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Prepublished online as a Blood First Edition Paper on November 27, 2002; DOI 10.1182/blood-2002-05-1300.
HEMOSTASIS, THROMBOSIS, AND VASCULAR BIOLOGY
From the Critical Care Medicine Department, Warren G. Magnuson Clinical Center, National Institutes of Health, Bethesda; and
Laboratory of Molecular Carcinogenesis, Division of Basic Sciences,
National Cancer Institute, Bethesda, MD.
Systemic inflammation because of sepsis results in endothelial cell
activation and microvascular injury. High-mobility group protein-1
(HMGB1), a novel inflammatory molecule, is a late mediator of endotoxin
shock and is present in the blood of septic patients. The receptor for
advanced glycation end products (RAGE) is expressed on endothelium and
is a receptor for HMGB1. Here we examine the effects of HMGB1 on human
endothelial cell function. Recombinant human HMGB1 (rhHMGB1) was cloned
and expressed in Escherichia coli and incubated with human
microvascular endothelium. rhHMGB1 caused a dose- and time-dependent
increase in the expression of intercellular adhesion molecule 1 (ICAM-1), vascular cell adhesion molecule 1 (VCAM-1), and RAGE. rhHMGB1
induced the secretion of tumor necrosis factor- Systemic inflammation is one of the hallmarks of
septic shock, and the microvascular endothelium provides an important
site of regulation and amplification of these inflammatory
responses.1 Endothelial cell activation leads to
alterations in hemostasis, increases in vascular permeability, cell
swelling and loss of barrier function, leukocyte adherence with cell
clumping, and microthrombi formation.2,3 Microvascular
injury is one of the characteristics of sepsis-associated tissue damage
that may be manifested by single (eg, acute respiratory distress
syndrome) or multiple organ failure syndromes.4 Many
inflammatory mediators of sepsis contribute to the development of an
activated endothelium (eg, tumor necrosis factor- HMGB1 is secreted by macrophages stimulated with endotoxin, TNF HMGB1 is a multifunctional protein; its earliest functions were
described as a nonhistone DNA-binding nuclear protein. HMGB1 binds to
DNA in a sequence-independent manner and modifies DNA structure to
facilitate transcription, replication, and repair.9,10 These functions are essential for survival, as HMGB1-deficient mice die
of hypoglycemia within 24 hours after birth.11 In the developing nervous system, membrane-associated HMGB1 localizes to
growth cones and promotes neurite outgrowth and extension by binding to
the receptor for advanced glycation end products
(RAGE).12,13 RAGE, a member of the immunoglobulin
superfamily of receptors, is expressed on endothelium, smooth muscle
cells, monocytes/macrophages, neurons, and in several malignant and
transformed cells.14 RAGE interacts with a variety of
ligands, including advanced glycation end products and
HMGB1.15 HMGB1 binding to RAGE on neuroblastoma cells
activates nuclear factor The presence of HMGB1 in the circulation of septic patients and the
widespread expression of RAGE on endothelium suggests that HMGB1 might
interact with endothelial cells and contribute to the inflammatory
responses to infection. We demonstrate that human microvascular
endothelial cells stimulated in vitro with HMGB1 increase their
expression of intercellular and vascular adhesion molecules (ICAM-1 and
VCAM-1) and RAGE receptor as well as secretion of a proinflammatory
cytokine (TNF Production of recombinant human HMGB1
Cell culture
Effects of rhHMGB1 on endothelial cells
