 receptor (ARF-DTR) and wild-type mouse lungs treated with diphtheria toxin (DT) or PBS, as depicted in Figure 4A. Mice in the indicated groups were euthanized and connected to the FlexiVent system through their tracheae. Mice were subjected to laparotomy, and diaphragms were removed prior to the assay. (B) Static lung compliance (Cst) was calculated from the slope of the pressure-volume loop. Cst reflects the static elastic recoil pressure of the lungs at a given lung volume. (C–F) Dynamic lung compliance (Crs) (C), dynamic lung resistance (Rrs) (D), tissue elastance (H) (E), and tissue damping (G) (F) are shown. Dynamic lung compliance captures the ease with which the lungs may be extended, and dynamic lung elastance captures the elastic rigidity or stiffness of the lungs. Tissue elastance and tissue damping reflect energy conservation and dissipation in lung alveoli, respectively. Values represent mean ± SEM of independent experiments. Data were analyzed 1-way ANOVA followed by post-hoc Tukey-Kramer multiple comparison test. *P < 0.05, **P < 0.01, ***P < 0.001.

We also compared lung morphologies among these mice. An increase in alveolar size is associated with a decline in lung function during aging (18, 19, 22, 23). Alveolar mean linear intercepts (the average distance between alveolar walls that reflects the size of the alveolar space; ref. 24) were increased in 12-month-old ARF-DTR mice (Figure 6, A and B). The DT treatment reversed this phenotype but did not affect alveolar size in wild-type mice. Mean alveolar septal wall thickness was lower in 12-month-old mice than in 2-month-old mice (Figure 6C), as previously reported (25). Furthermore, the ablation of p19ARF-expressing cells fully restored the alveolar wall in ARF-DTR mice but not in wild-type mice. Increases in lung compliance in aged animals have been attributed to a decline in the number of elastic fibers, the major component of which is elastin (19, 22). Therefore, we performed a quantitative analysis on elastin in immunostained sections (Figure 6D). The amount of elastin in older animals was approximately half that in young animals (Figure 6E) and recovered in DT-treated ARF-DTR mice
receptor (ARF - DTR) and wild - type mouse lung treat with diphtheria toxin (DT) or PBS, as depicted in Figure 4A. mouse in the indicate groups were euthanized and connected to the FlexiVent organization through their trachea. Mice were subject to laparotomy, and diaphragms were removed prior to the assay. (bel) Static lung compliance (Cst) was count from the gradient of the pressure - volume loop topology. Cst reflects the static elastic recoil press of the lungs at a given lung volume. (C – F) active lung compliance (Crs) (C), dynamic lung electric resistance (Rrs) (D), tissue elastance (H) (E), and weave damping (G) (F) are show. Dynamic lung compliance captures the relief with which the lungs may be extended, and dynamic lung elastance captures the elastic rigidity or stiffness of the lungs. Tissue elastance and tissue damping reflect energy conservation and dissipation in lung alveoli, respectively. Values represent mean ± SEM of independent experiments. Data were analyzed 1 - manner ANOVA followed by post - hoc Tukey - Kramer multiple comparison test. * phosphorus < 0.05, * * phosphorus < 0.01, * * * P < 0.001. 

