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Sympathetic nervous system regulation of liver repair.

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THE ANATOMICAL RECORD PART A 280A:874 – 883 (2004)
Sympathetic Nervous System
Regulation of Liver Repair
JUDE A. OBEN AND ANNA MAE DIEHL*
Department of Medicine, Johns Hopkins University, Baltimore, Maryland
ABSTRACT
This chapter reviews recent evidence that the sympathetic nervous system (SNS) regulates liver repair by modulating the phenotypes of hepatic stellate cells (HSCs), the liver’s
principal fibrogenic cells, and hepatic epithelial progenitors, i.e., oval cells. SNS nerve fibers
touch HSCs and these cells express adrenoceptors, suggesting that HSCs may be targets for
SNS neurotransmitters. HSCs also contain catecholamine biosynthetic enzymes, release
norepinephrine (NE), and are growth-inhibited by adrenoceptor antagonists. In addition,
HSCs from mice with reduced levels of NE grow poorly in culture and exhibit inhibited
activation during liver injury. Finally, growth and injury-related fibrogenic responses are
rescued by adrenoceptor agonists. Thus, certain SNS inhibitors (SNSIs) protect experimental
animals from cirrhosis. Conversely, SNSIs enhance the hepatic accumulation of oval cells
(OCs) in injured livers. This response is associated with improved liver injury. Because SNSIs
do not affect the expression of cytokines, growth factors, or growth factor receptors that are
known to regulate OCs, and OCs express adrenoceptors, it is conceivable that catecholamines
influence OCs by direct interaction with OC adrenoceptors. Given evidence that the SNS
regulates the viability and activation of HSCs and OCs differentially, SNSIs may be novel
therapies to improve the repair of damaged livers. © 2004 Wiley-Liss, Inc.
Key words: hepatic oval cells; hepatic stellate cells; cirrhosis; liver regeneration
Hepatic stellate cells (HSCs) and hepatic oval cells
(OCs) are important cell types in liver repair. HSCs, the
liver’s principal fibrogenic cells, are activated by liver
injury of any cause to move from a quiescent to an activated myofibroblastic phenotype. This myofibroblastic
phenotype is proliferative, expresses ␣-smooth muscle actin, and synthesizes fibrogenic matrix proteins that accumulate during cirrhosis (Friedman, 2000). Activated myofibroblastic HSCs are also contractile and therefore may
contribute to the pathogenesis of portal hypertension
(Reynaert et al., 2002). Conversely, OCs are liver resident
progenitor cells that help to regenerate the hepatic epithelial compartment. OC populations are activated when
mature hepatocytes reach a critically low number, such as
after severe liver injury, or when mature hepatocytes are
prevented from dividing by hepatotoxic drugs (Evarts et
al., 1996). Therefore, OCs promote the regeneration of
damaged livers. Emerging evidence suggests that the
sympathetic nervous system (SNS) effects the function of
both HSCs and OCs and therefore directly regulates liver
repair.
SNS REGULATION OF HSC:
HISTORICAL EVIDENCE
HSCs Express Neuronal Markers
HSCs may function as hepatic neuroglia cells, receiving
and integrating commands from the CNS, because SNS
©
2004 WILEY-LISS, INC.
fibers abut HSCs (Moghimzadeh et al., 1983; Ohata, 1984;
Bioulac-Sage et al., 1990), which possess functional ␣-adrenoceptors (Athari et al., 1994). That HSC may be neural
crest-derived hepatic neuroglial cells is further supported
by their expression of classical neuroglial proteins, such as
glial acidic fibrillary protein (GFAP), nestin, neural cell
adhesion molecule (NCAM), synatophysin, and neurotrophins, as well as by evidence that they contain synaptic
vesicles (Buniatian et al., 1996; Cassiman et al., 1999,
2001; Niki et al., 1999).
