1 Division of Gastroenterology, We investigated
prostanoid biogenesis in human colonic fibroblasts (CCD-18Co and 5 primary fibroblast cultures) and epithelial cell lines (NCM460, T84,
HT-29, and LS 174T) and the effect of PGE2 on fibroblast morphology.
Cytokine-stimulated PGE2
production was measured. PGH synthase-1 and -2 (PGHS-1 and -2) protein
and mRNA expression were evaluated. Basal
PGE2 levels were low in all cell
types (0.15-6.47 ng/mg protein). Treatment for 24 h with interleukin-1
colorectal adenocarcinoma; primary fibroblast cultures; colon
cancer cell lines; prostaglandin H synthases
ASPIRIN AND OTHER nonsteroidal anti-inflammatory drugs
(NSAIDs) inhibit the cyclooxygenase enzymes, PGH synthase-1 and -2 (PGHS-1 and -2; EC 1.14.99) (33).
PGE2 and other prostanoids are
generated through activity of these bifunctional enzymes, which are
usually rate limiting (14). Clinical (9) and experimental (27) evidence
of inhibition of colonic neoplastic progression by NSAIDs is consistent
with the epidemiological evidence that aspirin can reduce colorectal
cancer mortality (10).
Although PGHS-1 is constitutively expressed in a wide range of tissues
and there is little modulation in the levels of expression, PGHS-2 is
highly inducible by cytokines and other participants in inflammatory
and neoplastic processes (16). PGHS-2 mRNA and protein expression are
increased in colorectal adenocarcinomas compared with levels in
adjacent normal tissue (17). A causal role for PGHS-2 in murine
colorectal carcinogenesis was demonstrated experimentally in vivo, and
the biological effects of PGHS-2 upregulation were mediated
predominantly through increased
PGE2 production (24). However, the
cells from which prostanoids emanate in normal or neoplastic colorectal
tissue have not been defined.
DuBois et al. (7) predicted that transformed epithelial cells will
prove to be a major site of colorectal PGHS-2 expression and PG
synthesis. In support of this hypothesis, immunohistological localization of PGHS-2 in colorectal adenocarcinoma tissue sections was
most intense in the carcinoma cells themselves (28), and PGE2 synthesis is detectable in
some, but not all, colon cancer cell lines (15). Suggestive of an
alternative source, PGHS-2 was expressed essentially in interstitial
cells rather than in the intestinal epithelium of mice from a knockout
strain that develops numerous intestinal adenomas due to a truncation
mutation of the Apc gene (25).
Uncertainty about the source of prostanoids in benign or malignant
colorectal tissue prompted us to consider the synthetic capacity of
resident cell types other than epithelial cells. New concepts
concerning the functions of fibroblasts have been defined recently
(30). Fibroblasts express many receptors for cytokines, growth factors,
and hormones (8). Those from a single tissue do not constitute a
homogeneous phenotypic or functional population, and fibroblasts from
different tissues have characteristic phenotypes. For example, cultured
orbital fibroblasts were shown to differ metabolically from dermal
fibroblasts with regard to extracellular matrix production and
morphological changes elicited by
PGE2 (31).
Fibroblasts, regardless of their tissue of origin, express PGHS-1 at
relatively high levels, accounting for basal
PGE2 in these cells (30, 34). In
contrast, the tissue of origin is a crucial determinant of fibroblast
PGHS-2 expression induced by cytokines and other agents. Pulmonary,
synovial, and orbital fibroblasts express highly inducible PGHS-2, but
induction in dermal fibroblasts is much less robust. Colonic
fibroblasts have not previously been examined for this property.
Intestinal fibroblasts were shown to synthesize and release somatomedin
(insulin-like growth factor) that functioned by paracrine pathways to
influence the growth and differentiation of a number of adjacent cell
types (12). In a further example of the importance of
mesenchymal-epithelial interactions, fibroblasts induced organization and differentiation of colonic epithelial (T84) cells in vitro (11).
