Medical Service, Department of Veterans Affairs Medical Center and Department of Medicine, University of Michigan, Ann Arbor, Michigan 48109
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ABSTRACT |
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Endogenous
gut-derived bacterial lipopolysaccharides have been implicated as
important cofactors in the pathogenesis of liver injury. However, the
molecular mechanisms by which lipopolysaccharides exert their effect
are not entirely clear. Recent studies have pointed to proinflammatory
cytokines such as tumor necrosis factor- as mediators of hepatocyte
injury. Within the liver, Kupffer cells are major sources of
proinflammatory cytokines that are produced in response to
lipopolysaccharides. This review will focus on three important
molecular components of the pathway by which lipopolysaccharides activate Kupffer cells: CD14, Toll-like receptor 4, and
lipopolysaccharide binding protein. Within the liver,
lipopolysaccharides bind to lipopolysaccharide binding protein, which
then facilitates its transfer to membrane CD14 on the surface of
Kupffer cells. Signaling of lipopolysaccharide through CD14 is mediated
by the downstream receptor Toll-like receptor 4 and results in
activation of Kupffer cells. The role played by these molecules in
liver injury will be examined.
endotoxins; tumor necrosis factor; macrophages; CD14
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INTRODUCTION |
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LIPOPOLYSACCHARIDES (LPS) are glycolipids found in abundance on the outer membrane of all gram-negative bacteria and have the ability to incite a vigorous inflammatory response. In humans, nanograms of LPS injected into the blood stream can result in all the physiological manifestations of septic shock (70, 118). Given that gram-negative bacteria normally colonize the colon, the body has developed strong defensive mechanisms that tightly regulate the entry and processing of LPS. The liver plays a central role in this process by virtue of its dual ability not only to clear LPS, but to respond energetically to LPS. The vast majority of LPS that enters the host in normal and pathological states does so through the gastrointestinal (GI) tract. Strategically and uniquely located at the gateway of the portal blood flow draining the GI tract, the liver is the final barrier to prevent GI bacteria and bacterial products, such as LPS, from entering the systemic blood stream (74). In experimental studies on healthy animals, LPS is cleared from the circulation within a few minutes of intravenous injection, and the majority is traced to the liver (65, 128). The liver's primary role in clearing LPS can be demonstrated in patients with liver failure. Endotoxemia is frequently found in patients with cirrhosis, and the degree of endotoxemia is correlated with the degree of liver failure (12, 58).
In addition to its ability to clear LPS, the liver also responds to LPS
with production of cytokines (63) and reactive oxygen intermediates (7, 8). Both ex vivo and in vitro studies with isolated liver perfusion and liver slice models have demonstrated that tumor necrosis factor (TNF)- and interleukin (IL)-1 are released in response to LPS, primarily by Kupffer cells
(63). Most of the toxicities of LPS, both in the liver and
in the systemic circulation, have been related to the release of these
inflammatory cytokines and mediators (35, 111). In
baboons, passive immunization with murine monoclonal anti-TNF
antibodies protects against multisystem organ failure caused by lethal
doses of live gram-negative bacteria (111).
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ROLE OF LPS IN LIVER INJURY |
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Multiple lines of evidence point to LPS as a cofactor in liver injury (80, 81). In animal models of liver injury, including carbon tetrachloride toxicity, choline deficiency, and alcohol-induced liver disease, injury to the liver is mitigated or prevented by the addition of oral nonabsorbable antibiotics, colectomy, or germ-free conditions (2, 54, 62, 90). In contrast, the addition of LPS augments liver injury caused by these hepatotoxins (11, 14). Furthermore, in carbon tetrachloride- and galactosamine-induced liver injury, the addition of anti-LPS antibody E-5 significantly reduces hepatotoxicity caused by these agents (19). These studies strongly support a role for endogenous LPS in toxin-induced liver injury.
