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(Circulation. 1997;96:2144-2148.)
© 1997 American Heart Association, Inc.

Articles

Chlamydia Pneumoniae, Cytomegalovirus, and Herpes Simplex Virus in Atherosclerosis of the Carotid Artery

B. Chiu, MD, FRCPC; E. Viira, BSc; W. Tucker, MD, FRCPC; ; I.W. Fong, MB, BS, FRCP(C)

From the Departments of Medicine (I.W.F.), Pathology (B.C.), and Surgery (Neurosurgery) (W.T.), St Michael's Hospital, University of Toronto, Toronto, Ontario, Canada.

	Abstract

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Background Chlamydia pneumoniae and the herpes viruses cytomegalovirus (CMV) and herpes simplex virus type 1 (HSV-1) have been associated with human atherosclerosis in seroepidemiological and separate histopathological studies. We investigated the concurrent presence of these microorganisms in patients undergoing carotid endarterectomy.

Methods and Results Endarterectomy specimens from 76 patients with carotid artery stenosis were stained for C pneumoniae, CMV, and HSV-1 particles with specific IgG monoclonal antibodies by the avidin-biotin-peroxidase method. IgG antibodies to CMV and C pneumoniae were also measured in the serum. These were correlated with plaque morphology and the presence of the microorganisms in the atherosclerotic plaques. C pneumoniae was detected in 54 (71%) (95% confidence interval [CI], 59.5% to 80.9%), CMV was detected in 27 (35.5%) (CI, 24.9% to 47.3%), and HSV-1 was detected in 8 (10.5%) (CI, 4.7% to 19.7%) versus none of 20 (0%) control normal carotid artery and aortic tissue (autopsy) specimens (CI, 0% to 16.8%) (P<.001 for CMV and C pneumoniae). At least one microorganism was detected in 59 of the specimens (77.6%) (CI, 66.6% to 86.4%), with a single microorganism present only in 35 (46%), two microorganisms present in 18 (23.7%) (CI, 14.7% to 34.8%), and all three present in 6 (7.9%) (CI, 3.0% to 16.4%). Atherosclerotic plaques with thrombosis were more likely to have C pneumoniae (80.4%) or CMV (57.8%) than were plaques without thrombosis (56.7% and 16.7%, respectively; P=.04 and .007). There was no correlation between the presence of CMV and C pneumoniae in the atherosclerotic vessels and serum antibody titers.

Conclusions C pneumoniae and CMV are commonly detected in atherosclerotic plaques of the carotid arteries, but their presence cannot be predicted by measuring serum antibodies. The presence of these microorganisms may predispose to a greater risk of thrombosis in the plaques, but further studies are needed to confirm this observation.

Key Words: atherosclerosis • immunohistochemistry • carotid arteries

	Introduction

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There is accumulating evidence that certain infectious agents play a role in the pathogenesis of atherosclerosis. Infectious agents are well established causes of heart disease such as pericarditis, myocarditis, and endocarditis. Moreover, an infectious cause for atherosclerosis fits into the currently accepted response-to-injury model of atherogenesis. A viral role in atherogenesis was proposed >20 years ago.^{1 2 3} There also is circumstantial evidence that cytomegalovirus (CMV) and herpes simplex virus (HSV) can contribute to the pathogenic events associated with atherosclerosis. These lines of evidence include seroepidemiological links between CMV infections in normal hosts^{3 4} and cardiac transplants^{5 6} and histopathological evidence (immunocytochemistry, DNA hybridization, or polymerase chain reaction [PCR]) of the presence of CMV or HSV particles in the atherosclerotic vessels of humans.^{7 8 9 10 11 12} The strongest evidence of a pathogenic role of CMV in atherosclerosis, however, is in cardiac transplantation.^{5 6} Moreover, in experimental animals, an avian herpes virus (Marek's disease virus) can induce atherosclerosis in chickens,^{2 13} and immunization of the chickens can prevent atherosclerosis. However, no human virus has been shown to produce atherosclerosis in an animal model.

