The role of Ca2+ -dependent cell-cell adhesion mol ecules, E- and P-cadherins, in the histogenesis of mouse embryonic lung was studied. All epithelial cells of the lung express both E- and P-cadherin at the early developmental stage. P-cadherin, however, gradually disappears during development, initially from the main bronchi and eventually from all epithelial cells. When a monoclonal antibody to E-cadherin (ECCD-1) was added to monolayer cultures of lung epithelial cells, it induced a partial disruption of their cell-cell adhesion, while a monoclonal antibody to P-cadherin (PCD-1) showed a subtle effect. A mixture of the two antibodies, however, displayed a synergistic effect. We then tested the effect of the antibodies on the morphogenesis of lung primordia using an organ culture system. In control media, the explants formed typical bronchial trees. In the presence of ECCD-1, the explants grew up at the same rate as in the control, but their morphogenesis was affected. The control explants formed round epithelial lobules with an open luminal space at the tips of the bronchial trees, whereas the lobules of explants incu bated with ECCD-1 tended to be flat and devoid of the luminal space. PCD-1 showed a similar but very small effect. A mixture of the two antibodies, however, showed a stronger effect: the branching of epithelia was partially suppressed and the arrangement of epithelial cells was distorted in many places. These results suggest that E and P-cadherin have a synergistic role in the organiz ation of epithelial cells in lung morphogenesis.

The epithelia grow in a tissue-specific pattern under the guidance of the mesenchyme (Wessels, 1977). The mechanism of epithelial morphogenesis, however, is largely unknown. For example, it remains to be eluci dated how the epithelial cells are organized into two dimensional sheets or tubes, how the epithelium branches, and what kinds of signals are exchanged between the epithelium and the mesenchyme.

It is probable that cell adhesion molecules regulate, at least in part, the pattern formation of the epithelial cell sheets, since the cell-cell or cell-substratum con nection is fundamental in multicellular organization. Cadherins are a grou of genetically related glyco proteins involved in Ca2+ -dependent cell-cell adhesion (see Takeichi, 1988 for review). Two subclasses of cadherins, E- and P-cadherin, are detected in various epithelial tissues of mouse embryos (Nose & Takeichi, 1986). These molecules are localized at the cell-cell boundaries (Hirano et al. 1987), and it has been shown that E-cadherin is concentrated in the adherens junctions in certain cell types (Boller et al. 1985). Since the inhibition of E-cadherin activity induces the dis sociation of epithelial sheets (Yoshida-Naro et al. 1984), these molecules probably play a major role in the maintenance of the intercellular junctions in epithelial cells.

In the present study, we examined a role of cadherins in the morphogenesis of lung epithelia, which express both E- and P-cadherin. We cultured the lung primor dia in the presence or absence of antibodies that block the activity of E- or P-cadherin. The results show that abnormal morphogenesis is induced in these explants, especially when the activity of the two cadherins was blocked simultaneously.

Antibodies

Rat monoclonal antibodies ECCD-1 (Yoshida-Norn et al. 1984) and ECCD-2 (Shirayoshi et al. 1986) to E-cadherin, a rat monoclonal antibody PCD-1 to P-cadherin (Nose & Takeichi, 1986) and a rabbit antiserum to E-cadherin (anti-E cadherin; Nagafuchi et al. 1987) were used. ECCD-1 and PCD-1 block the activity of E-cadherin and P-cadherin, respectively. ECCD-2 has no such effect and was therefore used as a control antibody in some experiments.

Solutions of monoclonal antibodies were prepared as fol lows: hybridomas were cultured in high densities in a serum free 1: 1 mixture of Dulbecco’s modified Eagle MEM and Ham’s F12 medium (DH). Ammonium sulphate was added to the culture supernatant to make a 50 % saturated solution and the resulting precipitate was dissolved in a small volume of saline buffered with 10mm-Hepes (pH7·4). Antibodies were purified by gel filtration through a TSK gel G300 SW column (Toso) and the protein concentration in the final antibody solutions was determined using the Bio-Rad Protein Assay.

