The impact of ocular blood flow in glaucoma.

Raman spectroscopy is a potentially important clinical tool for real-time diagnosis of disease and in situ evaluation of living tissue. The purpose of this article is to review the biological and physical basis of Raman spectroscopy of tissue, to assess the current status of the field and to explore future directions. The principles of Raman spectroscopy and the molecular level information it provides are explained. An overview of the evolution of Raman spectroscopic techniques in biology and medicine, from early investigations using visible laser excitation to present-day technology based on near-infrared laser excitation and charge-coupled device array detection, is presented. State-of-the-art Raman spectrometer systems for research laboratory and clinical settings are described. Modern methods of multivariate spectral analysis for extracting diagnostic, chemical and morphological information are reviewed. Several in-depth applications are presented to illustrate the methods of collecting, processing and analysing data, as well as the range of medical applications under study. Finally, the issues to be addressed in implementing Raman spectroscopy in various clinical applications, as well as some long-term directions for future study, are discussed.

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Prospects for in vivo Raman spectroscopy.

The present work relates the turbidity of the eye to microscopic spatial fluctuations in its index of refraction. Such fluctuations are indicated in electron microscope photographs. By examining the superposition of phases of waves scattered from each point in the medium, we provide a mathematical demonstration of the Bragg reflection principle which we have recently used in the interpretation of experimental investigations: namely, that the scattering of light is produced only by those fluctuations whose fourier components have a wavelength equal to or larger than one half the wavelength of light in the medium. This consideration is applied first to the scattering of light from collagen fibers in the normal cornea. We demonstrate physically and quantitatively that a limited correlation in the position of near neighbor collagen fibers leads to corneal transparency. Next, the theory is extended to predict the turbidity of swollen, pathologic corneas, wherein the normal distribution of collagen fibers is disturbed by the presence of numerous lakes-regions where collagen is absent. A quantitative expression for the turbidity of the swollen cornea is given in terms of the size and density of such lakes. Finally, the theory is applied to the case of the cataractous lens. We assume that the cataracts are produced by aggregation of the normal lens proteins into an albuminoid fraction and provide a formula for the lens turbidity in terms of the molecular weight and index of refraction of the individual albuminoid macromolecules. We provide a crude estimate of the mean albuminoid molecular weight required for lens opacity.

Theory of Transparency of the Eye

The organization of collagen fibrils in the human cornea and sclera was studied by scanning electron microscopy, after digestion of cellular elements by sodium hydroxide, and by conventional transmission electron microscopy. The collagen fibrils in the cornea had a uniform diameter of about 25 nm. In Bowman's layer, individual collagen fibrils were interwoven densely to form a felt-like sheet. In the stroma, most of the collagen fibrils ran abreast in lamellae, with varying widths and thickness. These lamellae were arranged basically parallel to the corneal surface but often communicated with those of adjacent layers by interchanging their fibrils. In the innermost stromal region adjacent to Descemet's membrane, collagen fibrils were oriented in various directions and interlaced, forming loose fibrillar networks. The sclera, however, was composed of collagen fibrils with various diameters ranging from 25-230 nm. Although these collagen fibrils formed bundles, they were not parallel but were entangled in individual bundles. These collagen bundles varied in width and thickness, often gave off branches, and intertwined with each other.

The three-dimensional organization of collagen fibrils in the human cornea and sclera.

Numerical Techniques in Electromagnetics is designed to show the reader how to pose, numerically analyze, and solve electromagnetic (EM) problems. It gives them the ability to expand their problem-solving skills using a variety of available numerical methods. Topics covered include fundamental concepts in EM; numerical methods; finite difference methods; variational methods, including moment methods and finite element methods; transmission-line matrix or modeling (TLM); and Monte Carlo methods. The simplicity of presentation of topics throughout the book makes this an ideal text for teaching or self-study by senior undergraduates, graduate students, and practicing engineers.

Numerical Techniques in Electromagnetics, Second Edition

The physical basis for the transparency of the cornea to visible light is investigated theoretically in terms of the molecular structure as depicted by electron microscopy. Electron micrographs show that the major portion of the cornea contains long cylindrical fibrils arranged in a quasi-random fashion, with local order extending over distances comparable to the wavelength of light. Heretofore, the generally accepted explanation of transparency has been in terms of a supposed crystalline arrangement of the fibrils, because this was the only distribution that could ensure transparency on a simple theoretical basis. Thus, the non-crystalline structure shown by the electron microscope has been widely regarded as an artifact due to the fixation procedure. In the present work, the light scattering from the fibrils is formulated in terms of their radial distribution function, which is determined by numerical analysis of electron micrographs. Comparison of theoretical results and experimental values for transmittance through rabbit cornea shows that the quasi-regular quasi-random structure revealed by the electron microscope is not in conflict with transparency.

Light scattering in the cornea.

