Experimentally Induced Emphysema

A number of insults, including oxides of nitrogen, cadmium salts,
whole cigarette smoke, ozone exposure, and proteolytic enzymes,
have been used to induce or augment emphysematous lesions in a
variety of experimental animals. The reader interested in a compre-
hensive review of this topic is referred to the well-documented article
by Karlinsky and Snider (1978). The evidence for each of these
insults, as they pertain to cigarette smoke, is reviewed below.

Oxides of Nitrogen

Oxides of nitrogen, present in the gas phase of smoke, appear to
induce or potentiate emphysema-like lesions in some animals
(Karlinsky and Snider 1978).

Nitrogen dioxide (NO,) exposure causes airway narrowing and an
increase in the proteolytic burden within the lungs of experimental-
animals. Airway narrowing is postulated as a contributing factor in
the pathogenesis of pulmonary emphysema (Juhos et al. 1980).-
Following exposure to NO*, rats develop bronchiolar stenosis. The
nitrogen dioxide exposure also induces an influx of alveolar macro-
phages (AM) and polymorphonuclear leukocytes (PMNs) (cells
known to contain proteolytic enzymes) into the lungs (Juhos et al.
1980). Kleinerman et al. (1982) demonstrated an increased number of
alveolar macrophages and PMNs in lung lavages from hamsters
exposed to NO,. Although there is no detectable increase in the
elastolytic activity from lung lavages of NO,-exposed animals, the
cell-free culture medium from macrophage cultures of NO,-exposed
animals does show a twofold to fivefold increase in elastolytic
activity during the first 2 weeks of exposure (Kleinerman et al.
1982).

Nitrogen dioxide exposure has not been shown to cause alveolar
septal disruption, an essential feature of centrilobular emphysema,
but it does result in a significant reduction in the internal surface
area in the lungs of hamsters exposed for 12 to 14 months
(Kleinerman and Niewoehner 1973).

Cadmium Salts

Animals exposed to cadmium, a constituent found in the particu-
late phase of cigarette smoke, develop a number of histologic and-
biochemical changes that may lead to emphysematous lesions.

Exposed animals develop pulmonary edema, vascular congestion,
intraparenchymal hemorrhages, and a loss of Type I pneumocytes
(Palmer et al. 1975; Strauss et al. 19761, and PMNs and mononuclear
cells influx into the lungs (Snider et al. 19731. The animals develop
acute peribronchial damage followed by the accumulation of granu-
lation tissue near the respiratory bronchioles, thickened alveolar

276


septa, and distortion and distention of neighboring alveoli Snider et
al. 1973). These histologic changes are more suggestive of the scar or
paracicatricial form of emphysema than the centrilobular form
reported in some industrial workers exposed to CdCl, (Princi 1947).
This disparity in response to CdCl, could be related to species
differences or the interaction of CdCl, with other factors.

When hamsters are exposed to CdCl, plus beta-amino proprioni-
trile (f3-APN), an inhibitor of lysyl oxidase, they develop thin-walled
subpleural bullae and airspace enlargements resembling panlobular
emphysema (Niewoehner and Hoidal 1982). The mean linear dis-
tance between alveolar intercepts is significantly increased; pres-
sure-volume studies show overinflation and increased compliance of
the lungs (Niewoehner and Hoidal 1982). This study suggests that
CdClp, perhaps in conjunction with some other as yet undetermined
agent, may be important in the pathogenesis of pulmonary emphyse-
ma. The fact that CdCl, is a constituent of cigarette smoke (Randi et
al. 1969) lends support to this hypothesis.

Cigarette Smoke

Cigarette smoking has been clearly identified as a major causal
factor in the development of pulmonary emphysema in humans
(Auerbach et al. 1972; 1974; Petty et al. 1967; Andersen et al. 1967;
Niewoehner et al. 1974; USDHEW 1979). However, an animal model
for the development of emphysema using the inhalation of cigarette
smoke alone has not been convincingly demonstrated. Parenchymal
disruption resembling human emphysema has been reported in some
dogs following prolonged cigarette exposure, but this histologic
pattern is not uniformly present (Hernandez et al. 1966; Auerbach et
al. 1967a; Zwicker et al. 1978).

This difficulty in developing an animal model for cigarette-induced
emphysema may relate to the reluctance of animals to inhale smoke
and the relatively long duration of exposure required to produce
emphysema in humans. However, it may also result from the need
for a combination or sequence of effects to induce emphysematous
change. That is, an increased elastase burden might be necessary
isecondary to the cellular response to smoke) before the oxidant
damage of smoke to a,AT, or to repair mechanisms, results in
emphysema. Hoidal and Niewoehner (1983) examined this question
in hamsters exposed to low doses of smoke and elastase. Neither
exposure alone resulted in significant emphysematous change, but
the combined exposure did cause change. This suggests that an
increased elastase burden may be a precondition for smoking-
induced emphysematous lung injury, and may also explain the long
exposure period required in humans prior to the demonstration of an
increased prevalence of emphysema in smokers.

277


Experimental studies have shown that cigarette smoke can induce
a number of cellular, biochemical, and metabolic changes within the
lungs that may be causally related to the development of emphyse-
ma. Macrophages and leukocytes, cells known to contain proteolytic
enzymes, are recruited to the lungs of hamsters (Kilburn and
McKenzie 1975) and guinea pigs (Flint et al. 1971) following exposure
to cigarette smoke, thereby increasing the proteolytic burden of the
lungs. Conversely, the a,AT activity decreases in rats after inhala-
tion of cigarette smoke (Janoff et al. 1979a). The increased proteolyt-
ic burden within the lungs coupled with the concomitant diminu-
tion in inhibitory capacity tends to create a protease-antiprotease
imbalance and a situation whereby unrestrained connective tissue
destruction may occur.

The Effects of Smoking on Cellular and immune Defense
Mechanisms

There are important functional differences between macrophages
from smokers and those from nonsmokers (Table 1). For example,
Warr and Martin (1977) demonstrated that receptors for the third
component of complement (C3b) are decreased in number or function
on the surface of smokers' alveolar macrophages. The receptors for
the Fc portion of IgG, however, are normal on these cells (Warr and
Martin 1977). An important function of the C3b receptor is to
augment the attachment and phagocytosis of microorganisms and
particulates by the macrophages. It is not clear whether this subtle
defect in cell function results in a significant alteration in phagocyto-
sis or clearance of particulates or microorganisms by these cells. In
this regard, the phagocytosis and killing of a variety of microorga-
nisms by smokers' alveolar macrophages have been shown to be
normal by Harris et al. (1970) and Cohen and Cline (1977). One
report by Martin and Warr (19771, however, suggests that the
capacity of alveolar macrophages to kill bacteria is decreased in
smokers.

The observation that human alveolar macrophages from cigarette
smokers function normally to kill microorganisms appears to differ,
at first glance, from a number of animal studies demonstrating that
the capacity of alveolar macrophages to phagocytose and kill
bacteria is impaired following exposure to cigarette smoke (Halt and
Keast 1973; Rylander 1971,1973). In these animal studies, there was
an initial decrease in the numbers of alveolar macrophages and a
decrease in their bactericidal function following exposure to ciga-
rette smoke. With prolonged exposure, however, the number of
macrophages increased and their ability to kill microorganisms
returned to normal (Rylander 1973, 1974). These observations
suggest that cigarette smoke, initially, is toxic to alveolar macro-


phages. However, it is likely that the macrophages, with time, adapt
to the presence of cigarette smoke. In addition, a subpopulation of
macrophages that are more resistant to cigarette smoke may
increase in number in the lung. The macrophages isolated from the
lungs of smokers resemble those isolated from animals following
prolonged exposure to cigarette smoke. The acute effects of cigarette
smoking on the number and functions of alveolar macrophages in
man has not been systematically evaluated.

