Cytochrome Aa3 Concentration Research Articles

Near-infrared Spectroscopy: Neonatal and Perinatal Applications

After completing this article, readers should be able to: 1. Delineate the clinical benefits of near-infrared spectroscopy. 2. Describe the primary clinical applications of near-infrared spectroscopy. ### Development of Light-based Diagnostic Systems Transillumination, or the passage of light through the body, has been studied since the early 1800s. Attempts to image organs and tissues for the purpose of diagnosis and treatment have developed to the point that light-based monitoring in the form of pulse oximetry is used daily in hospitals and clinics. Transillumination of the head, first described in 1831 by Richard Bright, eventually would be recognized as the first light-based diagnostic technique to identify hydrocephalus. With modification, although still crude, it was used to diagnose intracranial hemorrhage in the neonate at a time before head ultrasonography was available and when computed tomography (CT) was extremely expensive and not widely available. In early devices, light in the visible part of the spectrum was employed; more recently, light over a narrower band of wavelengths has been used. In 1977, Jobsis showed that tissue absorption of near-infrared light at a wavelength of 700 to 1,000 nm was low enough to allow spectral measurements across the head of an animal with a diameter of 5 to 6 cm. The absorption of near-infrared light by particular pigments in the body changes as the oxygenation state of these pigments change. Thus, deoxyhemoglobin (Hb) absorbs light differently after it has bound oxygen and becomes oxyhemoglobin (HbO2). Cytochrome aa3, the terminal enzyme in the mitochondrial electron transport chain, also can be detected using near-infrared spectroscopy (NIRS). Measurement of cytochrome aa3 may allow assessment of oxygenation at the cellular level, with changes in oxidized and reduced cytochrome aa3 concentrations (redox state) indicating changes in molecular oxygen availability. The relative absorbances for HbO2 and Hb are unique at each wavelength (Fig. …

Changes in subcutaneous interstitial fluid pressure, tissue oxygenation, and skin red cell flux during venous congestion plethysmography in men.

Venous congestion plethysmography enables noninvasive assessment of microvascular filtration capacity (Kf) in limbs. However, increases in fluid filtration might alter the balance of Starling forces: for example, progressive increases in interstitial fluid pressure (Pi) would reduce net fluid flux, thus underestimating Kf. Furthermore, elevation of cuff pressure to values close to diastolic blood pressure, as used in the protocol, may be itself impair tissue perfusion with unknown effects on the microvascular parameters investigated. Pi was measured in healthy volunteers (n = 14) with a modified "Wick in needle" technique during small (8 mm Hg) cumulative increases in venous pressure (0-95 mm Hg). Changes in the hemoglobin (Hb) concentration, oxygenated hemoglobin (HbO2) concentration and oxidized cytochrome aa3 concentration were assessed in the calf using noninvasive near-infrared spectroscopy. Skin red blood cell flux close to the strain gauge was evaluated by laser Doppler fluxmetry. Pi at control was -0.89 +/- 0.8 mm Hg and during elevation of venous pressure remained constant until a cuff pressure of 30 mm Hg was reached. It rose thereafter to 1.57 +/- 1.3 mm Hg (mean +/- SD). Skin red cell flux was significantly reduced when cuff pressure exceeded 30 mm Hg and following cuff deflation, evidence of reactive hyperemia was obtained. Hb concentration increased significantly as a result of venous pressure elevation. No change in either HbO2 or cytochrome aa3 concentration was observed as long as cuff pressure remained under diastolic blood pressure. The small increase in Pi together with an absence of impaired tissue oxygenation during the venous congestion plethysmography protocol described by Gamble et al. supports the contention that this protocol enables accurate assessment of filtration capacity.

