Optical palpation for tumor margin assessment in breast-conserving surgery.

Intraoperative margin assessment is needed to reduce the re-excision rate of breast-conserving surgery. One possibility is optical palpation, a tactile imaging technique that maps stress (force applied across the tissue surface) as an indicator of tissue stiffness. Images (optical palpograms) are generated by compressing a transparent silicone layer on the tissue and measuring the layer deformation using optical coherence tomography (OCT). This paper reports, for the first time, the diagnostic accuracy of optical palpation in identifying tumor within 1 mm of the excised specimen boundary using an automated classifier. Optical palpograms from 154 regions of interest (ROIs) from 71 excised tumor specimens were obtained. An automated classifier was constructed to predict the ROI margin status by first choosing a circle diameter, then searching for a location within the ROI where the circle was ≥ 75% filled with high stress (indicating a positive margin). A range of circle diameters and stress thresholds, as well as the impact of filtering out non-dense tissue regions, were tested. Sensitivity and specificity were calculated by comparing the automated classifier results with the true margin status, determined from co-registered histology. 83.3% sensitivity and 86.2% specificity were achieved, compared to 69.0% sensitivity and 79.0% specificity obtained with OCT alone on the same dataset using human readers. Representative optical palpograms show that positive margins containing a range of cancer types tend to exhibit higher stress compared to negative margins. These results demonstrate the potential of optical palpation for margin assessment.

Tissue engineered autologous cartilage-bone grafts for temporomandibular joint regeneration.

Joint disorders can be detrimental to quality of life. There is an unmet need for precise functional reconstruction of native-like cartilage and bone tissues in the craniofacial space and particularly for the temporomandibular joint (TMJ). Current surgical methods suffer from lack of precision and comorbidities and frequently involve multiple operations. Studies have sought to improve craniofacial bone grafts without addressing the cartilage, which is essential to TMJ function. For the human-sized TMJ in the Yucatan minipig model, we engineered autologous, biologically, and anatomically matched cartilage-bone grafts for repairing the ramus-condyle unit (RCU), a geometrically intricate structure subjected to complex loading forces. Using image-guided micromilling, anatomically precise scaffolds were created from decellularized bone matrix and infused with autologous adipose-derived chondrogenic and osteogenic progenitor cells. The resulting constructs were cultured in a dual perfusion bioreactor for 5 weeks before implantation. Six months after implantation, the bioengineered RCUs maintained their predefined anatomical structure and regenerated full-thickness, stratified, and mechanically robust cartilage over the underlying bone, to a greater extent than either autologous bone-only engineered grafts or acellular scaffolds. Tracking of implanted cells and parallel bioreactor studies enabled additional insights into the progression of cartilage and bone regeneration. This study demonstrates the feasibility of TMJ regeneration using anatomically precise, autologous, living cartilage-bone grafts for functional, personalized total joint replacement. Inclusion of the adjacent tissues such as soft connective tissues and the TMJ disc could further extend the functional integration of engineered RCUs with the host.

Diagnostic Accuracy of Quantitative Micro-Elastography for Margin Assessment in Breast-Conserving Surgery.

Inadequate margins in breast-conserving surgery (BCS) are associated with an increased likelihood of local recurrence of breast cancer. Currently, approximately 20% of BCS patients require repeat surgery due to inadequate margins at the initial operation. Implementation of an accurate, intraoperative margin assessment tool may reduce this re-excision rate. This study determined, for the first time, the diagnostic accuracy of quantitative micro-elastography (QME), an optical coherence tomography (OCT)-based elastography technique that produces images of tissue microscale elasticity, for detecting tumor within 1 mm of the margins of BCS specimens. Simultaneous OCT and QME were performed on the margins of intact, freshly excised specimens from 83 patients undergoing BCS and on dissected specimens from 7 patients undergoing mastectomy. The resulting three-dimensional images (45 × 45 × 1 mm) were coregistered with postoperative histology to determine tissue types present in each scan. Data from 12 BCS patients and the 7 mastectomy patients served to build a set of images for reader training. One hundred and fifty-four subimages (10 × 10 × 1 mm) from the remaining 71 BCS patients were included in a blinded reader study, which resulted in 69.0% sensitivity and 79.0% specificity using OCT images, versus 92.9% sensitivity and 96.4% specificity using elasticity images. The quantitative nature of QME also facilitated development of an automated reader, which resulted in 100.0% sensitivity and 97.7% specificity. These results demonstrate high accuracy of QME for detecting tumor within 1 mm of the margin and the potential for this technique to improve outcomes in BCS. SIGNIFICANCE: An optical imaging technology probes breast tissue elasticity to provide accurate assessment of tumor margin involvement in breast-conserving surgery.

