EDITORIAL FOCUS
Bronchial endothelial cell phenotypes and the form:function
relationship
Troy
Stevens
Department of Pharmacology, Center for Lung Biology,
University of South Alabama College of Medicine, Mobile, Alabama 36688
 |
ARTICLE |
CULTURED SYSTEMS HAVE PROVEN highly
useful for generating insight into normal and abnormal cell function,
as illustrated by Brown et al. (1-3) in
identification of mechanism(s) underlying familial
hypercholesterolemia. In studies by Brown and coworkers, the
isolation and culture of fibroblasts were made with relative ease,
whereas other cell types are not readily accessible. Indeed, the
generally poor accessibility of bronchial endothelial cells has
hampered their in vitro purification. The study by Moldobaeva and
Wagner, one of the current articles in focus (Ref. 11a, see p. L520 in this issue), reports successful isolation and
culture of endothelial cells from the bronchial artery and
microcirculation and ascribes the cell's origin to unique barrier
regulatory properties.
To isolate bronchial artery endothelial cells (BAEC), the
bronchoesophageal artery was perfused and incubated with collagenase. Dissociated cells were collected and plated on gelatin-coated culture
dishes. After 4-5 days, colonies displaying the typical cobblestone morphology were transferred using cloning disks for expansion. To isolate bronchial microvascular endothelial cells, the
mainstem bronchus was dissected, and its epithelium was
removed. The microvessels to the cartilage were then dissected,
incubated with collagenase, and filtered through a nylon mesh. Cells
collected were grown on gelatin-coated dishes, and, after 4-5
days, were selected for expansion based on their typical cobblestone
morphology. Both bronchial artery and microvascular cells
displayed appropriate endothelial markers, including uptake of
low-density lipoprotein, factor VIII/von Willebrand-associated antigen,
and platelet-endothelial cell adhesion molecule-1 immunoreactivity, and
were therefore judged to be representative of endothelia from the
respective vascular sites.
It is now clear that not all endothelial cells are alike. For some
time, it was evident that endothelial cells differed on the basis of
their intercellular junctions and could be generally classified as
"tight," "continuous," or "fenestrated." In addition, high
endothelial venules importantly regulate white blood cell recruitment
(4, 6), and postcapillary venules form intercellular gaps
in response to inflammatory agonists (18), both highly specific attributes of very specific cells. There is even evidence that
endothelial cells immediately adjacent exhibit unique functions, some
with so-called pacemaker activity that regulates cytosolic calcium
concentrations (19). The cause of such disparate
endothelial cell function between and within vascular beds is not fully
understood but likely reflects environmental and epigenetic causes
(16).
Moldobaeva and Wagner (11a) describe unique bronchial macro- and
microvascular endothelial cell function in two aspects, including cell
proliferation and barrier function. Their proliferation studies resolved that BAEC grow faster than their microvascular counterparts. The macrovascular cells grew equally well in any of six different media
supplements. Remarkably, microvascular endothelial cells did not grow
equally well in all media supplements; use of MCDB 131 essentially
abolished growth. This medium was designed to optimize
serum-free growth and has been successfully utilized to enhance conduit
artery and microvascular endothelial cell proliferation (11, 14,
17). Absence of a growth response in bronchial microvascular
endothelial cells using this medium provides important information
regarding differences between these and BAEC and also regarding
mechanisms controlling microvascular endothelial cell growth. It will
be important to identify which component(s) of the MCDB 131 medium
arrests microvascular endothelial cell growth. Similar observations
have previously been extended to isolate multiple smooth muscle cell
phenotypes from the pulmonary artery wall (7-9, 15).
Because microvascular endothelial cells were isolated from the
bronchial mucosa, they were expected to possess greater permeability responses than BAEC, particularly in response to bradykinin. Basal dextran permeability was greater in microvascular cells than in macrovascular cells. Bradykinin transiently increased dextran (9.5 kDa)
transfer across both cell types, although the magnitude of this effect
was greater in microvascular than in macrovascular cells. Bradykinin
did not increase the transfer of 77-kDa dextrans, suggesting a size
restriction in gaps that formed. Microvascular endothelial cells also
responded to thrombin with an increase in permeability, whereas
macrovascular endothelial cells did not. Water permeability was not
measured in these studies. In the future, it may be important to
determine whether microvascular endothelial cells isolated from the
bronchial mucosa possess enhanced water permeability, which contributes
to airway humidification (12).
It is interesting that BAEC exhibit increased growth and permeability
responses compared with bronchial microvascular endothelial cells.
These parameters have been assessed in other vascular beds, but perhaps
the most intriguing comparison is with endothelial cells from the
pulmonary circulation. In this case, lung microvascular endothelial
cells grow faster than their macrovascular counterparts (unpublished
observations), and they also exhibit a more restrictive barrier
function (5, 10, 13). Macro- and microvascular endothelial
cells isolated from the bronchial circulation, therefore, exhibit
exactly opposite behavior than do respective cell types isolated from
the pulmonary circulation. These findings illustrate the need to
determine influences, environmental and epigenetic, that underlie
site-specific endothelial phenotype. Ultimate success of therapeutic
strategies to target endothelia will require an ability to discern
between organ- and site-restricted phenotypes.
In summary, Moldobaeva and Wagner (11a) report an important advance in
airway and endothelial cell biology. Evidence for distinct behavior of
bronchial macro- and microvascular endothelial cells, even under
similar environmental conditions, suggests the cell types possess
fundamentally different phenotype and function. Successful isolation
and culture of these cells may now reveal mechanistic insight into the
phenotype of these cell populations for rigorous comparison with their
behavior in vivo.
 |
FOOTNOTES |
Address for reprint requests and other correspondence: T. Stevens, Dept. of Pharmacology, Center for Lung Biology, Univ. of South
Alabama College of Medicine, Mobile, AL 36688 (E-mail:
tstevens{at}jaguar1.usouthal.edu).
