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(Circulation. 1999;99:1560-1566.)
© 1999 American Heart Association, Inc.
Clinical Investigation and Reports |
Endothelial Cytotoxicity Mediated by Serum Antibodies to Heat Shock Proteins of Escherichia coli and Chlamydia pneumoniae
Immune Reactions to Heat Shock Proteins as a Possible Link Between Infection and Atherosclerosis
From the Institute for Biomedical Aging Research, Austrian Academy of Sciences, Innsbruck (M.M., B.M., G.S., Q.X., G.W.); Department of Neurology, University Clinic, Innsbruck (S.K., J.W.); and Institute for General and Experimental Pathology, University of Innsbruck, Medical School, Innsbruck, Austria (M.M., G.W.).
Correspondence to Dr Georg Wick, Institute for Biomedical Aging Research, Austrian Academy of Sciences, Rennweg 10, A-6020 Innsbruck, Austria. E-mail IBA{at}oeaw.ac.at
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Abstract |
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Background—Growing evidence suggests that an immunological reaction against heat shock proteins (HSPs) may be involved in atherogenesis. Because HSPs show a high degree of amino acid sequence homology between different species, from prokaryotes to humans, we investigated the possibility of "antigenic mimicry" caused by an immunological cross-reaction between microorganisms and autoantigens.
Methods and Results—Serum antibodies against the Escherichia coli HSP (GroEL) and the 60-kDa chlamydial HSP (cHSP60) from subjects with atherosclerosis were purified by use of affinity chromatography. Western blot analyses and competitive ELISAs confirmed the cross-reaction of the eluted antibodies with human HSP60 and the bacterial counterparts. The cytotoxicity of anti-GroEL and anti-cHSP60 antibodies was determined on human endothelial cells labeled with 51Cr. A significant difference (40% versus 8%) was observed in the specific 51Cr release of heat-treated (42°C for 30 minutes) and untreated cells, respectively, in the presence of these anti-HSP antibodies and complement. This effect was blocked by addition of 100 µg/mL recombinant GroEL. In addition, seropositivity against specific non-HSP60 Chlamydia pneumoniae antigens is more prominent among high–anti-HSP titer sera than low-titer sera.
Conclusions—Serum antibodies against HSP65/60 cross-react with human HSP60, cHSP60, and GroEL; correlate with the presence of antibodies to C pneumoniae and endotoxin; and mediate endothelial cytotoxicity. These findings suggest that humoral immune reactions to bacterial HSPs, such as cHSP60 and GroEL, may play an important role in the process of vascular endothelial injury, which is believed to be a key event in the pathogenesis of atherosclerosis.
Key Words: stress • immunology • antibodies • C pneumoniae • atherosclerosis
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Introduction |
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TopAbstractIntroductionMethodsResultsDiscussionReferences |
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Recent evidence indicates that the first steps of atherosclerosis are inflammatory in nature.1 The discovery of macrophages, T lymphocytes, dendritic cells, and mast cells in atherosclerotic lesions; the detection of HLA class II antigen expression; and the finding of secretion of several cytokines point to the involvement of immune inflammatory mechanisms in the pathogenesis of atherosclerosis.1 2 Furthermore, atherosclerotic lesions contain immunoglobulin deposits and complement, including the lytic "membrane attack complex" C5b-C9, strongly suggesting the involvement of complement activation in atherogenesis.3 Bacterial and viral infections have been implicated as potential initiating factors. Infections are known to increase blood viscosity, cause hypercoagulability, and influence the serum lipid profile. In infections by Gram-negative bacteria, endotoxin may also contribute to endothelial cell production of free radicals, which may oxidize LDL.4 With respect to potential infectious causes involved in atherosclerosis development, attention has focused on Helicobacter pylori, Chlamydia pneumoniae, and cytomegalovirus.5
A putative antigen maintaining this inflammatory process in the arterial wall must be ubiquitous and present at young age, explaining the overall prevalence of this disease even in young people.6 Heat shock proteins (HSPs) serve as a promising target for inducing an immunological attack on endothelial cells and may constitute the missing link between the incriminated microorganisms and autoimmunity. HSP65/60 is a major antigen recognized during various bacterial infections,7 but stress protein induction also occurs in eukaryotic cells after viral infections.8 Data from our laboratory have shown that atherosclerotic lesions can be induced in normocholesterolemic rabbits by immunization with recombinant mycobacterial HSP65 (mHSP65)9 and inhibited by simultaneous immunosuppressive treatment (Dr Metzler, unpublished data, 1998). Interestingly, mHSP65-reactive T cells were enriched in atherosclerotic lesions compared with peripheral blood even in animals that had received a high cholesterol diet without mHSP65 immunization.10 Furthermore, levels of serum antibodies to mHSP65 were significantly increased in clinically healthy human subjects with carotid atherosclerosis compared with those without lesions, and these increased antibody levels were independent of common risk factors.11
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Methods |
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TopAbstractIntroductionMethodsResultsDiscussionReferences |
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Definition of Clinical and Laboratory Methods
Human sera were derived from the Bruneck Study, a large, population-based study on the epidemiology and origin of atherosclerosis and arterial disease. The Bruneck Study population comprises an age- and sex-stratified random sample of men and women who are 40 to 79 years old (baseline evaluation in 1990). Carotid atherosclerosis was quantified by means of high-resolution duplex ultrasound as extensively described previously.11 12 13 Sera for the current evaluation were drawn as part of the first follow-up study in 1995.
