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C-reactive protein (CRP) is the classic human acute-phase protein. Its concentration can cover a 10,000-fold range, from less than 50 µg/l to greater than 500 mg/l, during the nonspecific response to most forms of tissue injury, infec-tion and inflammation.1 The first proposal of an association between CRP and atherosclerosis was made by my group in 1982, when we discovered specific binding of LDL and VLDL by aggregated CRP.2 The avalanche of current interest in this field was, however, triggered by our finding that even modest increases in baseline CRP values are associated with an increased risk of future atherothrombotic events.3 In the same report, we also showed a similar predictive association between serum amyloid A, another extremely sensitive acute-phase protein, and future athero-thrombotic events. We explicitly discussed the nonspecific nature of the CRP response and the different possible reasons for the associa-tion.3 These considerations have largely been ignored; attention has inappropriately focused almost exclusively on CRP and it has been widely assumed—without any supporting evidence—that baseline CRP values specifically reflect atheroma-related inflammation in the arterial wall.

CRP is a stable analyte for which commer-cial clinical chemistry assays are routinely available. Such assays are standardized to the WHO’s International Reference Standard for CRP Immunoassay, which I produced in 1984. Measurement of CRP within the normal range of up to 3 mg/l requires more sensitive methods than those used for monitoring the acute-phase response.3 Badging of the CRP assays used in cardiovascular disease studies as high- sensitivity CRP (hs-CRP) tests has created a false but, unfortunately, enduring impression that this measurement is somehow new and distinct from previous CRP assays, and that it has specific relevance to cardiovascular disease. In fact, sensitive assays for CRP have been avail-able for over 30 years and there is nothing new or different about hs-CRP tests. It is essential to

C-reactive protein is neither a marker nor a mediator of atherosclerosisMark B Pepys

MB Pepys is Professor of Medicine and Head of the Department of Medicine at the Royal Free Campus of University College London and Honorary Consultant Physician at the Royal Free Hospital, London, UK.

CorrespondenceCentre for Amyloidosis and Acute Phase ProteinsDivision of MedicineRoyal Free CampusUniversity College LondonRowland Hill StreetLondon NW3 2PFUKm.pepys@medsch.ucl.ac.uk

Received 26 November 2007Accepted 11 January 2008Published online 4 March 2008

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understand that the CRP response is not specific, that CRP values alone can never be diagnostic, and that these values can be usefully interpreted only with comprehensive information about the individual patient at the time of sampling.1

The strength of the relationship between baseline CRP values and future risk of athero-thrombotic events in studies of the general population was initially exaggerated, leading to the widely repeated mantra that CRP is a “strong, independent marker of cardiovascular disease risk”. Now that the total events in these studies are approaching epidemiologically robust numbers, the strength of this association is much attenuated,4 and it is clear that at least 70% of the variance in baseline CRP is attrib-utable to the established causative risk factors for cardiovascular disease.5 Exaggeration of the epidemiological association, coupled with blindness to the nonspecific nature of the CRP response, has led to a misplaced focus on CRP. Indeed, other non specific systemic markers of low-grade inflammation (such as serum albumin concentration and total leukocyte count) show similar associations with cardio-vascular disease risk4,5 and are cheaper to measure. There is no compelling evidence that measurement of CRP, or of other non specific markers, adds useful information to a cardio-vascular risk assessment that is based on the classical causative risk factors.5 Studies of atherosclerotic burden, rather than of actual atherothrombotic events, have had varying results but overall provide no compelling evidence for an association with CRP values.5

Nevertheless, there has been an inappropriate and misleading conflation of association with causality. Enthusiasm for the potential patho-genicity of CRP has been fueled by multiple in vitro cell culture studies that used commercial recombinant human CRP produced in bacteria. This reagent is heavily contaminated with endotoxin, and possibly other bacterial prod-ucts, and is inappropriate and unsuitable for functional work with cells.6 Despite claims to the

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Competing interestsThe author has declared an association with the following company: Pentraxin Therapeutics. See the article online for full details of the relationship.

contrary, it is not possible to completely remove these potent bacterial molecules from solutions of proteins or to block all of their biological activity. None of the studies that used recom-bi nant CRP produced in bacteria has included adequate controls, and none of the observed direct pro inflammatory and proatherogenic effects has been reproducible with authentic pure human CRP or with recombinant CRP produced in mammalian cells.7 In the early work in this unfortunate saga, authors did not even bother to dialyze out the toxic bacteriostatic preservative sodium azide from the commercial CRP, so that there is now a controversial body of literature reporting on which of the ‘CRP effects’ were or were not due to azide!

