Improved interchangeability with different corneal specular microscopes for quantitative endothelial cell analysis

Introduction: During our clinical practice and research, we encountered an interchangeability problem when using the SP-2000P and SP-3000P TopCon corneal specular microscopes (CSMs) (TopCon Medical Systems, Tokyo, Japan) regarding the endothelial cell count (ECC). We describe a method to improve interchangeability between these CSMs. Methods: Five consecutive good-quality endothelial cell photographs were obtained in 22 eyes of 11 subjects. An ECC comparison between the two CSMs was performed after (I) gauging and calibration by the manufacturer, (II) adjustment of the magnification, (III) correction after external horizontal and vertical calibration. Results: There was a statistically significant difference between the ECC of the SP-2000P and SP-3000P at the start. The SP-2000P counted an average of 500 cells/mm2 more than the SP-3000P (p=0.00). After correction for magnification and determining a correction factor based on external calibration, the difference between the ECC of the SP-2000P and the SP-3000P was then found to be 0.4 cells/mm2 and was not statistically significant (p=0.98). Discussion: We propose a method for improving interchangeability, which involves checking magnification settings, re-checking magnification calibration with external calibration devices, and then calculating correction factors. This method can be applied to various specular or confocal microscopes and their associated endothelial cell analysis software packages to be able to keep performing precise endothelial cell counts and to enable comparison of ECCs when a CSM needs to be replaced or when results from different microscopes need to be compared

Neutrophil-Mediated Experimental Metastasis Is Enhanced by VEGFR Inhibition in a Zebrafish Xenograft Model

Inhibition of VEGF signalling effectively suppresses localized tumour growth but accelerates tumour invasiveness and micrometastasis by unknown mechanisms. To study the dynamic and reciprocal interactions between tumour cells and their microenvironment during these processes, we established a xenograft model by injecting tumour cells into the blood circulation of transparent zebrafish embryos. This reproducibly results in rapid simultaneous formation of a localized tumour and experimental micrometastasis, allowing time-resolved imaging of both processes at single-cell resolution within 1 week. The tumour vasculature was initiated de novo by remodelling of primitive endothelial cells into a functional network. Roles of myeloid cells in critical tumourigenesis steps such as vascularization and invasion were revealed by genetic and pharmaceutical approaches. We discovered that the physiological migration of neutrophils controlled tumour invasion by conditioning the collagen matrix and forming the metastatic niche, as detected by two-photon confocal microscopy and second harmonic generation. Administration of VEGFR inhibitors blocked tumour vascularization and a localized tumour growth but enhanced migration of neutrophils, which in turn promoted tumour invasion and formation of micrometastasis. This demonstrates the in vivo cooperation between VEGF signalling and myeloid cells in metastasis and provides a new mechanism underlying the recent findings that VEGFR targeting can promote tumour invasiveness

Silent volumetric multi-contrast 7 Tesla MRI of ocular tumors using Zero Echo Time imaging.

Magnetic Resonance Imaging (MRI) has become a valuable imaging modality in ophthalmology, especially for the diagnosis and treatment planning of patients with uveal melanoma, the most common primary intra-ocular tumor. We aim to develop and evaluate the value of silent Zero Echo Time (ZTE) MRI to image patients with ocular tumors at 7Tesla. Therefore, ZTE and different types of magnetization-prepared ZTE (FLAIR, SPIR, T2 and Saturation recovery), have been developed. After an initial validation with 7 healthy subjects, nine patients with an eye tumor have been evaluated. The ZTE scans were compared to their Cartesian equivalent in terms of contrast, motion-sensitivity, diagnostic quality and patient comfort. All volunteers and especially the patients reported a more comfortable experience during the ZTE scans, which had at least a 10 dB lower sound pressure. The image contrast in the native ZTE was poor, but in the different magnetization-prepared ZTE, the eye lens, cornea and retina were clearly discriminated. Overall the T2-prepared scan yielded the best contrast, especially between tumor and healthy tissue, and proved to be robust against eye motion. Although the intrinsic 3D nature of the ZTE-technique provides an accurate analysis of the tumor morphology, the quality of the ZTE-images is lower than their Cartesian equivalent. In conclusion, the quality of magnetization-prepared ZTE images is sufficient to assess the 3D tumor morphology, but insufficient for more detailed evaluations. As such this technique can be an option for patients who cannot comply with the sound-levels of Cartesian scans, but for other patients the conventional Cartesian scans offer a better image quality

Equilibrium shape of the aqueous humor-vitreous substitute interface in vitrectomized eyes

Purpose: To predict the shape of the interface between aqueous humor and a gas or silicone oil (SO) tamponade in vitrectomized eyes. To quantify the tamponated retinal surface for various eye shapes, from emmetropic to highly myopic eyes. Methods: We use a mathematical model to determine the equilibrium shape of the interface between the two fluids. The model is based on the volume of fluids (VOF) method. The governing equations are solved numerically using the free so ware OpenFOAM. We apply the model to the case of idealized, yet realistic, geometries of emmetropic and myopic eyes, as well as to the real geometry of the vitreous chamber reconstructed from magnetic resonance imaging (MRI) images. Results: The numerical model allows us to compute the equilibrium shape of the interface between the aqueous humor and the tamponade fluid. From this we can compute the portion of the retinal surface that is effectively tamponated by the fluid. We compare the tamponating ability of gases and SOs. We also compare the tamponating effect in emmetropic and myopic eyes by computing both tamponated area and angular coverage. Conclusion: The numerical results show that gases have better tamponating properties than SOs. We also show that, in the case of SO, for a given filling ratio the percentage of tamponated retinal surface area is smaller in myopic eyes. The method is valuable for clinical purposes, especially in patients with pathological eye shapes, to predict the area of the retina that will be tamponated for a given amount of injected fluid

