KEYWORDS: Optical coherence tomography, Skin, Angiography, Tissues, Control systems, Vascular diseases, Capillaries, Analog to digital converters, 3D image processing, Tunable filters
Systemic sclerosis (SSc) is a complex autoimmune disease characterized by skin and internal organ fibrosis, with vascular dysfunction playing a critical role in its pathogenesis. This study utilizes optical coherence tomography angiography (OCTA) to explore vascular abnormalities in SSc patients. We imaged 26 SSc patients and 17 age- and sex-matched healthy volunteers. High-resolution OCTA images of the forearm, hand, finger, and nailbed skin vasculature were obtained using a swept-source OCT system, with a central wavelength of 1300 nm, a scan range of 108 nm, a scan rate of 100 kHz. Post-processing was achieved using Matlab and QuPath, where a Hessian filter-based approach was utilized to enhance blood vessel contrast and connectivity in 2D projections. These tools were also employed for vessel lumen width (VLW) calculation, with group comparisons made using Mann-Whitney U-test. The results showed a significant decrease in VLW in SSc patients compared to healthy controls across all imaged regions (p<0.001 for all locations: finger, hand, forearm, and nailbed). These findings suggest that OCTA is a valuable tool for detecting and quantifying vascular abnormalities in SSc skin, potentially offering new insights for disease management.
Zebrafish are a widely used developmental model because of their transparent embryos and external development. These distinctive characteristics provide valuable insights into embryonic development. Optical coherence tomography (OCT) offers label-free structural imaging and has emerged as a preferred tool for embryonic imaging. On the other hand, light sheet fluorescence microscopy (LSFM) enables time-lapse molecular imaging of multi-hour to multi-day developmental processes due to its low phototoxicity and photobleaching compared to traditional confocal fluorescence microscopy. We developed a multimodal imaging system to obtain concurrent structural and molecular information by combining OCT and LSFM for embryonic imaging. A Michelson-type swept-source OCT system with a central wavelength of 1050 nm, the bandwidth of 100 nm, and sweep rate of 100 kHz captured the structural information with a lateral resolution of ~15 μm and an axial resolution of ~7 μm. The LSFM system captured the molecular information with a transverse resolution of ~2.1 μm and an axial resolution of ~13 μm. The optically co-aligned OCT and LSFM beams were scanned through the same scan head for trivial co-registration of the multimodal images. We imaged 1-5 μm green fluorescence microbeads to show the capability of this system. We then conducted imaging of zebrafish vasculature development with a transgenic line, Tg(kdrl:EGFP), where the erythroblasts express GFP. The results show that the multimodal system enables us to provide co-registered zebrafish structural and functional imaging.
Systemic sclerosis (SSc) is a rare, chronic autoimmune disease characterized by fibrosis and vascular abnormalities in multiple organs, including skin, lungs, and gastrointestinal tract. Early and accurate diagnosis of SSc is essential for timely intervention and improved patient outcomes. More than 90% of patients with SSc have fibrosis in the skin that manifests as mechanical changes in the skin. The modified Rodnan Skin Score (mRSS) is the gold standard for assessing skin involvement in SSc but has high inter-observer variability. The other widely used method is ultrasound imaging. Ultrasound elastography is emerging as a useful technique for assessing SSc, but its accuracy is still under investigation. Even though the results from these studies provide a quantitative assessment of SSc, ultrasound elastography has many limits, such as the lack of high-resolution performance to detect SSc-affected skin structural and elastic characteristics simultaneously. Optical coherence elastography (OCE) is an established technique to assess the mechanical properties of tissues noninvasively and quantitatively with high resolution. We measured the mechanical properties of skin in 45 patients (36 SSc and 9 matched controls) using a home-built swept source OCE setup that uses air-coupled acoustic radiation force for tissue excitation. The measurements were performed at three locations on each arm: the proximal phalanx of the third finger, the second intermetacarpal space, and the dorsal forearm midline. The OCE results were compared to mRSS and clinical ultrasound elastography, which a trained physician performed. Our results show that OCE outperformed ultrasound elastography with a higher correlation with mRSS.
