Department of Medicine, University of California, San Diego, California 92103
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ABSTRACT |
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We have
previously shown that Ca2+-dependent Cl
secretion across intestinal epithelial cells is limited by a signaling
pathway involving transactivation of the epidermal growth factor
receptor (EGFR) and activation of ERK mitogen-activated protein kinase (MAPK). Here, we have investigated a possible role for p38 MAPK in
regulation of Ca2+-dependent Cl
secretion.
Western blot analysis of T84 colonic epithelial cells revealed that the muscarinic agonist carbachol (CCh; 100 µM)
stimulated phosphorylation and activation of p38 MAPK. The p38
inhibitor SB-203580 (10 µM) potentiated and prolonged short-circuit
current (Isc) responses to CCh across
voltage-clamped T84 cells to 157.4 ± 6.9% of those
in control cells (n = 21; P < 0.001).
CCh-induced p38 phosphorylation was attenuated by the EGFR inhibitor
tyrphostin AG-1478 (0.1 nM-10 µM) and by the Src family kinase
inhibitor PP2 (20 nM-2 µM). The effects of CCh on p38
phosphorylation were mimicked by thapsigargin (TG; 2 µM), which
specifically elevates intracellular Ca2+, and were
abolished by the Ca2+ chelator BAPTA-AM (20 µM), implying
a role for intracellular Ca2+ in mediating p38 activation.
SB-203580 (10 µM) potentiated Isc responses to
TG to 172.4 ± 18.1% of those in control cells (n = 18; P < 0.001). When cells were pretreated with
SB-203580 and PD-98059 to simultaneously inhibit p38 and ERK MAPKs,
respectively, Isc responses to TG and CCh were
significantly greater than those observed with either inhibitor alone.
We conclude that Ca2+-dependent agonists stimulate p38 MAPK
in T84 cells by a mechanism involving intracellular
Ca2+, Src family kinases, and the EGFR. CCh-stimulated p38
activation constitutes a similar, but distinct and complementary,
antisecretory signaling pathway to that of ERK MAPK.
G protein-coupled receptor; epidermal growth factor receptor; intestinal secretion
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INTRODUCTION |
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FLUID TRANSPORT ACROSS intestinal epithelial cells is a finely balanced process with net absorption predominating under normal circumstances. However, a basal level of fluid secretion that is necessary for adequate hydration of the intestinal mucosal surface and that enables appropriate mixing of food particles with digestive enzymes and diffusion of digested nutrients to the epithelial layer normally occurs. Epithelial fluid secretion may be upregulated in response to a wide range of physiological stimuli, such as distension of the intestinal wall and the presence of antigenic material in the lumen. Under normal circumstances, upregulation of fluid secretion is advantageous because it aids in the smooth passage of digested materials through the intestine and in the expulsion of noxious substances from the intestinal tract before they can gain access to the systemic circulation. However, certain pathological conditions such as inflammatory bowel diseases and bacterial infections can result in an overexpression of epithelial fluid secretion, leading to the clinical manifestation of secretory diarrhea.
In the intestine, the secretion of water is a passive process driven by
the active secretion of ions, predominantly Cl (2,
28). Increases in chloride secretion are usually brought about
by hormones, neurotransmitters, and immune cell mediators, which act on
specific receptors on the surface of epithelial cells to increase
intracellular levels of second messengers, such as cyclic nucleotides
and Ca2+. In turn, increases in the levels of intracellular
messengers activate the transport proteins that comprise the chloride
secretory mechanism. However, even though both Ca2+ and
cyclic nucleotides are capable of promoting epithelial secretion, these
two classes of intracellular messenger stimulate distinctive secretory
responses. Responses to cyclic nucleotide-mediated agonists are
sustained, whereas those to Ca2+-mediated agonists are
transient even though levels of intracellular Ca2+ can
remain elevated after the secretory response has resolved (2). This implies that negative signals exist within
intestinal epithelial cells that limit, or "switch off," epithelial
secretory responses to Ca2+-mediated agonists. Such
antisecretory signaling pathways may represent physiological
"braking" mechanisms to prevent excessive Cl
and
fluid secretion when mucosal levels of neuroimmune mediators are elevated.
