Tissue sections from aggressive human intraocular (uveal) and metastatic cutaneous melanomas generally lack evidence of significant necrosis and contain patterned networks of interconnected loops of extracellular matrix. The matrix that forms these loops or networks may be solid or hollow. Red blood cells have been detected within the hollow channel components of this patterned matrix histologically, and these vascular channel networks have been detected in human tumors angiographically. Endothelial cells were not identified within these matrix-embedded channels by light microscopy, by transmission electron microscopy, or by using an immunohistochemical panel of endothelial cell markers (Factor VIII-related antigen, Ulex, CD31, CD34, and KDR[Flk-1]). Highly invasive primary and metastatic human melanoma cells formed patterned solid and hollow matrix channels (seen in tissue sections of aggressive primary and metastatic human melanomas) in three-dimensional cultures containing Matrigel or dilute Type I collagen, without endothelial cells or fibroblasts. These tumor cell-generated patterned channels conducted dye, highlighting looping patterns visualized angiographically in human tumors. Neither normal melanocytes nor poorly invasive melanoma cells generated these patterned channels in vitro under identical culture conditions, even after the addition of conditioned medium from metastatic pattern-forming melanoma cells, soluble growth factors, or regimes of hypoxia. Highly invasive and metastatic human melanoma cells, but not poorly invasive melanoma cells, contracted and remodeled floating hydrated gels, providing a biomechanical explanation for the generation of microvessels in vitro. cDNA microarray analysis of highly invasive versus poorly invasive melanoma tumor cells confirmed a genetic reversion to a pluripotent embryonic-like genotype in the highly aggressive melanoma cells. These observations strongly suggest that aggressive melanoma cells may generate vascular channels that facilitate tumor perfusion independent of tumor angiogenesis.

Vascular Channel Formation by Human Melanoma Cells in Vivo and in Vitro: Vasculogenic Mimicry

Photoacoustic (PA) imaging, also called optoacoustic imaging, is a new biomedical imaging modality based on the use of laser-generated ultrasound that has emerged over the last decade. It is a hybrid modality, combining the high-contrast and spectroscopic-based specificity of optical imaging with the high spatial resolution of ultrasound imaging. In essence, a PA image can be regarded as an ultrasound image in which the contrast depends not on the mechanical and elastic properties of the tissue, but its optical properties, specifically optical absorption. As a consequence, it offers greater specificity than conventional ultrasound imaging with the ability to detect haemoglobin, lipids, water and other light-absorbing chomophores, but with greater penetration depth than purely optical imaging modalities that rely on ballistic photons. As well as visualizing anatomical structures such as the microvasculature, it can also provide functional information in the form of blood oxygenation, blood flow and temperature. All of this can be achieved over a wide range of length scales from micrometres to centimetres with scalable spatial resolution. These attributes lend PA imaging to a wide variety of applications in clinical medicine, preclinical research and basic biology for studying cancer, cardiovascular disease, abnormalities of the microcirculation and other conditions. With the emergence of a variety of truly compelling in vivo images obtained by a number of groups around the world in the last 2–3 years, the technique has come of age and the promise of PA imaging is now beginning to be realized. Recent highlights include the demonstration of whole-body small-animal imaging, the first demonstrations of molecular imaging, the introduction of new microscopy modes and the first steps towards clinical breast imaging being taken as well as a myriad of in vivo preclinical imaging studies. In this article, the underlying physical principles of the technique, its practical implementation, and a range of clinical and preclinical applications are reviewed.

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Biomedical photoacoustic imaging

Objective: To demonstrate optical coherence tomography for high-resolution, noninvasive imaging of the human retina. Optical coherence tomography is a new imaging technique analogous to ultrasound B scan that can provide cross-sectional images of the retina with micrometer-scale resolution. Design: Survey optical coherence tomographic examination of the retina, including the macula and optic nerve head in normal human subjects. Settings Research laboratory. Participants: Convenience sample of normal human subjects. Main Outcome Measures: Correlation of optical coherence retinal tomographs with known normal retinal anatomy. Results: Optical coherence tomographs can discriminate the cross-sectional morphologic features of the fovea and optic disc, the layered structure of the retina, and normal anatomic variations in retinal and retinal nerve fiber layer thicknesses with 10- μm depth resolution. Conclusion: Optical coherence tomography is a potentially useful technique for high depth resolution, cross-sectional examination of the fundus.