Alveolar macrophages of cigarette smokers appear to interact in
an abnormal fashion with lymphocytes (Table 1). In this regard, the
alveolar macrophages from cigarette smokers function poorly as
accessory cells in presenting antigen to autologous lymphocytes
(Laughter et al. 1977). This latter defect may be further magnified by
the observation that lymphocytes from cigarette smokers also
respond poorly to mitogens (Neher 1974; Daniele et al. 1977).
Additional evidence for an abnormal interaction of macrophages and
lymphocytes in lungs of cigarette smokers is a decreased response of
alveolar macrophages to the lymphokine, macrophage migration
inhibitory factor (Warr 1979). These observations suggest that
cigarette smoking may have broad effects on the ability of the lung
to generate a cellular immune response.

In Vitro Effects of Cigarette Smoke on Inflammatory and
Immune Effector Cells

The most readily demonstrable effect of cigarette smoke, in vitro;
is a decrease in cell viability (Holt et al. 1974; Holt and Keast 1973;
Nulsen et al. 1974; Weissbecker et al. 1969). At relatively low
concentrations, cigarette smoke and its constituents rapidly kill
alveolar and peritoneal macrophages in vitro. Lymphocytes and
polymorphonuclear leukocytes are also very susceptible to these
agents (Holt et al. 1974; Blue and Janoff 1978).

When sublethal amounts of cigarette smoke are employed, a
number of metabolic and functional changes occur in macrophages.
Phagocytosis is depressed, as is the function of a number of
macrophage enzymes (Vassallo et al. 1973; Green 1968a, b, c, 1969,
1970; Green and Carolin 1966, 1967; Green et al. 1977; Powell and
Green 1971; Hurst and Coffin 1971). In addition; protein synthesis is
also depressed (Yeager 1969; Low 19741, and stimulatory effects have
been noted. While cigarette smoke depresses phagocytosis and
intracellular killing, nitrogen dioxide increases the metabolic activi-
ty of macrophages (Vassallo et al. 1973). Similar effects have been
observed under some conditions with cigarette smoke (Holt and
Keast 1973; Leuchtenberger and Leuchtenberger 1971). Results from
a number of investigators suggest that the balance between stimula-
tion and inhibition of macrophage activity is determined by dosage,
with stimulation occurring at low exposure levels and inhibition at

279


higher concentrations (Halt and Keast 1973; Lentz and DiLuzio 1974;
York et al. 1973). The most potent stimulation occurs after prolonged
exposure to low levels of the agent.

These in vitro effects of cigarette smoke are also seen acutely in
vivo following exposure to smoke. The immediate effect of exposure
to cigarette smoke, and to agents present in the smoke, is a decrease
in viability of the pulmonary alveolar macrophages (Halt and Keast
1973a, b; Rylander 1971, 1973; Coffin et al. 1968; Dowel1 et al. 1970;
Holt and Nulsen 1975; Gardner et al. 1969). Although not well
studied, it is also likely that cigarette smoke is toxic to polymorpho-
nuclear leukocytes (Blue and Janoff 1978). In support of these
observations are studies demonstrating that acute exposure of
experimental animals to tobacco smoke, or to components of
cigarette smoke, also lowers their resistance to bacterial infection
(Rylander 1969, 1971; Acton and Myrick 1972; Gardner et al. 1969;
Goldstein et al. 1971; Huber and LaForce 1971; Huber et al. 1971).
Short-term exposure to components of cigarette smoke, particularly
nitrogen oxide, has also reduced resistance to viral infection,
probably by inhibiting interferon production by macrophages (Va-
land et al. 1970). As noted above, these in vitro and acute in vivo
effects of cigarette smoke are not seen following long-term in vivo
exposure in animals; very little work has been done on the effects of
cigarette smoke in vitro, using human cells. However, several studies
have shown that cigarette smoking and nicotine, at levels compara-
ble to those encountered in the circulation of smokers, produce a
slight but significant depression of PHA-stimulated DNA synthesis
in human peripheral blood lymphocytes (Neher 1974; Silverman et
al. 1975; Vos-Brat and Rumke 1969).

The Effect of Cigarette Smoke on Antibody Production

The available data on antibody production in human smokers
suggest that cigarette smoking may depress these responses. The
production of antibodies was investigated in a large study involving
influenza vaccination. Smokers in the population exhibited in-
creased susceptibility to infection during an influenza outbreak
(Finklea et al. 1969, 1971). Prior to immunization with influenza
vaccine, smokers exhibited significantly lower titers of specific
antibodies than did nonsmokers. Immediately following vaccination
of both groups, the smokers developed levels of antibodies compara-
ble to those of nonsmokers. However, the antibody titer in the
smokers fell below their nonsmoking counterparts within a few
weeks, and by a year after vaccination the smokers exhibited
markedly depressed levels of circulating antibodies.

The capacity of cigarette smoking to alter antibody production was
also studied by evaluating the capacity of a fetus to stimulate
lymphocytotoxic antibodies against HLA antigens in the mother


(Nyman 1974). Sera from a large number of pregnant women were
tested for the presence of lymphocytotoxic antibodies against a 4&
donor panel. The smokers exhibited a significantly lower incidence
of these antibodies than did nonsmokers, and the divergence
between groups increased with the number of deliveries. Infections
during pregnancy were observed significantly more often in the
smokers in this trial.

In several animal models, acute exposure to whole cigarette smoke
or components of cigarette smoke depressed the numbers of anti-
body-forming cells in the spleen and the serum levels of antibodies in
animals exposed to a variety of antigens (Miller and Zarkower 1974;
Zarkower and Marges 1972; Zarkower et al. 1970). The depression
was greatest when the antigen was administered by an aerosol
(rather than by systemic inoculation), indicating t,hat smoke appears
to exert an effect close to the point of entry. Prolonged exposure
ultimately resulted in severe depression both in local and in systemic
antibody responses (E&r et al. 1973; Holt et al. 1976; Thomas et al.
1973,1974a, 1974b, 1975).

Although it is tempting to relate these abnormalities in immune
response to the known association between cigarette smoking and
increased incidence of upper respiratory infection, it is not clear
whether the subtle defects in immune functions can entirely account
for the infections present in cigarette smokers. Clearance of bacteria
from the respiratory tract is a complex process that involves
interplay between a variety of different mechanisms, only some of
which include the function of alveolar macrophages and the capacity
of the lung to mount cellular and humoral immune responses. Other
abnormalities present in cigarette smokers that could account for
this increased incidence of infection include a markedly abnormal
tracheal bronchial clearance of particulates and an increased
adherence of bacteria to airway epithelium (reviewed in USPHS
1971, 1973, 1974).

