(CANCER RESEARCH 50, 5481-5487. September 1. 1990]
Peripheral Airway Cell Differentiation in Human Lung Cancer Cell Lines
Adi F. Gazdar,1 R. Ilona Linnoila, Yukio Kurita, Herbert K. Oie, James L. Mulshine, Jean C. Clark, and
Jeffrey A. Whitsett
NCI-Navy Medical Oncology Branch, National Cancer Institute and Naval Hospital, Bethesda, Maryland 20814 [A. F. G., R. 1. L., Y. K., H. K. O., J. L. M.J, and
Pediatrics INeonatologo Division, University of Cincinnati College of Medicine, Cincinnati, Ohio 45267 [J. C. C., J. A. W.]
ABSTRACT
Clara cells and type II pneumocytes are the progenitor cells of the
bronchioles and alveoli, respectively. These peripheral airway cells (PAC)
contain characteristic cytoplasmic structures and express surfactant as
sociated proteins. PAC cell markers are expressed by many pulmonary
adenocarcinomas having papillary and/or lepidic growth patterns, which
are characteristics of the bronchioloalveolar and papillary subtypes. We
investigated the expression of PAC markers in a panel of 41 lung cancer
cell lines. Ultrastructural studies demonstrated the presence of cyto
plasmic structures characteristic of Clara cells or of type II pneumocytes
in 9 of 34 (26%) non-small cell lung cancer cell lines, including 7 of 17
(41%) adenocarcinomas, one squamous cell carcinoma, and one large cell
carcinoma. Of interest, the cytoplasmic structures were present in 5 of 6
(83%) cell lines initiated from papillolepidic adenocarcinomas. In addi
tion, we examined the lines for expression of the surfactant associated
proteins SP-A, SP-B, and SP-C. Eight of the nine cell lines containing
cytoplasmic inclusions characteristic of PAC cells also expressed protein
and/or RNA of SP-A, the major surfactant associated protein. Five of
these lines expressed SP-B RNA (either constitutively or after dexamethasone induction), while a single line expressed SP-C only after
dexamethasone induction. None of six small cell lung cancer cell lines
examined expressed any of the PAC markers. Thus, PAC markers are
expressed frequently (but not exclusively) in pulmonary adenocarcinoma
cell lines, especially in those initiated from tumors having papillolepidic
growth patterns. The establishment and identification of multiple cell
lines expressing PAC features provide an important new resource for
biological and preclinical therapeutic studies.
INTRODUCTION
The World Health Organization classification of lung cancers
includes four major types (squamous cell, large cell, small cell,
and adenocarcinomas) as well as several subtypes and rare
tumors (1). While squamous cell carcinoma has long been
regarded as the commonest type occurring worldwide, several
reports have noted an apparent increase in the incidence of
adenocarcinoma in some countries including the United States
(2-6). The WHO classification includes only light microscopic
examination. A study that combined light and ultrastructural
methodologies indicated that dual adenosquamous differentia
tion was the commonest form of lung carcinoma (7).
The WHO classification of adenocarcinomas consists of four
subtypes, including acinar (gland forming), solid with mucin,
bronchioloalveolar, and papillary (1). It is presumed that acinar
carcinomas arise more centrally from bronchi, while solid
adenocarcinomas represent relatively poorly differentiated tu
mors. BAC2 and papillary carcinomas constitute about one-half
of all pulmonary adenocarcinomas (6). Both of these subtypes
have overlapping pathological features and share common histogenetic origins. BAC tumors, especially when in an invasive
or metastatic mode, cannot always be differentiated from papReceived1/24/90;revised6/1/90.
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1To whom requests for reprints should be addressed.
1 The abbreviations used are: BAC, bronchioloalveolar carcinoma; PAC, pe
ripheral airway cells; cDNA, complementary DNA; NSCLC, non-small cell lung
carcinoma; SCLC, small cell lung carcinoma.
illary carcinomas (6). The cellular origins of both of these
subtypes (in humans and rodents) are heterogeneous and in
clude mucin secreting cells, Clara cells, and type II pneumocytes
(8-16). For these reasons, we prefer to regard BAC and papil
lary tumors as a single entity and refer to them as papillolepidic
tumors, after their main histopathological features.
Because Clara and type II pneumocytes are the progenitor
cells of the bronchioles and alveoli, respectively, we refer to
them as peripheral airway cells. Each of these cell types contains
characteristic cytoplasmic secretory structures. The Clara cells
have dense granules in the apical cytoplasm (17), while type II
pneumocytes contain multi hiniellar bodies (18). The function
of the Clara cell secretions have not been determined, while the
multilamellar bodies of type II cells represent the storage site
of surfactant.