Optical Coherence Tomography of the Human Retina

QUITE apart from the academic consideration that vital and medical statistics now form an obligatory part of the education of students seeking the University of London's diploma in public health, the demand for information about the methods of vital and medical statistics is increasing. The most casual reader of the newspapers is now aware that population problems are of serious practical importance and that the publications of the General Register Office cannot be ignored. It is scarcely an exaggeration to say that the recently issued preliminary report on the census of 1931 is one of the most sensational documents which has appeared for years, and that he who reads it intelligently will understand what is meant by saying that civilisation is in the melting pot. An Introduction to Medical Statistics. By Hilda M. Woods William T. Russell. Pp. x + 125. (London: P. S. King and Son, Ltd., 1931.) 7s. 6d.

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An Introduction to Medical Statistics

Human corneal thickness and its impact on intraocular pressure measures: a review and meta-analysis approach.

PURPOSE—To characterize the in vivo epithelial thickness profile in a population of normal eyes. METHODS—An epithelial thickness profile was measured by Artemis 1 (Ultralink LLC) very highfrequency (VHF) digital ultrasound scanning across the central 10-mm diameter of the cornea of 110 eyes of 56 patients who presented for refractive surgery assessment. The average, standard deviation, minimum, maximum, and range of epithelial thickness were calculated for each point in the 10×10mm Cartesian matrix and plotted. Differences between the epithelial thickness at the corneal vertex and peripheral locations at the 3-mm radius were calculated. The location of the thinnest epithelium was found for each eye and averaged. Correlations of corneal vertex epithelial thickness with age, spherical equivalent refraction, and average keratometry were calculated. RESULTS—The mean epithelial thickness at the corneal vertex was 53.4±4.6 μm, with no statistically significant difference between right and left eyes, and no significant differences in age, spherical equivalent refraction, or keratometry. The average epithelial thickness map showed that the corneal epithelium was thicker inferiorly than superiorly (5.9 μm at the 3-mm radius, P<.001) and thicker nasally than temporally (1.3 μm at the 3-mm radius, P< 001). The location of the thinnest epithelium was displaced on average 0.33 mm temporally and 0.90 mm superiorly with reference to the corneal vertex. CONCLUSIONS—Three-dimensional thickness mapping of the corneal epithelium demonstrated that the epithelial thickness is not evenly distributed across the cornea; the epithelium was significantly thicker inferiorly than superiorly and significantly thicker nasally than temporally with a larger inferosuperior difference than nasotemporal difference. The human corneal epithelium has five to seven cell layers and an accepted central thickness of approximately 50 to 52 μm. 1 Bowman’s layer, a dense collagenous layer approximately 8 to 10 μm thickness lies between the epithelium and stroma. The anterior margin of Bowman’s

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Epithelial Thickness in the Normal Cornea: Three-dimensional Display With Very High Frequency Ultrasound

The human corneal stroma represents approximately 90% of the total corneal thickness and has an accepted central thickness of approximately 478 to 500 μm.1-3 Knowledge of the stromal thickness profile is of interest in the area of corneal refractive surgery, as changes in corneal refractive power are achieved through changes in the stromal thickness profile. Measurements of stromal thickness profile pre- and postoperatively allow stromal ablation rate to be quantified and changes in thickness profile to be determined.4,5 Stromal thickness mapping could help further understand corneal biomechanics. Stromal thickness profiles could also be useful in identifying corneal disorders such as keratoconus; corneal thickness profile has previously been suggested for the early diagnosis of keratoconus6 following characterization of corneal thickness progression by Mandell in 1969.7

Different methods have been used to measure stromal thickness: optical coherence tomography,8,9 confocal microscopy,1-3,10 and through focusing confocal microscopy.11,12 All studies measured the average central stromal thickness. Only two studies provided stromal thickness measurements in the peripheral cornea; however, the number of points measured was limited to one point in the temporal cornea.1,2

Very high-frequency (VHF) digital ultrasound is, to date, the only published method measuring the stromal thickness profile in vivo continuously over a 10-mm diameter, measured from the anterior surface of Bowman’s layer to the posterior surface of the endothelium. Very high-frequency digital ultrasound technology has gradually improved both in precision and in area of acquisition. The repeatability of corneal thickness measurements in 10 consecutive examinations of 1 eye using the Artemis I VHF digital ultrasound arc-scanning system (ArcScan Inc, Morrison, Colo) has been shown to be less than 8 μm within the central 8-mm diameter, with a central repeatability of 1.5 μm.13 The repeatability of epithelial thickness measurements has been shown to be less than 1.30 μm within the central 8-mm diameter, with a central repeatability of 0.5 μm.13

We have previously described the use of VHF digital ultrasound to measure stromal thickness before and after LASIK4,13-16 to investigate stromal changes induced by laser refractive surgery and compare the intended ablation depth to the achieved stromal change. We have also used VHF digital ultrasound to examine the stromal layer before and after implantation of intracorneal ring segments and demonstrated anatomical changes that were induced.17

We have previously described the thickness profile of the corneal epithelium in a population of normal eyes with no ocular pathology other than refractive error.18 The purpose of this study was to characterize the thickness profile of the corneal stroma in the same population.

