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Hndaw Pblshn CopoatonPlmonay MdcnVolm 2013, Atcl ID 618576, 9 pashttp://dx.do.o/10.1155/2013/618576

Research ArticleComparison Study of Airway Reactivity Outcomes due toa Pharmacologic Challenge Test: Impulse Oscillometry versusLeast Mean Squared Analysis Techniques

Elena Rodriguez,1,2,3 Charrell M. Bullard,4 Milena H. Armani,1

Thomas L. Miller,5 and Thomas H. Shaffer1,2,5

1 Nemours Research Lung Center, Nemours/Alred I. duPont Hospital or Children, Wilmington, DE 19803, USA2 Nemours Biomedical Research, Nemours/Alred I. duPont Hospital or Children, Wilmington, DE 19803, USA3 Division o Clinical Pharmacology, Tomas Jeferson University, Philadelphia, PA 19107, USA4 Division o Neonatology, Tomas Jeferson University Hospital, Philadelphia, PA 19107, USA5 Department o Pediatrics, Tomas Jeferson University, Philadelphia, PA 19107, USA

Cospondnc shold b addssd to Elna Rodz; mod@nmos.o

Rcvd 31 Dcmb 2012; Rvsd 6 Mach 2013; Accptd 7 Mach 2013

Acadmc Edto: S. L. Johnston

Copyht 2013 Elna Rodz t al. Ts s an opn accss atcl dstbtd nd th Catv Commons AttbtonLcns, whch pmts nstctd s, dstbton, and podcton n any mdm, povdd th onal wok s poplyctd.

T tchnq o masn tansplmonay pss and spatoy afow wth manomty and pnmotachoaphy sn thlast man sqad analyss (LMS) has bn sd boadly n both pclncal and clncal sttns o th valaton o nonatal s-patoy ncton dn tdalvolm bathno lntss and away ctonal mchancal popts masmnts.Whasth tchnq o masn spatoy ncton sn th mpls oscllaton tchnq (IOS) nvolvs th assssmnt o th la-tonshp btwn pss andfow sn an mpls snalwth a an o qncs, qs lss coopatonand povds monomaton on total spatoy systm sstanc (chst wall, ln tss, and aways). T psnt stdy psnts a pclncalanmal stdy to dtmn whth ths spatoy ncton tchnqs (LMS and IOS) a compaabl n dtctn chans nspatoy sstanc dvd om a dct phamacolocal challn.

1. Introduction

T s o anmal modls o stdyn spatoy mchancs

nd away challn tsts has ld to a sddn ncaso nomaton adn th bhavo o th dnt aaso th spatoy systm. Dspt th la amont o -sach n th adlt and pdatc ops, w stll lack sn-cant knowld n th nonatal sbop. T psnt stdypsnts a pclncal anmal stdy to dtmn whthtwo spatoy ncton tchnqs a compaabl n d-tctn chans n spatoy sstanc dvd om a dctphamacolocal challn.

T tchnq o masn tansplmonay pssand spatoy afow wth sophaal manomty, awaymanomty, and pnmotachoaphy has bn pvoslydscbd [1]. ansplmonay pss dvd om

poxmal away psss and ntaplmonay sophaalpss dtctd om a wat-lld catht [2, 3] amasd by dntal pss tansdcs. T afow s

masd wth a low dad-spac volm pnmotachomtanda dntal psstansdc. T last man sqadanalyss (LMS) has bn sd boadly n both pclncal andclncal sttns o th valaton o nonatal ln nctondn tdal volm bathn [1, 4, 5].

