| HOME | HELP | FEEDBACK | SUBSCRIPTIONS | ARCHIVE | SEARCH | TABLE OF CONTENTS |
Regular Articles |
From the Medical Service, Department of Veterans Affairs Medical Center, Long Beach, California; and the Department of Medicine, University of California, Irvine, California
 |
Abstract |
|---|
 Top Abstract IntroductionMaterials and MethodsResultsDiscussionReferences |
|---|
|
Introduction |
|---|
|
TopAbstractIntroductionMaterials and MethodsResultsDiscussionReferences |
|---|
(TGF-), have been shown to participate in the repair of tissue
injury by stimulating the cell proliferation and migration necessary
for reepithelialization and ulcer healing.4-7
Immunohistochemical studies have shown overexpression of EGF and its
receptor (EGF-R) in epithelial cells lining ulcer margins and
regenerating glands,8-10
indicating that these cells are
major targets for the proliferation-stimulating action of EGF. Our recent study demonstrated that gastric ulceration triggers increased receptor tyrosine kinase activity, EGF-R phosphorylation, and extracellular signal regulated kinase 1 and 2 (ERK1 and ERK2) activity in epithelial cells of the ulcer margins.11 Biochemical and genetic studies in various cell systems (other than gastric mucosa) have demonstrated that Raf-1 functions downstream of activated tyrosine kinases and Ras, but upstream of mitogen-activated protein kinase (MAPK) and MAPK kinase (MEK).12,13 Raf-1 activity can be modulated by both Ras-dependent and Ras-independent pathways.14 In addition to Ras, Raf-1 activators may include protein kinase C (PKC), activated tyrosine kinases, or as yet unidentified serine threonine kinases and phosphatases.14 The intermediate steps linking activated receptor tyrosine kinase to MAP kinases (ERK1 and ERK2) during gastric ulcer healing in vivo remain unknown.
In some cellular sytems other than gastric cells (eg, Rat1, A431, and
Her14 cells), EGF-R activation has been shown to cause binding of the
SH2-containing adapter protein (Shc) to growth factor receptor-bound
protein (Grb2), leading to recruitment of Son of sevenless (Sos) to the
plasma membrane and Ras activation.15-18
Phospholipase
C-
(PLC-) is another signaling protein that contains SH domains
and is activated by tyrosine kinase.19
PLC-1 can act
either as an enzyme that generates diacylglycerol and IP3,
leading to PKC activation,20,21
or as an adapter protein
by binding to activated growth factor receptors via its SH2 domains and
a downstream molecule via its SH3 domains.22
PKC can
activate MAPK, which in turn activates the gene transcription involved
in cell proliferation and differentiation.23-27
Growth
factor-mediated activation of the Raf-1–MAPK/ERK cascade may therefore
involve either Ras, PKC, or both.13,23-27
Previous
studies have shown increased tyrosine kinase activity and PLC-1
phosphorylation during the early phase of acute injury repair in rat
gastric mucosa.28,29
However, involvement of Ras or PKC in
signaling pathways during chronic gastric ulcer healing is not known.
The present study was aimed at determining whether during experimental gastric ulcer healing Raf-1 is activated and to identify potential activators of Raf-1 by examining the Ras and PKC protein levels, PKC activity, and Shc-Grb2, Grb2-Sos complex levels. To determine whether activation of the Raf-1–ERK cascade during gastric ulcer healing occurs predominantly in the epithelial component, we studied the effect of EGF on Ras activation, and PKC and ERK activities in an in vitro model using a rat gastric epithelial cell line (RGM1) derived from normal rat gastric mucosa.