Immunohistochemical procedures

Fluorescence immunohistochemistry using PCD-1 and ECCD-2 was carried out as described previously (Nose & Takeichi, 1986). Briefly, cryostat sections of tissues fixed with 2 % paraformaldehyde were treated successively with a monoclonal antibody to cadherins, biotinylated anti-rat lg (Amersham) and fluorescein-streptavidin (Amersham). When the polyclonal anti-E-cadherin was used to stain the specimens, this primary antibody was detected using rhoda mine-labelled anti-rabbit lg (Cappel). For the double-staining of E- and P-cadherin, the specimens were treated with a mixture of the rabbit anti-E-cadherin and PCD-1, followed by the treatment using the above-mentioned probes to detect each of the primary antibodies. The stained sections were mounted in 90 % glycerol containing 0·1 % paraphenylene diamine. Photographs were taken with Tri-X film (Kodak) and a Zeiss 18FL fluorescence microscope.

The immunoperoxidase reaction was carried out for stain ing E-cadherin in the whole mount of organ cultures. Tissues on a Nuclepore filter were fixed in 3·5 % paraformaldehyde for one hour and permeabilized by treatment with 100 % methanol at -20°C for 2h. The specimens were then incu bated in the following solutions successively, each for one hour: Tris-buffered saline (pH 7·5) supplemented with 1 mm CaCl2 (TBS) containing 5 % skim milk (STBS), anti-E cadherin rabbit antiserum diluted 1: 250 with STBS, peroxi dase-conjugated anti-rabbit lg (Amersham) diluted 1:100 with STBS. After each incubation, the samples were thoroughly washed with TBS. They were then treated in a 20: 1 mixture of 0·1 M-acetate buffer (pH 5·4) and 0·4 % 3-amino-9-ethyl-carbazole in N,N-dimethylformamide, sup plemented with a small amount of hydrogen peroxide. When the epithelia were stained sufficiently, the samples were washed several times in TBS containing 0·1 % Triton X-100.

Organ cultures

ICR mice were used for all experiments. Lung rudiments were isolated from 11-day fetuses. (The day when the vaginal plug is found is counted as day 0.)

Nuclepore filters (SN110419, 8 μm pore size and 13 mm diameter) were coated with collagen by dipping in a 1: 1 mixture of a collagen solution (Cellmatrix Type I-A, Nitta Gelatin) and ethanol kept at 4°C, and then drying by means of u.v. irradiation. A few tissue rudiments were placed together on a piece of the collagen-coated filter with a small amount of the culture medium, and this explant assembly was floated on 2 ml of the culture medium in a 35 mm plastic dish. Dishes were placed on a Bellco rocker platform moving at 3 strokes min- and incubated at 37°C in a water-saturated atmosphere of 5 % CO2 in air. The DH medium supplemented with 10 % FCS (DH10) was used for the culture. Medium was changed every day.

When necessary, purified ECCD-1 or PCD-1 antibodies were added to the culture medium from the stock solution, so as to make the final concentration 300 μgm1-1. This concen tration was sufficient to completely block the activity of E-cadherin in teratocarcinoma cells and of P-cadherin in PSA5E cells, as determined previously (Yoshida-Norn et al. 1984; Nose & Takeichi, 1986). The control medium contained the same amount of the γ-globulin fraction of rat serum as above, unless otherwise noted.

Monolayer culture of lung epithelial cells

Lungs obtained from 15-day fetuses were treated with 3 % trypsin (DIFCO 1: 250) in Hepes-buffered (pH 7-4) saline containing 10 mm-CaCI2 at 37°C for 30 min. Tissues were then pipetted to separate the epithelial components from mes enchymal cells. Under these conditions, mesenchymal tissues were dissociated into single cells but the epithelia were not. The cell suspension was spun at 400 revs min-1 for one min using a Tomy CD-100R rotor. The resulting pellets were composed mainly of epithelial cell fragments. By repeating this centrifugation several times, the epithelial cells were for the most part purified. These epithelial cells were incubated with DH10 in Falcon culture dishes for one day before use.

lmmunohistological localization of E- and P-cadherin in the lung

E-cadherin was detected in all epithelial cells of the lung rudiments (Fig. lA), and this expression persisted in the lung epithelia throughout development (Fig. lC). P-cadherin was also detected in these cells, but its expression was spatiotemporally regulated. P-cadherin was detected in all epithelial cells of the early lung rudiments (Fig. lB) and, in the later developmental stages, it was detected in the distal regions of bronchial trees but diminished in the main bronchi (Fig. 1D). In new-born mice, this molecule was not detected in all epithelial cells of the lung (data not shown). The mesenthelium surrounding the lung also expresses P-cadherin but not E-cadherin. Mesenchymal cells have neither E-nor P-cadherin. The presence of the two cadherins in the lung was confirmed by immunoblot analysis (Fig. 2).