Blood samples were taken from the antecubital vein after subjects had
fasted and abstained from smoking for 
12 hours. Clinic and laboratory
parameters were examined by standard methods as described
previously.12 13 Antibodies against mHSP65 and
Escherichia coli lipopolysaccharides (L 2880, Sigma
Chemical Co) were determined by ELISAs following an established
protocol.11 All "high-titer sera" (HTS) had
anti-mHSP65 antibody titers of 1:1280, whereas "low-titer sera"
(LTS) did not exceed 1:160 (cutoff point for optical density, 0.2 at
410 nm).
Affinity Chromatography of Anti-HSP
Antibodies
Purification of serum anti-HSP antibodies was performed
following an established method.14 Briefly,
immunoglobulins of pooled HTS were precipitated by a standard
(NH4)2SO4
procedure and incubated in a chromatography column with
2 mL agarose gel beads (Affi-Gel Kit, Biorad) coupled with 3 mg
recombinant mHSP65, GroEL, or cHSP60. Specific immunoglobulins were
recovered by 20 mmol/L HCl acid elution, pooled, and equilibrated
with PBS, pH 7.2. Anti-HSP antibody titers of purified immunoglobulins
were similar to original HTS (1:1280), whereas unbound immunoglobulin
had no measurable HSP antibody titer (<1:20).
Western Blotting
This procedure was also described previously.14 In
brief, samples were diluted 1:20 (vol/vol) in sample buffer containing
5% 2-mercaptoethanol, 15% glycerol, 3% SDS, and 0.1 mol/L Tris, pH
6.8, and separated on a 12% polyacrylamide gel under reducing
conditions. Blots were probed with anti-HSP antibodies (5 µg/mL),
including purified anti-HSP antibodies, HTS, and the monoclonal
antibody II-13 (a gift from Dr R.S. Gupta, Hamilton, Canada). Reactions
were visualized by an enhanced chemiluminescence detection kit
(Amersham) after incubation with a peroxidase rabbit anti-human or
anti-mouse immunoglobulin conjugate.
Endothelial Cytotoxicity Assays
Human umbilical vein endothelial cells
(HUVECs) were cultured and identified as described
previously.14 Human aortic endothelial
cells were purchased from Cascade Biologics. For antibody-mediated
cytotoxicity, endothelial cells were heat stressed at
42°C for 30 minutes to induce HSP60 expression, whereas control cells
remained at 37°C. The assay was performed following an established
protocol14 and is described in brief in the legend of
Figure 5
. Supernatant was analyzed for
51Cr-radioactivity (Amersham) in a gamma counter
(Wallac-Wizzard Automatic Gamma Counter). Both human, including LTS and
unbound immunoglobulin fraction, and monoclonal mouse antibodies,
including mAb anti–
-actin (catalog No. 1148818, Boehringer,
Mannheim, FRG), mAb anti-CD3 (catalog No. M756, Dakopatts), and mAb
anti-ßHCG 58 prepared in our laboratory, were used as controls in the
cytotoxicity tests. Blocking experiments were performed by addition of
anti-HSP antibody preincubated with 100 µg/mL soluble recombinant
GroEL for 1 hour at room temperature. Specifically released
radioactivity was determined by calculation according to the following
formula: 51Cr release in the presence of
antibodies minus spontaneous release divided by maximal release minus
spontaneous release. Maximal release was obtained by adding 5% Triton
X-100 to the cultures. Spontaneous release was determined in the
presence of complement without antibodies and did not exceed 15% of
maximal release.
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Statistical Analysis
The association between anti-HSP antibody titers (category 
160
versus 1280) and
atherosclerosis/cardiovascular disease
was assessed by means of unconditional logistic regression
analysis. ANOVA was performed when >2 groups were compared.
Paired Student's t test was used to assess differences
between 2 groups. A value of P<0.05 was considered
significant.
Specific Tests for C pneumoniae
IgG antibodies to C pneumoniae were
measured by 3 different test systems: SeroCP-IgG (Savyon
Diagnostics Ltd),
microimmunofluorescence (MIF) antibody kit
(Labsystems Oy), and chlamydial IgG recombinant lipopolysaccharide
(LPS) ELISA (Medac). The analyses were performed and
calculated according to manufacturers' instructions. In contrast to
the former test systems that used elementary bodies, a recombinant
chlamydial LPS antigen was used for precoating in the last ELISA
kit.
