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Clinic for Gastroenterology, Hepatology and Infectiology, Heinrich-Heine-Universität, Düsseldorf, Germany
1Correspondence: Medizinische Einrichtungen der Heinrich-Heine Universität, Klinik für Gastroenterologie, Moorenstrasse 5, D-40225 Düsseldorf, Germany. E-mail: Freimut.Schliess{at}uni-duesseldorf.de
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ABSTRACT |
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 TOP ABSTRACT INTRODUCTIONMATERIALS AND METHODSRESULTSDISCUSSIONREFERENCES |
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Key Words: MAP kinase · JNK · heat · c-Jun · Hsp70 · MKP-1 · cell volume · arsenite
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INTRODUCTION |
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TOPABSTRACTINTRODUCTIONMATERIALS AND METHODSRESULTSDISCUSSIONREFERENCES |
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).
Whereas cell shrinkage supports a catabolic situation in the liver,
cell swelling exerts growth factor-like effects, for example, by
stimulation of protein- and glycogen synthesis and activation of the
mitogen-activated protein (MAP)2
kinases Erk-1 and Erk-2 (extracellular signal-regulated kinases)
(2
3
4)
. Also, the tolerance of the liver against different
forms of stress is modified by the hepatocellular hydration state. For
example, liver injury induced by reactive oxygen intermediates was
amplified by hyperosmolarity whereas hypo-osmolarity protected against
oxidative damage (5
, 6)
. This was due in part to
anisosmotic modulation of the stress-induced Kupffer cell activation
(6)
. However, there is evidence for a direct role of
parenchymal cell hydration for hepatoprotection: hypo-osmotic liver
perfusion was shown to increase canalicular bile excretion and the
generation of NADPH via stimulation of the pentose phosphate shunt
(6
, 7)
, whereas hyperosmolarity was suggested to create
some oxidative stress by itself in rat liver parenchymal cells
(5)
and H4IIE hepatoma cells (8)
.
The heat shock response can be elicited by a wide range of noxious
conditions such as heat, arsenite, ethanol, infections and heavy metals
and includes alterations in gene expression, MAP kinase activation,
cellular metabolism, and stress tolerance (9
, 10)
.
Heat-induced accumulation of HSP70 (inducible heat shock protein 70)
participates in the development of thermotolerance but also contributes
to cellular protection against many other types of stress by
stabilization of native protein structures (9
, 11)
.
Conflicting data exist with respect to a HSP70 response to osmotic
stress: a hyperosmotic induction of HSP70 was demonstrated in renal
cells (12
13
14
15)
and in 3T3 fibroblasts (16)
.
In HeLa cells, both hypo- and hyperosmotic treatment induced binding
activity of the heat shock transcription factor; however, this was not
accompanied by HSP70 mRNA and protein expression (17)
. In
rat hepatocytes, anisosmotic exposure did not elicit a HSP70 response
(18)
.
A very immediate response to heat is the activation of the c-Jun
amino-terminal kinases (JNKs), which belong to the family of MAP
kinases and play a crucial role in the cellular response to different
forms of environmental stress, including hyperosmolarity and heat shock
(for review, see ref 10
). The JNK pathway was suggested to
confer cellular injury after heat shock and cells defective in the JNK
response to heat acquired thermotolerance (19)
. Recently,
the balance between the activities of extracellular signal-regulated
kinases (Erk-type MAP kinases) and JNKs was reported to be a critical
determinant for cell survival after oxidative challenge (19
, 20)
. Phosphorylation of c-Jun by JNKs leads to AP-1-mediated
induction of c-jun mRNA expression (21)
. MAP
kinase activity can be antagonized by the MAP kinase phosphatase-1
MKP-1, which is an immediate early gene-encoded protein phosphatase and
is induced by growth factors, oxidative stress, or heat shock
(22)
.
An osmotic regulation of the heat shock response in primary rat
hepatocytes was found recently: hyperosmolarity led to a decreased heat
resistance, paralleled by an impaired HSP70 and MKP-1 expression and a
markedly increased JNK response (18)
. Hyperthermia
combined with chemotherapeutic drugs or radiation, respectively, is an
established protocol in tumor treatment (23)
, but little
is known about a potential influence of cell hydration on the response
of tumor cells to hyperthermia. In the present study, therefore,
anisosmotically exposed H4IIE hepatoma cells were chosen as a model to
evaluate the influence of cell hydration on the heat shock response.
