Erlangen Micro-lightguide Spectrophotometer Research Articles

Effect of nitric oxide synthase (NOS) inhibition on macro- and microcirculation in a model of rat endotoxic shock

Treatment of hemodynamic instability in septic shock often demands the administration of vasopressor agents, although these may have deleterious effects on microcirculatory homeostasis. Inhibition of nitric oxide synthase (NOS) has been suggested as an alternative therapeutic approach, as NO formation may be excessively increased in sepsis. To compare the effects of epinephrine titration, non-selective NOS inhibition by L-NMMA and selective inhibition of inducible NOS (iNOS) by 1400W on hemodynamics and on the regulation of microcirculation in a rat model of endotoxic shock, we intravenously injected endotoxin (LPS) or saline to male Wistar rats and after 2 hours randomized LPS treated rats into four different groups that received either saline, norepinephrine, L-NMMA or 1400W (n = 6 per group). Three hours after LPS administration, rats presented with severe systemic arterial hypotension (64 +/- 3 vs. 115 +/- 4 mmHg, p < 0.001), unresponsiveness to volume treatment, lactate acidosis and a marked increase in plasmatic nitrite and nitrate levels (15 +/- 8 vs. 263 +/- 47 microM, p < 0.001). Measurement of the tissue oxygenation in the ileum mucosal layer by the Erlangen micro-lightguide spectrophotometer (EMPHO) technique demonstrated marked heterogeneity of hemoglobin saturation, with appearance of low oxygenated areas. Norepinephrine, usually stabilizing blood pressure (99 +/- 7 vs. 67 +/- 4 mmHg 60 min after infusion, p < 0.01), increased lactate formation (7.9 +/- 0.2 vs. 3.7 +/- 0.5 mM, p < 0.001) and drastically increased low oxygenated regions in the ileum mucosal layer. L-NMMA similarly increased blood pressure (92 +/- 6 vs. 67 +/- 4 mmHg 60 min after infusion, p < 0.05), but did not enhance lactate acidosis. However, some further deterioration of mucosa oxygenation was again noted. 1400W forwarded stabilization of blood pressure (88 +/- 5 vs. 67 +/- 4 mmHg 60 min after injection, p < 0.05), reduced plasmatic nitrite and nitrate levels similar to L-NMMA, without an aggravation of lactate acidosis. In addition, mucosal oxygenation did not deteriorate in response to this agent. Thereby, we conclude that in a rat model of endotoxic shock selective iNOS inhibitors are superior to non-specific NOS inhibitors and in particular to norepinephrine for the treatment of macro- and microcirculatory abnormalities in experimental septic shock.

Effects of Elevated Intraocular Pressure on Haemoglobin Oxygenation in the Rabbit Optic Nerve Head: A Microendoscopical Study

Intraocular pressure dependent reactions of optic nerve head vasculature and intracapillary haemoglobin oxygenation (HbO2; oxygen saturation) were studied in the center and at the rim of the rabbit optic nerve head (ONH) as well as in the choroid, by a new combination of microendoscopy and simultaneous haemoglobin spectrophotometry.In 13 anesthetized albino rabbits the vasculature and the intracapillary Hb-oxygenation were studied by a microendoscope which was introduced into the eye bulb. Photometric measurements were performed via a beam splitter with the Erlangen micro-lightguide spectrophotometer (EMPHO) from the center of the endoscopic picture. The haemoglobin oxygenation was calculated by real time analysis of the spectral curves. Intraocular pressure was elevated stepwise from 20–80 mmHg.At the rim of the optic nerve head the vascular diameters as well as the intracapillary HbO2-values were stable till an intraocular pressure of 60 mmHg and decrease after IOP elevation to 70 and 80 mmHg. In contrast, in the center of the optic nerve head and in the choroid these parameters decline already from 40–50 mmHg on. At an IOP of 60 mmHg (P<0.01) and 70 mmHg (P<0.05) HbO2is significantly lower in the ONH center than at the rim. In the center and the choroid HbO2is well maintained between 20 and 40 mmHg. After pressure release at the end of the experiment HbO2increased to 94.3±4.6% (rim) and 98.8±1.5% (center) of the initial value at 20 mmHg (difference not significant).By the high spatial resolution of this new optical method we were able to demonstrate that the center of the optic nerve head is more sensitive to changes in intraocular pressure than the optic nerve head rim. Thus, tissue damage after critical haemodynamic and oxygenation parameters seems more probable in the relatively poor perfused center of the ONH than in the overperfused rim.