281


EFFECTS OF CIGARETTE SMOKE ON AIRWAY
MUCOCILIARY FUNCTION

Introduction

There is extensive literature on the effects of cigarette smoke on
mucociliary clearance in the airways, with the majority of the
reports appearing between 1965 and 1975. Different experimental
approaches have been used, including in vivo measurement of
mucociliary function in animal models and in human subjects. The
results of some of these studies have been contradictory, presumably
because of differences in experimental technique or the influence on
mucociliary function of factors other than cigarette smoking. For
example, tracheal mucociliary transport appears to decline with age
in normal subjects (Goodman et al. 19781, an important phenomermn
to consider when assessing the effects of long-term cigarette smok-
ing. Another complicating factor is the clearly demonstrated impair-
ment of mucociliary function produced by chronic bronchitis even in
nonsmokers, such as in patients with cystic fibrosis (Wood et al.
1975) or immunoglobin deficiency (Mossberg et al. 1982). Therefore,
it is difficult to separate the direct effects of cigarette smoke on
mucociliary function from those of smoking-associated chronic
bronchitis. Finally, in vitro bioassays for ciliotoxicity may not
reliably reflect the effects of cigarette smoke on the mucociliary
apparatus in the intact airways. Thus, Dalhamn et al. (1967) found
that smoke produced by cigarettes containing a high concentration
of hydrogen cyanide was more ciliotoxic in vitro than that produced
by cigarettes containing a low concentration of hydrogen cyanide,
and the two types of cigarettes caused a comparable reduction of
mucus transport in vivo.

This review is divided into three parts. The first part summarizes
the normal structure and function of the mucociliary system in the
airways. The second part deals with the direct effects of short-term
and long-term cigarette smoke exposure on mucociliary function,
and the third part discusses mucociliary function in chronic bronchi-
tis.

Normal Mucociliary Function

The principal function of the airway mucociliary system is its
contribution to host defenses. This is accomplished by physical
removal of inhaled foreign material from the ciliated airways by
mucous transport and by biochemical and immunological processes
that protect against invasion of the mucosa by infectious agents.
Normal mucociliary clearance depends upon an optimal interaction
between cilia and mucus.

283


Cilia

The respiratory mucosa from the proximal trachea to the terminal
bronchioles consists of a pseudostratified epithelium with cilia
protruding from the luminal surface of columnar cells (Figure 1).
The larynx contains a mucus-secreting squamous epithelium over
most of its surface, and cilia are present only in the posterior
commissure (Wanner 1977). The major cell types in the respiratory
mucosa are basal cells, intermediate cells, nonciliated columnar
cells, ciliated columnar cells, and goblet cells. In the larger airways,
the major part of the epithelial surface is ciliated. The ratio of
ciliated columnar cells to goblet cells is approximately 5:l in the
trachea, with a relative decrease in the number of both cell types
toward the peripheral airways. The surface of each ciliated columnar
cell contains approximately 200 cilia with an average length of 6 pm
and diameter of 0.2 urn. Both ciliated and nonciliated columnar cells
are characterized by microvilli on their luminal surface. These
measure 0.3 pm in length and 0.1 pm in diameter. The ultrastruc-
ture of cilia in lower animals and mammals is remarkably similar.
Each cilium contains longitudinal microtubules that appear to
represent contractile elements. Two single microtubules form a
central core, and nine microtubules with a doublet structure are
arranged in a circular fashion in the periphery of the cilium. A basal
body in the apex of the cell corresponds to each cilium. Circular and
radial bridges have been demonstrated between the peripheral
microtubules and between the peripheral and central microtubules.
These bridges (dynein arms, nexin links, radial spokes) appear to be
crucial for ciliary bending.

Mucus

Respiratory secretions consist of mucus produced by submucosal
glands and goblet cells and tissue fluid. The total volume of all
mucus-producing structures has been estimated at approximately 4
ml in human lungs; submucosal glands make up most of this volume
(Wanner 1977). The submucosal glands are under parasympathetic
nervous control, with an estimated daily volume of respiratory
secretions between 10 and 100 ml. FIuman respiratory secretions
contain approximately 95 percent water. `I'he rest consists of
micromolecules (electrolytes and amino acids) and macromolecules
(lipids, carbohydrates, nucleic acid, mucins, immunoglobulins, en-
zymes, and albumin). In situ, the respiratory secretions take the
form of two layers, i.e., periciliary fluid (sol phase), and mucus (gel
phase), as shown in Figure 1. Mucus has been clearly identified as
the product of submucosal glands and goblet cells; the origin of the
periciliary fluid has not been definitely established, although
transepithelial water transport appears to be the most likely source.
Tn central airways, the mucus layer is 5 to 10 pm deep and may be

284


FIGURE l.-Schematic representation of normal mucosa
      (central airway) with the components of the
      mucociliary apparatus

N(YTE: From top (airway lumen) to bottom, note IIIUCUE layer. periclliary tluld layer, eplthelxum mrh a
predominance of ciliated mlumnar cells and an mterapemed goblet cell, basement membrane, and aubmucoaal
gland.
SOURCE. Wanner (1979)

discontinuous. In peripheral airways where submucosal glands are
absent and goblet cells are rare, mucus is either absent or present
only in small quantities. Regeneration of injured ciliated respiratory
epithelium takes approximately 2 weeks in animals; the exact
regeneration time of damaged human tracheobronchial ciliated
epithelium is not known (Wanner 1977). It appears that regeneration
does not begin until 2 to 3 days following mechanical injury.

Mucociliary Interaction

Mucociliary interaction depends on ciliary activity, the rheologic
properties and depth of the mucus layer, and the depth of the
periciliary fluid layer. The viscoelastic properties of mucus are
determined by its biochemical characteiistics (&sulfide cross-linking
and hydrogen bonding between glycoprotein molecules and water)
(Wanner 1977). Cilia beat in one plane, with a fast effective stroke
(power stroke) in the cephalad direction and a recovery stroke that is
two to three times slower. Adenosine triphosphate has been identi-
tied as the energy source for ciliary bending. In mammals, the
average normal ciliary beat frequency is approximately 1,000 beats
per minute, with coordinated motion in adjacent cilia on an
individual cell and in cilia of adjacent cells. This interciliated


TABLE 2.-Measurement of airway mucociliary function in
       humans

Method                 Reference

Clearance of inhaled radioactive
aerceols from the lungs

Morrow et al. (1967)

Transport of discrete markers

Friedman et al. (1977)
Sackner et al. (1973)

Central airway clearance of inhaled
boli of radioactive micrarpheree

Yeatea et al. (1975)

pattern of motion by which adjacent cilia beat one after another to
generate a wave of ciliary motion is called metachronism.

The "normal" thickness of the periciliary fluid layer is less than,
or at best equal to, the length of the ciliar shafts, which measure
approximately 6 pm in the central airways. The luminal surface of
the mucus layer appears smooth, whereas the surface in contact with
the cilia is irregular, and the mucus penetrates between the ciliary
shafts. This penetration and the claw-like projections at the cilia tips
may further facilitate the mechanical interaction between cilia and
mucus. In the trachea, the "normal" surface mucus transport
velocity is between 5 and 20 mm per minute depending upon the
method of measurement. Mucous transport velocity decreases
toward the peripheral airways.

The three principal methods for the measurement of tracheobron-
chial mucociliary function in humans are listed in Table 2. All of
these have been used in studies of the effects of cigarette smoke on
mucociliary function.

