Absolute quantification (ml blood/sec ∗ mm2 tissue) of normal vs. diabetic foot skin microvascular blood perfusion: Feasibility of FM-PPG measurements under clinical conditions.

Fluorescence-mediated photoplethysmography (FM-PPG) is the first routine clinical methodology by which to quantifiably measure tissue blood perfusion in absolute terms (mL blood/sec ∗ mm2 tissue). The FM-PPG methodology has been described in detail previously in this journal (MVR 114, 2017, 92-100), along with initial proof-of-concept measurements of blood perfusion in both ocular and forearm skin tissues. The motivation for the current study was to investigate whether FM-PPG can be used readily and routinely under realistic clinical conditions. The vehicle for doing this was to measure medial foot capillary blood flow, i.e., tissue perfusion, in 7 normal subjects, mean = 6.76 ±â€¯2.29 E-005 mL/(sec ∙ mm2), and lesion-free areas of 8 type-2 diabetic patients with skin ulceration, mean = 4.67 + 3.15 E-005 mL/(sec ∙ mm2). Thus, perfusion in the diabetics was found to be moderately lower than that in the normal control subjects. Earlier skin perfusion measurements in medial forearms of 4 normal subjects, mean = 2.64 + 0.22 E-005 mL/(sec ∙ mm2), were lower than both the normal and diabetic foot perfusion measurements. Variability in the heartbeat-to-heartbeat blood perfusion pulses in the skin capillaries, defined as the ratio of the standard deviation among beat-to-beat pulses divided by the mean perfusion of those pulses, was determined for each subject. Average variability in foot skin was 21% in the diabetic population, versus 16% for normal subjects; and it was 18% in forearm skin. We conclude that absolute quantitative FM-PPG measurement of skin blood perfusion at the level of nutritive capillaries is feasible routinely under clinical conditions, allowing for quantitative measurement of skin tissue blood perfusion in absolute terms.

Prostaglandins as mediators of vasotonia in the immature retina.

In vivo observations of the immature beagle puppy retina indicates that in general O2-breathing induces vasoconstriction whereas CO2 or CO2 combined with O2 induces vasodilation. It was the goal of this study to determine whether a specific correlation could be found between this pattern of vascular response to changes in blood gas content and production of prostaglandins in the eye. Prostaglandin production by puppy choroidal and retinal tissues and platelets exposed in vitro to these gas combinations was such that it appears platelet-derived thromboxane could have a significant influence in determining retinal vasotonia.

Extraction of choriocapillaris hemodynamic data from ICG fluorescence angiograms.

PURPOSE: There are conflicting views about the organization of the posterior pole choriocapillaris, particularly concerning blood flow through it, and there are difficulties associated with attempting to obtain such information using histologic techniques, sodium fluorescein angiography, or both. The present study uses a method of analysis based on high-speed angiograms to investigate posterior pole choroidal blood flow. METHOD: The analysis method employed is based on the premises that dye filling of the choriocapillaris is more rapid (because it is pulsatile) than dye filling of the underlying larger diameter vessels, and that fluorescence from these two overlapping layers is additive. The described analysis algorithm was applied to high-speed ICG fluorescence angiograms to emphasize information about choriocapillaris hemodynamics. RESULTS: The analysis method was demonstrated in rhesus monkeys, and results indicate that the posterior pole choriocapillaris does not behave as a homogeneous structure, consisting of discrete lobular segments. In general, the cycle of dye filling of the choriocapillaris begins in the macular area and progresses radially toward the periphery in a wavelike manner, the filling cycle beginning with and being completed during one cycle of the intraocular pressure pulse. CONCLUSIONS: It is possible to extract information related to choriocapillaris blood flow from high-speed ICG angiograms when a fairly well-defined dye bolus wavefront is present. Interpretation of the results suggests that the choriocapillaris lobules fill in a pulsatile manner, out of phase with each other, and may act thereby to dissipate the blood volume entering the choroid during each cardiac cycle in such a way that the retinal macular is not significantly displaced by expansion of the choroidal vascular volume. Depending upon the distribution of pressure gradients across a group of lobules, blood may flow from one lobule into or even through an adjacent one.

Variability in choriocapillaris blood flow distribution.