Cerebral Monitoring by Near-Infrared Spectroscopy

Near-infrared spectroscopy (NIRS) is a noninvasive optical monitoring technology that can provide information on relative cerebral oxyhemoglobin, deoxyhemoglobin, and oxidized cytochrome aa3concentrations. Unlike other bedside clinical monitoring technologies, NIRS provides direct information on cerebral hemoglobin oxygenation and blood volume changes. NIRS recordings have documented changes in cerebral oxygenation in fetuses during labor, critically ill premature infants, cardiopulmonary bypass patients, and adult surgical patients. This information may improve understanding of the causes of brain injury and allow detection of inadequate oxygen delivery before brain injury occurs. Future developments in NIRS technology are likely to produce practical, quantitative bedside monitors of global and regional cerebral oxygenation.

Apparent cerebral cytochrome aa3 reduction during cardiopulmonary bypass in hypoxaemic children with congenital heart disease. A critical analysis of in vivo near-infrared spectrophotometric data

The purpose of this analysis was critically to examine the changes of cerebral cytochrome oxidase as detected by near-infrared spectrophotometry (NIRS) during induction of cardiopulmonary bypass (CPB) in 14 children undergoing open heart surgery. Five children were hypoxaemic (arterial oxygen saturation 49%-84%) before the switch to CPB and 9 children were not. In the hypoxaemic children, the total cerebral haemoglobin concentration (tHb) decreased rapidly and markedly, reaching a plateau after 2 min. Surprisingly, cytochrome aa3 concentration in its oxygenated form (CytO2) decreased in parallel while the cerebral haemoglobin oxygenation index (HbO2)-(Hb) increased gradually. In the 8 normoxaemic children, changes in the NIRS signals were insignificant. When a standard NIRS algorithm was used, the magnitude of the change in (CytO2) was closely associated with the magnitude of the change in (tHb) (p<0.0001), the time courses of the (CytO2) and (tHb) signals were parallel in 11 of the 14 children and the error of fit in the NIRS multicomponent analysis increased 10- to 100-fold over system noise. A new NIRS algorithm, using wavelength specific optical pathlengths, did not improve the error of fit but produced smaller estimates of (CytO2) changes, which were unrelated to the (tHb) changes. In the authors' opinion the interesting possibility of monitoring cytochrome oxidation by NIRS requires further validation.

Delayed decreases in specific brain mitochondrial electron transfer complex activities and cytochrome concentrations following anoxia/ischemia

Hyperglycemic, but not normoglycemic cats exposed to anoxia develop neurologic signs following reoxygenation including fasciculations, focal and tonic-clonic seizures and coma after a symptom-free period. These symptomatic hyperglycemic cats may develop brain edema and will show diffuse neuronal injury or brain infarction depending on length of survival. Brain mitochondria isolated from symptomatic but not asymptomatic cats have decreased ADP- and uncoupler-stimulated oxygen consumption rates. Since impaired respiration could result from altered electron transport chain function, we measured cytochrome c, b, and aa3 concentrations and the activities of the five electron transfer complexes in isolated brain mitochondria. In symptomatic cats marked alterations were present in particular in complex IV, cytochrome oxidase, with a 57% reduction in activity and a 45% reduction in prosthetic group (cytochrome aa3) concentrations. Less marked reductions in other segments of the chain included 27% and 41% decreases, respectively, in cytochrome c concentrations and in electron transfer complex II, succinate:ubiquinone oxidoreductase activity. Cytochrome b concentrations and complex I, II and V activities were unchanged. Small but significant decreases in cytochrome aa3 concentrations (18%) and cytochrome oxidase activity (20%) were also present in mitochondria from postanoxic hyperglycemic cats prior to appearance of neurologic signs. These results indicate that delayed decreases in the activities of specific electron transfer complexes are correlated with impaired mitochondrial respiration and neurologic deterioration in postanoxic hyperglycemic cats. However, it is presently unclear if these postanoxic brain mitochondrial alterations are primary or secondary events in the development of brain injury.