In alcoholic liver disease, the importance of LPS has also been demonstrated. Rats fed the Leiber-DeCarli liquid ethanol diet develop only fatty liver unless subsequent injections of LPS are given. Only the combination of LPS and chronic ethanol will produce a histological picture of hepatic necrosis and inflammation consistent with alcoholic hepatitis (11, 34). In contrast, with the Tsukamoto and French model of rat alcoholic hepatitis, in which rats are fed ethanol intragastrically in conjunction with a high-fat diet, the pathological picture of alcoholic hepatitis can be produced without further LPS challenge (114). Part of the discrepancy may be due to the presence of endogenous endotoxemia after the addition of alcohol to the diet. The level of endotoxemia present correlates with pathological injury (76, 78). Furthermore, pathological injury can be ameliorated by decreasing the level of endogenous endotoxemia with intraluminal lactobacillus or antibiotics (2, 75).
Endotoxin or LPS itself is not hepatotoxic at low concentrations.
However, its ability to stimulate an inflammatory response may account
for its pathogenicity in the liver. Although many parameters of the
inflammatory response contribute to liver injury (72), one
well-studied pathway is the production of TNF-. In many models of
liver injury elevated TNF-
levels are present and correlate with
injury (17, 48, 68). Inhibition of TNF-
activity can
decrease liver injury. The addition of soluble TNF receptors that
diminish the biological effect of TNF-
will significantly decrease
liver enzymes, improve liver histology, and decrease mortality acutely
after acute carbon tetrachloride administration (18).
Similar results are seen in chronic alcohol-induced liver disease
(44) in rats. In humans, elevated levels of TNF-
are seen in alcoholic hepatitis and are associated with increased mortality
(24). As potent producers of inflammatory cytokines such
as TNF-
, Kupffer cells have been implicated in the pathway leading
to liver injury (106). Accordingly, inhibition of Kupffer cells with gadolinium decreases injury in alcohol-induced liver disease
(1). Although the greatest attention has been focused on
the role of TNF-
in liver injury, other cytokines have been shown to
play important pathogenic roles, including proinflammatory cytokines
IL-6 and IL-18 (52, 115) as well as anti-inflammatory cytokines such as IL-10 (13). In alcoholic liver disease,
it is not just a marked elevation of one proinflammatory cytokine that
results in liver injury, but more likely an alteration in the balance
of pro- and anti-inflammatory factors that result in disease
(77).
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CD14 |
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Given the critical importance of LPS and Kupffer cells in the pathogenesis of many forms of liver injury, increasing attention has focused on the mechanisms by which LPS activates Kupffer cells. In peripheral blood monocytes, LPS activation is mediated through LPS binding protein (LBP), CD14, and Toll-like receptor (TLR)4. LPS binds to LBP, a 60-kDa acute-phase protein produced predominantly by the liver and secreted into the circulation (93, 121). Although LBP can be expressed extrahepatically, the majority of LBP is produced by the liver (102). Present in normal and acute-phase serum, LBP binds with high affinity to the lipid A portion of LPS and catalyzes the transfer of LPS to cell surface receptors such as membrane CD14 (31). In the presence of LBP, markedly less LPS is needed to activate peripheral blood monocytes. Although LBP is not required for interactions of LPS with CD14, its presence significantly decreases the concentration of LPS sufficient for cellular activation (92). Thus the LBP-CD14 pathway is critical at the low concentrations of LPS found under physiological condition (109).
Because of its location in the liver sinusoids, which drain blood from the GI tract, Kupffer cells are chronically exposed to higher concentrations of LPS than circulating peripheral blood monocytes. Thus it is likely that Kupffer cells have evolved specialized different mechanisms to interact with LPS. Unlike peripheral blood monocytes, Kupffer cells have relatively low baseline expression of CD14 (6, 103, 127). However, expression of CD14 on Kupffer cells can be upregulated with multiple stimuli including LPS (67). By immunohistochemical staining, low levels of CD14 are detected in livers of unstimulated mice. The liver CD14 levels rapidly increase and peak 6 h after intraperitoneal injection with LPS (67). CD14 expression in the liver is also increased in many types of liver disease, including alcoholic and cholestatic liver injury in rodents (20, 98, 112). In the intragastric model of alcohol-induced liver injury in rats, the level of hepatic inflammation varies with alterations in the fat composition of the diet (76). Animals fed more unsaturated fats have greater liver injury; the degree of injury correlates with the level of endotoxemia and with the level of Kupffer cell CD14 expression (100). In human disease, CD14 expression on Kupffer cells is low in normal human liver but increases with different inflammatory liver diseases (110). Kupffer cell CD14 expression also varies with different stages of the same disease. In biliary atresia, Kupffer cell CD14 increases in early stages but declines in the later stages (4).