We have previously identified a signaling pathway in intestinal
epithelia that appears to be involved in rapidly downregulating Ca2+-dependent Cl secretory responses
(25). In this study, we found that treatment of
T84 colonic epithelial cells with the muscarinic
Gq protein-coupled receptor (GPCR) agonist carbachol (CCh)
rapidly (<2 min) stimulated transactivation of the epidermal growth
factor receptor (EGFR) and subsequent activation of the ERK isoforms of
mitogen-activated protein kinase (MAPK). Because pharmacological
inhibition of either ERK or the EGFR resulted in potentiation of
CCh-stimulated secretory responses, we proposed that, in vivo,
recruitment of this EGFR/ERK signaling pathway may serve as a
physiological braking mechanism that limits the extent of
agonist-induced secretory responses.
Although our studies demonstrated that ERK MAPK plays a role in
downregulating epithelial Ca2+-dependent Cl
secretory responses, the role of other members of the MAPK family in
this regard was unknown. The stress-activated protein kinase p38 MAPK
has been shown to be expressed in intestinal tissues and is activated
by a variety of cellular stresses and cytokines (1, 9,
40). p38 MAPK has been shown to play a role in regulating a
variety of intestinal processes, including pancreatic amylase
secretion, epithelial restitution, and cytoskeletal organization (13, 16, 39, 46). Recent studies demonstrate that p38 MAPK
may also play a role in regulating intestinal epithelial ion transport.
For example, p38 has been shown to be involved in regulating
Na+/H+ exchange in Caco-2 cells and to inhibit
acid secretion in cultured parietal cells (35, 41).
Therefore, to further our understanding of the role of MAPKs in
regulating intestinal function, in the present study we have examined
the potential role for p38 in regulating Ca2+-dependent
Cl
secretion in colonic epithelial cells.
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EXPERIMENTAL PROCEDURES |
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Materials. CCh was obtained from Sigma Chemical, St. Louis, MO. Tyrphostin AG-1478, SB-203580, PD-98059, BAPTA-AM, and PP2 were obtained from Calbiochem, San Diego, CA. Epidermal growth factor (EGF) was obtained from Genzyme, Cambridge, MA. Thapsigargin (TG) was obtained from LC Laboratories, Lexington, MA. Rabbit anti-phospho-p38 antibodies were obtained from New England Biolabs, MA. Tris-glycine electrophoresis gels were obtained from Bio-Rad, Hercules, CA. All other reagents were of analytical grade and were obtained commercially.
Cell culture. Methods for maintenance of T84 cells in culture were as previously described (44). Briefly, T84 cells were grown in DME/F12 media (JRH, Lexena, KS) supplemented with 5% newborn calf serum. Cells were passaged by trypsinization. For Ussing chamber/voltage-clamp experiments, ~5 × 105 cells were seeded onto 12-mm Millicell transwell polycarbonate filters. For Western blotting experiments and MAPK assays, 106 cells were seeded onto 30-mm Millicell transwell polycarbonate filters. When grown on polycarbonate filters, T84 cells are known to retain the polarized phenotype of native colonic epithelia. Cells seeded onto filters were cultured for ~15 days before use.
Electrophysiological studies.
Monolayers of T84 cells were mounted in Ussing chambers
(window area = 0.6 cm2) and bathed in oxygenated (95%
O2-5% CO2) Ringer's solution at 37°C. The
composition of the Ringer's solution was (in mM) 140 Na+,
5.2 K+, 1.2 Ca2+, 0.8 Mg2+, 120 Cl, 25 HCO
Isc) in response
to agonists are wholly reflective of electrogenic chloride secretion
(4).