However, little is known of the colonic fibroblast and its potential
contribution to diseases such as colorectal cancer and inflammatory
bowel disease (IBD).
In the present study, we investigated PGHS-2 mRNA and protein induction
by cytokines in cultured human colonic fibroblasts. We report that
interleukin-1 Reagents.
IL-1 Cell culture.
CCD-18Co, a human colonic fibroblast strain derived from normal tissue,
and HT-29, LS 174T, and T84 colon cancer cell lines were obtained from
the American Type Culture Collection (Manassas, VA). NCM460, an
immortalized line established from normal human colonic epithelium, was
a generous gift of Dr. M. P. Moyer (University of Texas, San Antonio,
TX) (23).
ABSTRACT
Top
Abstract
Introduction
Materials & Methods
Results
Discussion
References
(IL-1
; 10 ng/ml) or tumor necrosis factor-
(50 ng/ml), respectively, elicited maximal 25- and 6-fold inductions of
PGE2 synthesis in CCD-18Co
cultures and similar results in primary fibroblast cultures; maximal
inductions with IL-1
in colonic epithelial cell lines were from zero
to fivefold. Treatment of CCD-18Co fibroblasts with IL-1
caused
maximal 21- and 53-fold increases, respectively, in PGHS-2 protein and
mRNA levels without altering PGHS-1 expression.
PGE2 (0.1 µmol/l) elicited a
dramatic shape change in selected fibroblasts. Colonic fibroblasts are potentially important as cytokine targets and a source of and target
for colonic prostanoids in vivo.
INTRODUCTION
Top
Abstract
Introduction
Materials & Methods
Results
Discussion
References
(IL-1
) in particular can induce dramatic PGHS-2
expression, with a consequent 25-fold increase in
PGE2 production. The response of
PGHS-2 and PGE2 synthesis to
IL-1
in colonic fibroblasts was considerably more robust than that
observed in neoplastic and nonneoplastic colonic epithelial cells.
These findings indicate the potentially prominent role of fibroblasts
in prostanoid synthesis in the colon and define another cellular target
for the beneficial actions of NSAIDs in colorectal neoplasia.
MATERIALS AND METHODS
Top
Abstract
Introduction
Materials & Methods
Results
Discussion
References
, Klenow enzyme, random primer, and oligonucleotides were
purchased from Boehringer Mannheim (Indianapolis, IN). Tumor necrosis
factor-
(TNF-
), dexamethasone, and indomethacin were from Sigma
(St. Louis, MO). SC-58125 was kindly provided by Dr. P. Isakson of
Searle (Skokie, IL). Cesium chloride and guanidinium isothiocyanate
were purchased from Bethesda Research Laboratories (Bethesda, MD).
DMEM, fetal bovine serum (FBS),
L-glutamine, sodium pyruvate,
penicillin, D-glucose, HEPES,
streptomycin, amphotericin, and nonessential amino acids were purchased
from GIBCO Laboratories (Grand Island, NY). Kits for measurement of
PGE2 levels by RIA were purchased
from Amersham (Arlington Heights, IL).
PGE2 assay.
Fibroblasts were seeded at a density of 5 × 105
cells/cm2 and grown to confluence
in 24-well plastic culture plates in medium supplemented with 10% FBS.
It is well recognized that serum is a potent inducer of PGHS-2 and thus
PGE2 synthesis (18). Therefore, confluent cultured fibroblasts were shifted from medium supplemented with 10% FBS to medium containing 1% FBS for the 48 h before
harvesting that included the period of treatment with test compounds.
To determine the effects of cytokines, cultures were incubated without addition of a cytokine or in medium supplemented with IL-1 (10 ng/ml) or TNF-
(50 ng/ml). The following compounds, added at the
same time as IL-1
, were used in some experiments at the final concentrations indicated: dexamethasone (10 nmol/l), indomethacin (10 µmol/l) and SC-58125 (5 µmol/l).