|
Materials and Methods |
|---|
|
TopAbstractIntroductionMaterials and MethodsResultsDiscussionReferences |
|---|
, ß, and isoforms), anti-Ras, rabbit polyclonal anti-ERK2, and
anti-Sos antibodies were purchased from Santa Cruz Biotechnology (Santa
Cruz, CA). Rabbit polyclonal anti-Shc antibody, mouse monoclonal
anti-Grb2, and anti-c-Raf-1 antibodies were purchased from Transduction
Laboratories (Lexington, KY). A Protein kinase C Assay System was
purchased from Gibco BRL (Gaithersburg, MD). [-32P]ATP
was purchased from Dupont NEN Research Products (Boston, MA), and all
other molecular biology-grade chemicals were purchased from Sigma
Chemical Company (St. Louis, MO). Animals
This study was approved by the Subcommittee for Animal Studies of the VA Medical Center (Long Beach, CA). Male Sprague-Dawley rats (Crl:CD(SD)BR; Charles River Laboratories; Wilmington, MA) were fasted for 12 hours and underwent laparotomy under Nembutal anesthesia (60 mg/kg body weight). One hundred percent acetic acid (50 µl) was applied to the serosa of lower gastric corpus at the posterior wall through a polyethylene tube (4.0 mm i.d.) for 90 seconds. The serosal area was then washed with isotonic saline and the abdomen closed. Sham-operated rats underwent a similar procedure without acetic acid administration. Rats with gastric ulcers (n = 54) and sham-operated rats (n = 36) were euthanized 3 and 7 days after operation. Ulcer margins were carefully dissected from granulation tissue, snap-frozen in liquid nitrogen, and stored at -80°C.
Raf-1 Kinase Assay
Raf-1 kinase activity was determined using the method previously
described.30
In brief, Raf-1 was immunoprecipitated from
protein normalized tissue lysates (0.03 mg), using protein A
Sepharose-antibody complex, and washed four times in lysis buffer.
After the final wash, the immunoprecipitated Raf-1 complexes were
resuspended and incubated for 20 minutes at 25°C in 40 µl kinase
buffer containing 30 mmol/L HEPES (pH 7.4), 7 mmol/L manganese
chloride, 5 mmol/L magnesium chloride, 1 mmol/L dithiothreitol, 15
µmol/L ATP, 10 µCi of [-32P]ATP (3000 Ci/mmol;
Dupont NEN), and 4 units of kinase-inactive MEK. To terminate the
assay, 15 µl of 4x Laemmli sample buffer was added and heated for 5
minutes at 100°C and analyzed by sodium dodecyl
sulfate-polyacrylamide gel electophoresis (SDS-PAGE) and
autoradiography. Quantification was performed with a phosphorimager
(Molecular Dynamics, Sunnyvale, CA).
Effect of Exogenous EGF Treatment on Raf-1 Activity in Ulcer Margins
To determine whether exogenous EGF could modulate Raf-1 activity during the healing process, ulcer margins from 3-day ulcers were carefully dissected and incubated with 10 ng/ml of EGF in Dulbecco’s minimum essential medium (DMEM)/F12 serum-free medium for 15 and 30 minutes in a humidified chamber with 5% CO2. After incubations, tissue samples were snap-frozen in liquid nitrogen and homogenized in lysis buffer, and Raf-1 activity was determined, following the procedure outlined above.
Protein Kinase C Assay
PKC activity was determined using an assay system described in our previous study.31 In brief, frozen tissue samples were lysed in 0.5 ml PKC homogenization buffer (20 mmol/L Tris-HCl, pH 7.4; 2 mmol/L EGTA; 2 mmol/L EDTA; 1% NP-40; 0.33 mol/L sucrose; 0.2 mmol/L sodium orthovanadate; 100 mmol/L sodium fluoride; 10 mmol/L sodium pyrophosphate; 10 µg/ml aprotinin; 10 µg/ml leupeptin; 1 mmol/L phenylmethylsulfonyl fluoride). The protein-normalized lysates were diluted in ice-cold dilution buffer (20 mmol/L Tris-HCl, pH 7.4; 0.2 mol/L NaCl; 0.5 mmol/L EGTA; 0.5 mmol/L EDTA; 10 mmol/L ß-mercaptoethanol), and 25 µl (1 µg of protein) was assayed using the PKC assay system according to the manufacturer’s instructions (Gibco BRL, Gaithersburg, MD).
