Adaptive optics SLO/OCT for 3D imaging of human photoreceptors in vivo

We present a new instrument that is capable of imaging human photoreceptors in three dimensions. To achieve high lateral resolution, the system incorporates an adaptive optics system. The high axial resolution is achieved through the implementation of optical coherence tomography (OCT). The instrument records simultaneously both, scanning laser ophthalmoscope (SLO) and OCT en-face images, with a pixel to pixel correspondence. The information provided by the SLO is used to correct for transverse eye motion in post-processing. In order to correct for axial eye motion, the instrument is equipped with a high speed axial eye tracker. In vivo images of foveal cones as well as images recorded at an eccentricity from the fovea showing cones and rods are presented.OCIS codes: (170.3890) Medical optics instrumentation, (110.1080) Active or adaptive optics, (170.4470) Ophthalmology, (330.5310) Vision - photoreceptors, (110.4500) Optical coherence tomography     

Noninvasive OCT angiography-based blood attenuation measurements correlate with blood glucose level in the mouse retina

In this study, we investigated the correlation of the blood optical attenuation coefficient (OAC) and the blood glucose concentration (BGC). The blood OAC was measured in mouse retina in vivo by analyzing the depth attenuation of backscattered light under the guidance of OCT angiography (OCTA) vascular mapping, and then its correlation to the BGC was further investigated. The optical attenuation of the blood components presented a more reliable correlation to BGC than that of the background tissues. The arteries and veins presented a blood OAC change of ∼0.05-0.07 mm^-1 per 10 mg/dl and a significant (P < 0.001) elevation of blood OAC in diabetic mice was observed. Furthermore, different kinds of vessels also presented different performances. The veins had a higher correlation coefficient (R=0.86) between the measured blood OAC and BGC than that of the arteries (R=0.73). Besides, the blood OAC changes of the specific vessels occur without any obvious change in the vascular morphology in the retina. The blood OAC-BGC correlation suggests a concept of non-invasive OCTA-based glucometry, allowing a fast assessment of the blood glucose of specific vessels with superior motion immunity. A direct glucometry of the retina would be helpful for accurately monitoring the progression of diabetic retinopathy.     

Photon efficient double-helix PSF microscopy with application to 3D photo-activation localization imaging

We present a double-helix point spread function (DH-PSF) based three-dimensional (3D) microscope with efficient photon collection using a phase mask fabricated by gray-level lithography. The system using the phase mask more than doubles the efficiency of current liquid crystal spatial light modulator implementations. We demonstrate the phase mask DH-PSF microscope for 3D photo-activation localization microscopy (PM-DH-PALM) over an extended axial range.     

Plastic embedding for precise imaging of large-scale biological tissues labeled with multiple fluorescent dyes and proteins

Resin embedding of multi-color labeled whole organs is the primary step to preserve structural information for visualization of fine structures in three dimensions. It is essential to study the morphological characteristics, spatial and positional relationships of the millions of neurons, and the intricate network of blood vessels with fluorescent labels in the brain. However, the current resin embedding method is inadequate because of incompatibilities with fluorescent dyes, making it difficult to reconstruct a variety of structures for the interpretation of their complex spatial relationships. We modified the resin embedding method for large biological tissues labeled with multiple fluorescent dyes and proteins through different labeling strategies. With TrueBlack as the background fluorescence inhibitor in the glycol methacrylate (GMA) embedding, we referred to the method as GMA-T (Glycol methacrylate with TB). In the GMA-T embedded mouse brains, structures labeled with fluorescent proteins and dyes were visualized in millimeter-scale networks with sub-cellular resolution, allowing quantitative analysis of different anatomical structures in the same brain, including neurons and blood vessels. In combination with high-resolution whole-brain imaging, it is possible to obtain a variety of fluorescence labeled structures in just a few days. We quantified the distribution and morphology of the tdTomato-labeled vasoactive intestinal polypeptide (VIP) neurons and the BSA-FITC labeled blood vessels in the same brain. These results demonstrated that VIP neurons and blood vessels have their own unique distribution patterns and morphological characteristics among cortical regions and different layers in cerebral cortex, and there was no significant correlation between VIP neurons and vessels. This approach provides a novel approach to study the interaction among different anatomical structures within large-volume biological samples labeled with multiple fluorescent dyes and proteins, which helps elucidating the complex anatomical characteristics of biological organs.     

