Characterization of neutralization epitopes of simian immunodeficiency virus (SIV) recognized by rhesus monoclonal antibodies derived from monkeys infected with an attenuated SIV strain.

A major limitation in the simian immunodeficiency virus (SIV) system has been the lack of reagents with which to identify the antigenic determinants that are responsible for eliciting neutralizing antibody responses in macaques infected with attenuated SIV. Most of our information on SIV neutralization determinants has come from studies with murine monoclonal antibodies (MAbs) produced in response to purified or recombinant SIV envelope proteins or intact SIV-infected cells for relatively short periods of time. While these studies provide some basic information on the potential immunogenic determinants of SIV envelope proteins, it is unclear whether these murine MAbs identify epitopes relevant to antibody responses elicited in monkeys during infection with either wild-type or attenuated SIV strains. To accomplish maximum biological relevance, we developed a reliable method for the production of rhesus monoclonal antibodies. In the present study, we report on the production and characterization of a unique panel of monoclonal antibodies derived from four individual monkeys inoculated with SIV/17E-CL as an attenuated virus strain at a time when protective immunity from pathogenic challenge was evident. Results from these studies identified at least nine binding domains on the surface envelope glycoprotein; these included linear determinants in the V1, V2, cysteine loop (analogous to the V3 loop in human immunodeficiency virus type 1), and C5 regions, as well as conformational epitopes represented by antibodies that bind the C-terminal half of gp120 and those sensitive to defined mutations in the V4 region. More importantly, three groups of antibodies that recognize closely related, conformational epitopes exhibited potent neutralizing activity against the vaccine strain. Identification of the epitopes recognized by these neutralizing antibodies will provide insight into the antigenic determinants responsible for eliciting neutralizing antibodies in vivo that can be used in the design of effective vaccine strategies.

Production and characterization of SIV envelope-specific rhesus monoclonal antibodies from a macaque asymptomatically infected with a live SIV vaccine.

Five rhesus monoclonal antibodies (RhMAbs) were produced by rhesus EBV transformation of peripheral blood B cells from a rhesus macaque that had been asymptomatically infected with an attenuated, macrophage-tropic SIV strain, 17E-Cl. These MAbs recognized conformation-dependent epitopes on SIV gp120 and could not be mapped using synthetic peptides. All five RhMAbs were able to neutralize the vaccine strain and a heterologous isolate, SIV/DeltaB670. The RhMAbs did not cross-react with HIV-2; by contrast, four human MAbs derived from an HIV-2-infected person were broadly cross-reactive with both SIV and HIV-2 gp120s. Cross-competition analysis indicated that the five RhMAbs could be placed in two groups recognizing two nonoverlapping epitopes; while the HMAbs were placed in two additional competition groups. Binding of the three group I RhMAbs (1.7F, 3.11B, and 1.10A) as well as HMAb 17A was shown to be sensitive to specific amino acid alterations in V4 occurring in natural env variants. The results of this study demonstrate that RhEBV transformation provides a means to probe rhesus antibody responses to SIV infection at the monoclonal level. RhMAbs will allow structural and functional studies of envelope glycoprotein determinants that elicit protective immune responses against SIV.

Effects of a single saturation dive on lung function and exercise performance.

We studied the effects of an experimental saturation dive to 360 and 450 m in a simulation chamber on spirometric lung function, diffusing capacity, pulmonary compliance, and exercise performance in eight professional divers (age 22-40 years). To assess intraindividual variability, all parameters were measured on 2 days before and on 2 consecutive days immediately after the dive. For the group as a whole there was a significant increase in vital capacity and alveolar volume, and a decrease in Krogh factor and specific compliance (P < 0.01). These changes were reduced on the 2nd day after the dive. All subjects showed lowered exercise performance after the dive. Arterial pressure of oxygen and ventilation during exercise increased (P < 0.01), whereas arterial pressure of carbon dioxide, oxygen uptake, and anaerobic threshold decreased (P < 0.01). Exercise parameters showed only a slight trend towards pre-dive values on the 2nd day after a dive. The individual analysis revealed that after the dive two subjects showed a marked decrease in diffusing capacity and a more than average decrease in Krogh factor (TLCO/VA). One of them had signs of mild decompression sickness and the other, signs of pre-existing obstructive airways disease. Our data are compatible with the hypothesis that the effects of a single deep saturation dive on pulmonary function and exercise performance are the results of counteracting mechanisms. We suggest that lung volumes increase due to the enhanced work of breathing during a deep saturation dive and that these changes could mask an impairment in gas exchange. Furthermore, a saturation dive can induce an apparent deterioration of pulmonary function.

A simple emergency underwater breathing aid for helicopter escape.