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Stromal Thickness in the Normal Cornea: Three-dimensional Display With Artemis Very High-frequency Digital Ultrasound

Keratoconus is a non-inflammatory corneal disease characterized by a cone-like–shaped protrusion of the corneal surface. It is associated with progressive corneal thinning, steepening of corneal curvature, and irregular astigmatism, resulting in mild to marked impaired quality of vision. It is a bilateral but often asymmetric disorder.

Total corneal thickness changes associated with keratoconus are due to changes in both stromal thickness and epithelial thickness. Although central total corneal thickness and total corneal thickness profile have been widely reported, only a limited number of studies have reported central stromal or epithelial thickness, and there are no published studies showing pachymetric maps of the stroma or epithelium.

Central corneal thinning in keratoconus has been reported using different methods such as ultrasound pachymetry,1–6 Orbscan (Bausch & Lomb, Salt Lake City, Utah),2,5–8 Pentacam (Oculus Optikgerate, Wetzlar, Germany),4,9,10 specular microscopy,6 ultrasound biomicroscopy,11,12 and optical coherence tomography (OCT).5,13 Total corneal thickness profile in keratoconus has been measured previously using Orbscan,7 Pentacam,9,10,14 and Visante OCT (Carl Zeiss Meditec AG, Jena, Germany)15 whereas other studies have provided total corneal thickness measurements at specific locations in the peripheral cornea.1,3,5,12,16–18

In keratoconus, the epithelium is known to thin in the area overlying the cone, and in advanced keratoconus, there may be excessive epithelial thinning leading to a breakdown in the epithelium. Epithelial thinning over the cone has been demonstrated using histopathologic analysis of keratoconic corneas,19 and the central epithelial thickness was found to be thinner in keratoconus than in normal corneas using OCT.5 Transmission electron microscopy has demonstrated that the epithelial thickness was irregular in keratoconus.20

The stromal thickness profile is also of interest to study as stromal thinning occurs during the progression of the disease.5,21 Early changes in the stromal thickness progression may be helpful in the early diagnosis of keratoconus.

Artemis very high-frequency (VHF) digital ultrasound (ArcScan, Morrison, Colo) is to date the only published method of accurately measuring individual corneal layers, including the epithelium and stroma, continuously over a large area.22,23 We have previously used the Artemis to describe the average epithelial thickness profile24 and stromal thickness profile25 in a population of normal eyes. The repeatability of total corneal thickness measurements in 10 consecutive examinations of 1 eye using the Artemis 1 VHF digital ultrasound arc-scanning system has been shown to be less than 8 µm within the central 8-mm diameter, with a central repeatability of 1.5 µm.23 The repeatability of epithelial thickness measurements in the normal population has been shown to be less than 1.30 µm within the central 8-mm diameter, with a central repeatability of 0.5 µm.23

The purpose of this study was to qualitatively and quantitatively characterize the thickness profile of the epithelium, stroma, and full cornea in a population of keratoconic eyes using the Artemis VHF digital ultrasound arc-scanning system.

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Epithelial, Stromal, and Total Corneal Thickness in Keratoconus: Three-Dimensional Display with Artemis Very-High Frequency Digital Ultrasound

Chapter 1: Physics of Ultrasound Chapter 2: Ultrasonic Systems Chapter 3: Ocular Diagnosis Chapter 4: Very High Frequency digital ultrasound scanning in LASIK and Phakic Intra-ocular Lenses Chapter 5: Orbital Diagnosis Appendices Index

D. Jackson Coleman

Papers

Epithelial Thickness in the Normal Cornea: Three-dimensional Display With Very High Frequency Ultrasound

Stromal Thickness in the Normal Cornea: Three-dimensional Display With Artemis Very High-frequency Digital Ultrasound

Epithelial, Stromal, and Total Corneal Thickness in Keratoconus: Three-Dimensional Display with Artemis Very-High Frequency Digital Ultrasound

Ultrasonography Of The Eye And Orbit

Unified Model for Accommodative Mechanism