Similarly, there is increasing evidence that Chlamydia pneumoniae, a common respiratory tract pathogen, may play a role in atherogenesis. C pneumoniae has been associated with coronary and carotid artery disease in seroprevalence epidemiological studies^{14 15 16 17 18 19 20} and in one prospective cohort study.²¹ In one seroepidemiological study, both Helicobacter pylori and C pneumoniae infections were associated with coronary heart disease.¹⁸ In addition, C pneumoniae elementary bodies have been detected in the atherosclerotic plaques and fatty streaks of the aorta and coronary arteries of autopsy cases,^{22 23 24} from atherectomy specimens of coronary arteries,²⁵ and from endarterectomy specimens of carotid arteries.^{26 27} Although C pneumoniae has been found in atherosclerotic plaques by several investigators, H pylori has not been detected in atherosclerotic arteries.²⁸ We examined the role of concurrent infection with the herpes viruses and C pneumoniae in atherosclerotic carotid arteries.

	Methods

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This study was approved by our ethics review committee, and informed consent was obtained from each patient participating in the study.

Population
Patients with significant carotid artery stenosis (>60% narrowing of the lumen) who required surgical endarterectomy were enrolled in the study. Patients with familial hyperlipidemia were excluded. Demographic information included history of hypertension, diabetes mellitus, angina, heart attack, or cerebrovascular accidents; obesity; cigarette smoking; chronic bronchitis or respiratory infection; hypercholesterolemia; and a family history of heart disease or stroke.

Serology
Serum was obtained before surgery for the measurement of antibodies to C pneumoniae and CMV. C pneumoniae antibodies (IgG, IgA, IgM) were measured by the microimmunofluorescence test, with formalin-fixed whole elementary bodies of C pneumoniae (AR-39) as antigen²⁹ (MRL Diagnostics). Immunoglobulin and serum antibody fractions were measured by using fluorescein isothiocyanate–conjugated goat anti-human IgG, IgA, and IgM (Sigma Chemical). Sera were screened for antibody at a 1:16 dilution, and those positive were titered by twofold dilutions to 1:512. Sera were screened for CMV IgG antibodies with an ELISA (IMX CMV MEIA assay, Abbott Labs), and the positive samples were quantified by serial dilutions to 1:64 using the CMV scan latex agglutination (Becton Dickinson).

Pathology
Carotid endarterectomy specimens obtained at surgery were fixed in 10% buffered formalin, decalcified, embedded in paraffin, sectioned, and stained with hematoxylin and eosin for histological examination. Movat's pentachrome stain was also used in the study of plaque morphology. Plaque ulcers were defined as depressions in the plaque with discontinued endothelial lining. Thrombus was defined as arising from the ulcerated plaque or adherent to the discontinued endothelial lining of the intima as defined by Van Damme et al,³⁰ excluding those formed within intraplaque hemorrhage. Immunocytochemical staining was performed on paraffin-embedded sections according to the avidin-biotin-peroxidase method.²³ Briefly, the sections were deparaffinized, rehydrated, and treated with 3% hydrogen peroxidase for 10 minutes to block endogenous peroxidase activity. The sections were then digested with 0.5% pronase (Sigma) for 10 minutes at room temperature, rinsed, and incubated with normal sera of both rabbit and mouse for 20 minutes. After draining of the excess serum, the sections were incubated overnight at 4°C with the following antisera (supplier and dilution): C pneumoniae–specific monoclonal IgG antibodies (Chlamydia-Cel-Pn; no dilution) and TT-401 (IgG) (Washington Research Foundation; 1:100), CMV IgG monoclonal antibody to the early antigen (DAKO; 1:30), HSV-1 IgG monoclonal antibody (DAKO; 1:250), factor VIII for endothelial cells (DAKO; 1:500), smooth muscle actin for smooth muscle cells (DAKO; 1:150), and CD68 for macrophages (DAKO; 1:70). A biotinylated rabbit anti-mouse secondary antibody (DAKO) was used followed by avidin-biotin complex (Vector Laboratories) for 30 minutes. The chromogen was 0.06% 3,3'-diaminobenzidine. Twenty control samples from normal carotid arteries and aorta without atherosclerosis (autopsy samples) were examined in parallel. A negative control substituting phosphate-buffered saline for the primary antibody was used in each case. The immunocytochemical stains for Chlamydia-Cel-Pn, CMV, and HSV-1 were performed on adjacent sections to the hematoxylin and eosin, whereas the TT- 401 was performed at separate times on different microtome sections of the specimens.

