Ultrahigh Sensitive Optical Microangiography Research Articles

Neovascularization in Alkali-Burned Rabbit Cornea

Objective: To study the neovascularization in regenerating and proliferating corneal cells following a standard alkali injury in rabbit eye. Methods: Three and six weeks after the creation of an alkali burn in the center of the right cornea of six albino rabbits, the animals were killed and histological sections from the cornea of both eyes were stained, photographed and studied for a possible formation of a neovascularization. The photographs were examined using the Quantimet image analyzer (Leica) and statistical analysis of the data was performed. Results: Sections of the injured cornea showed the formation of neovessels in the epithelial and superficial stromal layers. The neovascularization is present after 3 weeks of the corneal injury. After 6 weeks from the corneal alkali burn, neovessels are increased. Conclusions: There is a growing body of evidence suggesting that vascular abnormalities may play a crucial role in several ocular diseases. To improve our knowledge of the vascular involvement in these conditions, there is a need for a non-invasive imaging modality capable of assessing microcirculation within ocular tissue beds both in vitro and in vivo. This study shows that ultra-high sensitive optical microangiography, associated with other experimental techniques, is an adequate technique to visualize the eye surface microcirculations and to quantify microvascular vessel density under both normal and physio-pathological conditions.

Application of thinned-skull cranial window to mouse cerebral blood flow imaging using optical microangiography.

In vivo imaging of mouse brain vasculature typically requires applying skull window opening techniques: open-skull cranial window or thinned-skull cranial window. We report non-invasive 3D in vivo cerebral blood flow imaging of C57/BL mouse by the use of ultra-high sensitive optical microangiography (UHS-OMAG) and Doppler optical microangiography (DOMAG) techniques to evaluate two cranial window types based on their procedures and ability to visualize surface pial vessel dynamics. Application of the thinned-skull technique is found to be effective in achieving high quality images for pial vessels for short-term imaging, and has advantages over the open-skull technique in available imaging area, surgical efficiency, and cerebral environment preservation. In summary, thinned-skull cranial window serves as a promising tool in studying hemodynamics in pial microvasculature using OMAG or other OCT blood flow imaging modalities.

Volumetric cutaneous microangiography of human skin in vivo by VCSEL swept-source optical coherence tomography

We demonstrate volumetric cutaneous microangiography of the human skin in vivo that utilises high-speed swept-source optical coherence tomography (SS-OCT). The swept source is based on a micro-electro-mechanical (MEMS)-tunable vertical cavity surface emission laser (VCSEL) that is advantageous in terms of long coherence length over and spectral bandwidth, which enables the visualisation of microstructures within a few mm from the skin surface. We show that the skin microvasculature can be delineated in 3D SS-OCT images using ultrahigh-sensitive optical microangiography (UHS-OMAG) with a correlation mapping mask, providing a contrast enhanced blood perfusion map with capillary flow sensitivity. 3D microangiograms of a healthy human finger are shown with distinct cutaneous vessel architectures from different dermal layers and even within hypodermis. These findings suggest that the OCT microangiography could be a beneficial biomedical assay to assess cutaneous vascular functions in clinic.

In vivo OCT microangiography of rodent iris.

We report on the functional optical coherence tomography (OCT) imaging of iris tissue morphology and microcirculation in living small animals. Anterior segments of healthy mouse and rat eyes are imaged with high-speed spectral domain OCT (SD-OCT) utilizing ultrahigh sensitive optical microangiography (UHS-OMAG) imaging protocol. 3D iris microvasculature is produced by the use of an algorithm that calculates absolute differences between the amplitudes of the OCT interframes. We demonstrate that the UHS-OMAG is capable of delineating iris microvascular beds in the mouse and rat with capillary-level resolution. Furthermore, the fast imaging speed enables dynamic imaging of iris micro-vascular response during drug-induced pupil dilation. We believe that this OCT angiographic approach has a great potential for in situ and in vivo monitoring of the microcirculation within iris tissue beds in rodent disease models that have microvascular involvement.

