MINIREVIEW
Osmosensing and signaling in the regulation
of mammalian cell function
Freimut Schliess, Roland Reinehr and Dieter Ha
¨
ussinger
Clinic for Gastroenterology, Hepatology and Infectiology, Heinrich-Heine-University, Du
¨
sseldorf, Germany
Introduction
Sudden exposure of cells to hypo- or hyperosmotic
solutions induces a rapid osmotic swelling or shrink-
age, respectively. Extensive swelling or shrinkage is
counteracted by induction of a regulatory volume
decrease (RVD) or increase, respectively [1–3]. Most
hypoosmotically swollen cells perform RVD by a
release of inorganic ions, including K
+
,Na
+
,Cl
–
, and
HCO
3
–
and organic osmolytes (e.g. taurine, betaine).
Hyperosmotic regulatory volume increase (RVI) at a
short-term time scale is performed by activation of
electrolyte uptake (e.g. via Na
+

⁄ 2Cl
–
sym-
port and the Na
+
⁄ K
+
-ATPase [5].
In the early 1990s, it was recognized, that cell vol-
ume changes trigger signals involved in the regulation
of metabolism, gene expression and the susceptibility
to different kinds of stress [6]. For example, the inhibi-
tion of autophagic proteolysis by insulin, glutamine
and ethanol in the perfused liver critically depends on
the degree of hepatocyte swelling induced by these
stimuli and can be mimicked by hypoosmotic swelling
Keywords
apoptosis; bile acids; CD95; cell volume;
epidermal growth factor; insulin; integrins;
osmolytes; oxidative stress; proliferation
Correspondence
F. Schliess, Heinrich-Heine-Universita
¨
t,
Universita
¨
tsklinikum, Klinik fu
¨
r
Gastroenterologie und Infektiologie,

catabolism and insulin resistance and sensitizes cells to
apoptotic stimuli.
The influence of cell volume on cell function requires
structures that register volume changes (‘osmosensing’)
and trigger signaling pathways towards effector sites
(‘osmosignaling’). Anisoosmotically exposed cells and
tissues were frequently used as a model in order to
study osmosensing and osmosignaling. By this
approach, the activation of signaling pathways by cell
volume changes could be linked to specific functional
outcomes [8]. Here, we summarize some recent pro-
gress concerning the understanding of how ‘osmosen-
sing’ and ‘osmosignaling’ integrate into the overall
context of signal transduction, which is activated by
growth factors, substrates or (pro)-apoptotic stimuli in
mammalian cells with some focus on the hepatocyte. A
more general treatize on osmosensing and osmosignal-
ing is provided elsewhere [9].
Osmosensing in mammalian cells
The investigation of osmosensing processes and struc-
tures in mammalian cells considers, among others,
macromolecular crowding, stretch-activated ion chan-
nels, cholesterol-enriched microdomains of the plasma
membrane (caveolae), intracellular organelles, ligand-
independent activation of growth factor- and cytokine
receptors and autocrine stimulation of signal transduc-
tion by release of mediators such as ATP [10,11].
Recent studies have identified the integrin system as
one major sensor of hepatocyte swelling [12–14].
Integrin-inhibitory peptides exhibiting an arginine-gly-

+
-ATPase, probably
driven by an increase in intracellular Cl
–
concentration
due to osmotic water loss and Cl
–
accumulation in the
course of a RVI, respectively [15]. The endosomal
compartment acidified by hyperosmolarity colocalized
with the acidic sphingomyelinase [15]. Hyperosmolarity
in hepatocytes triggers a rapid production of reactive
oxygen species (ROS), which critically depends on ser-
ine phosphorylation of the NADPH oxidase regulatory
subunit p47
phox
, which again depends on a acid sphin-
gomyelinase-catalyzed ceramide production and subse-
quent activation of the PKCf [15]. Bafilomycin A1 (an
inhibitor of vacuolar-type H
+
-ATPases) and the anion
channel blocker 4,4¢-diisothiocyanostilbene-2,2¢-disulf-
onic acid disodium salt not only prevent endosomal
acidification by hyperosmolarity, but also block the
hyperosmotic increase of ceramide, p47
phox
phosphory-
lation and ROS [15]. The findings localize endosomal
acidification most upstream in the signaling cascade