Fig. 1.

Immunofluorescent localization of E-cadherin (A,C) and P-cadherin (B,D) in the sections of embryonic lungs. (A,B) In a 13-day embryo; (C,D) in a 15-day embryo. The two cadherins were double-stained on each section using FITC and rhodamine-labelled second antibodies. Arrowhead indicates a tubule of the main bronchi in which P-cadherin expression is diminishing. Bar, 50 μm.

Fig. 1.

Immunofluorescent localization of E-cadherin (A,C) and P-cadherin (B,D) in the sections of embryonic lungs. (A,B) In a 13-day embryo; (C,D) in a 15-day embryo. The two cadherins were double-stained on each section using FITC and rhodamine-labelled second antibodies. Arrowhead indicates a tubule of the main bronchi in which P-cadherin expression is diminishing. Bar, 50 μm.

Fig. 2.

Immunoblot detection of the 124×103 Mr E-cadherin (lane A) and the 118×103 Mr P-cadherin (lane B) polypeptide in the lung epithelium of 15-day embryos. The immunoblot analysis was performed using ECCD-2 and PCD-1 for detecting E-cadherin and P-cadherin, respectively, as described previously (Nose & Takeichi, 1986).

Fig. 2.

Immunoblot detection of the 124×103 Mr E-cadherin (lane A) and the 118×103 Mr P-cadherin (lane B) polypeptide in the lung epithelium of 15-day embryos. The immunoblot analysis was performed using ECCD-2 and PCD-1 for detecting E-cadherin and P-cadherin, respectively, as described previously (Nose & Takeichi, 1986).

Effect of cadherin antibodies on monolayer cultures of lung epithelial cells

Lung epithelial cells form colonies in which cells are tightly packed. When the monoclonal antibody ECCD-1 to E-cadherin was added to the cultures, the cell-cell association in the colonies became looser and some cells migrated out of the periphery of the colonies (Fig. 3B,E), while the monoclonal antibody PCD-1 to P-cadherin was only slightly effective (Fig. 3A,D). When the cultures were treated with a mixture of ECCD-1 and PCD-1, an additive effect was observed; the cell-cell connections were more strongly disrupted in these treated colonies (Fig. 3C,F).

Fig. 3.

Effect of cadherin antibodies on the colonies of lung epithelial cells. (A,D) 300 μgm1-1 PCD-1; (B,E) 300 μgm1-1 ECCD-1; (C,F) 300 μg ml-1 PCD-1 + 300 μg ml-1 ECCD-1. The upper panels represent cells before the addition of antibodies. The lower panels represent cells 3 h after the addition of antibodies. Bar, 70 μm.

Fig. 3.

Effect of cadherin antibodies on the colonies of lung epithelial cells. (A,D) 300 μgm1-1 PCD-1; (B,E) 300 μgm1-1 ECCD-1; (C,F) 300 μg ml-1 PCD-1 + 300 μg ml-1 ECCD-1. The upper panels represent cells before the addition of antibodies. The lower panels represent cells 3 h after the addition of antibodies. Bar, 70 μm.

Effect of cadherin antibodies on the morphogenesis of lung primordia

In organ cultures, explants of lung rudiments developed normally showing a typical branching pattern in the control medium, and the same developmental pattern was observed with the medium containing ECCD-2 monoclonal antibody which bound to E-cadherin with out inhibiting its function (Figs 4A,C and 5A,B). We then tested the effect of ECCD-1 or PCD-1 on the morphogenesis of lung rudiments in this system. In order to check the penetration of antibodies into the explants, we incubated the lung primordia with ECCD-2 for one day and stained their sections with a fluor escent anti-rat lg to detect these monoclonal anti bodies. We found that epithelial cells were positively stained after such a treatment (Fig. 4B), indicating that the antibodies penetrate into the tissues.

Fig. 4.

Organ cultures of lung rudiments. (A) An 11-day lung rudiment at the beginning of culture. (B) A test of penetration of antibodies. An explant was incubated with 30011g ml-1 ECCD-2 for one day, sectioned and stained with a FITC-labelled anti-rat IgG. (C-F) Cultured for 4 days in the presence of the control antibody ECCD-2 (C), PCD-1 (D), ECCD-1 (E) and PCD-1 + ECCD-1 (F), Concentrations of antibodies used were the same as in Fig. 3. Bar, 50 μm for Band 300 μm for others.