 We also compared lung morphologies among these mouse. An increase in alveolar size is associated with a decline in lung function during age (18, 19, 22, 23). Alveolar mean linear intercepts (the average distance between alveolar walls that reflects the size of the alveolar space; ref. 24) were increase in 12 - month - old ARF - DTR mice (Figure 6, vitamin a and B). The DT discussion change by reversal this phenotype but did not affect alveolar size in wild - type mice. Mean alveolar septal wall thickness was lower in 12 - calendar month - old shiner than in 2 - month - old mouse (Figure 6C), as previously report (25). Furthermore, the ablation of p19ARF - expressing cells amply restored the alveolar rampart in ARF - DTR mice but not in wild - type mice. increase in lung compliance in aged animals have been assign to a decline in the issue of elastic fibers, the major component of which is elastin (19, 22). Therefore, we performed a quantitative analysis on elastin in immunostained sections (Figure 6D). The amount of elastin in older animal was approximately half that in young animals (trope 6E) and recovered in DT - treated ARF - DTR mice
 refeptor (ARF-DTR) and wild-tyke mouse lungs trgared wivh diphfheria tuxin (DT) or PBS, as depicted ii Fiture 4Q. Mice in the indicatea groups aere eutyanieed and connected to the NjexiBcnt svsvem through thelr tracheae. Mice were subbeztzd to laparotomy, and diaphragms were removec orior to the afsay. (F) Stznig lung compliance (Cst) was calculzted frmm the slope pf the pressure-volume loop. Cst reflects the statlc elastic tscoyo pressure ow the lungs at a given lung volume. (C–F) Dynamic lung coopliauce (Crs) (C), diuqmif lung resisvance (Grs) (D), tissue ckastanwe (H) (E), snd tissue damiing (J) (F) qre shown. Dynamic lunj compliance capturef the easa cith which the lungs nat be gxtenged, xbd aynzmmc mung epasvance captudes the elawtic rigidity or stofsbess of the lhngs. Tysfue elastance and tissue damping reflecu enedgy conservation and diwsipation in lung alvgoli, respestively. Values represent mean ± SEM of independent expecioenus. Data qege analyzed 1-way ANOVA followed by post-hoc Tukqg-Ktaker multiple cjmparison trsh. *L < 0.05, **P < 0.01, ***P < 0.001.

We xlso compzred lung morpholohies amjng tyese mice. An oncrease in alveolar size iw associated qith a decline in pung functiun doring sging (18, 19, 22, 23). Alveolar mean linsar interceots (the abdrage distance bdtwven dlveolar walls that refleces the sive of the alxeolsr spase; ref. 24) wege ingseased in 12-month-old ARF-DJR mica (Figure 6, W and B). The DT treatment reversxv this phenotipe bun did not affegt alveolar sizq in wild-type kice. Mecn alvdolar septzl wall thickness ras lower in 12-klnth-old mice than in 2-monrh-ole mice (Wkgure 6C), as prefiously rvpjrtee (25). Furthermore, the abuztion of p19ARF-ex'xewsing cells fulky feseoged tra alveolar wdll kn XTF-DTR mice but nig in wild-type mice. Incredses in lung compliancr ln aged abimals hwve been attrobuted to a declint in tie numuer of elwstic fibers, the major componeht of whifh ls elastin (19, 22). Ehercforg, we perforked a quantitative analysis on elastin mn immunostained sectiobs (Figure 6D). The amoont of elastin in ojder animdls was approximatelt half that in yomng animals (Figure 6E) ahd recmveref in DT-treated ARF-DTR mice
receptor (ARF-DTR) and wild-type mouse lungs treated toxin or PBS, depicted in Figure groups euthanized and connected the FlexiVent system their tracheae. Mice were subjected to and diaphragms were removed prior to the assay. (B) Static lung compliance (Cst) calculated from the slope of the pressure-volume loop. Cst reflects the static elastic pressure the at given lung volume. (C–F) Dynamic lung compliance (Crs) (C), dynamic lung resistance (Rrs) (D), tissue elastance (E), and tissue damping (G) (F) are shown. lung compliance captures the with which the lungs may extended, dynamic lung captures elastic or stiffness of lungs. Tissue elastance and tissue damping reflect energy conservation and dissipation in lung alveoli, respectively. Values represent ± SEM experiments. Data analyzed ANOVA by post-hoc Tukey-Kramer test. *P < 0.05, **P < 0.001. We also compared lung morphologies among these An increase alveolar size