Theoretically, mucociliary dysfunction can result from alterations
in ciliary beat frequency and coordination, the quantity and visco-
elastic properties of mucus, and the thickness of the periciliary fluid
layer. In addition, focal destruction of the respiratory epithelium,
producing areas without cilia or mucus, is also associated with
impaired or absent mucociliary transport.

Effects of Cigarette Smoke on Mucociliary Function

The irritant effects of cigarette smoke on mucociliary clearance
were recognized by Mendenhall and Shreeve (1937). They observed a
decrease in the transport rate of carmine particles on the mucosa of
excised cat tracheas after bringing them in contact with cigarette
smoke, either directly or by dissolving it in the solution in which the
t.racheas were immersed (Mendenhall and Shreeve 1937, 1940).
These early findings were confirmed by Hilding (1956) and Dalhamn


(1959) approximately 25 years ago. Since then, investigations to
assess the effects of cigarette smoke on ciliary activity and mucocili-
ary transport have proliferated. Because cigarette smoke can impair
mucociliary transport by interfering with ciliary activity or mucus
secretion, the studies relating to these two component functions are
discussed separately and are complemented by a review of experi-
ments involving mucociliary transport, the ultimate expression of
mucociliary function.

The effects of cigarette smoke on mucociliary function have been
extensively studied in vitro, in intact animal models, and in human
subjects. Comparison among the results of these experimental
approaches is difficult, as there are major differences in inhalation
patterns even between animal models and human subjects. Cigarette
smoke is modified during its passage through the upper airways, and
this may vary depending upon the mode of inhalation. By using
smoke produced by radio-tracer spiked cigarettes, it has been shown
that mice, who are obligatory nose breathers, retain 50 percent of
the inhaled radioactivity in the nasal passages, 20 percent in the
lungs, and the rest in the esophagus, stomach, and other organs
(Page et al. 1973). Using artificial airways to bypass the nasopharynx
in experimental animals eliminates the problem of nasal cigarette
smoke absorption, but also prevents the oral modification of ciga-
rette smoke that is a typical feature of human smoking. In subjects
inhaling cigarette smoke from a smoke dosage apparatus that
delivers standard puffs, 86 to 99 percent of most components of gas
and particulate phases of cigarette smoke are retained, with the
exception of carbon monoxide (CO), of which only 54 percent is
retained (Dalhamn et al. 1968). Much of the smoke appears to be
retained in the mouth. In human subjects who hold cigarette smoke
in their mouth for 2 seconds, 60 percent of the water-soluble
components of the gas phase, 20 percent of the water-insoluble
components of the gas phase, and 16 percent of particulate matter
are absorbed or retained in the upper airways (Frances et al. 1970;
Stupfel et al. 1974). This marked modification of cigarette smoke
might decrease its ciliotoxic effect in the lower airways. Passing
unfiltered cigarette smoke through a chamber with wet surfaces
before bringing it in contact with ciliated epithelium decreased the
cilioinhibitory capacity of the cigarette smoke (Kaminski et al. 1968).
When filtered cigarette smoke was used, the wet surfaces had no
additional effect on ciliotoxicity, indicating that the mucosa of the
upper airway may serve as a filter (Kaminski et al. 1969). Similar
observations have been made when cigarette smoke was passed
through a water trap (Albert et al. 1969).

Because of the differences in inhalation pattern between humans
and animals, it might be argued that nasal mucociliary function
should be measured to assess the effects of inhaled cigarette smoke

287


TABLE S.-Effects of cigarette smoke on airway
      mucociliary system

EXposW?

                     Impaired mucociliary
Clliary dysfunction Mucus hypereecretion     clearance

Short term        Yes             ?             ?
Lang term        Yes            Ye8            YeS

in animal models. However, it has been clearly shown that nasal
mucociliary transport is not a good marker of tracheobronchial
mucociliary transport because of differential responses at the two
sites. For example, exposure to the whole cigarette smoke of up to 30
cigarettes does not impair nasal mucociliary transport in donkeys
(Frances et al. 19701, whereas the same number of cigarettes clearly
alters tracheobronchial deposition and clearance of radioactive
aerosols (Albert et al, 1969). Likewise, Hilding (1965) has concluded
from his studies that the nose is not an acceptable organ for the
study of the effects of cigarette smoke on mucociliary transport.

Realizing the problems with experimental models of ciliary
function in response to cigarette smoke inhalation, Dalhamn (1969)
postulated that a proper experimental design should fulfill the
following requirements: (1) the exposure pattern and level of
cigarette smoke inhalation should simulate that of natural smoking
in human subjects, (2) cigarette smoke should be delivered in air and
not as an aqueous solution, (3) the components of inhaled cigarette
smoke should be analyzed, and (4) exposure should be of long
duration. Although these criteria are obviously not met by many of
the studies quoted in this review, understanding these principles
allows a more critical assessment of the reported results as shown in
Table 3.

Short-Term Exposur&

The effects of short-term cigarette smoke exposure on the mor-
phology of the respiratory mucosa have not been investigated in
man. Cigarette smoke residue has been shown to cause ciliary
damage in cultured rabbit tracheal epithelium with a contact-time-
dependent effect (Kennedy and Allen 1979). The most consistent
abnormalities were cellular desquamation and alterations in mito-
chrondria, cilia, and microvilli, some of which occurred as early as 1
hour after exposure commenced.

Cytotoxicity has also been observed after short-term exposure of
ciliated epithelium to aqueous extracts of cigarette smoke conden-
sate in vitro (Donnelly 19691, but it is difficult to extrapolate data
from these in vitro studies to the in vivo conditions that occur during
cigarette smoking.

288


Cilia

With a few exceptions, e.g., Proetz (1939), most investigators have
demonstrated an irritant effect of smoke on ciliated epithelium,
usually characterized by ciliostasis. Residues of cigarette smoke
passed through an aqueous medium have been shown to produce
ciliostasis in protozoa (Weiss and Weiss 1964; Wang 1963) and in
fragments of human respiratory epithelium (Ballenger 1960). In
fragments of rat trachea, brief exposure to whole cigarette smoke
appears to elicit a biphasic response, with a short period of
stimulation during 1 to 2 minutes followed by a marked decrease in
ciliary beat frequency (Guillerm et al. 1961, 1972). In an excised
rabbit trachea model, 71 l-ml puffs or 35 lo-ml puffs of whole
cigarette smoke were necessary to produce ciliostasis; similar
relationships were demonstrated in the tracheas of living cats
(Dalhamn 1970; Dalhamn et al. 1968). Several investigators have
established a stimulus-response relationship between dilutions of
aqueous cigarette smoke extract and the time of exposure required
for total stoppage of ciliary beat frequency in different experimental
models (Donnelly 1969, 1972; Das et al. 1970; Donnelly et al. 1981).
The mechanism by which cigarette smoke acutely depresses ciliary
function is not clearly known, but may involve enzyme inhibition of
adenylate kinase, thereby reducing adenosine triphosphate (ATP),
the energy source for ciliary bending (Mattenheimer and Mohr 1975:
Schabort 1967). Ciliary function in response to short-term cigarette
smoke inhalation has not been studied in man.