The heat resistance of H4IIE cells remained largely unaffected by the
ambient osmolarity, although the heat shock response was osmolarity
dependent at the levels of HSP70, MKP-1, c-jun, and MAP
kinases. Moreover, a heat-induced sensitization to arsenite was
osmoregulated in H4IIE cells.
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MATERIALS AND METHODS |
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TOPABSTRACTINTRODUCTIONMATERIALS AND METHODSRESULTSDISCUSSIONREFERENCES |
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[32P]ATP and
[32P]dCTP were from Amersham (Braunschweig,
Germany). All other chemicals were of the highest quality, available
from commercial sources.
Cell culture and treatment of the cells with anisosmolarity and
heat
H4IIE-C3 rat hepatoma cells (A.T.C.C. CRL 1600) were maintained
in DMEM-F12/5% CO2/5 mM glucose at 37°C, pH
7.4, supplemented with 10% fetal calf serum. When cells had reached
90% confluency, they were washed with Dulbecco's phosphate-buffered
saline and the culture was continued in serum-free medium for an
additional 24 h. Extracellular osmolarity was adjusted by dilution
of the media with the appropriate volume of the respective NaCl-free
medium leading to hypo-osmolarity (205 mosmol/l) or with medium of
elevated NaCl content leading to hyperosmolarity (405 mosmol/l),
respectively. In the normo-osmotic control (305 mosmol/l), the same
volume of normo-osmotic medium was added. For heat shock, cells were
incubated at 43°C
(95%O2/5%CO2) for 1 h. After heat shock, cells were allowed to recover during an additional
culture period at 37°C. During heat shock and the recovery period,
the respective osmotic conditions remained unchanged.
Western blot analysis
At the end of the incubations, the medium was removed and the
cells were immediately lysed at 4°C using 50 mmol/l Tris/HCl buffer
(pH 7.2) containing 150 mmol/l NaCl, 40 mmol/l NaF, 5 mmol/l EDTA, 5
mmol/l EGTA, 1 mmol/l vanadate, 0.5 mmol/l
phenylmethyl-sulfonylfluoride, 0.1% aprotinin, 1% Nonidet-P40, 0.1%
sodium deoxycholate, and 0.1% sodium dodecyl sulfate (SDS). The
homogenized lysates were centrifuged at 20,000 x g at
4°C and the supernatant was added to an identical volume of gel
loading buffer containing 200 mmol/l dithiothreitol (DTT, pH 6.8).
After heating to 95°C for 5 min, the proteins were subjected to
SDS-polyacrylamide gel electrophoresis (PAGE; 50 µg protein/lane;
7.5% gel for HSP70 analysis, 12,5% gel for MKP-1 analysis). After
electrophoresis, gels were equilibrated with transfer buffer (39 mmol/l
glycine, 48 mmol/l Tris/HCl, 0.1% SDS, 20% methanol). Proteins were
transferred to nitrocellulose membranes using a semi-dry transfer
apparatus (Pharmacia, Freiburg, Germany). Blots were blocked in 5%
bovine serum albumin containing TBST (20 mmol/l Tris/HCl pH 7.5, 150
mmol/l NaCl, 0.1% Tween 20) and then incubated for 2 h with the
1:100,000-diluted monoclonal antibody recognizing both HSP70 and HSC70
(constitutive heat shock protein 70)or the 1:1000 dilution of the
polyclonal anti MKP-1 antibody, respectively. After washing with TBST
and incubation with horseradish peroxidase-coupled anti-mouse
immunoglobulin G antibody (Bio-Rad, Munich, Germany) or horseradish
peroxidase-coupled anti-rabbit immunoglobulin G antibody (Amersham,
Braunschweig, Germany), respectively, diluted 1:10,000 at room
temperature for 1 h, the blots were washed three times and
developed using enhanced chemiluminescent detection (Amersham,
Braunschweig, Germany).
Immune complex kinase assays
The immune complex assay was performed according to ref
24
. Briefly, aliquots of cell lysate containing 100 µg
protein were incubated with 1.0 µg of the antibody against Erk-2 or
JNK-1, respectively, for 2 h at 4°C. Immune complexes were
collected by using protein A-Sepharose 4B (Pharmacia), washed three
times with RIPA buffer, four times with kinase buffer (10 mmol/l
Tris/HCl pH 7.4, 150 mmol/l NaCl, 10 mmol/l
MgCl2, 0.5 mmol/l DTT), and incubated with 1
mg/ml MBP (Erk-assay) or GST c-Jun peptide (JNK-assay), 25 µmol/l ATP
and 5 µCi -[32P]ATP for 30 min at 37°C.