T tchnq o masn spatoy ncton snocd oscllaton tchnq (FO), o mpls oscllomty,nvolvs th assssmnt o th latonshp btwn pssand fow sn a ocd/mpls snal composd wth aan o qncs. T spons to ths snal s calld thspatoy mpdanc, whch s th qncy-dpndnt -latonshp btwn tansspatoy pss and fow. Tmpdanc o th spatoy systm nclds th spatoy

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montod and mantand at 37-38C on a adant wambd (Rsscta; Hll-Rom A-Shlds, Hatboo, PA, USA).Onc th physoloc stablty o th anmal was conmdby th analyss o th atal blood chmsty (Stat pol;Nova Bomdcal, Waltham, MA, USA), ECG, and bloodpss monton; th anmals spatoy masmnts

w codd as dscbd blow. Intavnos mdazolamwas sd as an anxolytc to as th wok o bathn. Fol-lown th complton o th potocol, anmals w tha-nzd wth pntobabtal (50 m/k) and satatd potassmchlod (2 mEq). All pocds o th anmal ppaatonw appovd by th Instttonal Anmal Ca and UsCommtt. Fnctonal spatoy paamts o ntstw codd a th anmal was stablzd. T plt thnwas connctd to th PEDS cct o basln mas-mnts. Immdatly ollown LMS basln assssmnt(ava o 5 mn), th plt wasconnctd to th IOS systmo basln oscllatoy masmnts, and tstn was p-omd as soon as th tansspatoy pss tnd to10cmH

2O as dscbd blow. Also, F 1 llstats th

tst paadm that was ollowd n od to compa s-patoy ncton assssmnt btwn th LMS and IOSmthods both bo and a phamacolocally ndcdaway constcton.

2.2. Respiratory Function Assessment-Least Mean SquaredAnalysis. Fo th convntonal tdal bathn sstancmasmnts, th E was connctd to th pnmota-chomt pot. A th-valv stopcock was sd to block thpss pot. T natv fow pot was manally occlddto pvnt fow atact snc th was no addtonal fow va

vntlato tbn. A wat-lld balloon was placd va th

moth nto th sophas to mas th ntathoacc ps-s dn spaton. Optmm placmnt o th balloonwas conmd by al-tm monton o pss tacns,sn cta o maxmm natv dfcton dn nsp-aton wth mnmm cadac atacts. Only spontanosbaths (at last 10 baths) w analyzd, and baths wthdstoton o th snal w xcldd om th ava data.

Plts w obsvd o 30 sconds on th cct. Rs-patoy volms w dtmnatd by lctonc ntatono fow snals, and th ollown paamts w calclatd:dynamc plmonay sstanc () n cmH

2O/L/s, spa-

toy at, tdal volm, and mnt vntlaton. Rsstancvals w codd whl postv pss was appld

to ndc opposton to fow and calclatd by last mansqad (LMS) alothms ncopoatd nto th comptsystm.

2.3. Respiratory Function Assessment-Impulse OscillometryAnalysis. T mpdanc o th total spatoy systm wasmasd sn a commcally avalabl IOS that has bndscbd pvosly. Dn tdal bathn thoh an E,an mpls nato dlvd b plss at ntvals o0.2 sc, spmposd on th spontanos bathn pattn.T dtalzd pss and fow snals w d nto th astFo tansomaton, wh 32 sampls w consdd.Fo xpmntal poposs, no systm cocton was sd.

Pharmacologic challenge

Baseline PFT

LMS IOSLMS IOS

Post PFT

1 mg/kg/hr

BL = 7 0 2Time (hr)

Pig preparation andstabilization period

(1 hr)

Bethanechol1 mg/kg twice

infusion

Test paradigm

Figure 1: A schmatc daam llstatn th cnt tst paa-dm. LMS, last man sqas; IOS, mpls oscllatoy systm;PF, plmonay ncton tstn; , nmb o anmals; BL, bas-ln; Post, a bthanchol admnstaton. m btwn th stand scond bthanchol doss was 10 mn.

Daly calbaton sn a calbaton pmp (3.0 0.01 L SD,Ja; Hochb, Gmany) and a nc mpdanc o10cmH

2O/L/s w pomd, and a maxmm o o 10%

was pmttd. Plts w connctd va E adapt to thpnmotach o thIOS, w tstd o 30 sconds on th c-ct wth a tansspatoy pss o 10 cmH

2O, and w

savd o lat valaton; poch o masmnts was ds-cadd th tm fow and pss pattn n th pmaydata tm and tm slts tnds showd ntpton o thoscllatoy snal, whch occd occasonally (appox-matly 1015%) n th cnt stdy. In th valaton phas,th mnmm tm pod o potn ths masmntswas 2 sconds. T mpdanc o th spatoy systm (s )and s n cmH2O/L/s w calclatd at 3, 5, 10, 15, 20, 25,and 35 Hz. Rplcat oscllatoy vals w pomd byJLAB 4.65.1.0 sowa (Ech Ja GmbH, Wzb, Gmany) and analyzd by JLAB 5.20.0.52 sowa (VIASYSHalthca GmbH, Hochb, Gmany). Snc paamtsw assssd wthn th sam plt ov tm/dlta chans,a cocton o th mpdanc o th E was not ndd.