Immunoprecipitation
Frozen tissues were homogenized in ice-cold lysis buffer (20 mmol/L Tris-HCl (pH 7.5), 50 mmol/L NaCl, 50 mmol/L sodium fluoride, 30 mmol/L sodium pyrophosphate, 5 mol/L EGTA, 10% glycerol, 1% Triton X-100, 1 mmol/L phenylmethylsulfonyl fluoride, 1 mmol/L sodium orthovanadate, and 5 µg/ml aprotinin) and clarified by centrifugation at 14,000 rpm for 10 minutes. The protein concentration of the lysate was determined using a bicinchoninic acid protein assay kit (Pierce, Rockford, IL). Equal amounts of proteins were incubated with specific primary antibody immobilized onto protein A Sepharose for 2 hours at 4°C under gentle rotation. Beads were washed extensively with lysis buffer, and immune complexes were eluted by heating for 5 minutes at 95°C in Laemmli buffer and microcentrifuged. The supernatant was subjected to SDS-PAGE, followed by immunoblotting with specific antibodies (listed in Materials and Methods).
Western Blot Analysis
Western blot analysis was performed following the method previously described.11 Tissue lysates containing equal amounts of proteins (0.15 mg) were subjected to SDS-PAGE and transferred onto a nitrocellulose membrane. Blots were stained with Ponceau Red to ensure equal loading and complete transfer of proteins. The membrane containing the transferred proteins was incubated with blocking buffer, subsequently washed, and incubated with specific primary antibodies (listed in Materials and Methods) for 1 hour at room temperature. Blots were washed and incubated with specific peroxidase-conjugated secondary antibodies for 1 hour at room temperature. After washing, bound antibody was visualized with the ECL detection system (Amersham Corp, IL) according to the manufacturer’s instructions. The density of the protein bands was analyzed using a laser densitometer (UltroScan XL Laser Densitometer; Pharmacia LKB Biotechnology, Uppsala, Sweden).
Ras Assay
Because there is no currently available method allowing determination of Ras activation in vivo, we examined Ras activation in rat gastric epithelial cells. RGM1 cells were plated in 100-mm tissue culture dishes and grown until ~80% confluent in DMEM/F12 supplemented with 20% fetal bovine serum. The cells were serum-starved for 16 hours and metabolically labeled for an additional 8 hours in serum-free, phosphate-free DMEM containing 200 µCi/ml32 PO4. To optimize the dose and time of EGF treatment on Ras activation, cells were treated with various concentrations of EGF (1–100 ng/ml) for 5–60 minutes. Ras activation was determined according to a previously described method.32 Briefly, Ras proteins contained in the cell lysates were immunoprecipitated with rat monoclonal anti-Ras antibody (Y13–259; Santa Cruz Biotechnology, Santa Cruz, CA). The guanine nucleotides bound to the Ras proteins were eluted in 16 µl of 2 mmol/L EDTA, 5 mmol/L dithiothreitol, 1 mmol/L GTP, 1 mmol/L GDP, 0.2% SDS at 68°C for 20 minutes and fractionated by thin-layer chromatography. Quantification was performed with a phosphorimager (Molecular Dynamics). The percentage of GTP bound to Ras (as an indicator of Ras activation) was calculated as cpm in GTP/(cpm in GTP + cpm in GDP) normalized for moles of phosphate in each nucleotide.
