By. Williams et A. Schonbrunn, BOMBESIN RECEPTORS IN A HUMAN DUODENAL TUMOR-CELL LINE - BINDING-PROPERTIES AND FUNCTION, Cancer research, 54(3), 1994, pp. 818-824
The bombesin family of peptides elicit numerous biological responses i
n the gut, including stimulation of cell proliferation. and have been
implicated as growth factors in a variety of gastrointestinal tumors.
Even though these peptides and their receptors are distributed through
out the gastrointestinal tract, there are few cell lines available as
model systems to study bombesin action in gastrointestinal cells. In t
his study, we have characterized functional bombesin receptors in a hu
man duodenal cancer cell line, HuTu-80. The binding of [I-125-Tyr4]bom
besin to intact cells at 4-degrees-C reached equilibrium by 6 h. Scatc
hard analysis of [I-125-Tyr4]bombesin binding showed that HuTu-80 cell
s contained a single class of high affinity binding sites (5900 +/- 19
60/cell; K(d) = 80 +/- 20 pM). [I-125-Tyr4]bombesin binding was inhibi
ted by bombesin receptor agonists and antagonists with the following o
rder of potencies: gastrin-releasing peptide (GRP) = GRP(14-27) = bomb
esin > [DPhe6]bombesin(6-13)ethylamide > [Leu13psi-CH2NH)Leu 14]bombes
in > neuromedin B. Photoaffinity cross-linking studies, in which N-5-a
zido-2-nitrobenzoyloxysuccinimide was used to covalently couple [I-125
]GRP(14-27) to cells at 4-degrees-C, resulted in the specific labeling
of a broad band with an apparent molecular mass of 66,000 daltons. Co
nsistent with the presence of high affinity receptors, bombesin increa
sed the formation of inositol phosphates in HuTu-80 cells in a dose-de
pendent manner (concentration eliciting half-maximal effect, 290 +/- 7
0 pm). However, under conditions where both insulin and serum increase
d [H-3]thymidine incorporation into DNA, 10 nM bombesin had no effect
either alone or in the presence of insulin. Bombesin also had no effec
t on colony formation by HuTu-80 cells in soft agar. Furthermore, the
bombesin receptor antagonist, [Leu13psi(CH2NH)Leu14]bombesin, did not
inhibit [H-3]thymidine incorporation or clonal growth either in the ab
sence or in the presence of serum. Together, these results show that H
uTu-80 cells contain high affinity bombesin receptors of the GRP subty
pe. These receptors are functionally coupled to second messenger produ
ction but do not stimulate cell proliferation.