Western analysis of PGHS-2 protein expression. Relative levels of the cyclooxygenase proteins were determined as described (34). Briefly, confluent cultured fibroblasts were shifted from medium supplemented with 10% FBS to medium containing 1% FBS for a total of 48 h during treatment with test compounds and then harvested. Lysates were subjected to gel electrophoresis, and the separated proteins were transferred to a membrane. Membranes were incubated with primary monoclonal antibodies specific for PGHS-2, generously provided by Dr. J. Maclouf (IFR Circulation-Lariboisiere, INSERM 348, Paris, France) or purchased from Cayman (Ann Arbor, MI). Bound antibodies were detected by chemiluminescence using ECL reagents purchased from Amersham. The resulting bands were analyzed densitometrically.
Isolation of fibroblast RNA and Northern analysis. Isolation of total cellular RNA, gel electrophoresis, Northern blotting, hybridization with cDNA probes, and quantitation of mRNA levels were performed as described (29). Fibroblasts were cultivated to confluence in 100-mm-diameter plastic culture plates, treated with test compounds, and harvested for isolation of total RNA. The human PGHS-1 and -2 cDNAs utilized for hybridizations were generously supplied by Drs. D. Young (University of Rochester, Rochester, NY) and T. Hla (American Red Cross, Washington, DC). The densities of the DNA-RNA hybrids were determined by scanning the resulting radiographs, and results were normalized for intensity of staining with ethidium bromide.
Effects of PGE2 on fibroblast morphology. Fibroblast cultures were grown to confluence in 24-well plates in medium supplemented with 10% FBS. Confluent cultures were shifted to medium containing 1% FBS 24 h before addition of PGE2 at a final concentration of 0.1 µmol/l. Phase-contrast analysis of cellular morphology was performed using a Zeiss Axiovert microscope (Carl Zeiss, Thornwood, NY) equipped with a 35-mm camera. Photographs were taken on Kodak Technical Pan film (Eastman Kodak, Rochester, NY) at a final magnification of ×200.
![]() |
RESULTS |
---|
![]() ![]() ![]() ![]() ![]() ![]() ![]() |
---|
Treatment of human colonic fibroblast strains with
IL-1 or TNF-
results in substantial
increases in PGE2 synthesis.
IL-1 plays a widespread and potent role in mediating immune and
inflammatory responses (21) and is produced by a wide variety of cells,
including fibroblasts (5). The stimulatory effects of proinflammatory
cytokines, such as IL-1
and TNF-
, on
PGE2 synthesis in various cell
types, including fibroblasts, are well documented (4, 13, 34). We
examined PGE2 synthesis in the CCD-18Co human colonic fibroblast strain (Fig.
1). The mean basal PGE2 level from seven separate
experiments was 1.40 ± 0.27 ng/mg protein. In an additional
experiment to characterize PGE2
synthesis in unstimulated cells, levels at 8 and 24 h, respectively,
were 2.4 and 2.3 ng/mg protein. The
PGE2 level increased 25-fold in response to IL-1
at the physiologically relevant concentration of 10 ng/ml (5). Increased PGE2
synthesis was evident at 2 h and peaked at 24 h. TNF-
(50 ng/ml)
elicited an almost sixfold induction in
PGE2 levels (data not shown).
|
|
Comparison of effects of IL-1 on
PGE2 synthesis in epithelial cell lines and
fibroblasts.
PGE2 synthesis in colonic
fibroblasts (CCD-18Co) in response to IL-1
was compared with the
levels in colonic nonneoplastic (NCM460) and malignant (HT-29, LS 174T,
and T84) epithelial cell lines and an orbital fibroblast strain (Fig.
3). Basal levels of
PGE2 in the epithelial cell lines
were 0.15-1.3 ng/mg protein. The basal level in the orbital
fibroblast strain was 0.64 ± 0.05 ng/mg protein.
|
IL-1 increases PGHS-2 protein levels in normal
colonic fibroblasts.
