An Evaluation of the Multiple-breath Nitrogen Washout as a Pulmonary Function Test in Horses G.J. Gallivan, L. Viel and W.N. McDonell
ABSTRACT
appeared to be the most sensitive index in detecting the horses with COPD. In the horses with COPD, salbutamol caused a decrease in FRC and FETN2,fb indicating increased ventilation of the lung. However, LCI increased suggesting a less efficient distribution of ventilation.
Multiple-breath nitrogen washouts (MBNW) were performed with 29 light horses. Seven normal horses were used to examine the reproducibility, and 22, ranging from normal to severely diseased, were used to examine the changes in chronic obstructive pulmonary disease (COPD) and the effect of a bronRESUME chodilator, salbutamol, on the distribution of ventilation. The MBNW Une etude des lavages pulmonaires a were analyzed using the functional l'azote au cours de respirations residual capacity (FRC), end-tidal N2 repetitives (LPAR) fut entreprise sur 29 concentration of the final breath of chevaux. Sept chevaux sains furent the MBNW (FETN2,fb), end-tidal N2 utilises pour confirmer la reproductibiconcentration when the cumulative lite des tests, 22 autres chevaux parmi expired volume was equal to body lesquels certains etaient sains et certains weight (FETN2,bw), lung clearance etaient malades servirent a examiner les index (LCI), Becklake index (BI), changements present lors de maladies mixing ratio (MR), index of distribu- respiratoires obstructives chroniques tion of inspiration (IDI), pulmonary ainsi qu'a etudier un bronchodilatateur N2 clearance delay (PCD) and (salbutamol) et ces effets sur la ventilatory efficiency (EFF). The ventilation pulmonaire. Le LPAR fut LCI, MR, IDI, PCD and EFF were analyse en tenant compte de la capacite calculated at end-tidal N2 concentra- residuelle fonctionnelle (CRF), de la tions (FETN2), equal to 3%, 2% and concentration en azote du volume courant a la fin du LPAR, de la FETN2,fb. The EFF was also calculated concentration finale en azote lorsque le at FETN = 8%, as was BI. The' within day coefficient of volume expiratoire cumulatif egalait le variation for most indices of the poids du cheval, de l'ndice de clerance MBNW was 10-15%. None of the pulmonaire (ICP), de l'indice de indices varied significantly from day- Becklake (I.B), du rapport des to-day. The FETN2,fbg FETN2,bW, MR, IDI, melanges gazeux (RMG), de llndice de PCD, and EFFfb, EFF3% and EFF2% distribution inspiratoire (IDI), du delai differed between the horses with de clerance pulmonaire en azote COPD and the normal horses, and all (DCPN2) et de l'efficacite ventilatoire of the indices, except FRC, were (E.V.). Ces derniers parametres furent correlated with a histopathological calcules lorsque la concentration du volume courant en azote egalait 3 et 2 score of the small airways. The FETN2,bW
a la fin du LPAR. L'efficacite ventilatoire et lI.B. furent aussi calculees lorsque la concentration finale en azote egale 8%. Le coefficient de variation pour la plupart des indices du LPAR obtenu au cours de la meme journee etait de 10-15%. Aucuns parametres ne variaient d'une facon significatives si les tests etaient repetes de jour en jour. La concentration en azote du volume courant a la fin du LPAR, la concentration finale en azote lorsque le volume expiratoire cumulatif egale le poids du cheval, le RMG, l'IDI, le DCPN2 et l'efficacite ventilatoire a 3%, 2% et a la fin du LPAR demontrerent des changements significatifs entre les chevaux souffrant de maladies respiratoires obstructives chroniques et les chevaux normaux. Tous les parametres, exception faite du CRF demontrerent une bonne correlation avec les donnees histopathologiques des petites voies respiratoires. La concentration finale en azote lorsque le volume cumulatif egale le poids du cheval semblerait etre lindice le plus sensible pour identifier les chevaux souffrant de maladies pulmonaire obstructives chroniques (MPOC). Chez les chevaux souffrants de MPOC, une baisse du CRF fut observee suivant l'administration du salbutamol, alors que la concentration finale en azote du volume courant demontrait une amelioration de la ventilation pulmonaire. Cependant, l'indice de clerance pulmonaire augmenta, ce qui indiquait une distribution ventilatoire beaucoup moins efficace.
pourcents et
Department of Clinical Studies, Ontario Veterinary College, University of Guelph, Guelph, Ontario N1G 2W I. Reprint requests to Dr. W.N. McDonell. Funding for this research was provided by the Medical Research Council of Canada and the Ontario Racing Commission. Submitted March 3, 1989.
