Dongyao Cui

Dual spectrometer system with spectral compounding for 1-μm optical coherence tomography in vivo.

1 μm axial resolution spectral domain optical coherence tomography (OCT) is demonstrated for in vivo cellular resolution imaging. Output of two superluminescent diode sources is combined to provide near infrared illumination from 755 to 1105 nm. The spectral interference is detected using two spectrometers based on a Si camera and an InGaAs camera, respectively. Spectra from the two spectrometers are combined to achieve an axial resolution of 1.27 μm in air. Imaging was conducted on zebra fish larvae to visualize cellular details.

In vivo imaging of airway cilia and mucus clearance with micro-optical coherence tomography

We have designed and fabricated a 4 mm diameter rigid endoscopic probe to obtain high resolution micro-optical coherence tomography (µOCT) images from the tracheal epithelium of living swine. Our common-path fiber-optic probe used gradient-index focusing optics, a selectively coated prism reflector to implement a circular-obscuration apodization for depth-of-focus enhancement, and a common-path reference arm and an ultra-broadbrand supercontinuum laser to achieve high axial resolution. Benchtop characterization demonstrated lateral and axial resolutions of 3.4 μm and 1.7 μm, respectively (in tissue). Mechanical standoff rails flanking the imaging window allowed the epithelial surface to be maintained in focus without disrupting mucus flow. During in vivo imaging, relative motion was mitigated by inflating an airway balloon to hold the standoff rails on the epithelium. Software implemented image stabilization was also implemented during post-processing. The resulting image sequences yielded co-registered quantitative outputs of airway surface liquid and periciliary liquid layer thicknesses, ciliary beat frequency, and mucociliary transport rate, metrics that directly indicate airway epithelial function that have dominated in vitro research in diseases such as cystic fibrosis, but have not been available in vivo.

Depth extension and sidelobe suppression in optical coherence tomography using pupil filters

We demonstrate a new focus engineering scheme to achieve both extended depth of focus (DOF) and sidelobe suppression in spectral-domain optical coherence tomography (SD-OCT) system. Each of the illumination pupil function and the detection pupil function is modulated using an annular pupil filter implemented by center obscuration. The two pupil filters are arranged in a dark-field configuration such that the first sidelobe of the illumination point-spread function (PSF) matches the first minimum of the detection PSF in the lateral focal plane. We tested the feasibility of the proposed scheme numerically, and then constructed a dark-field OCT (DF-OCT) system to further verify its effectiveness experimentally. Simulation results show that a DOF gain of 4.2 can be achieved compared with a full aperture OCT (FA-OCT) system, with a suppression ratio of 2.9 dB for the first sidelobe compared with an annular-aperture bright-field OCT (BF-OCT) system. Experimental results show that the constructed DF-OCT extends the DOF by three-fold compared with the constructed FA-OCT, and suppresses the first sidelobe by 3.1 dB compared with the BF-OCT. The penalty for the extended DOF is an ~11.6 dB drop in sensitivity compared with the FA-OCT system.

Flexible, high-resolution micro-optical coherence tomography endobronchial probe toward in vivo imaging of cilia

We report the design and fabrication of a flexible, longitudinally scanning high-resolution micro-optical coherence tomography (μOCT) endobronchial probe, optimized for micro-anatomical imaging in airways. The 2.4 mm diameter and flexibility of the probe allows it to be inserted into the instrument channel of a standard bronchoscope, enabling real-time video guidance of probe placement. To generate a depth-of-focus enhancing annular beam, we utilized a new fabrication method, whereby a hollow glass ferrule was angle-polished and gold-coated to produce an elongated annular reflector. We present validation data that verifies the preservation of linear scanning, despite the use of flexible materials. When utilized on excised, cultured mouse trachea, the probe acquired images of comparable quality to those obtained by a benchtop μOCT system.

Spectral estimation optical coherence tomography for axial super-resolution.

The depth reflectivity profile of Fourier domain optical coherence tomography (FD-OCT) is estimated from the inverse Fourier transform of the spectral interference signals (interferograms). As a result, the axial resolution is fundamentally limited by the coherence length of the light source. We demonstrate that using the autoregressive spectral estimation technique instead of the inverse Fourier transform, to analyze the spectral interferograms can improve the axial resolution. We name this method spectral estimation OCT (SE-OCT). SE-OCT breaks the coherence length limitation and improves the axial resolution by a factor of up to 4.7 compared with FD-OCT. Furthermore, SE-OCT provides complete sidelobe suppression in the depth point-spread function, further improving the image quality. We demonstrate that these technical advances enables clear identification of corneal endothelium anatomical details ex vivo that cannot be identified using the corresponding FD-OCT. Given that SE-OCT can be implemented in the FD-OCT devices without any hardware changes, the new capabilities provided by SE-OCT are likely to offer immediate improvements to the diagnosis and management of diseases based on OCT imaging.