PURPOSE: To investigate variability of choriocapillaris blood flow patterns. METHODS: After the intravenous injection of indocyanine green, angiograms were recorded at 30 images per second in rhesus monkey eyes using a fundus camera equipped with a pulsed laser diode light source, synchronized with a gated (5 msec), intensified charge-coupled device, or CCD, video camera. Images of choriocapillaris filling alone were extracted. Plastic corrosion casts were made of two of the monkey's choroidal vasculatures for subsequent scanning electron microscopy examination. RESULTS: Pulsed laser indocyanine green fluorescence excitation produced better definition of choriocapillaris filling than had been achieved using continuous illumination. No correlation was found between the choriocapillaris plexus architecture revealed by the plastic corrosion casts and the observed choriocapillaris lobular filling. Overall posterior pole choriocapillaris dye-filling patterns were relatively stable for periods of days, but they changed gradually for periods of weeks. Localized minor pattern changes occurred on a much shorter time scale. Choriocapillaris filling patterns were altered by acutely elevating intraocular pressure, by O2 and CO2 breathing, and by argon laser retinal photocoagulation of adjacent areas. CONCLUSIONS: Choriocapillaris filling patterns appear to be determined by the network of perfusion pressure gradients that exist among the interspersed feeding arterioles and draining venules connected to the choriocapillaris plexus. Changes in intraocular pressure and in blood PO2 and PCO2 levels can produce marked changes in the distribution of choriocapillaris blood flow. Retinal laser photocoagulation of adjacent fundus areas alters choriocapillaris blood flow to the extent that the altered flow might be an important factor in the beneficial results attributed to retinal laser treatment.

The effect of blood on ocular fundus reflectance and determination of some optical properties of retinal blood vessels.

Light reflected from 50 micrometer diameter spots on the fundi of two adult rhesus monkeys was measured as a function of wavelength in 10 nm jumps from 400 to 900 nm. The areas measured were a retinal artery and vein, areas of the disk, macula, and retina devoid of visible blood vessels, and the foveola. The eyes of the two monkeys were then exsanguinated, the blood vessels were filled with normal saline, and reflectances of the same spots on the fundi were again measured. The pairs of reflectance curves were compared and demonstrated that blood is not the major determinant of the characteristic shapes of the retinal tissue spectral reflectance curves. From the pre-exsanguination and postexsanguination data, the isolated retinal artery and vein wall reflectances were determined to be 0.020 and 0.009, respectively, their transmittances 0.837 and 0,977, and the fractions of light absorbed by them 0.143 and 0.014, respectively, in the visible spectral region below 500 nm.

An image processing approach to characterizing choroidal blood flow.

Indocyanine green (ICG) dye angiography has made possible routine visualization of choroidal blood flow in the human eye; however, to date, its clinical utility has been limited. An overlying layer of densely pigmented tissue and the complex, multilayered vascular structure of the choroid combine to produce angiographic images of low contrast which are difficult to interpret. Conventional image processing can enhance individual images of the blood vessels, but this approach contributes no information about the dynamics of blood flow. Using relatively inexpensive, commercially available personal computer hardware, angiographic image processing algorithms were developed which appear to characterize uniquely a subject choroid in terms of various blood flow parameters. We believe this to be the first successfully demonstrated approach to routinely characterizing the human choroidal circulation in a way that conserves spatial distribution of blood flow dynamics across the entire observed choroidal area. The computer system allows acquisition of digital images from photographic film negatives; alternatively, real-time direct digitization of images from a high-resolution video camera is possible. Once acquired, the digitized data are manipulated according to various algorithms that employ time-sequence analysis to generate two-dimensional curves or three-dimensional surfaces which characterize the choroidal circulation. The unique correspondence of each three-dimensional surface to the subject choroidal circulation from which it was derived is demonstrated. Grouping the characteristic three-dimensional surfaces according to various topographic features in common may provide a basis for discriminating between normal and abnormal choroidal circulations.

Reflection of light by small areas of the ocular fundus.

Light reflected from 50 micrometer spots on the fundi of two rhesus monkeys and three human volunteers was measured as a function of wavelength in 10 nm. jumps from 400 to 900 nm. The areas measured were a retinal artery and vein, areas of the disk, macula, and retina devoid of visible blood vessels, and the fovea.

The mechanism of optic nerve damage in experimental acute intraocular pressure elevation.

We produced intraocular pressure (IOP) elevations in 32 primate eyes and studied retinal ganglion cell rapid axonal transport with autoradiography and electron microscopy. Animals breathing room air at sea level pressure were compared to animals breathing 100% oxygen at 3 atm pressure in a hyperbaric chamber. Despite major increases in arterial oxygen levels in the hyperbarically oxygenated animals, both groups had axonal transport blockade at the optic nerve head. Anoxia appears not to be the most important cause of acute axonal damage induced by elevated IOP. The pattern of axonal abnormality within individual fiber bundles at the optic nerve head provides support for mechanical compression as a more likely alternative cause for induced neural damage.

Postnatal retinal vascular development of the puppy.