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Results |
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TopAbstractIntroductionMethodsResultsDiscussionReferences |
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Clinical Characterization of Serum Samples
A previous analysis in the Bruneck Study population has shown that increased antibody titers to HSPs were independent of classic risk factors for atherosclerosis (n=867).11 On comparison of subjects with antibody titers
160 and those with titers 1280, the latter group faced a 2- to
3-fold increased risk of prevalent carotid
atherosclerosis (unadjusted OR, 2.44; 95% CI, 1.56 to
3.80; P<0.0001; adjusted OR, 2.48; 95% CI, 1.58 to 3.89;
P<0.0001). Similar results were obtained when prevalent
cardiovascular disease was used as the outcome
variable (unadjusted OR, 3.63; 95% CI, 1.66 to 7.94;
P=0.0013; adjusted OR, 4.35; 95% CI, 1.93 to 9.84;
P=0.0004). In this analysis,
cardiovascular disease encompasses myocardial
infarction, ischemic stroke, transient ischemic attack,
and symptomatic peripheral artery disease.
Forty sera with known high or low anti-mHSP65 titers (HTS 1280/LTS
160) were selected from this population for the present study. No
significant difference in classic risk factors for
atherosclerosis was observed between these 2 groups
(Table 1). In addition, HSP
antibody titers did not show a significant correlation with C-reactive
protein and blood granulocyte count.
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Cross-Reactivity of Serum Antibodies Against HSPs
The specimens (HTS/LTS) were tested for their reactivity
against the E coli HSP60 GroEL and the HSP60 of C
trachomatis, which is nearly identical to the HSP of C
pneumoniae (amino acid sequence homology, 
97%). Comparison of
these antibody titers to the anti-mHSP65 titer revealed an overall
correlation of 90% for GroEL and 75% for cHSP60. The correlation
coefficient between the titers of anti-GroEL and anti-cHSP60 antibodies
was 0.7 (Figure 1).
|
) or low (
) anti-mHSP65
titers were tested for their reactivity to GroEL and cHSP60 in standard
ELISA procedure. First 2 scatter plots show correlation
analyses of these latter antibody titers to anti-mHSP65 titer.
Data revealed correlation of
We then purified these polyclonal anti-HSP antibodies from pooled HTS
by affinity chromatography. The eluted antibodies were
capable of recognizing their human and bacterial counterparts, as
demonstrated by Western blot analyses (Figure 2). However, the strongest reaction was
consistently found with the HSP of E coli, GroEL.
Ovalbumin, free of HSP but with an identical molecular weight
of 60 kDa, served as negative control and did not interact with the
purified antibodies, although it was still recognized by HTS because of
anti-ovalbumin antibodies, as confirmed by ELISA experiments.
As positive control, a monoclonal antibody to HSP60 (II-13) stained all
types of HSPs but not ovalbumin. The cross-reactivity and
specificity of the various purified antibodies were confirmed by
competitive ELISA experiments (Figure 3).
Complete inhibition was achieved only with the HSP used for antibody
purification. For anti-GroEL and anti-mHSP65 antibodies, inhibition
with cHSP60 was significantly less effective compared with the
bacterial counterparts (P<0.05).
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), GroEL (
), respectively, for 1 hour at room temperature and
tested on ELISA plates coated with 1 µg/mL of same protein used for
antibody purification. BSA (x), determined to be free of
HSPs, served as negative control.
Reactivity of Serum Antibodies to Non-HSP60 Bacterial
Antigens
Serum samples were assayed for C pneumoniae IgG
antibodies by use of an MIF test and 2 commercial ELISA kits. The
purified anti-HSP antibodies did not appear to cross-react with these
specific C pneumoniae antigens. Data from all tests revealed
that seropositivity against non-HSP60 C pneumoniae antigens
was more prominent among HTS than LTS (Table 2). The seroprevalence of IgG
antibodies among LTS was 77% all together, but only 31% within this
group were considered highly positive in the SeroCP ELISA system. In
contrast, all HTS had detectable IgG antibodies; surprisingly, 77%
were also highly reactive against C pneumoniae elementary
bodies. The mean optical density for LTS was calculated as 2.07,
interpreted as positive. The mean optical density for HTS reached 2.88,
considered highly positive. MIF data afforded a quantitative
determination of IgG antibody responses in both groups. Sera with
undetectable IgG titers (<1:32) occurred more often among LTS, whereas
medium- (1:32) and high- (1:128) titer sera were more frequent among
HTS. Acute, primary infections with C pneumoniae could be
ruled out, because none of the serum samples showed a positive reaction
for IgM antibodies after absorption with anti-human IgG. Data of the
recombinant chlamydial LPS ELISA correlated well with the MIF test and
provided clear-cut results: 54% of LTS were negative but about 55% of
HTS reached titers of 1:200. In parallel to elevated anti-GroEL
antibodies, we also detected a higher antibody prevalence against
E coli LPS in HTS (Figure 4). Statistical
analysis revealed a significant correlation between anti-LPS
antibodies and anti-HSP titers (r=0.59,
P<0.001).