Western analysis was performed to determine whether exposure of colonic
fibroblasts to IL-1
led to increased expression of PGHS-2 protein
(Fig. 4). PGHS-2 protein was almost
undetectable in untreated cells. A maximal 21-fold increase in PGHS-2
enzyme level was seen after exposure to IL-1
(10 ng/ml) for 16 h.
PGHS-1 protein levels, in contrast, were unaffected by cytokine
treatment (data not shown).
|
Treatment with IL-1 or TNF-
increases PGHS-2 mRNA levels in normal colonic fibroblasts.
To characterize further the cyclooxygenase pathway and its regulation
in colonic fibroblasts, Northern blots were performed to analyze basal
expression and upregulation of PGHS-1 and PGHS-2 mRNA in response to
IL-1
(Fig. 5) in CCD-18Co cells. With
IL-1
(10 ng/ml), PGHS-2 mRNA expression, barely detectable in
cellular mRNA from untreated CCD-18Co fibroblasts, increased 10-fold at 2 h (data not shown) and reached a level at least 50-fold greater than
baseline after exposure to IL-1
for 8 h. The level of induction remained 48-fold at 24 h. In contrast, PGHS-1 mRNA levels were unaffected by IL-1
treatment. TNF-
(50 ng/ml) produced a 15-fold induction of the PGHS-2 mRNA level at 18 h (data not shown).
|
PGE2 elicits a shape change in colonic fibroblasts. We reported previously that human orbital fibroblasts undergo a morphological change when incubated with PGE2 (32). Figure 6 shows typical microscopic fields of untreated CCD-18Co colonic fibroblasts and fibroblasts that were incubated with PGE2 (0.1 µmol/l) for 4 h. As with orbital fibroblasts, when the culture medium of colonic fibroblasts was supplemented with PGE2, some cells underwent a profound change in appearance; the central nucleus-containing region was condensed, and prominent spidery cytoplasmic processes developed. Other cells were unaffected. Increasing the concentration to 10 µmol/l did not appear to further increase the shape-changing properties of PGE2.
|
![]() |
DISCUSSION |
---|
![]() ![]() ![]() ![]() ![]() ![]() ![]() |
---|
In conjunction with experimental evidence of a causal relationship between increased PGHS-2 expression and colorectal carcinogenesis, immunohistological localization of the enzyme was reported in the interstitial but not the epithelial cells of murine colonic adenomas (24). We have demonstrated that, fully consistent with the murine immunohistological data, human colonic fibroblasts not only are highly sensitive and more responsive to proinflammatory cytokine-mediated induction of PGHS-2 expression than are epithelial cell lines under identical experimental conditions in vitro but are capable of much higher levels of PGE2 production.
Our findings indicate that colonic fibroblasts share with pulmonary, synovial, and orbital fibroblasts the property of expressing highly inducible PGHS-2. Further, we report that colonic fibroblasts, like orbital fibroblasts, are able to undergo dramatic morphological changes in vitro in response to concentrations of PGE2 that have physiological relevance. Whether these responses to PGE2 are relevant to the normal function of colonic tissue or to any disease state remains to be determined.
Clearly, establishment of cultures in vitro could have selected and accentuated cellular behavior patterns that may not accurately reflect the patterns of fibroblast gene expression in vivo. Nonetheless, five of six fibroblast strains utilized in this study (the American Type Culture Collection strain and four of five primary fibroblast cultures established in our laboratory) were initiated from histologically normal epithelium. Colorectal adenocarcinomas are clonal in origin and develop through accumulated malfunction of multiple genes. The capacity for PGHS-2 expression exhibited by the normal colonic fibroblast strains on which we report suggests that clones of neoplastic epithelial cells in the earliest stages of histological development (aberrant crypt foci and early adenomas) could be exposed to prostanoid products from adjacent cells, especially those found in the interstitium, with consequent potential for enhancement of neoplastic progression. Consistent with the potentially important influence of fibroblasts and other interstitial components in colorectal carcinogenesis, intestinal microadenomas in mice carrying a truncated Apc gene originated from the antiluminal aspect of crypt epithelium as outpockets growing into the intravillous space that is populated by fibroblasts and other interstitial cells (25).