Very little is known about the quantity and rheologic properties of
airway secretions after short-term cigarette smoke exposure. A brief
exposure of slugs to cigarette smoke has been-reported to stimulate
the production of mucus containing an increased number of acid
glycoprotein fibers (Wilde 1981). The significance of this observation
with respect to the human respiratory tract is not clear, except that
an increased number of acid glycoprotein fibers has also been
demonstrated in sputum obtained from cigarette smokers.

Mucociliary Interaction

Using a variety of different techniques in animal experiments,
ciliary dysfunction and impairment of mucociliary transport by
short-term exposure to cigarette smoke have been demonstrated in
rats (Iravani 1972; Dalhamn 1964; Ferin et al. 19661, rabbits
(Dalhamn 1964; Holma 19691, cats (Carson et al. 1966; Dalhamn
1964, 1969; Kaminski et al. 1968), dogs (Guillerm et al. 1972;
Sakakura and Proctor 1972; Isawa et al. 1980), donkeys (Albert et al.
1974, 19691, chickens,.and sheep (Stupfel et al. 19743. A few reports


have not demonstrated that short-term exposure to smoke depresses
mucociliary function in animal models (La Belle et al. 1966; Bair and
Dilley 1967). The reasons for the discrepancy between these and the
previously listed studies are not clear, but may be related to
methodology and dose of exposure. Stimulus response curves be-
tween dose of cigarette smoke and the degree of mucociliary
inhibition have been shown in airways of chickens and dogs (Battista
and Kensler 1970b; Sakakura and Proctor 1972; Isawa et al. 1980). In
one study, for example, 9 puffs of nonfiltered cigarette smoke had
variable effects on tracheal mucociliary transport in intact dogs, but
tracheal mucociliary transport was consistently inhibited by 12 puffs
(Sakakura and Proctor 1972). Likewise, the number of 4-second
exposures to cigarette smoke (separated by 1 minute) required to
reduce mucus transport in intact chicken tracheas by more than 90
percent has been shown to increase with increasing dilutions (from
50 to 3 percent) of smoke in air (Battista and Kensler 1970b).

Measurements of mucociliary clearance in man immediately after
smoking one or more cigarettes have shown conflicting results, with
either increased (Albert et al. 1973; Camner et al. 1971; Albert et al.
1975; Camner and Philipson 1971,1974), inconsistent, or unchanged
rates (Yeates et al. 1975; Pavia et al. 1971; Goodman et al. 1978) or
decreased (Nakhosteen et al. 1982) rates. Transient effects on
mucociliary clearance have been reported in both smokers and
nonsmokers (Hilding 1956; Pavia et al. 1971). Such differences
between human subjects may reflect a difference in the dose and
inhalation pattern of cigarette smoke.

Long-Term Exposure

In dogs inhaling cigarette smoke through a tracheostomy, histolog-
ic changes have been observed in the bronchi after 229 to 421 days of
exposure (Auerbach et al. 1967b). These changes consisted of
epithelial hyperplasia, decreased number of ciliated cells, and areas
of squamous metaplasia. This may be criticized as a poor model of
cigarette smoking in humans because the upper airway, which
absorbs part of the smoke and decreases its toxicity, is bypassed. The
irritant effect of cigarette smoke on the tracheobronchial mucosa
could be enhanced in this model. However, inflammatory changes in
the airways have also been observed in animals that inhaled
cigarette smoke via their upper airways (Leuchtenberger et al. 1958;
Rylander 1974; Mattenheimer and Mohr 1975; Park et al. 1977;
Basrur and Basrur 1976; Jones et al. 1973; Iravani 1973). Various
types of lesions have been observed, including tracheal and bronchial
epithelial hyperplasia (Park et al. 1977: Frasca et al 19741, goblet
cell proliferation and submucosal gland hypertrophy (Jones et al.
1973; Park et al. 19771, bronchiolar metaplasia of mucus-secreting
cells (Basrur and Basrur 19761, increased quantities of airway mucus

290


that appear to be adherent to submucosal gland openings (Iravani
19731, and a decreased number of ciliated epithelial cells (Basrur and
Harada 1979). One study suggested a dose-dependence of the mucosal
lesions when comparing hamsters exposed to either four cigarettes
per day or eight cigarettes per day for 2 weeks (Basrur and Basrur
1976). The pathologic changes produced by long-term cigarette
smoke exposure appear to be reversible if the exposure time is not
excessive. Thus, inflammatory changes in the airways of hamsters
exposed to cigarette smoke for 4 weeks showed marked reversibility
with a recovery time of several weeks (Basrur and Harada 1979).

The histologic changes in the airways of cigarette smokers are
similar to those produced by cigarette smoke in animal models, and
consist of varying degrees of denudation of the ciliated epithelium,
an increase in the number of goblet cells, submucosal gland
hypertrophy, and squamous metaplasia (Regland et al. 1976; Jones
1981). Morphometric studies have demonstrated an increased quan-
tity of mucus in the airway lumen without histologic evidence of
coexistent emphysema or a history of obstructive lung disease,
whereas this is not observed in the lungs of healthy nonsmokers
(Niewoehner et al. 1974; Matsuba and Thurlbeck 1971). Electron
microscopic examination of ciliated epithelium in surgical lung
specimens obtained from cigarette smokers has revealed ciliary
abnormalities consisting of compound cilia, single axoneme, intra-
cyctoplasmic microtubular doublets, and cilia within periciliary
sheaths (McDowell et al. 1976). If bronchial biopsy material is used to
detect ciliary abnormality in cigarette smokers, the results must be
interpreted with caution, for a single biopsy may be misleading
owing to the focal nature of the lesions (Fox et al. 1981).

The morphologic changes of the respiratory mucosa in animals
exposed to cigarette smoke for prolonged periods and in human
cigarette smokers strongly suggests the presence of mucociliary
dysfunction. This has been clearly demonstrated, particularly with
respect to the production and clearance of mucus.

Cilia

Ciliary function after long-term cigarette smoke exposure has not
been extensively studied. Iravani and Melville (1974) demonstrated a
decrease in ciliary beat frequency in the airways of hamsters
exposed to cigarette smoke for 1 year; however, in rats also exposed
for 1 year under almost identical conditions, ciliary frequency was
generally increased, although there were zones of ciliary inactivity
or discoordination. A sustained inhibition of adenylate kinase
activity in ciliated tracheal cells of hamsters exposed to cigarette
smoke for up to 9 months has also been reported (Mattenheimer and
Mohr 1975). Because inhibition of this enzyme leads to a decreased
generation of adenosine triphosphate. the energy source of ciliary

291


bending, a decreased ciliary activity might be expected (Mattenheim-
er and Mohr 1975).

Mucus

Mucus hypersecretion has been clearly demonstrated in the
airways of several animal species exposed to cigarette smoke for
prolonged periods of time (Battista and Kensler 1970a; Iravani and
Melville 1974). Rheologic measurements of airway mucus have not
been reported in such animal experiments, but biochemical analysis
has revealed the presence of serum proteins that might have
cilioinhibitory effect (Dalhamn and Pira 1979; Battista 1980). Mucus
hypersecretion may occur as early as 1 month after beginning a
smoke inhalation equivalent to as little as one cigarette per day
(Battista and Kensler 197Oa). Rheologic and biochemical examina-
tions of airway secretions in healthy smokers have not been carried
out, primarily because these subjects do not have a productive cough.
Once a smoker develops chronic productive cough, he or she is no
longer considered healthy, but by definition, has chronic bronchitis.