SNS Activity Correlates With Liver Fibrosis
A growing body of indirect evidence supports the possibility that SNS overactivity may be involved in the aetiopathogenesis and progression of cirrhosis. First, the severity of carbon tetrachloride (CCL4)-induced fibrosis is
greater in the spontaneously hypertensive rat, which has
a hyperactive SNS compared with normotensive control
*Correspondence to: Anna Mae Diehl, Duke University Medical
Center, GSRB-1, 595 La Salle Street, Durham, NC 27710. Fax:
919-684-4183. E-mail: diehl004@mc.duke.edu
Received 11 March 2004; Accepted 20 June 2004
DOI 10.1002/ar.a.20081
Published online 3 September 2004 in Wiley InterScience
(www.interscience.wiley.com).
SYMPATHETIC NERVOUS SYSTEM REGULATION
Wistar-Kyoto rats (Hsu, 1992). Second, in livers pretreated ex vivo with D-galactosamine, the degree of injury
is markedly enhanced by electrical stimulation of the attached sympathetic nerves. This effect is mimicked by
infusions of low doses of norepinephrine (NE) that do not
compromise perfusate flow to the liver (Iwai and Shimazu,
1996). Third, animals with low catecholamine levels and
low SNS tone (e.g., leptin-deficient ob/ob mice) (Young and
Landsberg, 1983; Liang and Cincotta, 2001) are resistant
to liver fibrosis (Honda et al., 2002). Together, these findings suggest that activation of sympathetic nerves increases circulating cathecholamines that exacerbate liver
injury. Conversely, chemical sympathectomy with 6-hydroxydopamine, or ␣1-adrenoceptor antagonism with prazosin, inhibits CCL4-induced liver fibrosis in rats (Dubuisson et al., 2002). Evidence that parasympathetic
cholinergic agonists modulate local matrix production to
regulate wound healing in other epithelial tissues (Heeschen et al., 2002; Jacobi et al., 2002) also supports the
concept that the autonomic nervous system regulates liver
injury and repair. Finally, activation of the SNS with
increased levels of NE and its cotransmitter neuropeptide
Y is well documented in patients with cirrhosis (Henriksen et al., 1984; Esler et al., 1992), particularly when the
hepatorenal syndrome develops (Uriz et al., 2002). However, whether altered SNS activity is primary to the
pathogenesis of cirrhosis or a secondary compensation for
the circulatory disturbances that accompany cirrhosis is
unknown.
SNS REGULATION OF LIVER FIBROSIS:
NEW EVIDENCE FOR DIRECT
SNS-HSC INTERACTIONS
Despite the cited indirect evidence suggesting that the
SNS regulates the development of fibrosis, until recently
there was no proof that neurotransmitters regulated liver
fibrogenesis directly or what cell types might be targeted.
Over the last 2 years, our group has been systematically
investigating interactions between the SNS and HSCs
(Oben et al., 2003d, e, f, 2004). Our overarching hypothesis
is that HSCs are hepatic neuroglia that are regulated
directly by the SNS and that provide a local source of
cathecholamines. We evaluated our hypothesis by addressing the following questions (Oben et al., 2003f, 2004).
Do HSCs contain NE synthesizing enzymes? Do HSCs
release NE? And do HSC change function in response to
exogenous or endogenous NE? The results of these studies
are summarized subsequently.
HSCs Synthesize and Release NE and Other
Neurotransmitters
The key enzymes in the synthesis of NE are tyrosine
hydroxylase (which converts dihydroxyphenylalanine to
dopamine) and dopamine-␤-hydroxylase (which converts
dopamine to NE). We evaluated the expression of these
catecholamine biosynthetic enzymes in primary HSCs cultured from normal mice. Western blot analysis of HSC cell
protein confirmed that HSCs express both dopamine-␤hydroxylase (Dbh) and tyrosine hydroxylase. Moreover,
high-pressure liquid chromatography analysis of HSCconditioned medium showed that normal HSCs also release NE. In contrast, NE was not detected in conditioned
medium from Dbh⫺/⫺ HSCs that are not able to synthesize
NE because of a targeted deletion of the Dbh gene. In
875
Fig. 1. HSCs express multiple adrenoceptor subtypes. RNA obtained by pooling HSCs from six normal mice was analyzed by RT-PCR.