Western blotting. Polarized T84 cell monolayers grown on Millicell filters were washed (2×) with Ringer's solution, allowed to equilibrate for 30 min at 37°C, and then stimulated with agonists (±antagonists) for the times indicated. The reaction was stopped by washing in ice-cold phosphate-buffered saline, and the cells were then lysed in ice-cold lysis buffer (1% Triton X-100, 1 µg/ml leupeptin, 1 µg/ml pepstatin, 1 µg/ml antipain, 100 µg/ml PMSF, 1 mM Na+-vanadate, 1 mM NaF, and 1 mM EDTA in PBS) for 45 min. Cells were then scraped into microcentrifuge tubes and spun at 12,000 rpm for 10 min, and the pellet was discarded. Samples were assayed for protein content (Bio-Rad protein assay kit; Hercules, CA) and adjusted so that each sample contained an equal amount of protein. Samples were then mixed with an equal volume of 2× gel loading buffer (50 mM Tris, pH 6.8, 2% SDS, 200 mM dithiothreitol, 20% glycerol, and 0.2% bromophenol blue), boiled for 2 min, and loaded onto a polyacrylamide gel, and proteins were separated by electrophoresis. Resolved proteins were transferred overnight at 4°C onto a polyvinylidene (PVDF) membrane (New England Nuclear, Boston, MA). After transfer, the membrane was preblocked with a 1% solution of blocking buffer (Upstate Biotechnology, Lake Placid, NY) for 30 min, followed by a 2-h incubation with antiphospho-p38 antibodies in 2% blocking buffer. After being washed (4 × 10 min) in Tris-buffered saline with 1% Tween (TBST), membranes were then incubated for 30 min in horseradish peroxidase-conjugated secondary antibody (anti-rabbit IgG; Transduction Laboratories, Lexington, KY) in 1% blocking buffer. This was followed by four 10-min washes in TBST. Proteins were then detected using an enhanced chemiluminescence detection kit (Amersham Pharmacia Biotech, UK). Densitometric analysis of Western blots was carried out using NIH Image software.
p38 MAPK assays. p38 MAPK assays were performed using a commercially available kit per the manufacturers' instructions. Briefly, cells were stimulated with CCh and lysed as described above. Lysates were normalized for protein concentration and then immunoprecipitated overnight with agarose-conjugated antiphospho-p38 antibodies. Immunoprecipitates were washed twice in lysis buffer and twice in p38 MAPK assay buffer. Washed immunoprecipitates were then incubated for 30 min at 30°C with 100 µM ATP and 1 µg ATF-2 substrate in 50 µl of assay buffer. Reactions were stopped by the addition of 50 µl 2× gel loading buffer, and samples were boiled for 3 min before SDS-PAGE. Phosphorylation of ATF-2 by p38 MAPK was determined by Western blotting (as described above) with antibodies specific for the phosphorylated form of ATF-2.
Statistical analyses. All data are expressed as means ± SE for a series of n experiments. Student's t-tests or analysis of variance (ANOVA) with the Student-Newman-Keuls post test were used to compare mean values as appropriate. P values <0.05 were considered to represent significant differences.
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RESULTS |
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CCh stimulates p38 MAPK phosphorylation and activation in
T84 cells.
The muscarinic M3 receptor agonist CCh was used as a
prototypical Ca2+-dependent secretagogue in these studies.
In our first experiments, we set out to determine whether CCh
stimulates activation of p38 MAPK in T84 cells. Cells grown
on permeable supports were stimulated basolaterally with CCh (100 µM)
for periods ranging from 0.5-15 min, and cell lysates were
analyzed by Western blotting with an antibody specific for the
phosphorylated form of p38 MAPK. We found that CCh stimulated p38 MAPK
phosphorylation in a time-dependent manner (Fig.