Retinal vascular development during the first three postnatal weeks was studied in 63 purebred beagle puppies. Use of a positive enzyme histochemical reaction for adenosine triphosphatase in the nuclei and nucleoli of vascular cells made visualization of the retinal vasculature possible. Animals were killed by decapitation. Thus, artifacts resulting from use of anesthetics or tracer substances were avoided. In general, this study demonstrates important similarities between canine and human retinal vascular development, and this gives further reason to use of the puppy retina as a superior model for studying retrolental fibroplasia pathogenesis. This staining technique demonstrates undifferentiated cells in the avascular retina that appear to be vascular precursors or angioblasts. Primordial vessels form by organization of differentiating angioblasts that exist in peripheral retinal cystic spaces at birth, or by addition of fully differentiated endothelium; they form unlike neovascularization. Müller cell processes appear to provide a structural matrix throughout the avascular puppy retina on which differentiated angioblasts organize into a vascular network. Arteries develop in beds of primordial capillaries lying near the leading edge of the developing vasculature. This precedes vein formation which occurs through a process involving coalescence of embryonic capillaries which themselves were derived from primordial capillaries. Preliminary examination of eight mongrel kitten retinas prepared by this method clearly indicates that the puppy retina is much more completely vascularized at birth than that of the newborn kitten. Moreover, the rate of postnatal retinal vascularization is significantly faster in the kitten. The kitten vasculature does appear to form by the organization of angioblasts as in the puppy, but kitten angioblasts have a different appearance from those in the puppy.

Foreign body granulomas in normal ovaries.

In 100 consecutive cases in which grossly normal ovaries were removed at the time of pelvic surgery, 9% were found to contain crystalline foreign particles. An additional 9% contained cortical granulomas. In four of six cases, computer-assisted x-ray analysis of the crystalline foreign particles was successful and revealed magnesium and silicon.

Experimental studies of indocyanine green dye-enhanced photocoagulation of choroidal neovascularization feeder vessels.

PURPOSE: To report a model of choroidal neovascularization feeder vessels that reconciles current histologic, angiographic, and clinical data, and to report experimental studies that investigate the potential of indocyanine-green-dye-enhanced photocoagulation to improve feeder-vessel treatment. METHODS: A model of choroidal neovascularization feeder vessels was conceived to account for current histologic and angiographic data. Based on that model, experimental studies of the efficacy of indocyanine green-dye-enhanced photocoagulation were performed, using pigmented rabbit eyes as a model system. A Zeiss fundus camera was modified to permit visualization of choroidal blood flow by high-speed indocyanine green angiography and to permit simultaneous delivery of 810-nm-wavelength diode laser photocoagulation pulses to specific choroidal vascular targets during indocyanine green-dye bolus transit. RESULTS: Choroidal neovascularization feeder vessels appear to originate in the Sattler layer (that is, that portion of the choroidal vasculature consisting of medium-diameter vessels) and enter the choriocapillaris in close proximity to the small capillary-like vessels that penetrate Bruch membrane and communicate with the choroidal neovascularization. The rabbit eye experiments demonstrated that the presence of high indocyanine green dye concentration in circulating blood enhances uptake of near-infrared laser energy (three eyes); injection of sequential indocyanine green dye boluses results in gradually decreased efficiency of dye-enhanced photocoagulation (two eyes); and by application of laser energy during the initial transit of small-volume, high-concentration indocyanine green dye boluses, dye-enhanced photocoagulation of large diameter choroidal arteries can be accomplished with relatively little concomitant retinal tissue damage (three eyes). CONCLUSIONS: Although future trials will be necessary to substantiate these initial findings in the clinical arena, it appears that the efficiency of choroidal neovascularization feeder-vessel photocoagulation may be enhanced, while minimizing concomitant damage to overlying retinal tissue, by delivery of 810-nm wavelength laser energy immediately upon arrival of a high-concentration indocyanine green dye bolus in a targeted feeder vessel. However, molecules of dye adhering to vessel walls or lying in tissue interstitial spaces appear to divert laser energy from the photocoagulation process, so efficiency of indocyanine green dye-enhanced photocoagulation gradually diminishes as the number of injected dye boluses increases.

Electroretinographic changes and choroidal defects in a case of central retinal artery occlusion.

A 62-year-old white woman with hypertensive diabetes had central retinal artery occlusion of her right eye. Serial electroretinographic (ERG) studies demonstrated reversible changes in the scotopic components but permanent reduction of the photopic b-wave. Serial simultaneous fluorescein and indocyanine green angiographic studies demonstrated a perfusion defect in the choroid involving part of the macula which subsided after reopening the retinal circulation. Differential hypoxic susceptibility of the ERG photopic and scotopic b-waves was demonstrated, and changes in the patient's retinal tissue oxygenation were postulated on the basis of retinal and choroidal angiographic studies performed during and after central retinal artery occlusion to account for the observed changes in serial ERGS.