Can J Vet Res 1990; 54: 99-105
99
INTRODUCTION The multiple-breath nitrogen washout (MBNW) is a relatively simple, noninvasive pulmonary function test for studying the distribution of ventilation. It is often useful in the detection of pulmonary diseases and in assessing the degree of functional impairment (1,2). The use of the MBNW as a pulmonary function test in horses was first described by Muylle et al (3) who suggested that the endtidal N2 concentration (FETN2) after breathing 600 L of 02 was useful in the detection and diagnosis of ventilatory abnormalities. Willoughby and McDonell (4) suggested using the FETN when the cumulative expired volume (EXVE) was equal to body weight for the detection of small airway disease in the horse. In the present paper we evaluate the MBNW as a pulmonary function test in horses, examining different methods of interpretation, their reproducibility, the changes in horses with chronic obstructive pulmonary disease (COPD), and the effect of a bronchodilator, salbutamol, in the horses with COPD. MATERIALS AND METHODS
This study was conducted with 29 light horses weighing from 338-580 kg, and ranging from 2-14 yr in age. Seventeen of the horses were classed as normal on the basis of a thorough clinical examination of the respiratory system, and gross and microscopic examination of the lungs at necropsy. The other 12 had COPD, ranging from mild to severe. The horses were housed in the Veterinary Teaching Hospital, Ontario Veterinary College during the measurements. There was no seasonal pattern in the measurements. The MBNW were performed as described previously (2,5). Briefly, the horse was restrained in 1 x 2 m stock. A mask was placed over the muzzle and the distal end sealed so the horse inspired and expired through a nonrebreathing valve (6). The inspiratory port of the nonrebreathing valve was connected to an 02 supply, and the expiratory port was connected to a 600 L spirometer (Warren E. Collins, Inc., Braintree, Massachusetts). At
100
was equal to body weight (FETN2,bw), lung clearance index (LCI), Becklake index (BI), mixing ratio (MR), pulmonary N2 clearance delay (PCD), index of distribution of inspiration (IDI) and ventilatory efficiency (EFF). The formulae used in calculating the FRC, LCI, BI, MR, PCD, IDI and EFF have been described by Gallivan and McDonell (2). The only difference was the use of a physiological dead space to tidal volume ratio (VD/ VT) of 0.62, the value for normal horses using this system (7). The VD/ VT ratio was not measured in the present study, but was needed for the calculation of alveolar ventilation used in the calculation of MR, PCD, IDI and EFF. The LCI, MR, PCD and IDI were calculated at FETN2 equal to 3% and 2%, and at FETN, fb (subscripts 3%, 2% MBNW (FETN2 fb), the FETN when Z VE and fb respectively). The BI was
end-inspiration the horse was given 100% 02 to breathe and the expired air was collected in the spirometer. The expired N2 concentration was measured using a rapid response N2 analyzer (Nitroverter, VR-3500, Vertek, Burlington, Vermont) and recorded as a function of the cumulative expired volume using a photographic recorder (VR-6, Electronics for Medicine, White Plains, New York). The expired volume was measured using a heated pneumotachograph (Fleisch #4, Instrumentation Associates, New York, New York) placed between the mask and the nonrebreathing valve. The washout was continued until FETN was less than 1%, or until the spirometer was filled. The MBNW were analyzed using the functional residual capacity (FRC), FETN of the final breath of the
B
A
80
score = 82
= 33
score
10:
iZlo: L-LL.
I
1 1 1
0
80
30
1
1
60
1
,
5
90
~I
60
120
90
120
90
120
D
c
score
score = 222
173
10
10:
AL
0
30
60 BREATH
90
120
0
30
60 BREATH
Fig. 1. Representative MBNW curves from a normal horse (A) and three horses with COPD (B-D). The two lines in A are two MBNW on the same day. The score is the histopathological score for the lungs (10).
