© 2000 American Heart Association, Inc.
Letters to the Editor |
Deposition of Modified or Native C-Reactive Protein in Atherosclerotic Arteries?
Abteilung Rheumatologie und Klinische Immunologie, Medizinische Unversitätsklinik, Universitäts Freiburg Klinikum, Freiburg, Germany
Next Era Therapeutics, Inc, Vernon Hills, Illinois
To the Editor:
Accumulating evidence suggests a predictive role of elevated serum concentrations of C-reactive protein (CRP) for atherosclerosis and its thrombotic complications.1 These findings apparently reflect an inflammatory component of the multifactorial atherosclerotic process. Furthermore, it is increasingly recognized that CRP may not merely represent an indicator of inflammation but may also, because of its known functional properties, be actively involved in the initiation or perpetuation of local inflammatory reactions.2 3 A direct approach in the study of the latter hypothesis is the search for CRP in atherosclerotic lesions. [banner]
In this context, the recent publication of Torzewski et al4 in Arteriosclerosis, Thrombosis, and Vascular Biology is of actual relevance. By means of immunohistochemistry, the authors clearly demonstrate deposits of CRP beside terminal complement proteins in the arterial wall of patients with early atherosclerotic lesions. Because ligand-bound CRP activates the classical pathway of complement, the authors suggest that the colocalization of terminal complement proteins with CRP might reflect complement activation by CRP in situ. Even if we share the hypothesis that CRP may contribute to local complement activation, this conclusion is not unequivocally supported by the presented findings. The monoclonal antibody against CRP used in that study (clone CRP-8 from Sigma) exclusively recognizes a modified form of CRP (mCRP) and not the native CRP molecule. This restricted specificity of clone CRP-8 against mCRP described by the manufacturer (Sigma) was recently confirmed experimentally by us.5 Differentiating between both CRP variants may be functionally relevant, because mCRP, contrary to its native counterpart, is not able to activate complement.5 6 Additionally, the denatured reduced form of CRP (mCRP) differs from native CRP in many biological and physicochemical properties,7 which may be important for the local prevalence and function of the CRP molecule. One might reconcile the findings and hypotheses of that study4 by the assumption that native CRP may reach an inflammatory focus, may activate complement after binding to appropriate ligands (phospholipids, lipoproteins, or nuclear debris), and may finally be transformed into mCRP under acidic conditions of an inflammatory microenvironment. The significance of mCRP is under investigation. Our data have identified mCRP as a natural component of normal blood vessel intima.8 Its presence and in vitro activities suggest that it may be a part of the physical signaling pathway of the extracellular matrix.9
References
1.
Koenig W, Sund M, Fröhlich M, Fischer
HG, Löwel H, Döring A, Hutchinson WL, Pepys MB. C-reactive
protein, a sensitive marker of inflammation, predicts future risk of
coronary heart disease in initially healthy middle-aged men.
Circulation. 1999;99:237–242.
2. Thompson D, Pepys MB, Wood SP. The physiological structure of human C-reactive protein and its complex with phosphocholine. Structure. 1999;7:169–177.[Medline] [Order article via Infotrieve]
3. Hack CE, Wolbink GJ, Schalkwijk C, Speijer H, Hermens WT, van den Bosch H. A role for secretory phospholipase A2 and C-reactive protein in the removal of injured cells. Immunol Today. 1997;18:111–115.[Medline] [Order article via Infotrieve]
4.
Torzewski J, Torzewski M, Bowyer DE, Frohlich M,
Koenig W, Waltenberger J, Fitzsimmons C, Hombach V. C-reactive protein
frequently colocalizes with the terminal complement complex in the
intima of early atherosclerotic lesions of human coronary
arteries. Arterioscler Thromb Vasc Biol. 1998;18:1386–1392.
5. Vaith P, Prasauskas V, Potempa L, Peter HH. Complement activation by C-reactive protein on the HEp-2 cell substrate. Int Arch Allergy Immunol. 1996;111:107–117.[Medline] [Order article via Infotrieve]
6. Potempa L A, Zeller J M, Fiedel B A, Kinoshita C M, Gewurz H. Stimulation of human neutrophils, monocytes, and platelets by modified C-reactive protein (CRP) expressing a neoantigenic specificity. Inflammation. 1988;12:391–405.[Medline] [Order article via Infotrieve]
7. Motie M, Brockmeier S, Potempa LA. Binding of model soluble immune complexes to modified C-reactive protein. J Immunol. 1996;156:4435–4441.[Abstract]
8. Diehl EE, Haines III GK, Radosevich JA, Potempa LA. Immunohistochemical localization of modified C-reactive protein antigen in normal vascular tissue. Am J Med Sci. In press.