The major function of type II pneumocytes is the production
of pulmonary surfactant, which exerts a detergent-like action
essential for maintaining the patency of the alveoli (for review
see Ref. 19). While phospholipids comprise the major compo
nent of surfactant, several surfactant associated proteins have
been identified which confer important physical properties to
the phospholipids and which are necessary for function. The
most abundant of these proteins is a M, 28,000-36,000 glycoprotein known as SP-A (previously referred to as SAP-35) (2022). The cDNA for human SP-A codes for a smaller polypeptide, which undergoes extensive posttranslational modification.
Recently, two other surfactant proteins, SP-B and SP-C, have
been identified (23). They are M, 5,000-14,000 hydrophobic
proteins and are components of active surfactant extracts uti
lized for the therapy of newborn infants suffering from hyaline
membrane disease. The functions of Clara cells are not as well
defined. They may include secretion of surfactant-like sub
stances, water and salt transport, and mixed-function oxidation
(24-26).
In this report, we describe the expression of morphological,
biochemical, and molecular properties of PAC by several mem
bers of a panel of human lung carcinoma cell lines established
by us. Most of the cell lines expressing PAC features were
derived from adenocarcinomas, especially those having papil
lolepidic growth patterns.
MATERIALS
AND METHODS
Cell Lines. The lung adenocarcinoma cell line A549 (27) was obtained
from the American Type Culture Collection, Rockville, MD. All other
cell lines were established by us. Cell lines were established from
pathologically proved primary or metastatic lung carcinomas as de
scribed previously (28). Solid tumors were finely minced with scissors
and disassociated into small aggregates by pipeting. Malignant effusions
were collected, pelleted, washed, and resuspended in growth medium.
Hemorrhagic effusions also underwent a Ficoll gradient separation
procedure in order to decrease the number of RBC. Approximately
1-5 x IO6cells were seeded into 25-cm2 flasks. While multiple types of
media were utilized, cell lines were usually initiated, established, and
maintained either in RPMI 1640 supplemented with 10% heat inacti
vated fetal bovine serum (RIO) or ACL-4, a serum-free medium we
have devised for the selective culture of NSCLC and other tumors (285481
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PERIPHERAL AIRWAY CELL DIFFERENTIATION
Table 1 Expression of PAC markers in lung cancer cell lines
Forty cell lines were examined for ultrastructural evidence of PAC differentiation. Of these 37 were examined for SP-A protein expression and 30 were examined
for SP RNAs (including all the lines containing cytoplasmic granules characteristic of PAC cells).
Cytoplasmic
granulesTumor
typeAdenocarcinoma
cell4/17°2/22/4°0/110/31/5°0/30/10/30/20/65/40Type
II4/17°0/22/4°2/111/31/5°0/30/10/30/20/66/40SP-Aprotein5/152/23/40/90/20/60/20/10/20/30/65/37SP-A
RNA7/152/23/42/91/20/30/30/1NT*NT0/68/30SP-B
RNA5/15*0/23/42/90/20/30/30/1NTNT0/65/30SP-C
RNA1/15*0/21/40/90/20/30/30/1NTNT0/61/30To
(all)BACPapillaryOtherSquamous
carcinomaLarge
cell
carcinomaAdenosquamous
cell
carcinomaMucoepidermoid
carcinomaCarcinoidMesothcliomaSmall
carcinomaTotalClara
cell
"'One papillary and one large cell carcinoma cell line contained both types of granules (see Table 2).
* Constitutive or dexamethasone induced expression (see Table 2).
c NT, not tested.
Table 2 Lung cancer cell lines expressing PAC cell lines
dehyde, postfixed in 1% osmium tetroxide, and stained with 1% uranyl
SP-A protein
SP-A
acetate, and thin sections were examined by an electron microscope.
