Document pGL97xjV9y66kqbE2Mrraq6E

JOURNAL OF AEROSOL MEDICINE Volume 1, Number 4, 1988 Mary Ann Liebert, Inc., Publishers infant, child, and )r. The journal is c, immunolc s the impac, ___________ I j I M.D. y Syndrome id Exerciseani, temic Lupus Rheumatoid nunology. npetence in I i I i Lung Clearance of Inhaled Insoluble and Soluble Particles G. OBERDORSTER Department o f Biophysics, Environmental Health Sciences Center, University o f Rochester, School o f Medicine and Dentistry, 601 Elmwood Avenue, Rochester, NY 14642 Table of Contents Introduction Overview of Clearance Mechanisms Tracheobronchial Clearance Alveolar Clearance Insoluble Particles Soluble Particles Concluding Remarks References Page 289 29L 293 298 298 307 317 320 INTRODUCTION E f fic ie n t elim ination of inhaled soluble and insoluble p articu late compounds deposited in the resp irato ry tra c t is necessary to keep it s mucosal surfaces clean and fu n ctio n a lly in ta c t. In addition, the c h a ra c te ris tic clearance processes and kinetics p revailing in d iffe re n t regions of the resp irato ry tra c t determine the retained dose of an inhaled substance in structures of the respirato ry system. Thus, whether deposition and subsequent retention of inhaled p a rtic le s is considered in c lin ic a l situ atio ns involving diagnostic or therapeutic aerosols or in toxicology with occupational or environmental pollutant aerosols, knowledge of the clearance of these substances is cru cial in understanding th e ir resp irato ry tra c t dosimetry. Clearance mechanisms are not only d iffe re n t for d iffe re n t regions of the resp irato ry tr a c t, but also depend on physico-chemical ch a ra c te ris tic s of the deposited p a rticu la te m aterial. Several recent review a rtic le s provide detailed descriptions and discussions of d iffe re n t processes governing the removal of inhaled solid and solute p a rtic le s a fte r deposition in the resp irato ry tra c t (Lauweryns and Baert, 1977; Morrow, 1977; Camner, 1980; Jones f i i a l . , 1982; Effros and Mason, 1983; Jones, 1984; Pavia, 1984; Morrow and Yu, 1985; Schlesinger, 1985; Brain, 1985, 1986; Cuddihy and Yeh, 1988). Because of the multitude and complexity of the diverse resp irato ry tra c t clearance processes, the present a r t ic le w ill s e le c tiv e ly review aspects of soluble and insoluble p a rtic le removal from tracheobronchial and deep lung regions. In addition, the a r t ic le emphasizes c lin ic a l and toxicological im plications of these removal processes and considers some unresolved issues related to lung clearance. Key words: p a rtic le s ; tracheobronchial clearance; pulmonary clearance; s o lu b ilit y ; a lveo lar macrophage; phagocytosis; p a rtic le overload; e p ith e lia l permeabi1i ty 289 Figure 1 is a schematic overview of clearance mechanisms, separated by those for insoluble and for soluble p a rtic le s . This scheme is sim plified insofar as i t assumes that insoluble p a rtic le s are not dissolved in the lung (no in vivo s o lu b ilit y ) and that inhaled soluble p a rtic le s w ill not remain in p a rtic u la te form a fte r deposition in the lung (high in vivo s o lu b ilit y ). One might predict that compounds of low water s o lu b ility are cleared from the lung more slowly than compounds of high water s o lu b ility . However, many "insolu b le" p a rtic le s show in vivo dissolution in the lung, very lik e ly within the phagolysosomes of alve o la r macrophages due to the low pH (Lundborg et al-> 1984). Thus, many solid p a rtic le s are cleared from the lungs involving mechanisms for both Insoluble p a rtic le s and solutes. Water s o lu b ilit y of an Fig. 1: Elimination of Inhaled Particles: Clearance Mechanisms Schematic representation of clearance mechanisms of Inhaled p a rtic le s deposited in the lung. "In solu b le" and "soluble" refers to in vivo s o lu b ility rather than s o lu b ility 1n water. Since most "In solu b le" p a rtic le s undergo dissolution in the lung to some degree, the dissolved material from "insolu b le" p articles and from solid p a rtic le s with rapid i_n vivo dissolution w ill be cleared by pathways lis te d under soluble p a rtic le s . Dissolution of p a rtic le s of low in vivo s o lu b ilit y can become an important clearance mechanism a fte r retention of months and years 1n the alve o la r and in t e r s t it ia l compartment. The trachea and lymph/blood c irc u la tio n are the fin a l pathways for elim ination out of the lung. Numbers 0-23 re fe r to respective generations in W eibel's lung model. 290 inhaled co lung. A compounds of fats re less than with a- r (Oberdorst very low h alftim e, a l . , 1981 mechani cal d is so lved mechanisms low water at a fast rats (Obe the lung nor in yi rate of a Mercer as a fun< particle r material J pointed oi for some needed be p a rtic le s quite d if layer of c e lls , or i ncreased possibly I The Radiologi i ncorpora lung and Y compou consi dere compounds (Class W) With this to thei r The I mechani ca general r following (Morrow, fn is tl bn the c correlate The rr region 1; ted by fied 1ung lain in ). One le lung oluble" in the Lt - . volving of an It c -Q 2 #s i'S . 3 i erst cells) ding l cells) posited rather olution r t i cl es ired by low i_n tion of hea and of the 1. inhaled compound alone is not a good predictor of it s clearance behavior in the lung. A few examples on the pulmonary retention kin e tic s of heavy metal compounds may illu s t r a t e th is : Water soluble N iCl2 is cleared from the lungs of rats rap id ly (English fit a l, 1981) corresponding to a retention halftim e of less than one day, whereas water soluble CdCl2 is retained in the rat lung with a retention halftim e of about two months due to chemical binding (Oberdorster i 1 ., 1979a). As one might expect, CdO and Ni0 p a rtic le s of very low water s o lu b ility are retained in the rat lung with a long retention halftim e, which is also about two months (Oberdorster i a l . , 1979a; English et a l . , 1981). However, whereas NiO lung retention r e fle c ts probably mainly mechanical lung clearance of the NiO p a rtic le s , the CdO p a rtic le s are rapidly dissolved in the lung (Hadley fit al_., 1980) and are then cleared by the same mechanisms and at the same slow rate as CdCl2- In contrast, ZnO p a rtic le s of low water s o lu b ilit y , which are also rapidly dissolved in the lung, are cleared at a fa st rate corresponding to a pulmonary retention halftim e of six hours in rats (Oberdorster et a l . , 1979b). Thus, i_n vivo dissolution of a p a rtic le in the lung can be d is tin c t ly d iffe re n t from water s o lu b ilit y , but neither water nor in vivo s o lu b ilit y are unequivocal predictors of the pulmonary clearance rate of a compound. Mercer (1967) proposed a clearance model based on dissolution of p a rtic le s as a function of th e ir surface area. In his model, knowledge about the p a rtic le mass d istrib u tio n and the nonequilibrium s o lu b ilit y c o e ffic ie n t of the material in lung flu id is required to predict it s clearance from the lung. As pointed out by Morrow (1974), such predictions turned out to be quite accurate for some p a rticu la te compounds, e .g .,UO2, yet much more information is needed before th is s o lu b ility concept can be applied generally to inhaled p a rtic le s . In p a rtic u la r, dissolution of p a rtic le s in the lung is conceivably quite d iffe re n t depending on whether the p a rtic le s are in the liq u id surface layer of the epithelium, phagocytized 1n alve o la r macrophages, in e p ith e lia l c e lls , or in the in terstitiu rn . For example, Lundborg fit al_. (1984) found increased dissolution of p a rtic le s of metal oxides in a lveo lar macrophages, possibly due to the low pH in th e ir phagolysosomes. The Task Group on Lung Dynamics of the International Commission of Radiological Protection (ICRP, 1966) proposed a lung clearance model which incorporates three classes of compounds with d iffe re n t s o lu b ilitie s in the deep lung and respective retention halftimes of days, weeks, and years for D, W, and Y compounds. Both absorptive and mechanical clearance mechanisms were considered, and resultin g pulmonary retention halftimes for very soluble compounds (Class D) were given as 0.5 days, for intermediate soluble compounds (Class H) as 50 days and for highly insoluble compounds (Class Y) as 500 days. With th is sim p lified clearance model, compounds could be c la s s ifie d with respect to th e ir pulmonary clearance, based so le ly on th e ir physico-chemical nature. The ICRP clearance model assumes that each clearance pathway (absorptive or mechanical) clears a p a rtic u la r compound at a constant rate. Thus, an ICRP general retention equation for the tracheobronchial or pulmonary compartment following a single inhalation exposure can be w ritten as: At - Aq Z f p exp (-bnt ) n (1) (Morrow, 1977) where A^ and A0 is the amount retained at time t and time 0, f n is the fra ctio n of A0 cleared with each of n clearance pathways, and bn the clearance rate for each of n clearance pathways; bn is inversely correlated with the retention halftime T-|/2 : OVERVIEW OF CLEARANCE MECHANISMS The main clearance pathway for insoluble p a rtic le s in the tracheobronchial region is the m ucociliary escalator consisting of the c ilia t e d epithelium f jnoving a mucous layer on top of i t . This layer is composed of a sol phase of low v is c o s ity (hypophase) in which the c i l i a beat, and a blanket of an overlaying gel phase of high v is c o s ity (epiphase) which is thought to be moved by c i l i a r y motion towards the pharynx. Although i t is generally believed that Ithis mucous blanket is continuous throughout the resp irato ry t r a c t, contrasting observations in rats have been made which led to the suggestion that th is layer may be discontinuous under normal conditions and becomes more widespread only under conditions of bronchitis (Ira v a n i and van As, 1972; van As and Webster, 1974; van As, 1980). In support of his observation of mucus is le ts in the conducting airways, van As (1980) reasoned that extremely high v e lo c itie s of tracheal mucus would be necessary to accommodate clearance of a continuous mucous blanket orig in atin g in the terminal bronchioles with th e ir large aggregated circumference. Obviously, such discontinuous mucous layer would make m ucociliary clearance of p a rtic le s deposited in the tracheobronchial region less e ff ic ie n t . Other investigators could not confirm the discontinuous mucous blanket, but did find a lack of the high v is c o s ity epiphase in the bronchioles (G il and Weibel, 1971; Luchtel, 1982) which would also reduce the e ffic ie n c y of m ucociliary movement in these generations. Clearance of p a rticle s depositing in the tracheobronchial region also occurs through phagocytosis by airway macrophages which are eith er a lv e o la r macrophages moving up with the m ucociliary escalator (Lehnert and Sanz-Rodriguez, 1988) or macrophages entering the airways v ia bronchial and bronchiolar mucosa (Morrow, 1974; Robertson, 1980). The importance of these macrophages fo r p a rticu la te clearance from the conducting airways as well as th e ir origin needs further in vestigatio n (B ra in , 1988; Gehr et a1., 1988). Another clearance mechanism, which is less important in terms of the amount being cleared in the tracheobronchial region, is penetration of Insoluble p a rtic le s of submicronic size through the epithelium (Gore and P a tric k , 1982), very lik e ly due to endocytosis by e p ith e lia l c e lls of the conducting airways (Sorokin and Brain, 1974). In contrast to the results with small submicronic p a rtic le s , no evidence was found that bigger p a rtic le s (7.9 pm) penetrated the bronchial epithelium (Velasquez and Morrow, 1984). F in a lly , cough can be a very e ff ic ie n t clearance mechanism in the conducting airways (L e ith , 1977; Kohler e l 4l . , 1986), but is probably lim ited to the upper generations of the conducting airways (Mossberg, 1980). However, formation of a flow lim iting segment ( FLS) in the trachea or upper generation bronchi due to repeated frequent coughing maneuvers could a c tu a lly lead to a decrease in m ucociliary clearance (Smaldone l I . , 1979; Smaldone, 1986). This observation corroborates c lin ic a l experience in patients with chronic obstructive pulmonary disease and asthma who show regional defects in clearance associated with the location of FLS (Smaldone, 1986). Soluble p a rtic le s depositingin the tracheobronchial tree are mainly cleared by absorptive mechanisms consisting of tra n se p ith e lia l permeation via in te r c e llu la r pathways (tig h t in te r c e llu la r junctions) or by active and passive tra n s c e llu la r transport (B h alla and Crocker, 1986). In addition, mechanical clearance along the m ucociliary escalator or cough can contribute to tracheobronchial clearance of inhaled solutes. Chemical reactions can also influence the rates of clearance of soluble substances from the tracheobronchial tree, i . e . , reactions with and binding to c e llu la r and e x tra c e llu la r components. The most e ffe c tiv e clearance mechanism for insoluble p a rtic le s in the alve o la r region 1s phagocytosis by alveolar macrophages (AM) (Fe rin al_., 1965; Green, 1973; Morrow, 1973; Hocking and Golde, 1979; Robertson, 1980; Herscowitz, 1985; van Furth, 1985; Brain, 1986) and transport in turn to the m ucociliary escalator for removal towards the larynx. Some AM may penetrate back into the in te rstitiu m (H o lt, 1980; Corry e l 1 ., 1984), from where the p a rtic le laden AM may reach the