Identification and quantification of fibrotic areas in the human retina using polarization-sensitive OCT

Subretinal fibrosis is one of the most prevalent causes of blindness in the elderly population, but a true gold standard to objectively diagnose fibrosis is still lacking. Since fibrotic tissue is birefringent, it can be detected by polarization-sensitive optical coherence tomography (PS-OCT). We present a new algorithm to automatically detect, segment, and quantify fibrotic lesions within 3D data sets recorded by PS-OCT. The algorithm first compensates for the birefringence of anterior ocular tissues and then uses the uniformity of the birefringent optic axis as an indicator to identify fibrotic tissue, which is then segmented and quantified. The algorithm was applied to 3D volumes recorded in 57 eyes of 57 patients with neovascular age-related macular degeneration using a spectral domain PS-OCT system. The results of fibrosis detection were compared to the clinical diagnosis based on color fundus photography (CFP), and the precision of fibrotic area measurement was assessed by three repeated measurements in a sub-set of 15 eyes. The average standard deviation of the fibrotic area obtained in eyes with a lesion area > 0.7 mm² was 15%. Fibrosis detection by CFP and PS-OCT agreed in 48 cases, discrepancies were only observed in cases of lesion area < 0.7 mm². These remaining discrepancies are discussed, and a new method to treat ambiguous cases is presented.     

荧光定量聚合酶链反应法检测光感受器细胞凋亡大鼠视网膜中半胱氨酸蛋白酶3 mRNA的表达

目的:检测半胱氨酸蛋白酶3mRNA在光感受器凋亡大鼠视网膜中的表达,探讨其在光感受器细胞凋亡的发生、发展中的意义。方法:实验于2004-03/12在中山眼科中心眼科学教育部重点实验室完成。选择雌性SD大鼠36只,随机数字表法分为正常对照组6只,N-甲基-N-亚硝脲组30只,后者又分为造模后12h,1,2,3,5d5个时相点,每个时相点6只。正常对照组大鼠经腹腔注射生理盐水;N-甲基-N-亚硝脲组大鼠按体质量一次性腹腔注射N-甲基-N-亚硝脲40mg/kg,分别于造模后12h,1,2,3,5d处死动物,摘除右眼球,立即分离视网膜,提取总RNA,用荧光定量聚合酶链反应法检测视网膜中半胱氨酸蛋白酶3mRNA的含量。结果:纳入动物36只,均进入结果分析。正常对照组、N-甲基-N-亚硝脲组造模后12h,1,2,3,5d5个时相点视网膜中半胱氨酸蛋白酶3mRNA表达量分别为1.52×105,18.35×105,25.14×105,29.25×105,13.72×105,12.24×105。N-甲基-N-亚硝脲腹腔注射后12h视网膜中半胱氨酸蛋白酶3mRNA表达量上升,第2天时最高,达正常对照组表达量的19倍,此后渐下降,第5天表达量仍为正常对照组的8倍。结论:视网膜中半胱氨酸蛋白酶3mRNA表达与光感受器细胞的存亡有关,可作为光感受器细胞凋亡发生、发展的预测指标之一。     