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Endothelial Cytotoxicity
All purified anti-HSP antibodies exerted effective
complement-mediated endothelial cytotoxicity on
stressed HUVECs. A specific dose-dependent release of
51Cr from heat-treated cells was observed in the
presence of anti-GroEL and anti-cHSP60 antibodies, with a dramatic
increase after antibody concentrations rose >25 µg/mL (Figure 5
). Lysis reached 70% in stressed
endothelial cells, whereas an average of only 10% of
total radioactivity was released from unstressed cells in the presence
of anti-HSP antibodies. This striking effect on stressed
endothelial cells could be blocked by the addition of
100 µg/mL recombinant GroEL (Figure 6).
Even the anti-cHSP60 antibody lost its cytotoxic potential after
preincubation with GroEL. Unbound immunoglobulins from the affinity
chromatographic procedure, LTS, and unrelated monoclonal
antibodies evoked no significant cytotoxic effect. In preliminary
experiments, anti-GroEL and anti-cHSP60 antibodies were also able to
lyse stressed endothelial cells in the presence of high
concentrations of normal human peripheral blood mononuclear
cells at an effector-to-target ratio of 1:50 via antibody-dependent
cellular cytotoxicity (data not shown). To determine whether such an
antibody-mediated cytotoxicity also occurs in arterial
endothelial cells, human aortic cells and HUVECs were
cultivated and treated in parallel. Although no significant difference
in the cytotoxic lysis between both types of
endothelial cells was found, anti-mHSP65 antibodies
markedly induced 51Cr release from heat-stressed
cells (Figure 7). In addition, heat
stress treatment at 42°C for 30 minutes did not result in
endothelial cell necrosis or apoptosis (data
not shown).
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Discussion |
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TopAbstractIntroductionMethodsResultsDiscussionReferences |
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We previously demonstrated a strong association between sonographically detectable carotid atherosclerosis and high levels of serum antibodies to mHSP65 in the Bruneck Study population (n=867).11 Bacterial and viral infections can induce immune reactions against HSPs, which may serve as targets for autoimmune reactions.15 The anti-mHSP65 antibodies react not only with mHSP65 but also with human HSP60 and the bacterial counterparts of E coli and C pneumoniae. Herein, we demonstrate that several epitopes of bacterial HSPs seem to exhibit cross-reactive antibody binding. These findings could signify a possible involvement of infections in atherogenesis, because the cross-reactive epitopes may contribute to the elevated anti-mHSP65 antibodies in subjects with atherosclerosis. The natural microbial flora may also influence such immune reactions to various degrees, eg, GroEL of E coli. However, competitive ELISAs indicate that some additional specific epitopes must exist.
Antibodies to mHSP65 can be induced by several different mechanisms in vivo. First, infectious agents containing homologous HSP60 proteins could induce an anti–self-immune response through molecular mimicry in susceptible individuals.16 Additionally, vaccination (particularly with heat-inactivated bacteria) may contribute to the development of anti-HSP antibodies. Second, viral infection might result in incorporation of HSP60, or at least of HSP-derived peptides, into the envelope of the budding virus.17 Arising from viral infections, HSP60 could also become immunogenic as a result of structural alteration or posttranslational modification.18 Third, exposure to endogenous HSPs or mimicry proteins might induce immune reactions against HSP60, or endogenous HSPs might interact with other immunogenic proteins.19
Proteins of the HSP60 family were considered to be located intracellularly in mitochondria only, where they facilitate protein translocation and act as chaperones protecting proteins from harmful enzymatic attacks during folding. Accumulating evidence now points to an additional surface location of HSP60 proteins on not only eukaryotic20 21 but also prokaryotic cells, such as H pylori.22 Immune reactions against HSPs could therefore provide a general basic level of protection and limit the spread of infections by acting as an initial defense mechanism that immediately interacts with microbial invaders. In contrast to this benefit, the risk of autoimmunity must be taken into consideration, because HSPs are also induced, eg, on endothelial