First lane, DNA ladder (500 –200 bp, arrowed). Each subsequent pair of
lanes is a replicate analysis of adrenoceptor genes. The 18S band (324
bp) serves as a control.
addition to NE, normal murine HSC lysates contain DA,
serotonin (5-hydroxytryptamine; 5-HT), cathecholamine
metabolites (dihydroxyphenylacetic acid; DOPAC), homovanillic acid (HVA), and the 5-HT metabolite, 5-hydroxyindoleacetic acid (5-HIAA), demonstrating that
HSCs can function as neuroglia to provide catecholamines
locally within injured livers (Oben et al., 2004).
HSCs Express Adrenoceptors
Although HSCs were known to express ␣1-adrenoceptors (Athari et al., 1994), which ␣-adrenoceptors subtypes
are expressed, or if HSCs express ␤-adrenoceptors, was
not known until our recent studies (Oben et al., 2004).
Using RT-PCR analysis, we demonstrated that primary
HSCs express ␣1B-, ␣1D-, ␤1-, and ␤2-adrenoceptor mRNAs
(Fig. 1). A similar expression profile was noted with Western blot analysis.
NE Regulates HSC Growth in Culture
To determine if endogenous NE is important for HSC
growth, we cultured normal HSCs in the presence and
absence of an ␣1-adrenoceptor antagonist, prazosin (PRZ;
10 ␮M), or a ␤-adrenoceptor antagonist, propranolol (PRL;
10 ␮M), and assessed their growth (Mosmann, 1983; Isobe
et al., 1999; Frank et al., 2000; Matsuoka et al., 2000;
Saxena et al., 2002). PRZ and PRL each reduced HSC
numbers by ⬃ 20%, and the combination of PRZ ⫹ PRL
decreased HSC growth by ⬃ 50%, showing that the
growth-inhibitory actions of the ␣- and ␤-adrenoceptor
antagonists are additive. Studies of HSCs cultured from
Dbh⫺/⫺ (NE-deficient) and Dbh⫹/⫺ (control) mice prove
that NE is an autocrine growth factor for HSCs. HSCs
from Dbh⫹/⫺ mice proliferate to become nearly confluent
by 4 days in culture, and their proliferative activity is
inhibited significantly by PRZ. Proliferative activity is
also significantly reduced in HSC from Dbh⫺/⫺ mice (Fig.
2), while addition of exogenous NE rescues their growth.
Together, these findings confirm the importance of NE for
HSC growth in culture (Oben et al., 2004). Moreover, NE
increases the growth of HSCs in a dose-dependent man-
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OBEN AND DIEHL
Fig. 2. HSCs that are genetically incapable of producing NE grow
poorly in culture and exogenous NE rescues proliferative activity. Representative photomicrographs of pooled HSCs isolated from six Dbh⫹/⫺
mice and cultured without (a) or with (b) PRZ (10 ␮M) for 4 days and
4-day-old cultures of HSCs pooled from six control Dbh⫺/⫺ mice (c).
HSCs from six additional Dbh⫺/⫺ mice were cultured in control medium
or medium ⫹ NE (10 ␮M) for 4 days and HSC numbers were quantified
(d). Asterisk, P ⬍ 0.05 vs. control.
ner. Neuropeptide Y, an SNS cotransmitter neuropeptide
that is released with NE from sympathetic nerve terminals, also promotes HSC growth (Oben et al., 2003f).
During culture, HSCs normally become activated and
proliferate at a greater rate than they die. Therefore,
increases in proliferative activity normally drive HSC
growth in culture. To determine to what extent, if any, the
NE-related differences in cell number might also reflect
differences in apoptotic activity, HSCs were harvested,
incubated with annexin V, and analyzed by flow cytometry. After 1 day in culture, slightly greater numbers of
apoptotic HSCs were detected in cultures from Dbh⫺/⫺
mice compared to Dbh⫹/⫺ mice. Thus, the antiapoptotic
effects of NE may help explain why endogenous NE is
required for optimal HSC growth. In any case, our data
show that cultured HSCs express NE synthesizing enzymes, release NE, and grow in response both to exogenous and endogenous NE.