Noninvasive Imaging of Retinal Morphology and Microvasculature in Obese Mice Using Optical Coherence Tomography and Optical Microangiography

To evaluate early diabetes-induced changes in retinal thickness and microvasculature in a type 2 diabetic mouse model by using optical coherence tomography (OCT)/optical microangiography (OMAG). Twenty-two-week-old obese (OB) BTBR mice (n = 10) and wild-type (WT) control mice (n = 10) were imaged. Three-dimensional (3D) data volumes were captured with spectral domain OCT using an ultrahigh-sensitive OMAG scanning protocol for 3D volumetric angiography of the retina and dense A-scan protocol for measurement of the total retinal blood flow (RBF) rate. The thicknesses of the nerve fiber layer (NFL) and that of the NFL to the inner plexiform layer (IPL) were measured and compared between OB and WT mice. The linear capillary densities within intermediate and deep capillary layers were determined by the number of capillaries crossing a 500-μm line. The RBF rate was evaluated using an en face Doppler approach. These quantitative measurements were compared between OB and WT mice. The retinal thickness of the NFL to IPL was significantly reduced in OB mice (P < 0.01) compared to that in WT mice, whereas the NFL thickness between the two was unchanged. 3D depth-resolved OMAG angiography revealed the first in vivo 3D model of mouse retinal microcirculation. Although no obvious differences in capillary vessel densities of the intermediate and deep capillary layers were detected between normal and OB mice, the total RBF rate was significantly lower (P < 0.05) in OB mice than in WT mice. We conclude that OB BTBR mice have significantly reduced NFL-IPL thicknesses and total RBF rates compared with those of WT mice, as imaged by OCT/OMAG. OMAG provides an unprecedented capability for high-resolution depth-resolved imaging of mouse retinal vessels and blood flow that may play a pivotal role in providing a noninvasive method for detecting early microvascular changes in patients with diabetic retinopathy.

Capillary blood flow imaging within human finger cuticle using optical microangiography.

We report non-invasive 3D imaging of capillary blood flow within human finger cuticle by the use of Doppler optical microangiography (DOMAG) and ultra-high sensitive optical microangiography (UHS-OMAG) techniques. Wide velocity range DOMAG method is applied to provide red blood cell (RBC) axial velocity mapping in capillary loops with ranges of ±0.9 mm/s and ±0.3 mm/s. Additionally, UHS-OMAG technique is engineered to acquire high resolution image of capillary morphology. The presented results are promising to facilitate clinical trials of treatment and diagnosis of various diseases such as diabetes, Raynaud's phenomenon, and connective tissue diseases by quantifying cutaneous blood flow changes within human finger cuticle.

Super-resolution spectral estimation of optical micro-angiography for quantifying blood flow within microcirculatory tissue beds in vivo

In this paper, we propose a super-resolution spectral estimation technique to quantify microvascular hemodynamics using optical microangiography (OMAG) based on optical coherence tomography (OCT). The proposed OMAG technique uses both amplitude and phase information of the OCT signals which makes it sensitive to the axial and transverse flows. The scanning protocol for the proposed method is identical to three-dimensional ultrahigh sensitive OMAG, and is applicable for in vivo measurements. In contrast to the existing capillary flow quantification methods, the proposed method is less sensitive to tissue motion and does not have aliasing problems due fast flow within large blood vessels. This method is analogous to power Doppler in ultrasonography and estimates the number of red blood cells passing through the beam as opposed to the velocity of the particles. The technique is tested both qualitatively and quantitatively by using OMAG to image microcirculation within mouse ear flap in vivo.

Optical microangiography provides correlation between microstructure and microvasculature of optic nerve head in human subjects

It is demonstrated that optical microangiography (OMAG) is capable of imaging the detailed microstructure and microvasculature of the in vivo human optic nerve head (ONH), including the prelaminar tissue, the lamina cribrosa, the scleral rim and the vessels in the region of the circle of Zin-Haller. For demonstration, an ultrahigh sensitive OMAG system operating in the 850 nm wavelength region and a 500 kHz A-scan rate resulting in a spatial resolution of ∼6 μm were used. It was shown that OMAG provides superior results for three-dimensional imaging of the ONH compared to conventional optical coherence tomography by simultaneously recording both the microstructure and the functional microcirculation. The blood supply to the tissues of the ONH is an essential physiologic parameter needed for clinical assessment of the health of the nerve.