The physiological significance of these observed differences in Kupffer cell CD14 expression is not entirely clear. However, it is tempting to hypothesize that the changes in CD14 expression could be the underlying mechanism that determines the liver's sensitivity to LPS toxicity. This line of reasoning is supported indirectly by studies showing that CD14 expression is linked with LPS responsiveness in the monocytic cell line THP-1 (64). Immature THP-1 cells respond poorly to LPS in the presence and absence of serum. Treatment with the maturational agent calcitriol causes dose- and time-dependent increases in CD14 mRNA and surface CD14 expression. This increased expression is associated with enhanced responsiveness of these THP-1 cells to LPS (64). Similarly, transfection of CD14-negative 70Z/3 cells, a murine pre-B cell line, results in heightened sensitivity to LPS (55). In vivo, CD14 transgenic mice that overexpress CD14 on monocytes have increased sensitivity to LPS (25). In contrast, genetically engineered CD14-deficient mice are insensitive to LPS (37). It should be noted that although CD14-deficient mice are insensitive to LPS, their responses to whole bacteria remain intact (71). Isolated peritoneal macrophages from CD14-deficient animals have deficient responses to LPS, but their response to whole bacteria remains intact (71) and appears to be mediated, in part, by other LPS receptors such as CD11b/CD18.
Because "resting" Kupffer cells have low basal expression of CD14,
one may hypothesize that Kupffer cells, unlike peripheral blood
monocytes, react to LPS in a CD14-independent manner (10, 57). Although this is an attractive hypothesis, the data support a more complicated paradigm. Although Kupffer cells express little CD14, CD14 is still critical for LPS activation (98).
Isolated human Kupffer cells can respond to low concentrations of LPS
(10 ng/ml) with the production of TNF-
. Preincubation of the
Kupffer cells with a monoclonal mouse anti-human CD14 antibody MY4
inhibits production of TNF-
and supports an essential role for the
CD14 receptor in Kupffer cell activation by LPS under
physiological conditions. In addition, Kupffer cells isolated from
CD14-deficient mice are significantly less sensitive to LPS than
wild-type animals (98).
Discovered as a myeloid differentiation marker, CD14 was previously
thought to be expressed exclusively by myeloid cells
(127). Both a membrane and soluble form of CD14 exist in
serum. The soluble form is identical to the membrane, with the
exception of the glycophosphatidyl inositol (GPI) anchor, which is not
present in the soluble form of the protein. sCD14 also binds LPS and
can facilitate interactions with CD14-negative cells such as
endothelial cells (26, 39, 87). The source of sCD14 was
previously attributed to shedding from monocytic cells bearing CD14 or
from direct secretion by monocytic cells (9, 15). However,
it has become clear that other cells can also secrete soluble CD14. In
situ hybridization and immunohistochemistry for CD14 show induction of
CD14 mRNA and protein in mouse hepatocytes after LPS injection in vivo
(22). In the intragastric ethanol model of rat alcoholic
liver disease, isolated hepatocytes from dextrose-fed animals do not
have significant expression of CD14 mRNA. However, with the addition of
ethanol in the diet, the level of CD14 mRNA increases and is correlated with the degree of liver injury observed (100). Isolated
hepatocytes from in vivo LPS-treated rats have upregulation of both
cell-associated and soluble CD14 protein production when plated in
vitro (59). Basal expression of hepatocyte CD14 is
present, but upregulation after inflammatory stimuli appears to be
mediated, in part, by IL-1 and TNF- (23, 59, 61).