regional lymphnodes (Harmsen e l 1 ., 1985) although this is disputed (Adamson and Bowden, 1978; Lehnert t 1 ., 1986). AM - p a rtic le encountersin the deep lung could be f a c ilita t e d through the a ctiva tio n of serum components by deposited p a rtic le s which then act as chemoattractants for AM (Warheit, et a l . , 1985, 1988). Likewise, chemotropism may be responsible for migration of AM towards the m ucociliary escalator, 292 al tho Baert epi th clear fiber Baert nonfi acros remov mecha surfa respi Brai n study and b T subst the trans rate hydre 1972a for Schar deper affec vol un epi U 1atei sol ui mol e< ( cl eat read i ntei (Fig endoi endo lympl fibe Seba j wi 11 lympi cl ea resu onco lymp and ! i nte Inde' more more via Depei i ntei 1 depo: although such chemotactic movement has not been demonstrated (Lauweryns and Baert, 1977). Endocytosis by type I - and to a lesser degree by type I I - e p ith e lia l c e lls and subsequent exocytosis into the in te rstitiu m is another clearance pathway in the deep lung by which deposited smaller p a rtic le s arid fib ers can reach In t e r s t it ia l site s (Greenberg i a l . , 1972; Lauweryns and Baert, 1974; Sorokin and Brain, 1974; Brody, 1979; 1981), whereas larger nonfibrous p a rticle s (>9 are probably less lik e ly to be translocated across the epithelium (Snipes and Clem, 1981; Snipes fit a l- , 1984). For the removal of free p a rticle s from resp irato ry bronchioles and alve o la r ducts a mechanical mechanism caused by a drag or flu id flux supported by continuous surfactant production, cranial movement of the m ucociliary layer and the tid a l resp irato ry movement of the lung was suggested (Macklin, 1955; Casarett, 1960; Brain, 1970; Robertson, 1980) but never proven experimentally. The resu lt of a study by Faridy (1976) showing cranial movement of surfactant into bronchioli and bronchi during ve n tila tio n of rat lungs support th is suggestion. Transepithel1a 1 transport is the major mechanism for clearance of soluble substances from the alveolar region. In te rc e llu la r d iffu sio n al transport in the region of tig h t junctions as well as a ctiv e and passive tra n s c e llu la r transport is involved (Lauweryns and Baert, 1977). Major determinants for the rate at which these absorptive processes occur are the 1ipophi1i c i t y and h yd ro p h ilicity of the solutes and th e ir molecular size (Enna and Schanker, 1972a,b). Saturable, c a rrie r type transport processes have also been described for some anionic compounds including the amino acid glycoleucine (Lin and Schanker, 1981),and for inorganic cations such as Cd a calcium channel dependent uptake into c e lls can exist (Hinkle i a l- , 1987). Other factors affectin g pulmonary clearance rates of hydrophilic substances include lung volume, e p ith e lia l surface area and d istrib u tio n of the substance in the e p ith e lia l surfactant layer (E ffro s and Mason, 1983) which w ill be discussed la te r. Endocytosis by AM and Type I e p ith e lia l c e lls can also contribute to solute clearance from the a lveo lar space of the lung, in p a rtic u la r for large molecular weight hydrophilic compounds (B h alla and Crocker, 1987). Clearance mechanisms 1n the a lveo lar space provide an e ffe c tiv e means to cleanse the a lveo lar epithelium of insoluble p a rtic le s . Only a small fraction reaches the in te rstitiu m under normal conditions. Once they have reached the In te rs titiu m , several mechanisms can contribute to th e ir further removal (F1g. 1). These include phagocytosis by In t e r s t it ia l macrophages (IM ), endocytosis and luminal exocytosis of small (< 0.1 pm) p a rtic le s by endothelial c e lls , directed fluid flux via cle fts of in te rce llu la r junctions of lymph c a p illa r ie s , and d ire c t mechanical movement (fo r example of mineral fib e rs ) through the in te rstitiu m (Morrow, 1972, Lauweryns and Baert, 1977; Sebastien fii 1 ., 1979; Leak, 1980). Water-soluble substances that reach the in te rstitiu m , on the other hand, w ill be cleared by diffusion into blood c a p illa r ie s and to a lesser degree into lymph c a p illa r ie s depending on th e ir molecular size. The major determinants of clearance of hydrophilic solutes Into c a p illa r ie s are the S ta rlin g forces which resu lt from the balance between in travascu lar and in t e r s t it ia l hydrostatic and oncotic pressures (S ta rlin g , 1896). Since the in te r c e llu la r junctions of the lymph c a p illa r ie s are less tig h t than those of the blood c a p illa r ie s (Lauweryns and Baert, 1969) one would predict that larger sized hydrophilic solutes in the in t e r s t it ia l space - lik e proteins - are cleared p r e fe re n tia lly into lymphatics. Indeed, Meyer fii a l. (1969) calculated lymphatic uptake to be almost 40 times more e ffe c tiv e than absorption v ia blood c a p illa r ie s ; however, q u a n tita tiv e ly more of such solutes were found to be cleared via the blood c irc u la tio n than via lymph due to the high blood flow to lymph flow ra tio (Meyer fii a l . , 1977). Depending on the chemical nature of solutes, binding to e p ith e lia l or in t e r s t it ia l ce ll structures can occur and delay th e ir clearance. TRACHEOBRONCHIAL CLEARANCE Figure 2 shows a hypothetical retention curve of Inhaled insoluble p a rticle s deposited in the lower resp irato ry tra c t, i . e . , from trachea to a lv e o li. Two 293 FIGURE 2 Hypothetical Retention of P a rtic le s in the Lower Respiratory Tract I t is assumed in this case that 80X of the deposited p a rtic le mass clears rap id ly from the tracheobronchial tree (retention halftime 0.3 days) and 201 more slowly from the a lveo lar region (retention halftime 50 days). Alveolar deposition is assumed to be represented by the intercept of the slow clearance phase with the R-axis. Both tracheobronchial and a lve o la r retention could probably be described more accurately by Increasing the numbers of exponential terms in the retention equation. d is tin c t phases can be distinguished, a rapid phase which in th is case clears 801 of the deposited material with a retention halftim e of 0.3 days and a slow phase with a retention halftim e of 50 days for 201 of the deposit. The r e la tiv e amount being cleared in eith er phase depends on the amount deposited in the tracheobronchial and in the pulmonary region, which in turn is a function of the c h a ra cte ris tics of the p a rtic le and of breathing (Heyder, 1982). The rapid phase of clearance is usually regarded as re fle c tin g tracheobronchial clearance, which is presumed to be e s s e n tia lly complete 24 hours a fte r the exposure. As pointed out by Morrow and Yu (1985), a better f i t could be achieved by a three exponential term, and additional exponents in the general equation ( 1) would improve the f i t s t i l l more. Therefore, the numbers of exponents one could use is a rb itra ry since there appears to be no physico-chemical or physiologic basis for selecting a certain number. In this context, Morrow and Yu (1985) discussed also a time dependent tracheobronchial retention for which a power function could be applied to describe the retention k in e tic s . The use of exponential terms, however, is very convenient and appealing since they allow clearance kin e tics to be expressed in terms of clearance rates and retention halftim es which are e a s ily understandable. Describing tracheobronchial clearance by several clearance rates may have a physiological basis, since c i l i a r y beat frequency increases from terminal bronchioles to trachea resultin g in increasing v e lo c itie s of the mucus (Ira va n i and van As, 1972). Indeed, Hilkey et 1. (1980) estimated from studies in humans that bronchial clearance of inhaled 7.9 pm p a rtic le s was fastest in the central zone, corresponding approximately to generations 1-5, and slowest for the peripheral zone, approximately generations 14-15. Respective retention 294 h a iftlme experi me clearanc proposec experime dfchoton lines ti vivo CO bronchic when th veloci t: trachea noni nva veloci t partici i* . 19. Yu and cle i ndepen and in t veloci t to gene a veloc Using 1 predict Bot 14-15) Veloci i human (1979). mucoci' region of muc and di halftimes were 1.97 hrs, and 2.62 hrs. The intermediate zone in th e ir experiment showed a s lig h t ly shorter retention halftim e (1.70 h rs). Predicted clearance of inhaled 7.9 pm p a rtic le s based on a tri-zonal clearance model proposed by the same group (Lee i s i . , 1979) agreed reasonably well with th e ir experimental findings. Their mathematical model is based on W eibel's symmetric dichotomous lung model, a d d itio n a lly assuming that the mucous blanket which lines the airways 1s uniformly thick throughout the e n tire lung. Simulating in vivo conditions, transport rates of mucus decrease from trachea to terminal bronchioles 1n th e ir model and gave the best agreement with experimental data when the v e lo c ity of mucus in the trachea was set at 5.5 mm/min, resultin g in a v e lo c ity in the terminal bronchioles of 4.6 pm/min. The applied v e lo c ity for tracheal mucus is well within the range of rates measured experimentally with nonlnvasive radioaerosol boli techniques (Yeates e i a l- , 1975). Transport v e lo c itie s of tracheal mucus measured a fte r in tratracheal in s t illa t io n of test p a rtic le s in dogs were about 10 mm/m1n and in rats about 2 mm/min ( F e lic e t t i et a l- , 1981). Yu al- (1986) proposed a multicompartmental tracheobronchial deposition and clearance model by considering each airway generation as equivalent to an independent m ucociliary escalator with it s own length and transport v e lo c ity and in teractin g with the other escalators as a linked serie s. The resulting v e lo c itie s of mucus are sim ilar to the model by Lee et al- (1979) for trachea to generation 5, but are faster for the rest of the tracheobronchial tree with a v e lo c ity of the mucus in the terminal bronchioles of 30 pm/min (F ig . 3). Using th e ir model, Yu a a l. (1986) could improve the agreement between model prediction and the experimental resu lt by Wilkey a i a l. (1980). Both models predict the longest retention halftimes 1n zone I I I (generations 14-16) to be between 2 and 8 hours, apparently in agreement with the prevailing FIGURE 3 Mucociliary Escalator V e lo c itie s of mucus in generations 0 (trachea) to 16 (term inal bronchioles) of human conducting airways were modeled by Yu i al- (1986) and Lee et al(1979). Both models simulate reasonably well experimental results of m ucociliary escalator mediated p a rtic le clearance from the tracheobronchial region (modified a fte r Yu a i a l- , 1986). V0 and d0 re fe r to the v e lo c ity of mucus and the diameter, resp e ctive ly, of generation 0 (tra ch e a), and and di to v e lo c ity and diameter of the i-th generation. 295 view that tracheobronchially deposited p a rticle s are e s s e n tia lly a ll cleared within 24 hours. However, recent results by Stahlhofen Al- (1986) seem to suggest that a substantial portion of tracheobronchially-depos1ted p a rtic le s may remain 1n the respirato ry tra c t of humans beyond 24 hours: When a bolus of uniform 2.9 pm p a rtic le s was Inhaled with a breathholding maneuver to a volumetric depth in the lung of ~50 cm3, i . e ., corresponding to the upper generations of the respirato ry t r a c t , a s ig n ific a n t retention of 40X of the deposited p a rtic le s was observed beyond 24 hours (F ig . 4 ). This can be explained by e ith er a much longer retention of p a rtic le s in c ilia t e d structures of the tracheobronchial tree or else by penetration of 40X of the p a rticle s into nonciHated airways due to mechanisms of flow-dynamics and d istrib u tio n not yet understood (e .g ., axial core flow, nonuniform asymmetric f i l l i n g ) - in which case postulation of extended tracheobronchial retention is not required. Although prolonged tracheobronchial retention of some p a rtic le s could be explained by factors such as endocytosis of p a rtic le s by e p ith e lia l c e lls , a discontinuous layer of the mucous blanket forming site s of in e ffic ie n t clearance, or lack of a mucous epiphase in the bronchioles, the magnitude of the p a rtic le s with prolonged retention (>40X of the deposited mass) is surprising and cannot e a s ily be explained by such mechanisms. Retention due to p a rtic le phagocytosis by airway macrophages which adhere to e p ith e lia l c e lls underneath the mucous layer could possibly be a more lik e ly explanation since the d is trib u tio n of these c e lls along the airways with regard to surface area FIGURE 4 Tracheobronchial Retention of P a rtic le s a 2 coe 0,6 0J 20 0 cm3 TIME . h Retention of 2.9 pm p a rticle s in human lower resp irato ry tra c t a fte r th e ir in halation as boli to volumetric depth of 50, 70 and 200 cm3 (Stahlhofen et a i . , 1986). S ig n ific a n t long-term retention beyond 24 hours occurred even a fte r inhalation to volumetric depth of only 50 cm3, presumably reaching the upper airway generations only. The retention of the rad ioactive tracer p a rtic le s was measured by noninvasive external gamma-counting (courtesy of Dr. Stahlhofen). Vp is the depth of the aerosol pulse between larynx and the front of the aerosol bolus at end expiration, given in cm3. 