Characterization of the fluorescent morphological structures in human arterial wall using ultraviolet-excited microspectrofluorimetry.

In this study, the fluorescent morphological structures in normal coronary artery, normal aorta, and atherosclerotic aorta were histochemically identified and spectroscopically characterized in situ using ultraviolet-excited microspectrofluorimetry. Excitation wavelengths of 290 nm and 310/312 nm were employed to observe two distinct fluorescence bands, with peak emission wavelengths near 335 nm and 380 nm, respectively. Emission of the short wavelength 335 nm band, previously assigned to tryptophan residues in tryptophan-containing proteins, was observed from all the morphological structures in the vessel walls and was isolated in groups of smooth muscle cells in aorta and coronary artery media. The long wavelength 380 nm band was assigned to distinct fluorophores associated with the structural proteins collagen and elastin and was observed in collagen fibers and elastic fibers, respectively. The corresponding morphological structures in normal aorta, normal coronary artery, and atherosclerotic aorta exhibited similar fluorescence lineshapes. In atherosclerotic plaque, a distinct fluorescence band, peaking near 370 nm, was observed in the emission from both ceroid granules and necrotic core. Using a simple, quantitative model, differing contributions of collagen, elastin, and tryptophan-containing protein fluorescence were shown to account for over 95% of the emission from the intima, media, and adventitia layers of non-necrotic aorta and coronary artery.

Photon-induced dissociation with a four-sector tandem mass spectrometer.

The feasibility of using photodissociation of protonated peptide molecules to sequence specific fragment ions with a 193-nm pulsed laser beam in a magnetic deflection tandem mass spectrometer of EBEB configuration was demonstrated. Although the short pulse (15 ns) and low repetition rate (100 Hz) of the excimer laser permitted the irradiation of only ∼ 0.02% of the (M + H)(+) ions exiting MS-1, a photon-induced decomposition spectrum of the heptapeptide angiotensio III (M r 930.5) was produced that was practically the same (but with better signal-to-noise ratio) as that generated by collision-activated dissociation at the same low duty cycle. Because of the low and pulsed fragment ion currents, an array detector was used to record the spectra. A dependence between laser power and abundance of fragment ions was observed (increased power increases the relative abundance of ions of low mass). Laser power was varied from 6 to 80 mJ. Formation of fragment ions from a large peptide (melittin, M, 2844.75) was also observed. The results permit the design of modifications that may increase the fragment ion yield to 10% or higher, which would make photon-induced decomposition a useful method for magnetic deflection mass spectrometers.

Removal of surgically induced fibrous arterial plaques by argon ion laser angiosurgery using a multifiber delivery system. An experimental study in the dog.

Removal of intravascular atherosclerotic obstructions by laser irradiation has gained the attention of many investigators, but has proven to be considerably more difficult to accomplish than initially envisioned. We tested, in an animal model, an argon ion laser delivery system that permits control of (1) laser power, (2) exposure time, and (3) laser beam spot size. The study was conducted on surgically, induced focal fibrous plaques in the carotid arteries of nine dogs. Plaque removal, vessel patency, and healing were evaluated angiographically and by light and electron microscopy at intervals up to 60 days after treatment. Results showed that intravascular obstructions could be removed, healing occurred, and vessels remained patent for up to 60 days.

Plasma shielding by Q-switched and mode-locked Nd-YAG lasers.

Q-switched and mode-locked Nd-YAG lasers depend on beam divergence and plasma formation to reduce energy transmission below the retinal damage threshold. In a model system of physiologic saline, we investigated plasma "shielding" and found that plasma formation markedly reduces further energy transmission along the beam path once plasma formation occurs. The protective effect of plasma shielding appears similar for a Q-switched pulse and a train of mode-locked pulses in a physiologic saline medium.

Alteration of spectral characteristics of human artery wall caused by 476-nm laser irradiation.