3D自由呼吸与屏气扫描磁共振心肌延迟成像技术对比

目的比较对心肌梗死患者采用3D自由呼吸(3DFB)采集与单次屏气(BH)采集反转恢复梯度回波序列心肌延迟成像的图像质量。方法对20例心肌梗死患者分别行呼吸导航回波触发、3DFH反转恢复梯度回波扫描和3DBH反转恢复梯度回波扫描,以"优=4、良=3、一般=2、差=1"四个级别评判心肌图像质量,计算两组左室心肌面积均值、梗死心肌面积均值、延迟强化心肌信噪比和对比噪声比,测量心肌梗死透壁程度。采用SPSS12.0软件对两组结果进行t检验分析。用相关回归法评价两种采集方法在计算左室活性心肌百分比方面的一致性。结果FB组图像质量评分高于BH组。两组图像信噪比和对比噪声比差异有统计学意义,左室心肌面积均值、梗死心肌面积均值差异无统计学意义,两组透壁厚度分别为(5.13±2.74)mm和(6.81±3.15)mm(P<0.05)。结论呼吸导航回波触发反转恢复梯度回波扫描可有效应用于3D磁共振心肌延迟成像,与单次屏气采集3D反转恢复梯度回波序列相比,可显著提高图像质量和分辨率,对坏死心肌能作出更为精确的量化评价。     

荧光纳米粒子在小鼠胃癌皮下瘤模型近红外成像中的实验研究

目的 合成特异性靶向胃癌纳米粒子,借助自主搭建的近红外荧光腹腔镜,探讨该纳米粒子在胃癌精确诊断及完整切除中的价值。方法 将吲哚菁绿（ICG）通过自主装方式包覆于双亲性聚合物二硬脂酰基磷脂酰乙醇胺-聚乙二醇-马来酰亚胺（DSPE-PEG-Mal）,同时,用精氨酸-甘氨酸-天冬氨酸（RGD）多肽修饰,制备靶向纳米粒子DSPE-PEG-RGD@ICG。通过核磁谱图验证产物,吸收光谱检测光学性质。建立小鼠胃癌皮下瘤模型,在自主搭建的近红外荧光腹腔镜下检测及切除肿瘤。结果 核磁谱图验证产物与预期相符,并表现出与游离ICG相似的吸收光谱。在体内实验中,DSPE-PEG-RGD@ICG可特异性靶向胃癌,指导胃癌的完整切除。静脉给药4 h后,荧光腹腔镜可清楚显示肿瘤位置。小鼠胃癌皮下瘤检出率为91.8%,检出肿瘤的最小直径为4.9 mm。结论 成功制备了特异性靶向胃癌的纳米粒子DSPE-PEG-RGD@ICG,其光学特性与ICG相似,在近红外荧光腹腔镜引导下,可有效地协助外科医师对小鼠胃癌皮下瘤进行定位及完整切除。     

荧光定量聚合酶链反应法检测光感受器细胞凋亡大鼠视网膜中半胱氨酸蛋白酶3mRNA的表达

目的：检测半胱氨酸蛋白酶3mRNA在光感受器凋亡大鼠视网膜中的表达，探讨其在光感受器细胞凋亡的发生、发展中的意义。方法：实验于2004—03／12在中山眼科中心眼科学教育部重点实验室完成。选择雌性SD大鼠36只，随机数字表法分为正常对照组6只，N-甲基-N-亚硝脲组30只，后者又分为造模后12h，1，2，3，5d5个时相点．每个时相点6只。正常对照组大鼠经腹腔注射生理盐水；N-甲基-N-亚硝脲组大鼠按体质量一次性腹腔注射N-甲基-N-亚硝脲40mg／kg，分别于造模后12h，1，2，3，5d处死动物，摘除右眼球，立即分离视网膜，提取总RNA，用荧光定量聚合酶链反应法检测视网膜中半胱氨酸蛋白酶3mRNA的含量。结果：纳入动物36只，均进入结果分析。正常对照组、N-甲基-N-亚硝脲组造模后12h，1，2，3，5d5个时相点视网膜中半胱氨酸蛋白酶3mRNA表达量分别为1．52&;#215;10^5，18．35&;#215;10^5，25．14&;#215;10^5，29．25&;#215;10^5，13．72&;#215;10^5．12．24&;#215;10^5。N-甲基-N-亚硝脲腹腔注射后12h视网膜中半胱氨酸蛋白酶3mRNA表达量上升，第2天时最高，达正常对照组表达量的19倍，此后渐下降，第5天表达量仍为正常对照组的8倍。结论：视网膜中半胱氨酸蛋白酶3mRNA表达与光感受器细胞的存亡有关，可作为光感受器细胞凋亡发生、发展的预测指标之一。     