The reactions were stopped by adding gel loading buffer and activities
of Erk-2 and JNK-1 were monitored via autoradiography after SDS-PAGE
(12.5% gel).
Nothern blot analysis
Total RNA from H4IIE cells was isolated by using guanidinium
thiocyanate solution as described in ref 25
. RNA samples
(10 µg) were electrophoresed in 0.8% agarose/3% formaldehyde, then
blotted onto Hybond-N nylon membranes with 20xSSC (3 mol/l NaCl, 0.3
mol/l sodium citrate). After brief rinsing with water and cross-linking
(Hoefer UV-cross-linker 500; Hoefer, San Francisco, Calif.), the
membranes were observed under UV light to determine RNA integrity and
location of the 28 S and 18 S rRNA bands. Blots were then subjected to
a 3 h prehybridization at 43°C in 50% de-ionized formamide in
sodium phosphate buffer (0.26 mol/l, pH 7.2) containing 0.25 mol/l
NaCl, 1 mmol/l EDTA, 100 µg/ml salmon sperm DNA, and 7% SDS.
Hybridization was carried out in the same solution with ~
106 cpm/ml
[32P]dCTP-labeled random-primed rat MKP-1
(675 bp cDNA fragment (8)
, the 1.8 kb
EcoRI-Pst-I mouse c-jun c-DNA fragment
(26)
, HSP70, or GAPDH (2.3 kb and 1.0 kb human cDNA
fragments; Clontech, Palo Alto, Calif.), respectively. Membranes were
washed three times in 2xSSC/0.1% SDS for 15 min, twice in sodium
phosphate buffer (25 mmol/l, pH 7.2)/EDTA (1 mmol/l)/0.1% SDS for 10
min, and twice in sodium phosphate buffer (25 mmol/l, pH 7.2)/EDTA (1
mmol/l)/1% SDS for 10 min at 53°C. Blots were then exposed to Kodak
AR X-Omat film at -70°C with intensifying screens.
Determination of cellular heat resistance and arsenite tolerance
Conversion of sodium
3'-[1-(phenylaminocarbonyl)-3,4-tetrazolium]-bis(4-methoxy-6-nitro)
benzene sulfonic acid (XTT) to the formazan by the mitochondrial
succinate-tetrazolium reductase system reflects the metabolic capacity
of the cells and was taken as a measure of heat resistance as described
(19)
. The reagents were purchased from Boehringer
(Mannheim, Germany). H4IIE cells were cultured using the 96-well plate
format (Costar, Cambridge, Mass.) at 1 x
104 cells/well 48 h prior to the treatment.
After treatment of the cells (1 h 43°C or 37°C for control under
the respective osmotic conditions), 50 µl of 5 mg/ml XTT was added to
each well. H4IIE cells were incubated for an additional 24 h
before detection of the optic densities (450 nm) on an enzyme-linked
immunoassay reader (Anthos Labotec Instruments). Lactate dehydrogenase
(LDH) activity was taken as a measure of arsenite tolerance after a
priming heat shock and was determined in the culture medium and cell
lysates as described (27)
. LDH release was expressed as
the percentage of total enzyme activity released into the supernatant
of the H4IIE cells.
Statistics
Results from n independent experiments are
expressed as means ± SE. Results were compared
using the Student's t test; P < 0.05
was considered statistically significant.
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RESULTS |
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A, the HSP70 mRNA transcripts (3.1 and 2.8 kb) were
detectable immediately after heat shock in cells exposed to
hypo-osmolarity and in the normo-osmotic control (305 mosmol/l). A
similar level of HSP70 mRNA was found only after 1–2 h of
recovery under hyperosmotic conditions. Heat-induced
HSP70mRNA levels were maximal after 2 h recovery of hypo-
or normo-osmotically exposed cells. Maximal HSP70 mRNA levels in
hyperosmotically treated cells were expressed after a recovery period
of 3–4 h and were comparable to the maximal expression levels observed
under hypo- and normo-osmotic conditions. A clear signal of both HSP70
mRNA transcripts was also found after 4 h recovery, whereas the
2.8 kb transcript was nearly undetectable at this time point under
hypo- and normo-osmotic conditions. Irrespective of the medium
osmolarity, HSP70 mRNA expression returned toward the basal level after
8 h recovery. The osmosensitivity of the heat-induced HSP70 mRNA
expression was largely mirrored at the level of HSP70 protein
(Fig. 2
); a weak HSP70 expression was found after 2 h recovery and became
maximal after 6–8 h recovery independent of the respective osmolarity.