Fluorescence spectroscopy is a promising new technique for discrimination of normal and atherosclerotic arterial tissues. It has been suggested that this technique be used as a guidance system for laser angiosurgery catheters; however, irradiation by 476-nm light can change the spectroscopic properties of arterial tissue. We present studies that establish intensity levels and exposure times at which alterations in tissue spectral properties are minimal. We also investigate the nature of spectral alterations following exposure of normal human aorta to high intensities of 476-nm laser light. Changes in laser-induced fluorescence (LIF) are characterized by two prominent features: the peak fluorescence intensity decreases permanently, and the fluorescence lineshape changes in a largely reversible way. We relate these changes to alterations in individual tissue chromophores: permanent changes in absolute fluorescence intensity are due to irreversible changes in tissue fluorophores, reversible changes in fluorescence lineshape are due alterations in tissue absorbers. A simple kinetic model is used to describe the decrease in absolute fluorescence intensity.

Gas volume quantitation during argon ion laser ablation of atheromatous aorta in blood and 0.9% saline media with an optically shielded catheter.

Using an optically shielded fiber optic laser catheter, the amount of gas produced when firing an argon ion laser into 0.9% saline solution or blood alone and into atheromatous aorta in either a blood or 0.9% saline medium was quantitated. Energies from 0.25 to 4 joules (J) were used at powers of 2, 5, and 8 W. We found that total volume of gas produced is small not only at equilibrium (0.3 +/- 0.1 microliter/J when firing in blood alone and also when ablating aorta in blood or saline media) but also at peak (2.5 +/- 0.2 microliters/J firing in blood alone and 1.0 +/- 0.1 microliter/J or 0.9 +/- 0.1 microliter/J when ablating aorta in saline or blood, respectively). Because these volumes are small, a clinically significant event from a gas embolus is unlikely during intravascular laser ablation of atheromatous plaque in the energy and power range studied. No gas was quantitated when firing the argon ion laser into 0.9% saline solution alone. The peak gas volume when firing in blood alone was significantly greater than that produced in the other chamber environments. This is thought to be due to increased absorption of argon laser light by hemoglobin. The gas volumes produced by lasing aorta in 0.9% saline or blood were not statistically different.

Laser induced fluorescence spectroscopy of normal and atherosclerotic human aorta using 306-310 nm excitation.

Ultraviolet excited laser induced fluorescence (LIF) was studied in normal and atherosclerotic human arterial wall in vitro. Using excitation wavelengths from 306 to 310 nm, two distinct emission bands were observed in the LIF of both normal and pathologic aorta: a short wavelength band, peaking at 340 nm emission, which was attributed to tryptophan; and a long wavelength band, peaking at 380 nm emission, which was assigned to a combination of collagen and elastin. The intensity of the short wavelength band was quite sensitive to the choice of excitation wavelength, while the long wavelength band was not, so that the relative contributions of the bands could be controlled by the precise choice of excitation wavelength. A valley in the spectra at 418 nm was attributed to fluorescence reabsorption by oxy-hemoglobin. By using 308 nm excitation to observe emission simultaneously from both the short and long wavelength bands, normal and atherosclerotic aorta were spectrally distinct. Two LIF emission intensity ratios were defined to characterize both the relative tryptophan fluorescence content as well as the ratio of elastin to collagen fluorescence in each spectrum. The differences in these two emission ratios among the various histologic tissue types correlated qualitatively with the histologic and biochemical compositions of these tissues. By combining these parameters in a binary classification scheme, normal and atherosclerotic aorta were correctly distinguished in 56 of 60 total cases. Furthermore, atherosclerotic plaques, atheromatous plaques, and exposed calcifications could be classified individually with sensitivities/predictive values of 90%/90%, 100%/75%, and 82%/82%, respectively.

Remote biomedical spectroscopic imaging of human artery wall.

We discuss a general technique, laser spectroscopic imaging (LSI), remote acquisition of spectroscopic images of biological tissues and tissue conditions. The technique employs laser-induced spectroscopic signals, collected and transmitted via an array of optical fibers, to produce discrete pixels of information from which a map or image of a desired tissue characteristic is constructed. We describe a prototype LSI catheter that produces spectral images of the interior of human arteries for diagnosis of atherosclerosis. The diagnostic is based on the fact that normal artery wall and atherosclerotic plaque exhibit distinct fluorescence spectra in the 500-650 nm range when excited by 476-nm laser light; the fluorescence from blood is minimal. The catheter is composed of 19 optical fibers enclosed in a transparent, protective shield. Argon ion laser radiation is used for excitation, and an optical multichannel spectral analyzer is used for detection. Sequential sampling is used to minimize crosstalk among fibers and reduce blurring of the image. Computer-processed 19-pixel spectroscopic images are produced of fresh cadaver artery in vitro. Regions of normal tissue, plaque, and blood are identified, and the diagnoses are confirmed histologically and by direct spatial correlation. The results demonstrate the concept of using this laser catheter system for real-time imaging.