Development of a Primary Human Co-Culture Model of Inflamed Airway Mucosa

Neutrophil breach of the mucosal surface is a common pathological consequence of infection. We present an advanced co-culture model to explore neutrophil transepithelial migration utilizing airway mucosal barriers differentiated from primary human airway basal cells and examined by advanced imaging. Human airway basal cells were differentiated and cultured at air-liquid interface (ALI) on the underside of 3 µm pore-sized transwells, compatible with the study of transmigrating neutrophils. Inverted ALIs exhibit beating cilia and mucus production, consistent with conventional ALIs, as visualized by micro-optical coherence tomography (µOCT). µOCT is a recently developed imaging modality with the capacity for real time two- and three-dimensional analysis of cellular events in marked detail, including neutrophil transmigratory dynamics. Further, the newly devised and imaged primary co-culture model recapitulates key molecular mechanisms that underlie bacteria-induced neutrophil transepithelial migration previously characterized using cell line-based models. Neutrophils respond to imposed chemotactic gradients, and migrate in response to Pseudomonas aeruginosa infection of primary ALI barriers through a hepoxilin A3-directed mechanism. This primary cell-based co-culture system combined with µOCT imaging offers significant opportunity to probe, in great detail, micro-anatomical and mechanistic features of bacteria-induced neutrophil transepithelial migration and other important immunological and physiological processes at the mucosal surface.

Illuminating dynamic neutrophil trans-epithelial migration with micro-optical coherence tomography

A model of neutrophil migration across epithelia is desirable to interrogate the underlying mechanisms of neutrophilic breach of mucosal barriers. A co-culture system consisting of a polarized mucosal epithelium and human neutrophils can provide a versatile model of trans-epithelial migration in vitro, but observations are typically limited to quantification of migrated neutrophils by myeloperoxidase correlation, a destructive assay that precludes direct longitudinal study. Our laboratory has recently developed a new isotropic 1-μm resolution optical imaging technique termed micro-optical coherence tomography (μOCT) that enables 4D (x,y,z,t) visualization of neutrophils in the co-culture environment. By applying μOCT to the trans-epithelial migration model, we can robustly monitor the spatial distribution as well as the quantity of neutrophils chemotactically crossing the epithelial boundary over time. Here, we demonstrate the imaging and quantitative migration results of our system as applied to neutrophils migrating across intestinal epithelia in response to a chemoattractant. We also demonstrate that perturbation of a key molecular event known to be critical for effective neutrophil trans-epithelial migration (CD18 engagement) substantially impacts this process both qualitatively and quantitatively.

Single-camera full-range high-resolution spectral domain optical coherence tomography

We developed spectral domain optical coherence tomography using a dual-channel spectrometer for complex conjugate artifacts (CCA) suppression. We used a three-line charge coupled device to simultaneously detect two interferometric spectra with 2π/3 phase difference. The complex interferometric signal was reconstructed by trigonometric manipulation of two real interferometric spectra, and then full-range images were obtained by use of inverse Fourier transform. Artifacts at direct current (DC) and the ghost remnant of the CCA are common issues with the previously reported two-spectrometer method because the slight mismatching between two spectral detection channels had strong negative effects on CCA suppression and appeared to be the limiting factor on system performance. This novel dual-channel spectrometer uses the same spectrometer optics for the two spectral detection channels and, therefore, achieves better matching between two spectral detection channels and consequently better performance in CCA suppression as compared with the dual spectrometer solution. Full-range imaging with CCA suppression up to ∼25  dB was demonstrated when imaging an attenuated reflector. The efficacy of both CCA and DC suppressions also was validated by imaging the anterior segment of a rat eye ex vivo. The quality of CCA-suppressed images was significantly improved with regard to those obtained with the dual-spectrometer design.

Modeling of Mechanical Stress Exerted by Cholesterol Crystallization on Atherosclerotic Plaques

Plaque rupture is the critical cause of cardiovascular thrombosis, but the detailed mechanisms are not fully understood. Recent studies have found abundant cholesterol crystals in ruptured plaques, and it has been proposed that the rapid expansion of cholesterol crystals in a limited space during crystallization may contribute to plaque rupture. To evaluate the effect of cholesterol crystal growth on atherosclerotic plaques, we modeled the expansion of cholesterol crystals during the crystallization process in the necrotic core and estimated the stress on the thin cap with different arrangements of cholesterol crystals. We developed a two-dimensional finite element method model of atherosclerotic plaques containing expanding cholesterol crystals and investigated the effect of the magnitude and distribution of crystallization on the peak circumferential stress born by the cap. Using micro-optical coherence tomography (μOCT), we extracted the cross-sectional geometric information of cholesterol crystals in human atherosclerotic aorta tissue ex vivo and applied the information to the model. The results demonstrate that (1) the peak circumference stress is proportionally dependent on the cholesterol crystal growth; (2) cholesterol crystals at the cap shoulder impose the highest peak circumference stress; and (3) spatial distributions of cholesterol crystals have a significant impact on the peak circumference stress: evenly distributed cholesterol crystals exert less peak circumferential stress on the cap than concentrated crystals.

Multifiber angular compounding optical coherence tomography for speckle reduction

We report on an integrated fiber optic design to implement multifiber angular compounding optical coherence tomography, which enables angular compounding for speckle reduction. A multi-facet fiber array delivers three light beams to the sample with different incident angles. Back-reflective/back-scattered signals from these channels were simultaneously detected by a three-channel spectrometer. The axial and lateral resolution was measured to be ∼3 and ∼3.5  μm, respectively, in air with ∼100  dB sensitivity. We conducted ex vivo experiments on a rat esophagus to demonstrate a contrast to noise improvement of 1.58.