Quantitative flow cytometry Endothelial cells were harvested with 0.25% trypsin and 1 mM EDTA (ethylenediaminetetraacetic acid; Clonetics), washed twice in PBS, and immediately fixed in 2% paraformaldehyde in PBS. After 15 minutes of incubation in 100 µL blocking solution (PBS, 0.1% bovine serum albumin [BSA], 10% normal mouse serum), 2 × 106 cells were incubated for 30 minutes with a phycoerythrin (PE)-mouse antihuman ICAM-1 monoclonal antibody (1 µg/mL; Caltag, Burlingame, CA), fluorescein isothiocyanate (FITC)-mouse antihuman VCAM-1 (2 µg/mL; Serotec, Raleigh, NC), or the corresponding immunoglobulin isotypes (IgG2a-PE and IgG1-FITC; Serotec). For detection of the RAGE, endothelial cells were blocked for 15 minutes in PBS with 0.1% BSA and 10% normal donkey serum and then incubated for 30 minutes with a goat antihuman polyclonal anti-RAGE antibody (1 µg/mL; Research Diagnostics) or an irrelevant goat IgG. Cells were washed, and an FITC-conjugated donkey antigoat polyclonal antibody (2 µg/mL; Molecular Probes, Eugene, OR) was added for 30 minutes. Cells were thoroughly washed and analyzed in a fluorescence-activated cell sorter scan (FACScan) system using CELLQuest software (Becton Dickinson, San Jose, CA). Fluorescence emission of 104 cells/sample was recorded as peak channel number on a logarithmic scale. To calculate the number of ICAM-1 and VCAM-1 receptors per cell, the peak channel of each cell population was transformed into antibody binding sites per cell using the Quantum Simply Cellular Microbeads and Quickcal software (Bangs Laboratories, Fishers, IN) as previously described.27,28 The number of antibody binding sites corresponding to the isotopic IgG was subtracted from each cell sample. For RAGE quantification, peak channel of each cell populations was transformed into molecules of equivalent soluble fluorochrome (MESFs) using the Quantum FITC MESF kit (Bangs Laboratories). MESFs corresponding to the irrelevant goat IgG were subtracted from each cell sample. Results are expressed as number of MESFs per cell.29Secreted protein assays Immunoreactive TNF , IL-1 , MCP-1, IL-8, soluble ICAM-1
(sICAM-1), and soluble VCAM-1 (sVCAM-1), were measured in HMEC-1 and HMLEC supernatants by enzyme-linked immunosorbent assay
(ELISA) according to the manufacturer's instructions (R&D Systems).
PAI-1 and tPA were measured by ELISA according to the manufacturer's instructions (American Diagnostica, Greenwich, CT).
Quantitative real-time PCR Changes in mRNA levels for TNF and IL-8 were quantified by
real-time PCR (Taqman PCR detection; Applied Biosystems, Foster City,
CA) in a 2-step reverse transcription-polymerase chain reaction (RT-PCR). Confluent monolayers of HMEC-1 cells were incubated with
rhHMGB1 (100 ng/mL) or medium alone for 2 hours (n = 4), 6 hours
(n = 4), and 24 hours (n = 3). Cells were lysed with RLT buffer and
total RNA was isolated (Rneasy; Qiagen). The reverse transcription of
RNA (2 µg/reaction) to cDNA was performed using random hexamers from
the TaqMan reverse transcription reagents according to the
manufacturer's instructions (Applied Biosystems). PCR products were
synthesized from cDNA samples using the TaqMan universal PCR master mix
and TaqMan predeveloped assay reagents for TNF and IL-8 according to
the manufacturer's instructions. RNase P (Applied Biosystems) was used
as an internal control. Amplification and detection were carried out
using the ABI PRISM 7900HT Sequence Detection System (Applied
Biosystems). All values were normalized to levels of RNase P and were
expressed as fold changes compared with unstimulated
samples.30
Activation of mitogen-activated kinases Mitogen-activated protein (MAP) kinase activation was determined by immunoblotting endothelial cell lysates. Confluent monolayers of HMEC-1 cells were stimulated with rhHMGB1 (100 ng/mL). At baseline, 5, 15, 30, 60, and 90 minutes, cells were washed with cold PBS and lysed (M-Per Mammalian Protein Extraction Reagent; Pierce, Rockford, IL). After centrifugation (14 000g for 10 minutes), supernatants were recovered, and the protein content was quantified by the Bradford method (BCA; Pierce). Total protein (10 or 20 µg) was fractionated by electrophoresis on 4% to 20% SDS-PAGE gels (Novex, Carlsbad, CA) under reducing and denaturing conditions and was transferred to nitrocellulose membranes. The double-phosphorylated forms of the MAP