296 densi Sanzthe nonm "(bac deto et i part They ini t carne norm litt peri prol requ part trac (198 an asse more regi to dedi funr s iti SZ! phy: proi muci muc< Cou Phy Inh S02 03 h2s Ci g Pha (Me 198 For density is sim ilar to that of the alveolar compartment (Lehnert and Sanz-Rodriguez, 1988). I f such macrophage mediated p a rticu la te retention in the airways can be demonstrated these c e lls then may appear as in h ib ito rs of a normally fa st bronchial clearance. However, phagocytosis of p a rtic le s (b a cte ria , antigens, e tc .) by airway macrophages serves at the same time as a detoxifying and protective mechanism, as pointed out by Brain (1988). Smaldone i t a h (1988) also reported s ig n ific a n t retention of ra d io a c tiv e ly tagged p a rticle s presumably in central airways of humans at 24 hours post inhalation. They believe that the p a rtic le s had fa ile d to cle a r from central sites of i n i t i a l deposition since the 24 hour retention pattern observed with a gamma camera correlated with the i n it i a l central deposition pattern. In addition, normal persons who deposited i n i t i a l l y more in the periphery of the lung showed l i t t l e retention in central airways at 24 hours so that movement of p erip h e rally deposited material into central regions cannot account for a prolonged central retention (Smaldone et a l . , 1988). Further studies are required to examine the i n it i a l location and disposition of the deposited p a rtic le s within the tracheobronchial tree to resolve the controversy of tracheobronchial retention beyond 24 hours. In a recent e d ito r ia l, Foster (1988) dealt in more d e tail with the question of the 24 hour lung retention as an index of alve o la r deposition. He concluded also that a more accurate assessment of aerosol deposition within central bronchi is needed to determine more p recisely the effectiveness of m ucociliary function within s p e cific lung regions. Several investigators have studied a lteratio n s of m ucociliary clearance due to physiologic, pharmacologic or toxicologic stim u li. In general, as can be deduced from the preceding discussion, the rate of m ucociliary clearance is a function of c i l i a r y a c t iv it y , the v is c o e la s tic properties of mucus and of the s ite of deposition of the inhaled p a rtic le s - which in turn depends on p a rtic le size, breathing parameters, and airway geometry. Changes in biochemical and physical properties of airway secretions and in vagal tone can a ffe c t the properties and thickness of mucus as well as the c ilia r y a c t iv it y and m ucociliary transport (P a via , 1984). Table 1 summarizes factors known to a lte r mucociliary clearance. TABLE 1 Factors Influencing Mucociliary Clearance Increase Decrease Cough Physical exercise Inhalation of inocuous p a rtic le s (high concentration, short duration) S02 O3 (man) HjSO^low concentration) Cig. smoking ( i n i t i a l l y ) Pharmaceutlcals: beta-adrenergi cs choli nergi cs ami nophyl1i n amiloride m uco lytics(?) histamine Sleep Age Inhaled gases and p a rtic le s : 02; 03(r a t ) ;<high concentration) cig. smoke (chronic) Diseases: bronchi t i s bronchi ectasi s asthma bronch. carcinoma cystic fibrosis (?) chronic flow lim itation primary dyskinesia syndrome Pharmaceuti c a ls : anticholinergics (?) anesthetics aspi ri n (Mossberg, 1980; Kenoyer et a i . , 1981; Matthys et a l .. 1983; Schlesinger et a h , 1984; Pavia, 1984; Khler et a h , 1986; Smaldone, 1986; Foster et a h , 1987). For more detailed information, see Pavia, 1984. 297 ALVEOLAR CLEARANCE I channels. ea rly stut Insoluble Particles (Ludwig, penetrate As previously discussed (F ig . 1), the most e ff ic ie n t and prominent alveolar clearance mechanism for insoluble p a rtic le s involves phagocytosis by alveolar macrophages (AM). AM are large mononuclear c e lls , representing more than 95-98% of the normal free c e ll population in the a lveo lar area. They orig in ate from bone marrow precursor c e lls and are carried as monocytes to the lungs where they can mature and m ultiply in the lung in te rstitiu m (P in k e tt fit al_., 1966; Brain et f il. , 1977; Blusse van Oud Alblas and van Furth, 1979; Johnson e i al_., 1980). AM can also m ultiply in the a lveo lar space (Blusse van Oud Alblas e i a l . , 1983; Evans fit a l . , 1986). Depending on the p a rtic le load in the a lv e o li a fte r inhalation exposure, there can be an immediate influx of monocytes from blood into the a lve o li followed by a delayed influx of macrophages from in t e r s t it ia l spaces (Adamson and Bowden, 1981). Endocytosis of p a rtic le s by AM involves the recognition of the p a rtic le , adhesion and active phagocytosis (Ueda et a].., 198i ; Ito fii f il., 1981; Lehnert and Tech, 1985). A directed movement of AM to the s ite of deposition of p a rticle s and th e ir subsequent phagocytosis by AM can be f a c ilita t e d by chemotactic movement, probably involving a ctiva tio n of the complement cascade in serum by p a rtic le s (Warheit fit a l . , 1985, 1988). Although there is no evidence that this early chem otactically fa c ilita t e d AM-particle encounter influences the long term AM mediated clearance of p a rtic le s i t may prevent free p a rtic le s from entering the in te rstitiu m via endocytosis by e p ith e lia l c e lls . Phagocytosis of p a rtic le s leads to metabolic a ctiva tio n of AM (re sp ira to ry burst) which in turn release a battery of mediators of d iffe re n t biological a c t iv it ie s including oxygen metabolites, neutrophil chemotactic factors, lysosomal hydrolases, other proteinases, prostaglandins, plasminogen activato rs and fib ro b la st growth factors (Bitterman high degre Fibrous, pa the postr (Oberdorst (Godwin an lymphnodes burden is subsequent Ferin on it s re high inha' in a high translocat lymphnodes burdens of completely 1986; Wolf Measured i nsoluble d iffe r cot halftimes hundred d a l . , 1983 clearance can better l 41., 1982; Nathan, 1987). The rate of phagocytosis seems to depend on p a rtic le size, with an optimum size around 1.5-3 pm and a slower rate of phagocytosis occurring for smaller and for larger p a rtic le s (Holma, 1967; Hahn et 41., 1977). AM which have phagocytized p a rtic le s w ill normally reach the m ucociliary escalator, whether by random or by directed movement is not known. Investigatio n of clearance mechanisms of the tra n sitio n a l zone, i. e . the in teractio n of alve o la r and m ucociliary transport mechanisms, need c e rta in ly more attention . The old concept that AM can migrate back into the in te rstitiu m a fte r phagocytosis of p a rticle s (Cummins and Sladden, 1930; Cole, 1944) was disputed by Lauweryns and Baert (1977) in th e ir comprehensive review of alve o la r clearance mechanisms and by Lehnert et a l. (1986); however, this confcept was recently re v ita liz e d by Harmsen et fil. (1985) who found in dogs c o oo that a fte r intrabronchial in s t illa t io n of AM containing eith er green or red fluorescent microspheres that individual macrophages in regional lymphnodes contained almost only p a rticle s of one fluorescent color, ra re ly both. Thus, the p o s s ib ility of alve o la r clearance of p a rtic le s within AM to regional lymphnodes should not be excluded but needs additional confirmation. Several authors suggested that AM which have entered the in te rstitiu m re-enter the "cVU `o V QC airways at the level of the c ilia te d airways (Green, 1973; Tucker fit a l- , 1973, Kilburn, 1974; Holt, 1982). However, unequivocal proof of such migratory behavior of p a rtic le laden macrophages is not ava ila b le . Free p a rticle s entering the in te rstitiu m can be carried to regional lymphnodes, entering the lymphatic c a p illa r ie s through th e ir in te r c e llu la r c le fts or a fte r phagocytosis by the lymphatic endothelium (Lauweryns and Baert, 197`7 ; Leak, 1980). There appears to be a lim ita tio n in diameter for p a rticle s which can be moved along those channels before they accumulate in the lymphnodes. For example, studies with 3-15 pm p a rtic le s in rats and dogs in d icate that 3 and 7 pm p a rtic le s in s t ille d into the lungs can be translocated to regional lymphnodes, but not 9-15 pm p a rtic le s (Snipes and Clem, 1981; Snipes et a l . , 1984). I t remains to be studied whether the lim itin g factor for transport of larger sized p a rtic le s to lymphnodes lie s in the tra n se p ith e lia l tran sfer or in the translocation within the lymphatic Mechanica particles pulmonary 1983, 198 was estim. were adjus 298 channels. Penetration of p a rtic le s through lymphnodes was reported in the e a rly studies of Drinker fit 1. (1934), and from this and other studies (Ludwig, 1971) i t appears that p a rtic le s above 10 pm in diameter hardly penetrate the nodes. Even smaller p a rtic le s of a few pm are retained to a r high degree 1n the lymphnodes (Morrow, 1972), but some w ill penetrate them. r Fibrous p a rticle s were also found to penetrate the lymphnodes and to appear in ,t the postnodal lymph c irc u la tio n , thus reaching the blood circu la tio n im (Oberdrster fit L . , 1988) and leading to a widespread systemic d istrib u tio n 'y it (Godwin and dagatic, 1970). Although the fraction of p a rtic le s penetrating the lymphnodes is minor, i t may become more important when a lung p a rticu la te burden is highly increased under conditions of high p a rtic u la te exposures which subsequently increase p a rticu la te burden of the lymphnodes. r Ferin (1972) investigated the e ffec t of the absolute lung burden of a dust >d on it s removal rate . He found in this and other studies (F e rin , 1977), that il high inhaled and in s t ille d concentrations of TO2 p a rtic le s in rats resulting ie in a high p a rticu la te burden of several mg/g lung led to a highly increased :o translocation of those supposedly innocuous p a rtic le s to the thoracic lymphnodes. In addition, several investigators reported that at such high lung in burdens of p a rtic le s , a lveo lar clearance is severely retarded or can even cease e completely (Fe rin and Feldstein , 1978; Muhle fit a l . , 1987; Bellmann et ai-, i . 1986; W olff fii a l . , 1987). This aspect w ill be discussed in more d etail la te r. id Measured pulmonary clearance rates due to mechanical clearance of highly >f insoluble p a rtic le s (1-4 ym radiolabeled fused alum ino-silicate p a rtic le s ) ia d if fe r considerably in d iffe re n t animal species (F ig . 5). Respective retention i c halftimes range from about 50-100 days in rats, mice and hamsters to several 3f hundred days in dogs, guinea pigs, and man (B a ile y et a l . , 1985a,b; Snipes et > 4i - , 1983; Snipes and McClellan, 1985). I t appears that mechanical lung 5 clearance cannot always be described by monoexponential clearance k in e tic s , but an can better be expressed as a multiphasic process. This is best illu s tr a te d by on FIGURE 5 Lung Clearance of Highly Insoluble P a rtic le s ry n. he iy um as of is gs ed es s, al al :he '3, >ry tal ar t, es he >gs Mechanical lung clearance of respirable radiolabeled fused alumino-si 1icate be p a rtic le s - excluding dissolution - was estimated in d iffe re n t species by ind pulmonary retention measurements over several hundred days (Snipes et a l . , :he 1983, 1985; B a ile y fit a l . , 1985a,b). Low i_n vivo dissolution of the p a rticle s i n was estimated from urinary excretion data of the la b e l, and retention curves :i c were adjusted for th is in vivo dissolution. 299 resulits of Snipes i a l- (1983) in rats (F ig . 5) which shows estimated mechanical clearance of inhaled 134Cs-labeled fused alum inosilicate p a rtic le s from retention measurements of up to 850 days. About 5% of the in it i a l lung burden of a rai: was estimated to be cleared mechanically with a long-term retention halftime of about 700 days, whereas the retention halftim e for most of the pulmonary partijculate burden changed with time from about 35 days during the f i r s t week a fte r exposure to about 120 days a fte r 6 months and to over 450 days a fte r more than one year. A sim ilar prolongation in retention halftimes in rats with time was reported by B a ile y i a l- (1985a), with halftimes of 32 days in the early phase of clearance and of 173 days a fte r one year. Reported monophasic retention halftimes in rats of about 50-70 days for tracer p a rtic le s in other studies (F e rin , 1978; Oberdorster t a l . , 1984; Lehnert and Morrow, 1985; Bellmann et a l . , 1986) might be due to the fa ct that clearance was followed only for 120 days or less by those in vestig ato rs. As can be seen from Fig. 5, a constant clearance rate (monoexponential) with a corresponding retention halftim e of about 60 days w ill very well describe the pulmonary retention c h a ra cte ristics of insoluble p a rticle s in the rat up to day 120. Multiphasic alve o la r clearance curves, although with d iffe re n t parameters and rates changing not as much as in the ra t, were also found to f i t observed data in dogs, guinea pigs, hamsters and mice (Snipes e i a l . , 1983; B a ile y et a l . , 1985a; Kreyling e l a l . , 1986, 1988). Likewise, Baile y e i a l. (1985b) estimated from th e ir studies in humans pulmonary mechanical clearance rates which changed with time (F ig . 