ERK2 Assay
ERK activity in RGM1 cells was determined as in our previous
study.11
In brief, serum-starved cells were treated with
various concentrations of EGF (1–100 ng/ml) for 5–60 minutes. After
incubation, cells were washed in cold phosphate-buffered saline and
lysed in lysis buffer (100 mmol/L NaCl, 50 mmol/L HEPES (pH 7.5), 1
mmol/L EDTA, 1% NP-40, 1 µmol/L pepstatin, 0.2 mmol/L
phenylmethylsulfonyl fluoride, 2 µg/ml aprotinin, 1 µg/ml
leupeptin, 0.2 mmol/L sodium orthovanadate, 40 mmol/L paranitrophenyl
phosphate). ERK2 was immunoprecipitated from protein-normalized cell
lysates (0.03 mg), using protein A Sepharose-antibody complex. Beads
were washed twice with lysis buffer and twice with wash buffer (10
mmol/L HEPES, 10 mmol/L magnesium acetate, pH 7.5). Kinase reactions
were performed in 30 µl of tracer buffer (4.0 µCi/tube
[-32P]ATP, 50 µmol/L ATP, 10 mmol/L magnesium
acetate, 7.5 mmol/L HEPES, pH 7.5) and 10 µl of myelin basic protein
(MBP) (3.0 mg/ml) for 30 minutes at 30°C. Then 20 µl of 2x Laemmli
sample buffer was added to each tube, and the samples were boiled for 5
minutes and subjected to 10% SDS-PAGE. The gel was stained with
Coomassie blue R250, dried, and autoradiographed. After this,
individual bands were cut out and counted for isotope labeling by
liquid scintillation spectrometry.
Statistical Analysis
All data are presented as mean ± SD. Student’s t-test was used to determine the statistical significance between ulcers and gastric tissues of sham-operated rats. One-way analysis of variance followed by Bonferroni correction was used for multiple comparisons. A P value of <0.05 was considered to be statistically significant.
|
Results |
|---|
|
TopAbstractIntroductionMaterials and MethodsResultsDiscussionReferences |
|---|
Because our previous study showed induction of the ERK cascade
during gastric ulcer healing,11
in the present study we
attempted to determine the activators of this cascade. We determined
the Raf-1 kinase activity by in vitro phosphorylation of
inactive MEK, using immunoprecipitated Raf-1 kinase from normal (sham
operated) and ulcerated gastric mucosa (Figure 1)
. In ulcerated gastric mucosa there was
about a twofold increase in Raf-1 activity versus controls
(P < 0.01).
|
Protein Kinase C Activity and Protein Level Are Down-Regulated during Gastric Ulcer Healing
PKC has been shown to modulate ERK activity in several cell
systems.23-27
To investigate whether PKC is involved in
ERK activation during gastric ulcer healing, we determined PKC activity
and protein levels in normal and ulcerated gastric mucosa. In ulcerated
gastric mucosa, PKC activity was significantly reduced at 3 days
(2.1-fold decrease; P < 0.002) and 7 days (1.4-fold
decrease) compared to controls (sham operated) (Figure 2)
. PKC protein levels determined by
Western blot analysis (Figure 3)
showed a
significant decrease at 3 days (1.9-fold decrease; P <
0.0001).
|
|
Because in some cell systems in vitro, growth factors
such as EGF can induce activation of Ras through the recruitment of
adapter proteins containing SH2 and SH3 domains such as Shc and
Grb2,15-17
in the present study we examined whether
activation of Raf-1 during ulcer healing in vivo involves
these adapter proteins. Immunoprecipitation of Shc protein followed by
immunoblotting with anti-Grb2 antibody revealed that gastric ulceration
significantly increases Shc binding to Grb2 at both 3 and 7 days
(Figure 4)
. At 3 days there was a
1.4-fold increase (P < 0.0001) and at 7 days
there was a 1.2-fold increase (P < 0.0002) in
Grb2 bound to Shc protein versus controls (sham operated).
|
Because we found increased binding of Grb2 to Shc during ulcer
healing, and because this association has been shown to result in
recruitment of Sos and Ras activation,15-18
we next
examined whether the guanidine exchange factor Sos was bound to Grb2.
This was analyzed by immunoprecipitation of Sos followed by
immunoblotting with Grb2 antibody. In ulcerated gastric mucosa, the
Grb2-Sos complex level was increased by 1.8-fold
(P < 0.0001) at 3 days and by 2.2-fold
(P < 0.002) at 7 days versus
controls (sham operated) (Figure 5)
.
|
To determine whether the above increase in Shc-Grb2-Sos
association reflects an increase in respective protein levels during
ulcer healing, we performed Western blot analysis for Shc, Grb2, and
Sos proteins. As shown in Figure 6A
, all
three isoforms of Shc protein (corresponding to 46, 52, and 66 kd) were
expressed almost equally in both normal and ulcerated gastric mucosa.