MRI of Uveal Melanoma

Uveal Melanoma (UM) is the most common primary malignant ocular tumor. The high soft tissue contrast and spatial resolution, and the possibility of generating 3D volumetric and functional images, make Magnetic Resonance Imaging (MRI) a valuable diagnostic imaging technique in UM. Current clinical MRI protocols, however, are not optimized for UM and therefore lack the quality for accurate assessments. We therefore developed a dedicated protocol at a 3 Tesla MRI, using an eye coil, consisting of multi-slice 2D sequences, different isotropic sequences and diffusion and perfusion-weighted images. This protocol was prospectively evaluated in 9 uveal melanoma patients. The multi-slice 2D sequences had the highest in-plane resolution, being the most suited for lesion characterization and local extension evaluation. The isotropic 3D Turbo-Spin Echo (TSE) sequences were the most suitable for accurate geometric measurements of the tumor and are therefore important for therapy planning. Diffusion and perfusion-weighted images aid in differentiating benign from malignant lesions and provide quantitative measures on tumor hemodynamics and cellularity, which have been reported to be effective in predicting and assessing treatment outcome. Overall, this dedicated MRI protocol provides high-quality imaging of UM, which can be used to improve its diagnosis, treatment planning, and follow-up

MR imaging characteristics of uveal melanoma with histopathological validation

Purpose: To evaluate the magnetic resonance imaging (MRI) characteristics of uveal melanoma (UM), to compare them with fundoscopy and ultrasound (US), and to validate them with histopathology. Methods: MR images from 42 UM were compared with US and fundoscopy, and on 14 enucleated cases with histopathology. Results: A significant relationship between the signal intensity on T1 and pigmentation on histopathology was found (p=0.024). T1 hyperintense UM were always moderately or strongly pigmented on histopathology, while T1-hypointense UM were either pigmented or non-pigmented. Mean apparent diffusion coefficient (ADC) of the UM was 1.16 ± 0.26 × 10−3 mm2/s. Two-thirds of the UM had a wash-out and the remaining a plateau perfusion time-intensity curve (TIC). MRI was limited in evaluating the basal diameter of flat tumors. US tends to show larger tumor prominence (0.5mm larger, p=0.008) and largest basal diameter (1.4mm larger, p<0.001). MRI was good in diagnosing ciliary body involvement, extrascleral extension, and optic nerve invasion, but limited on identifying scleral invasion. An increase of tumor prominence was associated with lower ADC values (p=0.030) and favored a wash-out TIC (p=0.028). An increase of tumor ADC correlated with a plateau TIC (p=0.011). Conclusions: The anatomical and functional MRI characteristics of UM were comprehensively assessed. Knowing the MRI characteristics of UM is important in order to confirm the diagnosis and to differentiate UM from other intra-ocular lesions and because it has implications for treatment planning. MRI is a good technique to evaluate UM, being only limited in case of flat tumors or on identifying scleral invasion

Automatic Three-Dimensional Magnetic Resonance-based measurements of tumour prominence and basal diameter for treatment planning of uveal melanoma

Background and Purpose: Three-dimensional (3D) Magnetic Resonance Imaging (MRI) is increasingly used to complement conventional two-dimensional ultrasound in the assessment of tumour dimension measurement of uveal melanoma. However, the lack of definitions of the 3D measurements of these tumour dimensions hinders further adaptation of MRI in ocular radiotherapy planning. In this study, we composed 3D MR-based definitions of tumour prominence and basal diameter and compared them to conventional ultrasound. Materials and methods: Tumours were delineated on 3DT2 and contrast-enhanced 3DT1 (T1gd) MRI for 25 patients. 3D definitions of tumour prominence and diameter were composed and evaluated automatically on the T1gd and T2 contours. Automatic T1gd measurements were compared to manual MRI measurements, to automatic T2 measurements and to manual ultrasound measurements. Results: Prominence measurements were similar for all modalities (median absolute difference 0.3 mm). Automatic T1gd diameter measurements were generally larger than manual MRI, automatic T2 and manual ultrasound measurements (median absolute differences of 0.5, 1.6 and 1.1 mm respectively), mainly due to difficulty defining the axis of the largest diameter. Largest differences between ultrasound and MRI for both prominence and diameter were found in anteriorly located tumours (up to 1.6 and 4.5 mm respectively), for which the tumour extent could not entirely be visualized with ultrasound. Conclusions: The proposed 3D definitions for tumour prominence and diameter agreed well with ultrasound measurements for tumours for which the extent was visible on ultrasound. 3D MRI measurements generally provided larger diameter measurements than ultrasound. In anteriorly located tumours, the MRI measurements were considered more accurate than conventional ultrasound