6), startin g at about 4 x 10-3 per day (equivalent to a retention halftim e of about 170 days) and decreasing to a constant rate of about 1 x 10~3 per day (about 700 day retention halftim e) at 300 days post inhalation. Since dissolution of the highly insoluble p a rtic le s used in th e ir study and subsequent absorption of the dissolved material was estimated to occur at rates approaching the mechanical long-term clearance rates, this was taken into account in those studies by measurement of in v itro s o lu b ility and in vivo urinary excretion rates. In addition, an estimated clearance due to transport into regional lymph nodes was considered in the calculations of B a ile y i a l- (1985). I t appears that several hundred days a fte r inhalation of even highly insoluble p articles dissolution becomes a major clearance mechanism in the lung (Kreyling et a l . , 1988'). However, lack of knowledge of exact in vivo dissolution rates and the p o s s ib ility of chemical reactions of the dissolved labels in the lung are factors of Uncertainty in the estimates of mechanical clearance rates. -Bohning s a l. (1982) could describe e ffe c tiv e clearance rates (including mechanical clearance and disso lu tio n) of Sr-85-labeled polystyrene p a rtic le s in normal humans by two exponential phases, with 27X of the p a rtic le s being cleared with a corresponding retention halftime of 30 days and the rest with a halftime of 296 days. In th e ir opinion, this suggests the existence of two different mechanisms of AM-mediated clearance, which could also be consistent with the data of B a ile y fit a l. (1985b). Based on the resu lts of Baile y t a l- (1985b) and sim ila r results in humans with labeled teflon p a rticle s by Philipson et a l- (1985), Cuddihy and Yeh (1988) proposed a mathematical model which makes use of variab le clearance rates that are expressed in terms of the changing fraction s of the remaining lung tissue burdens cleared per day. They suggested that the mechanical clearance rates for insoluble p a rtic le s in man, M (t), vary with time according to: M (t) - 0.005e-0-02t + o.OOl (3 ) (Cuddihy and Yeh, 1988). As one can see, at late time points a fte r inhalation (>2po days) the clearance rate approaches a constant corresponding to a retention halftim e of about 700 days (see also Fig. 6). A comparison of retention data of d iffe re n t p a rticu la te m aterials (B a ile y et a l . , 1985b) and the studies described in the preceding paragraph support the view that in a given animalspecies AM mediated clearance changes with time and is independent of the p a rticu la te compound as long as the compound exhibits very low in vi vo s o lu b ility and cy to to x ic ity . Results of some other studies do not seem to f i t this concept. For example, studies in rats, dogs and humans with inhaled m etallic iron, Fe203, Fe304, UO2 orsoot p a rtic le s showed a 300 4x1 C VoLac. G 3x1 C -oe Q. 07 Co 2x 1( oo `c -oDcC 1x1 2 oco E D CL Fractions < d a ily from correction 1985b). A p a rtic le s particles t estimated d by the aut both p artic particles i as 6.9 x 1 pm p a rtic i be 3.5 x Radiologica 1 - 2 x 10- pulmonary r and Arnett l . , 1979; dissolution of the obs retention particles : with low affected t however, n mediated c toxicity a common mecf The c inhalation ,. cal from in of ftime onary week more time early ntion udies a l., is or ranee days 1ubl e > and ta i n 985a; from wi th ition 10-3 Si nee quent chi ng those etion lymph that es , I , 1 the ctors udi ng !S i n ed me of erent data umans 8) t are rdens 1ubl e ation ntion ey et view id i s ) not wi th d a FIGURE 6 A lveolar Mechanical Clearance Rates Days after Exposure Fractions of 1 and 4 pm labeled fused alum ino-silicate p a rtic le s cleared d a ily from the pulmonary region of the human lung were estimated a fte r correction for dissolution and transport to lymph nodes (a f te r B a ile y et aj.., 1985b). A s lig h t ly slower mechanical clearance rate estimated for the 1 pm p a rtic le s up to day 100 and a s lig h tly slower clearance rate for 4 pm p a rtic le s beyond day 200 was found; y e t, in view of the uncertainties in the estimated dissolution rates, these differences were not regarded as s ig n ific a n t by the authors (B a ile y et a l . , 1985b). Thus, an averaged clearance rate for both p a rtic le s is shown in this fig ure. Fractional dissolution rates for these p a rtic le s in the human lung were estimated from urinary excretion of the label as 6.9 x 10-4 per day for 1 ,pm p a rtic le s and as 2.2 x 10-4 per day for 4 pm p a rtic le s ; fra ctio n a l mechanical transport to lymph nodes was assumed to be 3.5 x 10-4 per day as proposed by the International Commission on Radiological Protection (ICRP, 1966), which is in the same range as values of 1 - 2 x 10-4 per day estimated for th is translocation pathway from dog studies. pulmonary retention halftim e in a ll three species of about 60-100 days (A lbert and Arnett, 1955; LaBelle and Brieg er, 1961; Gibb and Morrow, 1962; Cohen et a l . , 1979; Chan a l . , 1984). However, i t is not known to what extent p a rtic le dissolution contributed to these retention halftim es; in addition, the length of the observation period may not have been long enough to determine a longer retention halftim e as discussed above. Other studies with inhaled UO2 p a rticle s showed a high correlatio n of pulmonary clearance rates in rats even with low UO2 lung burden (Downs et a l . , 1967). Whether AM function was affected by the low UO2 lung burdens (45-410 pg/g lung) is not known, however, no pathological effects in the lungs were observed. Possibly, AM mediated clearance is sp e c ific for some p a rtic u la te compounds of very low t o x ic ity and of low in vivo s o lu b ilit y thus making i t d if f ic u lt to apply a common mechanical pulmonary clearance rate to a ll p a rtic le s of this kind. The clearance k in e tics discussed above were observed a fte r a single inhalation exposure to p a rtic le s of low to x ic ity (b io lo g ic a lly " in e r t " ) leading to a low p a rticu la te lung burden of a few micrograms per gram lung tissue. However, for b io lo g ic a lly a ctive (or toxic) p a rtic le s - e .g ., s ilic a - lung clearance could be quite d iffe re n t. A d d itio n a lly, e ffec ts of such toxic pairticles on AM could lead to an impairment of lung clearance of other "in e rt" p a rtic le s , as demonstrated by the prolongation of Fe20j p a rtic le (" in e r t " ) clearance a fte r long-term exposure to low concentrations of NiO and CdO (to x ic ) aerosols was observed (Oberdorster and Hochrainer, 1980a,b). As mentioned before, exposure even to so called "in e r t " or "nuisance" p a rtic le s at high concentrations resulting in lung burdens in the mg/g lung ti'ssue range can lead to s ig n ific a n t retardation or even to stasis of p a rtic u la te lung clearance. For example, i t was reported that the elim ination of p a rtic le s from the alve o la r compartment was decreased in rats exposed c h ro n ica lly to high concentrations of TiO2 p a rtic le s (Bellmann et a l . , 1986; Muhle e i a l . , 1988). The in a b ilit y of AM to elim inate the deposited p a rtic le s resulted in a much higher lung burden of inhaled TiO2 p a rtic le s than predicted friom retention data derived from lower le ve ls of exposure. Concurrently, re d is trib u tio n of p a rticle s into in te rstitiu m and lymphatic tissue was also found, confirming e a r lie r results by Ferin (1977) that a high p a rticu la te lung burden w ill lead to an increased accumulation of p a rtic le s in regional lymph nodes. As evidenced by many long-term studies in rats in which d iffe re n t dust m aterials were used, e .g ., TiO2, coal dust, carbon p a rtic le s , f ly ash, PVC p a rtic le s , diesel p a rtic le s , photocopier toner p a rtic le s , SiO2 and asbestos (Le Bouffant, 1971; Ferin, 1972; Davis e i a l . , 1978; Bolton et 4I . , 1983; Wehner fii a l.., 1983; Chan fit a l . , 1984; Bellmann et a l . , 1986; W olff fit a l . , 1987; Lee e l a l . , 1987; Muhle fit a l . , 1987, 1988), the prolongation of dust retention in the lungs of laboratory animals exposed for many months to high dust exposure le vels appears to be a nonspecific phenomenon seen with many d iffe re n t kinds of p a rtic le s including so-called b io lo g ic a lly "in e r t " dusts. Impaired p a rtic le clearance was ty p ic a lly accompanied by an inflammatory response in the resp ira to ry tra c t, macrophage recruitment and e p ith e lia l c e ll p ro life ra tio n . From such observations of nonspecific in h ib itio n of pulmonary clearance mechanisms the concept of the "overloading" of the pulmonary clearance has emerged, i . e . , the accumulation and persistence of excessive amounts of a npn-biodegradable p a rticu la te material in the lung retards lung clearance and even tu ally induces lung in ju ry . This includes - in addition to inflammatory reactions and macrophage aggregations - the development of fib ro tic changes and even lung tumors (Campbell, 1937; Martin et f i i . , 1977; McClellan fit a l . , 1986; Heinrich fii a l .. 1986; Vostal, 1986; Lee fit a l- 1985). This concept of overload implies that b io lo g ic a lly "in e rt" p a rtic le s do not e x is t, but that any p a rtic le at s u f fic ie n tly high lung burdens w ill eventually cause lung damage. Underlying biochemical mechanisms of damage include the release of mediators from activated AM and chem otactically recruited neutrophils affectin g tissue fbnctions (e .g ., released oxygen metabolites a ffectin g a lveo lar membrane in te g r ity ; Welsh fii gj.., 1986; P it t fii a l . , 1987). I t appears, that a threshold concentration of the dust in the lungs exists above which the e ffects of "overloading" occurs, and th is concentration seems to be above 1 mg of dust per gram lung tissue (Bolton fit a l . , 1983; Morrow and Mermelstein, 1988), possibly as high as 2 or 3 mg/g. Although high dust loads of "in e r t " p a rtic le s in the a lveo lar space lead to adaptive responses including influx of blood-borne macrophages and neutrophils (e a rly phase) and of p ro life ra tin g in t e r s t it ia l macrophages (la t e r phase) into the a lv e o li (Adamson and Bowden, 1981), physical "overloading" of the individual macrophages may eventually occur when they are no longer able to cope with the high dust burden, and the dust clearance function subsides (Bowden, 1987). Lauweryns and Baert (1977) described this as AM reaching an exhaustion stage dependent on the maximal capacity of the plasmalemma, and they report a value of 50% of the c e ll membrane to be capable of incorporating ingested p a rticle s CChapman-Andresen, 1963, cited in Lauweryns and Baert, 1977). Recently, Morrow (1988) hypothesized that i t is the volume of the 0hagocytized p a rtic le s per AM rather than the mass burden per gram lung which in h ib its AM function under conditions of "overload". According to this hypothesis, a maximum phagocytized volume would incapacitate AM movement. 302 Morrow (19 detailed d volume is functional 1 recrui tment of a rat that phago could lea phagocytizf overloadi nc retention at low par was deterr particles period of retention retention particles parti cles recent pil particles that AM fi highly imm hardly ca Rheologic for AM fun 1 c oC oa 4co o a) cc C ( The p artic (50 pg < particles subsequent of the re in the lu translocat lymphatic Morrow (1988) described this as the "volumetric lim it" of the AM in his detailed discussion on dust overloading mechanisms. He suggested that this volume is about 60% of the normal macrophage volume which leaves the AM fu n ctio n a lly impaired, a value which he estimated from published data on AM recruitment, lung dust burdens and dust clearance. Considering a normal volume of a rat AM of about 1000 pm3 (Lehnert and Morrow, 1984), this would mean that phagocytosis of 74 p a rtic le s with a diameter of 2.5 pm by a single AM could lead to it s volumetric overloading. A lte rn a tiv e ly , one single phagocytized p a rtic le of 10.5 pm diameterwould also re s u lt in volumetric overloading. Indeed, Snipes and Clem (1981) found in rats that the pulmonary retention halftimes (T 1/2) increased s ig n ific a n tly for large p a rtic le s even at low p a rticu la te lung burdens (F ig . 7): Whereas a normal T1/2 of 70 days was determined for 3 pm p a rtic le s , i t increased to 580 days for 9 pm p a rtic le s and was not measurable for 15 pm p a rtic le s over the observation period of 106 days. Sim ilar results were obtained in dogs where the normal retention halftime of 820 days for 3 pm p a rtic le s increased to very long retention halftimes not measurable over a 120 day period for 7and 13 pm p a rtic le s (Snipes et 1 ., 1984). No proof was provided that the bigger p a rtic le s were a c tu a lly phagocytized by AM. However, we could demonstrate in a recent p ilo t experiment in rats (unpublished re s u lts ) that 10.3 pm polystyrene p a rtic le s were re a d ily phagocytized by AM (F ig . 