Grb2 showed substantial increases in protein levels in ulcerated mucosa
versus controls at both 3 days (1.2-fold increase;
P < 0.03) and 7 days (1.5-fold increase;
P < 0.0001) (Figure 6B)
. Gastric ulceration did not
significantly affect Sos protein levels (Figure 6C)
.
|
Because we observed significant increases in Shc-Grb2-Sos complex
formation during ulcer healing, we attempted to determine whether Ras
protein levels were affected. Immunoblot analysis showed that gastric
ulceration significantly increased the Ras protein level at 3 days
(P < 0.025) and sustained an increased level at
7 days (Figure 7)
versus
controls.
|
Previous studies showed that gastric ulceration triggers increased
EGF-R expression8
and ERK activity in epithelial cells of
the ulcer margins.11
In the present study we observed
increased Ras protein and Raf-1 kinase activity levels in the ulcer
margins. These findings prompted us to hypothesize that activation of
the Ras–Raf-1–ERK cascade occurs predominantly in the epithelial
component of the ulcer margins. To test this, we studied the effect of
EGF on Ras activation and PKC and ERK activities in an epithelial cell
line (RGM1) derived from normal rat gastric mucosa. EGF treatment (10
ng/ml; 5 minutes) did not alter PKC activity in rat gastric epithelial
cells versus untreated controls (Figure 8)
. However, treatment with 50 nM PMA for
5 minutes (positive control) significantly increased PKC activity.
Prolonged treatment with 50 nM PMA (24 hours; negative control)
significantly decreased PKC activity in RGM1 cells versus
untreated controls, showing the efficacy of the signaling system in
these cells. Dose-response studies for EGF-stimulated Ras activation
and ERK2 activity (EGF 1–100 ng/ml; 5 minutes) demonstrated a maximum
response at 10 ng/ml (Figures 9A and 10A)
versus untreated
controls. Time response studies for EGF-stimulated Ras activation and
ERK2 activity (5–60 minutes; EGF 10 ng/ml) showed a maximum response
at 5 minutes (Figures 9B and 10B)
versus untreated controls.
|
|
|
|
Discussion |
|---|
|
TopAbstractIntroductionMaterials and MethodsResultsDiscussionReferences |
|---|
, and basic
fibroblast growth factor, have been shown to accelerate gastric injury
repair and ulcer healing by stimulating cell migration and
proliferation.4-8
However, the cellular signaling
pathways involved in these two processes in the in vivo
condition are not fully elucidated. Our previous study demonstrated
increased receptor tyrosine kinase activity and induction of the ERK
cascade during ulcer healing.11
Furthermore, it showed
that treatment with Tyrphostin A46, a potent inhibitor of EGF-R kinase
and EGF-R kinase-dependent cell proliferation,33
dramatically delays ulcer healing,11
clearly demonstrating
the importance of the EGF-R–ERK signaling pathway in the healing
process. Our present study shows that gastric ulceration increases
Raf-1 kinase activity and Grb2 and Ras protein levels and triggers
Shc-Grb2-Sos association. PKC activity and protein levels were
down-regulated in ulcer margins. In a rat gastric epithelial cell line
(RGM1), EGF significantly increased Ras activation and ERK activity,
without affecting PKC activity, supporting our contention that
activation of the Raf-1–ERK cascade occurs predominantly in the
epithelial component of the ulcer margins. While EGF has been shown to
play a crucial role in ulcer healing by activation of the ERK signaling
pathway,11
activation of this pathway by other growth
factors may also be important. The gut epithelial motility itself (even
in the absence of growth factors) might be expected to be associated
with intracellular signals,34
perhaps via
integrin-mediated altered cell-matrix interaction.35,36 The Ras and Raf-1 proto-oncogene products are key proteins in the transmission of a variety of intracellular proliferative and differentiation signals. Raf-1 and Ras serve as intermediates in these signaling pathways by connecting upstream tyrosine kinases with downstream serine/threonine kinases such as MAPK (or ERK) and MAPK kinase (MEK).12,13 This phosphorylation cascade leads to activation of transcription factors involved in cell growth and differentiation.12,13 Raf-1-dependent activation of ERK1 and ERK2 has been demonstrated in wounded intestinal epithelial monolayers,37 in mechanically stretched rat cardiac myocytes,38 and in shear stress exposed human endothelial cells,39 but has not been