Statistical Analysis
Comparison of proportions were made with ² analysis or Fisher's exact test. Confidence intervals (CI) at the 95% level were calculated for values.

	Results

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Seventy-six patients were enrolled in the study, and their demographic features are summarized in Table 1. All endarterectomy specimens demonstrated histopathological evidence of severe atherosclerosis with atheromatous lesions, with a core containing pools of extracellular lipid/necrotic and calcific debris and a fibrous cap. On the atheromatous plaques, there were 46 (60.5%) containing thrombus and 16 (21%) containing ulceration.

View this table: [in this window] [in a new window]   Table 1. Demographic Features of Patients With and Without C pneumoniae in Atherosclerotic Plaques

Immunocytochemical stains were positive for C pneumoniae in 30 samples (39.5%) with the Chlamydia-Cel-Pn antibody and 41 samples (54%) with TT-401 antibody; the combined results of the two antibodies detected C pneumoniae in 54 samples (71%) (CI, 59.5% to 80.9%). CMV was detected in 27 samples (35.5%) (CI, 24.8% to 47.3%), and HSV was detected in 8 samples (10.5%) (CI, 4.7% to 19.7%; see Fig 1). None of the controls (0 of 20) were positive for C pneumoniae, CMV, or HSV (P<.001 for CMV and C pneumoniae) (Table 2). At least one of the microorganisms was detected in 59 of the diseased arteries (77.6%) (CI, 66.6% to 86.4%), with a single microorganism in 35 of the samples (46%), two microorganisms in 12 (23.7%) (CI, 14.7% to 34.8%), and all three microorganisms in 6 (7.9%) (CI, 3.0% to 16.4%). Fig 2 illustrates the immunocytochemical stain positive for C pneumoniae in a carotid intimal plaque.

View larger version (34K): [in this window] [in a new window]   Figure 1. Rates of detection of C pnuemoniae, CMV, and HSV-1 singly and concurrently in 76 carotid atheromatous plaques.

View this table: [in this window] [in a new window]   Table 2. Presence of C pneumoniae, CMV, and HSV in Atherosclerotic vs Control (Normal) Arteries

View larger version (102K): [in this window] [in a new window]   Figure 2. Immunocytochemical staining (through ABC method) of carotid intimal atheromatous plaque with (arrowhead) strong positivity for Chlamydia-Cel-Pn in macrophage and endothelial cell. Small arrowheads indicate plaque ulceration with hemorrhage and thrombus formation. Hematoxylin counterstained. Original magnification, 250x.

The correlation of thrombosis and ulceration of the atherosclerotic plaques with the presence of the microorganisms is shown in Table 3. The presence of C pneumoniae or CMV independently was associated with a greater risk of thrombosis on the plaques but not plaque ulceration.

View this table: [in this window] [in a new window]   Table 3. Correlation of Thrombosis and Ulceration of Atherosclerotic Plaques With C pneumoniae and CMV

Serology
C pneumoniae IgG serum antibodies were present in 63 of 76 patients (82.9%); IgA antibodies were present in 26 (34.2%); and IgM antibodies were present in only 2 (2.6%). In patients with C pneumoniae in atherosclerotic plaques, 32 (59.3%) had high IgG titers (1:128), 12 (22.2%) had low titers (1:16 to 64), and 10 (18.5%) had no detectable antibodies (<1:16). Similarly, in patients with C pneumoniae absent in the blood vessels, 14 of 22 (63.6%) had high IgG titers (1:128), 5 (22.7%) had low titers (1:16 to 64), and 3 (13.6%) had no detectable antibodies. The distribution of the IgG quantitative antibodies was not significantly different between patients with or without C pneumoniae in the endarterectomy specimens. Only 9 of the patients (30%) with C pneumoniae in the atherosclerotic plaques had IgA antibodies versus 17 (37%) without C pneumoniae (not statistically significant).

CMV IgG antibody was present in 50 of the patients (65.8%). In the 27 patients with CMV antigen detected in the carotid artery, 8 (29.6%) had high titers (1:32), 7 (25.9%) had low titers (1:2 to 1:16), and 12 (44.4%) had no detectable antibodies. Similarly, in 49 patients without CMV in the carotid artery, 15 (30.6%) had titers of 1:32, 20 (40.8%) had low titers (1:4 to 1:16), and 14 (28.6%) had no detectable IgG antibodies. There was no correlation between presence of thrombosis on the atherosclerotic plaque and serum antibodies to C pneumoniae or CMV.