1A). Increases in levels of
p38 phosphorylation in response to CCh were not due to alterations in
expression of the protein because levels of total p38 in cells
stimulated with CCh (100 µM) for 15 min were 85.0 ± 15.3% of
those in control cells (n = 4). The time course for the
effects of CCh on p38 MAPK phosphorylation was slightly slower than for
CCh-stimulated ERK phosphorylation (Fig. 1B) with a response
occurring 2-5 min after agonist addition and a maximal effect
observed between 5-15 min. The time course of p38 phosphorylation
also lagged behind that of CCh-induced Isc
responses across voltage-clamped monolayers of T84 cells
(Fig. 1C), with p38 phosphorylation increasing toward
maximal levels as Isc responses returned to
baseline. CCh-stimulated p38 MAPK phosphorylation was closely
accompanied by an increase in activity of the enzyme, as determined by
the ability of CCh-stimulated antiphospho-p38 immunoprecipitates to
phosphorylate the p38 MAPK substrate ATF-2 (Fig. 1D).
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SB-203580 potentiates CCh-stimulated
Cl secretion in T84 cells.
Next, to determine whether stimulation of p38 MAPK plays a role in
regulation of CCh-stimulated Cl
secretion, we examined
the effects of the p38 MAPK inhibitor SB-203580 on
Isc responses to CCh across
voltage-clamped monolayers of T84 cells mounted in
Ussing chambers. Pretreatment of T84 cells with SB-203580
(10 µM) significantly potentiated and prolonged subsequent
Isc responses to CCh (Fig.
2A). Maximal responses to CCh
in control cells were 27.7 ± 3.0 µA/cm2 compared
with 43.0 ± 4.7 µA/cm2 in SB-203580-pretreated
cells (n = 21; P < 0.001). Responses to CCh remained significantly greater in SB-203580-treated cells up to
7 min after addition of CCh. SB-203580 (10 µM) did not alter baseline
Isc (data not shown). Concentration-response
analysis revealed that SB-203580 was maximally effective in
potentiating CCh-induced Isc responses between
concentrations of 3 and 10 µM (Fig. 2B). At this
concentration, SB-203580 is known to be specific for p38 MAPK
(10, 14, 30), and we found that it did not alter
CCh-induced phosphorylation of ERK MAPK in T84 cells. ERK phosphorylation in SB-203580-pretreated cells was found to be 119 ± 23.9% of that in cells stimulated with CCh alone (n = 6).
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TG, but not PMA, stimulates p38 phosphorylation in T84
cells.
Having demonstrated a role for p38 MAPK in negative regulation of
CCh-stimulated Cl secretion, we next went on to examine
possible signal transduction mechanisms that might underlie
CCh-stimulated p38 activation. Because GqPCRs such as the
M3 muscarinic receptor for CCh on T84 cells are
coupled to both activation of protein kinase C (PKC) and elevations in
intracellular Ca2+, we examined whether selective
activation of either of these pathways alone could mimic the effects of
CCh on p38 phosphorylation. PKC was stimulated using the phorbol ester
PMA, whereas the endoplasmic reticulum Ca2+-ATPase
inhibitor TG was employed to elevate intracellular Ca2+. We
found that treatment of T84 cells with TG (2 µM), but not with PMA (100 nM), resulted in increased phosphorylation of p38 MAPK
(Fig. 3, A-B).
The lack of effect of PMA on p38 MAPK phosphorylation was not due to
inactivity of the compound because, like TG, and as we have previously
shown (25), PMA effectively stimulated ERK phosphorylation
by 31.1 ± 14.7-fold over controls within 5 min after addition to
T84 cells (n = 3). These data suggest that CCh-stimulated p38 MAPK activation likely occurs by a mechanism involving elevations in intracellular Ca2+. To confirm
this, we examined the effects of the intracellular Ca2+
chelator BAPTA-AM on CCh-stimulated p38 phosphorylation and found that
pretreatment of T84 cells with BAPTA-AM (20 µM)
practically abolished CCh-stimulated p38 MAPK phosphorylation (Fig. 3,
C-D).
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CCh-stimulated p38 MAPK phosphorylation is mediated by a mechanism
involving Src family kinases and transactivation of the EGFR.
