Effects of high and low barometric pressures on susceptibility and resistance to infectio

Bond, S.

Gillmore, J. D.

Gordon, F. B.

Zebovitz, E.

English

NASA Technical Reports Server

Former designations: L-97, 464 
R-21-010-010 
Quarterly Status Report No. 20 to 
National Aeronautics and Space Administration 
1 April - 30 June, 1970 
"Effects of High and Low Barometric Pressures on Susceptibility and 
Resistance to 
Francis B. Gordon, James D. GBBBmore, 
Eugene Zebovitz and Sheila B o d  
Department of Microbiohgy 
Naval Medical Research Institute 
Bethesda, Marylland 20014 
https://ntrs.nasa.gov/search.jsp?R=19700031577 2020-03-11T22:44:08+00:00Z
Abstract 
&xperiments have continued directed toward a solution. of the problem 
of rnain@&&ng the body temperature of small e q e r h e n t d  
helium atmosphere, and toward an mce  of a lowered 
body temperature in susceptibility to ion. Maintenance 
of the gas phase (1% 02 in Heb 500 psig) at 35'6 enabled mice to preserve 
a body temperature within 2-3 C of normal. 
toward an adverse effect of a normoxic O2-He atmosphere, at normal OF 
increased barometric (95 psig) pressures, even when the temperature of 
the gaa phase is raised to a point (3OOC) that enables mice to maintain 
a normal, or nearly normal, body temperature. 
Additional experiments point 
An experiment utilizing 11% 0 2  kt N2 at one atmosphere for exposure 
of mice following inoculation with Coxsackie virus has confirmed the earlier 
observation of a more severe infection due to hypoxia 8s provided by an 
atmosphere of air at 7 . 3  psia. 
An -- in vitro cell culture system has been set up by which the trans- 
forming effect of SV40 virus will be studied under psrabarosis. Although 
definitive tests on the incidence of transformation in altered atmospheres 
a re  not yet complete, an increased resistance of transformed cells to O2 
toxicity has been observed. 
An additional experiment has been completed, designed to determine the 
effect of an environmental temperature of 3 5 ' ~  on rectal temperatures of 
mice exposed to a 1% O2 in He atmosphere at 500 psig. 
compared to a similar experiment in which the environmental temperature 
was 3OoC9 as summarized in Fig, 3, QSR No. 99 (1 January - 80 March, 
1970). With the chamber elevated to 35OC, rectal temperatures of the 
lightly restrained mice became fairly well stabilized between 35.5 and 
36,5OC after a sharp initial rise during compression when the internal 
chamber temperature increased to 57OC (Fig. 1). 
posure interval in 1% O2 - He atmosphere at 500 psig (34 atm), decompmssion 
was initiated. 
decompression to a condition of ambient pressure with normal p02 in He, 
even though the environmental chamber temperature was decreased to slightly 
below 35OC during the 80 minute decompression interval. 
This is to be 
After the four hour ex- 
Rectal temperatures rose steadily to original values during 
The results depicted in Fig. 1 may be compared with the rectal 
and subcutaneous temperatures recorded in mice mainkxi ned in a 2.8% 
O2 in He atmosphere at 95 psig as presented in Fig. 5 of QSR No, 14 
(1 October - 31 December, 1968). 
perature of 33-35OC in the 1968 experiments was able to maink& mouse 
subcutaneous and rectal temperatures at approximately 3%"C, in the present 
experiment utilizing 1% O2 in He, atmosphere at 500 psig, the rectal tem- 
Although a compensating chamber tem- 
res stabilized only about 1°C lower when the endl rom 
ature was maintained at 35oC. These ~ ~ s e r v a t i o ~ s  w 
that elevation of the envlromentsl temperature of mice to 35%, under rn 
essentially normoxic p02 in He environment at 508 psig, is &Be to 
compensate for a large fraction of the body ~ e m ~ e r ~ ~ r e  loss due to  the 
compressed helium atmo e in the rang from 95-580 psig. 
heat Boss, and will increase the possibility of identi kmig variayones 