	Discussion

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Human atherogenesis appears to be of multifactorial etiology, and no single entity can fully explain the pathogenesis. There is little doubt that risk factors such as genetic predisposition, hypercholesterolemia, hypertension, smoking, and diabetes mellitus are major predisposing conditions for atherosclerosis. There is substantial evidence, albeit circumstantial, that infectious agents are associated with atherosclerosis, but their exact role in the pathogenesis of atherosclerosis is unknown. The most compelling evidence to date is the presence of infectious agents in the arterial wall, particularly in diseased vessels or within atherosclerotic plaques.

Our study is the first report to assess the presence of herpes viruses (CMV and HSV) and C pneumoniae together in the same tissue samples. C pneumoniae and CMV are commonly present in atherosclerotic carotid arteries (71% and 35.5%, respectively), but HSV is less frequently found (10%). Of interest is that 77.6% of the diseased arteries have at least one of the microorganisms detectable by immunocytochemistry and that 23.7% of the vessels have two microorganisms. Previous studies assessing human herpes viruses (particularly CMV) in latent infection of arterial walls have reported incidences ranging from 10% to 90%,^{7 8 9 10 11 12} depending on the technique used. Benditt et al,⁸ using in situ hybridization techniques, documented the presence of herpes virus nucleic acid in 10 of 60 tissue samples from patients who underwent coronary artery bypass graft surgery. Gyorkey et al⁷ reported the detection of herpes virus particles in electron microscopy sections from the aorta of 10 of 60 patients with atherosclerosis. Yamahiniroya et al,¹¹ using DNA hybridization and immunocytochemical techniques, identified HSV and CMV in 8 of 20 coronary artery specimens from young trauma victims. More recently, Hendricks et al¹² detected CMV nucleic acid through PCR in 90% of samples with severe atherosclerosis and in 50% of patients with minimal or no atherosclerosis. Similarly, Melnick et al³¹ detected CMV DNA through PCR by using E2 primers in 90% of 47 atherosclerotic tissues and 93% of 13 uninvolved aorta. Together, these studies indicate that CMV commonly causes latent infection of the arterial wall, but the presence of the virus is not specific for atherosclerotic vessels. In cardiac transplant recipients, however, graft atherosclerosis is more severe and more frequent in patients with CMV infection than in the uninfected patients.^{5 6} Our findings with immunocytochemical techniques are compatible with the results of other studies on CMV and HSV using similar techniques,^{7 11} but CMV was much less frequently detected in control normal arterial walls in our study than in other studies.^{12 31}

C pneumoniae has now been detected in atherosclerotic plaques in several different arterial sites (coronary arteries, aorta, and carotid arteries) and in early lesions (fatty streaks) and through the use of various independent techniques (immunocytochemical stains, PCR, and electron microscopy^{22 23 24 25 26 27} ), with immunocytochemistry being the most sensitive.^{24 25} However, C pneumoniae has not been detected in normal arterial walls.

Our findings with the TT-401 antibody are similar to those of the recent report of Grayston et al,²⁷ who detected C pneumoniae in 32 of 56 carotid endarterectomy tissues (57%), but the combined results with Chlamydia-Cel-Pn antibody are even higher (71%). The difference in the results between the two antibodies tested may be related to differences in cellular tissue avidity. However, the ability of the Chlamydia-Cel-Pn antibody to stain elementary bodies in some specimens in which the TT-401 stain was negative may be related to different sections (or cells) of the same tissues that are affected.

There are little data in the literature on the correlation between serum antibodies and the presence of either the herpes viruses or C pneumoniae in the arterial wall. It was surprising to note that 44% of our patients with CMV in the carotid arteries had no detectable IgG antibodies. This is in contrast to the findings of Melnick et al,³¹ who reported that serum antibody was detected in 86% of patients whose tissue demonstrated CMV DNA. Similarly, in our study, C pneumoniae antibodies were not detected in 20% of the patients in whom the bacterium was present in the artery, nor was there any correlation with the level of the antibody titer. These findings are compatible with a previous report on a smaller number of patients.²⁴ In that study, 6 of the 20 autopsy cases with C pneumoniae detectable in atherosclerotic plaques had no measurable antibodies by microimmunofluorescence in postmortem sera. The discrepancy between the immunocytochemical findings and serum antibodies may be in part explained by the presence of immune complexes and the binding of the antibodies, resulting in undetectable serum levels. Other possibilities might be a lack of sensitivity of the antibody assays or an obscure immune defect in some patients preventing the production of adequate specific antibodies, thus predisposing them to greater invasion of the arterial wall by these microorganisms.