Theoretical investigation of the role of choriocapillaris blood flow in treatment of subfoveal choroidal neovascularization associated with age-related macular degeneration.

PURPOSE: To investigate the relationship between choriocapillaris blood flow and blood flow through an overlying choroidal neovascularization, as it relates to photocoagulation-induced changes in the choriocapillaris circulation. METHODS: A theoretical model that simulates the blood flow in the choriocapillaris and choroidal neovascularization of the human eye was developed, based on histologically determined vascular geometry and experimentally measured blood pressure gradients. The choriocapillaris blood pressure and blood flow were examined before and after simulated photocoagulation of various Sattler layer vessels entering the choriocapillaris in the vicinity of the choroidal neovascularization. (The Sattler layer is the inner layer of medium-sized choroidal vessels that includes both arterioles and venules that supply the choriocapillaris.) RESULTS: The theoretical model showed that both partial and complete occlusion of either Sattler arteriole or venous vessels in the vicinity of the capillary-like vessels connecting a choroidal neovascularization to the underlying choriocapillaris results in significant choroidal neovascularization blood flow reduction. These theoretical results are similar to clinically observed changes induced by laser photocoagulation of feeder vessels. (In this discussion, the term "feeder vessels" refers to those vessels in an indocyanine green angiogram image that appear to supply blood to a choroidal neovascularization; these vessels appear to be Sattler layer vessels, rather than the histologically demonstrated short, capillary-like vessels that form choriocapillaris-choroidal neovascularization communications.) CONCLUSIONS: Reduction of choriocapillaris blood flow underlying a choroidal neovascularization may be sufficient to reduce the blood flow rate in the choroidal neovascularization and thereby reduce the associated retinal edema. The results also suggest that reduction of choriocapillaris blood flow may be the common hemodynamic event associated with the successful application of several currently practiced methods of choroidal neovascularization treatment, including feeder vessel photocoagulation, photodynamic therapy, transpupillary thermotherapy, and prophylactic drusen photocoagulation. Ultimately, this model may be useful in determining optimal placement of laser photocoagulation burns to achieve a desirable perturbation in choroidal blood flow distribution and thereby reduce choroidal neovascularization blood flow to the extent necessary to obliterate associated retinal edema.

Platelet-fibrin embolism in a rhesus macaque: angiographic and pathologic studies comparing fluorescein and indocyanine green.

A male rhesus macaque was found to have what appeared to be numerous platelet-fibrin emboli in retinal vessels in the perimacular area. Indocyanine green (ICG) dye fluorescence and fluorescein angiograms of the fundus demonstrated leakage of fluorescein, but not ICG, from the involved arterioles. Histopathologic changes in the eyes included occlusion of retinal and choroidal vessels with platelet-fibrin emboli, inner retinal ischemia, ischemic injury to the parafoveal capillary bed distally to occlusion of precapillary arterioles, and retinal exudate limited to the regions of capillary damage. Differential leakage of fluorescein may be explained by the difference in binding affinities of the 2 dyes to blood protein: 20% to 40% of the circulating fluorescein is unbound, and 98% of ICG is bound to serum albumin. Simultaneously or serially performed angiograms with fluorescent probes of different sizes might be used to obtain a qualitative measure of vascular integrity in persons with embolism, diabetic retinopathy, sickle cell retinopathy, vasculitis, and other disorders known to produce focal retinal and choroidal vascular occlusion.

An ICG angiogram-based clinical method for characterizing the choroidal circulation used to assess the hemorrheologic effects of pentoxifylline.

PURPOSE: To demonstrate an indocyanine green (ICG) angiography-based clinical method for characterizing choroidal blood flow and for detecting changes in choroidal circulation patterns, and by use of that method, to demonstrate that pentoxifylline affects choroidal blood flow. METHODS: High-speed ICG angiography was performed in rhesus monkeys before and after intravenous administration of pentoxifylline or saline (which served as a control) while monitoring blood pressure and heart rate. From these data, three-dimensional surface maps indicating the instantaneous relative distribution of choroidal blood flow during the peak of intra-ocular pressure pulse systole in a 30 degrees field, centered on the macula, were generated to characterize the state of the choroidal circulation at various times during the experiments. RESULTS: Comparisons of the 3-dimentional surface maps consistently indicated an increase in sub-macular choroidal blood flow occurring within 5 to 10 minutes post-pentoxifylline injection, with a gradual return to baseline level 20-40 minutes later. Injection of equal volumes of saline produced no changes in choroidal blood flow. CONCLUSIONS: Posterior-pole choroidal blood flow can be characterized as by a three-dimensional surface representing the instantaneous relative distribution of choroidal blood flow during the peak of intra-ocular pressure pulse systole. Pentoxifylline does, at least transiently, increase sub-macular choroidal blood flow.