However, accumulation of HSP70 protein was higher and more sustained
during hyperosmotic exposure compared with the normo- and hypo-osmotic
conditions. Expression of HSP70 persisted during a recovery period of
18 h in cells challenged with hyperosmolarity, whereas HSP70 was
no longer detectable after 12 h of recovery in normo- or
hypo-osmotically maintained cells. Also, the cellular levels of HSC70
protein were increased after heat shock; however, the HSC70 expression
was largely independent of the ambient osmolarity. No HSP70 mRNA- or
protein could be detected when H4IIE cells were challenged with
anisosmolarity or maintained normo-osmotically for up to 24 h in
the absence of heat shock (not shown).
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Osmosensitivity of MAP kinase activation and MKP-1 induction by
heat shock
Hyperosmolarity triggered in H4IIE cells an activation of Erk- and
JNK-type MAP kinases, which was maximal between 80 and 120 min of
hyperosmotic exposure and declined toward basal levels within 4 h,
whereas hypo-osmolarity per se was without effect on Erks
and JNKs in these cells (8
, 28)
. Figure 3
shows the effect of superimposed anisosmolarity and heat shock on Erk-2
and JNK-1 activity. When anisosmotically pretreated (1 h) H4IIE cells
were heat shocked under the respective osmotic condition,
hyperosmolarity markedly amplified the JNK-1 response to heat compared
with the hypo- or normo-osmotically incubated cells. Further,
heat-induced JNK activation under hyperosmotic conditions was more
sustained than the JNK activation triggered by hyperosmolarity
per se (Fig. 3)
. Immediately after the heat treatment, a
pronounced activation of Erk-2 was found in hypo- and normo-osmotically
maintained cells, which declined toward basal levels within a recovery
period of 2 h. Under hyperosmotic conditions the
heat-induced activation of Erk-2 was delayed (2 h recovery) and weak
compared with the JNK-1 activation (Fig. 3)
.
|
The MKP-1 is an immediate early gene-encoded phosphatase that is
responsive to heat and contributes to deactivation of MAP kinases
(29)
. As shown in Fig. 1A
, heat shock induced
MKP-1 mRNA expression in H4IIE cells, which was detectable immediately
after the heat treatment under hypo- and normo-osmotic conditions and
with some delay in cells exposed to hyperosmolarity. In
hypo-osmotically challenged cells and in the normo-osmotic control,
heat-induced MKP-1 mRNA accumulation was maximal after 2 h of
recovery and declined to basal niveau within another hour at 37°C.
Under hyperosmotic conditions the heat-induced MKP-1 mRNA accumulation
was more pronounced, peaked after a 3 h recovery period, and then
declined toward basal niveau. According to the observed mRNA levels,
the maximal heat-induced MKP-1 protein expression was delayed but most
pronounced under hyperosmotic conditions (Fig. 4
, 2 h recovery), whereas heat induced a much weaker MKP-1 protein
expression under hypo-osmotic conditions. In the normo-osmotic control,
MKP-1 occurrence after heat shock was maximal after 1 h recovery
(Fig. 4
, large arrow). Possibly MKP-1 contributes to
down-regulation/suppression of heat-induced MAP kinase activation, but
evidence for a simple reciprocal relation between MKP-1 expression
levels and MAP kinase activities cannot be derived from the data
(compare Figs. 3
and 4
). Heat shock transiently suppressed the
expression of a protein recognized by the antibody against MKP-1, which
runs in SDS gel electrophoresis above the MKP-1 protein (Fig. 4
, small
arrow). The expression of this protein was restored after 4 h
recovery; it was not cross-reactive with an antibody raised against
MKP-2 (not shown). A potential relationship of this protein to the
MKP-1 was not further investigated in this study. Hyperosmolarity
per se also triggered MKP-1 mRNA expression in H4IIE cells
(Fig. 1B
) (8)
. However, hyperosmotic MKP-1 mRNA
accumulation was weak compared with the induction in response to heat
(Fig. 1A, B
) and no MKP-1 protein was recognized in Western
blot in cells challenged with hyperosmolarity at 37°C. (Fig. 4)
.
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The osmosensitivity of the heat-induced c-jun mRNA
expression is similar to that found for the MKP-1 mRNA (Fig. 1)
.