kinase were detected with the following antibodies: 0.5 µg/µL rabbit polyclonal anti-ERK1&2/MAPK (extracellular signal-related kinase 1 and 2; pTpY185/187) phosphospecific antibody, 0.5 µg/µL rabbit polyclonal anti-JNK1&2/SAPK (Jun N-terminal kinase 1 and 2 and stress-activated protein kinase; pTpY183/185) phosphospecific antibody, and 1 µg/µL rabbit polyclonal anti-p38 (pTpY180/182) phosphospecific antibody (all Biosource International, Camarillo, CA). Total MAP kinase proteins were detected with the following antibodies: 100 ng/mL rabbit polyclonal antibody anti-ERK1/2 (Promega), and 1/1000 dilution of rabbit polyclonal antibodies anti-SAPK/JNK and anti-p38 (Calbiochem, San Diego, CA). After incubation with peroxidase-conjugated donkey antirabbit secondary antibody (1/10 000 dilution; Jackson Immunoresearch, West Grove, PA), immunoreactive proteins were visualized with a chemiluminescence detection system (ECL+plus, Amersham Biotech). Band optical density corresponding to phosphorylated isoforms was analyzed by densitometry (Molecular Dynamics 301, Piscataway, NJ) using Image Quant software.To assess the contribution of p38 MAPK and ERK pathways to
HMGB1-stimulated cytokine responses, we used specific inhibitors of p38
MAPK (SB203580; Calbiochem) and ERK1/2 (PD98059; Calbiochem). The
inhibitors were dissolved in dimethyl sulfoxide (DMSO) and added individually or together 2 hours prior to cell stimulation with
HMGB1 (50 ng/mL). The final concentration of the inhibitors was
SB203580, 10 µM dissolved in 0.1% DMSO, and PD98059, 25 µM dissolved in 0.04% DMSO.31 Cells were stimulated for 6 hours, and IL-8 and TNF Electrophoretic mobility shift assay (EMSA) Nuclear and cytoplasmic extracts from HMEC-1 cells were prepared (NE-PER Nuclear and Cytoplamic Extraction Reagents; Pierce) after 4-hour incubation with rhHMGB1 (100 ng/mL), TNF (10 ng/mL), or
control medium, and they were stored at  80°C until use. Protein content was assayed using the Bradford reagent and BSA as a standard (Pierce). End labeling of double-strand oligonucleotides containing the
NF- B binding motif (5'-AGT TGA GGG GAC TTT CCC AGC-3') and the Sp1
consensus DNA binding motif (5'-ATT CGA TGC GGG CGG GGC GAG C-3') was
performed using  32P-dATP (deoxyadenosine
triphosphate; Amersham Pharmacia, Piscataway, NJ) and
T4-polynucleotide kinase (Promega) at 37°C for 30 minutes. The
removal of unincorporated nucleotide was done using G-25 Sephadex columns (Pharmacia Biotech, Piscataway, NJ). Equal amounts of nuclear
extract protein (10 µg) were incubated in a 10-µL reaction mixture
of 4% glycerol, 1 mM MgCl2, 0.5 mM EDTA, 0.5 mM
DTT (dichlorodiphenyltrichloroethane), 50 mM NaCl, 10 mM Tris
(tris(hydroxymethyl)aminomethane)-HCl, and 50 µg/mL poly (dI-dC)
with or without a 100 × unlabeled oligonucleotide for 10 minutes at
room temperature. 32P-labeled oligonucleotide probes were
then added and incubated at room temperature for 20 minutes. In
supershift assays, 1 µg antibody against NF-B subunits p50 or p65
(Santa Cruz Biotechnology, Santa Cruz, CA) was added after the binding
reaction and incubated for 1 hour at 4°C. Electrophoresis of samples
was performed on 6% DNA retardation gels run in 0.5 × TBE buffer.
Gels were vacuum-dried, and autoradiography was performed for 12 to 36 hours at 80°C.
Statistics The data were analyzed using an analysis of variance (ANOVA) (SAS System 8.0; SAS Institute, Cary, NC).32 Depending on the experimental conditions, the model contained the following effects: time, cell, condition (HMGB1, negative and positive control), and level of antibody. All remaining interactions were pooled to form the error term. An estimate of the difference of the HMGB1 and negative control was made at the final time point. A linear and a quadratic model were used to examine the dose response curves, and the model with the best fit is reported. Variables were transformed where appropriate. Data are presented as mean ± standard error of the mean.
