Simultaneous fluorometry and phosphorometry of Langendorff perfused rat heart: ex vivo animal studies

Fluorescence imaging of intrinsic fluorophores of tissue is a powerful method to assess metabolic changes at the cellular and intracellular levels. At the same time, exogenous phosphorescent probes can be used to accurately measure intravascular tissue oxygenation. Heart failure is the leading cause of death in America. A rat heart can potentially model the human heart to study failures or other abnormalities optically. We report simultaneous fluorescence and phosphorescence measurements performed on a rat heart. We have used two different optical systems to acquire fluorescence signals of flavoprotein and nicotinamide adenine dinucleotide—the two intrinsic fluorophores of mitochondria—and the phosphorescence signal of an intravascular oxygen probe to extract intracellular and intravascular metabolism loads, respectively

Respiratory muscle deoxygenation during exercise in patients with heart failure demonstrated with near-infrared spectroscopy

AbstractExertional dyspnea in patients with heart failure may be due, in part, to respiratory muscle underperfusion. Near-infrared spectroscopy is a new technique that permits noninvasive assessment of skeletal muscle oxygenation by monitoring changes in nearinfrared light absorption. With use of near-infrared spectroscopy, serratus anterior muscle oxygenation during maximal bicycle exercise was compared in 10 patients with heart failure (ejection fraction 16 ± 5%) and 7 age-matched normal subjects. Oxygen consumption (VO2), minute ventilation (VE) and arterial saturation were also measured. Changes in difference in absorption between 760 and 800 nm, expressed in arbitrary units, were used to detect muscle deoxygenation.Minimal change in this difference in absorption occurred in normal subjects during exercise, whereas patients with heart failure exhibited progressive changes throughout exercise consistent with respiratory muscle deoxygenation (peak exercise: normal 3 ± 6, heart failure 12 ± 4 near-infrared arbitrary units, p < 0.001). At comparable work loads patiente with heart failure had significantly greater minute ventilation and respiratory rate but similar tidal volume when contrasted with normal subjects. However, at peak exercise normal subjects achieved significantly greater minute ventilation and tidal volume with a comparable respiratory rate. No significant arterial desaturation occurred during exercise in either group.These findings indicate that respiratory muscle deoxygenation occurs in patients with heart failure during exercise. This deoxygenation may contribute to the exertional dyspnea experienced by such patients

Time-resolved reflectance measurements on layered tissues with strongly varying optical properties

Most biological tissues consist of layers with different optical properties. A few examples are the skin, the esophagus, the stomach and the wall of arteries. An understanding of how the light propagates in such layered systems is a prerequisite for any light based therapy or diagnostic scheme. In this study we investigate the influence of different kinds of layers on time resolved reflectance measurements. Experiments were performed on layered gel phantoms and the results compared to Monte Carlo simulations and diffusion theory. It is shown that when a low absorbing medium is situated on top of a high absorbing medium, the absorption coefficient of the lower layer is accessible if the differences in the absorption coefficient are only small. In the case of large difference the optical properties of the upper layer dominate the signal and shield information on the lowest layer. The degree of this shielding effect depends on layer thickness as well as optical properties. In the case of an almost absorption and scattering free layer in between two normal tissues, an overall increase of the signal is visible. However, the overall shape of the curve is about preserved. The apparent scattering coefficient is slightly decreased, while the apparent absorption coefficient is unaltered

Near-infrared spectroscopy of a heterogeneous turbid system containing distributed absorbers

In most biological tissues, absorbers such as blood in the blood vessels are localized within a low-absorbing background medium. To study the effect of distributed absorbers on the near infrared reflectance, we developed a Monte Carlo code and performed time-domain measurements on heterogeneous tissue-vessel models. The models were made of low absorbing polyester resin mixed with TiO_2 as scatters. A series of tubes with diameters of 3.2 or 6.4 mm were made in the resin sample. The volume ratio of the tubes to the total sample is about 20%. During the measurement, these tubes were filled with turbid fluids with different absorption coefficients to simulate blood in various oxygenation states. We found that the apparent absorption coefficient of the resin/tube system, determined by using the diffusion equation fit, can be approximated by a volume-weighted sum of the absorption coefficients of the different absorbing components. This approximation has to be replaced by a more complex expression if the difference in absorption between the absorbers and background is very large (approximately 20 times). The results of the tissue phantom study are supported by the Monte Carlo simulation. Possible explanations for the photon migration in this kind of heterogeneous system is also presented

Determination of blood oxygenation in the brain by time-resolved reflectance spectroscopy: influence of the skin, skull, and meninges

