Epithelial, fibroblast and intermediate cell lines were employed to examine the mechanism(s) essential for heterocellular gap junction intercellular communication in vitro. These cell lines were characterized extensively for cell type based on morphology, intermediate cytoskeletal proteins, cell adhesion molecules and their associated proteins, tight junction proteins as well as functional differentiation. All cell types expressed connexin43 and were dye-coupled in homocellular culture. Epithelial and intermediate cells or fibroblasts and intermediate cells readily assembled heterocellular connexin43-positive gap junction plaques when co-cultured, while gap junction plaques in mixed cultures of epithelial cells and fibroblasts were rare. Dye microinjection studies were used to show that there was little gap junction intercellular communication between epithelial cells and fibroblasts. However, intermediate cells were able to communicate with epithelial cells and, to a lesser extent, fibroblasts and could transfer dye to both epithelial cells and fibroblasts when all three cell types were cultured together. Fibroblasts that were stably transfected with a cDNA encoding E-cadherin had a greater tendency to aggregate and exhibited a more epithelial-like phenotype but heterocellular gap junction intercellular communication with epithelial cells, which endogenously express E-cadherin, was not enhanced. These results suggest that mutual expression of E-cadherin is insufficient to stimulate gap junction formation between epithelial cells and fibroblasts. Moreover, our results also demonstrate that communication gaps between epithelial cells and fibroblasts can be bridged by intermediate cells, a process that may be important in mammary gland development, growth, differentiation and cancer.

REFERENCES

Bassnett
S.
,
Kuszak
J. R.
,
Reinisch
L.
,
Brown
H. G.
,
Beebe
D. C.
(
1994
).
Intercellular communication between epithelial and fiber cells of the eye lens.
J. Cell Sci
107
,
799
811
Berga
S. E.
(
1984
).
Electrical potentials and cell-to-cell dye movement in mouse mammary gland during lactation.
Am. J. Physiol
247
,
20
–.
Birkedal-Hansen
H.
,
Moore
W. G. I.
,
Bodden
M. K.
,
Windsor
L. J.
,
Birkedal-Hansen
B.
,
DeCarlo
A.
,
Egler
J. A.
(
1993
).
Matrix metalloproteinases: a review.
Crit. Rev. Oral. Biol. Med
4
,
197
250
Bruzzone
R.
,
White
T. W.
,
Paul
D. L.
(
1996
).
Connections with connexins: The molecular basis of direct intercellular signaling.
Eur. J. Biochem
238
,
1
27
Cyr
D. G.
,
Blaschuk
O. W.
,
Robaire
B.
(
1992
).
Identification and developmental regulation of cadherin messenger ribonucleic acids in the rat testis.
Endocrinology
131
,
139
145
Cyr
D. G.
,
Hermo
L.
,
Laird
D. W.
(
1996
).
Immunocytochemical localization and regulation of connexin43 in the adult rat epididymis.
Endocrinology
137
,
1474
1484
Fentiman
I.
,
Taylor-Papadimitriou
J.
,
Stoker
M.
(
1976
).
Selective contact-dependent cell communication.
Nature
264
,
760
762
Goodenough
D. A.
,
Dick
J. S. B.
II
,
Lyons
J. E.
(
1980
).
Lens metabolic cooperation: A study of mouse lens transport and permeability visualized with freeze-substitution autoradiography and electron microscopy.
J. Cell Biol
86
,
576
589
Goodenough
D. A.
,
Goliger
J. A.
,
Paul
D. L.
(
1996
).
Connexins, connexons, and intercellular communication.
Annu. Rev. Biochem
65
,
475
502
Guelstein
V. I.
,
Tchypysheva
T. A.
,
Ermilova
V. D.
,
Ljubimov
A. V.
(
1993
).
Myoepithelial and basement antigens in benign and malignant human breast tumors.
Int. J. Cancer
53
,
269
277
Huynh
H. T.
,
Robitaille
G.
,
Turner
J. D.
(
1991
).
Establishment of bovine mammary epithelial cells (MAC-T): an in vitro model for bovine lactation.
Exp. Cell Res
197
,
191
199
Jongen
W. M. F.
,
Fitzgerald
D. J.
,
Asamoto
M.
,
Piccoli
C.
,
Slaga
T. J.
,
Gros
D.
,
Takeichi
M.
,
Yamasaki
H.
(
1991
).
Regulation of connexin43-mediated gap junctional intercellular communication by Ca2+in mouse epidermal cells is controlled by E-cadherin.
J. Cell Biol
114
,
545
555
Kam
E.
,
Melville
L.
,
Pitts
J. D.
(
1986
).
Patterns of junctional communication in skin.
J. Invest. Dermatol
87
,
748
753
Kanno
Y.
,
Sasaki
Y.
,
Shiba
Y.
,
Yoshida-Noro
C.
,
Takeichi
M.
(
1984
).
Monoclonal antibody ECCD-1 inhibits intercellular communication in teratocarcinoma PCC3 cells.
Exp. Cell Res
152
,
270
274
Laird
D. W.
(
1996
).
The life cycle of a connexin; Gap junction formation, removal, and degradation.
J. Bioenerg. Biomembr
28
,
311
318
Laird
D. W.
,
Revel
J.-P.
(
1990
).
Biochemical and immunochemical analysis of the arrangement of connexin43 in rat heart gap junction membranes.
J. Cell Sci
97
,
109
117
Laird
D. W.
,
Castillo
M.
,
Kasprzak
L.
(
1995
).
Gap junction turnover, intracellular trafficking and phosphorylation of connexin43 in brefeldin-A treated rat mammary tumor cells.
J. Cell Biol
131
,
1193
1203
Lee
S. W.
,
Tomasetto
C.