The finding of hepatocellular sCD14 production is also supported by studies of transgenic mice generated using an 80-kb human CD14 genomic DNA fragment (40). These mice have high expressions of CD14 in both monocytic cells and hepatocytes. Further analysis of these transgenic mice demonstrated that monocyte and hepatocellular expression of CD14 are differentially regulated. An upstream regulatory element beyond a 24-kb region of the genomic DNA but within the 33-kb region of human CD14 was critical for hepatocellular CD14 expression but not monocytic expression (84). Furthermore, the 33-kb transgenic mice produced a soluble form of human CD14 in serum, confirming that hepatocytes are a source of sCD14 in vivo.
Isolated human hepatocytes from normal livers express CD14 mRNA and protein in culture, predominantly the soluble form (97). This sCD14 is biologically active and able to facilitate LPS responses in the CD14-negative epithelial cell line SW620, resulting in activation and production of IL-8 in response to low concentrations of LPS. It is important to note, however, that sCD14 can exert widely varying effects on cellular responses to LPS. The balance between activation vs. inhibition of LPS responses by sCD14 is dependent on its concentration. At what are thought to be supraphysiological concentrations, sCD14 can compete with mCD14 and inhibit LPS activation of CD14-positive cells (29, 38, 96, 113). The exact concentrations of mCD14 and sCD14 present in the microenvironment of the liver are not known. Whether local hepatocyte expression of sCD14 can lead to high enough concentrations to suppress Kupffer cell reactivity to LPS is not known. However, interpretations of studies in liver injury should take into account the relative contributions of Kupffer cell and hepatocyte CD14.
In CD14 knockout mice that are deficient in both membrane and soluble
CD14 expression, the predominant phenotype is that of LPS insensitivity
(37). Yin et al. (126) demonstrated that after 4 wk of continuous enteral ethanol delivery, CD14-deficient animals had less liver injury than wild-type BALB-c controls. These
studies in the rodent model of ethanol-induced liver disease suggest
that increases in CD14 expression contribute to the pathogenesis of
liver injury. In human alcoholic disease, a promoter polymorphism that
is associated with increased CD14 expression was found to be more
common in men with alcoholic hepatitis and cirrhosis (47). A polymorphism at 159 (cytosine to thymidine) of the human CD14 gene,
which lies within the Sp1 transcriptional binding site known to affect
CD14 expression in monocytes, was examined in autopsy specimens of men
with documented alcohol consumption. The thymidine variants of
the
159 polymorphism promote CD14 gene expression and cause higher
expression of CD14 on monocytes (43). In this study, the
thymidine allele was found to be associated with alcoholic hepatitis
and cirrhosis but not with fatty liver, periportal fibrosis, or
bridging fibrosis. These findings suggest that CD14 expression may
directly influence human susceptibility to alcohol-induced liver injury
(47).
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TLRS |
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Because membrane CD14 is a GPI-anchored protein without a transmembrane component, downstream partners for this receptor have been long sought. Recent studies in TLRs have been instrumental in elucidating the pathway by which ligand binding to membrane CD14 results in cellular activation. The Toll receptors are evolutionarily well conserved and were first described in Drosophila with biological functions in development and immunity (89). A series of TLRs, which are human homologs of the Toll receptors in Drosophila, have been cloned (69). To date, at least 10 TLRs have been identified in mammalian cells, although the ligands for most of them have not been identified (3). These receptors are typified as type I transmembrane proteins, with a cytoplasmic domain resembling that of the IL-1 receptor. In contrast to the IL-1 receptors, the extracellular domain of TLRs contains leucine-rich repeats rather than immunoglobulin-like domains.
Of particular interest as putative LPS receptors are TLR2 and TLR4.
Both of these receptors are highly expressed in cells that respond to
LPS, such as macrophages and monocytes. Both receptors can induce
activation of cells through nuclear factor- (NF-
B). Initially,
cellular transfection experiments suggested TLR2 as the LPS receptor
for CD14/LBP-dependent activation of cells (49, 125).