In pathogenic terms, the presence of these microorganisms in diseased vessels or atherosclerotic plaques clearly does not necessarily imply a causal relationship. They might simply be present or carried there by mononuclear phagocytes without playing an active role. The presence of multiple infectious agents may also suggest nonspecific trapping of these microorganisms in areas of tissue damage, such as the atherosclerotic plaques. Nevertheless, their presence, although as yet unexplained, is intriguing, especially because C pneumoniae is not found in normal arterial wall. The recovery of viable C pneumoniae organisms through culture of coronary artery specimens of heart transplant recipients³² and carotid endarterectomy specimens³³ supports an active role of C pneumoniae. In addition, our ability to produce early changes in atherosclerosis (fatty streaks and complex spindle cell lesions) in a rabbit model infected with C pneumoniae and fed a noncholesterol standard rabbit diet³⁴ partly fulfills Koch's postulate and strongly suggests a causal role of C pneumoniae in the pathogenesis of atherosclerosis.

Furthermore, our data suggest that the presence of CMV or C pneumoniae may independently influence plaque morphology and predispose the patient to a greater risk of thrombosis on the plaques. However, further studies are needed to confirm this observation.

	Acknowledgments

 
We are grateful to Dr R. Bannatyne for his assistance in obtaining the CMV serology, Dr J. Romagnuolo for his assistance in data collection, D. Noda and M. Laurel for their technical assistance, Jianli Li for his assistance in the statistical analysis, and Dawn Bajhan for her assistance in preparation of the manuscript.

	Footnotes

 
Reprint requests to Dr I.W. Fong, Division of Infectious Diseases, St Michael's Hospital, 30 Bond St, Room 4137C, Toronto, Ontario, Canada M5B 1W8.

Received November 26, 1996; revision received April 7, 1997; accepted May 20, 1997.

	References

Top Abstract Introduction Methods Results Discussion References

 

1. Fabricant CG, Knook L, Gillespie JH. Virus-induced cholesterol crystals. Science. 1973;181:566-567.[Abstract/Free Full Text]
   
2. Fabricant CG, Fabricant J, Litrenta MM, Minick CR. Virus-induced atherosclerosis. J Exp Med. 1978;148:335-340.[Abstract/Free Full Text]
   
3. Hajjar DP. Viral pathogenesis of atherosclerosis. Am J Pathol. 1991;139:1195-1211.[Abstract]
   
4. Adam E, Melnick JL, Probesfield JL, Petrie BL, Burek J, Bailey KR, McCollum CH, DeBakey ME. High levels of cytomegalovirus antibody in patients requiring vascular surgery for atherosclerosis. Lancet. 1987;2:291-293.[Medline] [Order article via Infotrieve]
   
5. Gratton MT, Moreno-Cabral CE, Stames VA, Oyer PE, Stinson EB, Shumway NE. Cytomegalovirus infection is associated with cardiac allograft rejection and atherosclerosis. JAMA. 1989;261:3561-3566.[Abstract]
   
6. MacDonald K, Rector TS, Braunlan EA, Coubo SH, Olivori MT. Association of coronary artery disease in cardiac transplant recipients with cytomegalovirus infection. Am J Pathol. 1989;64:359-362.
   
7. Gyorkey F, Melnick JL, Guinn GA, Gyorkey P, DeBakey ME. Herpes viridae in the endothelial and smooth muscle cells of the proximal aorta of atherosclerotic patients. Exp Mol Pathol. 1984;40:328-339.[Medline] [Order article via Infotrieve]
   
8. Benditt EP, Barrett T, McDougall JK. Viruses in the etiology of atherosclerosis. Proc Natl Acad Sci USA. 1983;80:6386-6389.[Abstract/Free Full Text]
   
9. Melnick JL, Petrie BL, Dreesman GR, Burek J, McCollum CH, DeBakey ME. Cytomegalovirus antigen within human arterial smooth muscle cells. Lancet. 1983;2:644-647.[Medline] [Order article via Infotrieve]
   
10. Hajjar DP, Pomerantz KB, Falcone DJ, Weksler BB, Grant AJ. Herpes simplex virus infection in human arterial cells: implications in atherosclerosis. J Clin Invest. 1987;80:1317-1321.
    