ob/ob mice known to have low levels of NE (Knehans and
Romsos, 1982; Young and Landsberg, 1983), but which are
resistant to fibrosis, despite clear evidence of chronic liver
injury (Ikejima et al., 2001; Honda et al., 2002; Leclercq et
al., 2002; Oben et al., 2003c). Hepatic expression of GFAP
was then analyzed as a readout of quiescent and activated
HSCs (Cassiman et al., 2002a). We found that control
ob/ob mice have significantly fewer GFAP⫹ HSCs than
their lean littermates and NE replacement markedly
stimulates the in vivo proliferation of HSCs, such that the
numbers of GFAP⫹ HSCs in ob/ob mice approach that
seen in lean controls (Oben et al., 2004).
Because leptin deficiency might have confounded the
effects of NE in ob/ob mice, we next studied Dbh⫺/⫺ mice,
which also have reduced NE but are not leptin-deficient
(Thomas and Palmiter, 1997a, 1997b). Dbh⫺/⫺ mice and
Dbh⫹/⫺ mice were fed an antioxidant-depleted hepatotoxic
and fibrogenic diet (Leclercq et al., 2002). After 4 weeks of
treatment, control Dbh⫹/⫺ mice exhibit a striking accumulation of HSCs that express ASMA, an accepted
marker of HSC activation (Fig. 3a and c). In contrast,
ASMA⫹ HSCs could not be demonstrated in Dbh⫺/⫺ mice
SNS Regulates HSCs in Intact Mice
To study the effects of NE on HSC function in intact
animals, NE was infused chronically into leptin-deficient
SYMPATHETIC NERVOUS SYSTEM REGULATION
877
Fig. 3. HSC activation is reduced in NE-deficient Dbh⫺/⫺ mice.
Twelve Dbh⫺/⫺ and six of their control Dbh⫹/⫺ littermates were fed
methionine-choline-deficient (MCD) diets. Half of the Dbh⫺/⫺ mice were
also infused with isoprenaline (ISO) for 4 weeks. a: Photomicrograph
from representative Dbh⫹/⫺ mice. Arrows indicate typical ASMA⫹ HSCs
stained brown. b: Photomicrograph from typical Dbh⫺/⫺ mice. c: ␣
smooth muscle actin (ASMA)⫹ sinusoidal cells were counted in five
randomly selected fields/liver section from each mouse. Mean ⫾ SD
results of one experiment are graphed. Asterisk, P ⬍ 0.05 for Dbh⫺/⫺ vs.
Dbh⫹/⫺ control; number sign, P ⬍ 0.05 for Dbh⫺/⫺ ⫹ ISO vs. Dbh⫺/⫺
control. Identical results were obtained in a second experiment that
studied 12 additional mice (4 mice/group). [Color figure can be viewed in
the online issue, which is available at www.interscience.wiley.com].
(Fig. 3b and c), while blood vessel walls in these mice
clearly express ASMA⫹, arguing against this effect being
a staining artifact (Fig. 3a). Moreover, compared to
Dbh⫹/⫺ controls, Dbh⫺/⫺ mice also exhibit significantly
less hepatic expression of collagen and TGF-␤1, two other
indicators of HSC activation (Fig. 4). Ribonuclease protection analysis also demonstrates a small (⬃ 30 – 40%) but
statistically significant (P ⬍ 0.05) reduction in the induction of tissue inhibitor of metalloproteinase (TIMP)-2
transcripts in the Dbh⫺/⫺ group.
To verify that it was reduced adrenergic activity that
prevented HSC activation, we implanted osmotic
minipumps containing vehicle or isoprenaline (ISO), a
␤-adrenoceptor agonist, into Dbh⫺/⫺ mice and repeated
the feeding experiment. Infusion of ISO rescues HSC activation in Dbh⫺/⫺ mice and returned numbers of ASMA⫹
HSCs to levels exhibited by Dbh⫹/⫺ mice also fed the
hepatotoxic diet (Fig. 3c). ISO infusion similarly normalizes induction of TGF-␤ in Dbh⫺/⫺ mice (Fig. 4b). In parallel experiments, we examined the effect of NE, an ␣-adrenoceptor agonist, on HSC activation in ob/ob mice.