PGHS-2 expression, as reflected by PGE2 synthesis, was responsive to cytokines in all the colonic fibroblast strains examined, but the degree of inducibility varied considerably. Although increased PG levels have been widely reported in colon cancer specimens, the stage of neoplastic transformation at which increased PGE2 is first apparent is debated. Oshima et al. (24) did not detect PGHS-2 protein in polyps <2 mm, but Boolbol et al. (2) reported increased levels of the enzyme in histologically normal colonic epithelium from Min mice compared with normal littermates lacking the Apc mutation.
To reiterate, five of the six fibroblast strains that were examined in this study were from normal mucosa. It will be necessary to study fibroblasts from multiple additional subjects without colonic disease and from patients with adenomas, adenocarcinomas, and other colonic diseases to define the range of colonic fibroblast responsiveness to cytokine stimulation and the temporal relationship of increased PG synthesis to the stage of colon carcinogenesis. It may be that the distinct phenotypic attributes of an individual's colonic fibroblasts, such as relative sensitivity to cytokine stimulation, can have a protective role or contribute to the pathogenesis of large bowel disease.
Knowledge of the behavior of colonic fibroblasts in patients with IBD is limited. Although intestinal mucosal PG levels are elevated in IBD (20), indomethacin and other cyclooxygenase inhibitors are clinically ineffective in these diseases. In the absence of evidence that prostaglandins are important mediators of inflammation in IBD, interest has focused on arachidonic acid metabolites of the 5-lipoxygenase pathway. Leukotriene B4, in particular, has been incriminated as a potential inflammatory mediator in the intestine, but elucidation of the pathophysiological mechanisms of IBD is far from complete. In light of their profound responsiveness to proinflammatory cytokines, further investigations of colonic fibroblasts from patients with IBD may provide useful new insights.
The finding that fibroblast PGHS-1 mRNA and protein levels were
unaffected by any of the cytokine treatments is consistent with the
view that this cyclooxygenase is constitutively expressed in a wide
variety of tissues and not necessarily involved in the inflammatory
response (14). However, evidence from experiments utilizing knockout
mice with disruptions of PGHS-1 or -2 gene expression raises the
possibility that PGHS-2 expression and inducibility alone may not
entirely account for prostanoid-dependent inflammatory responses (19);
PGHS-1 expression may also be involved. This possibility is supported
by evidence for differential regulation of PGHS-1 expression by
cytokines, including IL-1 and TNF-
, through alternative mRNA
splicing (6).
It is increasingly evident that fibroblasts and their products are crucial determinants of epithelial cell behavior. We suggest that fibroblasts could be important cytokine targets in the colon. Furthermore, they may both represent a potentially important site of colonic prostanoid biogenesis in vivo and be targets for PGE2.
![]() |
ACKNOWLEDGEMENTS |
---|
We thank H. Hameer for immunohistochemical characterization of primary fibroblast cultures.
![]() |
FOOTNOTES |
---|
This work was supported by National Institute of Diabetes and Digestive and Kidney Diseases Grant DK-43649, Veterans Affairs Central Office Merit Review, and a grant from the Margaret Duffy and Robert Cameron Troup Memorial Fund for Cancer Research of the Buffalo General Hospital (to P. Lance) and by National Eye Institute Grants RO1-EY-08976 and RO1-EY-11708 and Veterans Affairs Central Office Merit Review (to T. J. Smith).
A portion of this work was presented at the annual meeting of the American Gastroenterological Association in May 1997 and published in abstract form (17a).