Our previous studies showed that CCh-stimulated ERK MAPK activation
occurs by a pathway involving transactivation of the EGFR, secondary to
increases in intracellular Ca2+. We therefore examined the
possibility that CCh-stimulated p38 MAPK activation might also occur
via a similar mechanism. We first examined whether stimulation of
T84 cells with a bona fide ligand for the EGFR, EGF, could
also stimulate p38 activation. Similar to CCh, basolateral treatment of
T84 cells with EGF (100 ng/ml) stimulated a time-dependent,
albeit slower, increase in phosphorylation of the enzyme (Fig. 5,
A and B). Next, to
determine whether the EGFR mediates the effects of CCh on p38
phosphorylation, we examined the effects of the EGFR inhibitor
tyrphostin AG-1478 on phosphorylation of the enzyme (Fig. 5,
C and D). Pretreatment of T84 cell
monolayers with tyrphostin AG-1478 (10 nM-1 µM) inhibited
CCh-stimulated p38 phosphorylation in a concentration-dependent manner.
We have previously found that, at these concentrations, tyrphostin
AG-1478 effectively inhibits transactivation of the EGFR in response to CCh (25). Thus, similar to its effects on ERK activation,
CCh appears to stimulate p38 MAPK activation by a pathway involving transactivation of the EGFR.
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DISCUSSION |
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The secretion of fluid and electrolytes across intestinal
epithelial cells in response to Ca2+-dependent
secretagogues is a tightly regulated process that is subject to both
positive and negative influences. Although the mechanisms by which
agonist-stimulated elevations in intracellular Ca2+
stimulate Cl secretion are relatively well defined
(2, 28), the mechanisms underlying its negative regulation
are only beginning to be understood. However, over the past several
years, work from this laboratory has succeeded in identifying several
intracellular signals that appear to be involved in downregulation of
this physiologically important process. Agonists of either receptor
tyrosine kinases or GqPCRs appear to have the ability to
evoke these antisecretory mechanisms. Growth factors, such as EGF,
insulin, and heregulin, inhibit chloride secretory responses via
stimulation of a phosphatidylinositol 3-kinase-dependent signaling
pathway (6, 7, 23, 42). GqPCR agonists appear
to have the ability to evoke two types of antisecretory signal. One is
mediated by the generation of inositol (3,4,5,6)
tetrakisphosphate and serves to chronically downregulate epithelial
responsiveness to subsequent challenge with Ca2+-dependent
agonists (20, 43). The other is mediated by rapid transactivation of the EGFR and ERK MAPK and serves to limit the extent
of ongoing Ca2+-dependent secretory responses
(25). Our present data suggest that this rapidly acting
antisecretory pathway may also involve p38 MAPK.
We found that CCh, a GqPCR agonist that acts via the
muscarinic M3 receptor, stimulates phosphorylation of p38
MAPK in T84 colonic epithelial cells. Phosphorylation of
p38 was accompanied by an increase in activity of the enzyme as
measured by the ability of CCh-stimulated antiphospho-p38
immunoprecipitates to phosphorylate the p38 MAPK substrate ATF-2. Of
note, the onset of p38 MAPK activation occurs 2-5 min after
stimulation with CCh, which temporally corresponds to that time at
which CCh-stimulated chloride secretion is in decline. Thus a temporal
relationship exists between CCh-stimulated p38 activation and the
termination of CCh-stimulated chloride secretory responses. That p38
MAPK is involved in negative regulation of CCh-stimulated chloride
secretion is supported by experiments in which we examined the effects
of the specific p38 MAPK inhibitor SB-203580 on CCh-stimulated
Isc responses across voltage-clamped monolayers
of T84 cells. This agent inhibits p38 by blocking the ATP-binding site of the enzyme and, at the concentrations employed in
this study, has no activity toward other members of the MAPK family
(10, 14, 30, and data presented herein). Pretreatment of T84 cells with SB-203580 significantly potentiated
subsequent secretory responses to CCh compared with control cells.