11. Yamashiroya HM, Ghosh L, Yang R, Robertson AL. Herpes viridae in the coronary arteries and aorta of young trauma victims. Am J Pathol. 1988;130:71-79.[Abstract]
    
12. Hendricks MGR, Salimens MMM, Vanboven CPA, Bruggeman CA. High prevalence of latently present cytomegalovirus in arterial walls of patients suffering from grade III atherosclerosis. Am J Pathol. 1990;136:23-28.[Abstract]
    
13. Minick CR, Fabricant CG, Fabricant J, Litrenta MM. Atherosclerosis induced by infection with herpes virus. Am J pathol. 1979;96:673-706.[Abstract]
    
14. Saikku P, Mattila K, Nieminen MS, et al. Serological evidence of an association of a novel chlamydia, TWAR, with chronic coronary heart disease and acute myocardial infarction. Lancet. 1988;2:983-986.[Medline] [Order article via Infotrieve]
    
15. Thom DH, Grayston JT, Siscovitch DS, Wang SP, Weiss NS, Daling JR. Association of prior infection with Chlamydia pneumoniae and angiographically demonstrated coronary artery disease. JAMA. 1992;268:68-72.[Abstract]
    
16. Linnanmaki E, Leinonen M, Matilla K, Nieminen MS, Valtonen V, Saikku P. Chlamydia pneumoniae–specific circulating immune complexes in patients with chronic coronary heart disease. Circulation. 1993;87:1130-1134.[Abstract/Free Full Text]
    
17. Mendall MA, Carrington D, Strachan D, et al. Chlamydia pneumoniae: risk factor for seropositivity and association with coronary artery disease. J Infect. 1995;30:121-128.[Medline] [Order article via Infotrieve]
    
18. Patel P, Mendall MA, Carrington D, et al. Association of Helicobacter pylori and Chlamydia pneumoniae infection with coronary heart disease and cardiovascular risk factor. BMJ. 1995;311:711-714.[Abstract/Free Full Text]
    
19. Thom DH, Wang SP, Grayston JT, et al. Chlamydia pneumoniae strain TWAR antibody and angiographically demonstrated coronary artery disease. Arterioscler Thromb. 1991;11:547-551.[Abstract/Free Full Text]
    
20. Melnick S, Shakar E, Folsom A, Grayston JT, Sonlie P, Wang SP, Szklo M. Past infection by Chlamydia pneumoniae strain TWAR and asymptomatic carotid atherosclerosis. Am J Med. 1993;95:499-504.[Medline] [Order article via Infotrieve]
    
21. Saikku P, Leinonen M, Tenkanen L, Linnanmaki E, Ekman MR, Manninen V, Manttari M, Frick MH, Huttunen JK. Chronic Chlamydia pneumoniae infection as a risk factor for coronary heart disease in the Helsinki Heart Study. Ann Intern Med. 1992;116:273-278.
    
22. Shor A, Kuo CC, Patton DL. Detection of Chlamydia pneumoniae in coronary artery fatty streaks and atheromatous plaques. S Afr Med J. 1992;82:158-161.[Medline] [Order article via Infotrieve]
    
23. Kuo CC, Gown AM, Benditt EP, Grayston JT. Detection of Chlamydia pneumoniae in aortic lesions of atherosclerosis by immunocytochemical stain. Arterioscler Thromb. 1993;13:1500-1504.
    