10.1152/ajplung.00103.2002
 |
REFERENCES |
1.
Brown, MS,
Dana SE,
and
Goldstein JL.
Regulation of 3-hydroxy-3-methylglutaryl coenzyme A reductase activity in cultured human fibroblasts. Comparison of cells from a normal subject and from a patient with homozygous familial hypercholesterolemia.
J Biol Chem
249:
789-796,
1974[Abstract/Free Full Text].
2.
Brown, MS,
and
Goldstein JL.
Expression of the familial hypercholesterolemia gene in heterozygotes: mechanism for a dominant disorder in man.
Science
185:
61-63,
1974[ISI][Medline].
3.
Brown, MS,
and
Goldstein JL.
Familial hypercholesterolemia: defective binding of lipoproteins to cultured fibroblasts associated with impaired regulation of 3-hydroxy-3-methylglutaryl coenzyme A reductase activity.
Proc Natl Acad Sci USA
71:
788-792,
1974[Abstract].
4.
Cavender, DE.
Organ-specific and non-organ-specific lymphocyte receptors for vascular endothelium.
J Invest Dermatol
94:
41S-48S,
1990[Abstract].
5.
Chetham, PM,
Babal P,
Bridges JP,
Moore TM,
and
Stevens T.
Segmental regulation of pulmonary vascular permeability by store-operated Ca2+ entry.
Am J Physiol Lung Cell Mol Physiol
276:
L41-L50,
1999[Abstract/Free Full Text].
6.
Colditz, IG.
Margination and emigration of leucocytes.
Surv Synth Pathol Res
4:
44-68,
1985[ISI][Medline].
7.
Dempsey, EC,
Das M,
Frid MG,
and
Stenmark KR.
Unique growth properties of neonatal pulmonary vascular cells: importance of time- and site-specific responses, cell-cell interaction, and synergy.
J Perinatol
16:
S2-S11,
1996[Medline].
8.
Dempsey, EC,
Frid MG,
Aldashev AA,
Das M,
and
Stenmark KR.
Heterogeneity in the proliferative response of bovine pulmonary artery smooth muscle cells to mitogens and hypoxia: importance of protein kinase C.
Can J Physiol Pharmacol
75:
936-944,
1997[ISI][Medline].
9.
Frid, MG,
Moiseeva EP,
and
Stenmark KR.
Multiple phenotypically distinct smooth muscle cell populations exist in the adult and developing bovine pulmonary arterial media in vivo.
Circ Res
75:
669-681,
1994[Abstract].
10.
Kelly, JJ,
Moore TM,
Babal P,
Diwan AH,
Stevens T,
and
Thompson WJ.
Pulmonary microvascular and macrovascular endothelial cells: differential regulation of Ca2+ and permeability.
Am J Physiol Lung Cell Mol Physiol
274:
L810-L819,
1998[Abstract/Free Full Text].
11.
Knedler, A,
and
Ham RG.
Optimized medium for clonal growth of human microvascular endothelial cells with minimal serum.
In Vitro Cell Dev Biol
23:
481-491,
1987[ISI][Medline].
11a.
Moldobaeva, A,
and
Wagner EM.
Heterogeneity of bronchial endothelial cell permeability.
Am J Physiol Lung Cell Mol Physiol
283:
L520-L527,
2002.
12.
Nielsen, S,
King LS,
Christensen BM,
and
Agre P.
Aquaporins in complex tissues. II. Subcellular distribution in respiratory and glandular tissues of rat.
Am J Physiol Cell Physiol
273:
C1549-C1561,
1997[Abstract/Free Full Text].
13.
Schnitzer, JE,
Siflinger-Birnboim A,
Del Vecchio PJ,
and
Malik AB.
Segmental differentiation of permeability, protein glycosylation, and morphology of cultured bovine lung vascular endothelium.
Biochem Biophys Res Commun
199:
11-19,
1994[ISI][Medline].
14.
Stein, GH,
and
St. Clair JA.
Human microvascular endothelial cells: coordinate induction of morphologic differentiation and twofold extension of life span. In Vitro
Cell Dev Biol
24:
381-387,
1988.
15.
Stenmark, KR,
and
Frid MG.
Smooth muscle cell heterogeneity: role of specific smooth muscle cell subpopulations in pulmonary vascular disease.
Chest
114:
82S-90S,
1998[Free Full Text].
16.
Stevens, T,
Rosenberg R,
Aird W,
Quertermous T,
Johnson FL,
Garcia JG,
Hebbel RP,
Tuder RM,
and
Garfinkel S.
NHLBI workshop report: endothelial cell phenotypes in heart, lung, and blood diseases.
Am J Physiol Cell Physiol
281:
C1422-C1433,
2001[Abstract/Free Full Text].
17.
Terramani, TT,
Eton D,
Bui PA,
Wang Y,
Weaver FA,
and
Yu H.
Human macrovascular endothelial cells: optimization of culture conditions.
In Vitro Cell Dev Biol Anim
36:
125-132,
2000[ISI][Medline].
18.
Thurston, G,
Baluk P,
and
McDonald DM.
Determinants of endothelial cell phenotype in venules.
Microcirculation
7:
67-80,
2000[ISI][Medline].
19.
Ying, X,
Minamiya Y,
Fu C,
and
Bhattacharya J.
Ca2+ waves in lung capillary endothelium.
Circ Res
79:
898-908,
1996[Abstract/Free Full Text].
Am J Physiol Lung Cell Mol Physiol 283(3):L518-L519
1040-0605/02 $5.00
Copyright © 2002 the American Physiological Society