Venous gas embolism as a predictive tool for improving CNS decompression safety.

A key process in the pathophysiological steps leading to decompression sickness (DCS) is the formation of inert gas bubbles. The adverse effects of decompression are still not fully understood, but it seems reasonable to suggest that the formation of venous gas emboli (VGE) and their effects on the endothelium may be the central mechanism leading to central nervous system (CNS) damage. Hence, VGE might also have impact on the long-term health effects of diving. In the present review, we highlight the findings from our laboratory related to the hypothesis that VGE formation is the main mechanism behind serious decompression injuries. In recent studies, we have determined the impact of VGE on endothelial function in both laboratory animals and in humans. We observed that the damage to the endothelium due to VGE was dose dependent, and that the amount of VGE can be affected both by aerobic exercise and exogenous nitric oxide (NO) intervention prior to a dive. We observed that NO reduced VGE during decompression, and pharmacological blocking of NO production increased VGE formation following a dive. The importance of micro-nuclei for the formation of VGE and how it can be possible to manipulate the formation of VGE are discussed together with the effects of VGE on the organism. In the last part of the review we introduce our thoughts for the future, and how the enigma of DCS should be approached.

The effect of endurance training on the rate of nitrogen elimination in women.

INTRODUCTION: The rate of nitrogen elimination may be an important factor in evaluating the risk of DCS following dives. The present study determined the reproducibility of a method for evaluating nitrogen elimination (series I), and the effect of chronic training on the nitrogen elimination in healthy young women (series II). METHODS: Nitrogen elimination was determined with subjects wearing an AGA full-face mask breathing pure oxygen. To evaluate the reproducibility of the method for nitrogen elimination, three tests were performed in six subjects in series I. Nitrogen elimination in series II was measured before and after the training period. The training protocol (series II) consisted of interval training, three times per week for eight weeks. Four repeated intervals alternated between four minutes at 90-95% of maximum heart rate and three minutes at 50-60%. RESULTS: There was no significant difference between the three repeated tests. Interval training for eight weeks increased maximum oxygen uptake by 22.1%. Endurance training did not influence the total nitrogen elimination at rest. CONCLUSION: The method for evaluating nitrogen elimination at rest was found to be reproducible. Improved aerobic capacity does not increase the rate of nitrogen elimination at rest.

Evaluation of cerebral gas retention and oedema formation in decompressed rats by using a simple gravimetric method.

OBJECTIVE: The objective of the study is twofold: first, to develop a specific gravity method for distinguishing between bubbles and oedema following decompression, and, second, to evaluate the extent to which the change in specific gravity is due to retained gas in cerebral tissue. METHODS: A brombenzene/kerosene gradient column was used to measure changes in brain specific gravity at 100 and 300 kPa, respectively. This study was performed on 23 rats. Non-exposed rats constituted the control group A (n=6). The exposed animals were divided into two groups according to the number of bubbles they developed upon decompression; group B (bubble grade 0-2, n=9) and group C (bubble grade 3-5, n=8). RESULTS: Cerebral gas retention was determined by increasing the pressure on the gradient column from 100 to 300kPa. Median specific gravity of the brain at 300kPa bar was significantly higher compared to 100 kPa for the decompressed groups B (p= 0.018) and C (p=0.012), thus implying gas retention. The cerebral gas volume was significantly higher for rats with a high bubble score compared to rats with a low bubble score (p=0.043). However, the major contribution to the change in specific gravity was due to oedema formation. CONCLUSION: The brombenzene/kerosene gradient column was found to be a sensitive method for distinguishing between gas retention and oedema formation in decompressed animals. There was a higher gas retention in rats with a high bubble score compared to rats with a low bubble score. The major contribution to the change in specific gravity in decompressed animals is due to oedema formation.

Hyperbaric oxygen and neutrophil accumulation/tissue damage during permanent focal cerebral ischaemia in rats.

The use of hyperbaric oxygenation (HBO) for the treatment of severe brain ischaemia remains controversial. The HBO may interfere with destructive neutrophil (PMN) infiltration following ischaemia/reperfusion. The effects of HBO on PMN accumulation and the area of ischaemic tissue damage were investigated in rats having permanent focal ischaemia (4 h). The right middle cerebral arteries of a group of Wistar rats were permanently occluded. The rats were then randomly divided into those ( n=7) to be treated with HBO at 2 atm for 230 min and those ( n=8) to breathe air at atmospheric pressure for an equivalent period. The HBO had no effect on permanent ischaemia, as there was no significant difference in the area of ischaemic tissue damage between HBO-treated [mean (SD)] [331 (88) mm(3)] and non-treated animals [322 (111) mm(3)]. Moreover, the increase in myeloperoxidase [5.4 (4.1) compared to 2.4 (1.2) pg x g(-1) wet weight of brain] was not significantly different. The results indicate that HBO did not reduce tissue damage during 4 h of permanent focal ischaemia.

The effect of air bubbles on rabbit blood brain barrier.