Systemic Sclerosis (SSc) is a chronic disease of autoimmune etiology that causes vasomotor disturbance, fibrosis, and atrophy of skin, underlying tissue, muscles, and internal organs. It manifests in about 30 people per million every year, and there are an estimated three million cases worldwide. Currently, SSc is assessed using the modified Rodnan Skin Score (mRSS), which is a manual palpation test on 17 sites that requires extensive physician training and experience for accurate assessments. Unfortunately, mRSS has very high inter-observer variability and is subjective. Optical coherence elastography (OCE) is a well-established technique for assessing the mechanical properties of tissues with sub-millimeter spatial resolution. In this work, OCE was used to non-invasively assess the mechanical properties of mouse skin in vivo. OCE measurements were performed on 3 groups of mice, (1) control group that was injected with PBS, (2) SSc group that was injected with bleomycin (BLM) to induce SSc, and (3) treatment group that was first injected with BLM and then injected with imatinib, which is postulated to reduce disease in SSc. The wave speed in BLM-SSc skin was significantly higher than that of normal skin (p<0.05). The wave speed in murine skin in the treatment group was slightly lower as compared to the BLM-SSc skin, but the difference was not significant. These results demonstrate the ability of OCE to monitor SSc disease and treatment response and support further evaluation of this platform in monitoring SSc in the clinic.
Fluorescent two-photon selective-plane illumination microscopy (2P-SPIM) enables deep imaging of cellular information such as proliferation, type identification, and signaling using fluorescence. Optical coherence tomography (OCT) can capture complementary structural information based on intrinsic optical scattering. We developed a specialized multimodal high-resolution embryonic imaging system combining the benefits of OCT with 2P-SPIM. The OCT and 2P-SPIM beams were optically co-aligned and scanned using the same scanners and the same objective lens. The resulting light sheet thickness was ~13 µm with a transverse resolution of ~2.1 µm. The OCT system was based on a 1050 nm centered swept source laser with a bandwidth of ~100 nm and a sweep rate of 100 kHz. The OCT system utilized a Michelson-style interferometer and had a lateral resolution of ~15 µm and an axial resolution of ~7 µm. The capabilities of the multimodal imaging system were demonstrated using images of fluorescent microbeads and a fluorescently tagged mouse embryo at gestational day 9.5. Due to the co-alignment of the OCT and 2P-SPIM systems, image registration was simple and allowed for high-throughput multimodal imaging without the use of sophisticated registration methods.
Optical coherence tomography (OCT) and light sheet fluorescence microscopy (LSFM) are well-established imaging techniques preferred in developmental biology, e.g., embryonic imaging. However, each technique has its own drawbacks, such as resolution and molecular specificity with OCT and field-of-view (FOV) and speed with LSFM. To overcome these limitations for small animal embryo imaging, we have developed a co-aligned multimodal imaging system combining OCT and LSFM. The OCT probe and LSFM excitation beams were combined and scanned with a galvanometer-mounted mirror through the same objective lens. The light sheet thickness was ~13 μm. The LSFM collection arm consisted of a 0.8 numerical aperture water immersion objective, tube lens, and CCD camera, resulting in a transverse resolution of ~2.1 μm. The OCT system was based on a 100 kHz swept-source laser with a central wavelength of 1050 nm and had a lateral resolution of ~15 µm and an axial resolution of ~7 μm. Images of fluorescent microbeads and a fluorescent-tagged mouse embryo at gestational day 9.5 showed the capabilities of the multimodal imaging system. Since the OCT system and LSFM system were co-aligned, image registration was straightforward and enabled high-throughput multimodal imaging without the need for complex registration techniques.
We developed a high-resolution multimodal system for mouse embryonic imaging that combines Optical Coherence Tomography (OCT) and Light Sheet Fluorescence Microscopy (LSFM). LSFM Illumination is restricted to fluorophores in the focal volume, and collecting the light using a microscope objective increases the signal from that plane and reduces the noise coming from outside of the plane. Colinearly aligning this modality with the OCT beam allows one to acquire the structural information from the same plane that is illuminated by the LSFM beam. A 3D image of 9.5 day mouse embryo was captured using this multimodal system.
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