Address for reprint requests: P. Lance, Division of Gastroenterology, Buffalo General Hospital, 100 High St., Buffalo, NY 14203.
Received 5 December 1997; accepted in final form 10 June 1998.
![]() |
REFERENCES |
---|
![]() ![]() ![]() ![]() ![]() ![]() ![]() |
---|
1.
Angel, J.,
F. Berenbaum,
C. Le Denmat,
T. Nevalainen,
J. Masliah,
and
C. Fournier.
Interleukin-1-induced prostaglandin E2 biosynthesis in human synovial cells involves the activation of cytosolic phospholipase A2 and cyclooxygenase-2.
Eur. J. Biochem.
222:
125-131,
1994.
2.
Boolbol, S. K.,
A. J. Dannenberg,
A. Chadburn,
C. Martucci,
X. Guo,
J. T. Ramonetti,
M. Abreu-Goris,
H. L. Newmark,
M. L. Lipkin,
J. J. DeCosse,
and
M. M. Bertagnolli.
Cyclooxygenase-2 overexpression and tumor formation are blocked by sulindac in a murine model of familial adenomatous polyposis.
Cancer Res.
56:
2556-2560,
1996[Abstract].
3.
Bradford, M.
A rapid and sensitive method for the quantitation of microgram quantities of protein utilizing the principle of protein-dye binding.
Anal. Biochem.
72:
248-254,
1976[Medline].
4.
Casey, M. L.,
K. Korte,
and
P. C. MacDonald.
Epidermal growth factor stimulation of prostaglandin E2 biosynthesis in amnion cells. Induction of prostaglandin H2 synthase.
J. Biol. Chem.
263:
7846-7854,
1988
5.
Crofford, L. J.,
R. L. Wilder,
A. P. Ristimäki,
H. Sano,
E. F. Remmers,
H. R. Epps,
and
T. Hla.
Cyclooxygenase-1 and -2 expression in rheumatoid synovial tissues.
J. Clin. Invest.
93:
1095-1101,
1994[Medline].
6.
Diaz, A.,
A. M. Reginato,
and
S. A. Jimenez.
Alternative splicing of human prostaglandin G/H synthase mRNA and evidence of differential regulation of the resulting transcripts by transforming growth factor 1, interleukin 1
, and tumor necrosis factor
.
J. Biol. Chem.
267:
10816-10822,
1993
7.
DuBois, R. N.,
A. Radhika,
B. S. Reddy,
and
A. J. Entingh.
Increased cyclooxygenase-2 levels in carcinogen-induced rat colonic tumors.
Gastroenterology
110:
1259-1262,
1996[Medline].
8.
Fries, K. M.,
T. Blieden,
R. J. Looney,
G. D. Sempowski,
M. R. Silvera,
R. A. Willis,
and
R. P. Phipps.
Evidence of fibroblast heterogeneity and the role of fibroblast subpopulations in fibrosis.
Clin. Immunol. Immunopathol.
72:
283-292,
1994[Medline].
9.
Giardiello, F. M.,
S. R. Hamilton,
A. J. Krush,
S. Piantadosi,
L. M. Hylind,
P. Celano,
S. V. Booker,
C. R. Robinson,
and
G. J. A. Offerhans.
Treatment of colonic and rectal adenomas with sulindac in familial adenomatous polyposis.
N. Engl. J. Med.
328:
1313-1316,
1993
10.
Giovannucci, E.,
E. B. Rimm,
M. J. Stampfer,
G. A. Colditz,
A. Ascherio,
and
W. C. Willett.
Aspirin use and the risk for colorectal cancer and adenoma in male health professionals.
Ann. Intern. Med.
121:
241-246,
1994
11.
Halttunen, T.,
A. Marttinen,
I. Rantala,
H. Kainulainen,
and
M. Maki.
Fibroblasts and transforming growth factor beta induce organization and differentiation of T84 human epithelial cells.
Gastroenterology
111:
1252-1256,
1996[Medline].