explored during healing of injured gastric mucosa. In our present study, we demonstrated that gastric ulceration triggers a significant increase in Raf-1 activity. During ulcer healing the Raf-ERK signaling pathway is significantly activated, most likely by endogenous EGF produced by regenerating epithelial cells (ulcer-associated lineage).9 Therefore, epithelial cells in the ulcer margin may have already attained submaximum stimulation due to overexpression of EGF and EGF-R8,9 and may render them less susceptible to exogenous EGF. This could explain why treatment of ulcer margins with exogenous EGF causes only a very modest (nonsignificant) increase in Raf-1 activity when compared to untreated ulcer margins.
Potential activators of Raf-1 include protein kinase C and activated Ras.14 While studies of a number of tissues have implicated PKC in the positive control of cell growth and transformation, some data also point to its involvement in cell growth inhibition and differentiation.40 PKC represents a family of phospholipid-dependent serine/threonine kinases consisting of 11 members, including the conventional, novel, and atypical isozymes, which differ in their structural and biochemical properties.41 Individual PKC isozymes are known to exhibit varying substrate specificities, tissue distributions, and subcellular localization. Although the understanding of biological functions of individual isozymes and the molecular regulatory pathways in which they participate remains limited, individual isozymes may play specific specialized roles in cell signaling.
Not only can PLC-1 activated by receptor tyrosine kinases generate
DAG and subsequently activate PKC; it can also associate with SH2
domains of activated EGF-R and with both dynamin and Sos (through its
SH3 domain) in EGF-stimulated Madin-Darby canine kidney
cells.22
Previous studies have shown increased tyrosine
kinase activity and PLC-1 phosphorylation during the early phase of
acute injury repair in rat gastric mucosa, which is a process that is
entirely different from chronic gastric ulcer
healing.28,29
Our present finding that gastric ulceration
down-regulates both PKC activity and protein levels indicates that ERK
activation during gastric ulcer healing is PKC-independent. The
observed reduction in both activity and protein levels of PKC could be
due to either a lack or reduced availability of a co-factor necessary
for PKC activation,42
tightly controlled regulation
preventing its inhibitory effect on cell cycle progression (as shown in
intestinal epithelial cells (IEC-18)),43
] or loss of a
particular cell type, eg, parietal cells, which have been shown to
express high levels of PKC.44
The latter possibility is
supported by our previous study demonstrating a substantial reduction
of the parietal cells in dilated glands lining the ulcer
margin.45
Furthermore, the substrate and the antibody used
in the present study to measure PKC activity and the protein level,
respectively, recognize only the conventional isoforms, -, ß-, and
-isozymes (which share a high sequence homology), and not the novel
isoforms, 
-, 
-, and 
-isozymes (which have lower sequence
homology). Therefore, a possibility that isozyme-specific PKC
activation (eg, -, -, -isozymes) may occur during chronic
gastric ulcer healing cannot be completely ruled out by our studies.
Studies of various cell systems have shown that Shc, Grb2, and Sos link activated tyrosine kinases to Ras.15,16 Previous studies have demonstrated increased receptor tyrosine kinase activity and increased EGF-R phosphorylation levels during gastric mucosal injury repair.11,28,29 Our data demonstrating increased binding of adapter protein Grb2 to Shc and to Sos indicate their involvement as mediators of receptor tyrosine kinase signaling via Ras during ulcer healing. In the present in vivo study, we examined Ras protein expression during ulcer healing but were unable to determine Ras activation, because currently there is no available method allowing for this in an in vivo condition. Because the method of Ras activation measurement in vitro is well established, we used gastric epithelial RGM1 cells to study the effect of EGF on Ras activation. In this model, EGF dramatically increased Ras activation and ERK2 activity but not PKC activity, clearly indicating that in gastric epithelial cells, EGF-triggered induction of ERK activity is mediated by Ras and not by PKC.