Responses to CCh were also prolonged by pretreatment with SB-203580 and remained significantly greater than those in control cells for up to 7 min after addition of the agonist. In vivo, such a
potentiating effect on Cl secretion would be expected to
increase the osmotic driving force for fluid secretion into the
intestine. Thus it seems that, similar to ERK (25), p38
MAPK may act as a physiological braking mechanism that serves to limit
the extent of chloride secretion in response to neurohumoral mediators
that act via increases in intracellular Ca2+.
It is interesting to note that other investigators have found that p38 MAPK may also be involved in limiting the extent of secretory responses in other parts of the intestine. Pausawasdi and coworkers (35) have recently shown that CCh stimulates p38 activity in canine parietal cells and that inhibition of p38 activation results in potentiation of CCh-induced acid secretion. Their data, which were generated in gastric epithelial cells, are analogous to those we present here in colonic epithelium and raise the possibility that MAPKs may mediate a generalized "switching off" system for secretory responses to Ca2+-dependent agonists throughout the intestine.
That signaling cross talk exists between GPCR signaling pathways and MAPK cascades has been known for several years. The mechanisms by which GPCR agonists activate ERK are highly heterogeneous and depend upon cell type, the subtype of GPCR in question, and the relative abundance of signaling proteins present (12, 17, 18, 32, 37). Although less is known about signaling mechanisms linking GPCRs to the p38 MAPK cascade, it is becoming apparent that these too are heterogeneous in nature. Depending on cell type, p38 activation can occur by mechanisms that are dependent on elevations in intracellular Ca2+, activation of PKC, or both (11, 22, 34-36). Whether tyrosine kinases are involved is also cell type-specific (15, 21, 29, 33, 34, 45). To begin investigating the signaling pathways underlying CCh-induced p38 activation in our colonic epithelial cell model, we first examined whether selective activation of either of the classical signaling pathways linked to GqPCR activation mimicked the effects of CCh on p38 phosphorylation. We found that TG, an agent that inhibits the endoplasmic reticulum Ca2+-ATPase pump and thereby elevates intracellular levels of Ca2+, was sufficient to stimulate p38 MAPK. In contrast PMA, an activator of PKC, did not alter phosphorylation of the enzyme over the time course studied. This indicates that activation of p38 by CCh more likely involves elevations in intracellular Ca2+ than activation of PKC. This hypothesis is supported by our findings with the intracellular Ca2+ chelator BAPTA-AM, which practically abolished p38 phosphorylation in response to CCh. These data are reminiscent of our previous work, in which we have found that CCh-stimulated ERK activation is also mediated by increases in intracellular Ca2+ and not by PKC (24), and imply that CCh-stimulated p38 and ERK activation may both occur by similar means.
Our present studies also reveal that, similar to ERK, p38 activation is mediated by Src-dependent EGFR transactivation. That the EGFR is involved is based on the finding that tyrphostin AG-1478 significantly attenuated CCh-induced p38 phosphorylation. At the concentrations employed in these studies, tyrphostin AG-1478 is specific for the EGFR over other members of the ErbB family of receptor tyrosine kinases (31) and effectively inhibits transactivation of the EGFR in response to CCh in T84 cells (25). That the EGFR is capable of transducing signals to the p38 MAPK cascade is supported by the observation that its cognate ligand EGF also stimulates phosphorylation of the enzyme, albeit with kinetics distinct from those of CCh. That Src-family kinases are involved in mediating CCh-stimulated p38 activation is based on the findings that, similar to its effects on EGFR and ERK phosphorylation (24), the Src family kinase inhibitor PP2 attenuated CCh-stimulated p38 phosphorylation. Interestingly, we found that the concentration-dependence of the effects of PP2 on CCh-stimulated ERK and p38 phosphorylation are somewhat different, with phosphorylation of p38 being more sensitive to inhibition by PP2 than that of ERK (24). These data might point to different members of the Src family of kinases being involved in mediating the effects of CCh on ERK and p38 activation. Although our previous studies have shown that CCh stimulates association of p60src with the EGFR with kinetics that coincide with EGFR phosphorylation (24), the possibility that other members of the Src family of kinases might also be involved has not yet been studied.
