Sean J. Kirkpatrick

Can laser speckle flowmetry be made a quantitative tool

The ultimate objective of laser speckle flowmetry (and a host of specific implementations such as laser speckle contrast analysis, LASCA or LSCA; laser speckle spatial contrast analysis, LSSCA; laser speckle temporal contrast analysis, LSTCA; etc.) is to infer flow velocity from the observed speckle contrast. Despite numerous demonstrations over the past 25 years of such a qualitative relationship, no convincing quantitative relationship has been proven. One reason is a persistent mathematical error that has been propagated by a host of workers; another is a misconception about the proper autocorrelation function for ordered flow. Still another hindrance has been uncertainty in the specific relationship between decorrelation time and local flow velocity. Herein we attempt to dispel some of these errors and misconceptions with the intent of turning laser speckle flowmetry into a quantitative tool. Specifically we review the underlying theory, explore the impact of various analytic models for relating measured intensity fluctuations to scatterer motion, and address some of the practical issues associated with the measurement and subsequent data processing.

Phase-sensitive optical coherence elastography for mapping tissue microstrains in real time

The authors present a phase-sensitive optical coherence elastography (PSOCE) approach to image instantaneous tissue deformations, strain rates, and strains of soft tissue in real time with sensitivity at the nanometer scale. This method exploits the phase information available in the complex optical coherence tomography images and measures the phase changes between the successive B scans to resolve the instantaneous tissue deformations. The PSOCE system described is capable of producing localized microstrain rate and strain maps of tissue subjected to a dynamic compression in real time. They show that this approach is capable of resolving deformations as small as 0.26nm.

Tissue Doppler optical coherence elastography for real time strain rate and strain mapping of soft tissue

The authors present a tissue Doppler optical coherence elastography (tDOCE) method to image tissue movements, strain rates, and strains of soft tissue in real time. The method exploits the Doppler effect in optical coherence interferograms induced by tissue motion and measures the phase changes between successive A scans to resolve the instantaneous tissue displacement. The tDOCE system is capable of displaying the strain rates and strain maps of tissue subjected to a dynamic compression in real time. The system is demonstrated by the use of a heterogeneous tissue phantom with known mechanical properties.

Detrimental effects of speckle-pixel size matching in laser speckle contrast imaging

Through a series of simulations and experiments, we demonstrate that the frequently cited criterion of matching speckle size to detector element (pixel) size in laser speckle contrast imaging (LSCI) has the detrimental effect of reducing the contrast and thereby decreasing the variation in the laser speckle contrast image. Unlike quasi-elastic light scattering, where this matching condition has been shown to maximize the signal-to-noise ratio, in LSCI, the minimum speckle size must exceed the Nyquist criterion in order to maximize the contrast of the speckle patterns.

OCT-Based Elastography for Large and Small Deformations

We present two approaches to speckle tracking for optical coherence tomography (OCT)-based elastography, one appropriate for small speckle motions and the other for large, rapid speckle motions. Both approaches have certain advantages over traditional cross-correlation based motion algorithms. We apply our algorithms to quantifying the strain response of a mechanically inhomogeneous, bi-layered polyvinyl alcohol tissue phantom that is subjected to either small or large dynamic compressive forces while being imaged with a spectral domain OCT system. In both the small and large deformation scenarios, the algorithms performed well, clearly identifying the two mechanically disparate regions of the phantom. The stiffness ratio between the two regions was estimated to be the same for the two scenarios and both estimates agreed with the expected stiffness ratio based on earlier mechanical testing. No single numerical approach is appropriate for all cases and the experimental conditions dictate the proper choice of speckle shift algorithm for OCT-based elastography studies.

Micromechanical behavior of cortical bone as inferred from laser speckle data.

We investigated the micromechanical behavior of porcine femoral cortical bone using a novel, nondestructive, noncontacting, laser-based strain measurement technique. The technique is based upon the well-known concept of tracking translating laser speckle with a linear array CCD camera, but employs a unique data-processing scheme based upon a two-dimensional frequency transform of the data. The method proved to be successful in evaluating strain rates in the bone samples. Measured strain rates ranged between 4.61 and 23.84 micro epsilon/s. Total strains recorded were between 3.7 and 19.1 micro epsilon. Estimated Youngs moduli averaged 9.01 +/- 3.93 GPa, which, considering the extremely slow strain rates, is an acceptable value for porcine cortical bone. General advantages of the technique include high sensitivity, insensitivity to zero-mean noise sources, compact design, and the fact that it is truly noncontact. A brief discussion of possible error sources within the method is also given.