Cutaneous hemoglobin oxygenation of different free flap donor sites.

Microsurgical skin flaps raised from various body regions show differences in vascular architecture, cutaneous capillary density, and skin perfusion. Therefore, it can be expected that oxygenation of the skin is different at the various free flap donor sites. To determine the cutaneous oxygen supply, intracapillary hemoglobin oxygenation was measured on the donor sites of the radial forearm flap, scapula, latissimus dorsi, rectus abdominis, anterolateral thigh, and osteocutaneous fibula flap on 50 healthy subjects (25 men and 25 women aged 20 to 40 years). Measurements were performed noninvasively with the Erlangen Microlightguide Spectrophotometer (EMPHO) on skin areas of 6 x 8 cm in each region under resting conditions. The hemoglobin oxygenation of the skin in all donor regions varied between a maximum of 43.94 to 58.94 percent in the scapula region and a minimum of 13.89 to 29.45 percent in the lateral calf. High oxygenation values were also found on the skin over the latissimus dorsi muscle (34.56 to 48.45 percent), followed by the distal volar forearm (29.78 to 40.30 percent), whereas paraumbilical skin and the donor sites of the lower extremities were less oxygenated. By using the Wilcoxon test, significant differences were found between all donor regions except for the anterolateral thigh and lateral calf (p = 0.05). There were no gender-specific differences. From these results, it is concluded that, on young healthy subjects, regional oxygen supply on different free flap donor sites varies significantly. This finding must be considered in the interpretation of intracutaneous or transcutaneous PO2 measurements for flap monitoring.

Spectrophotometric in vivo determination of local mitochondrial metabolism by use of a tetrazolium salt

The Erlangen micro-lightguide spectrophotometer (EMPHO) was used to determine local mitochondrial activity. The measurement was based on a commercially available test (WST1, Boehringer Mannheim, Germany). A tetrazolium salt is cleaved to formazan by the `succinate - tetrazolium reductase' system which belongs to the respiratory chain of the mitochondria and is active only in viable cells. of the reagent is placed in a recess at the tip of the lightguide. The tip of the lightguide is then placed at the tissue surface. The reagent diffuses into the cells and is cleaved to formazan. The absorption change is measured and analysed. This method was used in early chick embryos to determine the metabolic activity of the area vasculosa during the first 6 d of normoxic incubation. During this period of time the absorption change measured after 30 min increased from 0.001 units on day 2 to 0.262 units on day 6. This indicates an increase in local metabolic activity which may be due to an increase in metabolic activity of individual cells and/or an increase in the number of cells in the area under investigation. The results show that it is possible to determine local metabolic activity under in vivo conditions. Keywords: micro lightguide spectrophotometer, tetrazolium salt, formazan, mitochondrial activity, local metabolism

Measurements of oxygen saturation in normal human sural nerve by micro-lightguide spectrophotometry and endoscopy.

Micro-lightguide spectrophotometry is a new technique which allows continuous deter-mination of the intracapillary haemoglobin oxygen saturation (HbO2%) from the remission spectra of small volumes of tissue. We have used the Erlangen Micro-lightguide Spectro-photometer (EMPHO II) to measure the HbO2% in human sural nerve. The measurements are made 0.5–1 mm from the surface of the nerve through a small 10mm inscision under local an-aesthetic. The lightguide, which consists of a single 250 μm illuminating and six receiving fi-bres, can be attached to the eyepiece of a small (1.9 mm diameter) endoscope via a beam splitter so the exact site of measurement can be visualised.