However, subsequent experiments identifying naturally occurring mutations of TLR4 as the cause of LPS resistance in two strains of mice
cast doubt on TLR2 as the main LPS receptor. A missense mutation in the
TLR4 gene accounted for the endotoxin resistance found in C3H/HeJ mice,
and a null mutation of TLR4 accounted for the endotoxin resistance in
C57BL/10ScCr mice (86). Furthermore, TLR4 knockout mice
have been produced and have been found to have LPS resistance
(42) similar to the naturally occurring mutants. In
contrast, TLR2-deficient mice are as capable of responding to LPS as
their wild-type controls (104). This discrepancy in previously reported experimental results may have been due to the
purity of LPS preparations used. Repurification of LPS resulted in loss
of signaling through TLR2, suggesting that the initially observed TLR2
signaling may have occurred as a result of contaminants within LPS
preparations (41).
TLR 2 appears to mediate responses to a wide variety of bacterial
products including lipoproteins derived from gram-negative and
-positive bacteria as well as mycoplasma (5). In contrast, TLR4 is specific for LPS from gram-negative bacteria. Signaling through
TLR4 requires MD-2, a secreted protein that is closely associated with
the extracellular domain of TLR4 (94). LPS responsiveness in human kidney 293 cells requires overexpression of both TLR4 and MD-2
(94). Mutations in MD2 result in decreased LPS
responsiveness (91). Downstream of TLR4, signaling can
occur through the IL-1-receptor pathway, which is MyD88 dependent, or
alternatively via a MyD88-independent pathway (5). In the
MyD88-dependent pathway, the death domain of MyD88 associates with the
serine threonine kinase IL-1 receptor-associated kinase. Through the
subsequent association with TNF receptor-activated factor 6 and a
series of signaling pathways, IB degradation occurs. This liberates
NF-
B, allowing its translocation to the nucleus, where it can induce
target gene expression (Fig. 1)
(5). Although ample evidence supports TLR4 as an important
downstream partner for membrane CD14, a role for other coreceptors may
also exist (85). With the use of fluorescence resonance
energy transfer, Pfeiffer et al. (85) demonstrate
coclustering of several receptors including Fc
-RIIIa (CD16a), CD81,
and TLR4 into lipid rafts after binding of LPS to CD14.
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Within the liver, a critical role for TLR4 is suggested by the presence
of TLR4 on isolated Kupffer cells and their requirement for a
functional TLR4 protein to mediate Kupffer cell responses to low
concentrations of LPS (99). A role for TLR4 in liver injury is suggested by studies with intragastric ethanol administration in TLR4 mutant C3H/HeJ mice. After 4 wk of intragastric ethanol, these
mice have less liver injury histologically, lower alanine amino
transferase levels, and lower TNF- mRNA expression than wild-type
controls (116). Given the narrow specificity of
TLR4 for LPS and not for other immunogenic bacterial products such as
bacterial lipoproteins and bacterial CpG-DNA, this study suggests that
in this mouse ethanol model of liver injury, LPS rather than other
bacterial components is the specific pathophysiological agent. Whether
this will be true in other models of liver injury remains to be seen.
In addition to TLR4, TLR2 mRNA is expressed by rodent hepatoctyes
(60, 66) and can be upregulated by LPS in vivo. Regulation
of TLR2 mRNA appears to be mediated, in part, by IL-1 and TNF-
(60). The significance of TLR2 expression on hepatocytes
and its role in liver injury is, as yet, unknown.
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OTHER LPS RECEPTORS |
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The focus of this review has been on the CD14 and TLR4 pathway of
Kupffer cell activation by LPS, because this is the best characterized
high-affinity receptor system for LPS in monocytic cells. However,
other receptor systems may be present (56). Another
receptor system well described in the liver are the scavenger receptors, also known as Ac-LDL receptors (95). This
receptor system has specificity toward negatively charged ligands such as Ac-LDL, polyinosinic acid, and maleylated BSA. On macrophages such
as Kupffer cells, two types of scavenger receptor A have been
identified (types I and II), which are both products of a single gene.
The two gene products result from alternative splicing. Both subtypes
of scavenger receptor A are found in abundance on macrophages and have
very broad specificity toward negatively charged ligands such as
Ac-LDL, polyinosinic acid, and maleylated BSA. They also have a great
affinity for LPS, lipid A, and the bioactive precursor of lipid A,
lipid IVA. This receptor system has been found to
participate in the binding of LPS by macrophages before its metabolism
to a less active form. Binding of LPS to the scavenger receptors does
not result in Kupffer cell activation or production of TNF-. In
mice, ~75% of the total injected lipid IVA can be found
in the liver within 10 min. The amount of liver uptake can be inhibited
by competitors of the scavenger receptor such as Ac-LDL, polyinosinic
acid, and maleylated BSA, suggesting that scavenger receptors play a
role in the clearance of LPS by the liver (32).