Compared to control ob/ob mice, ob/ob mice treated chronically with NE minipumps have significantly increased
liver expression of TGF-␤1, collagen mRNA (Oben et al.,
2004), and histological evidence of fibrosis (Oben et al.,
2003c). Therefore, both ␣-predominant NE and ␤-predominant ISO adrenoceptor agonists affect HSC activation in
vivo. Despite this increase in HSC activation, alanine
aminotransferase (ALT) values in NE-treated ob/ob mice
were lower than their littermate controls. As such, NErelated increases in fibrogenesis are not easily attributed
to NE exacerbation of liver injury (Oben et al., 2003c).
These studies then confirm that HSCs express key
enzymes for cathecholamine biosynthesis, actually produce NE and other cathecholamines, and that HSCs are
directly regulated by the SNS both in vitro and in whole
animals. HSCs appear to use cathecholamines to autoregulate their growth, because increases in HSC number are significantly attenuated by culturing normal
HSCs with ␣- or ␤-adrenoceptor antagonists. HSCs from
NE-deficient animals also grow poorly in culture. In
vivo activation of HSCs, as judged by the expression of
ASMA, TGF-␤ and collagen along with histological evidence of fibrosis, is markedly reduced in NE-deficient
animals.
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OBEN AND DIEHL
likely that NE interacts with other peptides to modulate
HSC function. Indeed, our studies identify at least three
other factors, acetylcholine (Ach), neuropeptide Y
(NPY), and leptin, that may modulate SNS-HSC interactions to influence liver fibrosis.
Acetylcholine
Like NE, the parasympathetic neurotransmitter Ach
appears to function extraneuronally (Heeschen et al.,
2001, 2002; Jacobi et al., 2002). In keeping with this
concept, we demonstrated that Ach promotes proliferation
and induces collagen gene expression in activated HSCs
(Oben et al., 2003d). Whether HSCs also synthesize and
release Ach is unknown. More studies are also needed to
elucidate the role of the parasympathetic nervous system
in HSC biology.
Neuropeptide Y
Fig. 4. NE regulates hepatic expression of collagen and TGF-␤. Liver
RNA was isolated from six Dbh⫹/⫺ mice, six Dbh⫺/⫺ mice, and six of the
Dbh⫺/⫺ mice that were infused with ISO. All mice had been fed MCD diet
for 4 weeks. Hepatic expressions of collagen1-␣-1 (a) and TGF-␤ (b)
were evaluated by ribonuclease protection assay (20 ␮g RNA/assay). A
representative phosphoimage displays individual data from three mice/
group from the first of two RPA assay. Normalized mean (SD) collagen
(n ⫽ 12; 6 mice/group) and TGF-␤1 gene expression from all 18 mice (6
mice/group) is graphed. Asterisk, P ⬍ 0.05 Dbh⫹/⫺ vs. Dbh⫺/⫺ mice;
number sign, P ⬍ 0.05 Dbh⫺/⫺ vs. Dbh⫺/⫺ ⫹ ISO.
SNS REGULATION OF LIVER FIBROSIS: NE
INTERACTIONS WITH OTHER PEPTIDES
That NE may subserve functions other than its classically assigned role of neurotransmission is established
in other organs. For example, cardiac remodeling in
heart failure involves mitogenic and fibrogenic actions
of NE that are mediated via adrenoceptors (Fisher and
Absher, 1995; Xiao et al., 2001; Akiyama-Uchida et al.,
2002). The in vivo sources that might provide NE for
HSC regulation include HSCs themselves, SNS nerve
terminals that abut HSCs, and the adrenal medulla
that releases NE and adrenaline into the circulation
under stressful conditions, such as liver injury. The
relative importance of these three sources in the regulation of HSC function in vivo is as yet unclear. It is also
The sympathetic neurotransmitter NPY is coreleased
with NE from SNS terminals. We demonstrated that NPY
promotes the proliferation of cultured HSCs. Indeed, NPY
has much greater mitogenic potency than NE, because
peak HSC proliferation was observed at 1 ␮M NE, but
occurred at as little as 0.1 nM NPY (Oben et al., 2003f). As
has been shown in studies of other mesenchymal cell types
(Kanevskij et al., 2002), we noted that these two cotransmitters interact to influence HSC proliferation. Increasing
concentrations of NE attenuate NPY-induced proliferation of HSCs (Oben et al., 2003f). NE and NPY also appear
to have divergent effects on collagen gene expression by
HSCs. While NE increases the expression of collagen1-␣2,
neither low nor high concentrations of NPY alter collagen1-␣2 mRNA levels. Thus, although both NE and NPY
increase HSC proliferation, only NE induces collagen gene
expression (Oben et al., 2003f). The clinical significance of
NE-NPY interactions for regulating HSC biology remains
untested.