cells by other exogenous and endogenous stimuli. The endothelium, as a barrier between blood and underlying cells, is subjected to continuous mechanical stress from blood pressure and acute or chronic injury from toxins, such as LPS and importantly oxidized LDL, a known risk factor in atherosclerosis.23 These stressors and many others, such as cytokines24 and fever, all induce or augment HSP production by the arterial endothelium to prevent cell damage and maintain homeostasis in the vessel wall.25 Temperatures of 42°C, as used in our experiments, are indeed encountered in a setting of infection. They are rarely achieved at the body core but might be reached in an inflamed atherosclerotic lesion. Thus, preexisting antibodies and HSP60-specific T cells could react with overexpressed HSP60 components in atherosclerotic lesions,26 27 causing endothelial14 and macrophage injury28 and subsequently contributing to the initiation and perpetuation of inflammation.29 The in vitro phenomenon of anti-HSP serum antibodies also exerting a weak cytotoxic effect on unstressed endothelial cells may be explained by a basal level of HSP60 expression, particularly because in vitro culture is itself stressful to cells compared with in vivo conditions.14
Because persistent chlamydial infections are associated with decreased expression of protective antigens, such as the major outer membrane protein and LPS, but an abundance of the immunopathogenic HSP60,30 they may contribute to the elevated serum antibody levels against mHSP65 found in patients with clinically asymptomatic carotid atherosclerosis.11 In fact, our data provide evidence that sera with high titers to mHSP65 are highly reactive against C pneumoniae. Additionally, cHSP60 synthesis in infected cells is likely to be accompanied by an increase in human HSP60 production, because chlamydial infection extensively stresses host cells.31 cHSP60 appears to be the key antigen in eliciting a delayed hypersensitivity reaction of the host in blinding trachoma or fallopian tube obstruction, resulting in severe tissue damage from C trachomatis infections with interesting similarities to atherosclerosis.32 Immunopathology plays a decisive role in chlamydial disease, but anti-HSP60 antibody–mediated immunity may be a possible general link between infections and atherogenesis.
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Acknowledgments |
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This work was supported by grant P-12213-MED (Dr Wick) from the Austrian Science Fund.
Received July 1, 1998; revision received December 22, 1998; accepted January 8, 1999.
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 Q. Xiao, K. Mandal, G. Schett, M. Mayr, G. Wick, F. Oberhollenzer, J. Willeit, S. Kiechl, and Q. Xu Association of Serum-Soluble Heat Shock Protein 60 With Carotid Atherosclerosis: Clinical Significance Determined in a Follow-Up Study Stroke, December 1, 2005; 36(12): 2571 - 2576. [Abstract] [Full Text] [PDF]
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 E Toubi and Y Shoenfeld Predictive and protective autoimmunity in cardiovascular diseases: is vaccination therapy a reality? Lupus, September 1, 2005; 14(9): 665 - 669. [Abstract] [PDF]
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K Mandal, G Foteinos, M Jahangiri, and Q Xu Role of antiheat shock protein 60 autoantibodies in atherosclerosis Lupus, September 1, 2005; 14(9): 742 - 746. [Abstract] [PDF]
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 A G Pockley and J Frostegard Heat shock proteins in cardiovascular disease and the prognostic value of heat shock protein related measurements Heart, September 1, 2005; 91(9): 1124 - 1126. [Abstract] [Full Text] [PDF]
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C. Espinola-Klein, S. Blankenberg, and T. Munzel Editorial Comment--Is Heme Oxygenase-1 a Causal Player for Plaque Stability? Stroke, September 1, 2005; 36(9): 1901 - 1903. [Full Text] [PDF]
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G. Foteinos, A. R. Afzal, K. Mandal, M. Jahangiri, and Q. Xu Anti-Heat Shock Protein 60 Autoantibodies Induce Atherosclerosis in Apolipoprotein E-Deficient Mice via Endothelial Damage Circulation, August 23, 2005; 112(8): 1206 - 1213. [Abstract] [Full Text] [PDF]
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 M. Ghayour-Mobarhan, S. A New, D. J Lamb, B. J Starkey, C. Livingstone, T. Wang, N. Vaidya, and G. A Ferns Dietary antioxidants and fat are associated with plasma antibody titers to heat shock proteins 60, 65, and 70 in subjects with dyslipidemia Am. J. Clinical Nutrition, May 1, 2005; 81(5): 998 - 1004. [Abstract] [Full Text] [PDF]