It is widely appreciated that impaired vascular circulation in the eye plays a significant role in the pathology of the retina, the optic nerve and the choroid. Capillary closure, microaneurysm formation, shunts, neovascularization and breakdown of the blood retinal barrier are non-specific responses to ischemia, congestion and thrombosis. Examples include diabetic retinopathy, where the microaneurysms and neovascularization reflect an underlying focal hypoxia and impaired ocular blood flow (Ashton 1953,1969; Davis 1968; Kohner et al. 1969; Gamer & Ashton 1972). Unfortunately, the identification of patients with impaired ocular blood flow, and the assessment of treatments aimed at improving the ocular circulation have been precluded by the absence of a suitable routine measurement of ocular blood flow. This situation is now changing with the recent development of non-invasive procedures for the assessment of ocular blood flow in the retina and ciliary choroidal networks. The non-invasive procedure of laser doppler velocimetry was developed for the measurement of retinal blood flow (Riva & Feke 1981; Riva et al. 1981, 1985). In this procedure the retinal blood flow is derived from the velocity (V) of red cells and the cross sectional area (A) of a given artery or vein; blood flow equals VA (mm3 min-1) for an individual vessel, and the total blood flow is the sum of flows in all the retinal arteries or veins. The mean velocity of blood was measured using a bidirectional laser doppler system and the vessel diameter assessed from monochromatic fundus photographs. In 8 healthy human eyes the average total volumetric flow in the retina was 34 p1 min-1 with a range of 18-44 pl min-I. The values of the retinal blood flow in man are similar to those determined in primate eyes by the radioactive labelled microsphere entrapment procedure, namely 25 If: 9 ~l min-l (Alm & Bill 1973). The non-invasive quantitative assessment of the ciliary choroidal blood flow is based on the analysis of the intraocular pressure (IOP) pulse. The flow of blood into the eye is pulsatile and causes a rhythmic fluctuation of the steady-state IOP. It is the magnitude and the form of the IOP pulse that is used to assess the pulsatile component of ocular blood flow. The high fidelity recording of the IOP that is essential for the evaluation of the pulsatile blood flow (PBF) is obtained with a pneumatic tonometric probe (Langham 1987). In order to quantitate, automate and rapidly analyze the IOP pulsations, the electronic signal from the probe is digitalized and fed into a modified IBM PS2/30 computer. Representative recordings of the IOP in eyes of 4 normal subjects are shown in Fig.1. Each IOP measurement is completed within 10 microseconds and the measurement repeated at intervals of 30 milliseconds, giving a total of 30 IOP readings during each cycle of the IOP. The average of all the individual IOP measurements is the steady state value, Goldmann (1954) recognized the problem of

Blood flow in the human eye.

A relationship has been derived between intraocular pressure and pulsatile blood flow in the eye. Measurements of intraocular pressure show a time variation that is associated with the pulsatile component of arterial pressure. Experimental results provide a means of transforming intraocular pressure changes into ocular volume changes. The eye is represented by a chamber with elastic walls, a pulsatile incoming flow of incompressible fluid (blood), and a steady outgoing flow of blood. Under these conditions, the rate of pulsatile blood flow through the eye can be approximated from the instantaneous intraocular pressure measurements. Data from a healthy human eye are used to illustrate the analysis.

Estimation of pulsatile ocular blood flow from intraocular pressure.

1. A theoretical and experimental analysis of the relationship of the corneal stromal ultrastructure with light transmission has been made in an attempt to resolve recent contradictory explanations of corneal transparency.



2. The spatial distribution of collagen fibrils in electronmicrographs of rabbit corneal stroma has been analysed in terms of a radial distribution function. The results indicate the presence of local order extending to at least 200 nm from individual fibrils.



3. The observed spatial distribution of the collagen fibrils was used as a basis to compare the theoretically derived and the experimentally determined values of light transmission. It has been found that the transparency of the normal cornea may be explained by the quasi-random structure revealed by the electronmicroscope.



4. Histograms of the collagen fibril diameter in normal rabbit corneal stroma revealed the range to be 12·5–32·5 nm and the mean value to be approximately 20 ± 1·5 nm. Corneal swelling did not change the collagen fibril diameter significantly.



5. It is concluded that the size and distribution of collagen fibrils revealed in electronmicrographs are consistent with the observed transparency of normal stromas.



6. A marked heterogeneity in the spatial distribution of collagen fibrils was found in the swollen cornea. This is qualitatively consistent with the observed decrease in transparency.

https://physoc.onlinelibrary.wiley.com/doi/pdf/10.1113/jphysiol.1970.sp009230

The transparency of the mammalian cornea

1. The studies described herein involve the use of light scattering measurements to characterize the ultrastructural arrangement of the constituent collagen fibrils in rabbit corneal stromas.

2. Theoretical light scattering techniques for calculating the scattering to be expected from the structures revealed by electron micrographs are discussed, and comparison with the experimental light scattering tests the validity of these structures.

3. The wave-length dependence of light transmission and of angular light scattering from normal corneas is in agreement with the short range ordering of collagen fibrils depicted in electron micrographs.

4. The transmission measurements on oedematous rabbit corneas indicate that transmission decreases linearly with the ratio of thickness to normal thickness.

5. The wave-length dependence of transmission through cold swollen corneas indicates that the increased scattering is caused by large inhomogeneities in the ultrastructure. Electron micrographs do, indeed, reveal the presence of such inhomogeneities in the form of large regions completely devoid of fibrils.

Richard A. Farrell

Papers

Light scattering in the cornea.

Blood flow in the human eye.

Estimation of pulsatile ocular blood flow from intraocular pressure.

The transparency of the mammalian cornea

Wave‐length dependencies of light scattering in normal and cold swollen rabbit corneas and their structural implications