rhHMGB1 induces the expression of the adhesion molecules ICAM-1 and VCAM-1 and its putative receptor RAGE on microvascular endothelial cells ICAM-1 and VCAM-1 are adhesion receptors expressed on endothelial cells, and their up-regulation in response to proinflammatory cytokines is a sensitive marker of endothelial cell activation.1 RAGE is a receptor for extracellular HMGB1,16 and its up-regulation might provide an amplification system for rhHMGB1 effects on endothelial cells. A preliminary study was designed to select an appropriate dose of hrHMGB1 for the endothelial cell stimulation experiments, based on HMGB1 levels detected in the serum of septic patients (approximately 30-150 ng/mL).7 Confluent cultures of HMEC-1 cells were incubated for 24 hours with doses of rhHMGB1, ranging from 5 to 200 ng/mL, and ICAM-1 and VCAM-1 expression was analyzed by quantitative flow cytometry. rhHMGB1 increased the expression of ICAM-1 and VCAM-1 in a dose-dependent manner (25 ng/mL, 1.6- and 3.7-fold increase; 50 ng/mL, 12.1- and 20.9-fold increase; 100 ng/mL, 13.4- and 24.2-fold increase; 200 ng/mL, 11.3- and 22.3-fold increase compared with control, respectively, both P < .0001). HMGB1derived from A549 cells (100 ng/mL) increased ICAM-1 and VCAM-1 when incubated with the HMEC-1 cells (data not shown).Incubation of confluent cultures of HMEC-1 cells with rhHMGB1 (50 ng/mL × 24 hours) or TNF
rhHMGB1 induces the secretion of a proinflammatory cytokine
(TNF , IL-8, and MCP-1 rose significantly compared with control
conditions with plateau levels at 24 to 48 hours (all P < .0001, Figure 4A-C). We
found different secretion patterns for TNF and the chemokines.
TNF concentration rose sharply 3 hours after rhHMGB1 stimulation,
reached a peak at 24 hours, and then slightly declined, whereas IL-8
and MCP-1 concentrations continued to increase. Similar induction of
cytokine and chemokines release by HMGB1 was observed with HMLEC-1
(Table 1). Proteolysis of rhHMGB1 with trypsin abolished release of
IL-8 by HMEC-1 cells (data not shown). No IL-1 was detected
following rhHMBG1 or TNF stimulation of the HMEC-1 or HLMEC cells (2 experiments in HMEC-1 and 2 experiments in HMLEC over 48 hours).
rhHMBG1 also significantly increased the release of 2 pivotal
regulators of fibrinolysis, tPA and PAI-1 (at 48 hours,
P = .005 and .0006, respectively, Figure
5 and Table 1). Incubation with an
anti-RAGE antibody (80 µg/mL) diminished IL-8 production by 14%
(5477 ± 1158 versus 4720 ± 1097 pg/mL,
P = .02, t test) and TNF
production by 17% (479 ± 24 versus 365 ± 39 pg/mL,
P = .11). Incubation with anti-RAGE antibody alone or
murine IgG (alone or together with rhHMGB1) had no effect on IL-8 or
TNF production.
Quantitative PCR analysis of TNF gene expression using TaqMan quantitative PCR
revealed that TNF mRNA was highly up-regulated 2 hours after treatment with rhHMGB1 (403.9- ± 96.7-fold increase from baseline, P < .001), sharply declined by 6 hours (56.2 ± 13.7),
and returned to baseline levels at 24 hours (5.2 ± 1.9). IL-8 mRNA
levels were up-regulated at 2 hours (182.6 ± 55.9,
P = .03) and 6 hours (129. 2 ± 14.9,
P = .02) and returned toward baseline levels at 24 hours (36.4 ± 10.8). These patterns change in parallel with the levels of
TNF and IL-8 protein found in the HMEC-1 supernatants.
Proinflammatory activity of HMGB1 is amplified by local TNF production To investigate whether the proinflammatory activity of rhHMGB1 on endothelial cells is mediated in part by local release of TNF, we
incubated HMEC-1 cells with an anti-TNF-neutralizing antibody (5 or
10 µg/mL) 30 minutes prior to HMGB1 (100 ng/mL) and TNF (10 ng/mL)
stimulation. Cells were incubated for 24 hours, IL-8 concentration was
measured in the supernatants, and ICAM-1 and VCAM-1 expression was
quantified by flow cytometry. Both concentrations of anti-TNF
antibody significantly decreased TNF- and rhHMGB1-induced secretion
of IL-8 and expression of ICAM-1 and VCAM-1 (Figure 6A-C). The suppressive effects of the
anti-TNF antibody were greater for cells stimulated with TNF than
with rhHMGB1 (10 µg/mL anti-TNF antibody decreased IL-8 production
by 91% ± 2% versus 72% ± 2%, ICAM-1 by 78% ± 4% versus
65% ± 3%, and VCAM-1 by 81% ± 3% versus 74% ± 3%,
respectively, P = .004). The suppressive effects of the
anti-TNF antibody were incomplete; after coincubation with the
anti-TNF antibody, both TNF- and rhHMGB1-stimulated cells had
levels of IL-8 and adhesion molecules that remained greater than media
alone (P < .0001).