24. Kuo CC, Shor A, Campbell LA, Fukushi H, Patton DL, Grayston JT. Demonstration of Chlamydia pneumoniae in atherosclerotic lesions of coronary arteries. J Infect Dis. 1993;167:841-849.[Medline] [Order article via Infotrieve]
    
25. Campbell LA, O'Brien ER, Cappuccio AL, Kuo CC, Wang SP, Stewart D, Patton DL, Cummings PK, Grayston JT. Detection of Chlamydia pneumoniae (TWAR) in human atherectomy tissues. J Infect Dis. 1995;172:585-588.[Medline] [Order article via Infotrieve]
    
26. Chiu B, Tucker W, Fong IW, Bilbao JM, Cheng Z. Chlamydia pneumoniae in carotid endarterectomies: morphologic and immunohistochemical studies. International Congress of the International Academy of Pathology, Hong Kong, October 9-14, 1994. Abstract.
    
27. Grayston JT, Kuo CC, Coulson AS, Campbell LA, Lawrence RD, Lee MJ, Strandness ED, Wang SP. Chlamydia pneumoniae (TWAR) in atherosclerosis of the carotid artery. Circulation. 1995;92:3397-3400.[Abstract/Free Full Text]
    
28. Blasi F, Denti F, Erba M, Cosentini R, Raccanelli R, Rinaldi A, kFagetti L, Esposito G, Ruberti U, Allegra L. Detection of C pneumoniae but not Helicobacter pylori in atherosclerotic plaques of aortic aneurysms. J Clin Microbiol. 1996;34:2766-2769.[Abstract]
    
29. Wang SP, Grayston JT. Micro-immunofluorescence serological studies with the TWAR organism. In: Oriel JD, Ridgway G, Schlachter J, Taylor-Robinson D, Ward M, eds. Chlamydial Infections. Cambridge, UK: Cambridge University Press; 1986:329-332.
    
30. VanDamme H, Vivario M, Boniver J, Limet R. Histologic characterization of carotid plaques. Cardiovasc Pathol. 1994;3:9-17.
    
31. Melnick JL, Hu C, Burek J, Adam E, DeBakey ME. Cytomegalovirus DNA in arterial walls of patients with atherosclerosis. J Med Virol. 1994;42:170-174.[Medline] [Order article via Infotrieve]
    
32. Ramirez J, and the Chlamydia pneumoniae/Atherosclerotic Study Group. Isolation of Chlmaydia pneumoniae from the coronary artery of a patient with coronary atherosclerosis. Ann Intern Med. 1996;125:979-982.[Abstract/Free Full Text]
    
33. Jackson LA, Rodriguez DI, Lee A, Kuo C-C, Campbell LA, Grayston JT. Isolation of Chlamydia pneumoniae (TWAR) from carotid atherosclerotic plaque specimen obtained by endarterectomy. 36th Interscience Conference of Antimicrobial Agents Chemotherapy, September 15-18, 1996, New Orleans, La. Abstract.
    
34. Fong IW, Chiu B, Viira E, Fong MW, Jang D, Mahony J. A rabbit model for Chlamydia pneumoniae infection. J Clin Microbiol. 1997;35:48-51.[Abstract]
    

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				A. Chait, C. Y. Han, J. F. Oram, and J. W. Heinecke Thematic review series: The Immune System and Atherogenesis. Lipoprotein-associated inflammatory proteins: markers or mediators of cardiovascular disease? J. Lipid Res., March 1, 2005; 46(3): 389 - 403. [Abstract] [Full Text] [PDF]

				H. Chi, E. Messas, R. A. Levine, D. T. Graves, and S. Amar Interleukin-1 Receptor Signaling Mediates Atherosclerosis Associated With Bacterial Exposure and/or a High-Fat Diet in a Murine Apolipoprotein E Heterozygote Model: Pharmacotherapeutic Implications Circulation, September 21, 2004; 110(12): 1678 - 1685. [Abstract] [Full Text] [PDF]

				B. Ludewig, P. Krebs, and E. Scandella Immunopathogenesis of atherosclerosis J. Leukoc. Biol., August 1, 2004; 76(2): 300 - 306. [Abstract] [Full Text] [PDF]

				M. K. Froberg CMV Escapes! Ann. Clin. Lab. Sci., April 1, 2004; 34(2): 123 - 130. [Abstract] [Full Text] [PDF]

				A. Jain, E. L. Batista Jr., C. Serhan, G. L. Stahl, and T. E. Van Dyke Role for Periodontitis in the Progression of Lipid Deposition in an Animal Model Infect. Immun., October 1, 2003; 71(10): 6012 - 6018. [Abstract] [Full Text] [PDF]