8). I t is e a s ily conceivable that AM f ille d with a phagocytized p a rtic le volume as shown in Figure 8 w ill be highly immobile: the normally ru ffle d surface is stretched smoothly and seems hardly capable of forming pseudopodia or of engulfing additional p a rtic le s . Rheologic c e ll properties such as cytoplasmic v is c o s ity and e la s t ic it y important for AM function (Valberg and Feldman, 1987) might also be disturbed resultin g in FIGURE 7 Lung Clearance of Monodispersed Polystyrene P a rtic le s of D ifferen t Sizes in Rats (Snipes and Clem, 1981) Days after Instillation The p a rtic le s had been in s t ille d in tra tra c h e a lly as a mixture of a ll three sizes (50 pg of 141Ce labeled 3 pm p a rtic le s ; 5 pg of 85Sr labeled 9 pm p a rtic le s and 2 pg of 48Sc labeled 15 pm p a rtic le s ) on day 0, and subsequent pulmonary retention was followed for up to 106 days for determination of the retention halftim es. The label of the 3 ^ p a rtic le s only was found in the lung associated lymphnodes, indicating a size lim ita tio n for p a rtic la te translocation either across the alveolar epithelium or within pulmonary lymphatic vessels. FIGURE 8 A lveolar Macrophage in Broncho-Alveolar Lavage of a Rat with Phagocytized 10.3 pm Polystyrene P a r tic le Parti c The monodispersed 10.3 pm p a rticle s had been in s t ille d in tra tra c h e a lly 24 hr prior to the broncho-alveolar lavage. Almost a ll of them were found to be phagocytized by AM. C ells were fixed in 2X glutaraldehyde in 0.1 m phosphate buffer and prepared for SEM. a disruption of the cytoskeleton. Possibly, in less extreme cases as that shown in Fig. 8 - i . e . , with many small p a rticle s instead of one big p a rtic le disturbances of AM f lu id it y may not be as severe. So fa r, the phenomenon of chronic p a rtic le overload has only been reported in experiments with rats in which high a ir concentrations (>5 mg/m3) of in ert p a rtic le s inhaled over weeks and months resulted in a s ig n ific a n t reduction of AM mediated p a rticu la te clearance and eventually fib ro tic lung disease. Many questions remain unanswered at this point, e .g ., is the loss in AM function reve rsib le ? Does the i n it i a l adaptive increase in AM numbers seen in acute overload situ atio ns (Adamson and Bowden, 1978) p ersist in chronic overload conditions? What is the significance of the dust deposition rate vs. the total lung dust burden? Is there a common pathogenetic mechanism of chronic in t e r s t it ia l lung disease caused by overloading with low t o x ic ity dust and by lower lung loads of cytotoxic dust? and f in a lly : what are the im plications for human exposure to high dust le ve ls? Answers to such questions w ill not only shed lig h t on mechanisms involved in overload situations but are also important for the understanding of basic clearance mechanisms and the pathogenesis of lung diseases. For example, Rom et al_. (1987) and Bowden (1987) discuss the central role of activated AM in the pathogenesis of pneumoconioses a fte r chronic exposure to high concentrations of inorganic dusts since they found under such exposure conditions an increased release from AM of AM derived growth factor and fibronectin which are important for fib ro b last a ttra c tio n and re p lica tio n . Predicted k in e tics of dust retention in the lungs during chronic high exposure conditions are shown in Figure 9 for a rat lung based on the experimental findings of several groups (Chan e i a l . , 1984; Bellmann et a l . , 1986; Wolff t al.., 1987; Muhle et a l . , 1988). At low to moderate exposure 304 The kineti and high results of Muhle l a burden siP a rti cul att particles ; At low expc a steady s rates acc particle v k = alveol. volume (Vc clearance : At the ma> predicted 1 become equ: concentrti lung or AM is quivale the ra t. F burdens ha be impaire ("volu m etri P a r t ic le a: but may ev p a rtic le s , clearance particulate Althouc particles f rats as c; large part I I FIGURE 9 P a r tic le Accumulation in the Lung During Normal and Impaired Clearance 14 hr :o be phate that cle - orted inert on of Many ction acute rload total roni c id by s for only rtant lung ntral roni c such actor on. high the a l., osure Days The kin e tics of p a rtic le accumulation in the lungs of rats during chronic low and high exposure concentrations of "nuisance" p a rtic le s are derived from results of Chan at a l. (1984), Bellmann et a i. (1986), Wolff et a l. (1987) and Muhle s i 4l- (1988). The p a rtic le burden is expressed as volumetric p a rtic le burden since th is seems to co rrelate with AM m obility (Morrow, 1988). P a rtic u la te burden in whole lung compartment re fle c ts p a rtic le s in AM, free p a rticle s in the a lveo lar epithelium and p a rtic le s in the pulmonary lymph nodes. At low exposure concentrations, the p a rticu la te burdens in the lung and AM reach a steady sta te, which can be predicted by the d a ily deposition and clearance rates according to: Vq/K[ 1--( exp-Kt) ] , where = accumulated p a rtic le volume at time t ; Vq = d a ily deposited volumetric p a rtic le burden; k - alve o la r clearance rate. At high exposure concentrations, when a c r it ic a l volume ( V r r it ) of phagocytized p a rtic le s is reached, AM mediated p a rtic le clearance begins to slow and accumulation of p a rtic le s in the lung increases. At the maximum phagocytized volume (Vmax), AM mediated p a rtic le clearance is predicted to subside and the rate of accumulation of p a rtic le s in the lung may become equivalent to th e ir deposition rate. concentrations of p a rtic le s , the lung burden - here expressed as volumetric lung or AM burden - w i l l reach an equilibrium when the pulmonary clearance rate is equivalent to the pulmonary deposition rate , i . e . , a fte r about 12 months in the ra t. However, during chronic high exposure concentrations when AM p a rtic le burdens have reached a c r it ic a l value, normal AM mediated clearance begins to be impaired and eventually may cease when a maximum phagocytized volume ("volum etric lim it " ) of p a rtic le s is reached in the AM (Morrow, 1988). P a rtic le accumulation in the lung then does not follow the predicted pattern but may eventually become lin ea r re fle c tin g the deposition rate of the inhaled p a rtic le s . Muhle a i al_. (1988) determined e m p irically that the pulmonary clearance rate of p a rtic le s decreased lin e a r ly with the logarithm of p a rticu la te mass in the lung once a c r it ic a l mass burden had been reached. Although the existence of such overload related impaired lung clearance of p a rticle s has not been demonstrated in humans i t is probably not res tric te d to rats as can be deduced from the previously mentioned retention studies with large p a rticle s in dogs of Snipes e i aj_. (1984). Furthermore, Muhle (1988) 305 observed in hamsters an impairment of p a rtic le clearance a fte r exposure to high concentrations of photocopier-toner p articles. I t is conceivable, therefore, tliat humans occupationally exposed to high dust concentrations on a chronic basis might show signs of "overload" related impaired p a rtic le clearance and lung damage (Bowden, 1987). For example, i t is well known from studies in coal miners that exposure to excessive amounts of respirable coal mine dust w ill lead to an increased risk of progressive massive fib ro s is , chronic bronchitis or emphysema (Ruckley t aj_., 1984; M ille r and Jacobsen, 1985; Rom t a]_., 1987). In e a rlie r studies, Nagelschmidt (1965) reported that in coal workers extremely high amounts of lung dust of 9 g to over 60 g correlated with the se ve rity of pulmonary fib ro s is . However, they pointed out in addition that the rank of the coal is of importance for it s fibrogenic p o te n tia l. I t is in trig uing to view this response to chronically-inhaled high le vels of coal dust as a consequence of dust overloading of AM mediated clearance in humans analogous to results of rat studies. Indeed, Freedman and Robinson (1988) found in coal workers a decreased clearance of the inhaled coal dust as one in d icato r of an e ffect of chronic exposure. The demonstrated importance of coal rank for the fibrogenic response may indicate that c y to to x ic ity of the d iffe re n t coal dusts could also play a role. In contrast to the coal dust, much lower amounts of lung dusts of 2-6 g in fib r o tic coal workers' lungs were found when 30-501 of the retained dust consisted of the more cytotoxic quartz p a rtic le s (Nagelschmidt, 1965). In view of the results of the animal experiments i t is therefore prudent not to exclude the p o s s ib ility of dust overload effects in humans. Considering a recommended threshold lim it value (TLV) in the U.S. for occupational exposure to so-called "nuisance dust" of 10 mg/m3 (ACGIH, 1987), a chronic eight hour d a ily exposure to th is permissible level could lead to dust accumulation of several mg dust per gram lung. 1 This should a le rt us to an important issue in occupational health which c a lls for further attention and re-evaluation of present nuisance dust standards by health au th o rities. Figure 10 depicts a compartmental scheme of p a rtic le translocation in the peripheral lung region under conditions of normal low exposure concentrations and high exposure concentrations (overload). Differences in translocation pathways, r e la tiv e translocated amounts and translocation rates between "normal" and "overload" are indicated as well as gaps in our knowledge for such translocations. The scheme is sim plified in several respects; P a rtic le s in in te rstitiu m and lymph nodes are placed into the same compartment as representing a common pool of p a rticle s which have crossed the alveolar membrane. Obviously, biological consequences are quite d iffe re n t whether p a rtic le s are in the in te rstitiu m or in lymph nodes. I t is assumed, that released p a rticle s from dying AM (kg) w ill be taken up by other AM under normal conditions (k ;) - a repeatedly described concept (Heppleston, 1963; Heppleston and Young, 1973; Lehnert et a l.., 1986) which is supported by recent studies of Lehnert et a]_. (1988) - whereas they remain mostly in the free p a rtic le pool under conditions of overload. Based on the suggested immobility of overloaded AM i t is further assumed that overloaded AM do not tran sfer into the in te rstitiu m U 4) but that the reported in t e r s t it ia l dust accumulation occurs via the free p a rtic le pool (kg) (Lehnert et a l . , 1986; Bowden, 1987). C le a rly , more studies are needed to elucidate the mechanism of in t e r s t it ia l dust accumulation in overload situ atio ns. The scheme also sim p lifies by Assuming (1) these are respirable p a rtic le s (mass median aerodynamic diameter = 3.5 pm with geometric standard deviation of 1.5); (2) a (co nservatively low) pulmonary deposition of 15%; (3) an inhaled volume of 10 m^ per day; (4) a pulmonary retention halftime of 500 days; and (5) a human lung weight of 950 g ), the accumulated dust concentration in the lung would reach a steady state of 11 mg dust per gram lung, a level which is known from rat studies to cause lung damage due to overload as discussed in the preceding paragraphs. Even the legal 0SHA standard in the U.S. of 5 mg/m3 respirable p a rtic le s could lead to lung dust accumulation of over 5 mg/g which is s t i l l an excessive 1e v e l. 306 showi ng p a rtic le represer under n< movement overloac acute hi Bowden, become 1 dying. (1988) 1 diesel t In ; (1986) functior pulmona; concent; (1984) i ntrodut very lc (Bolton a small prolong sequest plaques which p Baert, Mos clearan mechani transfe dying , quanti f i ntrodu This mo normal after e as well was ret whi ch saturab by Bio: possibi wi thi n speci fi p a rtic l adjusta ki neti c attempt mechani normal conseqi Soluble The number exchanc macromc addi t ic ai rborr the pr< ;h d, onic and coal wi 11 i t 1s a l., kers the the : is coal ns 988) one 3 of the lust, were lartz not ng a isure hour n of i ssue n of the tions ' on I" such s in : as ?olar !ther that inder 963; .cent free 11it y into ition 987) . itia l s by neter t of / to showing the same designation for slow clearance processes of AM containing p a rtic le s (kg) for normal and overload conditions; y e t, th is pathway represents more than one clearance mechanism which are very lik e ly d iffe re n t under normal and overload conditions, i . e . , a ctive movement of AM and passive movement due to flu id fluxes. An adaptive increase in AM numbers under chronic overload conditions - as suggested in Fig. 10 - may reasonably be assumed from acute high p a rtic le load studies (B ra in , 1971; Adamson and Bowden, 1978, 1980;' Bowden, 1987) unless the in t e r s t it ia l macrophage pool and/or blood monocytes become exhausted and cannot replenish AM at the same rate at which they are dying. This seems, however, u n lik e ly since Strom (1984) and Henderson e i a l. (1988) found that a fte r two years of exposure of rats to high concentrations of diesel exhaust the numbers of lavagable AM were doubled compared to controls. In an attempt to model overload related clearance phenomena, Yu and Morrow (1986) created a nonlinear mathematical model in which the clearance rate is a function of mass burden in the lung and which could describe increasing pulmonary p a rtic le accumulation observed in rat studies with high exposure concentrations. Likewise, the nonlinear sequestration model by Chan et a l. (1984) and Strom and Chan (1988) provides a description of overload e ffects by introducing a sequestration compartment - represented by aggregated AM - with a very long retention halftim e. Introduction of a "sequestration1' compartment (Bolton s i a l . , 1983) could accommodate both a prolonged long-term retention of a small fra ctio n of p a rtic le s observed under normal conditions (F ig . 