Furthermore, recent studies in scavenger receptor type A knockout mice
show that these animals have a greater susceptibility toward endotoxin
shock that is correlated with higher serum levels of TNF-
and IL-6
(36). These findings suggest that scavenger receptors can
downregulate proinflammatory cytokine production by competing for LPS.
Recently, an interesting class of intracellular receptors for LPS has
been identified. Nucleotide binding domains (Nod)1 and -2 are members
of a family of intracellular proteins composed of an
NH2-terminal caspase recruitment domain and a centrally located Nod (45). The COOH-terminal portions have
leucine-rich repeats and serve as the sensors for intracellular
ligands. LPS has been identified as a ligand for Nod1 and Nod2. Binding
of LPS to Nod1 and Nod2 results in TLR4- and MYD88-independent
activation of NF-B activation, as demonstrated in experiments with
Nod1- and Nod2-transfected human embryonic kidney 293T cells. Not
surprisingly, the leucine-rich region of the protein appears to be
essential for LPS-induced NF
B activation (46). Whereas
Nod1 is found in a wide range of cell types, Nod2 appears to be
restricted to monocytic cells (83). A recent report of a
single nucleotide polymorphism in the Nod2 gene that is associated with
susceptibility to Crohn's disease has generated much interest in this
receptor. The frameshift mutation results in a truncated protein
lacking part of the leucine-rich region and deficient in LPS-mediated NF-
B activation (82). Whether this receptor will have
any role in susceptibility to liver injury remains to be seen.
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LBP |
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As noted earlier, LBP binds with a high degree of specificity and
affinity (dissociation constant = 109) to the lipid
A portion of bacterial LPS (107). The
NH2-terminal half of the protein is responsible for
specific binding to LPS, whereas the COOH-terminal half of LBP is
responsible for CD14 interactions. A truncated form of human LBP
composed of the 197 amino acids in the NH2 terminus of the
parent molecule effectively binds LPS but cannot transfer the LPS to
CD14 (33, 105). At low concentrations (3 ug/ml), which is
present is normal serum, LBP can facilitate the transfer of LPS to CD14
on isolated Kupffer cells ex vivo, with accelerated binding and more
TNF-
production at lower concentrations of LPS (98,
99). As an acute-phase protein, LBP is present in normal serum
at a concentration of 0.5-20 µg and can increase as much as
100-fold with an acute-phase response (16, 27, 73, 93,
108). Hepatocytes are the main producers of LBP, although LBP is
also produced locally in the lung and skin (50, 51, 88,
102). Within the liver, therefore, Kupffer cells can be exposed
to high concentrations of LBP.
Whereas the LPS-potentiating properties of LBP are well recognized, its ability to mediate widely varying functions at different concentrations has been underappreciated. With significant sequence homology to the lipid transfer proteins, cholesterol ester transfer protein and phospholipid transfer protein, LBP can transfer LPS to lipoproteins such as high-density lipoprotein (HDL), leading to neutralization of LPS (122). In fact, the ability of lipoproteins such as HDL to bind and neutralize LPS may depend on a transfer protein such as LBP (124). Reconstituted HDL (R-HDL) prepared from purified apolipoprotein A-I combined with phospholipid and free cholesterol were not sufficient to neutralize the biological activity of LPS, but the addition of LBP enabled prompt binding and neutralization of LPS by R-HDL (124). In serum, LBP is predominantly associated with lipoproteins, and its association strongly enhances the ability of lipoproteins to bind LPS (119). Given the role that lipoproteins play in neutralizing LPS in blood, LBP is well placed to participate in the neutralization of LPS.