Mucociliaty Interaction

Long-term effects of cigarette smoke on airway mucociliary
transport have been studied in different animal species, In purebred
beagle dogs exposed to cigarette smoke (100 cigarettes per week) for
13.5 months via a mask that administered cigarette smoke through
both the mouth and the nose for 1.5 hours twice daily, tracheal
mucus transport rate was decreased to approximately 30 percent of
that observed in control animals (Wanner et al. 1973). Pulmonary
function did not differ significantly between the two groups. It has
subsequently been shown that the abnormality in mucociliary
transport in beagles may already be present after 6 months of
cigarette smoke exposure (Park et al. 1977). An impairment of
mucociliary clearance with long-term cigarette smoke exposure has
also been demonstrated in rabbits, guinea pigs, rats, and chickens
(Okajima 1971; Rylander 1971b; Iravani and Melville 1974; Battista
and Kensler 1970a). In some of those experiments, impaired mucoci-
liar-y clearance was already observed 4 weeks after the beginning of
exposure.

The long-term effects of cigarette smoking on mucociliary function
in human subjects has been investigated by aerosol clearance
techniques and discrete marker transport techniques. Some of the
investigators using radioactive aerosols demonstrated no abnormali-
ty of overall clearance in habitual cigarette smokers, particularly in
those who already may have had symptoms of chronic bronchitis
(Sanchix et al. 1972; Yeates et al. 1975; Pavia et al. 1970; Pavia and
Thomson 19701. However, the deposition of the inhaled radioactive

292


aerosol is more central in normal smokers and in patients with
chronic bronchitis than in nonsmokers (Lippman et al. 1970).
Because clearance is faster in central airways than in peripheral
airways, this centralization of aerosol deposition may compensate for
the overall decrease in mucociliary clearance. Investigations that
have related mucociliary clearance to deposition pattern have
generally found an impairment of mucociliary clearance in cigarette
smokers (Lourenco et al. 1971; Camner et al. 1973a; Camner and
Philipson 1972, 1974). Camner and Philipson (19721, in a study of 10
pairs of twins discordant for cigarette smoking, showed a significant-
ly lower average clearance rate in smokers compared with nonsmok-
ers; in 5 pairs clearance was slower in the smoker than in the
nonsmoker, whereas in the remaining 5 pairs there was no differ-
ence. Analysis of regional clearance has produced further evidence
that overall clearance of inhaled radioactive aerosols may fail to
detect an abnormality in mucociliary clearance. Thus, Bohning and
co-workers (1975) studied the deposition and clearance of 7 pm
diameter particles in the tracheobronchial tree of six pairs of
monozygotic twins, four of whom were discordant for cigarette
smoking. They found comparable overall mucociliary clearance in
the smoking and nonsmoking pairs, but more central deposition and
slower central clearance in the smokers. Others have reported an
impairment of peripheral mucociliary clearance and alveolar clear-
ance as well (Matthys et al. 1983; Cohn et al. 1979).

Discrete particle techniques involving either bronchoscopy or
radiography have been used to assess mucus transport in central
airways, notably the trachea. Most investigators have reported a
decrease of tracheal mucus velocity in healthy smokers, with values
ranging between 20 percent and 80 percent of those of nonsmoking
controls (Goodman et al. 1978; Toomes et al. 1981; Nakhosteen et al.
1982).

The bulk of the evidence indicates that long-term cigarette
smoking alters mucociliary transport mechanisms and that these
changes can occur as early as 1 year after smoking onset. Partial
recovery of mucociliary transport has been observed in cigarette
smokers after cessation for 3 months or more, but not after 1 week of
cessation (Camner et al. 1973). These observations have also been
supported by animal experiments (Albert et al. 1971).

Fractionation and Filtering of Cigarette Smoke

Whole cigarette smoke is composed of voiatile elements and
particulate matter, and it has become customary to distinguish
between the gas phase and the particulate phase. The gas phase, by
definition, consists of the components that remain after cigarette
smoke has been "effectively" filtered by passing it through appropri-
ate filters (Dalhamn 1966; Kensler and Battista 1963; Falk et al.

293


1959). The major constituents of the particulate phase are nicotine,
phenols, hydrocarbons, aldehydes and ketones, organic acids, and
alcohols. Although 95 percent of the gas phase (approximately 300
ml per cigarette)~cogaists of combustion products and admixed air
(nitrogen, oxygen, carbon dioxide, carbon monoxide) in concentra-
tions that do not affect mucociliary transport, some trace gases are
important (Rat&a Qt al. 1962). These include nitrogen dioxide,
ammonia, cyanides, aldehydes, ketones, acrolein, and acids. As is
shown below, some controversies still remain about whether the gas
phase or the particulate phase of cigarette smoke is primarily
responsible for its depressant effect on mucociliary activity. This
problem is relevant when comparing the effects on mucociliary
clearance of low tar versus high tar cigarettes, low nicotine versus
high nicotine cigarettes, and filtered versus nonfrltered cigarettes.
Dalhamn (1966) reviewed the controversy over the separate effects
of the gas and the particulate phases of cigarette smoke on
mucociliary function. In protozoa, both phases of cigarette smoke
have been shown to possess ciliotoxic properties (Kennedy and
Elliott 1970). Falk and associates (19591 reported that exposure to
whole cigarette smoke for 30 seconds resulted in a biphasic response,
with an initial stimulation, followed by depression with a minimum
value at about 15 minutes and a tendency toward recovery 45
minutes after exposure. Removal of the particulate matter in
cigarette smoke by passing it through filters decreased its depressant
effect on mucus transport, indicating that the major effect on
mucociliary clearance was related to the particulate phase. Similar
observations have been made by others (Rylander 1970; Falk et al.
1959). In contrast, Kensler and Rattista (1963) incriminated the gas
phase of cigarette smoke; they exposed strips of rabbit trachea to
smoke from different cigarettes for 12 seconds and identified various
gas phase constituents as having a depressan t effect on mucus
transport. These findings have also been confirmed by others in in
vitro and in in vivo animal experiments (Kensler and Rattista 1963;
Hee and Guillerm 1973; Dalhamn 1956; Albert et al. 1974; Carson et
al. 1966).
The most comprehensive study of individual gas and semivolatile
constituents of cigarette smoke has been conducted by Petterson et
al. (1982). Using chicken tracheal organ cultures, they showed that
at a 5 mm concentration, 36 percent of 316 different compounds
caused ciliostasis after 15 seconds of exposure, but 50 percent were
without effect after an exposure time of 60 seconds. On the basis of
this criterion of separation, either alkylated phenylethers, benzoni-
triles, benxaldehydes, benxenes, napthalenes and indoles, or a-satu-
rated, &unsaturated ketones and aldehydes, or aliphatic alcohols,
aldehydes, acids, and nitrates were found to be ciliotoxic. Inactive
compounds included benxoic acids, esters, polyaromatic hydrocar-