⁄ 2Cl
–
cotransport and glutamine uptake [17].
Likewise, hepatocyte swelling due to the activation of
system A-type amino acid transporters was observed
Osmosensing and signaling in mammalian cell function F. Schliess et al.
5800 FEBS Journal 274 (2007) 5799–5803 ª 2007 The Authors Journal compilation ª 2007 FEBS
in vivo following partial hepatectomy, and inhibition of
cell swelling antagonized liver regeneration [21]. Hypo-
osmolarity in many cell types activates the MAPKs
Erk1 ⁄ Erk2 and the PI 3-kinase [8], which play a major
role in mitogenic signaling. Hypoosmotic exposure of
HepG2 cells potentiates proliferation by a PI 3-kinase-
meditated activation of the transcription factor activa-
tor protein AP-1 [22], corroborating a critical role of
cell swelling for cell cycle progression. Consistently,
the cell volume was increased in 3T3 cells expressing
oncogenic Ha-ras [23] and cell hyperhydration has
been linked to tumor growth [24].
A volume decrease resulting from osmolyte release
through specific transport proteins at the beginning of
apoptosis (apoptotic volume decrease) is an early pre-
requisite for the execution of apoptotic programs [25].
Signaling mechanisms upstream of apoptotic volume
decrease depend on the cell type and stimulus under
investigation and have been discussed previously [26–
28]. The contribution of apoptotic volume decrease to
apoptotic signal transduction is currently not well
understood. Using hyperosmotically treated cells as a
model, it was shown that efficient volume regulation

tors and the availability of antioxidants and osmolytes.
In rat hepatocytes, mild hyperosmolarity (405
mosmolÆL
)1
) activates the CD95 system, which local-
izes downstream of the ROS production triggered by
endosomal acidification mentioned above [15,38]. Hyp-
erosmotic CD95 activation includes trafficking of the
CD95 from inside the hepatocyte to the plasma mem-
brane [39], which depends on a ROS-mediated tyrosine
phosphorylation of the epidermal growth factor
(EGF)-receptor, the association of CD95 with the
EGF-receptor, and phosphorylation of CD95 on
Tyr232 and Tyr291 by the EGF-receptor tyrosine
kinase activity [15]. Although the appearance of CD95
at the plasma membrane was associated with death
inducing signaling complex formation and activation
of caspases 3 and 8, mild hyperosmolarity was not suf-
ficient to induce hepatocyte apoptosis [39], suggesting
that apoptotic signals under this condition are counter-
balanced by yet unknown survival signals. However,
more severe hyperosmolarity (‡ 505 mosmolÆL
)1
) shifts
the balance towards hepatocyte apoptosis [15].
Like hyperosmolarity, CD95 ligand in hepatocytes
via generation of ROS induced EGF-receptor tyrosine
phosphorylation, CD95 ⁄ EGF-receptor association,
CD95 tyrosine phosphorylation, trafficking of the
CD95 to the plasma membrane surface, and death

survival and death signals, thereby promoting execu-
tion of the apoptotic program.
Concluding remarks
It is well acknowledged that cell volume fluctuations
release signals of (patho)physiological relevance. The
understanding of how cell swelling integrates into the
cell cycle machinery and how cell shrinkage sensitizes
cells to apoptotic stimuli requires further scientific
effort. Cell hydration may markedly affect the action
of drugs. For example cell hydration changes may
switch the outcome of proteasome inhibitors from a
nontoxic or even protective one to injury and apopto-
sis [40]. Although routine monitoring of cell hydration
in patients would provide valuable information in clin-
ical medicine, this is currently limited by methodologi-
cal difficulties.
Acknowledgements
Our own studies were supported by Deutsche Fors-
chungsgemeinschaft through Sonderforschungsbereich
575 ‘Experimentelle Hepatologie’ (Du
¨
sseldorf).
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