,
Paul
D.
,
Keyomarsi
K.
,
Sager
R.
(
1992
).
Transcriptional down-regulation of gap junction proteins blocks junctional communication in human mammary tumor cell lines.
J. Cell Biol
118
,
1213
1221
Mege
R. M.
,
Matsuzaki
F.
,
Gallin
W. J.
,
Goldberg
J. I.
,
Cunningham
B. A.
,
Edelman
G. M.
(
1988
).
Construction of epitheloid sheets bytransfection of mouse sarcoma cells with cDNAs for chicken cell adhesion molecules.
Proc. Nat. Acad. Sci. USA
85
,
7274
7278
Mesnil
M.
,
Piccoli
C.
,
Klein
J. L.
,
Morand
I.
,
Yamasaki
H.
(
1993
).
Lack of correlation between the gap junctional communication capacity of human colon cancer cell lines and expression of the DCC gene, a homologue of a cell adhesion molecule (N-CAM).
Jpn J. Cancer Res
84
,
742
747
Meyer
R. A.
,
Laird
D. W.
,
Revel
J.-P.
,
Johnson
R. G.
(
1992
).
Inhibition of gap junction and adherens junction assembly by connexin and A-CAM antibodies.
J. Cell Biol
119
,
179
189
Miettinen
P.J.
,
Ebner
R.
,
Lopez
A. R.
,
Derynck
R.
(
1994
).
TGF-induced transdifferentiation of mammary epithelial cells to mesenchymal cells: involvement of type I receptors.
J. Cell Biol
127
,
2021
2036
Monaghan
P.
,
Perusinghe
N.
,
Carlile
G.
,
Evans
W. H.
(
1994
).
Rapid modulation of gap junction expression in mouse mammary gland during pregnancy, lactation, and involution.
J. Histochem. Cytochem
42
,
931
938
Mork
C.
,
van Deurs
B.
,
Petersen
O.
(
1990
).
Regulation of vimentin expression in cultured human mammary epithelial cells.
Differentiation
43
,
146
156
Musil
L. S.
,
Cunningham
B. A.
,
Edelman
G. M.
,
Goodenough
D. A.
(
1990
).
Differential phosphorylation of the gap junction protein connexin43 in junctional communication-competent and-deficient cell lines.
J. Cell Biol
111
,
2077
2088
Nose
A.
,
Nagafuchi
A.
,
Takeichi
M.
(
1988
).
Expressed recombinant cadherins mediate cell sorting in model systems.
Cell
54
,
993
1001
Pitelka
D. R.
,
Hamamoto
S. T.
,
Duafala
J. G.
,
Nemanic
M. K.
(
1973
).
Cell contacts in the mouse mammary gland. I. Normal gland in postnatal development and the secretory cycle.
J. Cell Biol
56
,
797
818
Pitts
J. D.
,
Burk
R. R.
(
1976
).
Specificity of junctional communication between animal cells.
Nature
264
,
762
764
Pitts
J. D.
,
Kam
E.
(
1985
).
Communication compartments in mixed cell cultures.
Exp. Cell Res
156
,
439
449
Pozzi
A.
,
Risek
B.
,
Kiang
D. T.
,
Gilula
N. B.
,
Kumar
N. M.
(
1995
).
Analysis of multiple gap junction gene products in the rodent and human mammary gland.
Exp. Cell Res
220
,
212
219
Rae
J. L.
,
Kuszak
J. R.
(
1983
).
The electrical coupling of epithelium and fibers in the frog lens.
Exp. Eye Res
36
,
317
326
Ronnov-Jessen
L.
,
Petersen
O. W.
,
Koteliansky
V. E.
,
Bissell
M. J.
(
1995
).
The origin of the myofibroblasts in breast cancer.
J. Clin. Invest
95
,
859
873
Rook
M. B.
,
Jongsma
H. J.
,
de Jonge
B.
(
1989
).
Single channel currents of homo-and heterologous gap junctions between cardiac fibroblasts and myocytes.
Pflugers Arch
414
,
95
98
Sakakura
T.
(
1991
).
New aspects of stroma-parenchyma relations in mammary gland differentiation.
Int. Rev. Cytol
125
,
165
202
Simon
A. M.
,
Goodenough
D. A.
,
Li
E.
,
Paul
D. L.
(
1997
).
Female infertility in mice lacking connexin 37.
Nature
385
,
525
529
Takeichi
M.
(
1991
).
Cadherin cell adhesion receptors as a morphogenetic regulator.
Science
251
,
1451
1455
Tomasetto
C.
,
Neveu
M. J.
,
Daley
J.
,
Horan
P. K.
,
Sager
R.
(
1993
).
Specificity of gap junction communication among human mammary cells and connexin transfectants in culture.
J. Cell. Biol
122
,
157
167
Valdimarsson
G.
,
De-Sousa
P. A.
,
Kidder
G. M.
(
1993
).
Coexpression of gap junction proteins in the cumulus-oocyte complex.
Mol. Reprod. Dev
36
,
7
15
Wang
Y.
,
Rose
B.
(
1997
).
An inhibition of gap-junctional communication by cadherins.
J. Cell Sci
110
,
301
309
Woodward
T. L.
,
Akers
R. M.
,
Turner
J. D.
(
1994
).
Lack of mitogenic response to EGF, pituitary and ovarian hormones in bovine mammary epithelial cells.
Endocrine
2
,
529
535
Woodward
T. L.
,
DuMont
N.
,
O'Connor-McCourt
M.
,
Turner
J. D.
,
Philip
A.
(
1995
).
Characterization of transforming growth factor beta, growth regulatory effects and receptors on bovine mammary cells.
J. Cell. Phys
165
,
339
348
This content is only available via PDF.