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 J.-P. Carralot, C. Dumrese, R. Wessel, R. Riessen, I. Autenrieth, S. Walter, O. Schoor, S. Stevanovic, H.-G. Rammensee, and S. Pascolo CD8+ T cells specific for a potential HLA-A*0201 epitope from Chlamydophila pneumoniae are present in the PBMCs from infected patients Int. Immunol., May 1, 2005; 17(5): 591 - 597. [Abstract] [Full Text] [PDF]
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 G. Pfister, C. M. Stroh, H. Perschinka, M. Kind, M. Knoflach, P. Hinterdorfer, and G. Wick Detection of HSP60 on the membrane surface of stressed human endothelial cells by atomic force and confocal microscopy J. Cell Sci., April 15, 2005; 118(8): 1587 - 1594. [Abstract] [Full Text] [PDF]
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 M. Ghayour-Mobarhan, D. J. Lamb, N. Vaidya, C. Livingstone, T. Wang, and G. A. A. Ferns Heat Shock Protein Antibody Titers Are Reduced by Statin Therapy in Dyslipidemic Subjects: A Pilot Study Angiology, January 1, 2005; 56(1): 61 - 68. [Abstract] [PDF]
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M. Mayr, K. Mandal, Q. Xu, J. S. Lindholt, J. Stovring, S. Urbonavicius, E. W. Henneberg, L. Ostergaard, B. Honore, and H. Vorum Pitfalls of Proteomics * Response Circulation, September 21, 2004; 110(12): e316 - e316. [Full Text] [PDF]
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J. Zhu, R. J. Katz, A. A. Quyyumi, D. A. Canos, D. Rott, G. Csako, A. Zalles-Ganley, J. Ogunmakinwa, A. G. Wasserman, and S. E. Epstein Association of Serum Antibodies to Heat-Shock Protein 65 With Coronary Calcification Levels: Suggestion of Pathogen-Triggered Autoimmunity in Early Atherosclerosis Circulation, January 6, 2004; 109(1): 36 - 41. [Abstract] [Full Text] [PDF]
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 S. Hirono, E. Dibrov, C. Hurtado, A. Kostenuk, R. Ducas, and G. N. Pierce Chlamydia pneumoniae Stimulates Proliferation of Vascular Smooth Muscle Cells Through Induction of Endogenous Heat Shock Protein 60 Circ. Res., October 17, 2003; 93(8): 710 - 716. [Abstract] [Full Text] [PDF]
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 E. Lalla, I. B. Lamster, M. A. Hofmann, L. Bucciarelli, A. P. Jerud, S. Tucker, Y. Lu, P. N. Papapanou, and A. M. Schmidt Oral Infection With a Periodontal Pathogen Accelerates Early Atherosclerosis in Apolipoprotein E-Null Mice Arterioscler Thromb Vasc Biol, August 1, 2003; 23(8): 1405 - 1411. [Abstract] [Full Text] [PDF]
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 R. Elhage, J. Jawien, M. Rudling, H.-G. Ljunggren, K. Takeda, S. Akira, F. Bayard, and G. K Hansson Reduced atherosclerosis in interleukin-18 deficient apolipoprotein E-knockout mice Cardiovasc Res, July 1, 2003; 59(1): 234 - 240. [Abstract] [Full Text] [PDF]
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L. M. Biasucci, G. Liuzzo, A. Ciervo, A. Petrucca, M. Piro, D. J. Angiolillo, F. Crea, A. Cassone, and A. Maseri Antibody Response to Chlamydial Heat Shock Protein 60 Is Strongly Associated With Acute Coronary Syndromes Circulation, June 24, 2003; 107(24): 3015 - 3017. [Abstract] [Full Text] [PDF]
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H. Perschinka, M. Mayr, G. Millonig, C. Mayerl, R. van der Zee, S. G. Morrison, R. P. Morrison, Q. Xu, and G. Wick Cross-Reactive B-Cell Epitopes of Microbial and Human Heat Shock Protein 60/65 in Atherosclerosis Arterioscler Thromb Vasc Biol, June 1, 2003; 23(6): 1060 - 1065. [Abstract] [Full Text] [PDF]
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G. G. L. Biondi-Zoccai, A. Abbate, G. Liuzzo, and L. M. Biasucci Atherothrombosis, inflammation, and diabetes J. Am. Coll. Cardiol., April 2, 2003; 41(7): 1071 - 1077. [Abstract] [Full Text] [PDF]
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 M. V. Kalayoglu, C. Galvan, O. S. Mahdi, G. I. Byrne, and S. Mansour Serological Association Between Chlamydia pneumoniae Infection and Age-Related Macular Degeneration Arch Ophthalmol, April 1, 2003; 121(4): 478 - 482. [Abstract] [Full Text] [PDF]
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M. Mayr, S. Kiechl, M. A. Mendall, J. Willeit, G. Wick, and Q. Xu Increased Risk of Atherosclerosis Is Confined to CagA-Positive Helicobacter pylori Strains: Prospective Results From the Bruneck Study Stroke, March 1, 2003; 34(3): 610 - 615. [Abstract] [Full Text] [PDF]