RNAAbsentAbsentAbsent1
RNAAbsentAbsentAbsentAbsentAbsent1
RNAType
granules
(ng/mg)
lineNCI-H226NCI-H322NCI-H358NCI-H441-4NC1-H726NC1-H820NCI-H920NCI-HI
Cell
Culture Characterization. Mycoplasma contamination was tested for
C"2
+
by the use of a rRNA hybridization method (Gen-Probe, San Diego,
11ClaraClaraClara
C1+
CA). Cell homogenates were tested for the human forms of the follow
-t-C2
ing enzymes by starch gel (32) using the Authentikit system (Corning
>type
C1+
I1+
Science Products, East Walpole, MA): purine nucleoside phosphorylase
IIType
(EC 2.4.2.1); glucose-6-phosphate dehydrogenase (EC 1.1.1.49); pep+C2
+C1
11Type
IAbsentAbsentAbsent
+
C1+
I1+
IIType
tidase B (EC 3.4.11.4); and láclatedehydrogenase (EC 1.1.1.27). Tu+CAbsent1 +CAbsent1
IIClara
morigenicity was tested for by inoculation of approximately 5 x IO6
334NC1-H1404Cytoplasmic
>type
cells s.c. into the flanks of male athymic nude mice of BALB/c back
11Clara052791700460001202
+CSPA-B +CSPA-C
ground.
°1+, weak expression; 2+, strong expression; C, constitutive expression; I,
Enzyme-linked Immunosorbent Assay for SP-A. SP-A protein was
dexamethasone induced expression.
30). Initial cell passages were performed whenever vigorous tumor cell
growth was observed. Established cell lines were passaged weekly.
Anchorage independent cultures were passaged by transfer of floating
multicellular aggregates. Anchorage dependent cultures were passaged
at subconfluence after trypsinization. If stromal cell growth was noted
in initial cultures, differential trypsinization (31) was used to obtain a
pure tumor cell population. RPMI 1640. trypsin, and sera were ob
tained from Grand Island Biological Co., Grand Island, NY. Growth
factors used in ACL-4 medium were obtained from Sigma Chemical
Co., St. Louis, MO, and Collaborative Research, Bedford, MA. Cul
tures were maintained in humidified incubators at 37°Cin an atmos
phere of 5% CO2 and 95% air.
Morphological Studies. Initial diagnostic materials (pathological and
cytological) were reviewed and compared to the appearances of the
corresponding cell culture and xenograft. Saccomanno fluid fixed cytospin preparations of floating cultures and trypsinized adherent cltures
and paraffin embedded sections of xenografts were stained with hematoxylin-eosin, Alcian blue, and mucicarmine.
For ultrastructural studies, cell pellets were fixed in 2.5% glutaral-
measured by an enzyme-linked immunosorbent assay as described
previously (33). This assay utilizes a goat anti-human SP-A IgG and a
rabbit anti-human SP-A in a capture assay. Lysates from tumor cell
lines were harvested and frozen at —20°C
until analyzed.
Northern Blot Analyses for Surfactant Protein RNAs. RNA was
extracted by the method of Chirgwin et al. (34). For Northern blot
analysis, three identical gels (1.2% agarose, 7% formaldehyde) were
run, loading 20 ¿igof total RNA per lane. After electrophoresing, the
RNA was transferred to Nytran filters (Schleicher and Scimeli. Keene,
NH). Filters were baked at 80°Cfor 2 h prior to hybridization as
described previously (35). Probes used for hybridization (-1 to 5 x IO6
cpm/ml) were partial cDNAs for SP-A (probe 7-1, a 0.9-kilobase clone);
SP-B (probe 4-3, a 1.8-kilobase clone); and SP-C (probe 2-1, a 0.8kilobase cDNA clone) (36-38). Probes were isolated from a Xgtll
expression library generated from adult human lung polyadenylated
RNA. The cDNA probes were labeled with |«-"P]dCTP using a nicktranslation kit.
Dexamethasone Induction of Surfactant Associated Proteins. All cell
lines were tested for SP-A, SP-B, and SP-C RNA while cultured in
their usual maintenance medium (RIO or ACL-4) (constitutive expres
sion). In addition, cell lines constitutively expressing SP-A RNA were
Table 3 Origin of lung cancer cell lines expressing PAC markers
dateMarch
growthAttachedAttachedAttachedAttachedFloatingFloatingFloatingAtt
lineNCI-H226NCI-H322NC1-H358NCI-H441-4NCI-H726NCI-H820NCI-H920NCI-HI
Cell
diagnosis(see
sitePleural
therapyNoneNoneNoneNoneNoneChemo.NoneNoneNoneCulture
mediumRIORIORIORIOACL-4ACL-4ACL-4RIOACL-4Culture
histologySquamousAdenoca.Ade
ef.°LungLungPericard.
1980Feb.
text)BACBACPapillaryAdenoca.PapillaryAdenoca.Large
1981Aug.
1981May
1982Apr.
ef.Pleural
1984Sept.
ef.LNLN1
1984Dec.
1984Feb.