Supercontinuum light source enables in vivooptical microangiography of capillary vessels within tissue beds

This Letter reports on the use of a supercontinuum light source to achieve ultrahigh resolution and ultrahigh sensitive optical microangiography (OMAG) imaging of microcirculations within tissue beds in vivo. After passing through a specially designed optical filter with a passband of 120 nm centered on 800 nm, the light source is coupled into an optic-fiber-based OMAG system that provides a measured axial resolution of ∼3 μm over a ranging distance of 2 mm. Within this ranging distance, the system gives an averaged signal-to-noise ratio of 87 dB and a sensitivity roll-off of 7 dB at an A-scan rate of 70 kHz. We demonstrate the capability of the system to visualize a detailed microvascular perfusion map, including the single red blood cells within the capillaries, by imaging a mouse ear flap in vivo.

Highly sensitive imaging of renal microcirculation in vivo using ultrahigh sensitive optical microangiography

Studying renal microcirculation and its dynamics is of great importance for understanding the renal function and further aiding the diagnosis, prevention and treatment of renal pathologies. In this paper, we present a potentially useful method to provide high-sensitive volumetric imaging of renal microcirculations using ultrahigh-sensitive optical microangiography (UHS-OMAG). The UHS-OMAG image system used here is based on spectral domain optical coherence tomography, which uses a broadband light source centered at 1300 nm with an imaging speed of 150 frames per second that requires ~6.7 sec to complete one 3D scan of ~2.5 × 2.5 mm2 area. The technique is sensitive enough to image capillary networks, such as peritubular capillaries within renal cortex. We show the ability of UHS-OMAG to provide depth-resolved volumetric images of capillary level renal microcirculation. We also show that UHS-OMAG is capable of monitoring the changes of renal microcirculation in response to renal ischemia and reperfusion. Finally, we attempt to show the capability of OMAG to provide quantitative analysis about velocity changes in a single capillary vessel (down to tens of microns per second) in response to the ischemic event.

Full range complex ultrahigh sensitive optical microangiography

This Letter presents a useful method that combines the full range complex Fourier domain optical coherence tomography (OCT) with the ultrahigh sensitive optical microangiography (OMAG) to achieve full range complex imaging of blood flow within microcirculatory tissue beds in vivo. We propose to use the fast scanning axis to realize the full range complex imaging, while using the slow axis to achieve OMAG imaging of blood flow. We demonstrate the proposed method by using a high speed 1310 nm OCT/OMAG system running at 92 kHz line scan rate to image the flow phantoms in vitro, and the blood flows in tissue beds in vivo.

In vivo volumetric imaging of microcirculation within human skin under psoriatic conditions using optical microangiography.

There is a growing body of evidence suggesting that vascular abnormalities may play crucial role in several dermatologic diseases, such as psoriasis, port wine stain, and skin cancer. To improve our understanding of vascular involvement in these skin conditions, there is a need for a non-invasive imaging modality capable of assessing 3D microcirculations within skin tissue beds in vivo. This study aims to demonstrate whether ultra-high sensitive optical microangiography (UHS-OMAG) is feasible to visualize skin microcirculations in 3D and to quantify microvascular vessel density under normal and psoriatic conditions in vivo. An UHS-OMAG system operating at 1,310 nm wavelength was used for in vivo imaging of microcirculation in human skin. The system has a spatial resolution of ∼10 µm × 20 µm (axial × lateral), running at 280 fps to acquire 3D imaging dataset to represent morphology and capillary level microvascular blood perfusion within the scanned skin tissue volume. The sensitivity of the system to the blood flow is as low as ∼4 µm/second. With this system, we performed the imaging experiments on the skin of a volunteer with stable plaque-type psoriasis. The microcirculation and structural information of normal and diseased skins were compared both qualitatively and quantitatively. The UHS-OMAG is capable of differentiating the microcirculation within the normal skins from that in the psoriatic skins. The 3D optical images show that the blood vessel elongation and the dense network in the psoriatic lesion skin, the appearance of which is not observed within the normal skin. Based on the results obtained from one subject, the statistical analyses show that higher blood vessel density presented within the psoriasis lesion skin than that of the normal skin. UHS-OMAG can be a valuable tool for imaging skin microcirculations non-invasively with high-speed and high-sensitivity, and therefore may have a useful role in future clinical diagnosis and treatment of dermatologic diseases such as psoriasis in human subjects.