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 M. V. Kalayoglu, P. Libby, and G. I. Byrne Chlamydia pneumoniae as an Emerging Risk Factor in Cardiovascular Disease JAMA, December 4, 2002; 288(21): 2724 - 2731. [Abstract] [Full Text] [PDF]
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Q. Xu Role of Heat Shock Proteins in Atherosclerosis Arterioscler Thromb Vasc Biol, October 1, 2002; 22(10): 1547 - 1559. [Abstract] [Full Text] [PDF]
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O. S. Mahdi, B. D. Horne, K. Mullen, J. B. Muhlestein, and G. I. Byrne Serum Immunoglobulin G Antibodies to Chlamydial Heat Shock Protein 60 but Not to Human and Bacterial Homologs Are Associated With Coronary Artery Disease Circulation, September 24, 2002; 106(13): 1659 - 1663. [Abstract] [Full Text] [PDF]
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R. Maron, G. Sukhova, A.-M. Faria, E. Hoffmann, F. Mach, P. Libby, and H. L. Weiner Mucosal Administration of Heat Shock Protein-65 Decreases Atherosclerosis and Inflammation in Aortic Arch of Low-Density Lipoprotein Receptor-Deficient Mice Circulation, September 24, 2002; 106(13): 1708 - 1715. [Abstract] [Full Text] [PDF]
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 J C Ranford and B Henderson Chaperonins in disease: mechanisms, models, and treatments Mol. Pathol., August 1, 2002; 55(4): 209 - 213. [Abstract] [Full Text] [PDF]
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W. Koenig, N. Khuseyinova, M. M. Hoffmann, W. Marz, M. Frohlich, A. Hoffmeister, H. Brenner, and D. Rothenbacher CD14 C(-260)->T polymorphism, plasma levels of the soluble endotoxin receptor CD14, their association with chronic infections and risk of stable coronary artery disease J. Am. Coll. Cardiol., July 3, 2002; 40(1): 34 - 42. [Abstract] [Full Text] [PDF]
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 C. Stollberger and J. Finsterer Role of Infectious and Immune Factors in Coronary and Cerebrovascular Arteriosclerosis Clin. Vaccine Immunol., March 1, 2002; 9(2): 207 - 215. [Full Text] [PDF]
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 L.C. von Hertzen Role of persistent infection in the control and severity of asthma: focus on Chlamydia pneumoniae Eur. Respir. J., March 1, 2002; 19(3): 546 - 556. [Abstract] [Full Text] [PDF]
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T. Huittinen, M. Leinonen, L. Tenkanen, M. Manttari, H. Virkkunen, T. Pitkanen, E. Wahlstrom, T. Palosuo, V. Manninen, and P. Saikku Autoimmunity to Human Heat Shock Protein 60, Chlamydia pneumoniae Infection, and Inflammation in Predicting Coronary Risk Arterioscler Thromb Vasc Biol, March 1, 2002; 22(3): 431 - 437. [Abstract] [Full Text] [PDF]
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A. G. Pockley Heat Shock Proteins, Inflammation, and Cardiovascular Disease Circulation, February 26, 2002; 105(8): 1012 - 1017. [Full Text] [PDF]
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 Y. Bulut, E. Faure, L. Thomas, H. Karahashi, K. S. Michelsen, O. Equils, S. G. Morrison, R. P. Morrison, and M. Arditi Chlamydial Heat Shock Protein 60 Activates Macrophages and Endothelial Cells Through Toll-Like Receptor 4 and MD2 in a MyD88-Dependent Pathway J. Immunol., February 1, 2002; 168(3): 1435 - 1440. [Abstract] [Full Text] [PDF]
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S. E. Epstein The Multiple Mechanisms by Which Infection May Contribute to Atherosclerosis Development and Course Circ. Res., January 11, 2002; 90(1): 2 - 4. [Full Text] [PDF]
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A. Ciervo, P. Visca, A. Petrucca, L. M. Biasucci, A. Maseri, and A. Cassone Antibodies to 60-Kilodalton Heat Shock Protein and Outer Membrane Protein 2 of Chlamydia pneumoniae in Patients with Coronary Heart Disease Clin. Vaccine Immunol., January 1, 2002; 9(1): 66 - 74. [Abstract] [Full Text] [PDF]
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G. K. Hansson Immune Mechanisms in Atherosclerosis Arterioscler Thromb Vasc Biol, December 1, 2001; 21(12): 1876 - 1890. [Abstract] [Full Text] [PDF]
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J. George, A. Afek, B. Gilburd, Y. Shoenfeld, and D. Harats Cellular and humoral immune responses to heat shock protein 65 are both involved in promoting fatty-streak formation in LDL-receptor deficient mice J. Am. Coll. Cardiol., September 1, 2001; 38(3): 900 - 905. [Abstract] [Full Text] [PDF]