In summary, this study demonstrates for the first time that during experimental gastric ulcer healing, Raf-1 activity is induced and that Raf-1 activation involves increased Shc-Grb2-Sos association. Furthermore, this study indicates that ERK activation during ulcer healing is not mediated by PKC. Similarly, treatment of rat gastric epithelial cells with EGF increased Ras and ERK activity but not PKC activity. Thus activation of the Raf-1–ERK cascade during gastric ulcer healing is Ras-mediated, independent of PKC, and is attributable to the epithelial component of ulcer margins.
|
Footnotes |
|---|
Supported by a Merit Review Award from the Department of Veterans Affairs to AST.
Accepted for publication July 8, 1999.
|
References |
|---|
|
TopAbstractIntroductionMaterials and MethodsResultsDiscussionReferences |
|---|
1 isoform with tyrosine kinase. Trends Biochem Sci 1991, 16:297-301[Medline]
in vitro by epidermal growth factor receptor. Biochem J 1993, 293:507-511
is a substrate for the PDGF and EGF receptor protein-tyrosine kinases in vivo and in vitro. Cell 1989, 57:1109-1122[Medline]
1 enzyme activity in growth factor-induced mitogenesis. Mol Cell Biol 1998, 18:590-597This article has been cited by other articles:
 |
|
 |
|
  N. J. Eungdamrong and R. Iyengar Compartment-Specific Feedback Loop and Regulated Trafficking Can Result in Sustained Activation of Ras at the Golgi Biophys. J., February 1, 2007; 92(3): 808 - 815. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 E. Kamoshita, Y. Ikeda, M. Fujita, H. Amano, A. Oikawa, T. Suzuki, Y. Ogawa, S. Yamashina, S. Azuma, S. Narumiya, et al. Recruitment of a Prostaglandin E Receptor Subtype, EP3-Expressing Bone Marrow Cells Is Crucial in Wound-Induced Angiogenesis Am. J. Pathol., October 1, 2006; 169(4): 1458 - 1472. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
D. Baatar, M. K. Jones, K. Tsugawa, R. Pai, W. S. Moon, G. Y. Koh, I. Kim, S. Kitano, and A. S. Tarnawski Esophageal Ulceration Triggers Expression of Hypoxia-Inducible Factor-1{alpha} and Activates Vascular Endothelial Growth Factor Gene : Implications for Angiogenesis and Ulcer Healing Am. J. Pathol., October 1, 2002; 161(4): 1449 - 1457. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 I. L. Szabo, R. Pai, M. K. Jones, G. R. Ehring, H. Kawanaka, and A. S. Tarnawski Indomethacin Delays Gastric Restitution: Association with the Inhibition of Focal Adhesion Kinase and Tensin Phosphorylation and Reduced Actin Stress Fibers Experimental Biology and Medicine, June 1, 2002; 227(6): 412 - 424. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
D. Baatar, M. K. Jones, R. Pai, H. Kawanaka, I. L. Szabo, W. S. Moon, S. Kitano, and A. S. Tarnawski Selective Cyclooxygenase-2 Blocker Delays Healing of Esophageal Ulcers in Rats and Inhibits Ulceration-Triggered c-Met/Hepatocyte Growth Factor Receptor Induction and Extracellular Signal-Regulated Kinase 2 Activation Am. J. Pathol., March 1, 2002; 160(3): 963 - 972. [Abstract] [Full Text] [PDF]
|
|
| |||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
| HOME | HELP | FEEDBACK | SUBSCRIPTIONS | ARCHIVE | SEARCH | TABLE OF CONTENTS |