Using ultrahigh sensitive optical microangiography to achieve comprehensive depth resolved microvasculature mapping for human retina

This paper presents comprehensive and depth-resolved retinal microvasculature images within human retina achieved by a newly developed ultrahigh sensitive optical microangiography (UHS-OMAG) system. Due to its high flow sensitivity, UHS-OMAG is much more sensitive to tissue motion due to the involuntary movement of the human eye and head compared to the traditional OMAG system. To mitigate these motion artifacts on final imaging results, we propose a new phase compensation algorithm in which the traditional phase-compensation algorithm is repeatedly used to efficiently minimize the motion artifacts. Comparatively, this new algorithm demonstrates at least 8 to 25 times higher motion tolerability, critical for the UHS-OMAG system to achieve retinal microvasculature images with high quality. Furthermore, the new UHS-OMAG system employs a high speed line scan CMOS camera (240 kHz A-line scan rate) to capture 500 A-lines for one B-frame at a 400 Hz frame rate. With this system, we performed a series of in vivo experiments to visualize the retinal microvasculature in humans. Two featured imaging protocols are utilized. The first is of the low lateral resolution (16 μm) and a wide field of view (4 × 3 mm(2) with single scan and 7 × 8 mm(2) for multiple scans), while the second is of the high lateral resolution (5 μm) and a narrow field of view (1.5 × 1.2 mm(2) with single scan). The great imaging performance delivered by our system suggests that UHS-OMAG can be a promising noninvasive alternative to the current clinical retinal microvasculature imaging techniques for the diagnosis of eye diseases with significant vascular involvement, such as diabetic retinopathy and age-related macular degeneration.

Optical microangiography provides an ability to monitor responses of cerebral microcirculation to hypoxia and hyperoxia in mice

In vivo imaging of microcirculation can improve our fundamental understanding of cerebral microhemodynamics under various physiological challenges, such as hypoxia and hyperoxia. However, existing techniques often involve the use of invasive procedures or exogenous contrast agents, which would inevitably perturb the intrinsic physiologic responses of microcirculation being investigated. We report ultrahigh sensitive optical microangiography (OMAG) for label-free monitoring of microcirculation responses challenged by oxygen inhalation. For the first time, we demonstrate that OMAG is capable of showing the impact of acute hypoxia and hyperoxia on microhemodynamic activities, including the passive and active modulation of microvascular density and flux regulation, within capillary and noncapillary vessels in rodents in vivo. The ability of OMAG to functionally image the intact microcirculation promises future applications for studying cerebral diseases.

Ultrahigh sensitive optical microangiography reveals depth-resolved microcirculation and its longitudinal response to prolonged ischemic event within skeletal muscles in mice

The primary pathophysiology of peripheral arterial disease is associated with impaired perfusion to the muscle tissue in the lower extremities. The lack of effective pharmacologic treatments that stimulate vessel collateralization emphasizes the need for an imaging method that can be used to dynamically visualize depth-resolved microcirculation within muscle tissues. Optical microangiography (OMAG) is a recently developed label-free imaging method capable of producing three-dimensional images of dynamic blood perfusion within microcirculatory tissue beds at an imaging depth of up to ∼2 mm, with an unprecedented imaging sensitivity of blood flow at ∼4 μm∕s. In this paper, we demonstrate the utility of OMAG in imaging the detailed blood flow distributions, at a capillary-level resolution, within skeletal muscles of mice. By use of the mouse model of hind-limb ischemia, we show that OMAG can assess the time-dependent changes in muscle perfusion and perfusion restoration along tissue depth. These findings indicate that OMAG can represent a sensitive, consistent technique to effectively study pharmacologic therapies aimed at promoting the growth and development of collateral vessels.