Leptin
In bone and fat, leptin modulates tissue remodeling by
regulating NE production. Our recent studies of Dbh⫺/⫺
mice and ob/ob mice suggest that a similar process applies
in the liver, at least during fibrogenesis. Dbh⫺/⫺ mice
(which are leptin-replete but lack NE) do not activate
their HSCs in response to injury. Conversely, supplemental NE normalizes HSC numbers and activation in leptindeficient ob/ob mice. Whether or not leptin induces production of NE by HSCs, as it does in adipocytes (Commins
et al., 1999), is not yet known. It is also conceivable that
leptin promotes HSC activation by inducing the expression or function of adrenoreceptors or components of the
postreceptor signaling pathways that mediate NE effects.
More work is required to delineate the exact nature of
leptin-NE interactions.
SNS EFFECTS ON HEPATIC OVAL CELLS
SNS Inhibition Promotes Liver Regeneration:
Historical Evidence
Oval cells are another major cell type that is involved
in liver repair. Hepatic oval cells (HOCs) are facultative
stem cells that are activated when mature hepatocytes
reach a critically low level, as after subtotal necrosis, or
when mature hepatocytes are prevented from dividing.
SYMPATHETIC NERVOUS SYSTEM REGULATION
879
Fig. 5. SNS inhibition increases the numbers of hepatic progenitors
in livers with diet-induced damage. a: Immunohistochemistry for oval
cells in representative mice that were fed control diet (CMCD; top left),
MCDE (top right), MCDE diet ⫹ PRZ (bottom left), or MCDE ⫹ 6-OHDA
(bottom right). Oval cells are stained brown. b: The numbers of oval cells
were increased in all MCDE-fed groups compared to CMCD controls
(asterisk, P ⫽ 0.0001). Both groups treated with SNS inhibitors had more
oval cells than mice that were fed MCDE diets alone (number sign, P ⫽
0.001).
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OBEN AND DIEHL
The latter occurs in animals and humans with various
chronic liver diseases (Roskams et al., 2003). To determine whether the SNS effects HOC-mediated liver regeneration, we investigated the effects of SNS inhibition on the HOC response to chronic liver injury.
Although earlier work from other groups had shown
that SNS inhibition enhanced hepatic regeneration after a partial hepatectomy (Kato and Shimazu, 1983;
Kiba et al., 1994), it was not known if (or how) the SNS
influenced HOCs themselves or if SNS-HOC interactions played any role in the recovery from chronic liver
damage.
SNS Inhibition Promotes Accumulation of
HOCs and Reduces Liver Injury: New Evidence
We used an established model of OC activation (methionine-choline-deficient diets plus ethionine) to injure
the liver, inhibit mature hepatocyte replication, and
induce HOC accumulation (Akhurst et al., 2001). We
hypothesized that SNS inhibition would promote further HOC accumulation and reduce liver damage. Compared to control mice that were fed only the antioxidantdepleted diets, mice fed the same diets with prazosin
(an ␣1-adrenoceptor antagonist) or 6-hydroxydopamine
(6-OHDA, an agent that induces chemical sympathectomy) had significantly increased numbers of HOCs
(Fig. 5). Increased HOC accumulation was accompanied
by less hepatic necrosis (Fig. 6a) and steatosis (Oben et
al., 2003a, b), lower serum aminotransferases (Fig. 6b),
and greater liver and whole body weights (Oben et al.,
2003a, 2003b).