Immune-mediated ECM depletion improves tumour perfusion and payload delivery.

High extracellular matrix (ECM) content in solid cancers impairs tumour perfusion and thus access of imaging and therapeutic agents. We have devised a new approach to degrade tumour ECM, which improves uptake of circulating compounds. We target the immune-modulating cytokine, tumour necrosis factor alpha (TNFα), to tumours using a newly discovered peptide ligand referred to as CSG. This peptide binds to laminin-nidogen complexes in the ECM of mouse and human carcinomas with little or no peptide detected in normal tissues, and it selectively delivers a recombinant TNFα-CSG fusion protein to tumour ECM in tumour-bearing mice. Intravenously injected TNFα-CSG triggered robust immune cell infiltration in mouse tumours, particularly in the ECM-rich zones. The immune cell influx was accompanied by extensive ECM degradation, reduction in tumour stiffness, dilation of tumour blood vessels, improved perfusion and greater intratumoral uptake of the contrast agents gadoteridol and iron oxide nanoparticles. Suppressed tumour growth and prolonged survival of tumour-bearing mice were observed. These effects were attainable without the usually severe toxic side effects of TNFα.

Wide-field quantitative micro-elastography of human breast tissue.

Currently, 20-30% of patients undergoing breast-conserving surgery require a second surgery due to insufficient surgical margins in the initial procedure. We have developed a wide-field quantitative micro-elastography system for the assessment of tumor margins. In this technique, we map tissue elasticity over a field-of-view of ~46 × 46 mm. We performed wide-field quantitative micro-elastography on thirteen specimens of freshly excised tissue acquired from patients undergoing a mastectomy. We present wide-field optical coherence tomography (OCT) images, qualitative (strain) micro-elastograms and quantitative (elasticity) micro-elastograms, acquired in 10 minutes. We demonstrate that wide-field quantitative micro-elastography can extend the range of tumors visible using OCT-based elastography by providing contrast not present in either OCT or qualitative micro-elastography and, in addition, can reduce imaging artifacts caused by a lack of contact between tissue and the imaging window. Also, we describe how the combined evaluation of OCT, qualitative micro-elastograms and quantitative micro-elastograms can improve the visualization of tumor.

Clinical feasibility of optical coherence micro-elastography for imaging tumor margins in breast-conserving surgery.

It has been demonstrated that optical coherence micro-elastography (OCME) provides additional contrast of tumor compared to optical coherence tomography (OCT) alone. Previous studies, however, have predominantly been performed on mastectomy specimens. Such specimens typically differ substantially in composition and geometry from the more clinically relevant wide-local excision (WLE) specimens excised during breast-conserving surgery. As a result, it remains unclear if the mechanical contrast observed is maintained in WLE specimens. In this manuscript, we begin to address this issue by performing a feasibility study of OCME on 17 freshly excised, intact WLE specimens. In addition, we present two developments required to sustain the progression of OCME towards intraoperative deployment. First, to enable the rapid visualization of en face images required for intraoperative assessment, we describe an automated segmentation algorithm to fuse en face micro-elastograms with OCT images to provide dual contrast images. Secondly, to validate contrast in micro-elastograms, we present a method that enables co-registration of en face images with histology of WLE specimens, sectioned in the orthogonal plane, without any modification to the standard clinical workflow. We present a summary of the observations across the 17 specimens imaged in addition to representative micro-elastograms and OCT images demonstrating contrast in a number of tumor margins, including those involved by invasive ductal carcinoma, mucinous carcinoma, and solid-papillary carcinoma. The results presented here demonstrate the potential of OCME for imaging tumor margins.

In vivo volumetric quantitative micro-elastography of human skin.

In this paper, we demonstrate in vivo volumetric quantitative micro-elastography of human skin. Elasticity is estimated at each point in the captured volume by combining local axial strain measured in the skin with local axial stress estimated at the skin surface. This is achieved by utilizing phase-sensitive detection to measure axial displacements resulting from compressive loading of the skin and an overlying, compliant, transparent layer with known stress/strain behavior. We use an imaging probe head that provides optical coherence tomography imaging and compression from the same direction. We demonstrate our technique on a tissue phantom containing a rigid inclusion, and present in vivo elastograms acquired from locations on the hand, wrist, forearm and leg of human volunteers.

Cell-matrix mechanical interaction in electrospun polymeric scaffolds for tissue engineering: Implications for scaffold design and performance.