5) and the prolonged retention under overload conditions. The anatomical co rrelate of the sequestration compartment in overload situations might be the so-called "dust plaques" (Gross and Hatch, 1962) which could be considered as a reservo ir from which p a rticle s are liberated into the a lveo lar in te rstitiu m (Lauweryns and Baert, 1977). Most recen tly, Stober (1988) proposed a multicompartmental a lveo lar clearance model for insoluble p a rtic le s which takes into account physiologic mechanistic features such as AM life tim e , decreasing macrophage mediated tran sfer rates with increasing lung dust burden, and p a rtic le release from dying AM with subsequent re-phagocytosis from the e p ith e lia l surface. The q u an tificatio n of th is la t t e r tran sfer rate with a ll it s consequences was f i r s t introduced into a model by Stober (1988) who termed i t "reverse tran sfer ra te ." This model may give reasonable predictions of both p a rtic le accumulation during normal and excessive loading and subsequent clearance or ir re v e rs ib le retention a fte r exposure cessation. A d iffe re n t model incorporating p a rtic le deposition as well as the d iffe re n t aspects of p a rtic le clearance discussed in this section was recen tly described by Smith (1985). This model features clearance rates which change with dust load due to a biological feedback system; 1t assumes saturable Michaelis-Menten kin etics for uptake of p a rtic le s into AM as described by Biozzi i a l. (1953) and takes into account dissolution of p a rtic le s . The p o s s ib ility of clearance of free p a rtic le s as well as p a rtic le sequestration within the in t e r s t it i a l compartment are also Included. Although this model was s p e c ific a lly developed for describing the clearance kin e tic s of quartz p a rtic le s and the related development of fib ro s is , it s parameters are e a sily adjustable to new data as they become availab le to model pulmonary clearance k in e tics of p a rtic le s of low c y to to x ic ity . However, these useful modeling attempts should not d is tra c t from the necessity that more k in e tic and mechanistic experimental studies and data on AM behavior and function under normal and overload conditions are needed to understand the pathogenetic consequences. Soluble Particle s (Solutes) The a lv e o la r- c a p illa ry membrane is extremely well endowed to f u l f i l l a number of d iffe re n t physiological tasks. Besides it s function of e f f ic ie n t ly exchanging oxygen and carbon dioxide, i t also provides transport of macromolecules 1n both d irectio ns. Two major physiological functions, in addition, are the formation of a b a rrie r against the penetration of harmful airborne p a rticu la te matter a fte r deposition in the alve o la r compartment and the prevention of leakage of solutes from c a p illa r ie s into a lveo lar spaces. Fig. 10: Translocation of highly insoluble particulate material of low cytotoxicity in the distal lung during chronic exposure to low (normal) and very high (overload) particle concentrations in the air. escalator Low particulate exposure Particulate burden in lung reaches steady state and is nearly equivalent to AM particulate burden (normal) mucociliary escalator blood circulation High particulate exposure Particulate burden in AM reaches steady state, particle build up in lung continues with exposure (overload) free particle pool particles in resident and newly arrived AM particles in interstitium and lymph nodes The scheme is complicated by the appearance of numerous PMN in overload conditions which also phagocytize particles yet have a much snorter life span in the alveolar space than AM. In addition, release of mediators from activated AM and PMN affects function of these cells and of the epithelial barrier. Dissolution of particles can also play a role, and the dissolution rate depends on the particulate material and on whether it is located in AM, on the epithelium or in the interstitium Translocation Pathway Flux of Particulate M a ss Fractional Transfer Rate in Mechanism of Translocation Normal Overload Overload Compared to Normal Input kn deposition of inhaled particles low high no change unless change in Jv- f j i i c i y li, particles yet have a much sfiorter life span in the . .olar space than AM. In addition, release of mediators from activated AM and PMN affects function of these cells and of the epithelial barrier Dissolution of particles can also play a role, and the dissolution rate depends on the particulate material and on whether it is located in AM, on the epithelium or in the interstitium 309 Translocation Pathway Flux of Particulate M a ss Mechanism of Translocation Fractional Transfer Rate in Normal Overload Overload Compared to Normal Input k0 deposition of inhaled particles on alveolar epithelium low high no change unless change in lung structure or respiration k 1 phagocytosis of free particles by resident AM low high lower k2 release of particles from dying AM into free particle pool low high higher Intrapulmonary k3 translocation transepithelial transport of free particles into interstitium and lymph nodes very low high no change (?) k4 migration of AM containing particles into interstitium and b lymph nodes low none (?) lower k5 fast clearance of mobile AM via mucociliary escalator high none lower Output k6 slow clearance of AM via mucociliary escalator low low lower k7 removal of free particles via very low low (?) mucociliary escalator no change (?) k8 W postnodal clearance of particles from lymph nodes low low (?) lower (?) Thus, the in te g r ity of the a lve o la r- e p ith e lia l b a rrie r is important for several physiological functions. Inhaled aerosolized soluble compounds which are deposited in the alveolar region are mainly cleared into the in te rstitiu m v ia tra n se p lth e lia l transport (F ig . 1). This transport and the resultin g pulmonary retention of these compounds are influenced by th e ir water or lip id s o lu b ilit y , th e ir molecular size and by chemical reactions (e .g ., binding to proteins, c e ll structures) in the lung. Absorption studies with non-reacting lip id - so lu b le and water soluble solutes showed that they are absorbed from a lv e o li by diffu sio n across a lipoid-pore membrane (Taylor and Gaar, 1970; Enna and Schanker, 1972). Lipid soluble substances of diverse chemical structure and degree of ionization penetrate the a lve o la r epithelium at rates which increase roughly with th e ir lip id water p a rtitio n c o e ffic ie n t, passing e a s ily through the b ilip id cell-membrane rather than the in te r c e llu la r junctions (Jones et a h , 1982). Since th is is a very rapid process the clearance rate of lip o p h ilic compounds from the a lve o la r space is lim ited mainly by blood perfusion which needs to be s u f f ic ie n t ly high to maintain a concentration gradient. However, pulmonary retention of lip o p h ilic compounds, and possibly hydrophilic compounds, can markedly be increased by adsorption onto p a rtic le s . For example, adsorption of benzo-a-pyrene (BaP) onto inhaled diesel p a rticle s increased the pulmonary retention of BaP one day a fte r inhalation from about IX,; when BaP was inhaled alone, to about 50X when i t was adsorbed onto inhaled diesel p a rtic le s (Bond a h , 1986) (F ig . 11). Since this longer retention means a prolonged exposure of lung target c e lls to BaP, i t may conceivably augment the adverse e ffe c t of this carcinogen as suggested by S a f f io t t i et a h (1968). In general, the prolongation of pulmonary retention of the solute FIGURE 11 E ffe c t of P a rtic le s on Lung Retention Pulmonary retention of a lip o p h ilic organic compound (Benzo-a-pyrene, BaP) is a ltere d depending on whether i t is Inhaled alone or adsorbed onto inhaled diesel p a rtic le s (a f te r Bond s i 41-, 1986). Although the in i t i a l dose rate of BaP delivered to target c e lls on day one is much higher when the compound is inhaled alone, adsorption onto the p a rtic le s results in a higher dose delivered d a ily to target c e lls a fte r day one. Since diesel p a rtic le s are phagocytized by AM the prolonged retention r e fle c ts e ith er BaP s t i l l on the p a rtic le s or retention of BaP a fte r desorption from the p a rtic le s in AM. 310 adsorbed the part i nteract i nip1i cat smoke physi-coenhance example, nasophar and reac In 1imited "effecti epi th eli lower rr normal . Gaar, V Nelson, derived nm and t assumes by inha endothel the reg 1976; C molecule rate lir and an exi sts via ini pinocytc particul i ntercel transcel tracheot 1986, H Ref 1 to desc barrier, and a r permeabl the cor pulmonar shows, hydrophi than th the pulr the alv contrast hydrophi impermea workers protein lung (7 protei n average transpor Base noninvas inhaled measurer ""T c - le la r ort ese llar ' in ib 1e sa ipid tion lei r ipid 32). jnds 3 be i bly 1e s . ces bout aled tion ably al-I ute 3) is haled te of nd is vered tized 3s or adsorbed onto p a rticle s depends on the physico-chemical nature of both the p a rticle s and the adsorbed compound (Bond fit i l - . 1986). Such physical interactions and resulting physiological consequences may have practical im plications for e ffects of compounds adsorbed onto p a rtic le s in cig arette smoke and in the urban and occupational environment. Likewise, physico-chemical reactions of inhaled gaseous compounds with p a rtic le s could enhance the e ffects of the gaseous substance due to a c a rrie r e ffe c t. For' example, inhaled SO2, which is normally e ffe c tiv e ly absorbed in the nasopharyngeal region, can penetrate the upper airways when i t is absorbed into and reacts with a wet aerosol (M cJilto n e i 1 ., 1976). In contrast to lip o p h ilic solutes, clearance of hydrophilic solutes is lim ited by th e ir diffu sio n rate. Several investigators have determined "e ffe c tiv e " pore sizes for water soluble substances in both the alveolar e p ith e lia l and the c a p illa ry endothelial layer by using macromolecules and lower molecular weight solutes given in tra tra c h e a lly or intravenously under normal and altered physiological conditions (Taylor et a i- , 1965; Taylor and Gaar, 1970; Enna and Schanker, 1972; Schneeberger, 1976; Jones e i a l- , 1978; Nelson, 1978; Gardiner, 1978; Egan, 1980). From results of such studies i t was derived that pore sizes of the alveolar epithelium are in the range of 0 . 6-1.5 nm and those of the endothelium in the range of 2-13 nm. The concept of pores assumes that hydrophilic solutes of low molecular weight deposited in the lung by inhalation w ill diffuse through the alve o la r epithelium and c a p illa ry endothelium via these pores, whose anatomical correlate is thought to lie in the region of the in te r c e llu la r tig h t junctions (Schneeberger and Karnovsky, 1976; Chopra fii 1979; Jones fit a h , 1982; Effros and Mason, 1983). The molecular size of the hydrophilic solutes deposited in the lung represents the rate lim iting facto r for d iffu sio nal clearance of those solutes from the lung, and an inverse relation ship between clearance rate and solute molecular weight exists (E ffro s and Mason, 1983). In addition to these d iffu sio n al processes via in te r c e llu la r tig h t junctions, transcel lu la r transport of solutes by pinocytotic a c t iv it y in e p ith e lia l c e lls occurs (Chinard, 1980). This is p a rtic u la rly important for larger solute molecules, such as proteins, for which in te r c e llu la r junctions are highly impermeable. An increased tran sfer by this tra n s c e llu la r route was reported following e p ith e lia l damage in the tracheobronchial and alve o la r region a fte r O3 exposure (B h a lla and Crocker, 1986, 1987). Reflection co e fficie n ts as a measure of osmotic effectiveness can be used to describe hydrophilic solute perm eability across the alveolar-endothelial b a rrie r, i . e . , a re fle ctio n c o e ffic ie n t of 0 means a fre e ly permeable membrane and a re fle c tio n c o e ffic ie n t of 1 represents an ideal semipermeable membrane, permeable for the solvent, but not the solute. Figure 12 shows schem atically the correlatio n between re fle c tio n co e ffic ie n ts for alve o la r epithelium and pulmonary c a p illa r y endothelium ford iffe re n t molecular solute r a d ii. I t shows, that the perm eability properties of the alve o la r c a p illa r y membrane to hydrophilic solutes 1s determined prim arily by the alve o la r epithelium rather than the c a p illa r y endothelium. Corresponding perm eability c o e ffic ie n ts for the pulmonary c a p illa r y endothelium are approximately 10 times greater than for the alve o la r epithelium (Staub, 1974). Thus, the a lveo lar epithelium, in contrast to the c a p illa ry endothelium, is only s lig h t ly permeable to hydrophilic solutes with molecular weights above 15000 daltons and e ss e n tia lly impermeable to molecules above 40,000 daltons (Jones fii aj.., 1982). Several workers suggested that the normal alve o la r b a rrie r is completely impermeable to protein (Egan fii 1977; Nelson fii al_., 1978; Egan, 1982) lik e in the fetal lung (Adams, 1966). P h y sio lo g ic a lly , however, a f in it e perm eability for protein is present, and the results of several studies in dogs showed an average clearance rate for protein of 1% per hour (Staub, 1983). Pinocytotic transport may contribute to this rate, as mentionedabove. Based on these physiological p rin cip les of e p ith e lia l perm eability, a noninvasive method of determining the in te g rity of the alve o la r epithelium with inhaled hydrophilic solutes has been developed. The technique makes use of the measurement of the pulmonary clearance rate of a small molecular weight tracer, 99mTC-iabeled diethylenetriam inepentaacetic acid (DTPA), a stable chelator 311 FIGURE 12 Reflection C oefficients of Alveolar Epithelium and Pulmonary C a p illa ry Endothelium for Hydrophilic Solutes 4 S 5` c t\Ao Solute Molecular Radius, nm The molecular radius for NaCl2 is -0.23 nm, for albumin -3.6 nm (modified a fte r Jones s 4! . , 1982). The re fle c tio n c o e ffic ie n t is the ra tio of the actual osmotic pressure and the ideal osmotic pressure for a given concentration of the solute: A re fle c tio n c o e ffic ie n t of 1 represents an ideal semipermeable membrane, and of 0 a fre e ly permeable membrane. The alve o la r retention halftim es, Tl/2, shown for the solute diethylenetriam inepentaacetic acid (DTPA) with a molecular radius of 0.57 nm, indicate a value of about 60 min for the normal a lve o la r epithelium which w ill decrease su b sta n tially for the damaged a lveq lar epithelium, shown here as a hypothetical dotted lin e . In theory, the alved lar epithelium could be damaged to a degree that solute clearance becomes lim ited by the endothelial b a rrie r of the pulmonary c a p illa r ie s , provided, the capi T1ary endothelium remains unaltered in such situ atio n s. (Chopra e i a l- . 