294


bons, amines, and N-heterocycles (except indoles). With respect to
aldehydes, the time to ciliostasis on tissues of rabbit trachea has
been reported shortest for formaldehyde, followed by acetaldehyde,
acrolein, crotonaldehyde, and methacrolein. The ciliotoxic effects of
aldehydes have been confirmed by others using different experimen-
tal approaches (Guillerm et al. 1968; Hee and Guillerm 1973;
Kensler and Battista 1963). It has also been shown that acute
acrolein inhalation causes denudation of ciliated cells, goblet cell
discharge, exfoliation of surface epithelial cells, and infiltration of
inflammatory cells in the lower airways of several different mam-
mals (Dahlgren et al. 1972). Another volatile constituent of cigarette
smoke with marked cilioinhibitory effects is hydrogen cyanide
(Wynder et al. 1965al.
Weissbecker et al. (1971) used a different approach to assess the
effects of several volatile cigarette smoke constituents on mucocili-
ary transport in the cat trachea. The addition of individual volatile
cigarette smoke components (isoprene, nitric oxide, and nitrogen
dioxide) to carbon-filtered cigarette smoke either aggravated the
impairment of tracheal mucus velocity produced by the filtered
smoke or abolished the protection afforded by the carbon filter.
When these constituents were added to whole cigarette smoke, no
further impairment of mucus transport velocity was observed,
indicating a saturation by whole cigarette smoke of receptors
responsible for mucociliary depression.
A direct relation has been reported between tar content and the
ciliotoxic effect of cigarette smoke (Dalhamn and Rylander 1967;
Falk et al. 1959). However, Falk and associates (1959) found no
difference between low tar and high tar cigarette residues with
regard to in vitro mucociliary transport. The effects of nicotine on
mucociliary transport are also controversial, although more investi-
gators have demonstrated a lack of effect (Falk et al. 1959; Guillerm
et al. 1972; Rakieten et al. 1952; Donnelly 1972) than a depression of
mucociliary transport (Carson et al. 1966). Indeed, a biphasic dose-
dependence has been suggested, with stimulation at lower concentra-
tions and depression at higher concentrations (Tsuchiya and Kensler
1959). The stimulation of mucociliary function may be related to
stimulation of nicotinic ganglionic receptors causing choline@
ciliostimulation. This is baaed on the observation that the stimulat-
ing effect of nicotinecontaining cigarettes on the metachronal wave
frequency in the maxillary sinus of anesthetized rabbits is blocked by
atropine and hexamethonium (Hybbinette 1982).
Among the different cigarette tobacco additives, menthol does not
interfere with mucociliary transport (Rakieten et al. 1952). With
respect to phenols, one investigator has reported that the ciliotoxici-
ty of cigarette smoke produced by freezedried tobacco is the same as
that produced by conventionally cured tobacco although the former


contains less phenol (Enzell et al. 1971). On the other hand, phenols
have been shown to impair mucociliary activity and mucus transport
both in vitro (Dalhamn and Lagerstedt 1966; Bernfeld et al. 1964;
Dalhamn 1968) and in vivo (Dalhamn 1968). Dalhamn and associates
[Dalhamn and Lagerstedt 1966; Dalhamn 1968) have even attempted
to relate the toxicity of various phenols to their boiling points.
Addition of the anti-inflammatory agents phenylvinyloxadiozole and
phenylmethyloxadiozole to tobacco has been shown to reduce the
ciliotoxicity of tobacco smoke (Dalhamn and Rylander 1971; Rylan-
der 1971b; Dalhamn 1969), and treatment of rats undergoing long-
term exposure to tobacco smoke with phenylmethyloxadiozole has
been shown to protect the animals against the cigarette-smoke-
induced increase in the number of goblet cells in the respiratory
mucosa (Jones et al. 1973).

It can be concluded from these studies that both the particulate
phase and the gaseous phase of cigarette smoke impair mucociliary
function, that a large number of volatile components are ciliotoxic,
that nicotine may or may not contribute to ciliotoxicity, and that the
additive phenol is ciliotoxic, but the anti-inflammatory agents
phenylmethyloxadiozole and phenylvinyloxadiozole afford partial
protection against the deleterious effects of cigarette smoke. The
mechanisms by which the various constituents of cigarette smoke
interfere with mucociliary transport are unknown. On the basis of
experiments in the fresh water mussel, it has been suggested that
ciliotoxicity depends on their pH in solution (Wynder et al. 1963). It
should be noted, however, that such in vitro experiments requiring
an aqueous medium do not necessarily reflect the type of exposure
occurring in smokers in whom contact between cigarette smoke and
the ciliated epithelium is made by impingement or bypass.

Effects of Filters

Because the toxic effect of cigarette smoke on mucociliary trans-
port mechanisms seems to reside both in the gas phase and in the
particulate phase, the filtering of cigarette smoke before inhalation
may be protective. It has been clearly shown that a longer exposure
time is needed for ciliostasis to occur with smoke from filtered
cigarettes than from unfiltered cigarettes, with respect to both
ciliary activity in vitro and mucociliary transport in vivo (Dalhamn
and Rylander 1964; Dalhamn 1964).

Four major types of filters have been evaluated: cellulose acetate
(Cambridge filter), charcoal, glass fiber, and aqua. The histologic
changes in the airways of guinea pigs exposed to unfiltered cigarette
smoke for 4 to 8 weeks were not seen when cigarette smoke was
passed through a Cambridge filter (Rylander 1974). Likewise,
Kaminski and coworkers (1968) have shown that cellulose-acetate
filters provide protection for the mucociliary activity in the cat

296


trachea. Similar results have been obtained in other experiments
involving in vitro and in vivo systems (Dalhamn and Rylander 1968;
Donnelly 1972; Wynder et al. 1965b), and cellulose-acetate filters
have been found to reduce the inhibitory effect of cigarette smoke on
tracheal epithelial adenylate kinase activity in hamsters exposed for
1 to 5 days (Mattenheimer and Mohr 1975). Charcoal filters are also
capable of reducing the ciliotoxicity of cigarette smoke (Kaminski et
al. 1968; Kensler and Battista 1963; Battista and Kensler 1970a, b).
In one study involving cat tracheas, short-term exposure to a
standardized dose of cigarette smoke decreased particle transport
rates by 50 percent when unfiltered smoke was used, by 40 percent
when the cigarette smoke was passed through a cellulose-acetate
filter, and by 20 percent when a carbon-cellulose filter was used
(Carson et al. 1966). In another comparison of different studies, a
charcoal filter was more effective than a cellulose-acetate filter in
reducing the metachronal wave frequency and mucus transport of
the eulamellibranch gill in vitro (Wynder et al. 1965b). Glass-fiber
and aqua filters were generally less effective (Isawa et al. 1980;
Wynder et al. 1965b).

As expected, better protection might be provided by combined
filters because they remove components of the particulate and the
gaseous phase of cigarette smoke more effectively. Thus, a combina-
tion of cellulose-acetate and charcoal filter has been found to be
more effective than either filter alone (Dalhamn 1966; Wynder et al.
1965bl.

Mucociliary Function in Chronic Bronchitis

Since chronic bronchitis is defined clinically as chronic productive
cough rather than by clearly defined morphologic or functional
abnormalities (American Thoracic Society 19621, some of the previ-
ously reviewed studies of mucociliary function in cigarette smokers
may have included patients with chronic bronchitis as well. Con-
versely, most patients with chronic bronchitis are cigarette smokers
or have been cigarette smokers in the past. Although it is very
difficult to separate the direct effects of cigarette smoke on mucocili-
ary transport from those related to the pathophysiologic changes of
chronic bronchitis, the discussion herein is limited to mucociliary
function in chronic bronchitis without considering the direct effects
of cigarette smoke on the mucosa.