Fig. 4.

Organ cultures of lung rudiments. (A) An 11-day lung rudiment at the beginning of culture. (B) A test of penetration of antibodies. An explant was incubated with 30011g ml-1 ECCD-2 for one day, sectioned and stained with a FITC-labelled anti-rat IgG. (C-F) Cultured for 4 days in the presence of the control antibody ECCD-2 (C), PCD-1 (D), ECCD-1 (E) and PCD-1 + ECCD-1 (F), Concentrations of antibodies used were the same as in Fig. 3. Bar, 50 μm for Band 300 μm for others.

Fig. 5.

Staining of organ cultures of lung rudiments for E-cadherin to visualize the epithelial structure. Lung rudiments were cultured in the absence (A,B) or presence of ECCD-1 (C,D) and PCD-1 + ECCD-1 (E,F) under the same conditions as in Fig. 4, and then stained with ECCD-2 using a peroxidase method. The right column represents a higher magnification of a portion of the corresponding photographs in the left column. Darker irregular clusters seen on C-F represent an endogenous peroxidase reaction in leukocytes. Bar, 300 μm for the left column; 100 μm for the right column.

Fig. 5.

Staining of organ cultures of lung rudiments for E-cadherin to visualize the epithelial structure. Lung rudiments were cultured in the absence (A,B) or presence of ECCD-1 (C,D) and PCD-1 + ECCD-1 (E,F) under the same conditions as in Fig. 4, and then stained with ECCD-2 using a peroxidase method. The right column represents a higher magnification of a portion of the corresponding photographs in the left column. Darker irregular clusters seen on C-F represent an endogenous peroxidase reaction in leukocytes. Bar, 300 μm for the left column; 100 μm for the right column.

In the presence of ECCD-1, the lung rudiments grew up at the same rate as in the control media, yet the branching pattern of the epithelium was found to be altered especially at the distal regions of the branches (Figs 4E and 5C,D). In control explants, the epithelia formed lobules with a round and smooth morphology, and the line corresponding to the luminal surface of the epithelium was visible with phase-contrast microscopy (Figs 4C and 5A,B). However, in the medium with ECCD-1, the lobules showed a rough and flatter appearance, and the line corresponding to the luminal surface of the epithelium was not clearly visible. In the transverse sections of the explants, the epithelial lobules or tubules in the control explants generally displayed a round or oval shape with a wide luminal space (Fig. 6A). In the presence of ECCD-1, however, they had a crushed appearance and little luminal space, and sometimes the epithelial arrangement was distorted (Fig. 6C). PCD-1 also had a similar but very small effect (Figs 4D and 6B).

Fig. 6.

Transverse sections of lung rudiments in organ cultures. (A) Control. (B) PCD-1. (C) ECCD-1. (D) PCD-1 + ECCD-1. Tissues were cultured for 4 days under the same conditions as in Fig. 4, sectioned and stained with hematoxylin. Arrowheads indicate tubules with the disordered epithelial arrangement. Bar, 50 μm.

Fig. 6.

Transverse sections of lung rudiments in organ cultures. (A) Control. (B) PCD-1. (C) ECCD-1. (D) PCD-1 + ECCD-1. Tissues were cultured for 4 days under the same conditions as in Fig. 4, sectioned and stained with hematoxylin. Arrowheads indicate tubules with the disordered epithelial arrangement. Bar, 50 μm.

We then tested a mixture of ECCD-1 and PCD-1 (one-to-one ratio), and found a more severe pertur bation in lung morphogenesis than above. Under these conditions, the branching of the epithelia did not normally occur and the lobules had a broader and more irregular appearance (Figs 4F and 5E,F). The histologi cal sections showed that the structure of the epithelial tubules was distorted in many places; the alignment of epithelial cells was disrupted and occasionally the lumen of the tubules was packed with randomly arranged epithelial cells (Fig. 6D). This strong effect of the mixture of two antibodies is not simply due to an increase in the total antibody concentration, since ECCD-1 or PCD-1 alone could not exhibit the same effect even when these antibodies were used in a concentration that was double that used in the above experiments.

Since the explants incubated in the presence of the antibodies grew up to the same size as those in the control medium, the antibodies are assumed to have had no effect on the growth of cells in the lung rudiments. The effect of the antibodies was reversible; if the antibodies were removed from the culture me dium after two days of incubation, a normal mor phology was generally restored to the explants within another two days (data not shown). As mentioned, ECCD-2 antibody, which is known to react with E cadherin without blocking its function, had no effect (Fig. 4C). This suggests that the binding of antibodies to the cell surface itself does not affect the morphogen esis of lung epithelium.