9. Lelievre S, Weaver VM, Bissell MJ. Extracellular matrix signaling from the cell membrane skeleton to the nuclear skeleton: a model of gene regulation. Recent Prog Horm Res. 1996;51:417–432.
Comment on ‘Deposition of Modified or Native C-Reactive Protein in Atherosclerotic Arteries?’
Department of Cardiology, Internal Medicine II, University of Ulm, Ulm, Germany
Department of Clinical Chemistry, University of Regensburg, Regensburg, Germany
In their letter "Deposition of Modified or Native C-Reactive Protein in Atherosclerotic Arteries?" Peter Vaith and Lawrence Potemka comment on our recent publication "C-Reactive Protein Frequently Colocalizes With the Terminal Complement Complex in the Intima of Early Atherosclerotic Lesions of Human Coronary Arteries."R1 In that article, we demonstrated deposition of C-reactive protein (CRP) in early human atherosclerotic lesions and its colocalization with the terminal complement complex C5b-9. On the basis of that finding, we suggested that CRP in early atherosclerotic lesions may promote inflammation in the arterial wall by activating the complement system.
The authors of this valuable letter pointed to the fact that CRP
in the arterial wall, owing to acidic conditions in
inflamed tissue, may be modified into a denatured and reduced form
(mCRP) displaying other biological and physiochemical properties than
the native form. In the light of increasing evidence for a pivotal role
of CRP in atherosclerosis, we agree with the authors’
notion that mCRP and its functional properties are most important to
study. However, we do not agree with their statement that the antibody
that we used for CRP staining (clone CRP-8 from Sigma Chemical Co)
"... exclusively recognizes a modified form of CRP (mCRP) and not
the native CRP molecule." At this point, we would like to cite the
specification sheet of the antibody provided by Sigma, which states
"monoclonal anti-human C-reactive protein (CRP) recognizes an epitope
located on the 24-kDa subunit of denatured reduced CRP applying the
immunoblotting technique. It does not cross-react with
human serum amyloid P component (SAP), human haptoglobin, human 
-1
acid glycoprotein, and human IgG, nor with CRP from
Limulus. The product displays its reactivity
against CRP (native and denatured-reduced) applying ELISA, dot-blot,
and immunoblotting technique)" (italics added by
us for emphasis). In addition, we have addressed the point
raised by Vaith and Potemka in a conversation with the manufacturer,
who confirmed the information given in the specification
sheet.
On the basis of this information, we suggest that the antibody used in our study is able to detect both the native and the modified form of CRP. In this context, we would like to emphasize that we were not able to demonstrate CRP staining in coronary arteries without any signs of atherosclerotic lesion development. The question remains which form of CRP is the dominating one in early atherosclerotic lesions. Although we did not address this question in our study, extensive costaining with C5b-9 suggests the presence of considerable amounts of native CRP. Furthermore, very recently we demonstrated that native CRP binds to degraded LDL, enhancing complement activation.R2 Immunohistochemical studies complemented these findings in showing that CRP colocalizes with enzymatically modified–LDL in early human atherosclerotic lesions.R2 However, it is most likely that CRP becomes modified in the lesion, at least in intermediate and advanced lesions with increasing acidic conditions, and the significance of this modified CRP needs to be investigated further.
References
1. Torzewski J, Torzewski M, Bowyer DE, Frohlich M, Koenig W, Waltenberger J, Fitzsimmons C, Hombach V. C-reactive protein frequently colocalizes with the terminal complement complex in the intima of early atherosclerotic lesions of human coronary arteries. Arterioscler Thromb Vasc Biol. 1998;18:1386–1392.
2.
Bhakdi S, Torzewski M, Klouche M, Hemmes M.
Complement and atherogenesis: binding of CRP to degraded, nonoxidized
LDL enhances complement activation. Arterioscler Thromb Vasc
Biol. 1999;19:2348–2354.
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