressure that can give rise to undesirable effects and complications. The comparison between reflected and transmitted laser light through the eye tissue has allowed to establish the main requirements for laser settings parameters responsible for efficacy and safety of the laser irradiation. The positive effect is achieved only by using relatively small intensity of the laser radiation. At high intensity, the hydraulic permeability decreases due to denaturation and tissue hardening. Atomic Force Microscopy (AFM) measurements with nanoindentation and optical coherence tomography (OCT) based compressional phase-sensitive optical coherence elastography (OCE). OCE measurements demonstrated laser-induced dilatation areas attributed to formation of ensembles of micro-and nano-pores in sclera providing increase in its hydraulic permeability. Much higher resolution AFM examinations directly demonstrated such individual irradiation-produced pores. At the same time, the collagen structure of the sclera is not destroyed, and tissue mechanical properties do not degrade under laser radiation. The process of pore formation is in good agreement with computer simulations of the dynamics of thermal stress fields induced by laser irradiation.

Soft biological tissues manifest strongly pronounced nonlinear elastic behavior. Namely, the Young modulus for some tissues strongly depends on the applied stress. This property can be evaluated by compressional OCT elastography. We demonstrate the evaluation of nonlinear elastic properties on the samples of coronary arteries, breast cancer and cornea. The developed technique can be used for further investigation of the nonlinear properties of healthy and pathological tissues.

Non-surgical thermo-mechanical reshaping of avascular collagenous tissues (cartilages and cornea) using moderate heating by IR-laser irradiation is an emerging technology that can find important applications in visioncorrection problems and preparation of cartilaginous implants in otolaryngology. To estimate both transient interframe strains and cumulative resultant strains produced by the laser irradiation of the tissue we use and improved version of strain mapping developed in our previous work related to compressional phase-sensitive optical coherence tomography. To reveal microstructural changes in the tissue regions where irradiation-produced strains do not disappear after temperature equilibration, we apply compressional optical coherence elastography in order to visualize the resultant variations in the tissue stiffness. The so-found regions of the stiffness reduction are attributed to formation of microscopic pores, existence of which agree with independent data obtained using methods of high-resolution microscopy.

OCT-based local strain relaxation/creep evaluation is an emerging tool for tissue viscoelasticity characterization. We present a tool for 2D visualization of local strain relaxation and creep time/rate inside the tissue.

Multimodal OCT is a promising tool for monitoring of individual tumor response to antitumor therapies. The changes of tumor cells, connective tissue, microcirculation and stiffness can be estimated simultaneously in real time with high resolution.

We discuss the application of compressional OCT-based elastography for measuring elastic response of biological tissues in the strain range from fractions of one per cent up to strains over ~10% and greater. Such fairly large strains are not typical of phase-sensitive OCT-based elastographic methods and can be measurable due to application of interframe strain-estimation method in which interframe phase gradients are estimated without the necessity of preliminary measuring total displacements of scattering particles in combination with summation of interframe strains. For estimating applied stresses, intervenient soft layer of translucent silicone is used a a reference fairly linear material. The obtained results based on comparison of strains in the reference layer and tissue demonstrate highly nonlinear character of elasticity of biological tissues even for rather moderate strains on order of several per cent.

Phase-sensitive optical coherence tomography (OCT) is used for visualizing dynamic and cumulative strains and corneashape changes during laser-produced tissue heating. Such non-destructive (non-ablative) cornea reshaping can be used as a basis of emerging technologies of laser vision correction. In experiments with cartilaginous samples, polyacrilamide phantoms and excised rabbit eyes we demonstrate ability of the developed OCT system to simultaneously characterize transient and cumulated strain distributions, surface displacements, scattering tissue properties and possibility of temperature estimation via thermal-expansion measurements. The proposed approach can be implemented in perspective real-time OCT systems for ensuring safety of new methods of laser reshaping of cornea.

The main goal of the study was a vasculature targeted PDT. A new approach named M-mode-like OCT (MML OCT) was applied to monitor early response to PDT. Due to the chosen filtering parameters (96 Hz filter threshold), this approach visualizes only vessels with flowing blood. Without flowing blood even filled vessels are not visualized because flow-less blood causes speckle variations with significantly lower frequencies (

Application of compressional optical coherence elastography (OCE) for delineation of tumor and peri-tumoral tissue with simultaneous assessment of morphological/molecular subtypes of breast cancer is reported. The approach is based on the ability of OCE to quantitatively visualize stiffness of studied samples and then to perform a kind of OCE-based biopsy by analyzing elastographic B-scans that have sizes ~several millimeters similarly to bioptates used for “gold-standard” histological examinations. The method relies on identification of several main tissue constituents differing in their stiffness in the OCE scans. Initially the specific stiffness ranges for the analyzed tissue components (adipose tissue, fibrous and hyalinized tumor stroma, lymphocytic infiltrate and agglomerates of tumor cells) are determined via comparison of OCE and morphological/molecular data. Then assessment of non-tumor/tumor regions and tumor subtypes is made based on percentage of pixels with different characteristic stiffness (“stiffness spectrum”) in the OCE image, also taking into account spatial localization of different-stiffness regions. Examples of high contrast among benign (or non-invasive) and several subtypes of invasive breast tumors in terms of their stiffness spectra are given.