Each plt povdd a matchd st o data om th Mas-tScn PF IOS (Ja, Hochb, Gmany) and thPEDS plmonay ncton nt (MAS, Hatld, PA) o th

oscllomty and LMS analyss, spctvly. Each plt wasmasd wth ach dvc nd two condtons: basln(p) and a ntvnton (post).

Follown basln mass,bthancholwas sd as thntvnton to phamacolocally ndc bonchoconstc-ton and sbsqnt lvatd away sstanc. Bthancholpowd (cabamyl--mthylcholn, Sma-Aldch) wasmxd wth saln (5 m o bthanchol : 1 mL o saln) andadmnstd as two ntavnos njctons at a dos o1 m/k o bthanchol chlod n 0.2 mL/k o saln st10 mn apat to achv an ncas n th spatoy ss-tanc. Dcay n th lvatd sstanc was compnsatd wthan nson o 1 m/k/h o bthanchol chlod statd

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thoh th vnos ln ollown th ntal bols. Ts dos-n mn was adaptd om a pvosly dscbd pltpotocol [30]. o compa th two mthodolos dscbdhn, w w ntstd n stablshn a la an o pl-monay chans (basln to maxmm dos spons wth-ot vokn sncant cadovascla chans) nstad o a

standad dos spons assssmnt wth small plmonaychans.Bthanchol bols and mantnanc dos w vn as

dscbd abov. Plts w obsvd ntl stabl boncho-constcton was assmd (35 mn a th scond dos)whn sstanc ncasd at last 40% om basln. Post-ntvnton LMS masmnts w obtand, ollowd byIOS masmnts, to pvnt any navoabl ct omth mplss on LMS paamts. Paamts o ntstw docmntd o ach tchnq, ach tm n th samsqnc.

2.4. Statistics. Contnos vaabls w smmazd ov-

all by man and standad o o th man (SEM), nlssndcatd othws. All assmptons w tstd o nomal-ty; n th cas o volaton o any assmpton, nonpaamtctsts w pomd. T dlta chans w compad snpad -tst o th dpndnt contnos nomal vaablso Wlcoxon snd ank tst o th nonnomal dpndnt

vaabls, tstn o postv man o mdan dncs,spctvly. Sncanc was st at alpha lvl o 0.05 (1-sdd). A compaatv analyss o amnt btwn th twomthods was dtald o th phamacolocal challnsn th Bland-Altman plot analyss; th bas, o th avao th dncs, and th 95% lmts o amnt (LOA)w potd o all qncs alon wth th lna -

sson analyss (2

, slop, and vals). Statstcal analysswas don sn a combnaton o statstcal sowa packas(GaphPad Psm vson 5.02 o Wndows; GaphPad So-wa, San Do, CA, USA, and SPSS vson 19; IBM, Ch-cao, IL, USA).

3. Results

In ths anmal modl, masmnts o spatoy mchan-cs w possbl, and dtcton o an ncas n sstancwas achvd wth both systms a bthanchol admns-taton n svn o th plts. On anmal dd dn thpostbthanchol masmnts (post) sconday to ca-

dac ast. Follown ntavnos (IV) admnstaton obthanchol, a small bt statstcally sncant dcas wassn n hat at (man SD, 15112 vss 1336, 12%, = 0.03) and atal pss (man SD, 79 5 vss675, 15%, = 0.002) o th mann 7 plts; althohths chans w not physolocally lvant o hmody-namc nstablty, th w no sncant chans n sp-atoy at, tdal volm, and mnt vntlaton ollownbthanchol admnstaton.