rhHMGB1 interaction with endothelial cells induces MAP kinase activation MAP kinase signaling pathways play a central role in endothelial cell activation in response to proinflammatory stimuli such as TNF,
IL-1, and thrombin.33-35 Kinase activation was
determined in HMEC-1 cells after rhHMGB1 stimulation by detection of
the active, phosphorylated forms of 3 different MAP kinases.
Representative blots of 5 independent experiments and densitometry
analysis are shown in Figure 7. The
phosphorylated form ERK1/2 was constitutively expressed in HMEC-1
cells, was strongly induced after 15 minutes of rhHMGB1 exposure, and
remained up-regulated up to 60 minutes. Phosphorylation of JNK was
detected at 5 minutes, peaked at 30 minutes of rhHMGB1 stimulation, and
declined after 60 minutes. p38 phosphorylation was detectable at 5 minutes and remained present up to 60 minutes after incubation with HMGB1.
Inhibition of p38 MAPK with SB203580 decreased rhHMGB1-stimulated IL-8
release (at 6 hours) by nearly two thirds (P = .03), whereas ERK1/2 inhibition with PD98059 had no effect. The combination of both SB203580 and PD98059 decreased IL-8 release by 87%
(P = .0003). TNF
HMGB1 activates NF- B and Sp1. NF-B is a
multisubunit molecule that belongs to the Rel family of transcription
factors and its activation is triggered by proinflammatory cytokine
stimulation of endothelial cells.36 RAGE-HMGB1 binding in
neuroblastoma cells enhances NF-B-dependent
transcription.16 Sp1 is a transcriptional factor that
binds to GC boxes of various cellular promoters with sequence
specificity.37 SP-1 binding elements in the RAGE promoter mediate HMGB1-induced RAGE expression in neuroblastoma
cells.38 Nuclear extracts from HMEC-1 cells were incubated
for 4 hours with rhHMGB1, TNF, or medium alone and analyzed by EMSA.
Nuclear extracts from HMEC-1 cells treated with rhHMGB1 and TNF
showed large increases in specific binding of the NF-B
oligonucleotide. Specificity was confirmed by competitive assay with a
100-fold molar excess of unlabeled oligonucleotide and a specific
supershift with a monoclonal antibody against the p65 subunit (Figure
8). Sp1 binding activity was present in
control cells but was increased in rhHMGB1-stimulated cells (Figure 8).
Thus, HMGB1 activates 2 nuclear transcription factors in the
endothelium: NF-B, a rapid-response factor for gene expression
during inflammation, and Sp1, a binding factor associated with the RAGE promoter.
HMGB1 is a novel inflammatory cytokine that contributes to the
lethality of sepsis.7 We demonstrate that rhHMGB1 induces a proinflammatory phenotype in human microvascular endothelial cells in
vitro characterized by up-regulation of leukocyte adhesion molecules
(ICAM-1 and VCAM-1), secretion of the neutrophil and monocyte
chemokines (IL-8 and MCP-1), production of a proinflammatory cytokine
(TNF Endothelial activation is required for the transmigration and accumulation of inflammatory cells into tissues. We show that rhHMGB1 is a proinflammatory mediator and induces expression of 2 key adhesion molecules, ICAM-1 and VCAM-1. ICAM-1 mediates the firm adhesion of leukocytes to the endothelium and subsequent transmigration to the inflammatory sites. VCAM-1 contributes to the adhesion of activated lymphocytes and monocytes to endothelial cells in acute inflammatory tissues.39 Accompanying the up-regulation of adhesion molecules is the secretion of 2 potent neutrophil and monocyte chemokines, IL-8 and MCP-1. Thus, HMGB1 activates components that provide the necessary substrate for recruitment, adhesion, and transmigration of leukocytes across an activated endothelium and possibly to a nidus of inflammation.3 Tumor necrosis factor is a pivotal early mediator that regulates and
amplifies the development of inflammation. Endothelial cells secrete
TNF