Engineered scaffolds produced by electrospinning of biodegradable polymers offer a 3D, nanofibrous environment with controllable structural, chemical, and mechanical properties that mimic the extracellular matrix of native tissues and have shown promise for a number of tissue engineering applications. The microscale mechanical interactions between cells and electrospun matrices drive cell behaviors including migration and differentiation that are critical to promote tissue regeneration. Recent developments in understanding these mechanical interactions in electrospun environments are reviewed, with emphasis on how fiber geometry and polymer structure impact on the local mechanical properties of scaffolds, how altering the micromechanics cues cell behaviors, and how, in turn, cellular and extrinsic forces exerted on the matrix mechanically remodel an electrospun scaffold throughout tissue development. Techniques used to measure and visualize these mechanical interactions are described. We provide a critical outlook on technological gaps that must be overcome to advance the ability to design, assess, and manipulate the mechanical environment in electrospun scaffolds toward constructs that may be successfully applied in tissue engineering and regenerative medicine. STATEMENT OF SIGNIFICANCE: Tissue engineering requires design of scaffolds that interact with cells to promote tissue development. Electrospinning is a promising technique for fabricating fibrous, biomimetic scaffolds. Effects of electrospun matrix microstructure and biochemical properties on cell behavior have been extensively reviewed previously; here, we consider cell-matrix interaction from a mechanical perspective. Micromechanical properties as a driver of cell behavior has been well established in planar substrates, but more recently, many studies have provided new insights into mechanical interaction in fibrillar, electrospun environments. This review provides readers with an overview of how electrospun scaffold mechanics and cell behavior work in a dynamic feedback loop to drive tissue development, and discusses opportunities for improved design of mechanical environments that are conducive to tissue development.

Investigation of optical coherence micro-elastography as a method to visualize micro-architecture in human axillary lymph nodes.

BACKGROUND: Evaluation of lymph node involvement is an important factor in detecting metastasis and deciding whether to perform axillary lymph node dissection (ALND) in breast cancer surgery. As ALND is associated with potentially severe long term morbidity, the accuracy of lymph node assessment is imperative in avoiding unnecessary ALND. The mechanical properties of malignant lymph nodes are often distinct from those of normal nodes. A method to image the micro-scale mechanical properties of lymph nodes could, thus, provide diagnostic information to aid in the assessment of lymph node involvement in metastatic cancer. In this study, we scan axillary lymph nodes, freshly excised from breast cancer patients, with optical coherence micro-elastography (OCME), a method of imaging micro-scale mechanical strain, to assess its potential for the intraoperative assessment of lymph node involvement. METHODS: Twenty-six fresh, unstained lymph nodes were imaged from 15 patients undergoing mastectomy or breast-conserving surgery with axillary clearance. Lymph node specimens were bisected to allow imaging of the internal face of each node. Co-located OCME and optical coherence tomography (OCT) scans were taken of each sample, and the results compared to standard post-operative hematoxylin-and-eosin-stained histology. RESULTS: The optical backscattering signal provided by OCT alone may not provide reliable differentiation by inspection between benign and malignant lymphoid tissue. Alternatively, OCME highlights local changes in tissue strain that correspond to malignancy and are distinct from strain patterns in benign lymphoid tissue. The mechanical contrast provided by OCME complements the optical contrast provided by OCT and aids in the differentiation of malignant tumor from uninvolved lymphoid tissue. CONCLUSION: The combination of OCME and OCT images represents a promising method for the identification of malignant lymphoid tissue. This method shows potential to provide intraoperative assessment of lymph node involvement, thus, preventing unnecessary removal of uninvolved tissues and improving patient outcomes.

Quantitative micro-elastography: imaging of tissue elasticity using compression optical coherence elastography.

Probing the mechanical properties of tissue on the microscale could aid in the identification of diseased tissues that are inadequately detected using palpation or current clinical imaging modalities, with potential to guide medical procedures such as the excision of breast tumours. Compression optical coherence elastography (OCE) maps tissue strain with microscale spatial resolution and can delineate microstructural features within breast tissues. However, without a measure of the locally applied stress, strain provides only a qualitative indication of mechanical properties. To overcome this limitation, we present quantitative micro-elastography, which combines compression OCE with a compliant stress sensor to image tissue elasticity. The sensor consists of a layer of translucent silicone with well-characterized stress-strain behaviour. The measured strain in the sensor is used to estimate the two-dimensional stress distribution applied to the sample surface. Elasticity is determined by dividing the stress by the strain in the sample. We show that quantification of elasticity can improve the ability of compression OCE to distinguish between tissues, thereby extending the potential for inter-sample comparison and longitudinal studies of tissue elasticity. We validate the technique using tissue-mimicking phantoms and demonstrate the ability to map elasticity of freshly excised malignant and benign human breast tissues.