Rspatoy mchancs masmnts showd a sn-cant ncas n dynamc plmonay sstanc masd byth LMS (+53%, = 0.003) ollown bthanchol admns-taton (F 2). Pss-volm loops also vd that,

whn compad wth basln, bthanchol admnstatonsltd n ncasd pss qmnts as wll as awdnn and dcas n th slop o th pss-volmloops.

Follown bthanchol admnstaton, a sncant ov-all ncas n th spatoy sstanc masd by IOS was

dmonstatd by postv chans n s om 10 to 25 Hz(+96%, = 0.018; F 3). Sncant ncass n spc-c spatoy sstancs w dmonstatd by postvchans n th ntmdat qncy spctm (qn-cs: s 10Hz [+106%, = 0.031], s 15Hz [+96%, = 0.013],s 20Hz [+95%, = 0.026], and s 25Hz [+85%, = 0.037]).No statstcally sncant dncs btwn th p-bthanchol (p) and postbthanchol (post) otcoms oany mann qncs w obsvd: s 3Hz (+67%, =0.131), s 5Hz (+67%, = 0.09), and s 35Hz (+95%, =0.084), and th sstanc spctm o s dmonstatdncasd postv qncy dpndnc (F 4).

T Bland-Altman analyss dmonstatd that 95% o thdncs btwn mthods (IOS and LMS) lay wthn 1.96SD om th man dnc ndcatd by th LOA. Ovall,th was a consstnt lna slopd tnd wth a systmatcbas (natv bas psntd at low vals and postvbas psntd at hh vals o th masmnt an)acossth Bland-Altmanplots, bt wthn thLOA (F 5);ths ct bcoms lss basd towads low qncs(Fs 5(a) and 5(b)) asdo th slops oth lnassonanalyss.

4. Discussion

In th psnt stdy, w dmonstatd that both spatoyncton tchnqs assssd th cts o ntavnosbthanchol on th spatoy systm. O slts showdthat dlta chans o oscllomtc sstancs n th mddlqncy an (s 10, 106%; s 15, 96%; s 20, 95%; s 25,85%) w hh than LMS dynamc plmonay sstanc(, 53%).

T smooth mscl o th spatoy tact has both paa-sympathtc and sympathtc nnvatons; cabamyl--mthylcholn (bthanchol) s a choln st and an aonsto mscanc cptos that stmlats ths cptos on thsmooth mscl o th spatoy tact, casn away na-own and phaps tss constcton. Bthanchol blonsto th sam op o ds as actylcholn, cabachol, and

mthacholn [39]. T plt modl sd n ths stdy waschosn basd on ts compaabl sz to th nonat/pdatchman and ts common s o th nvstaton o vntla-toy thaps [3137].

In addton, bthanchol-ndcd away challns havbn pvosly potd as an appopat xpmntalntvnton n th laboatoy [4044], and smla analos(.., mthacholn) hav bn sd o clncal sach p-poss [4547]. In hmans, bthanchol s appovd only ooal admnstaton s; n vtnay mdcn, t s avalablas an njcton o sbctanos (SC), ntamscla, o IVadmnstaton. Cadac ast s a sv, a advs ac-ton docmntd n hmans, and n anmals, t s a lkly

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40

30

20

10

0

Resistance

(c

mH2

O/L/s)

Pre Post

Bethanechol

= 0.003

Figure 2: Plots o man vals o dynamc plmonay sstanc(sstanc) bo (p) and a (post) bthanchol o ach plt. = 0.003, pad -tst on tald.

= 0.018

40

50

30

20

10

0 Pre Post

Rrs1025Hz

(cmH2

O/L/s)

Figure 3: Plots o man vals o spatoy sstanc (s ) om10 Hz to 25 Hz bo (p) and a (post) bthanchol o achplt. = 0.018, pad -tst on tald o th ava.

sv cholnc acton vn IV o n ovdosa. It scommndd that atopn b mmdatly avalabl whnvn IV o SC [4851].