The previous observations that the inhibition of cadher ins induces the dissociation of cell layers in monolayer cultures (Damsky et al. 1983; Yoshida-Noro et al. 1984; Behrens et al. 1985; Hatta et al. 1985; Gumbiner & Simons, 1986) imply that cadherins play a role in the morphogenetic assembly of cells in embryos. The pres ent results support this notion by demonstrating that the morphogenesis of the lung epithelium is perturbed by antibodies to cadherins. The abnormal morphogen esis was also induced by blocking the activity of cadherins in such morphogenetic systems as feather formation (Gallin et al. 1986), hair follicle differen tiation (Hirai et al. 1989) and neural retina histogenesis (Matsunaga et al. 1988).

How does the inhibition of cadherin-mediated cell junctions destroy the normal arrangements of epithelial cells? Antibodies to cadherins crushed and deformed the lobules or tubules of the lung epithelium. The primary cause for such deformation of the epithelial structure would be the loss or reduction in the energy for intercellular connection, but the following possi bility should also be considered. We and others suggested that cadherins may be anchored with cortical actin belts (Volk & Geiger, 1986a,b; Hirano et al. 1987). It is known that the cortical actin belt is one of the cytoskel etal components essential for maintaining the tubular structures of the epithelium (Wessels, 1977). It is thus likely that the cadherin-mediated junctions have a function in integrating the actin belts in individual cells into a supracellular ‘tissue-skeleton’. If such an archi tecture were destroyed, the epithelial tubules might lose their rigidity and consequently be crushed and deformed.

The above effect of antibodies was more clearly observed when E-cadherin was blocked, but was not as strong when P-cadherin was inhibited. The simultaneous inhibition of E- and P-cadherin, however, had a synergistic effect. Under these conditions, the arrangement of epithelial cells was severely distorted, and the epithelium could not undergo the normal branching process. This result suggests that both cadherins play an active role in lung epithelial morphogenesis. In contrast to the constitutive expression of E-cadherin, the ex pression of P-cadherin was spatiotemporally regulated. It is therefore possible that E-cadherin has the principal role in maintaining the lung epithelial structure, while P-cadherin may have a more regulatory role. It has been shown, however, that P-cadherin plays a more critical role than E-cadherin in organizing other tissues (Hirai et al. 1989).

As shown in the present study, a cell layer expresses multiple cadherin subclasses, although their combi nations differ with the cell type (Takeichi, 1988). If all cadherin molecules present in a tissue are functioning, the inhibition of the activity of a fraction of these molecules should not be sufficient to block all of the cadherin-mediated stages of histogenesis. From this point of view, the failure to affect metanephric morpho genesis with antibodies against uvomorulin in an exper iment by Vestweber et al. (1985) does not necessarily contradict the present results, as the metanephric tubules express multiple subclasses of cadherins during development (Nose & Takeichi, 1986; Hatta et al. 1987). In lung epithelial cells of the mouse, it is possible that they also express cadherin subclasses other than E and P-cadherin, since we showed that the chicken embryonic lung epithelia transiently express N-cadherin (Hatta et al. 1987). It is therefore likely that the cadherin system plays a more important role in lung epithelial morphogenesis than is indicated by the pres ent observations.

Part of this work was supported by research grants from the Ministry of Education, Science and Culture of Japan.