parabark conditions . 
eompensation will enim 
C2LUShg ~l'hW~tiQl'l~ in s ~ s ~ e ~ ~ ~ ~ i ~  fi;O G % ~ f X i I l l & X d  %LKff3CfhhX'W h miCX? UIIdC% 
2. 
Previous experiments, PR8-30, $1, a d  32i9 reported b QSR No. 19, 
gave evidence that chilling of mice by maintaining them in air at B5O6 results 
in a greater mor 
of the factors responsible for the increased mortality observed h- similar 
groups maintained in 
as described in QSR No. 12, expm5nxmts PR8-4 m d  5. While complete 
ema viral hfect io~.  ChiUihg may be one 
erbarie (95 paig) - normoxk 0 2  - He gas mixtures 
2 
%sollation of this factor was not achieved, the evidence derived indicated that 
a 5OC elevation of the ambient 20% 0 2  - He environment would decrease 
the observed mortality to a level equal to o r  slightly less than that observed 
in %I 25'6 ambient air atmosphere, Since a 10°C elevation of chamber en- 
vironment to 3506 has enabled mice to compensate for body heat loss when 
maintained in a normoxic (2e8% 0 2  - He) environment at 95 psig, QSR No. 14 
(I October - 31 December, 1968) the outcome of influenza viral infection 
of mice kept at 95 psig in O2 - He should not be influenced by loss of body 
heat per se if the ambient temperature is kept at 35OC. 
.. 
The fofollowing experiments were designed to test the effect of 8 
normoxic O2 - He mi-xture at 95 psig on influenza1 pneumonia in mice 
when loss of body heat was prevented by raising the temperature of the 
gas phase. Controls were maintained at ambient temperature (25OC) in 
either at 20% O2 - He o r  air atmospbere. 
Groups of 20-30 mice each, following aerosol challenge with influema 
virus, were held in the parabaric atmospheres described below for deter- 
mination of mortality rates. 
were also determined. For PR8-35, the three experimental groups consisted 
of 20 mice each and were exposed post-challenge to a 2.8% 0 
psig, atmosphere at 3OoC with one control group being expose$ to a 20% 0 2  - 
He zitmosphere at 25OC and ambient pressure and the second control group 
to ambient air at 25'C. In this experiment (not illustrated) the challenge 
aerosol from a newly prepared purified pool of PR-8 virus caused over 
95% mortality in all three experimental. groups and little difference could 
be discerned under the various environmental conditions 
In one instance, infectious titers of lung pools 
- He, 95 
In PR8-37, the experimental conditions consisted of e-xposure of 20 
mice to a 2.8% 02-He atmosphere at 35O6, an environment enabling the 
experiments1 mice to maintain inearly normal body temperatures. 
earlier and more severe 695%) mortality occurred in the hyperbaric mice 
than in the control group (%5%), ax shown h Figo 2. Similarlyy, in PR8-38 
(Fig- 31, the hyperbaric group (I95 psig, 2.8% O2 - He9 maintained in a 
35'C ewiroment ,  again demonstcated more than s 25% greater mortality 
than occurred in the corresponding control group held In an ambient 20% 
0 2  - He mixture at 25OC, even though this Batter atmosphere stresses the 
mice by causing an approximate 5OC reduction in intrarecbl or subcutaneous 
temperature as described in Fig. 3, QSR No. 114 (Il October - 31 December, 
1968). 
An 
The parabaric conditions for PR8-39 for both hyperbaric-normoxic 
0 2  - He and ambient O2 - He atmospheres were as described for the 
p rev i~us  experiment, BR8-38. In this instance, the parabaric and 
mortality groups were increased to 25 mice each with an aqlditional eight in 
each group similarly exposed and maintained for sacrifice and preparation 
of 10% lung pools from four mice each at the fifth and seventh day post- 
challenge exposure interval. Earlier deaths and a greater mortality rate 
(84%) was again observed (Fig. 4) in the 35'6 temperature compensated 
hyperbaric group, when compared with the 20% O2 - He group (at 25OC) 