A model for thermal ablation of biological tissue using laser radiation.

We present a theory of thermal laser ablation based on the heat equation and on an energy balance equation derived from it. Ablation is assumed to be brought about by the heating and evaporation of tissue water. The model is three-dimensional, and scattering and the water-steam phase transition are explicitly taken into account. The model predicts threshold parameters and a steady-state ablation velocity in terms of the optical and thermal properties of the tissue and the laser beam intensity and spot diameter.

Characterization of ultraviolet laser-induced autofluorescence of ceroid deposits and other structures in atherosclerotic plaques as a potential diagnostic for laser angiosurgery.

Unstained frozen sections of normal and atherosclerotic human aorta and coronary artery were examined using histochemical and fluorescence microscopic techniques to identify the structures responsible for autofluorescence under 351 to 364 nm laser excitation. These structures included elastin and collagen in normal and atherosclerotic specimens, calcium deposits in calcified plaques, and granular or ring-shaped deposits histochemically identified as ceroid found in both calcified and non-calcified plaques. Qualitatively, both the color and intensity of ceroid autofluorescence differed greatly from that of elastin or collagen. The emission spectra of elastin, collagen, and ceroid were examined by microscopic spectrofluorimetry, and were found to differ significantly as well. When compared with spectra of elastin and collagen, spectra of ceroid were broader, shifted to the red, and were somewhat resistant to bleaching. We conclude that detection of laser-induced ceroid autofluorescence may aid in identifying plaques for laser ablation.

Effects of varying argon ion laser intensity and exposure time on the ablation of atherosclerotic plaque.

Using continuous wave (CW) argon ion laser light, a total of 253 laser exposures of varying power (1.5, 3, 5, 8 or 10 W) and duration (20-1,333 ms) were delivered to four segments of human atheromatous aorta obtained at autopsy. Exposure conditions were controlled by using an optically shielded laser catheter that provided a 500 micron spot of light of known power. Two thresholds for consistently reproducible ablation could be defined-an intensity threshold at 25.5 W/mm2 and a fluence threshold at 3.2 J/mm2. Above threshold, a fluence of 5.1 J/mm2 was found to produce the most efficient ablation, ie, removed the greatest volume (mm3) per energy delivered (J) compared to other fluence levels employed (p less than 0.0001). Between aortic segments, however, considerable variability in efficiency (mm3/J) was observed, possibly owing to different optical properties and/or plaque composition. Low-intensity laser radiation produced inconsistent ablation and extensive coagulation effects to surrounding tissue. When a fluence of 5.1 J/mm2 was constructed with a high-intensity laser beam and a short exposure time, consistent and efficient tissue removal resulted without histologic evidence of coagulation necrosis.

Spectral diagnosis of atherosclerosis using an optical fiber laser catheter.

This communication demonstrates that fluorescence spectra of human aorta with good S/N ratios can be collected using an optical fiber laser catheter. The performance of this catheter is compared to a non-fiber optic collection system with an equivalent delivery/collection geometry. For a given sample, fluorescence lineshapes obtained using the two systems are identical; differences in peak fluorescence intensity are related to the different collection efficiencies of the two systems. It is shown that the fluorescence lineshape of arterial tissue depends on the delivery/collection geometry of the detection system, and that this is due to the interaction of absorption and fluorescence within the artery wall. This effect is investigated systematically using a specially designed collection system. Results are analyzed qualitatively using a simple, one-dimensional model of tissue fluorescence. With this analysis, we present design requirements for a collection system in which such geometric effects are eliminated, and show that our optical fiber laser catheter satisfies these requirements.