82 Relation between arterial carbon dioxide tension and cerebral oxidised cytochrome aa3, concentration in preterm infants

The aim of this study was to find out whether the intracerebral concentration of cytochrome aa3 ([CytO2]) (the terminal enzyme of the respiratory electron transport chain) altered in response to changes in arterial carbon dioxide tension (PaCO2).

80 Effect of indomethacin on cerebral oxidised cytochrome aa3 concentration in preterm infants

Using near infrared spectroscopy (NIRS) we have shown previously that the intravenous administration of indomethacin for treatment of patent ductus arteriosus (PDA) in sick preterm infants caused a reduction of approximately 40% in cerebral oxygen delivery1. We have used NIRS to investigate the effect of this change on the cerebral concentration of oxidised cytochrome aa3 ((CytO2]), the terminal enzyme in the mitochondrial electron transport chain. Studies by NIRS were performed on 15 infants born at 23-29 weeks gestation weighing 600-1620 grams, who required treatment for PDA aged 8-27 days. Infants received indomethacin 0.1-0.2 mg.kg−, seven by fast (30 sees) and eight by slow (30 mins) infusion. The expected haemodynamic changes were seen in all infants. In 5 infants an unequivocal fall in [CytO2] was observed (range 0.3-0.9 μmol.1−), while the remaining 10 infants showed no definite change. No significant difference between the fast and slow infusion was observed. We conclude that following indomethacin the close relationship between cerebral intracellular oxygen supply and cerebral energy metabolism was frequently maintained, but in 5 cases there was evidence of a relative intracellular oxygen deficit. 1. AD Edwards et al. Pediatr Res 1989:26:522

Delayed Onset of Neurologic Deterioration following Anoxia/Ischemia Coincides with Appearance of Impaired Brain Mitochondrial Respiration and Decreased Cytochrome Oxidase Activity

We previously demonstrated markedly inhibited brain mitochondrial respiration only in cats that (a) were hyperglycemic at anoxia and (b) had neurologic signs, i.e., fasciculations in tongue or facial muscles or focal seizures following reoxygenation. However, since the relationship between time of onset of mitochondrial dysfunction and neurologic signs was unclear, in the present study we killed postanoxic cats immediately when signs first appeared. Cerebrocortical homogenates and isolated brain mitochondria only from symptomatic cats showed markedly inhibited substrate-, ADP-, and uncoupler-stimulated respiration rates. Cytochrome oxidase activity and cytochrome aa3 concentrations were also markedly reduced in these mitochondria. Since brain mitochondrial function was impaired when neurologic signs first appeared, mitochondrial alterations are an important early organellar change correlated with development of neurologic deterioration following anoxia.

Streptomycin accumulation by Bacillus subtilis requires both a membrane potential and cytochrome aa3

Cytochrome aa3 concentrations in the cytoplasmic membrane of Bacillus subtilis were altered by growth conditions, and the effects on the membrane potential (delta psi) in whole cells were measured. When cytochrome aa3 was absent, the magnitude of delta psi was not diminished by comparison with the delta psi measured in cells containing normal cytochrome aa3 concentrations. In addition, the energy-dependent uptake of proline and glutamate was comparable at both cytochrome aa3 concentrations. However, in the cytochrome aa3-deficient cell preparation, accumulation of the aminoglycoside antibiotic streptomycin was much lower than that of the cytochrome aa3-sufficient cells. When cells were cultured under conditions that stimulated higher than normal concentrations of cytochrome aa3, delta psi was also increased, and enhanced streptomycin accumulation was observed. Phenazine methosulfate-ascorbate was used both in delta psi measurements and in uptake studies to provide high rates of electron transport and maximal delta psi values. These results, taken together with those previously published (A. S. McEnroe and H. W. Taber, Antimicrob. Agents Chemother. 26:507-512, 1984) suggest that the uptake of streptomycin by B. subtilis requires adequate levels both of delta psi and cytochrome aa3.

Correlation between cytochrome aa3 concentrations and streptomycin accumulation in Bacillus subtilis.