Near infrared light has been used for the determination of blood oxygenation in the brain but little attention has been paid to the fact that the states of blood oxygenation in arteries, veins, and capillaries differ substantially. In this study, Monte Carlo simulations for a heterogeneous system were conducted, and near infrared time-resolved reflectance measurements were performed on a heterogeneous tissue phantom model. The model was made of a solid polyester resin, which simulates the tissue background. A network of tubes was distributed uniformly through the resin to simulate the blood vessels. The time-resolved reflectance spectra were taken with different absorbing solutions filled in the network. Based on the simulation and experimental results, we investigated the dependence of the absorption coefficient obtained from the heterogeneous system on the absorption of the actual absorbing solution filled in the tubes. We show that light absorption by the brain should result from the combination of blood and blood-free tissue background

Discrepancy between cardiorespiratory system and skeletal muscle in elite cyclists after hypoxic training

BACKGROUND: The purpose of this study was to determine the effects of hypoxic training on the cardiorespiratory system and skeletal muscle among well-trained endurance athletes in a randomized cross-over design. METHODS: Eight junior national level competitive cyclists were separated into two groups; Group A trained under normoxic condition (21% O(2)) for 2 hours/day, 3 days/week for 3 weeks while Group B used the same training protocol under hypoxic condition (15% O(2)). After 3 weeks of each initial training condition, five weeks of self-training under usual field conditions intervened before the training condition was switched from NT to HT in Group A, from HT to NT in Group B. The subjects were tested at sea level before and after each training period. O(2 )uptake ([Image: see text] O(2)), blood samples, and muscle deoxygenation were measured during bicycle exercise test. RESULTS AND DISCUSSION: No changes in maximal workload, arterial O(2 )content, [Image: see text] O(2 )at lactate threshold and [Image: see text] O(2max )were observed before or after each training period. In contrast, deoxygenation change during submaximal exercise in the vastus lateralis was significantly higher at HT than NT (p < 0.01). In addition, half time of oxygenation recovery was significantly faster after HT (13.2 ± 2.6 sec) than NT (18.8 ± 2.7 sec) (p < 0.001). CONCLUSIONS: Three weeks of HT may not give an additional performance benefit at sea level for elite competitive cyclists, even though HT may induce some physiological adaptations on muscle tissue level

Fluorescence spectroscopy and imaging of myocardial apoptosis

Fluorometry is used to detect intrinsic flavoprotein (FP) and nicotinamide adenine dinucleotide NADH signals in an open-chest rabbit model of myocardial ischemia-reperfusion injury. Myocyte apoptosis has been shown clinically to contribute to infarct size following reperfusion of ischemic myocardium. A noninvasive means of assessing apoptosis in this setting would aid in the treatment of subsequent ventricular remodeling. We show that in vivo fluorometry can be useful in apoptosis detection in open-chest surgeries. Specific changes in myocardial redox states have been shown to indicate the presence of apoptosis. Two main mitochondrial intrinsic fluorophores, NADH and FP signals, were measured during normoxia, ischemia, and reperfusion experimental protocol. Ischemia was induced by occlusion of the largest branch of the circumflex coronary artery and fluorescence signals are collected by applying two different fluorescence techniques: in vivo fluorometry and postmortem cryoimaging. The first technique was employed to detect FP and NADH signals in vivo and the latter technique uses freeze trapping and lowtemperature fluorescence imaging. The heart is snap frozen while still in the chest cavity to make a snapshot of the metabolic state of the tissue. After freezing, the ischemic area and its surrounding border zone were excised and the sample was embedded in a frozen buffer for cryoscanning. These two data sets, in vivo fluorometry and low temperature redox scanning, show consistent extreme oxidation of the mitochondrial redox states (higher redox ratio) suggesting the initiation of apoptosis following reperfusion. This represents the first attempt to assess myocyte apoptosis in the beating heart

Quantifying Acute Myocardial Injury Using Ratiometric Fluorometry

Early reperfusion is the best therapy for myocardial infarction (MI). Effectiveness, however, varies significantly between patients and has implications for long-term prognosis and treatment. A technique to assess the extent of myocardial salvage after reperfusion therapy would allow for high-risk patients to be identified in the early post-MI period. Mitochondrial dysfunction is associated with cell death following myocardial reperfusion and can be quantified by fluorometry. Therefore, we hypothesized that variations in the fluorescence of mitochondrial nicotinamide adenine dinucleotide (NADH) and flavoprotein (FP) can be used acutely to predict the degree of myocardial injury. Thirteen rabbits had coronary occlusion for 30 min followed by 3 h of reperfusion. To produce a spectrum of infarct sizes, six animals were infused cyclosporine A prior to ischemia. Using a specially designed fluorometric probe, NADH and FP fluorescence were measured in the ischemic area. Changes in NADH and FP fluorescence, as early as 15 min after reperfusion, correlated with postmortem assessment infarct size (r=0.695, p\u3c0.01). This correlation strengthened with time (r=0.827, p\u3c0.01 after 180 min). Clinical application of catheter-based myocardial fluorometry may provide a minimally invasive technique for assessing the early response to reperfusion therapy