TABLE I. Within-day coefficients of variation of the indices of the MBNW at different end-points for seven horses on two days
End-pointb
Variablea FRC (mL kg-')
8%
3%
2%
-
-
-
10.4 (10.5)
14.5 (12.5)
fb 6.1 (3.5)c 28.8 (22.2) 10.9 (8.2)
10.6 (10.7) 31.1 (28.4) 10.4 (10.6) 10.4 (10.5)
14.6 (13.0) 37.4(32.3) 14.5 (12.5) 14.5 (12.5)
8.8 (6.9) 21.2(16.3) 8.7 (6.8) 8.6 (6.9)
FETN2(%)
LCI BI MR PCD(%) IDI EFF(%)
7.0 (5.1)
_
7.0 (5.1)
-
ajd
bw
-
29.2 (12.8)
10.6 (7.7) -
-
aFRC: functional residual capacity; FET,: end-tidal N2 concentration; LCI: lung clearance index; BI: Becklake index; MR: mixing ratio; PCD: pulmonary N2 clearance delay; IDI: index of distribution of inspiration; EFF: ventilatory efficiency bEnd-points are FET = 8%, 3%, 2% and the final breath (fb) of the MBNW; adj: adjusted to compensate for t&e percentage of the initial N2 remaining in the lung at the end of the MBNW (2); bw is the FETN2 when the cumulative expired volume equals body weight cMean (standard deviation)
calculated as FETN = 8% (subscript 8%) which represented the point at which 90% of the N2 had been washed out of the lung as the FETN2 at the beginning of the washout was approximately 80%. The EFF was calculated at FETN2 = 8%, 3%, 2% and FETN2Thb. Several of the normal horses and one of the horses with COPD reached the end-point of FETN2 = 1 % when EVE was less than body weight. For these individuals FETN2 fb was used as the value of FETN bw. EXPERIMENTAL PROTOCOL
All the MBNW were measured between 0800 and 1200 h, after the horse had been starved for 16-18 h to eliminate the reduction in lung volume associated with feeding (8). Two
MBNW were measured with each horse. Each MBNW required 5-7 min, and they were separated by an interval of 7-10 min during which the horse breathed room air. To examine the reproducibility of the MBNW, measurements were made on two days with seven normal horses. The interval between days varied between horses, ranging from 2-10 days. The results were analyzed using a nested analysis of variance (9) with horses and days within horses as the factors. The other 22 horses, 10 normals and 12 diseased, were used in a study of COPD. Measurements were made on one day with each horse. Shortly thereafter the horse was euthanized and a histopathological score for the
lungs was calculated based on the degree of airway occlusion, cellular infiltration, fibrosis, smooth muscle hypertrophy, and epithelial cell hyperplasia and metaplasia of the small airways (10). The indices of the MBNW were then compared to the histopathological score using regression analysis (11). To examine the sensitivity of the indices in a clinical situation, the results from individual horse with COPD were compared to the 95% confidence limits calculated for the population of ten normal horses. To examine the effect of a bronchodilator on the distribution of ventilation in COPD, 1.2-1.5 mg of a 132-agonist, salbutamol (Ventilin, Allen and Hanburys, Toronto, Ontario), was administered via aerosol into the mask after the second MBNW in the 12 diseased horses. The MBNW was then repeated. The changes in the indices before and after salbutamol were tested using a paired t-test, and were compared to the histopathological score and the values obtained prior to the administration of salbutamol using regression analysis.
RESULTS The typical MBNW was irregular (Fig. 1) with the most horses exhibiting a marked reduction in tidal volume (VT) after three to four breaths of 02. However, VT was also variable throughout the remainder of the washout causing some irregularity in the MBNW.
TABLE II. Indices of the MBNW in normal horses and horses with chronic obstructive pulmonary disease (COPD)
Correlation with Variablea FRC (mL-kg')
FETN2fb (%) FET 2,bw (%) LCtad' BI
MRf,
IDIfi
PCD(%)
Normals (n 10) 54.2 ± 7.4d 1.0 ± 0.4 1.1 ± 0.3 18.3
9.2 1.47 ± 1.54 ± 47.2
4.1
2.4 0.29 0.30 29.1 77.0± 17.1 67.4 ± 14.7
95% Confidence limits for normals 37.4; 70.9 0.2; 1.8 0.3; 1.9 9.0; 27.5
3.7; 14.7 0.81; 2.13 0.85; 2.23
COPD horses (n = 12) 60.1 ± 16.4 (1+, 3+)e 2.7 ± 1.6 (6+) 3.2 ± 1.8 (9+) 21.2 ± 4.0 (1+) 11.0 ± 3.3 (2+) 2.15 ± 0.45 (7t) 2.24 ± 0.48 (7+) 115.0 ± 45.2 (7+) 63.4± 15.2 (1+) 46.8 ± 10.7 (1+)
histopathological score pb
r
0.433 0.001 0.001
-0.03 0.54 0.58
0.104
0.49
0.010
0.173 0.001 0.001 0.001 0.105 0.001
0.39 0.75 0.74 0.75 -0.38 -0.66
0.037