Volumetric in vivo imaging of microvascular perfusion within the intact cochlea in mice using ultra-high sensitive optical microangiography.

Studying the inner ear microvascular dynamics is extremely important to understand the cochlear function and to further advance the diagnosis, prevention, and treatment of many otologic disorders. However, there is currently no effective imaging tool available that is able to access the blood flow within the intact cochlea. In this paper, we report the use of an ultrahigh sensitive optical micro-angiography (UHS-OMAG) imaging system to image 3-D microvascular perfusion within the intact cochlea in living mice. The UHS-OMAG image system used in this study is based on spectral domain optical coherence tomography, which uses a broadband light source centered at 1300 nm with an imaging rate of 47[Formula: see text] 000 A-scans/s, capable of acquiring high-resolution B scans at 300 frames/s. The technique is sensitive enough to image very slow blood flow velocities, such as those found in capillary networks. The 3-D imaging acquisition time for a whole cochlea is ∼4.1 s. We demonstrate that volumetric reconstruction of microvascular flow obtained by UHS-OMAG provides a comprehensive perfusion map of several regions of the cochlea, including the otic capsule, the stria vascularis of the apical and middle turns and the radiating arterioles that emanate from the modiolus.

Depth-resolved imaging of capillary networks in retina and choroid using ultrahigh sensitive optical microangiography

We demonstrate the depth-resolved and detailed ocular perfusion maps within retina and choroid can be obtained from an ultrahigh sensitive optical microangiography (OMAG). As opposed to the conventional OMAG, we apply the OMAG algorithm along the slow scanning axis to achieve the ultrahigh sensitive imaging to the slow flows within capillaries. We use an 840 nm system operating at an imaging rate of 400 frames/s that requires 3 s to complete one 3D scan of approximately 3 x 3 mm(2) area on retina. We show the superior imaging performance of OMAG to provide functional images of capillary level microcirculation at different land-marked depths within retina and choroid that correlate well with the standard retinal pathology.

Ultrahigh sensitive optical microangiography for in vivo imaging of microcirculations within human skin tissue beds

In this paper, we demonstrate for the first time that the detailed cutaneous blood flow at capillary level within dermis of human skin can be imaged by optical micro-angiography (OMAG) technique. A novel scanning protocol, i.e. fast B scan mode is used to achieve the capillary flow imaging. We employ a 1310nm system to scan the skin tissue at an imaging rate of 300 frames per second, which requires only ~5 sec to complete one 3D imaging of capillary blood flow within skin. The technique is sensitive enough to image the very slow blood flows at ~4 microm/sec. The promising results show a great potential of OMAG's role in the diagnosis, treatment and management of human skin diseases.

Label-free and highly sensitive optical imaging of detailed microcirculation within meninges and cortex in mice with the cranium left intact

We demonstrate for the first time that the detailed blood flow distribution within intracranial dura mater and cortex can be visualized by an ultrahigh sensitive optical microangiography (UHS-OMAG). The study uses an UHS-OMAG system operating at 1310 nm with an imaging speed at 150 frames per second that requires approximately 10 s to complete one 3-D scan of approximately 2.5 x 2.5 mm(2). The system is sensitive to blood flow with a velocity ranging from approximately 4 microms to approximately 23 mms. We show superior performance of UHS-OMAG in providing functional images of capillary level microcirculation within meninges in mice with the cranium left intact, the results of which correlate well with the standard dural histopathology.

Three-dimensional optical imaging of microvascular networks within intact lymph node in vivo

Sentinel lymph nodes (SLNs) are the first lymph nodes to drain wastes originated from cancerous tissue. There is a need for an in vivo imaging method that can image the intact SLN to further our understanding of its normal as well as abnormal functions. We report the use of ultrahigh sensitive optical microangiography (UHS-OMAG) to image functional microvascular and lymphatic vessel networks that innervate the intact lymph node in mice in vivo. The promising results show a potential role of UHS-OMAG in the future understanding and diagnosis of the SLN involvement in cancer development.