12.
Han, V. K.,
A. J. D'Ercole,
and
P. K. Lund.
Cellular localization of somatomedin (insulin-like growth factor) messenger RNA in the human fetus.
Science
236:
193-197,
1991.
13.
Herrmann, F.,
A. Lindemann,
J. Gauss,
and
R. Mertelsmann.
Cytokine-stimulation of prostaglandin synthesis from endogenous and exogenous arachidonic acids in polymorphonuclear leukocytes involving activation and new synthesis of cyclooxygenase.
Eur. J. Immunol.
20:
2513-2516,
1990[Medline].
14.
Herschman, H. R.
Regulation of prostaglandin synthase-1 and prostaglandin synthase-2.
Cancer Metastasis Rev.
13:
241-256,
1994[Medline].
15.
Hubbard, W. C.,
M. C. Alley,
T. L. McLemore,
and
M. R. Boyd.
Profiles of prostaglandin biosynthesis in sixteen established cell lines derived from human lung, colon, prostate, and ovarian tumors.
Cancer Res.
48:
4770-4775,
1988[Abstract].
16.
Jones, D. A.,
D. P. Carlton,
T. M. McIntyre,
G. A. Zimmerman,
and
S. M. Prescott.
Molecular cloning of human prostaglandin endoperoxide synthase II and demonstration of expression in response to cytokines.
J. Biol. Chem.
268:
9049-9054,
1993
17.
Kargman, S. L.,
G. P. O'Neill,
P. J. Vickers,
J. F. Evans,
J. A. Mancini,
and
S. Jothy.
Expression of prostaglandin G/H synthase-1 and -2 protein in human colon cancer.
Cancer Res.
55:
2556-2559,
1995[Abstract].
17a.
Kim, E. C.,
V. Andersen,
D. Sciaky,
H. J. Cao,
T. J. Smith,
and
P. Lance.
Cytokine-mediated cyclooxygenase and prostaglandin expression by colonic fibroblasts (Abstract).
Gastroenterology
112:
A591,
1997.
18.
Kujubu, D. A.,
S. T. Reddy,
B. S. Fletcher,
and
H. R. Herschman.
Expression of the protein product of the prostaglandin synthase-2/TS10 gene in mitogen-stimulated Swiss 3T3 cells.
J. Biol. Chem.
268:
5425-5430,
1993
19.
Langenbach, R.,
S. G. Morham,
H. F. Tiano,
C. D. Loftin,
B. I. Ghanayem,
P. C. Chulada,
J. F. Mahler,
C. A. Lee,
E. H. Goulding,
K. D. Kluckman,
H. S. Kim,
and
O. Smithies.
Prostaglandin synthase 1 gene disruption in mice reduces arachidonic acid-induced inflammation and indomethacin-induced gastric ulceration.
Cell
83:
483-492,
1995[Medline].
20.
Lauritsen, K.,
L. S. Laursen,
K. Bukhave,
and
J. Rask-Madsen.
In vivo profiles of eicosanoids in ulcerative colitis, Crohn's colitis, and Clostridium difficile colitis.
Gastroenterology
95:
11-12,
1988[Medline].
21.
Leung, K.,
J. C. Betts,
L. Xu,
and
G. J. Nabel.
The cytoplasmic domain of the interleukin-1 receptor is required for nuclear factor-B signal transduction.
J. Biol. Chem.
1994:
1579-1582,
1994.
22.
Meade, E. A.,
W. L. Smith,
and
D. L. DeWitt.
Differential inhibition of prostaglandin endoperoxide synthase (cyclooxygenase) isozymes by aspirin and other non-steroidal anti-inflammatory drugs.
J. Biol. Chem.
268:
6610-6614,
1993
23.
Moyer, M. P.,
L. A. Manzano,
R. L. Merriman,
J. S. Stauffer,
and
L. R. Tanzer.
NCM460, a normal human colon mucosal epithelial cell line.