334NC1-H1404Pathological
ilLNAge(yr)952995753445648SexMMMMFMMMMCulture
piilm.
1986Dec.
cellPapillaryTumor
1986Prior
" Pleural ef.. pleura! effusion; Pericard. fl., pericardial effusion; LN. lymph node; epidural. epidural mass; chemo., patient received prior chemotherapy; Adenoca.,
adenocarcinoma; adenosq. adcnosquamous carcinoma: NT. not tested.
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PERIPHERAL
AIRWAY CELL DIFFERENTIATION
b
d
Fig. 1. Comparison of tumor, culture, and xenograft morphologies of cell line NCI-H820. a, malignant pleural effusion, demonstrating papillary clusters of
malignant cells; l>.supraclavicular lymph node from same patient, with metastatic adenocarcinoma (most of the lymph node consisted of undifferentiated metastatic
tumor, with foci of papillary carcinoma (as illustrated)); c. cell line NCI-H820. initiated from lymph node metastasis, demonstrating floating clusters of tumor cells
forming papillary structures; (/. xenograft morphology, cell line NCI-H820. demonstrating tubulopapillary morphology.
tested for expression of SP-B and SP-C RNA after growth in the
presence of dexamethasone (induced expression). Because ACL-4 me
dium contains hydrocortisone (S x 10 " M), cells maintained in that
medium were cultured for at least 2 days in the ACL-4 medium lacking
hydrocortisone and then in fresh medium lacking hydrocortisone but
containing dexamethasone. Dexamethasone (Sigma) was prepared as a
10 /¿M
stock in 95% ethanol and diluted to the stated concentrations in
hydrocortisone-free medium.
RESULTS
Selection of Cell Lines. While we have initiated more than 75
NSCLC cell lines since 1977, 29 were available at the initiation
of these studies in 1985. An additional 6 adenocarcinoma cell
lines, initiated at a subsequent time point, also were studied. In
addition to the 35 NSCLC lines, six SCLC cell lines were
randomly selected from a panel of approximately 200 lines (39).
All of the cell lines studied expressed human forms of the
enzymes analyzed, and they were free of Mycoplasma contam
ination.
Expression of PAC Markers in Lung Cancer Lines. We ex
amined our panel of 41 lung cancer cell lines for ultrastructural,
biochemical, or molecular evidence of PAC differentiation.
While not all cell lines were examined by all techniques (Table
1), all cell lines identified as containing ultrastructural evidence
of PAC differentiation were examined for SP protein and RNA
expression. One or more markers were present in 9 of 35 (26%)
NSCLC lines (Tables 1 and 2). Multiple markers were present
in 7 of 17 (41%) adenocarcinomas (including 3 of 4 papillary
carcinomas and 2 of 2 BACs), 1 of 6 large cell and 1 of 3
squamous cell carcinomas. The latter cell line, NCI-H226, was
initiated from a malignant pleural effusion with an original
cytological diagnosis of "tumor cells, possibly mesothelioma."
However, examination of the cell line demonstrated multiple
biochemical and ultrastructural features of squamous cell car
cinoma (40). The large cell carcinoma line (which only had
ultrastructural evidence of PAC differentiation) was derived
from a tumor that lacked morphological evidence of differen
tiation. However, the corresponding cell culture contained a
gland forming subpopulation. PAC markers were not detected
in carcinoids, mesotheliomas, adenosquamous, mucoepidermoid, or SCLC lines.
Properties of Cell Lines Expressing PAC Markers. The origin
and properties of nine cell lines expressing one or more PAC
markers are presented in Tables 2 and 3. Two cell lines were
initiated from primary tumors, and seven were from métastases.
They had been cultured for periods of 2 to 8 years when
characterized. All but one of the patients were males, and only
one patient had received therapy prior to culture. The Five
cultures that had been initiated in serum containing RIO me
dium demonstrated substrate attachment and had an epithelioid
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PERIPHERAL
Fig. 2. Ultraslructural appearances of cell
lines expressing PAC features, a, ultrastruc
tural appearances of cell line NCI-H820. The
cells demonstrated polarity, with microvilli
being present on the apical surfaces. Many
multilamellar bodies are present in the apical
cytoplasm. Because of fixation artifact, the
phospholipid contents of many of the bodies
have been dissolved. Original magnification.
x7,050. Insel, a relatively well preserved mul
tilamellar body. A, a multilamellar body in the
process of being extruded from the apical cell
surface of cell line NCI-H726. Original mag
nification. X 22.400.