Thus the data presented in this study suggest that, in intestinal
epithelial cells, the EGFR acts as a central signaling molecule through
which GPCR agonists that elevate intracellular Ca2+ bring
about activation of both ERK and p38 MAPKs. Our previous data suggest
that transactivation of the EGFR occurs by a mechanism involving
activation of the Ca2+-dependent tyrosine kinase Pyk-2 and
Src family kinases that form a complex with the EGFR. Although
activation of both ERK and p38 appears to occur by a similar pathway,
and both MAPK isoforms appear to play a similar role in downregulation
of Ca2+-dependent secretory responses, it appears that p38
and ERK ultimately act independently of one another to exert their
inhibitory effects. This conclusion is based on the observation that
simultaneous inhibition of p38 and ERK resulted in greater potentiation
of Ca2+-dependent secretory responses than when either
enzyme was inhibited alone. It remains to be determined how ERK and p38
ultimately interact with the ion transport machinery of epithelial
cells to inhibit Cl secretion, but this is currently an
area of active research in our laboratory. A schematic representation
of our current understanding of the role of MAPKs in regulation of
Ca2+-dependent secretory responses is depicted in Fig.
7.
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The critical role of MAPKs in regulation of epithelial function is
gradually being uncovered. It is now well established that in addition
to their classical roles in mediating changes in cell growth and
differentiation in response to growth factors, MAPKs also play
important roles in modulating cellular responses to a broad panel of
physiological and pathophysiological stimuli, including cytokines,
bacteria, osmotic stress, monolayer disruption, and GPCR agonists
(1, 8, 9, 27, 35, 38-40, 45). In turn, MAPK
activation modulates an ever increasing number of cellular responses,
including mitogenesis, cytokine synthesis and release, alterations in
tight junction permeability, cytoskeletal organization, nutrient
absorption, and cell migration (5, 13, 16, 19, 26, 46).
The importance of MAPKs in regulating intestinal ion transport is also
becoming more recognized, and we now know that MAPKs may be involved in
regulation of gastric acid secretion, ileal Na+ absorption,
and colonic Cl secretion (3, 35, 41). The
present study furthers our current understanding of the role of MAPKs
in regulation of intestinal epithelial physiology and underscores the
complexity of signaling mechanisms that regulate
Ca2+-dependent Cl
secretory responses. It is
our hope that a greater understanding of the ways in which
antisecretory signaling mechanisms interact with the Cl
secretory machinery of epithelial cells will ultimately lead to the
development of new and more specific approaches for treatment of
intestinal disorders associated with dysregulated epithelial transport.
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ACKNOWLEDGEMENTS |
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These studies were supported by a Career Development Award from the Crohn's and Colitis Foundation of America to S. J. Keely and by a National Institute of Diabetes and Digestive and Kidney Diseases Grant DK-28305 to K. E. Barrett.
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FOOTNOTES |
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Portions of these studies were presented at the annual meeting of the American Gastroenterological Association (May 2000) and have appeared in abstract form (S. J. Keely and K. E. Barrett, Gastroenterology 118: A871, 2000).
K. E. Barrett is a member of the Biomedical Sciences Ph.D. Program at the University of California, San Diego, School of Medicine.
Address for reprint requests and other correspondence: S. J. Keely, Univ. of California, San Diego, Division of Gastroenterology, 200 W. Arbor Drive, San Diego, CA 92103-8414 (E-mail: skeely{at}ucsd.edu).
The costs of publication of this article were defrayed in part by the payment of page charges. The article must therefore be hereby marked "advertisement" in accordance with 18 U.S.C. Section 1734 solely to indicate this fact.
First published October 3, 2002;10.1152/ajpcell.00144.2002
Received 24 September 2002; accepted in final form 28 September 2002.
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