Endothelial cell cytoskeletal alignment independent of fluid shear stress on micropatterned surfaces

Endothelial cells (ECs) in athero-protective regions are elongated with actin and microtubule fibers aligned parallel to the direction of blood flow. Fluid shear stress (FSS) affects EC shape and functions, but little is known about shape-dependent EC properties that are independent of FSS. To evaluate these properties, ECs were elongated on micropatterned (MP) 25mum wide collagen-coated lanes (MPECs) and characterized by cell shape index, actin and microtubule alignment, and polarization of the microtubule-organizing center (MTOC). ECs on non-patterned surfaces were also exposed to FSS. MPEC elongation was microtubule-dependent (and actin-independent); shape indices and cytoskeletal alignment were comparable to FSS-elongated ECs. Cytoskeletal alignment was lost when MPECs were exposed to perpendicular FSS, but not parallel FSS. MTOC polarization was FSS-dependent. Thus, by isolating EC elongation and cytoskeletal alignment from FSS, micropatterning creates a platform for studying EC shape-related cellular functions that are independent of FSS.

Optical assessment of tissue mechanical properties

Many disease processes, such as scirrhous carcinoma of the breast and atherosclerosis physically manifest themselves as changes in the mechanical properties of the involved tissue. Optical evaluation of the mechanical properties of biological tissues offers methods that can directly provide information regarding the state of health of both hard and soft tissue. Clinically, this information may be used for diagnostic purposes. Since all biological tissues display viscoelastic behavior and therefore exhibits a time dependence of their mechanical behavior, the important quantities for evaluating the mechanics of tissue under load are those that reflect this time dependence. For example, by evaluating the retardation spectrum of the tissue, the underlying molecular processes that govern the mechanical behavior may be elucidated. Understanding these spectra may provide insight into pathological processes and also provide some guidance for the development of synthetic materials or engineered tissues to replace damaged or pathological tissue. Thus, there is a need for methods that directly assess the time rate of the mechanical response of tissue to an imposed load. Herein, a few such methods will be discussed and applications to biomaterials and medical diagnostics will be discussed.

Imaging the mechanical stiffness of skin lesions by in vivo acousto-optical elastography.

Optical elastography is an imaging modality that relies on variations in the local mechanical properties of biological tissues as the contrast mechanism for image formation. Skin lesions, such as melanomas and other invasive conditions, are known to alter the arrangement of collagen fibers in the skin and thus should lead to alterations in local skin mechanical properties. We report on an acousto-optical elastography (AOE) imaging modality for quantifying the mechanical behavior of skin lesions. The method relies upon stimulating the tissue with a low frequency acoustic force and imaging the resulting strains in the tissue by means of quantifying the magnitude of the dynamic shift in a back-reflected laser speckle pattern from the skin. The magnitude of the shift reflects the local stiffness of the tissue. We demonstrate AOE on a tissue-mimicking phantom, an in vivo mouse melanoma lesion and two types of in vivo human melanocytic nevi. The skin lesions we examined were found to have distinct mechanical properties that appear to correlate with the varying degrees of dermal involvement of the lesions.

Distinct extracellular matrix microenvironments of progenitor and carotid endothelial cells.

Endothelial cells (ECs) produce and maintain the local extracellular matrix (ECM), a critical function that contributes to EC and blood vessel health. This function is also crucial to vascular tissue engineering, where endothelialization of vascular constructs require a cell source that readily produces and maintains ECM. In this study, baboon endothelial progenitor cell (EPC) deposition of ECM (laminin, collagen IV, and fibronectin) was characterized and compared to mature carotid ECs, evaluated in both elongated and cobblestone morphologies typically found in vivo. Microfluidic micropatterning was used to create 15-microm wide adhesive lanes with 45-microm spacing to reproduce the elongated EC morphology without the influence of external forces. Both EPCs and ECs elongated on micropatterned lanes had aligned actin cytoskeleton and readily deposited ECM. EPCs deposited and remodeled the ECM to a greater extent than ECs. Since a readily produced ECM can improve graft patency, EPCs are an advantageous cell source for endothelializing vascular constructs. Furthermore, EC deposition of ECM was dependent on cell morphology, where elongated ECs deposited more collagen IV and less fibronectin compared to matched cobblestone controls. Thus micropatterned surfaces controlled EC shape and ECM deposition, which ultimately has implications for the design of tissue-engineered vascular constructs.