Investigation of Optical Coherence Microelastography as a Method to Visualize Cancers in Human Breast Tissue.

An accurate intraoperative identification of malignant tissue is a challenge in the surgical management of breast cancer. Imaging techniques that help address this challenge could contribute to more complete and accurate tumor excision, and thereby help reduce the current high reexcision rates without resorting to the removal of excess healthy tissue. Optical coherence microelastography (OCME) is a three-dimensional, high-resolution imaging technique that is sensitive to microscale variations of the mechanical properties of tissue. As the tumor modifies the mechanical properties of breast tissue, OCME has the potential to identify, on the microscale, involved regions of fresh, unstained tissue. OCME is based on the use of optical coherence tomography (OCT) to measure tissue deformation in response to applied mechanical compression. In this feasibility study on 58 ex vivo samples from patients undergoing mastectomy or wide local excision, we demonstrate the performance of OCME as a means to visualize tissue microarchitecture in benign and malignant human breast tissues. Through a comparison with corresponding histology and OCT images, OCME is shown to enable ready visualization of features such as ducts, lobules, microcysts, blood vessels, and arterioles and to identify invasive tumor through distinctive patterns in OCME images, often with enhanced contrast compared with OCT. These results lay the foundation for future intraoperative studies. Cancer Res; 75(16); 3236-45. ©2015 AACR.

Optical palpation in vivo: imaging human skin lesions using mechanical contrast.

We demonstrate the first application of the recently proposed method of optical palpation to in vivo imaging of human skin. Optical palpation is a tactile imaging technique that probes the spatial variation of a sample's mechanical properties by producing an en face map of stress measured at the sample surface. This map is determined from the thickness of a translucent, compliant stress sensor placed between a loading element and the sample and is measured using optical coherence tomography. We assess the performance of optical palpation using a handheld imaging probe on skin-mimicking phantoms, and demonstrate its use on human skin lesions. Our results demonstrate the capacity of optical palpation to delineate the boundaries of lesions and to map the mechanical contrast between lesions and the surrounding normal skin.

Three-dimensional optical coherence micro-elastography of skeletal muscle tissue.

In many muscle pathologies, impairment of skeletal muscle function is closely linked to changes in the mechanical properties of the muscle constituents. Optical coherence micro-elastography (OCME) uses optical coherence tomography (OCT) imaging of tissue under a quasi-static, compressive mechanical load to map variations in tissue mechanical properties on the micro-scale. We present the first study of OCME on skeletal muscle tissue. We show that this technique can resolve features of muscle tissue including fibers, fascicles and tendon, and can also detect necrotic lesions in skeletal muscle from the mdx mouse model of Duchenne muscular dystrophy. In many instances, OCME provides better or additional contrast complementary to that provided by OCT. These results suggest that OCME could provide new understanding and opportunity for assessment of skeletal muscle pathologies.

Optical palpation: optical coherence tomography-based tactile imaging using a compliant sensor.

We present optical palpation, a tactile imaging technique for mapping micrometer- to millimeter-scale mechanical variations in soft tissue. In optical palpation, a stress sensor consisting of translucent, compliant silicone with known stress-strain behavior is placed on the tissue surface and a compressive load is applied. Optical coherence tomography (OCT) is used to measure the local strain in the sensor, from which the local stress at the sample surface is calculated and mapped onto an image. We present results in tissue-mimicking phantoms, demonstrating the detection of a feature embedded 4.7 mm below the sample surface, well beyond the depth range of OCT. We demonstrate the use of optical palpation to delineate the boundary of a region of tumor in freshly excised human breast tissue, validated against histopathology.

Optical coherence micro-elastography: mechanical-contrast imaging of tissue microstructure.

We present optical coherence micro-elastography, an improved form of compression optical coherence elastography. We demonstrate the capacity of this technique to produce en face images, closely corresponding with histology, that reveal micro-scale mechanical contrast in human breast and lymph node tissues. We use phase-sensitive, three-dimensional optical coherence tomography (OCT) to probe the nanometer-to-micrometer-scale axial displacements in tissues induced by compressive loading. Optical coherence micro-elastography incorporates common-path interferometry, weighted averaging of the complex OCT signal and weighted least-squares regression. Using three-dimensional phase unwrapping, we have increased the maximum detectable strain eleven-fold over no unwrapping and the minimum detectable strain is 2.6 µÎµ. We demonstrate the potential of mechanical over optical contrast for visualizing micro-scale tissue structures in human breast cancer pathology and lymph node morphology.