Behrens
,
J.
,
Birchmeier
,
W.
,
Goodman
,
S. L.
&
Imhof
,
B. A.
(
1985
).
Dissociation of Madin-Darby canine kidney epithelial cells by the monoclonal antibody anti-Arc-1: mechanistic aspects and identification of the antigen as a component related to uvomorulin
.
J. Cell Biol
.
101
,
1307
1315
.
Boller
,
K.
,
Vestweber
,
D.
&
Kemler
,
R.
(
1985
).
Cell adhesion molecule uvomorulin is localized in the intermediate junction of adult intestinal epithelial cells
.
J. Cell Biol
.
100
,
327
332
.
Damsky
,
C. H.
,
Richa
,
J.
,
Solter
,
D.
,
Knudsen
,
K.
&
Buck
,
C. A.
(
1983
).
Identification and purification of a cell surface glycoprotein mediating intercellular adhesion in embryonic and adult tissue
.
Cell
34
,
455
466
.
Gallin
,
w. G.
,
Chuong
,
C.-M.
,
Finkel
,
L. H.
&
Edelman
,
G. M.
(
1986
).
Antibodies to liver cell adhesion molecule perturb inductive interactions and alter feather pattern and structure
.
Proc. natn. Acad. Sci. U.S.A
.
83
,
8235
8239
.
Gumbiner
,
S.
&
Simons
,
K.
(
1986
).
A functional assay for proteins involved in establishing an epithelial occluding barrier: Identification of a uvomorulin-like polypeptide
.
J. Cell Biol
.
102
,
457
468
.
Hatta
,
K.
,
Okada
,
T. S.
&
Takeichi
,
M.
(
1985
).
A monoclonal antibody disrupting calcium-dependent cell-cell adhesion of brain tissues: Possible role of its target antigen in animal pattern formation
.
Proc. natn. Acad. Sci. U.S.A
.
82
,
2789
2793
.
Hatta
,
K.
,
Takagi
,
S.
,
Fujisawa
,
H.
&
Takeichi
,
M.
(
1987
).
Spatial and temporal expression pattern of N-cadherin cell adhesion molecules correlated with morphogenetic processes of chicken embryos
.
Devi Biol
.
120
,
215
227
.
Hirai
,
Y.
,
Nose
,
A.
,
Kobayashi
,
S.
&
Takeichi
,
M.
(
1989
).
Expression and role of E- and P-cadherin adhesion molecules in embryonic histogenesis. II. Skin morphogenesis
.
Development
105
,
271
277
.
Hirano
,
S.
,
Nose
,
A.
,
Hatta
,
K.
,
Kawakami
,
A.
&
Takeichi
,
M.
(
1987
).
Calcium-dependent cell-cell adhesion molecules (cadherins): Subclass-specificities and possible involvement of actin bundles
.
J. Cell Biol
.
105
,
2501
2510
.
Matsunaga
,
M.
,
Hatta
,
K.
&
Takeichi
,
M.
(
1988
).
Roll of N cadherin cell adhesion molecules in the histogenesis of neural retina
.
Neuron
1
,
289
295
.
Nagafuchi
,
A.
,
Shirayoshi
,
Y.
,
Okazaki
,
K.
,
Yasuda
,
K.
&
Takeichi
,
M.
(
1987
).
Transformation of cell adhesion properties by exog nously introduced E-cadherin cDNA
.
Nature, Land
.
329
,
341
343
.
Nose
,
A.
&
Takeichi
,
M.
(
1986
).
A novel cadherin adhesion molecule: Its expression patterns associated with implantation and organogenesis of mouse embryos
.
J. Cell Biol
.
103
,
2649
2658
.
Shirayoshi
,
Y.
,
Nose
,
A.
,
Iwasaki
,
K.
&
Takeichi
,
M.
(
1986
).
N-linked oligosaccharides are not involved in the function of a cell cell binding glycoprotein E-cadherin
.
Cell Struct. Funct
.
11
,
245
252
.
Takeichi
,
M.
(
1988
).
The cadherins: cell-cell adhesion molecules controlling animal morphogenesis
.
Development
102
,
639
655
.
Vestweber
,
D.
,
Kemler
,
R.
&
Ekblom
,
P.
(
1985
).
Cell-adhesion molecule uvomorulin during kidney development
.
Devi Biol
.
112
,
213
221
.
Volk
,
T.
&
Geiger
,
B.
(
1986a
).
A-CAM: a l35kd receptor of intercellular adherens junction. I. Immunoelectron microscopic localization and biochemical studies
.
J. Cell Biol
.
103
,
1441
1450
.
Volk
,
T.
&
Geiger
,
B.
(
1986b
).
A-CAM: a 135kd receptor of intercellular adherens junction. II. Antibody-mediated modulation of junction formation
.
J. Cell Biol
.
103
,
1451
1464
.
Wessels
,
N. K.
(
1977
).
Tissue Interactions and Development
.
Philippines
:
W. A. Benjamin, Inc
.
Yoshida-NöRo
,
C.
,
Suzuki
,
N.
&
Takeichi
,
M.
(
1984
).
Molecular nature of the calcium-dependent cell-cell adhesion system in mouse teratocarcinoma and embryonic cells studied with monoclonal antibody
.
Devi Biol
.
101
,
19
27
.