AIRWAY CELL DIFFERENTIATION
i
••'-s
fM .
*££\ •'•
morphology. The four adenocarcinoma cultures that had been
initiated in defined ACL-4 medium (which lacks attachment
factors) lacked substrate attachment. They formed papillary
(Fig. 1) or inter- and intracellular gland-like structures, the
lumina of which were distended with fluid, not mucin. Adeno
carcinoma cells demonstrated polarity, with microvilli being
present on the apical surfaces, and they were joined by tight
junctions and desmosomes. All 5 cell lines tested were tumorigenie in athymic nude mice. In general, xenograft histology
resembled that of the original tumor (Fig. 1). However, the
xenograft histology of one BAC cell line was adenosquamous cell carcinoma.
Ultrastructural Evidence of PAC Differentiation. All 9 cell
lines expressing PAC markers demonstrated cytoplasmic gran
ules characteristic of Clara or type II pneumocytes (Fig. 2).
Both of these types of granules were located in the apical
cytoplasm. Clara cell granules consisted of round, oval, or
irregularly shaped membrane bound structures (about 0.3 urn
in diameter) with finely granular contents of variable electron
density. The type II cell inclusions consisted of larger, oval
membrane bound structures having whorled or lamellar osmiophilic structures. The contents of many granules appeared to
have been partially or completely dissolved by fixation. On
occasion, the limiting membrane of the lamellar bodies was
observed to be fused with the cell membrane, and the contents
were extruded into the extracellular space (Fig. 2). Clara cell
and type II cell granules were each observed in three cell lines,
while two cell lines contained predominantly Clara cell gran
ules, with a subpopulation of cells containing type II cell gran
ules.
Expression of Surfactant Proteins. SP-A RNA was detected
in 8 of the 9 PAC cell lines, and in 5 of these SP-A protein was
also demonstrated (Table 2; Fig. 3). SP-B RNA (constitutive
or induced) was detected in 5 of the lines demonstrating SP-A
RNA. In a single cell line, NCI-H820, SP-C RNA was detected.
When this cell line was maintained in hydrocortisone contain
ing ACL-4 medium, a faint band was occasionally visible on
Northern blots. However, SP-C RNA could be readily detected
after dexamethasone induction. In NCI-H820 cells, expression
of SP-A, SP-B, and SP-C RNAs was increased after dexameth-
5484
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PERIPHERAL
8cr>
I
CD
O
co LO
III
AIRWAY CELL DIFFERENTIATION
reported to express features of type II cells (27). While multilamellar cytoplasmic structures were readily found on ultrastructural examination, neither constitutive nor dexamethasone
induced expression of SP-A protein or SP-A, SP-B, or SP-C
RNAs were detected.
CD
CM 00 CO CO 00 LO
LO TÕ Tt
CO CM
CM
CM co
i- i- oo i- CO
CM CM CD
CO
I
I
I
11
I I I
DISCUSSION
SP-A
—2 2 Kb
SP-B
-2.0kb
Fig. 3. Northern blots, demonstrating constitutive expression of SP-A and
SP-B RNA by lung cancer cell lines. Top, SP-A RNA (2.20-kilobase band)
expressed by cell lines NCI-H920 (adenocarcinoma), NCI-H441 (BAC), NCIH358 (BAC), and NCI-H226 (squamous cell carcinoma); bottom, fainter 2.0kilobase (*e) band, representing SP-B expression, present in cell lines NCI-H920
andNCI-H441.
SP-A
—22
SP-B
-2.0kb
SP-C
•
—10
kb
kb
tures. In 1976 Lieber et al. (27) described ultrastructural fea
tures and phospholipid formation characteristic of type II pneu
mocytes in A549 cells and claimed that the line was initiated
from an "alveolar cell carcinoma." However, their illustration
0 0.1 1.0 10 50 100
Dexamethasone
^e f°un(ievidence of PAC differentiation in 9 lung cancer
cell lines, 7 of which were adenocarcinomas. Five of the ade
nocarcinoma lines were initiated from tumors that had papillolepidic growth features. All of the 9 cell lines had ultrastruc
tural characteristics of Clara cells or type II pneumocytes and
8 of them expressed SP-A RNA constitutively. In addition, five
lines expressed SP-B constitutively or after dexamethasone
stimulation, while a single line expressed SP-C RNA only after
dexamethasone stimulation. We have previously described
expression of surfactant proteins by two of the PAC cell lines
described in this report, NCI-H441-4 and NCI-H820 (35, 41,
42).