1979; Rinderknecht at a l - . 1980; Jones a t 41-, 1982; Coates and O'Brqdovich, 1986). As pointed out by Effros and Mason (1983), i t is necessary that solutes used in th is method, lik e DTPA, do not react or bind to sp ecific site s or cross a c tiv e ly the e p ith e lia l membrane. This tracer is inhaled as an aerosol of submicronlc p a rticle s so that they are deposited p rim arily in the alvecilar region of the lung. Upon deposition of the solute droplets, they mergq with the flu id lin in g the a lveo lar surface, a process which is possibly influenced by surface tension. The molecular size of DTPA of 492 daltons corresponds to a molecular radius of 0.57 nm (Jones a t a l . , 1982) and thus DTPA can e a s ily penetrate the endothelial layer (F ig . 12). However, i f unaltered, the tig h t a lveo lar epithelium represents a b a rrie r to DTPA diffu sio n so that it s Clearance occurs at a rate corresponding to a halftim e of about 60-85 minutes in humans (Mason et a l- , 1985; Dusser et a l - , 1984; Rees et a l - . 1985; Mark^ at a l - , 1985; Langford a t a l- , 1986; U te ll a t 4 l ; , 1985). This clearance rate can be determined noninvasively in vivo by gamma camera imaging of the aerosolized 99mjc_Qjp/\ deposited in the lung, representing generally f ir s t 312 order clea in the bl( i ndi cated the reflec lungs beco been demon origins-, di seases, (Chopra a t 1982; Dus: Kehrl a t a Alveol when no 1 volumes ( 1986). Tl alveolar : deposi tion Effros an soluble pa C where ct = c Co = 1 P =p S =c V =\ I f the ra changes ii i ndependei i ncrease alveolar ( Wool man a Figurf 99mTc-iab ( di fferent min there (Oberdorsi ncreased lung clea Two mi nut at normal change su clearance and clear Fig. 13). alveolar an increa However, additio n a ' could pos (E ffro s ar Such from resu DTPA d iffi lung volu: i nfl uence aerosol spreading di ffusion 1 i order clearance k in e tics since there is i n i t i a l l y l i t t l e build up of a c t iv it y in the blood compartment due to rapid elim ination of DTPA via the kidney. As indicated 1n Figure 12, DTPA retention halftimes w ill become much shorter when the re fle c tio n c o e ffic ie n t of the a lveo lar membrane decreases, i . e . , when the lungs become more "leaky" due to damaged a lveo lar epithelium. Such damage has . been demonstrated in acute and chronic lung in ju rie s of d iffe re n t pathogenetic o rig in s, including inflammatory reactions, edema, in t e r s t it ia l pulmonary diseases, pulmonary emboli, asthma, cig arette smoking, and ozone exposure (Chopra a l . , 1979; Rinderknecht et 1-, 1980; Mason, 1985; Dolovich s i a l- , 1982; Dusser i t a l - , 1984; Mason i t a l . , 1985; Buxton-Thomas e i a l . , 1986; Kehrl i t a l . , 1987). Alveolar clearance of DTPA can also increase under physiological conditions when no lung in ju ry is present, e .g ., during exercise and at increased lung volumes (Melgnan i t a l- , 1986; O'Brodovich s i a l . , 1986; Hoolman et a l- , 1986). Thus, DTPA clearance does not only r e fle c t the perm eability of the a lveo lar membrane since those and other factors including s ite s of aerosol deposition and surface area can s ig n ific a n tly influence DTPA clearance rates. Effros and Mason (1983) described the clearance kin e tic s of inhaled water soluble p a rticle s in analogy to CO d iffusion studies by the relation ship Ct - Coe-< PS/V)t (4 ) I where ct - concentration in lung at time t (pg/g) C0 - in i t i a l concentration in lung (pg/g) P - perm eability of the alve o la r epithelium (cm/sec) S e p ith e lia l surface area (cm2) V * volume of solute d istrib u tio n (cm 3 ; e p ith e lia l lin in g flu id volume) I f the ra tio S/V remains constant, the changes in solute clearance re fle c t changes in perm eability of the membrane only and clea'rance rates are r e la t iv e ly ed independent of changes in surface area (E ffro s and Mason, 1983). However, an UQ increase in surface area w ill also increase solute clearance across the alve o la r epithelium as seen in studies with increased lung volumes (Egan, 1982; Woolman s i a l . , 1986; Rizk s i a l . , 1984; Marks s i a l . , 1985). on Figure 13 shows results of an experiment in our laboratory in which 'A) 99mTC-labeled aerosols were delivered for two minutes to anesthetized dogs at he d iffe re n t lung volumes, and lung clearance of the solute was followed for 30 led min thereafter at eith e r spontaneous breathing or at increased lung volume he (Oberdorster s i a l - , 1985): When the solute aerosol was delivered at an les increased lung volume (tid a l volume 55 ml/kg body weight, b in Fig. 13) solute he lung clearance was increased s ig n ific a n tly over control levels (a in Fig. 13). Two minute v e n tila tio n at this same high lung volume prior to aerosol d e liv e ry at normal lung volume (tid a l volume 13 ml/kg body weight, c in Fig. 13) did not change subsequent solute clearance. The most dramatic increase in lung solute clearance was seen when the solute aerosol was delivered at normal lung volume .nd and clearance was measured under conditions of increased lung volume (d in ry Fig. 13). In this la t te r case, the ra tio S/V in Eq. (4 ) increased since the ic a lve o la r surface area, but not the volume of d istrib u tio n increased, and thus an an increased surface area may p a rtly explain the increased clearance rate. he However, results from flu id in s t ille d lungs suggest, that lung h yperinflation iey a d d itio n a lly increases the perm eability of the membrane, P in Eq. (4 ), which >iy could possibly be due to stretching of the in te re p ith e lia l tig h t junctions ms (E ffro s and Mason, 1983). PA Such junctional stretching during lung hyp erinflation is also suggested id, from results of studies in rats with i . v . administered DTPA where the amount of iat DTPA d iffu sin g into the alve o la r space was found to be increased with increased 85 lung volumes (F io r ic a s i a l . , 1988). In addition, high lung volumes might i5; influence the d iffu sio n c h a ra c te ris tic s of the solute - when delivered as ice aerosol - into the surface lin ing flu id , i . e . , i t may increase the d iffu sio n al :he st spreading and thus increase the surface area a va ila b le for tra n se p ith e lia l diffu sio n (O'Brodovich and Coates, 1987). Spreading on a f la t surface occurs 313 FIGURE 13 Lung Volume and Solute Clearance abed A lveo lar clearance rates in dogs of 99mTc-DTPA administered for 2 min as an aerosol show the influence of lung volume (Oberdorster et a l . , 1986b). a: DTPA aerosol d e live ry at normal lung volume, clearance measurement at normal lung volume, b: DTPA aerosol d e live ry at high lung volume, clearance measurement at normal lung volume (s ig n ific a n tly d iffe re n t from a, p < 0.05). c: Two minute v e n tila tio n at high lung volume prior to DTPA aerosol d e liv e ry at normal lung volume and clearance measurement at normal lung volume. d: DTPA aerosol d e liv e ry at normal lung volume and clearance measurement at high lung volume (s ig n ific a n t ly d iffe re n t from a, p < 0 .02). u su ally rap id ly , within a few seconds, and thus should g rea tly exceed the lung clearance rate of the solute across the alve o la r epithelium (E ffro s and Mason, 1983). However, the hydrophobic surface of the alve o la r lin in g flu id could conceivably slow down the d iffu sio n of hydrophilic solutes into the lin in g flu id (O'Brodovich and Coates, 1987), a facto r that needs further in vestig a tio n . I t is in terestin g to speculate that d e liv e ry of the solute aerosol at high lung volumes in our study described above (b in Fig. 13) might have led to deposition of the p a rtic le s in a lv e o li with less lin in g flu id (lower volume of d is trib u tio n , V in Eq. (4) since they are not in flated during normal v e n tila tio n , thus explaining the observed fa s te r lung clearance rate. Although applying this technique of noninvasively measuring an index of a lve o la r perm eability with 99mTc-DTPA aerosols c lin ic a lly is very appealing, several unresolved questions require further basic research for Interpreting resu lts from th is method before i t should be used ro u tin ely. O'Brodovich and Coates (1987) also emphasized in a recent review that this method remains an experimental in v e s tig a tiv e tool that is not yet ready for widespread c lin ic a l ap p licatio n . For example, c la r ific a t io n is needed on questions related to the sig n ifica n ce of lung surfactant; the s t a b ilit y of the 99mTc-DTPA complex (Nolop i & 1., 1986; Haldman et a l . , 1987; Dolovich al_., 1987); unexpected decreases in clearance rates in the beginning phases of an inflammatory reaction in the lung (Oberdorster et a l . , 1986a; P la in e r and Morrow, 1988); the Importance of DTPA build-up in the blood compartment during accelerated lung clearance (Jones a l al- 1982; Langford et a l . , 1986); the rep ro d u cib ility w ithin and between laboratories and standardization of the technique (O'Brodovich and Coates, 1987). 314 mark, voi u .meas larg such of ( stab phar (di f phys has wi th Ober half stud absc and i nha spec the sol i to betv Spe sol mou Res the lip (af ; an DTPA lung it at nute lung osol il ume lung son, ould ning ther lute ight luid ri ng : of ing, ting and ; an ical the pl ex :ted tory the lung lity i que I t is conceivable that the DTPA molecule is too small, representing a marker which is too sen sitive towards small physiological changes in lung volume thus making i t d if f ic u lt to use i t as a routine diagnostic tool for measuring an indicato r of alve o la r e p ith e lia l perm eability. A solute tracer of larger molecular weight might be better suited for th is purpose. The choice of such tra ce r depends on how it s clearance is influenced by factors such as: s ite of deposition in resp irato ry tra c t; hydrophi1i c i t y , 1ipophi1i c i t y ; chemical s t a b ilit y and reactions (binding, receptors); lung disorders and administered pharmaceuticals (see Jones, 1984 andNewhouse gt a l . , 1987); recircu la tio n (d iffu sio n gradient, e .g ., influenced by renal elim in a tio n ); lung volume; physical exercise. With regard to pulmonary solute clearance in d iffe re n t animal species i t has been found that clearance of inhaled DTPA in dogs is faster than in humans, with pulmonary retention halftimes o f about 30 minutes (Rizk et al_., 1984; Oberdorster g i 1 ., 1986b). On the other hand, respective DTPA retention halftimes were longer in rabbits (T ]/2 - 127 min, Je ffe r ie s e l a J . , 1984). A study in d iffe re n t species with a range of inhaled solutes showed that absorption of hydrophilic solutes was fastest in the mouse, followed by the rat and was slowest in the rabbit (Schanker gt a].., 1986). In contrast, a given inhaled lip o p h ilic solute was absorbed at about the same rate in a ll three species (F ig . 14). Thus, lung clearance of hydrophilic solutes depends on both the animal species and the solute molecular weight whereas for lip o p h ilic I solutes i t seems to depend only on th e ir Iipid/w ater p a rtitio n c o e ffic ie n ts and to be independent of animal species. Since the ra tio of clearance rates between species was found to be constant for hydrophilic solutes, re fle c tin g FIGURE 14 Lung Clearance of Inhaled Solutes in D ifferen t Species Species dependency was observed for hydrophilic, but not for lip o p h ilic solutes. Ratios of clearance rates of d iffe re n t hydrophilic compounds for m ouse:rat:rabbit were remarkably constant with approximately 0 .4 :1 :2 .5. Respective clearance rates for d iffe re n t hydrophilic compounds increase with the molecular size of the compound. In contrast, clearance rates for lip o p h ilic compounds increases with th e ir Iipid/w ater p a rtitio n c o e ffic ie n t (a f te r Schanker e i a l . , 1986). the species sp e cific porosity of the epithelium, i t may be possible to predict the rate of pulmonary absorption of a given drug in one species from data obtained for the same drug in another species (Schanker et 1-, 1986). A leaky a lveo lar epithelium in diseased lungs leading to increases of the clearance of hydrophilic solutes from the alveolar space Implies that lung retention of soluble compounds given as aerosols is decreased compared to a healthy lung, e .g ., a therapeutical aerosol could have a s ig n ific a n tly shorter residence time - and thus is less e ffe c tiv e - at a target s ite in the lung than would be expected from its kin etics under healthy conditions. Encapsulation of such therapeutic agents in p a rticle s with membranes of low in vivo s o lu b ility could increase th e ir pulmonary retention. Obviously, such modifications depend also on whether a systemic d istrib u tio n of an inhaled therapeutic agent or a more localized e ffe c t in the lung is desired (Lourenco and Cotromanes, 1982). Chemical reactions influencing pulmonary retention of solutes (F ig . 