Specific phosphorylation of c-Jun by the JNKs was reported to mediate
the induction of its own mRNA expression (21)
. Thus, the
pronounced JNK activation by heat shock in hyperosmotically maintained
cells (Fig. 3)
may lead to the markedly increased levels of c-jun mRNA
under this condition (Fig. 1A
).
Osmotic modulation of the stress tolerance of H4IIE cells
The effect of anisosmolarity (205 and 405 mosmol/l) and heat shock
(1 h 43°C) on the capacity of H4IIE cells to convert XTT to the
formazan was taken as a measure of cellular heat resistance.
Figure 5
shows that hypo-osmotic treatment per se was without effect
on sodium XTT conversion, whereas hyperosmolarity led to a slight but
significant decrease by ~10% compared with the control
(normo-osmolarity, 37°C). Heat shock decreased XTT conversion by
~10% under normo- and hypo-osmotic conditions. In hyperosmotically
exposed cells, heat shock did not significantly decrease XTT conversion
below the levels reached under hyperosmotic conditions at 37°C. The
data suggest that the heat treatment only slightly impaired H4IIE cell
metabolism and that hyperosmolarity did not lead to a significant
amplification of heat-induced injury on cellular metabolism.
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In another series of experiments, a potential role of H4IIE cell
hydration for the development of arsenite tolerance/sensitization after
a priming heat shock was examined. Therefore, H4IIE cells were
pretreated by anisosmotic incubation (1 h) and challenged with a
priming heat shock (1 h, 43°C) without change of the respective
osmotic condition. An additional time period of 18 h at 37°C
allowed the cells to recover from the priming heat shock, which leads
to HSP70 accumulation under hyperosmotic, but not hypo- or
normo-osmotic, conditions (see Fig. 2
). After the recovery period,
cells were exposed to sodium arsenite (1 h, 1 mmol/l). H4IIE cell
viability was determined by measuring the LDH release in response to
the arsenite treatment. In the absence of a priming heat shock,
arsenite-induced LDH release from hyperosmotically exposed cells was
amplified about threefold compared to the hypo- and normo-osmotic
conditions (Fig. 6
). The priming heat shock markedly sensitized the cells to arsenite
under normo-osmotic conditions (LDH release ~60%), whereas no
heat-induced sensitization was observed in hyperosmotically exposed
cells. The resistance to arsenite was improved under hyperosmotic
conditions compared with the normo-osmotic control (Fig. 6)
. Only a
small and not significant sensitization to arsenite was found in
heat-primed, hypo-osmotically maintained cells (Fig. 6)
. Thus, the
sustained HSP70 expression in hyperosmotically dehydrated cells may
counterbalance a heat-induced sensitization, which was found under
normo-osmotic conditions in the absence of HSP70. On the other hand, an
HSP70-independent protection mechanism is suggested for the
hypo-osmotic condition.
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DISCUSSION |
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TOPABSTRACTINTRODUCTIONMATERIALS AND METHODSRESULTSDISCUSSIONREFERENCES |
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Hyperosmolarity per se induced the expression of mRNAs for
MKP-1 and c-jun (Fig. 1B
) (8
, 30)
, which was
weak compared with the heat shock response (Fig. 1)
. Hyperosmolarity,
however, was unable to stimulate a HSP70 expression in H4IIE cells (not
shown). In renal cells (12
, 13)
, an elevated intracellular
ionic strength due to high salt exposure (500–900 mosmol/l) led to a
long-lasting expression of HSP70 (15)
. Proteotoxicity of a
similar extent may arise in H4IIE cells from the synergistic action of
milder hyperosmolarity (405 mosmol/l) and heat shock, leading to a
likewise sustained HSP70 expression (Fig. 2)
. The persistent
heat-induced expression of HSP70 under hyperosmotic conditions, even
after the return of HSP70 mRNA levels to basal niveau (compare Fig. 1A
and Fig. 2
), suggests a hyperosmolarity-induced
prolongation of the HSP70 half-life as an underlying regulatory
mechanism. In line with this, HSP70 expression in heat-shocked and
hyperosmolarity-exposed cells persisted when 6 h after recovery
from heat shock, the incubation was continued for another
12 h in the presence of the protein synthesis inhibitor
cycloheximide (1 µg/ml; data not shown). A marked prolongation of
HSP70 half-life was also induced in Drosophila cells by
continued heat shock, possibly due to an altered subcellular
distribution of HSP70 (31)
. Whether continued hyperosmotic
exposure prolongs HSP70 half-life by a similar mechanism is currently
unknown.