Accumulation of aminoglycosides by Bacillus subtilis appears to require specific components of the electron transport chain. These components include cytochromes and the lipophilic quinone vitamin K2. The present study concerns the importance of cytochrome aa3, a terminal oxidase, in the uptake of streptomycin. Growth conditions have been established such that the concentration of cytochrome aa3 can be modified over a wide range; on defined minimal salts agar, the wild-type strain (RB1) and an strC mutant (RB95) synthesized cytochrome aa3 only when adequate amounts of Casamino Acids (Difco Laboratories, Detroit, Mich.) were present. A positive correlation between cytochrome aa3 levels and streptomycin accumulation was observed. The same correlation was seen when cytochrome aa3 was measured in relation to growth susceptibility. These correlations suggest that cytochrome aa3 is necessary for accumulation of streptomycin by B. subtilis.

Effects of hypoxia and fasting on the cytochrome concentration in intestinal epithelial villous cell mitochondria. Role of changes in the life-span of the cells.

The effects of hypoxia in vivo (40.8 kPa barometric pressure up to 120 h) and fasting on the characteristics of intestinal epithelial villous cell mitochondria and the turnover of epithelial villous cells and mitochondria were studied in rats. Using cells and mitochondria isolated in the isotonic mannitol medium, it was found that 24-h hypoxia or fasting did not alter the mitochondrial cytochrome content, but 48-h hypoxia or fasting led to increases of 70% and 37% in the cytochrome aa3 concentration in the hypoxic and fasting animals respectively. The turnover of intestinal epithelial cells was studied by observing the labelling kinetics of the cells with 3H-thymidine and the turnover of the cell and mitochondrial proteins with (guanido-14C)-arginine or 3H-leucine. The decay in thymidine radioactivity obeyed exponential kinetics from which half-lives of 1.15, 1.31 and 1.53 days were calculated in the control, fasting and hypoxic animals respectively. The half-lives for total cellular protein were 1.31, 1.54 and 1.54 days respectively when calculated from the (guanido-14C)-arginine experiments, or 0.69, 0.75 and 0.99 days when calculated from the leucine experiments. The labelling experiments with (guanido-14C)-arginine indicated that the turnover of mitochondrial proteins in intestinal epithelial cells is the same as that of the cells themselves. Since the turnover of mitochondrial proteins in other tissues is shown to be a relatively slow process, the increase in the cytochrome concentration in the intestinal cells of the hypoxic rats must be due to the longer life of the cells, which allows for the synthesis of larger amounts of the mitochondrial components.

Effect of hypoxia on mitochondrial mass and cytochrome concentrations in rat heart and liver during postnatal development.

Cytochrome concentrations of rat heart and liver mitochondria were measured postnatally from newborn animals to young adults. Mitochondrial cytochrome aa3 concentration increased shortly after birth in both tissues, this concentration being in a newborn animal 0.149 +/- 0.027 nmol/mg protein in liver and 0.333 +/- 0.082 nmol/mg protein in heart. The respective values from a two week old animal were 0.216 +/- 0.031 and 0.583 +/- 0.100. Postnatally cytochromes c+c1 and b increased markedly in the heart, but in the liver of newborn animals the cytochrome content was more close to the adult values. The amount of mitochondrial protein increased after birth, too. In a newborn animal the mitochondrial protein values were 39.7 +/- 3.6 mg/g wet weight in liver and 26.6 +/- 6.5 mg/g wet weight in heart. In adult animals the respective values were 71.5 +/- 4.8 and 80.7 +/- 13.1. The effect of postnatal hypoxia (10% O2, 24 h and 48 h) on liver and heart mitochondrial cytochrome concentration and protein values of newborn animals were investigated. During hypoxia the amount of mitochondrial protein remained about at the level of a newborn animal. The postnatal increase in the mitochondrial cytochrome concentration, which was smaller in the liver than in the heart, was also inhibited by hypoxia.