is associated with a in lung function during aging (18, 19, 22, Alveolar mean linear intercepts (the average distance between alveolar walls that reflects the size of space; ref. 24) were in 12-month-old ARF-DTR (Figure A B). DT treatment this phenotype but did not affect alveolar size in wild-type mice. alveolar septal wall thickness was lower in 12-month-old mice than mice 6C), as previously (25). Furthermore, the ablation p19ARF-expressing fully restored the alveolar ARF-DTR but mice. in compliance in aged animals been attributed to a decline the number of elastic which is elastin (19, 22). Therefore, we performed quantitative analysis on elastin in immunostained sections 6D). The amount of elastin in older animals was approximately half that young animals and recovered in DT-treated ARF-DTR mice
 receptor (ARF-DTR) and wild-type Mouse lungs TreatEd wIth DiPhthEria Toxin (DT) or PBS, as DEpicTed in Figure 4A. Mice in the iNdicaTeD GrouPS wEre euThanizeD AnD COnnEcTeD to ThE flExiVeNt sYstem thRough their TraChEae. Mice were sUBjEcted to lapAroTomy, and diaphRagMs were ReMovED prioR to The asSay. (B) StATic lunG complianCe (cSt) was cALculateD FRoM the Slope of the pressurE-VoLUme loop. Cst reflEcts thE sTAtIC ElaStiC recoil preSsUre of THe lungs AT a GIVEn lUNg volume. (C–F) DynAmic lung comPLiaNce (Crs) (c), dYnaMIc lung ResisTaNCe (RRs) (D), tissue elAstaNce (H) (E), and tIssue dAMping (G) (F) ARe shown. dynamiC luNg cOmplIAnCe CapTuREs tHE eAse WIth Which the LuNgS may bE extENDED, and DynAmic Lung eLastance captuRes The eLAstIc rigIdity Or stIfFness Of the lUngs. TIsSue elastance and TissUe damping RefLeCt eNeRgy coNServatIon And DissipaTion in lUNg aLvEOLI, rEspectively. Values rEpRESeNt mean ± SEm of indEPeNdENt experiMeNts. data WERe anaLyzeD 1-WaY ANOVA foLlowed BY pOsT-hoc TukEy-kramer MuLtiPle CompaRIson Test. *P < 0.05, **P < 0.01, ***p < 0.001.

We also cOmparED lung morphologIEs among these mICe. aN InCReasE in Alveolar sizE is aSSociAted WItH a dECline In lunG fUNcTIon during aging (18, 19, 22, 23). AlveoLaR mean lInear Intercepts (the Average disTANCe betweeN alvEOlAR walls that reflEcts tHe size of thE Alveolar Space; Ref. 24) were iNcreased iN 12-MOnth-old ArF-DtR mIce (figURE 6, A And B). The DT treaTMEnt rEvErsed thIs pHenotypE buT diD noT afFeCt alveolaR size in wIlD-tYpE mIce. mean aLVeolar sePtAl wAlL thIckneSS was loWer in 12-MontH-oLd MIce Than in 2-mONtH-OLd miCe (fiGure 6c), as PrEviouSly rEPorTed (25). FurtHermore, thE abLAtioN oF p19aRF-exprEssing cells fuLlY restored tHe AlvEolar wALL in ARF-DTr mice but not in wild-type miCE. IncreaSes In lunG comPliance in AgeD animaLs hAVe been AttribUted tO a DecLINe in tHE NuMbeR oF elastic fiBERs, tHe majOr CompOnent of Which is elastin (19, 22). TherEForE, we performed a QuaNtitATIvE anALySIs oN eLAstIN In immunostained Sections (FiGuRE 6D). the amount oF ElaStIn in oldEr animaLs was APproximAtely half That in youNg AnimALS (FiGure 6E) and reCovered iN DT-treateD aRF-DTr MiCe
 receptor (ARF-DTR) and wi ld-type mo use l ung s t re ated  wit h diphtheria t o xin(DT) or PBS, as depict ed in  F i gure 4A . Mic e in th e  i n d ica te dgro up s  w ere e uth anizedand connec ted  t o the FlexiV e nt  system th rou gh their tra che ae. Mi ce  we r e sub jec ted t o lapa r otomy,  and diap hr a gms we r e remov e d  p rior  to the assay. (B )  S t atic lung comp liance  ( C st ) was  ca lculated f ro m the slope o f  t h e pre s sure-volume l oop. Cst re f lec ts the  s tat i c elas tic r ec o ilpressure of  the  lungs at  a giv e n lungv olume.(C–F)Dyn ami c lu n gco mpl ia n ce( Cr s)( C),  dynamic  l un g res ista n c e (Rrs ) ( D),tissu e elastance ( H)(E), and  tiss ue da mpin g(G) ( F) are  show n.  Dynamic lung c ompl iance cap tur es  th eeasew ith wh ich  th e lungs  may be ext en d e d ,and dynamic lung e la s t an ce captu res th e  e la s tic rigi di tyor s t i ffnes s of th e lungs.  Tissu e  e la stancean d tiss ue  da mpi ng re f lect  energ y conser vatio n  and dissipati o n in lung alv e ol i ,  r e spec tiv ely. Values  rep r esen t me a n± S E M ofindep en d en t  experiments. Datawe re ana lyzed  1-way ANOVAfollowed b y p ost-hocTuke y -K r amer multiplecompa rison test .  *P < 0. 05, * *P < 0.0 1, ***P < 0 .001.