Whereas hypo-osmotic exposure of H4IIE cells was without effect on XTT
conversion, hyperosmolarity per se led to a slightly
reduced XTT conversion (Fig. 5)
. However, the effect of heat shock on
XTT conversion was not significantly amplified under hyperosmotic
conditions (P>0.05; Fig. 5
). This may be attributable
to the delay of gene expression in response to heat, which helps
initially to avoid a potentially harmful energy drain due to an
immediate synthesis of stress proteins, thereby preventing energy
limitations for other vital metabolic processes (32)
. MAP
kinase activation in response to heat preceded the induction of HSP70,
MKP-1 and c-jun (compare Fig. 1
A and Fig. 3
). Thus, it
is conceivable that MAP kinases participate in the determination of
cellular impairment during and immediately after heat shock. Especially
the pronounced JNK-1 activation under hyperosmotic conditions is a
candidate for mediating the delayed onset of heat-induced gene
expression under this condition. Once induced, however, HSP70
accumulation in H4IIE cells after heat shock was most pronounced under
the hyperosmotic condition (Fig. 2)
, probably due to the
superimposition of two different types of stress. On the other hand,
HSP70 induction by heat was absent in hyperosmotically exposed rat
hepatocytes, but was restored by the hepatoprotective osmolyte taurine
(18)
or when long-term adaption (20 h) to hyperosmolarity
was allowed prior to the heat shock (F. Schliess and D.
Häussinger, unpublished results). This suggests that, to a high
degree, cellular adaption to osmotic stress plays a role in the osmotic
control of heat-induced HSP70 expression. An altered response of
signaling cascades to anisosmolarity in tumor-derived cells compared
with the respective nonmalignant cells is well established (30
, 33)
, and may contribute to the differential sensitivity of
cellular heat resistance to hydration changes.
Arsenite inactivates various enzymes and impairs proper folding of
peptides during de novo synthesis by binding to vicinal
cysteine residues; these effects were diminished under reducing
conditions (34)
. In H4IIE cells, hyperosmolarity by itself
was suggested to generate reactive oxygen intermediates
(8)
, which may explain the increased arsenite toxicity in
hyperosmotically exposed cells (Fig. 6
, light bars). Heat-induced
expression of HSP70 was suggested to confer tolerance to many types of
stress by stabilizing the native protein structures of mature proteins
and supporting a proper folding of nascent polypeptides in a
cotranslational fashion (9)
. In H4IIE cells, continued
HSP70 expression induced by heat shock under hyperosmotic conditions
(Fig. 2)
was paralleled by inhibition of a heat-induced sensitization
to arsenite (Fig. 6)
. Under normo-osmotic conditions, the HSP70
response to heat was transient (Fig. 2)
; after termination of HSP70
expression, cells were highly sensitized against arsenite (Fig. 6)
.
Arsenite-induced protein denaturation may have been ameliorated by
preexisting HSP70. H4IIE cell protection under hypo-osmotic conditions,
however, apparently occurred independent of HSP70 (Figs. 2
and 6)
. Such
protection could involve the recently reported hypo-osmotic stimulation
of the pentose phosphate pathway, which favors NADPH provision for
glutathione reductase, thereby providing tolerance to oxidative stress
(5)
. Thus, it could be speculated that hypo-osmolarity
protects proteins against arsenite by creating reducing conditions.
Together, a high degree of resistance to heat shock was found in H4IIE
cells irrespective of the ambient osmolarity. At the level of selected
molecular components, however, the heat shock response was strongly
modulated by the ambient osmolarity. This may reflect adaptive
processes in H4IIE cells leading to an osmolarity-independent heat
resistance. Despite the H4IIE cell resistance to a priming heat shock,
the response of heat-sensitized cells toward arsenite was
osmodependent. Keeping in mind that the cellular hydration state not
only is determined by the extracellular osmolarity, but also underlies
hormonal and nutritional regulation (35)
, a physiological
role of cellular hydration in modulating resistance against
chemotherapeutic drugs under conditions of hyperthermia appears
evident.
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FOOTNOTES |
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Received for publication November 6, 1998. Revised for publication March 19, 1999.
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REFERENCES |
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TOPABSTRACTINTRODUCTIONMATERIALS AND METHODSRESULTSDISCUSSIONREFERENCES |
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This article has been cited by other articles:
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  W. H. Miller Jr., H. M. Schipper, J. S. Lee, J. Singer, and S. Waxman Mechanisms of Action of Arsenic Trioxide Cancer Res., July 15, 2002; 62(14): 3893 - 3903. [Abstract] [Full Text] [PDF]
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