Experiments were undertaken to determine whether a simple rebreathing system, termed "Air Pocket" (AP), could, when integrated into an immersion dry suit, extend the underwater survival time of individuals when compared with their maximum breath hold time (BHTmax). Eight naive healthy male subjects undertook a series of resting submersions and simulated simple helicopter underwater escapes in water 25 degrees C and 10 degrees C. During the submersions the subjects breath-held maximally and then rebreathed using an otherwise empty AP. the BHTmax times of subjects and the total time they could remain underwater (RBT) were recorded. The results showed that the ability to rebreathe following a BHTmax extended the time all subjects could remain submerged, resting or exercising, in cold water by a factor of at least two. The average BHTmax during simulated helicopter underwater escapes in the cold water was 17.2 s. It is concluded that the ability of subjects to rebreathe immediately following maximum breath holding extends the time they can remain submerged in cold water to as much as 60 s. Further, if used unprimed, a simple rebreathing system will not introduce any additional dangers such as a pulmonary over-pressure accident.

Human initial responses to immersion in cold water at three temperatures and after hyperventilation.

The present investigation was designed to examine the influence of water temperature and prior hyperventilation on some of the potentially hazardous responses evoked by immersion in cold water. Eight naked subjects performed headout immersions of 2-min duration into stirred water at 5, 10, and 15 degrees C and at 10 degrees C after 1 min of voluntary hyperventilation. Analysis of the respiratory and cardiac data collected during consecutive 10-s periods showed that, at the 0.18-m/s rate of immersion employed, differences between the variables recorded on immersion in water at 5 and 10 degrees C were due to the duration of the responses evoked rather than their magnitude during the first 20 s. The exception to this was the tidal volume of subjects, which was higher on immersion in water at 15 degrees C than at 5 or 10 degrees C. The results suggested that the respiratory drive evoked during the first seconds of immersion was more closely reflected in the rate rather than the depth of breathing at this time. Hyperventilation before immersion in water at 10 degrees C did not attenuate the respiratory responses seen on immersion. It is concluded that, during the first critical seconds of immersion, the initial responses evoked by immersion in water at 10 degrees C can represent as great a threat as those in water at 5 degrees C; also, in water at 10 degrees C, the respiratory component of this threat is not influenced by the biochemical alterations associated with prior hyperventilation.

The effect of clothing on the initial responses to cold water immersion in man.

The protection provided by three clothing assemblies against the cold shock response was investigated. Nine healthy male volunteers each undertook three two minute head-out immersions into stirred water at 10 degrees C. The subjects wore a different clothing assembly for each immersion, these were: a) Swimming trunks only; b) Conventional clothing (equivalent to RN No 8s); c) Conventional clothing plus windproof/shower-proof clothing (RN foul-weather clothing Mk III). The cardiac, ventilatory and thermal responses of the subjects were examined before and during the immersions. No significant differences were found between the magnitude of the responses recorded on immersion when conventional clothing or foul-weather clothing were worn. Mean skin temperature was lower (P less than 0.05) and respiratory frequency and minute ventilation were higher (P less than 0.05) on immersion in swimming trunks compared to the other two conditions. It is concluded that when policies for the use of immersion protective clothing are being formulated, consideration should be given to all of the potentially hazardous responses associated with cold water immersion.

The thermal performance of partial coverage wet suits.

A wet-suit worn external to normal clothing and covering the trunk and arms only has been assessed as a method for providing short-term immersion protection for helicopter passengers in offshore oil field operations. Manikin measurements of effective insulation in water give a mean figure of 0.54 togs for the areas covered by the suit and 0.09 togs for uncovered areas. These figures were used to obtain model predictions of survival time for 'thin' and 'average' men which suggest that the suit can give adequate protection for 1 h at 5 degrees C subject to care in fitting. Direct measurements of heat flux have demonstrated the presence of water flushing beneath the suit and the potentially serious loss of insulation that can result.

Nitrogen-oxygen saturation therapy in serious cases of compressed-air decompression sickness.

Decompression sickness and arterial air embolism which follow exposure to raised environmental pressures of compressed air are usually adequately treated by accepted recompression procedures of relatively short durations. With serious cases, however, conventional treatment may not allow sufficient time at depth for the complete resolution of manifestations because of the need to avoid pulmonary oxygen toxicity which is associated with a prolonged period of breathing compressed air. Treatment by nitrogen-oxygen saturation at a pressure equivalent of 30 m (100 ft) sea water is proposed. Based upon the success of three refractory cases treated by this procedure, recommendation are made for the conversion of standard compressed-air chambers into an emergency saturation mode for therapy.

Mechanisms underlying spinal cord damage in decompression sickness.

Decompression sickness, which damaged the spinal cord, was produced in anesthetized dogs using a compression chamber. Cerebrospinal fluid pressure and several intravascular and intracardiac pressures were monitored during the course of the simulated dives. Manometric responses to forcible lung inflation and abdominal compression were measured both predive and postdive after signs of spinal cord damage were evident. Cinevenography of the epidural vertebral venous system was performed both predive and postdive. Histopathologic studies of the brains and cords of both predive and postdive. Histopathologic studies of the brains and cords of paretic animals were carried out. The results indicate that the epidural vertebral venous system becomes obstructed during spinal cord damaging decompression sickness and strongly suggests that spinal cord infarction in decompression sickness is caused by obstruction of cord venous drainage at the level of the epidural vertebral venous system.