An in vivo subject-specific 3D functional knee joint model using combined MR imaging

Purpose

We aim to quantitatively characterise the knee joint function in vivo under body-weight-bearing conditions via subject-specific models extracted from magnetic resonance (MR) data, in order to better understand the knee joint kinematic function in 3D.

Methods

Six healthy volunteers without any record of knee abnormality were scanned using a combined MR imaging strategy to record quasi-squatting motion and 3D knee anatomy. After a semi-automatic segmentation to delineate tibio-femoral articulation components, motion data were mapped to the anatomical data using a bi-rigid registration in order to achieve six degrees of freedom. The individual knee joint function was characterised by analysing the tibio-femoral articulation contact mechanism based on the reconstructed models in 3D and MR images in 2D. Contact points were extracted and their trajectory was plotted on the tibia plateau.

Results

The 3D models clearly show the relative rotation and gliding between tibia and femur during global flexion. Within the measured flexion arc, the contact points move less between 30 $^{\circ }$ and 100 $^{\circ }$ on both tibial plateaux as compared to that on the rest of the flexion arc. Four out of the six volunteers showed a global pattern of less moving extent of contact points on the medial tibial plateau than on the lateral tibial plateau in both 3D and 2D.

Conclusion

The proposed subject-specific model is able to characterise knee joint kinematic function. It provides a way to describe knee joint surface kinematics quantitatively, which may help to better understand the knee function and joint derangements.     

Vector of motion measurements in the living cochlea using a 3D OCT vibrometry system

Optical coherence tomography (OCT) has become an important tool for measuring the vibratory response of the living cochlea. It stands alone in its capacity to measure the intricate motion of the hearing organ through the surrounding otic capsule bone. Nevertheless, as an extension of phase-sensitive OCT, it is only capable of measuring motion along the optical axis. Hence, measurements are 1-D. To overcome this limitation and provide a measure of the 3-D vector of motion in the cochlea, we developed an OCT system with three sample arms in a single interferometer. Taking advantage of the long coherence length of our swept laser, we depth (frequency) encode the three channels. An algorithm to depth decode and coregister the three channels is followed by a coordinate transformation that takes the vibrational data from the experimental coordinate system to Cartesian or spherical polar coordinates. The system was validated using a piezo as a known vibrating element that could be positioned at various angles. The angular measurement on the piezo was shown to have an RMSE of ≤ 0.30° (5.2 mrad) with a standard deviation of the amplitude of ≤ 120 pm. Finally, we demonstrate the system for in vivo imaging by measuring the vector of motion over a volume image in the apex of the mouse cochlea.     

3D动脉自旋标记、弥散张量成像联合常规MRI鉴别诊断颅内血管外皮细胞瘤与血管瘤型脑膜瘤

目的 观察3D动脉自旋标记（3D ASL）、弥散张量成像（DTI）联合常规MRI鉴别诊断颅内血管外皮细胞瘤（HPC）与血管瘤型脑膜瘤（AM）的价值。方法 回顾性分析经手术病理证实的15例HPC（HPC组）与11例AM（AM组），分析2组临床表现、MRI（包括平扫、增强、DTI、3D ASL）征象及功能成像参数差异。结果 HPC组平均脑血流量（CBF）及各向异性分数（FA）均低于AM组（t=-8.99，P<0.01；t=-3.66，P<0.01），表观弥散系数（ADC）高于AM组（t=2.61，P=0.02）；HPC组高灌注区和低灌注区CBF均低于AM组（t=-15.13，P<0.01；t=-8.30，P<0.01）；HPC组平均发病年龄低于AM组（t=-2.39，P=0.02），性别差异无统计学意义（χ²=1.69，P=0.19）；相比AM组，HPC组更易出现分叶征（χ²=9.09，P<0.01）及囊变坏死（χ²=9.38，P<0.01），HPC病灶T1WI以等高信号为主（χ²=27.78，P<0.01）、T2WI以等低信号为主（χ²=16.33，P<0.01），瘤周水肿程度轻（χ²=19.25，P<0.01），血管流空影更常见且更粗大（χ²=9.02，P=0.01），多与硬脑膜多为窄基底相连（χ²=28.54，P<0.01）而脑膜尾征少见（χ²=25.00，P<0.01）。结论 HPC与AM的MRI征象及各功能成像参数值存在一定差异，可为鉴别诊断提供参考。     