Microvascular coagulation and fibrinolysis are important pathophysiologic events in inflammation and sepsis. We show that rhHMGB1 induces endothelial cells to secrete 2 key components of the fibrinolytic system, tPA and PAI-1 in endothelial cells. Under physiologic conditions, tPA is released from the endothelium, binds to the endothelial cell surface, and converts plasminogen to plasmin. Plasmin degrades fibrin and prevents clotting. PAI-1 inactivates tPA and limits any exaggerated fibrinolysis that might lead to bleeding.2 An excess of PAI-1 compared with tPA is commonly found during acute inflammation because of endotoxin, sepsis, or injury.45 In neural tissue, membrane-associated HMGB1 binds both tPA and plasminogen via the lysine-rich kringle domain, resulting in plasmin activation that degrades HMGB1.19 An additional source of HMGB1 during hemostasis is activated platelets; HMGB1 is exported to the surface of platelets on activation by thrombin.20 Thus, HMGB1 has complex interactions with the hemostatic system that lead to modulation of coagulation and fibrinolysis. Further, these interactions may result in the generation of plasmin that limits HMGB1 effects by proteolytic degradation.19 Therapies limiting the effects of HMGB1 may be useful to alter the development of the coagulopathic state associated with sepsis. Several lines of evidence demonstrate that the effects of HMGB1 are
mediated by binding to RAGE. In neural tissue and malignant cells, RAGE
activation by HMGB1 leads to MAP kinase activation and is associated
with enhanced tumor growth, metastases, and release of matrix
metalloproteinases.17 Rat smooth muscle cells undergo
chemotaxis and cytoskeleton reorganization following HMGB1 binding to
RAGE.46 We show that rhHMGB1 up-regulates RAGE on endothelial cells similar in degree to that of TNF We evaluated 2 similar but functionally distinct MAP kinase pathways;
ERK1/ERK2 kinase, activated by growth factors or mitogens, and the
stress-activated protein kinases, JNK and p38, that are activated in
response to diverse agonists (ie, cytokines, physical and chemical
agents).53 Within 5 to 15 minutes, concentrations of
rhHMGB1, similar to those found in the blood of septic patients, activate these 3 MAP kinase pathways in human microvascular endothelial cells. These results are similar to studies that demonstrate that several signaling pathways are activated by HMGB1 interaction with
target cells. In neuronal tissue, HMGB1 binding to RAGE simultaneously activates 2 distinct downstream signaling pathways: Rac and Cdc42 (members of the Rho family of small guanosine triphosphatases [GTPases]) and Ras and NF- MAP kinase inhibitors have been used to characterize key cell signaling
pathways resulting from HMGB1 cell activation. Increased smooth muscle
migration following HMGB1 binding to RAGE is mediated through a
pertussis toxin-sensitive pathway and blocked by PD98059, an inhibitor
of ERK1/ERK2 activation.46 The use of the p38 MAP kinase
inhibitor, SB203580, in our study significantly decreased IL-8
production and marginally decreased the release of TNF. Several factors
may have contributed to the diminished effect of p38 MAP kinase
inhibition on TNF secretion. SB203580 inhibits p38 Little data are available that describe the signaling pathways
associated with TNF secretion by endothelial cells. Human intestinal microvascular and umbilical vein endothelium constitutively express TNF
and IL-8 mRNA.41 We show that following HMGB1 stimulation, the temporal sequence of IL-8 and TNF mRNA expression in endothelial cells is different; TNF mRNA returns to baseline levels at 6 hours, whereas IL-8 mRNA returns to baseline at 24 hours. Activated MAP kinases may contribute to these different profiles by affecting transcription rates through NF- We show that 2 nuclear transcription factors, NF- rhHMGB1 is a potent proinflammatory molecule that activates endothelial
cells and smooth muscle cells at low concentrations (25-100 ng/mL).46 At higher concentrations (10-20 µg/mL), HMGB1 stimulates migration and growth of embryonic or malignant
cells.17,18,61 In vivo, 2 potential sources of circulating
HMGB1 may occur; HMGB1 secreted from monocyte-macrophages in response
to endotoxin, TNF
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Top
Abstract
Introduction
.002). Thus, rhHMGB1 elicits
proinflammatory responses on endothelial cells and may contribute to
alterations in endothelial cell function in human inflammation.
(Blood. 2003;101:2652-2660)
.