in which the mortality rate was 56%. Infectivity (EID50) titrations per- 
formed on the fifth and seventh day lung pool preparations, however, again 
showed only minimal differences between test and control mice, as calcu- 
lated by Karbers method and shown in Table 1. 
for lung consolidation were also hund to be almost identical. 
The gross pathology scores 
Table 1. Experiment PR8-39. Effect of parabaric conditions on mouse lung infection 
following aerosol challenge with PR-8 influenza virus. 
Day of NO. Infectivity 
b Environment sacrifice of Av.mouse Av.lung Lung titer of lung post-challenge mice wt. (g) wt. (g) consoLa (E ID50 ; Karber) 
107 .3 Group A 5 4 15.6 0.18 0.2 
2.8% O2 in (0.3,O. 3,O. 1,O. 1) 
He 
95 psig 7 4 12.9 0.23 2.1 lo5- 
35OC (3.0,2.5,2.0,1.0) 
Group B 5 4 15.9 0.20 0.15 loa= 
20% O2 in He (0.39 0.2,o. 1,o. 0) 
1 &trn a 4 14.0 0.24 2.1 io5* 
25°C (3.0,3.0,2.0,1.5) 
5 4 15.0 0.20 0.2 
7.1 
10 
(0.3,0.3,0.2,0.l) 
7 4 13.4 0.28 2.8 lo5* 
(3.5 3.0 2.5,2.0) 
~ ~~ ~~ __ 
a Arbitrary 0-5 scoring for degree of lung invalvernent: average and (individual) scores. 
Expressed at 50% egg infective dose per 0.1 ml inoculum (Kgrber) 
Aerosol exposure: 25 minn. using E:450 dilution Pool A (PR8), 0,25 ml/min. €3. H. 88% 
4 
One additional experiment, PR8-40, was performed to determine again 
whether the reported mortality increase in mice exposed to ambient 20% 
02 - He atmosphere at 25OC as compared to ambient air at the same tem- 
perature could be demonstrated. The experimental groups in this instance 
consisted of 30 mice each and again a 25% greater mortality was observed 
in the O2 - He mixture as compared with the air control group, as presented 
in Fig. 5. 
The data from these and earlier experiments indicate that in normobaric, 
normoxic helium (20% O2 - He) atmospheres at 25OC, the stress from loss 
of body heat in this artificial environment contributed significantly to a de- 
crease in resistance to influenzal infection in the mouse. The loss of body 
heat can be prevented by elevation of the environmental chamber tempera- 
ture to a sufficient level, i. e., 30°C. Elevation of environmental tem- 
perature to 35OC in a hyperbaric normoxic (2.8% 0 2  - He) environment, 
wU1 maintain mouse intrarectal o r  subcutaneous temperatures close to 
normal limits. But even with body temperatures approximately normal, 
such parabaric conditions still result in a greater mortality in influemd 
mice than in appropriate control groups, It is, therefore, reasonable to 
postulate that some other factor@) in the hyperbaric, normoxic - He environ- 
ment accounts for the increased mortality observed repeatedly in influenza 
virus infected mice kept under this condition. 
3. Effect of parabarosis on mouse lung influemal hfectiosn; microscopic study. 
In a single experiment PR8-34, the experimental conditions were designed 
to evaluate the influenzal process by measuring either altered mortality rates 
or  changes in the lung pathologic process by routine light microscopy section 
supplemented with electron microscopy sections if indicaded. The parabaric 
exposure was limited to 7'7% O2 at one atmosphere fol8owhg aerosol 
challenge with either PR-8 influema virus diluted in 0.5% bovine plasma a9- 
bumin in phosphate buffered saline, OF to the dihent alone ( ~ ~ n t r d s ) ~  
resultant mortality curves are depicted in Fig. 6 and may be observed to 
confirm previously reported experiments, PR8-111, 13, and 17, where in- 
creased mortalities were always seen ]in mice exposed to hyperoxic environ- 
menta. Conventional H & E staining has been performed on representative 
~sc t ions  prepared for light microscopy and their interpretation utilized to 
select the more promising areas to prepare for ultramicroscopic sectioning 
and interpretation to be reported later. 
%he
5 