Neither PRZ nor 6-OHDA affected the hepatic expression of granulocyte, granulocyte/macrophage, or macrophage colony stimulating factor (G-CSF, GM-CSF,
M-CSF), interleukin (IL)-6, IL-7, IL-11, leukemia inhibitory factor, stem cell factor, hepatocyte growth factor, or
vascular endothelial growth factor (VEFG) and its receptors VEGFR1 and -3 cytokines, growth factors, or growth
factor receptors that are known to regulate progenitor
cells (Oben et al., 2003a, 2003b). Hence, the SNS may
exert its inhibitory effects on HOCs directly, by interacting with HOC adrenoceptors (Fig. 7) (Oben et al., 2003a,
2003b). More work is required to evaluate this possibility.
Nevertheless, it is likely that autonomic control of hepatic
progenitors is important given recent evidence that the
parasympathetic nervous system, acting via the vagus
nerve, also regulates the accumulation of HOCs (Cassiman et al., 2002b).
SNS REGULATION OF HSC AND HOC:
THERAPEUTIC IMPLICATIONS
Our studies of the SNS and HSCs extend understanding
of the mechanisms by which cathecholamines regulate the
repair of injured livers. The aggregate data support the
notion that SNS activation during liver disease (Henriksen et al., 1984; Esler et al., 1992) promotes liver fibrosis
(by activating HSCs) and may simultaneously inhibit liver
regeneration (by reducing HOC accumulation). Our goal is
to exploit this knowledge to develop novel treatments for
liver disease. Because NE promotes HSC activation, targeted interruption of catecholamine signaling in HSCs
may be a useful therapeutic approach to constrain liver
fibrosis. Similarly, targeted inhibition of SNS actions on
Fig. 6. SNS inhibition reduces diet-induced liver injury. a: Necrosis
score. Compared to controls (CMCD), all HMCDE-fed groups had more
necrotic hepatocytes (asterisk, P ⫽ 0.01), but compared to mice that
were fed the HMCDE diet alone, the numbers of necrotic hepatocytes
were reduced in HMCDE ⫹ PRZ (number sign, P ⫽ 0.05) or HMCDE ⫹
6-OHDA (number sign, P ⫽ 0.05). b: Serum ALT. Serum levels of ALT, a
marker of liver injury, were increased in all HMCDE-fed groups compared to CMCD controls (asterisk, P ⫽ 0.01). Compared to HMCDE-fed
mice, mice treated with HMCDE ⫹ PRZ or HMCDE ⫹ 6-OHDA had lower
ALT levels (number sign, P ⫽ 0.03).
HOCs may enhance liver regeneration by negating NE’s
inhibitory actions on liver repair by HOCs (Fig. 8). The
recent delineation of the adrenoceptor subtypes that are
expressed by HSCs and OCs is a step toward this goal.
Remaining challenges include the development of adrenoceptor antagonists that are devoid of vascular effects that
would otherwise limit their utility in patients with liver
disease.
Fig. 7. Oval cells express ␣1-adrenoceptors. a: Immunohistochemistry for ␣1-adrenoceptors on bile duct-type cytokeratin-positive oval
cells in a liver section from representative mice fed HMCDE. Oval cells
expressing ␣1-adrenoceptors are stained brown. b: Immunofluores-
cence studies confirms the colocalization of ␣1-adrenoceptors on bile
duct-type cytokeratin-positive oval cells. Without the primary antibodies,
binding of the secondary antibodies was negligible (not shown). ␣1adrenoceptors, red; cytokeratins, green; colocalization, yellow.
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OBEN AND DIEHL
Fig. 8. Schematic representation of the effect of SNS inhibition on
liver repair via SNS regulation of HSCs and OCs. SNS inhibition either at
its origin in the hypothalamic ventral medial nucleus (VMH) or peripherally will lead to a reduction of HSC activation, an increase of progenitor
cell numbers, a consequent reduction of scarring, and perhaps return
the injured liver to health. [Color figure can be viewed in the online issue,
which is available at www.interscience.wiley.com].
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