In Vitro Cell. Dev. Biol.
32:
315-317,
1996.
24.
Oshima, M.,
J. E. Dinchuk,
S. L. Kargman,
H. Oshima,
B. Hancock,
E. Kwong,
J. M. Trzaskos,
J. F. Evans,
and
M. M. Taketo.
Suppression of intestinal polyposis in Apc716 knockout mice by inhibition of cyclooxygenase 2 (COX-2).
Cell
87:
803-809,
1996[Medline].
25.
Oshima, M.,
H. Oshima,
K. Kitagawa,
M. Kobayashi,
C. Itakura,
and
M. Taketo.
Loss of Apc heterozygosity and abnormal tissue building in nascent intestinal polyps in mice carrying a truncated Apc gene.
Proc. Natl. Acad. Sci. USA
92:
4482-4486,
1995[Abstract].
26.
Raz, A.,
A. Wyche,
and
P. Needleman.
Temporal and pharmacological division of fibroblast cyclooxygenase expression into transcriptional and translational phases.
Proc. Natl. Acad. Sci. USA
86:
1657-1661,
1989[Abstract].
27.
Reddy, B. S.,
J. Nayini,
K. Tokumo,
J. Rigotty,
E. Zang,
and
G. Kelloff.
Chemoprevention of colon carcinogenesis by concurrent administration of piroxicam, a nonsteroidal antiinflammatory drug with D,L-alpha-difluoromethylornithine, an ornithine decarboxylase inhibitor, in diet.
Cancer Res.
50:
2562-2568,
1990[Abstract].
28.
Sano, H.,
Y. Kawahito,
R. L. Wilder,
A. Hashiramoto,
S. Mukai,
K. Asai,
S. Kimura,
H. Kato,
M. Kondo,
and
T. Hla.
Expression of cyclooxygenase-1 and -2 in human colorectal cancer.
Cancer Res.
55:
3785-3789,
1995[Abstract].
29.
Shah, S.,
P. Lance,
T. J. Smith,
C. S. Berenson,
S. A. Cohen,
P. J. Horvath,
J. T. Y. Lau,
and
H. Baumann.
n-Butyrate reduces the expression of beta-galactoside alpha-2,6-sialyltransferase in Hep G2 cells.
J. Biol. Chem.
267:
10652-10658,
1992
30.
Smith, R. S.,
T. J. Smith,
T. M. Blieden,
and
R. P. Phipps.
Fibroblasts as sentinel cells: synthesis of chemokines and regulation of inflammation.
Am. J. Pathol.
151:
317-322,
1997[Abstract].
31.
Smith, T. J.,
R. S. Bahn,
and
C. A. Gorman.
Hormonal regulation of hyaluronate synthesis in cultured human fibroblasts: evidence for differences between retrooccular and dermal fibroblasts.
J. Clin. Endocrinol. Metab.
69:
1019-1023,
1989[Abstract].
32.
Smith, T. J.,
H. S. Wang,
M. G. Hogg,
R. C. Henrikson,
C. R. Keese,
and
I. Giaever.
Prostaglandin E2 elicits a morphological change in cultured orbital fibroblasts from patients with Graves opthalmopathy.
Proc. Natl. Acad. Sci. USA
91:
5094-5098,
1994[Abstract].
33.
Smith, W. L.,
R. M. Garavito,
and
D. L. DeWitt.
Prostaglandin endoperoxide H synthases (cyclooxygenases)-1 and -2.
J. Biol. Chem.
271:
33157-33160,
1996
34.
Wang, H. S.,
H. J. Cao,
V. D. Winn,
L. J. Rezanka,
Y. Frobert,
C. H. Evans,
D. Sciaky,
D. A. Young,
and
T. J. Smith.
Leukoregulin induction of prostaglandin-endoperoxide H synthase-2 in human orbital fibroblasts.
J. Biol. Chem.
271:
22718-22728,
1996