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H. J. Rupprecht, S. Blankenberg, C. Bickel, G. Rippin, G. Hafner, W. Prellwitz, W. Schlumberger, and J. Meyer Impact of Viral and Bacterial Infectious Burden on Long-Term Prognosis in Patients With Coronary Artery Disease Circulation, July 3, 2001; 104(1): 25 - 31. [Abstract] [Full Text] [PDF]
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G. Caligiuri, M. Rottenberg, A. Nicoletti, H. Wigzell, and G. K. Hansson Chlamydia pneumoniae Infection Does Not Induce or Modify Atherosclerosis in Mice Circulation, June 12, 2001; 103(23): 2834 - 2838. [Abstract] [Full Text] [PDF]
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T. Huittinen, D. Hahn, T. Anttila, E. Wahlstrom, P. Saikku, and M. Leinonen Host immune response to Chlamydia pneumoniae heat shock protein 60 is associated with asthma Eur. Respir. J., June 1, 2001; 17(6): 1078 - 1082. [Abstract] [Full Text] [PDF]
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K. Burian, Z. Kis, D. Virok, V. Endresz, Z. Prohaszka, J. Duba, K. Berencsi, K. Boda, L. Horvath, L. Romics, et al. Independent and Joint Effects of Antibodies to Human Heat-Shock Protein 60 and Chlamydia pneumoniae Infection in the Development of Coronary Atherosclerosis Circulation, March 20, 2001; 103(11): 1503 - 1508. [Abstract] [Full Text] [PDF]
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S. Kiechl, G. Egger, M. Mayr, C. J. Wiedermann, E. Bonora, F. Oberhollenzer, M. Muggeo, Q. Xu, G. Wick, W. Poewe, et al. Chronic Infections and the Risk of Carotid Atherosclerosis : Prospective Results From a Large Population Study Circulation, February 27, 2001; 103(8): 1064 - 1070. [Abstract] [Full Text] [PDF]
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J. Zhu, A. A. Quyyumi, D. Rott, G. Csako, H. Wu, J. Halcox, and S. E. Epstein Antibodies to Human Heat-Shock Protein 60 Are Associated With the Presence and Severity of Coronary Artery Disease : Evidence for an Autoimmune Component of Atherogenesis Circulation, February 27, 2001; 103(8): 1071 - 1075. [Abstract] [Full Text] [PDF]
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 S A Morre, W Stooker, W K Lagrand, A J C van den Brule, and H W M Niessen Microorganisms in the aetiology of atherosclerosis J. Clin. Pathol., September 1, 2000; 53(9): 647 - 654. [Abstract] [Full Text] [PDF]
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M. Mayr, S. Kiechl, J. Willeit, G. Wick, and Q. Xu Infections, Immunity, and Atherosclerosis : Associations of Antibodies to Chlamydia pneumoniae, Helicobacter pylori, and Cytomegalovirus With Immune Reactions to Heat-Shock Protein 60 and Carotid or Femoral Atherosclerosis Circulation, August 22, 2000; 102(8): 833 - 839. [Abstract] [Full Text] [PDF]
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Q. Xu, G. Schett, H. Perschinka, M. Mayr, G. Egger, F. Oberhollenzer, J. Willeit, S. Kiechl, and G. Wick Serum Soluble Heat Shock Protein 60 Is Elevated in Subjects With Atherosclerosis in a General Population Circulation, July 4, 2000; 102(1): 14 - 20. [Abstract] [Full Text] [PDF]
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C. Schmidt, J. Hulthe, J. Wikstrand, H. Gnarpe, J. Gnarpe, S. Agewall, and B. Fagerberg Chlamydia pneumoniae Seropositivity Is Associated With Carotid Artery Intima-Media Thickness Stroke, July 1, 2000; 31(7): 1526 - 1531. [Abstract] [Full Text] [PDF]
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S. E. Epstein, J. Zhu, M. S. Burnett, Y. F. Zhou, G. Vercellotti, and D. Hajjar Infection and Atherosclerosis : Potential Roles of Pathogen Burden and Molecular Mimicry Arterioscler Thromb Vasc Biol, June 1, 2000; 20(6): 1417 - 1420. [Abstract] [Full Text] [PDF]
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 T. Quaschning and C. Wanner The role of Chlamydia in coronary heart disease--fact or fiction? Nephrol. Dial. Transplant., December 1, 1999; 14(12): 2800 - 2803. [Full Text] [PDF]
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C. A. Gunnett, D. J. Berg, F. M. Faraci, and G. Feuerstein Vascular Effects of Lipopolysaccharide Are Enhanced in Interleukin-10-Deficient Mice • Editorial Comment Stroke, October 1, 1999; 30(10): 2191 - 2196. [Abstract] [Full Text] [PDF]
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Q. Xu, S. Kiechl, M. Mayr, B. Metzler, G. Egger, F. Oberhollenzer, J. Willeit, and G. Wick Association of Serum Antibodies to Heat-Shock Protein 65 With Carotid Atherosclerosis : Clinical Significance Determined in a Follow-Up Study Circulation, September 14, 1999; 100(11): 1169 - 1174. [Abstract] [Full Text] [PDF]
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