Irrespective of its ability to transfer LPS to lipoproteins, the
multiple functions of LBP can be illustrated by its dose-dependent effects on monocyte activation. Addition of low concentrations of
recombinant murine LBP (up to 1 ug/ml) augments RAW 267.4 cell responses to low concentrations of LPS (0.33 ng/ml). However, at higher
LBP concentrations (10 ug/ml), the previously observed enhancement of
responses to LPS is lost. At higher concentrations of LPS, the
previously effective concentration of LBP (10 ug/ml) is no longer
sufficient to inhibit responses to LPS. In vivo, the addition of
recombinant murine LBP diminishes LPS response (TNF- levels),
mortality, and ALT levels in the LPS-galactosamine model of liver
injury (53). Concentration-dependent effects of LBP have
also been described for priming of neutrophils by LPS (1 ng/ml) for
subsequent responses to
N-formyl-methionyl-leucyl-phenylalanine (113).
In this system, sCD14 can activate or inhibit the priming of
neutrophils. The end effect of sCD14 is dependent on the concentration of LBP. For LBP concentrations >3-10 ng/ml, the addition of sCD14 resulted in inhibition of neutrophil priming. At <3 ng/ml of LBP, the
addition of sCD14 enhanced priming (113). The mechanism by which LBP can exert opposing effects is not known, but these data suggest that a complex interplay of multiple concentration-dependent factors determines the biological effect of these proteins.
The phenotype expressed by the LBP knockout mice produced by Wurfel et
al. (123) supports the hypothesis that the primary role of
LBP in vivo may not be to augment LPS reactivity. These LBP-deficient
mice do not demonstrate any difference in TNF- production when
injected with LPS in vivo. In contrast, whole blood from these animals
was 1,000-fold less responsive to LPS as assessed by TNF-
production
ex vivo than that of wild-type controls. These findings suggest that in
vivo sources of TNF-
production such as the liver may not require
LBP for LPS responsiveness. In fact, the critical function of LBP in
vivo may be to protect animals from bacterial infections. This latter
supposition is supported by studies showing the exquisite sensitivity
of these LBP-deficient animals to a Klebsiella pneumoniae
challenge (21).
As an acute phase protein, hepatocyte LBP expression is upregulated in
many models of acute-phase response and liver injury including
endotoxemia, intramuscular turpentine injection, and alcoholic
hepatitis (28, 100). Like other acute phase proteins, LBP
production is regulated by proinflammatory cytokines such as IL-6,
IL-1, and TNF- (30, 120). The role LBP plays in liver injury when it is upregulated is not clear. Early studies with acute
carbon tetrachloride-induced liver injury in LBP-deficient animals
suggest that LBP-deficient animals have defective recovery from liver
injury (79, 101). However, in mice given intragastric ethanol for 1 mo, a reduction in liver injury is seen in LBP-deficient animals (117). These contrasting effects of LBP deficiency
suggests a more complex paradigm for liver injury that is dependent on relative concentrations of LBP and its cofactors (CD14 and TLR4) as
well as the timing and type of injury.
In summary, the liver and, in particular, the Kupffer cell play an important role in the innate immune response to bacteria and bacterial products that enter the portal system. In the setting of liver injury, this function is impaired. Evidence suggests that these endogenous bacterial products, LPS in particular, can exacerbate the ongoing liver injury. Recent investigations into the molecular mechanisms by which LPS interacts with Kupffer cells and other mononuclear cells shed light on the potential role played by LBP, CD14, and TLR4 in the pathogenesis of different forms of liver injury. Although binding of LPS to LBP with transfer to CD14/TLR4 on Kupffer cells can result in activation, a complex interplay of other factors, including the relative concentrations of LBP and sCD14, may ultimately determine the Kupffer cell's response to LPS. An understanding of these factors, which are altered in liver injury, can lead to better therapeutic options in the future.
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ACKNOWLEDGEMENTS |
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This work was supported by National Institutes of Health Grants DK-53296 and GM-60205 and a Veteran's Administration Merit Award.
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FOOTNOTES |
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Address for reprint requests and other correspondence: G. L. Su, Univ. of Michigan Medical Center, 1510C MSRB I, Box 0666, 1150 W. Medical Center Dr., Ann Arbor, MI 48109-0666 (E-mail: gsu{at}umich.edu).
10.1152/ajpgi.00550.2001
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