Role of fibronectin deposition in cystogenesis of Madin-Darby canine kidney cells.
BACKGROUND
Madin-Darby canine kidney (MDCK) cells cultured within collagen I gel exhibit clonal growth and form spherical multicellular cysts. The cyst-lining epithelial cells are polarized with the basolateral surface in contact with the collagen gel and the apical surface facing the lumen. To understand whether MDCK cysts construct the basal lamina, we characterized the composition of the extracellular matrix deposited by MDCK cysts. The cyst-lining cells produced an apparently incomplete basal lamina containing a discontinuous laminin substratum. In addition, the basal cell surface of the cyst was surrounded by a thick layer of fibronectin. This study was conducted to delineate the role of fibronectin deposition in cystogenesis.


METHODS
MDCK cells cultured in collagen gel were employed. We first used Arg-Gly-Asp (RGD) peptides containing disintegrin rhodostomin to disturb the interaction between fibronectin and the cell surface integrin. We then established several stable transfectants expressing the fibronectin antisense RNA and with which to directly examine the role of fibronectin in cystogenesis.


RESULTS
Rhodostomin markedly decreased the growth rates of the MDCK cyst, suggesting the importance of a normal interaction between fibronectin and integrins. The stable transfectants overexpressing the fibronectin antisense RNA exhibited relatively lower levels of fibronectin and markedly lower cyst growth rates than the control clone. The lower growth rate was correlated with an increase in collagen gel-induced apoptosis.


CONCLUSIONS
The results indicate that the deposition of fibronectin underlying the cyst-lining epithelium serves to prevent apoptosis induced by three-dimensional collagen gel cultures, and hence facilitates cyst growth of MDCK cells.
Department of Physiology, National Cheng Kung University Medical College, Tainan, Taiwan.
S T
ST
Jiang
H C
HC
Chiang
M H
MH
Cheng
T P
TP
Yang
W J
WJ
Chuang
M J
MJ
Tang
D000818:Animals; D001483:Base Sequence / Q000235:genetics; D002460:Cell Line; D003560:Cysts / Q000209:etiology* / Q000473:pathology; D004285:Dogs; D005353:Fibronectins / Q000235:genetics / Q000378:metabolism*; D007674:Kidney Diseases / Q000209:etiology* / Q000473:pathology; D008969:Molecular Sequence Data; D016376:Oligonucleotides, Antisense / Q000235:genetics / Q000378:metabolism; D010455:Peptides / Q000494:pharmacology; D012313:RNA / Q000378:metabolism; D014162:Transfection
D005353:Fibronectins; D016376:Oligonucleotides, Antisense; D010455:Peptides; C072026:rhodostomin; D012313:RNA