Nitrogen-doped fluorescent carbon dots used for the imaging and tracing of different cancer cells

Here, we report the synthesis of nitrogen-doped fluorescent carbon (C) dots using a one-pot hydrothermal method. Transmission electron microscopy results reveal that the particle size of the nitrogen-doped C-dots is very small, with an average diameter of 4.6 nm. After being kept in water for 10 days, the nitrogen-doped C-dots can still dissolve well in the water, showing good stability and compatibility in aqueous solution. The fluorescence spectra show that the nitrogen-doped C-dots exhibit emission-tunable color from blue to green upon excitation from 230 to 520 nm. Cell tests show that the C-dots are low in cytotoxicity and can be used for imaging, detecting and tracing between hepatoma and HeLa cells, because hepatoma and HeLa cells show different sensitivity to different fluorescent colors pumped at different excitation wavelengths.

Here, we report the synthesis of nitrogen-doped fluorescent carbon (C) dots using a one-pot hydrothermal method.     

Measurement of pulsatile total blood flow in the human and rat retina with ultrahigh speed spectral/Fourier domain OCT

We present an approach to measure pulsatile total retinal arterial blood flow in humans and rats using ultrahigh speed Doppler OCT. The axial blood velocity is measured in an en face plane by raster scanning and the flow is calculated by integrating over the vessel area, without the need to measure the Doppler angle. By measuring flow at the central retinal artery, the scan area can be very small. Combined with ultrahigh speed, this approach enables high volume acquisition rates necessary for pulsatile total flow measurement without modification in the OCT system optics. A spectral domain OCT system at 840nm with an axial scan rate of 244kHz was used for this study. At 244kHz the nominal axial velocity range that could be measured without phase wrapping was ±37.7mm/s. By repeatedly scanning a small area centered at the central retinal artery with high volume acquisition rates, pulsatile flow characteristics, such as systolic, diastolic, and mean total flow values, were measured. Real-time Doppler C-scan preview is proposed as a guidance tool to enable quick and easy alignment necessary for large scale studies. Data processing for flow calculation can be entirely automatic using this approach because of the simple and robust algorithm. Due to the rapid volume acquisition rate and the fact that the measurement is independent of Doppler angle, this approach is inherently less sensitive to involuntary eye motion. This method should be useful for investigation of small animal models of ocular diseases as well as total blood flow measurements in human patients in the clinic.     

Structure modulation on fluorescent probes for biothiols and the reversible imaging of glutathione in living cells

The detection of small molecular biothiols (cysteine, homocysteine and glutathione) is of great importance, as they involve in a series of physiological and pathological processes and are associated with many diseases. To realize the real-time monitoring of a specific biothiol, a rapid and reversible probe is required. Therefore, three probes, namely, o-MNPy, m-MNPy and p-MNPy, with pyridine substituted α, β-unsaturated ketone as the recognition site, were reported here, and the reactivity of the recognition site was finely tuned by the connection mode of the pyridine unit. To single out the optimal one, the response performances of three probes toward each biothiol were systemically studied, taking the differences of the intracellular contents of three biothiols into account during the evaluation. Biothiols reacted with the probes through Michael addition, and results showed that the slight structural variations could affect the performances of the probes obviously. p-MNPy with the pyridine unit connected to the recognition site through the para-position of the nitrogen atom, revealed the best sensing ability among the three probes. It demonstrated rapid response, good selectivity and sensitivity, excellent pH adaptability to Cys and GSH, and displayed reversible detection toward GSH. Finally, p-MNPy was successfully applied to track the GSH fluctuations under the oxidative stress stimulated by H₂O₂ in living cells.