Monitoring of flaps by measurement of intracapillary haemoglobin oxygenation with EMPHO 11: experimental and clinical study

Objective: To find out whether measurements of intracapillary haemoglobin oxygenation made with the non-invasive Erlangen microlightguide spectrophotometer (EMPHO II) provided accurate data about the oxygen supply as well as the blood flow to free flaps in rats and humans. Design: Experimental, and prospective open clinical study. Setting: University hospital, Germany. Animals and subjects: Thirty male Wistar rats and 20 patients who underwent microsurgical transplant of free flaps (radial forearm, n = 8; osteocutaneous fibula, n = 7; and myocutaneous from the lateral thigh, n = 5). Interventions: Measurement of the haemoglobin oxygenation of the skin before, during, and after transfer of the flap. Main outcome measures: Reproducibility and mean curves for haemoglobin oxygenation and haemoglobin concentration. Results: All values were reproducible, and there were only slight fluctuations. Uninjured skin in rats (baseline value) oxygenation of the total haemoglobin concentration ranged from 15% to 45% (mean 23%). After the flaps were raised there was a slight increase (to a mean of 37%), probably as a result of reactive hyperaemia. There was a rapid decrease within a few minutes of arterial occlusion, and residual oxygenation of up to 20% after perfusion stopped. The flap was totally deoxygenated after 1 h. Venous occlusion caused a similar pattern and all flaps were deoxygenated by 30 min. In the clinical study ligation of the vascular pedicle caused a massive reduction in values but after anastomosis there was significantly higher haemoglobin oxygenation ( P=0.05) and this continued to increase postoperatively. There were no complications and oxygenation gradually and continuously decreased from the base of the flap to the periphery. Conclusion: The non-invasive EMPHO II provides reliable and easily assessable data about the circulation and supply of oxygen to a transplanted free flap.

Anaemia and Haemoglobin Status: A New Concept and a New Method of Assessment

Traditionally, anaemia has been determined and interpreted by the magnitude and severity of iron deficiency and the impact of intervention strategies. Internationally, it is defined as a state in which the quality and/or quantity of circulating red cells are reduced below a normal level The body employs several mechanisms during the development of anaemia to maintain the oxygen supply to the tissues. Thus, applying any quantitative cut-off point as an indicator for anaemia may lead to misclassification, since haemoglobin concentration does not necessarily reflect the level of tissue oxygen supply. Ideally, an assessment strategy should be able to determine both the degree of haemoglobin oxygenation and the haemoglobin concentration at a tissue level. The Erlangen microlight-guide spectrophotometer is a non-invasive instrument that can assess both capillary blood oxygenation and relative haemoglobin concentration.

Measurements of tissue blood flow by the hydrogen clearance technique (HCT): A comparative study including laser Doppler flowmetry (LDF) and the erlangen micro‐lightguide spectrophotometer (EMPHO)

The purpose of this study was to compare the hydrogen clearance technique (HCT) with two different, well-established techniques, i.e., the laser Doppler flowmetry (LDF) and the Erlangen micro-lightguide spectrophotometer (EMPHO), for tissue blood flow measurements in an experimental setting. For the animal experiments, we chose a rat model for arterial and venous flap thrombosis, using the epigastric groin flap. Forty male dark Aguty rats were included in the study. The animals were divided into eight groups, each with a different vascular thrombotic model. HCT was used to collect 1,467 measurements, and 2,934 graphs were recorded; 27 measurements (54 graphs) had to be discarded due to faulty electrode placements and electronic noise. In 19 of the 27 discarded measurements the cause of failure was moving of the awaking animal with disruption of the decay signal. The LDF and EMPHO measurements were performed continuously during each measuring phase. Simple and multiple linear regression and paired t-tests were used to compare the three techniques. The reproducibility of registered blood flow values in phases 1, 3, 6 and 7 varied between 7.8% and 13.6% which is in about the same range as LDF and EMPHO. We could not find a significant difference between the three techniques in this regard. The correlation coefficient for HCT and LDF was r = 0.89. For HCT and EMPHO we found r = 0.67. Sensitivity and specificity values for HCT were both 1.0 in detecting arterial and venous flap thrombosis; for LDF we found values of 0.89 and 0.92, respectively. Sensitivity and specificity values for EMPHO were 0.92 and 0.95, respectively. After careful evaluation of three different techniques (HCT, LDF, and EMPHO) for measurements of local tissue blood flow we came to the conclusion that HCT must be favoured as a reliable tool for quantitative measurement of local tissue blood flow and early diagnosis of arterial and venous flap thrombosis.

