Publication: Morphogenesis of normal human salivary gland cells in vitro
Authors
Azuma, M. ; Sato, M.
item.page.secondaryauthor
item.page.director
Publisher
Murcia : F. Hernández
publication.page.editor
publication.page.department
DOI
item.page.type
info:eu-repo/semantics/article
Description
Abstract
Primary cultured human salivary gland cells
were transfected with ori-defective mutant DNA of
SV40. Following 2-3 weeks of transfection, slowly
expanding colonies consisting of small compact cells
emerged, while mock-transfected cells did not grow any
more and eventually entered crisis, followed by cell
death. Using limited dilution technique, we isolated 4
cell clones with distinct morphology from a single
colony. Morphological observation of cells cultured on
plastic dishes precisely revealed the characteristics of the
constituent cells of salivary gland; i.e., three cell clones
showing cuboidal- (NS-SV-DC), spindle- (NS-SV-MC),
and flattened morphology (NS-SV-SC) were similar to
duct-, myoepithelial-, and squamous phenotype,
respectively. A remaining cell clone, polygonal in shape
and with numerous secretory granules (NS-SV-AC),
resembled an acinar cell. Characterization of cell clones by ultrastructural
exarnination and search for specific antigens showed the
similarity of NS-SV-DC, NS-SV-MC, NS-SV-AC, and
NS-SV-SC to duct, myoepithelial, acinar, and squamous
cells, respectively. Anchorage-independent growth in
soft agar and tumorigenicity in nude mice were not
recognized in al1 cell clones. These results demonstrate
that establishment of cell clones with duct-, myoepithelial-,
acinar-, or squamous phenotype was
accomplished in the in vitro system, and that based on the
evaluation of colony-forming ability in soft agar and
tumorigenicity in nude mice these cell clones can be
considered to be non-tumorigenic. Using the above in
vitro system, we examined the effect of a reconstituted
basement membrane extract, Matrigel, on the morphogenesis
of cultured normal human salivary gland cells.
When NS-SV-DC or NS-SV-MC were seeded on
Matrigel in serum-free culture conditions, they formed
round or zona1 clusters on day 1; failing however, to
develop into a salivary gland morphogenesis. Semithin
sections of cell clones cultured on Matrigel exhibited multicellular aggregates on day 1, while on days 2 and 3
these cells lost both cell-cell and cell-Matrigel
interactions and eventually entered crisis. In an attempt
to understand the mechanism involved in this
phenomenon, we have investigated proteolytic enzymes
and their inhibitors secreted by cell clones. Although cell
clones produced almost identical levels of gelatinases,
they released increased amounts of plasminogen
activators (PAs) as compared with a neoplastic human
salivary gland cell line (HSG), which had already been
demonstrated to differentiate into acinar cells when
cultured on Matrigel. Obvious difference of expression
leve1 of tissue inhibitor of metalloproteinases-1 (TIMP-
1) was not observed in these cells. However, secretion of
PA inhibitors was elevated in NS-SV-MC when
compared to NS-SV-DC. Neutralization of excess PAs
by exogenously-added serine protease inhibitors
corrected the aberrant morphogenesis of NS-SV-DC, but
not that of NS-SV-MC, and allowed NS-SV-DC to form
glandular-like structures. Thus, these results may suggest
that a tightly regulated proteolytic activity is, in part,
essential for in vitro morphogenesis of normal salivary
gland duct cells.
publication.page.subject
Citation
item.page.embargo
Ir a Estadísticas
Sin licencia Creative Commons.