The histologic changes of the mucociliary apparatus in chronic
bronchitis include hypertrophy and hyperplasia of the submucosal
glands, an increase in the number and distribution of goblet cells,
and goblet cell metaplasia in smaller airways (Reid 19671. In
addition, atrophy of the columnar epithelium (Wright and Stuart
1965) and spotty squamous metaplasia (Kleinerman and Boren 19741

297

480-144 0 - 85 - 11


have been reported. A decrease in both the number of ciliated cells
and the mean ciliary length has been noted in the larger airways in
patients with chronic bronchitis (Wanner 1977), and electron micro-
scopic examinations of the airway epithelium show subtle abnormal-
ities in bronchial biopsy material (Miskovitz et al. 1974). Auerbach
and associates (19621, in a large postrmortem study of cigarette
smokers, reported epithellal lesions with loss of cilia in up to 30
percent of random sections, compared with approximately 15
percent of sections from nonsmokers. These ultrastructural changes
consisted of swelling and serration of the epithelium with transfor-
mation of the goblet cell granules. The capsule surrounding the cilia
was irregular, with areas of breakage and outward projections; some
cilia showed fibrillar degeneration or were fused to form compound
cilia.

The presence of visible respiratory secretions is a frequent
endoscopic fmding in patients with chronic bronchitis, and increased
amount of bronchial secretions can be seen on pathologic sections of
the lung (Kleinerman and Boren 1974; Hogg et al. 1968). Thus, the
morphologic changes of chronic bronchitis involve both the ciliary
apparatus and the mucus-producing structures.

cilia

In vitro examination of ciliated lower airway epithelial cells
obtained from chronic bronchitis patients by brushing has failed to
reveal an abnormality in beat frequency Wager et al. 1980).
However, in vitro study of ciliary function is of limited informative
value since the ciliated cells are suspended in an artificial medium
and are not exposed to their natural milieu. This may explain the
discrepancy between this study and one reported by Iravani and Van
As (19721, in which ciliary motion was observed in vivo with an
incident light technique. In the carefully dissected tracheobronchial
tree of rata with experimental chronic bronchitis, the ciliary system
showed discoordination and xonal akinesia. In addition, reversals of
transport direction, whirlpool formations, and inactive zones without
ciliary motion as large as 2 mm by several hundred pm were seen.

Mucus

The distribution, amount, and rheologic properties of mucus
within the airways have not been studied in chronic bronchitis, but
extensive literature exists on the biochemistry @oat and Mathews
1973) and rheology of expectorated sputum from patients with
chronic bronchitis. These results must be interpreted with caution,
partly because of contamination with saliva and the rapid physical
alteration of expectorated sputum, and partly because normal
respiratory secretions for comparison are virtually impossible to
obtain. Mucoid sputum of patients with chronic bronchitis is


biochemically similar to sputum of normal subjects induced by
hypertonic saline aerosol, with the exception of a slightly higher
fucose and neuraminic acid content in the former (Lopata et al.
1974). In purulent sputum from these patients, biochemical changes
typical of inflammatory conditions (increases in the dry weight and
deoxyribonucleic acid content and increased cross-linking by hydra
gen bonding) were observed. Reid (1966) showed that the neuraminic
acid content of sputum is increased in chronic bronchitis, suggesting
augmented secretion by the mucus-producing structures. This find-
ing is supported by histochemical studies indicating distended acini
of the submucosal glands in patients with chronic bronchitis
compared with normal subjects, along with an increase in the
volume of both the acid and the neutral mucopolysaccharide-produc-
ing acini (Reid 1968).
Impaired mucus transport in chronic bronchitis may, in part, be
related to the rheologic abnormalities of respiratory secretions.
Deviation from the ideal ratio between viscosity and elasticity may
prevent an optimal interaction between cilia and mucus, thereby
decreasing mucus transport rates (Dulfano and Adler 1975; Adler
and Dulfano 1976). Higher values of sputum viscosity and lower
values of sputum elasticity have been observed during exacerbations
of chronic bronchitis than during clinical stability (Dulfano et al.
1971). In addition, purulent sputum has a higher viscosity than
mucoid sputum (Charman and Reid 1972; Mitchell-Heggs et al.
19741, suggesting a relationship between the concentration of certain
mucus constituents and mucus rheology. Indeed, examination of
sputum obtained from patients with chronic bronchitis has shown
positive correlations between protein content (particularly IgA) and
mucus glycoprotein content on the one hand and viscosity on the
other (Harbitx et al. 1980, Lopez-Vidriero and Reid 1978). That
altered rheologic properties of airway secretions play a role in
abnormal mucociliary clearance has been suggested by an observed
relationship between in vivo mucociliary clearance, in vitro trans-
portability of expectorated sputum (using the frog palate), and the
viscoelastic properties of sputum (Puchelle et al. 1980).

Mucociliary Interaction

Mucus transport has been studied either by directly or indirectly
observing the motion of discrete particles placed on the tracheal
mucosa (Santa Crux et al. 1974; Goodman et al. 1978) or by the
deposition pattern and clearance rates of ingaled radioactive aerc+
sols (Lourenco 1970; Camner et al. 1973a, b; Luchsinger et al. 1968;
Patrick and Stirling 1977; Dulfano et al. 1971). In one study, a
marked slacking of tracheal mucus velocity -.found in 15 patients
with chronic bronchitis who were between 57 and 71 years of age
(Santa Crux et al. 1974). Clinical examination and pulmonary


FIGURE t.-Comparison of mean (S.E. in bracket) tracheal
      mucociliary transport velocity among young
      nonsmokers (n = lo), elderly nonsmokers (n = 71,
      healthy young exemokers (n=9), healthy
      young smokers (n=lS), and patients with
       chronic bronchitis (n= 14)
so- - et al. (1878).

function tests diagnosed these patients as having both chronic
bronchitis and emphysema. Mucociliary clearance of inhaled aero
sols is also altered in patients with chronic bronchitis. The clearance
of inhaled particles from the lung is influenced by the deposition
pattern, which in turn depends on particle size and flow regime in
the airways. Clearance rates, therefore, can be interpreted only if
particle deposition is carefully monitored (Pircher et al. 1965; Lopez-
Vidriero 1973). Coughing, which is difficult to control in such
patients, may also contribute to the clearance of particles CToigo et
al. 1963). For these reasons, it is not surprising that mucociliary
clearance has been reported to be increased (Muller et al. 1975;
Luchsinger et al. 1968), normal (Thomson and Short 1969), or
decreased (Lourenco 1970; Camner et al. 1973a, b; Tiogo et al. 1963;
Mossberg and Camner 1980; Agnew et al. 1982) in patients with
chronic bronchitis.

Once a subject has developed chronic bronchitis, cessation of
smoking does not reverse the effect on mucociliary function, and a
similar impairment of mucociliary transport has been reported in
smokers and ex-smokers with this disorder (Agnew et al. 1982; Santa
Cruz et al. 1974; Goodman et al. 1978). Persistence of mucociliary