A bthanchol admnstaton, a at pcnta ochan was sn n away sstanc manly psntd bys 10s 25 (+10685%) whn compad wth th ln sst-

anc mostly psntd bys 3s 5 (+6963%), ndcatna at nonpphal away constcton spons thanpphal away/ln tss spons. Radn th lntss contbton vss away contbtons to th totalspatoy sstanc, dncs n th d o tss v-ss away contbton xstbtwn spcs [52, 53], ncld-n hmans [54], and wthnndcd-physolocaland pha-macolocal condtons [44]; ths, caton shold b takn ntanslatn ths ndns btwn spcs.

It s wdly conzd that th s o an sophaal bal-loon o sstanc masmnts spaats th ln andchst wall componnt, tho, xcldn th chst wallsstanc [4], whl s nclds th sstanc o th chst

Post

Pre

Frequency (Hz)

3 5 10 15 20 25 35

80

60

40

20

0

= 0.0002

Rrs

(cm

H2

O/L/s)

Figure 4: Plots o man (SEM) vals o spatoy sstanc(s ), as a ncton o oscllatoy qncy, bo (p) and a(post) bthanchol

( = 7 ). As shown, sncant dncs n

th mddl oscllatoy qncs an a dsplayd (

< 0.05);pad -tst on tald at ach qncy and ovall ( = 0.0002).

wall (.., ln sstanc) and away sstanc. T lowth spcc oscllatoy qncy s, th mo snstv thmasmnt s to th pphy o th spatoy systm.Masmnt o th sstanc sn mthods that qplacmnt o an sophaal balloon o spatoy nctontstn pocds to assss th spatoy systm n no-nats s not asy. Nonnvasvspatoy ncton tchnqs(.., mpls oscllomty) q lss patnt coopatonand povd mo nomaton abot th total spatoysystm sstanc [19].

W also dmonstatd that no pas n bathn wasncssay o codn labl data at low qncs os sn th mpls oscllaton tchnq n nonatal pltsspontanosly bathn thoh an E. In ths ad, thHn-B nfaton fx and ac mask w not sdn o stdy n compason to pvosly ctd stds n n-sdatd nonats/nants [28, 29].

T Bland-Altman analyss showd ood amnt bt-wn th two tchnqs (IOS andLMS) omasn sst-anc popts, and th amnt btwn ths tchnqs

dpndd on th oscllaton qncy. Fom a mchanstcvw pont, masd by th LMS n th stdy psntssstanc o th ln tss and away ctonal mchancalpopts, whas s masd by IOS psnts total s-patoy sstanc, so th two tchnqs a not dctlyntchanabl at low qncs. Howv, as mntondn Scton 3, th bas and slop dcas towads low q-ncs. In ths ad, spatoy sstanc at 5 Hz (s 5)and spatoy sstanc at 3 Hz (s 3) may b closlyntchanabl n th appopat sttns. Howv, thdnc o 1.96 SD om th man dnc o s 35(F 5()) was dmonstatd to b naccptabl o nt-chanablty.

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28.75

Mean1.03

Slope = 2.36 0.72

= 0.67, value: 0.024

70605040302010

0

20

30

40

50Diferencein%change

(LMS-IOS)

10

10 5 0 5 10 15 20 25 30 35 40

Average in % change by LMS and IOS

30.75

+1.96 SD

1.96 SD

Rrs 3Hz

(a)

70605040302010

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20

30

40

50

10 5 0 5 10 15 20 25 30 35 40

Average in % change by LMS and IOS

Dif

erencein%change

(LMS-IOS)

25.67

Mean0.67

27.02

Slope = 2.2 0.82

= 0.56, value: 0.051

10

+1.96 SD

1.96SD

Rrs 5Hz

(b)

10 5 0 5 10 15 20 25 30 35 40

Average in % change by LMS and IOS

Mean

2

= 0.67, value: 0.024Slope = 2.39 0.7

25.96

33.43

3.73

70605040302010

010

20

30

40

50

Diferencein%change

(LMS-IOS)

+1.96 SD

Rrs 10Hz

1.96 SD

(c)

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20

30

40

50

10 5 0 5 10 15 20 25 30 35 40

Average in % change by LMS and IOS

Mean

31.01

4.33

22.34

2

= 0.62, value: 0.034Slope = 2.84 0.9

Dif

erencein%change

(LMS-IOS) +1.96 SD

Rrs 15Hz

1.96 SD

(d)