Needle optical coherence elastography for the measurement of microscale mechanical contrast deep within human breast tissues.

Optical coherence elastography (OCE) is an emerging imaging technique that probes microscale mechanical contrast in tissues with the potential to differentiate healthy and malignant tissues. However, conventional OCE techniques are limited to imaging the first 1 to 2 mm of tissue in depth. We demonstrate, for the first time, OCE measurements deep within human tissues using needle OCE, extending the potential of OCE as a surgical guidance tool. We use needle OCE to detect tissue interfaces based on mechanical contrast in both normal and malignant breast tissues in freshly excised human mastectomy samples, as validated against histopathology. Further, we demonstrate the feasibility of in situ measurements >4 cm from the tissue surface using ultrasound guidance of the OCE needle probe. With further refinement, our method may potentially aid in accurate detection of the boundary of the tumor to help ensure full removal of all malignant tissues, which is critical to the success of breast-conserving surgery.

Analysis of mechanical contrast in optical coherence elastography.

Optical coherence elastography (OCE) maps the mechanical properties of tissue microstructure and has potential applications in both fundamental investigations of biomechanics and clinical medicine. We report the first analysis of contrast in OCE, including evaluation of the accuracy with which OCE images (elastograms) represent mechanical properties and the sensitivity of OCE to mechanical contrast within a sample. Using phase-sensitive compression OCE, we generate elastograms of tissue-mimicking phantoms with known mechanical properties and identify limitations on contrast imposed by sample mechanics and the imaging system, including signal-processing parameters. We also generate simulated elastograms using finite element models to perform mechanical analysis in the absence of imaging system noise. In both experiments and simulations, we illustrate artifacts that degrade elastogram accuracy, depending on sample geometry, elasticity contrast between features, and surface conditions. We experimentally demonstrate sensitivity to features with elasticity contrast as small as 1.1∶1 and calculate, based on our imaging system parameters, a theoretical maximum sensitivity to elasticity contrast of 1.002∶1. The results highlight the microstrain sensitivity of compression OCE, at a spatial resolution of tens of micrometers, suggesting its potential for the detection of minute changes in elasticity within heterogeneous tissue.

Improved measurement of vibration amplitude in dynamic optical coherence elastography.

Optical coherence elastography employs optical coherence tomography (OCT) to measure the displacement of tissues under load and, thus, maps the resulting strain into an image, known as an elastogram. We present a new improved method to measure vibration amplitude in dynamic optical coherence elastography. The tissue vibration amplitude caused by sinusoidal loading is measured from the spread of the Doppler spectrum, which is extracted using joint spectral and time domain signal processing. At low OCT signal-to-noise ratio (SNR), the method provides more accurate vibration amplitude measurements than the currently used phase-sensitive method. For measurements performed on a mirror at OCT SNR = 5 dB, our method introduces <3% error, compared to >20% using the phase-sensitive method. We present elastograms of a tissue-mimicking phantom and excised porcine tissue that demonstrate improvements, including a 50% increase in the depth range of reliable vibration amplitude measurement.

Strain estimation in phase-sensitive optical coherence elastography.

We present a theoretical framework for strain estimation in optical coherence elastography (OCE), based on a statistical analysis of displacement measurements obtained from a mechanically loaded sample. We define strain sensitivity, signal-to-noise ratio and dynamic range, and derive estimates of strain using three methods: finite difference, ordinary least squares and weighted least squares, the latter implemented for the first time in OCE. We compare theoretical predictions with experimental results and demonstrate a ~12 dB improvement in strain sensitivity using weighted least squares compared to finite difference strain estimation and a ~4 dB improvement over ordinary least squares strain estimation. We present strain images (i.e., elastograms) of tissue-mimicking phantoms and excised porcine airway, demonstrating in each case clear contrast based on the sample's elasticity.

Needle optical coherence elastography for tissue boundary detection.

We incorporate, for the first time, optical coherence elastography (OCE) into a needle probe and demonstrate its ability to measure the microscopic deformation of soft tissues located well beyond the depth limit of reports to date. Needle OCE utilizes the force imparted by the needle tip as the loading mechanism and measures tissue deformation ahead of the needle during insertion. Measurements were performed in tissue-mimicking phantoms and ex vivo porcine trachea. Results demonstrate differentiation of tissues based on mechanical properties and highlight the potential of needle OCE for in vivo tissue boundary detection.