4. Preparation of influema virus pool. 
Experiments PR8-1 through 33 depleted the original PR-8 pool (A). 
An additional partially purified high titer PR-8 pool B has been prepared 
and stored as  aliquots for future experiments. Titrations have been per- 
formed to allow the selection of doses of aerosolized virus to induce greater 
o r  less severe infections, depending upon the objective of the experiments. 
Titrations have also been performed ta provide positive controls in the usual 
ED50 and hemagglutination tests. 
5. Experiments with Coxsackie virus. 
Experiment Coxsackie #12 has been completed with post-challenge 
parabaric exposure limited to h-yperoxia (77% 02, one atmosphere), 
hypoxia (11% 02, one atmosphere), in contrast to the hypoxic envimnment 
of air at 7.3 psia utilized in Coxsackie #lo, and to an ambient air control 
group. The results confirm the results of earlier experiments. 
Groups of 20 mice each were challenged i.p. as before with 0.25 ml 
of k500 dilution of pool A (equivalent to 2500 LD50 suckling mouse doses). 
Four mice from each group were sacrificed on day 3 post-challenge and 
three mice in each group on days 5 and 7 for preparation of 10% pan- 
creatic pool suspensions. Ths remaining 10 mice in each group were 
maintained under the parabaric atmospheres €or observations on mortality. 
A definite increase in pancreatic tissue viral titer, as determined by 
standard plaquing procedures using LLC-MK2, rhesus monkey kidney cells, 
was observed in animals exposed to the decreased pO2 at ambient pressure. 
As  presented in Table 2, B plaque titer s f  8800 x JlO'/gm pancreatic 
tissue was observed in hypoxic animals on day 3 post-challenge a s  com- 
pared to B l O O  x 103/gm for the hyperoxic group and 570 x 103/gm for am- 
bient air controls. 
sacrifice on day 5 post-challenge, the values for the hypoxic animals 
were still elevated (48 x 103/gm9 over either hyperoxic (5e 8 x BO /gm), or  
air in the ~ ~ n t r o l i  animals (3.3 x 103/gm). Plaque assays were not 
performed on the seventh day post-challenge pancreatic pools because 
of the abrupt decrease observed at day 5. No deaths occurred in any 
of the mouse groups during the 211 days of the experiment. 
Although the plaque titers were sharply reduced at 
3 
6. Effect of increased oxygen tensions on latent virus infections (by 
E. Zebovitz and S. Bond) 
The microbial fl.cPra of the hu:mam body is composed not S D I ~  of 
bacterial species, but of a number of viruses which are present as latent 
6 
or  masked Ln&xtio~.s within the cells of their host. Nosmd.ly, the host 
mima% does not exhibit disease symptoms aid :~ppsa:rs healthy, 
consequmm, the presence of the virus usually 1s unsuspected. Certain 
viruses which can establish latent infections in man are  oncogenic, i. e., 
they produce tumors in various a&nd species md  transform cells in 
tissue culture. 
Sable 2. (Coxsackie E-q. #B) Effect of p a r a b a r d s  OB susceptibility of adult 
mice to Coxsackie €3-1 ~ E ~ E X A ~ Q ~ ;  parabaric exposure followed challkeDge. 
.Observation on sacrificed mice 
Group and Viral titer 01 
(1 atm pressure) Mortality mcrificea pancreatic p o d  PFU/g x IO3 
environment Day of wt. (g) of paracreatic pool 
O/lOb 3 0.93 110 0 
0.63 5.8 
1.45 NDC 
5 0.32 48 
(hypoxia) 7 0.45 ND 
musual environmental conditions will  trigger a normally Bata&, virus b express its 
oncogenic capacity. From what is known about oxygen toxicity and other effects of 
abnormal oxygen tensions, it is reasonable to postdate that enviromental changes 
of this type can activate latent oncogenic viruses. 
A model system has been devised to study the effect of oxygen concentration 
The vims selected was SV$o on the trans€orming activity of an oncogenic virus. 
(Simian vacuolating) since i d  has the highest transforming activity of any known 
oncogenic virus. 