1, Table 2) are of p a rticu la r importance for inhalation toxicology. Many water soluble inhaled substances w ill not be cleared from the a lv e o la r space simply by d iffu sio nal processes but may react with lung flu id s or c e lls , e .g ., being hydrolyzed to less soluble compounds or bound to proteins. Resulting clearance rates are unpredictable and can also d iffe r quite s ig n ific a n tly among animal species. Conversely, substances with low water s o lu b ility can be cleared from the lungs quite rap id ly because th e ir i_n vivo dissolution in the lung is very rapid. As mentioned in the Introduction, examples both for a fa st lung clearance of water insoluble compounds (high in vivo s o lu b ilit y ) and of low lung clearance of water soluble compounds (chemical binding) are found a fte r inhalation of heavy metals. For instance, inhaled aerosols of ZnO and Pb (OH)2 (low water s o lu b ilit y ) and other lead compounds were found to have a monoexponential retention halftime of a few hours only (Bianco i a l- , 1974; Oberdorster i a i . , 1979; Morrow et a l . , 1980). Inhaled aerosols of CdCl2 (water so lu b le), on the other hand, showed a much longer pulmonary retention than predicted from th e ir water s o lu b ility and molecular size. Moreover, pulmonary retention of CdCl2 a fte r a single exposure is s ig n ific a n tly d iffe re n t between ra ts , dogs, and monkeys (F ig . 15, Oberdorster and Cox, 1988): rats exhibited a monoexponential clearance pattern corresponding to a halftime of 85 days, in dogs 70X of the deposited CdCl2 was cleared with a retention halftim e of 153 days and in monkeys a monoexponential clearance with a retention halftim e of 818 days was found over an observation period of 650 days. Although th is species dependent pulmonary clearance of water soluble Cd-compounds appears to be very sim ila r to the pulmonary clearance c h a ra cte ris tics of highly insoluble p a rtic le s (F ig . 5) - which is related to AM function - pulmonary clearance of cadmium is not mediated by AM. This was demonstrated in rats when during long-term inhalation of CdO aerosols the a lve o la r clearance of Fe203 test p a rtic le s was s ig n ific a n tly retarded, but not that of a CdCl2 test aerosol (Oberdorster and Hochrainer, 1980). Hater insoluble CdO p a rticle s are cleared with the same rates as CdCl2 . due to a rapid in i t i a l in vivo dissolution of those p a rtic le s , probably in the phagolysosomes of AM (Hadley i a l . , 1980; Lundborg 1 ., 1984). Following th is rapid in vi vo dissolution of CdO, subsequent elim ination occurs at the slow cllearance rate of soluble cadmium compounds. This slow clearance may involve binding to serum and c e llu la r proteins, most lik e ly including m etallothionein (Oberdorster, 1986) which is induced in the lung due to cadmium exposure (Post f i i i l . , 1982). The measured pulmonary retention halftimes of soluble p a rtic le s are undoubtedly the resu lt of several clearance processes whose r e la tiv e contributions change depending on the physico-chemical properties of a sp ecific p a rticu la te matter. These processes - whether reactions with s p e c ific ligands, d iffu sio n rates across membranes, dissolution rates in AM, or mechanical transport rates - are of d iffe re n t magnitude in d iffe re n t mammalian species and thus make i t d if f ic u lt or even impossible to extrapolate d ir e c t ly results of pulmonary retention measurements of a sp e cific compound in animals to humans. FIGURE 15 ta Pulmonary Retention of Inhaled Water Soluble CdCl2 P a rtic le s in D ifferen t Species (Oberdorster and Cox, 1988) he ng a er an of ty nd a 1, er iy ng ce ;al om ry ng ow er Pb a 4; :12 Retention measurements were started one day a fte r exposure to CdCl2 aerosol. on Pulmonary cadmium retention 1n rat and monkey could be described by a r, monoexponential term with biological retention halftimes of 85 days and 835 ]y days, resp e ctive ly; in dogs, a biological retention halftim e of 153 days plus a constant fitte d the data best. un a CONCLUDING REMARKS s. ile Basic differences exist in the clearance of insoluble and soluble inhaled ce p a rticle s from the respiratory tra c t. Conducting airway clearance of insoluble AM p a rtic le s is fa st and maybe e s s e n tia lly complete within one day, although 'as results of recent studies in d icate that retention in the conducting airways may he be su b sta n tially longer than generally presumed. Possibly, airway macrophages 1ut which phagocytlce p a rticle s in this region may act as "in h ib ito rs " of an :er otherwise fa st bronchial clearance of insoluble p a rtic le s . Inhaled soluble a p a rtic le s are cleared from both the conducting and the a lve o la r zone at a very :he rapid rate with corresponding retention halftimes of minutes or hours, depending ng on lipophf H c it y . h yd ro p h ilic ity , and molecular size. However, binding with and he to tissue and c e llu la r components and adsorption onto highly insoluble p a rticle s lay can increase retention halftimes s ig n ific a n tly to days and months. In vivo ng retention of an Inhaled hydrophilic solute 1s longer in the conducting airways urn than in the a lve o la r region (Brown and Schanker, 1983; Oberdorster e a l . , 1986b), although pore equivalents seem to be of larger size in bronchial than ire alve o la r epithelium (Gatzy and Boucher, 1982). This longer retention seen i_n ve vivo is possibly due to differences in the thickness and composition of the Me liq u id layer lin in g the e p ith e lia l surfaces-of a lv e o li and conducting airways. Is, Alveolar clearance mechanisms for solute p a rtic le s involve mainly d iffusion al and transport via in te r c e llu la r and tra n s c e llu la r pathways, whereas insoluble ind p a rtic le s are transported up the m ucociliary escalator mostly a fte r phagocytosis of by a lveo lar macrophages, and a minor fra ctio n may leave the lung via this pathway as free p a rtic le s . E ffe c tiv e pulmonary clearance rates of inhaled p a rticu la te compounds are determined by th e ir mechanical clearance rate , th e ir dissolution rate and the clearance rate of the dissolved compound. The la tte r can involve d iffu sio n , active transport or chemical reactions in the lung which 317 - lik e the other factors - can be d iffe re n t in d iffe re n t animal species. I t is therefore d if f ic u lt to determine or to predict mechanical clearance rates of p ir t ic le s with unknown in vivo s o lu b ility and r e a c t iv it y from in vivo retention measurements 1n a given animal species. Estimated mechanical pulmonary clearance rates of solid p a rtic le s showed a species dependency and became slower with time. However, some uncertainties remain with regard to the accuracy of the estimates of in vivo dissolution rates and transport rates of p a rtic le s into lymph nodes which need to be known for calcu latin g mechanical clearance rates. Although the mechanical clearance rate of insoluble p a rtic le s is very slow for the alve o la r compartment compared to the tracheobronchial region, i t is s t i l l very e ff ic ie n t in handling p a rtic u la te burdens resu ltin g from inhalation of typ ical dust concentrations in ambient a ir . However, studies in rodents showed that under extremely dusty conditions lastin g for an extended period of time AM mediated clearance becomes increasingly impaired resultin g in the development of chronic lung disease. Transepithe 1ia l transport of solid p a rtic le s across the alve o la r epithelium occurs normally at a low rate only, but can increase s u b sta n tially under conditions of high p a rtic u la te lung burdens. I t appears that whenever AM-mediated p a rtic le clearance is impaired, an increased penetration of p a rtic le s into the in te rstitiu m takes place and an increased build-up of p a rticu la te burden in the regional lymph nodes occurs (e .g ., Ferin and Feldstein , 1978). Conceivably, AM function may be affected s y n e rg is tic a lly by a combination of high exposure concentrations and c y to to x ic ity of the inhaled compounds, e .g ., in halation of high concentrations of quartz and asbestos (F e rin , 1972; Ferin and Leach, 1976) or of diesel exhaust with it s many chemicals of known and unknown tp x ic ity in both the gas and p a rtic u la te phase (Chan e i a l . , 1984; W olff s i 1 ., 1987). Under such conditions, AM clearance function should be affected at lower Pjarticulate lung burdens than at those with "nuisance" p a rtic le s of low to x ic ity lik e TiO2- The results of Wolff fit al_. (1987) seem to support this view since exposure of rats for 18 months to low concentrations of diesel exhaust resu ltin g 1n a lung burden of 320 pg soot/lung - below a concentration of p a rtic le s of low to x ic ity in the lung known to cause impaired lung clearance in rats - led to an accelerated accumulation of soot upon furth er exposure. Diseases of the lung can a ffe c t clearance of soluble and insoluble p a rtic le s d iffe re n tly : elim ination of inhaled insoluble p a rtic le s is retarded, whereas elim ination of inhaled solute p a rtic le s is accelerated. This is demonstrated 1n Figure 16 in which the e ffe c t of smoking on alve o la r lung clearance of highly Insoluble and highly soluble p a rtic le s 1s shown. Long-term lung clearance of trace amounts of magnetite test p a rtic le s was s ig n ific a n tly decreased 1n smokers (Cohen fit aj.., 1979), whereas " mTc-DTPA solute lung clearance in smokers was s ig n ific a n tly increased (Jones fit 4].., 1980). In addition to showing the q u a lita tiv e differences in the e ffe c t of smoking on a lve o la r lung clearance, differences in the rate of clearance between the two p a rtic le types become apparent from the time scale, months for the solid p a rtic le s and minutes for the liq u id p a rtic le s (F ig . 16). Obviously, such differences are re a d ily explained by the d iffe re n t underlying clearance mechanisms for these two types of inhaled p a rtic le s . However, in both cases AM probably play a central role in the mechanisms of altered alve o la r clearance in lungs of smokers and nonsmokers: Solid p a rtic le clearance decreased in smokers due to impaired AM function (Green, 1985), and solute clearance may have increased due to e ffects on the alve o la r e p ith e lia l b a rrie r of mediators Released from activated AM (Jones fii al.., 1983). As already mentioned, the im plications for diseased lungs - in this case, smokers' lungs - are that retention of water soluble therapeutic aerosols in the deep lung may be less than anticipated and that - in contrast - smokers accumulate s ig n ific a n tly more of deposited nuisance dust or other solid p a rtic le s in the alve o la r region than nbnsmokers. The la t te r was corroborated by findings of an increased retention o;f p a rticu la te material in the lungs of smokers (Churg and Wiggs, 1987). However, i t is possible that this greater retention and resultin g dust accumulation in lungs of smokers could somewhat be o ffs e t by a lower pulmonary deposition due to a greater central deposition in th e ir conducting airways i f chronic bronchitis is present. 100- 80 c o c 4*-)> 6 0 4) QL c 4oL_) 4 0 4) n. 20 0 In lung p a rtic l 1985); of oth T63O4 ni ne n conclud (Cohen si mi la r smokers solute alveola of thi: smokers and 20 nonsmok fit 4 L.. Kno lung is occupat doslmet humans determi bases c particu importa Is ; of tion nary ower / of i nto s. slow : is tion ents d of the ol id n ly , 1ung red, d an curs n of -9 -. and nown a l- , ower ci ty vi ew aust i of in uol e ded, is lung term ntly lung In on two olid such ance s AM 3 in kers have tors the that 1ess more than tion 37). dust nary ; if FIGURE 16 Smoking and Lung Clearance Smoking and Lung Clearance of Particles Solid Particles (Fe3 0 4 ) Liquid Particles ( 99mT c-D T P A ) In lung In ju ry caused by smoking, pulmonary clearance of highly insoluble test p a rticle s ^ 304) is retarded probably due to decreased AM function (Green, 1985); predictably, smokers w ill accumulate in addition to smoke p a rtic le s more of other inhaled p a rticle s in th e ir lungs than nonsmokers. Retention of Fe304 te s t p a rtic le s was measured noninvasively by magnetopneumography in nine nonsmoking and three heavy smoking male subjects, and the authors concluded that smoking was the cause of retardation of test p a rtic le clearance (Cohen e l 1 ., 1979). Bohning et al_. (1982) came to the same conclusion in a sim ilar study a fte r inhalation of radiolabeled polystyrene test p a rtic le s by smokers and nonsmokers. In contrast, pulmonary clearance of the hydrophilic solute DTPA inhaled as an aerosol is increased in smokers due to a more leaky alve o la r epithelium. The DTPA study (Jones et al_., 1980) was the f i r s t study of this kind measuring lung clearance of ^ mTc-DTPA with a gamma camera in smokers and nonsmokers. Mean retention halftimes were 59 min for nonsmokers and 20 min for smokers. The result of this study in ten smokers and ten nonsmokers was confirmed la te r in other studies (Dolovich s t 4].., 1982; Mason fii 41-, 1983; Mason, 1985). Knowledge of lung clearance processes of inhaled p a rtic le s in the healthy lung 1s essential for the understanding of e ffects of inhaled environmental, occupational and c lin ic a l aerosols. I t is also a necessary presupposition for dosimetric extrapolation modeling of results from animal inhalation studies to humans since retained doses of substances at pulmonary target site s are larg ely determined by th e ir clearance behavior. Unraveling of c e llu la r and molecular bases of lung clearance mechanisms requires considerable additional work. In p a rtic u la r, the mechanisms of clearance behavior in lung diseases are an important issue which needs further attention. ACKNOWLEDGEMENTS The author g ra te fu lly acknowledges the helpful review and valuable suggestions by Dr. P.E. Morrow, Dr. J . Ferin, Dr. M .J. U t e ll, and Dr. S.C. Soderholm. Special thanks goes to P.A. Morse and N.L. Corson for helping to prepare the figures of th is a r t ic le . 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