Effect of arterial PCO2 on local HbO2 and relative Hb concentration in the human brain-a study with the Erlangen micro-lightguide spectrophotometer (EMPHO)

Intraoperative investigations of local intracapillary haemoglobin oxygenation have been performed in 5 patients during neurosurgical interventions using the Erlangen micro-lightguide spectrophotometer (EMPHO). As the measurements can be performed in a very short period of time (approximately 60 s) the EMPHO enables rapid monitoring of local haemoglobin oxygenation in the brain. All patients received thiopental anaesthesia and were mechanically ventilated. The local measurements were performed during normoventilation (arterial PCO2-PaCO2 37.8+or-0.7 mm Hg) and hyperventilation (PaCO2 31+or-0.5 mm Hg). Hyperventilation caused a decrease in the mean SO2 from 47.8% to 41.1%. The number of SO2 values <50% increased from 58% to 77.8%. The possible significance of the low local SO2 values is discussed.

Spectrometric measurements in the anterior eye vasculature of the albino rabbit—A study with the EMPHO I

Intracapillary haemoglobin oxygenation (oxygen saturation) and haemoglobin concentration (corresponding with blood volume) were measured in different regions of the albino rabbit anterior eye segment using the Erlangen micro-lightguide spectrophotometer (EMPHO I). The tip of the microlightguide (active diameter 210 microns, cover diameter 0.5 mm) was placed in the vitreous by a goniometric mounting and controlled by a micromanipulator. After stepwise elevation of intraocular pressure (IOP) the haemoglobin oxygenation decreased slightly in the iris at an IOP-level of 60 mmHg, whereas in the three regions of the ciliary processes and in the peripheral choroid haemoglobin oxygenation did not decrease until values of 80 mmHg were applied. In contrast, haemoglobin concentration decreases when the IOP increases in all regions except in the pars plana where the haemoglobin concentration increased at 40 and 60 mmHg. The most pronounced reactive hyperaemia was found in the major ciliary processes. Quite passive changes of haemoglobin concentration were seen in the peripheral choroid. Effective regulatory mechanisms must exist which result in IOP-independent (20-60 mmHg) haemoglobin oxygenation, whereas pronounced changes in intracapillary haemoglobin concentration were observed. The decay of both parameters at IOP 80 mmHg is indicative of an exhaustion of their regulatory capacity. Epinephrine topically applied onto the conjunctival sac or injected into the common carotid artery led to a short-term decrease of haemoglobin concentration and, later, a hyperaemic response. A short-term haemoglobin deoxygenation was found especially in the major ciliary processes.

The Erlangen micro-lightguide spectrophotometer EMPHO I

The Erlangen micro-lightguide spectrophotometer EMPHO I was designed for fast diffuse reflection (remission) spectrophotometry in small tissue volumes. The aim was to construct a compact, modular instrument with a high repetition rate which can be adapted to moving organs, e.g. the beating heart in situ, by the use of highly flexible micro-lightguides. Focusing problems, which cannot be solved when conventional optical devices such as microscopes are used in moving tissues, become negligible. A bandpass interference filter disk, which is rotated by a motor serves as a monochromating unit. One diffuse reflection spectrum in a selected wavelength domain is recorded during each revolution of the motor. Special filter disks, with spectral ranges of 400-520, 500-630, 600-1200 nm can be used for different tasks. The monochromated light is transmitted by means of a flexible fluid-lightguide to a photomultiplier tube. The electrical signal, which is proportional to the light intensity is recorded by an IBM-compatible AT. An analogue to digital converter has been developed for the AD conversion.