inbred strain of mice, the AL/N strainn, which has a low susceptibility to the 
development of spontaneous tumors. 
An epithelial line of cells was derived in this laboratory from an 
Approximately 1-2% of cells exposed to SV40 virus were transformed under 
normal oxygen tensions. 
ment o r  suppression of the frequency sf transformation when infected cells were  
exposed to one or more 18 hr  pulses of 95% oxygen at 4 day intervals. Oxygen, 
however, is deleterious to normal cells and lowers their plating efficiency 5 to 
BO fold below normal. 
sistant to high levels of oxygen. 
Prellirnbary exper h e n t s  show no dramatic enhance- 
Transformed cells on the other hand appear to be re- 
The effect of prolonged exposure to high oxygen tensions on the frequency 
of viral induced transformations is being investigated further. 
8 
m E C T  OF INCREASED CHAM0ER TEMPERATURE ON RECTAL TEW€RATURE 
OF MICE MAINTAINED AT Sooprlg IN 1% OS IN HI. 
, ,COYPRLSwN 
-* AMBIENT * ~ - R c I u ' o N  - CIKSSlUIC 500 Dsia  
I 1 I I 1 
Figure 1, Ability of increased temperature (35OC) of gas phase 
to enhance maintenance of normal body of mice in normoxic (l% 
0 2 )  - He environment as compared to a 3OoC temperature of the 
gas phase as depicted in Fig. 3 of QSR-19. 
I 
A Group A-i?8% 02 M He. 95PW.35°c 
Group 0 TANK AIR, IATM. 25.c 
DAYS POST CHALLENGE I li I 
Figure 2. Exp, PR8-37. Failure of increased temperature (35OC) 
of gas phase to prevent increased mortality in influenza1 mice ex- 
posed to normoxic (2*8% 0 ) - He environment at 95 psig as com- 
pared with ambient air (25 8 6) influenza1 controls. 
9 
, 
AEROSOL CHALLENGE WITH PR-8 INFLUENZA VIRUS 
I 
A Group A-2.8% 0 2  inHe 
95 prig, 35% 
X Groq~ 840% 0 2  InHo 
I ATM. 25.C 
a 
3 
\x- X- x- x- a 
L 
2ol L g
t 
A l l = d - . J m m S E b r e  
I I I I I 1  I I 1 1 . 1  I I 
I 2 3 4 5 6 7 8 9 10 I I  12 1314 
DAY POST-CHALLENGE 
L 
Figure 3. Exp. PR8-38. Failure of increased temperature (35OC) 
of gas phase to prevent increased mortality in influenzal mice ex- 
posed to normoxic (2.8% 02) - He environment at 95 psig as com- 
pared with ambient 20% 02-He (25OC) influenzal controls. 
r 
il 
< ?  
i 
I '  
il- 
i 
EFFECT OF ALTERED ATMOSPHERE ON SURVIVAL OF WCE FOLLOWING 
bEROSOL CHALLENGE WITH PR-8 INFLUENZP. VIRUS 
A Group A-2.8% Or in He 
EO? 9 5  psIa.35.C 
x Erwp B-20% Or inHe 
I ATM, 2 5 X C  
\ ,  -X 
40 
20 
0 
DAY POST CHALLENGE 
. -. 
Figure 4. Exp. PR8-39. Failure of increased temperature (35OC) 
of gas phase to prevent increased mortality in influenzal mice ex- 
posed to normoxic (2.8% 02) - He environment at 95 psig as com- 
pared with ambient 20% 02-He (25OC) influenzal controls. Note mini- 
mal differences in lung consolidation and infectivity titration between 
test and control mice at sacrifice as presented in Table 1. 
10 
. 
EFFECT OF ALTERED ATMOSPHERE ON SURVIVAL OF MICE FOLLOWING 
AEROSOL CHALLENGE WITH PR-8 INFLUENZA VIRUS 
x Grow B-20% 02 in He 
I ATM, 25°C 
e Group 0- TANK AIR 
\ I ATM.25.C \ 
0 
b 
20- 8 
0 
I 2 3 4 5  6 7  8 9 1 0 1 1 1 2 I 3 1 4  
DAYS POST-CHALLENGE 
I 
Figure 5. Exp. PR8-40. Effect of chilling in ambient gas phase 
of 20% O2 in He (25OC) on mouse influenza1 pneumonia. 
OWING AEao$oL WITH PR-a INFLUENZA VIRUS 
A Group B -77% 02, I ATM. Vinis 
0 GrarpBr-77% 02,l ATM. Dlluant 
o GrarpC -AIR, I ATM.Virus 
e Group CI-AIR, IATM. Diluent 
DAYS POST CHALLENGE 
Figure 6. Exp. PR8-40. Effect of ambient hyperoxic or  
... .,- - 
normoxidq environmental atmospheres on mouse lung influenza 
as compared with uninfected diluent control mice. 
of sacrifice for preparation of lung histopathologic sections. 
Note intervals 
11 


Effects of high and low barometric pressures on susceptibility and resistance to infection Quarterly status report, 1 Apr. - 30 Jun. 1970

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