Surfactant proteins were expressed by lung cancer cell lines
demonstrating ultrastructural features consistent with type II
pneumocytes or Clara cells. On occasion, features of both cell
types were present in the same lines. These findings have been
described previously in human tumors (43, 44). While Clara
cells express a characteristic M, 10,000 protein (45), normal
and neoplastic Clara cells also may express surfactant proteins
independently of surfactant phospholipids (26, 46). Our study
and others discussed above indicate the interrelationship be
tween Clara and type II pneumocyte cells and suggest that PAC
tumors arise from bronchiolar or alveolar cells capable of
multipotent differentiation. In addition, PAC differentiation
may, occasionally, be present in more centrally arising NSCLC
tumors. We did not detect evidence of PAC differentiation in
SCLC cell lines. In another study, we failed to demonstrate SPA by immunohistochemical techniques in SCLC tumors or in
adenocarcinomas of nonpulmonary origin (46).
Previous reports of established cell lines that express PAC
features are very limited. A recent report in the Chinese litera
ture describes lamellar bodies in a subline of a pulmonary
carcinoma culture, but not in the parent cell line (47). Multilamellar bodies have also been described in a cell line established
from the lung of a sheep with jaagsiekte (ovine pulmonary
adenomatosis) (48). Jaagsiekte is a virall y induced ovine disease
characterized by hyperplastic or tumorous proliferation of type
II pneumocytes. We also studied A549 cells, a human lung
carcinoma cell line widely believed to be of BAC origin. The
A549 line was described by Giard and associates in 1973 as
being initiated from a tumor designated simply as "lung carcinoma" (49). Xenografts of the cell line contained acinar struc
(nM)
Fig. 4. Northern blots of cell line NCI-H820, derived from a papillary ade
nocarcinoma, demonstrating dose dependent dexamethasone induction of surfac
tant protein RNAs SP-A, SP-B, and SP-C. The cell line was cultured in steroid
free medium for 48 h and then exposed to varying concentrations of dexametha
sone for 4 h. kh. kilobases.
asonéstimulation in a dose dependent manner (Fig. 4).
Properties of A549 Cells. In addition to our cell lines, we
examined A549 cells, a pulmonary adenocarcinoma cell line
(and description) of the tumor histology is that of a gland
forming acinar carcinoma with mucin formation. Other pub
lished reports have suggested that A549 cells do not express
biochemical properties characteristic of type II cells (50, 51).
In addition, TGF-/3 induces goblet cell differentiation in A549
cells (52). While our studies confirmed previous reports that
A549 cells contained cytoplasmic lamellar structures (27), the
cells failed to express surfactant protein or RNAs. The ultrastructural appearances of Clara and type II granules are not
completely diagnostic, and other cell types may contain artifac-
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PERIPHERAL
AIRWAY CELL DIFFERENTIATION
tual or real inclusions with similar morphologies (53, 54). Thus,
we regard only those cell lines that express both ultrastructural
and molecular or biochemical markers as having unequivocal
evidence of PAC differentiation. Using this criterion, eight of
our cell lines appear to be the first continuous lines of any
species that demonstrate definite evidence of PAC differentia
tion.
While most of our cell lines expressing PAC features were
initiated from papillolepidic tumors, these tumors form a het
erogeneous group, as discussed previously. In addition, some
other NSCLC lung tumors may express markers of PAC dif
ferentiation (43, 50, 55). Also, métastasesfrom non-lung car
cinomas may have papillolepidic growth features (44, 56, 57).
Thus, papillolepidic morphology and expression of PAC mark
ers are suggestive, but not diagnostic, of PAC histogenesis.
Our PAC cell lines may be useful for several types of studies
including: (a) the physiology of surfactant and its proteins (41,
42); (b) the biology of Clara cells and their role in xenobiotic
metabolism; (c) in vitro testing of new therapeutic modalities
specifically directed at PAC cells, such as 4-ipomeanol (58);
and (d) preparation of human-derived materials potentially
useful for the treatment of the respiratory distress syndrome of
the newborn and other surfactant deficient states (21, 59). The
establishment of multiple human cell lines manifesting une
quivocal evidence of PAC differentiation provides a major new
resource for biological and preclinical therapeutic studies.
18.
19.
20.
21.
22.
23.
24.
25.
26.
27.
28.
29.
30.
31.
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Peripheral Airway Cell Differentiation in Human Lung Cancer
Cell Lines
Adi F. Gazdar, R. Ilona Linnoila, Yukio Kurita, et al.
Cancer Res 1990;50:5481-5487.
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