				P. J. Lindsberg and A. J. Grau Inflammation and Infections as Risk Factors for Ischemic Stroke Stroke, October 1, 2003; 34(10): 2518 - 2532. [Abstract] [Full Text] [PDF]

				A. J. Karter, D. H. Thom, J. Liu, H. H. Moffet, A. Ferrara, and J. V. Selby Use of Antibiotics Is Not Associated With Decreased Risk of Myocardial Infarction Among Patients With Diabetes Diabetes Care, July 1, 2003; 26(7): 2100 - 2106. [Abstract] [Full Text] [PDF]

				M. Benagiano, A. Azzurri, A. Ciervo, A. Amedei, C. Tamburini, M. Ferrari, J. L. Telford, C. T. Baldari, S. Romagnani, A. Cassone, et al. T helper type 1 lymphocytes drive inflammation in human atherosclerotic lesions PNAS, May 27, 2003; 100(11): 6658 - 6663. [Abstract] [Full Text] [PDF]

				Y. Agmon, B. K. Khandheria, I. Meissner, T. M. Petterson, W. M. O'Fallon, T. J. H. Christianson, D. O. Wiebers, T. F. Smith, J. M. Steckelberg, and A. J. Tajik Lack of association between Chlamydia pneumoniae seropositivity and aortic atherosclerotic plaques: A Population-Based transesophageal echocardiographic study J. Am. Coll. Cardiol., May 7, 2003; 41(9): 1482 - 1487. [Abstract] [Full Text] [PDF]

				P. Khairy, S. Rinfret, J.-C. Tardif, R. Marchand, S. Shapiro, J. Brophy, and J. Dupuis Absence of Association Between Infectious Agents and Endothelial Function in Healthy Young Men Circulation, April 22, 2003; 107(15): 1966 - 1971. [Abstract] [Full Text] [PDF]

				M. Fukui, Y. Kitagawa, N. Nakamura, and T. Yoshikawa Hepatitis C Virus and Atherosclerosis in Patients With Type 2 Diabetes JAMA, March 12, 2003; 289(10): 1245 - 1246. [Full Text] [PDF]

				P. Liuba, J. Persson, J. Luoma, S. Yla-Herttuala, and E. Pesonen Acute infections in children are accompanied by oxidative modification of LDL and decrease of HDL cholesterol, and are followed by thickening of carotid intima-media Eur. Heart J., March 2, 2003; 24(6): 515 - 521. [Abstract] [Full Text] [PDF]

				J. D. Spence and J. Norris Infection, Inflammation, and Atherosclerosis Stroke, February 1, 2003; 34(2): 333 - 334. [Full Text] [PDF]

				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]

				C. Espinola-Klein, H.-J. Rupprecht, S. Blankenberg, C. Bickel, H. Kopp, A. Victor, G. Hafner, W. Prellwitz, W. Schlumberger, and J. Meyer Impact of Infectious Burden on Progression of Carotid Atherosclerosis Stroke, November 1, 2002; 33(11): 2581 - 2586. [Abstract] [Full Text] [PDF]

				A. W. Haider, P. W. F. Wilson, M. G. Larson, J. C. Evans, E. L. Michelson, P. A. Wolf, C. J. O'Donnell, and D. Levy The association of seropositivity to Helicobacter pylori, Chlamydia pneumoniae, and cytomegalovirus with risk of cardiovascular disease: A prospective study J. Am. Coll. Cardiol., October 16, 2002; 40(8): 1408 - 1413. [Abstract] [Full Text] [PDF]

				I. W. Fong, B. Chiu, E. Viira, D. Jang, and J. B. Mahony Influence of Clarithromycin on Early Atherosclerotic Lesions after Chlamydia pneumoniae Infection in a Rabbit Model Antimicrob. Agents Chemother., August 1, 2002; 46(8): 2321 - 2326. [Abstract] [Full Text] [PDF]

				I. Baranova, T. Vishnyakova, A. Bocharov, Z. Chen, A. T. Remaley, J. Stonik, T. L. Eggerman, and A. P. Patterson Lipopolysaccharide Down Regulates Both Scavenger Receptor B1 and ATP Binding Cassette Transporter A1 in RAW Cells Infect. Immun., June 1, 2002; 70(6): 2995 - 3003. [Abstract] [Full Text] [PDF]