Several investigators have claimed that the blood brain barrier (BBB) may be broken by circulating bubbles, resulting in brain tissue edema. The aim of this study was to examine the effect of air bubbles on the permeability of BBB. Three groups of 6 rabbits were infused an isoosmotic solution of NaCl w/macrodex and 1% Tween. The solution was saturated with air bubbles and infused at rates of 50-100 ml hr(-1), a total of 1.6, 3.3, or 6.6 ml in each group, respectively. Two groups, each consisting of 6 rabbits, served as controls; one was infused by a degassed isoosmotic NaCl solution and one was sham-operated. All animals were left for 30 min before they were sacrificed. Specific gravity of brain tissue samples was determined using a brombenzene/kerosene gradient column, where a decrease in specific gravity indicates local brain edema. Specific gravity was significantly lower for left (P = 0.037) and right (P = 0.012) hemisphere white matter and left (P = 0.0015) and right (P = 0.002) hemisphere gray matter for the bubble-infused animals compared to the sham-operated ones. Infusion of degassed NaCl solution alone affected white left (P= 0.011) and right (P= 0.013), but not gray matter of both hemispheres. We speculate that insufficient degassing of the fluid may cause the effect of NaCl solution on the BBB of the white matter, indicating that the vessels of the white matter are more sensitive to gas bubbles than gray matter. Increasing the number of infused bubbles had no further impact on the development of cerebral edema, indicating that a threshold value was reached already at the lowest concentration of bubbles.

Lack of effect of anti-C5a monoclonal antibody on endothelial injury by gas bubbles in the rabbit after decompression.

Previous studies have shown that gas bubbles activate the complement system in vitro, generating C5a. The effect of anti-C5a monoclonal antibody 4B1C11 in preventing endothelial damage caused by decompression in the pulmonary artery of the rabbit was examined. The endothelial response was measured using tension measurements in the blood vessel wall. The mean bubble count for all rabbits (n = 24) was 4.2+/-3.1 bubbles x cm(-2), and ranged from 0 to 15 bubbles x cm(-2). Animals with many bubbles showed significantly more vascular damage than those with fewer bubbles. Anti-C5a monoclonal antibody could not prevent endothelial damage than that occurred after exposure to this level of gas bubbles. The maximum number of gas bubbles present is important for the endothelial damage. We speculate that the endothelial damage observed was mainly mechanical. A possible beneficial effect of anti-C5a antibody can thus be masked at a high degree of bubble generation. This study, together with a previous paper, demonstrates that gas bubbles cause endothelial damage from decompression both in the pig and in the rabbit.

Effect of anti-C5a antibody on blood-lung and blood-brain barrier in rabbits after decompression.

The complement activation product C5a may be an important mediator of tissue injury after decompression stress. This study investigated whether the administration of anti-C5a antibody may reduce changes after decompression in the lung and in the brain. Two groups of rabbits were used; one receiving anti-C5a monoclonal antibody (n = 7) and the other receiving a sham antibody as control (n = 7) before pressure exposure. Five rabbits (4 in the anti-C5a group and 1 in the control group) died during the 2-h observation period postdive due to massive bubbling. Polymorphonuclear leukocyte (PMN) infiltration of lung tissue and pulmonary edema was observed, but this accumulation was unaffected by anti-C5a pretreatment. However, a significant positive correlation was observed between PMN accumulation and survival time postdive. Brain-specific gravity was lower for the group treated with anti-C5a antibody compared to the control group. Further, it was lower for those rabbits that died early compared to the ones that survived the 2-h period. This study was unable to prove a protective effect on the blood-brain and blood-lung barrier by injecting anti-C5a antibody. A possible beneficial effect of anti-C5a antibody may be masked by the mechanical damage caused by the gas bubbles.

Complement activation in divers after repeated air/heliox dives and its possible relevance to DCS.

Plasma levels of the anaphylatoxin C5a were measured in 19 divers performing repeated air dives. Blood samples were collected immediately before the first dive and 2 h after the first and the second or third dive. Serum obtained at the same times was subjected to complement activation in vitro by air bubbles. Six divers developed symptoms of decompression sickness (DCS). Most intravascular bubbles were observed in divers with the lowest plasma levels of C5a. Postdive plasma levels of C5a did not increase compared with predive levels, nor were postdive levels significantly different after two or three dives compared with the first dive. Repeated dives did not influence the amounts of C5a generated in vitro. Neither plasma levels of C5a nor C5a generated in vitro were significantly different in divers who experienced symptoms of DCS vs. divers without symptoms of DCS. We conclude that plasma level of C5a and measurement of C5a generation in vitro cannot be used to predict DCS.

Variability over time of complement activation induced by air bubbles in human and rabbit sera.

Complement activation induced by air bubbles in rabbit and human sera was studied by measuring the generation of anaphylatoxin des-Arg-C5a. des-Arg-C5a was quantified by sandwich enzyme-linked immunosorbent assays based on neoepitope-specific anti-des-Arg-C5a monoclonal antibodies. Air bubbles were continuously introduced to serum via a calibrated microflowmeter, and the serum was incubated at 37 degrees C for 30 min. Air bubbles clearly generated increased amounts of des-Arg-C5a compared with corresponding levels in control serum, and a dose-dependent effect was also noted. Strong positive correlations between des-Arg-C5a concentrations in control sera and sera incubated with air bubbles at a flow of 0.5 ml/min were found. To study variation over time, serum was obtained at regular intervals from six rabbits and from six healthy humans during 66- and 196-day periods, respectively. A pronounced intraindividual variability over time was thus observed. The reason for the large variability is at present unknown. We conclude that the sensitivity of complement to activation by air bubbles is not an inherent, static feature of the complement system of an individual. Therefore single-point analysis of complement activation by air bubbles appears to be an inappropriate parameter by which to differentiate a "sensitive" or "insensitive" complement system between individuals.