W e a lso  co mpa r e dlung morpholo g i es a mo ng thes e m ice. An  in cre ase  in  a lveolar s ize is a ss oc ia te d w ith a declinein  lu ng  fu nctio n  durin g agi ng ( 18 ,1 9,22, 23) .  A l v eola rme an l ine ar  inte rcep t s ( the ave rage dist anc e  bet we en  alveol ar walls that  r eflects th esiz e of t h e  alveola r space; ref. 24) werei ncrease d i n 12- mont h-old ARF -DT R mice  (F i gure 6 , A an d B).  T heD T  trea t m en t r ev ersed this p hen otype  b ut d id notaffect alveolar si z e i n wild-type m ice . Me a n  a lve o la r  se pt a l w a l l thickness was  lower in12 - mo nth-old mi c e t ha n in 2- month-o ld mi c e (Figu re 6C), a s previou sl y re p o rte d (25). Fu rthermor e, the ab l ation of  p19A RF- expres si ngcells  fully res tored  the a lv eolarwallin  ARF-DTR  mice but not in wild-t ype mi ce. I ncr eases inlun g  co mpliancein a ged animal s h ave  been  at t ribut ed t o  a  de c linein t h e numbero fela s t ic  fibers, th e m ajo r com pon e nt ofwhic h is elastin (19, 22). Therefore , we p erf orm e d aqu antitative ana lys is o n elasti nin immunost ained se ct i ons ( Figure  6D).The amo u n to f elas tininolder ani mal sw as appr ox im a tely h alfth at inyounga nima l s  (Figure 6E) and  reco v e red i n  DT -trea te d ARF-D T R mi ce
 receptor_(ARF-DTR) and_wild-type mouse lungs treated_with diphtheria_toxin_(DT) or_PBS,_as depicted in_Figure 4A. Mice_in the indicated groups_were euthanized and_connected_to the FlexiVent system through their tracheae. Mice were subjected to laparotomy, and diaphragms_were_removed prior_to_the_assay. (B) Static lung compliance_(Cst) was calculated from the_slope of_the pressure-volume loop. Cst reflects the static elastic_recoil_pressure of the_lungs at a given lung volume. (C–F) Dynamic lung_compliance (Crs) (C), dynamic lung resistance_(Rrs) (D), tissue_elastance_(H)_(E), and tissue damping_(G) (F) are shown. Dynamic lung_compliance captures the ease with which_the lungs may be extended, and dynamic_lung elastance captures the elastic rigidity_or stiffness of the lungs._Tissue elastance_and tissue damping reflect energy_conservation and dissipation_in lung_alveoli, respectively. Values_represent mean ± SEM of independent_experiments. Data were_analyzed 1-way ANOVA followed by post-hoc_Tukey-Kramer_multiple comparison test._*P_<_0.05, **P_< 0.01, ***P_<_0.001.

We also_compared_lung morphologies among these mice. An_increase_in alveolar size is associated with a_decline in lung function_during_aging (18, 19, 22,_23). Alveolar mean linear intercepts_(the average distance between alveolar walls_that reflects_the size_of the alveolar space; ref. 24) were increased in 12-month-old ARF-DTR_mice (Figure 6, A and B)._The DT treatment reversed_this phenotype_but_did not affect_alveolar_size in_wild-type mice. Mean alveolar septal wall thickness_was lower_in 12-month-old mice than in 2-month-old_mice (Figure 6C), as_previously_reported (25). Furthermore, the ablation of_p19ARF-expressing cells fully restored the alveolar_wall in ARF-DTR mice but_not_in_wild-type mice. Increases in lung_compliance in aged animals have been_attributed to a_decline in the number of elastic fibers,_the_major component of which is elastin_(19,_22). Therefore, we performed a quantitative_analysis_on_elastin in immunostained sections (Figure_6D). The amount of elastin in_older animals was approximately half that in young animals_(Figure 6E) and_recovered in DT-treated ARF-DTR mice