Mean

Slope = 2.83 0.6

5.95

40.31

28.4

70605040302010

010

10

2030

40

50

5 0 5 10 15 20 25 30 35 40

Average in % change by LMS and IOS

Difer

encein%cha

llenge

(LMS-IOS)

2

= 0.81, value: 0.005+1.96 SD

Rrs 20Hz

1.96 SD

()

Mean

33.88

49.94

8.02

70605040302010

010

2030

40

50Diferencein%cha

llenge

(LMS-IOS)

10 5 0 5 10 15 20 25 30 35 40

Average in % change by LMS and IOS

2

= 0.87, value: 0.002+1.96 SD

Slope = 2.59 0.4Rrs 25Hz

1.96 SD

()

70605040302010

010

20

30

40

50Diferencein%cha

llenge

(LM

S-IOS)

10 5 0 5 10 15 20 25 30 35 40

Average in % change by LMS and IOS

Mean

67.92

9.88

48.15

Slope = 2.06 0.22

= 0.91, value: 0.0009+1.96 SDRrs 35Hz

1.96 SD

()

Figure 5: Bland-Altman plots constctd om th last man sqa (LMS) and mpls oscllomty (IOS) data at ach spcc qncy(pcnta o chan) wth man (bas), LOA, and bst-t vals o lnasson (slop, 2, val) a admnstaton o ntavnosbthanchol. LOA, lmt o amnt; Upp LOA, man dnc (bas) + 1.96 SD; Low LOA, man dnc (bas) 1.96 SD; 2, sqas. T val s tstn th nll hypothss that th ovall slop s zo.

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Plmonay Mdcn 7

Lk many plmnay stds, th psnt stdy hassom xpmntal lmtatons. Alon ths lns, th cntstdy s an xpmntal pclncal modl compan thIOS and th LMS appoach as an ndcato o away ac-tvty bhavo to a phamacoloc ntvnton n a smallnmb o anmals. Addtonal wok s qd n th po-

manc, podcblty, and saty o th oscllaton tchn-q, ncldn valdaton and qalcaton as a bomak,bo t can b poposd o standad clncal pactc n thnonatal poplaton.

5. Conclusions

In conclson, w docmntd sncant away actvtysn both spatoy ncton tchnqsand dmonstatdth mantd o th chan n away sstanc componntat basln and a admnstaton o ntavnos bthan-chol. In a smla scnao, th masmnts o spatoysstanc om th mpls oscllaton tchnq (om 3 Hz

to25 Hz) may b sdn l o th dynamc plmonayss-tanc masmnts om th LMS tchnq to dtmnsstanc chans sconday to ndcd away actvtydn spontanos baths.

T IOS tchnq as a nonnvasv mans o valatnth dnt componnts o th spatoy systm and pl-monay mchancsn th nonatal sttn s vy appaln; tvs th nomaton thoh a an o sstanc valspsntn th away, tss, and chst mchancal pop-ts. Howv, th modcaton n th tchnq stp,ncopoaton o th appopat ncs vals o thnonatal poplaton, and collaboaton wth xpncdphyscans and scntsts n spatoy physoloy/spa-

toy ncton tchnqs a qd bo boad clncals s psd n th nonatal poplaton.

Funding

Ts stdy was ndd thoh Nmos Rsach Po-ams/Nmos Fondaton, NIH COBRE Gant no. 8P20 GM103464 (. H. Sha), and NIH Gant no. 532GM008562-17 (E. Rodz) pdatc phamacoloy llowndd by NICHD nd th Bst Phamactcals o Chl-dn Act. Ts wok was pomd at th Nmos/Ald I.dPont Hosptal o Chldn.

Acknowledgment

T athos thank Ms. Ann Hsk oh assstanc wth thanmal scal ppaaton.

References

[1] V. K. Bhtan,E. M. Sv, S. Abbas, and.H. Sha, Evala-ton o nonatal plmonay mchancs and ntcs: a twoacto lastman sqa analyss, Pediatric Pulmonology,vol.4,no. 3, pp. 150158, 1988.

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