A reversible fluorescent probe for GSH was obtained through structure modulation, by which the intracellular GSH fluctuation was imaged.     

Paraumbilical vein in the cirrhotic patient: imaging with 3D CT angiography

The paraumbilical vein is a common venous collateral in patients with cirrhosis and portal hypertension. This pictorial essay demonstrates the use of computed tomographic angiography with three-dimensional volume rendering techniques to visualize the paraumbilical vein and its relationship to abdominal wall collaterals. The unique anatomy and embryology of this vessel is also discussed. Received: 25 March 1997/Accepted: 30 April 1997     

胎儿并腿畸形下肢骨骼异常三维超声骨骼成像技术、MRI与3D-HCT诊断研究

目的：探讨胎儿三维超声骨骼成像技术、胎儿磁共振成像（MRI）及引产后三维螺旋CT（3-HCT）在并腿畸形胎儿下肢骨骼异常诊断中的临床应用价值。方法应用胎儿超声骨骼成像技术与磁共振成像对7例并腿畸形胎儿进行分型诊断,并与引产后胎儿三维螺旋CT及病理检查结果对照,分析三维超声骨骼成像技术、MRI、三维螺旋CT在并腿畸形胎儿下肢骨骼异常产前与产后诊断中的应用结果。结果与终止妊娠后三维螺旋CT及病理检查结果对照,产前三维超声骨骼成像均正确显示7例并腿畸形胎儿（6例单胎,1例联体双胎）下肢股骨异常、胫腓骨异常及单足6例,无足1例。其中绒毛和（或）脐血染色体检查（5例）与生后病理检查提示男性3例,女性4例。根据Stocker并腿畸形分型标准,产前与引产胎儿标本影像学检查诊断并腿畸形Ⅲ型1例;Ⅳ型2例;Ⅴ型3例;Ⅵ型1例。7例胎儿MRI正确显示股骨异常4例、胫腓骨异常1例,足异常均未显示。结论产前三维超声骨骼成像技术与产后三维螺旋CT骨骼成像技术均为胎儿骨骼异常影像诊断新技术,可清晰显示胎儿骨骼声像,对下肢畸形胎儿正确诊断有重要临床应用价值。     

Motility imaging via optical coherence phase microscopy enables label‐free monitoring of tissue growth and viability in 3D tissue‐engineering scaffolds

As the field of tissue engineering continues to progress, there is a deep need for non‐invasive, label‐free imaging technologies that can monitor tissue growth and health within thick three‐dimensional (3D) constructs. Amongst the many imaging modalities under investigation, optical coherence tomography (OCT) has emerged as a promising tool, enabling non‐destructive in situ characterization of scaffolds and engineered tissues. However, the lack of optical contrast between cells and scaffold materials using this technique remains a challenge. In this communication, we show that mapping the optical phase fluctuations resulting from cellular viability and motility allows for the distinction of live cells from their surrounding scaffold environment. Motility imaging was performed via a common‐path optical coherence phase microscope (OCPM), an OCT modality that has been shown to be sensitive to nanometer‐level fluctuations. More specifically, we examined the development of human adipose‐derived stem cells and/or murine pre‐osteoblasts within two distinct scaffold systems, commercially available alginate sponges and custom‐microfabricated poly(d , l ‐lactic‐co‐glycolic acid) fibrous scaffolds. Cellular motility is demonstrated as an endogenous source of contrast for OCPM, enabling real‐time, label‐free monitoring of 3D engineered tissue development. Copyright © 2013 John Wiley & Sons, Ltd.