In addition to contamination artefacts, there are also critical technical and scientific considera-tions regarding the credibility of the wide-ranging claims of the biological effects of CRP. These considerations include the use of whole IgG antibodies to detect binding of CRP by Fc recep-tors, the fact that CRP itself is a ligand-binding protein that recognizes cell membrane struc-tures, the effect of covalent modification (such as coupling to biotin or fluorochromes) on the binding characteristics of CRP, the absence of essential controls such as blockade of CRP effects by anti-CRP antibodies and by inhibition of CRP ligand binding, and the lack of repro-ducibility of the results obtained with bacterial CRP when authentic pure human CRP is used.Furthermore, if the effects claimed for CRP on the basis of in vitro studies are to be credible, they must be consistent with the known behavior and actions of CRP in vivo. Infusions of huge doses of pure human CRP have no vascular or pro inflammatory effects in vivo in normal mice and rats. Patients whose endogenous CRP values vary by thousands of fold over a couple of days during the acute-phase response, or remain persistently elevated at hundreds of fold above normal for months or years, do not experience vascular or other effects compatible with CRP being a multifunctional, pro inflammatory, prothrombotic and proatherothrombotic cytokine that triggers its own production. In rigorous studies of mouse atherosclerosis models, transgenic human CRP either has no effect on atherogenesis8,9 or is atheroprotective.10 Preliminary genetic epidemio logical studies

based on the principle of Mendelian randomiza-tion, which should reduce confounding and avoid reverse causality, so far provide no support for a pathogenic role of CRP in hypertension, atherosclerosis or athero thrombosis.5

In contrast to the absence of rigorous evidence for a pathogenic role of human CRP in atherosclerosis, there is robust evidence that high concentrations of human CRP can exacer bate ischemic infarction via a complement-dependent mechanism.11 This process is completely different from the direct cellular signaling postulated by the proponents of CRP per se as a pathogenic mediator, and it could well operate in many different clinical situations in which high CRP concentrations are associated with pre-existing tissue damage. Abundant CRP is required, rather than just trivially increased baseline values, and the enhanced tissue damage is prevented by a specific drug that inhibits CRP ligand binding and thus inhibits complement activation by CRP.11

References1 Pepys MB and Hirschfield GM (2003) C-reactive

protein: a critical update. J Clin Invest 111: 1805–18122 de Beer FC et al. (1982) Low density lipoprotein and

very low density lipoprotein are selectively bound by aggregated C-reactive protein. J Exp Med 156: 230–242

3 Liuzzo G et al. (1994) The prognostic value of C-reactive protein and serum amyloid A protein in severe unstable angina. N Engl J Med 331: 417–424

4 Danesh J et al. (2004) C-reactive protein and other circulating markers of inflammation in the prediction of coronary heart disease. N Engl J Med 350: 1387–1397

5 Lowe GD and Pepys MB (2006) C-reactive protein and cardiovascular disease: weighing the evidence. Curr Atheroscler Rep 8: 421–428

6 Pepys MB et al. (2005) Proinflammatory effects of bacterial recombinant human C-reactive protein are caused by contamination with bacterial products, not by C-reactive protein itself. Circ Res 97: e97–e103

7 Taylor KE et al. (2005) C-reactive protein-induced in vitro endothelial cell activation is an artefact caused by azide and lipopolysaccharide. Arterioscler Thromb Vasc Biol 25: 1225–1230

8 Hirschfield GM et al. (2005) Transgenic human C-reactive protein is not proatherogenic in apolipoprotein E-deficient mice. Proc Natl Acad Sci USA 102: 8309–8314

9 Tennent GA et al. (2007) Transgenic human CRP is not pro-atherogenic, pro-atherothrombotic or pro-inflammatory in apoE-/- mice. Atherosclerosis 196: 248–255

10 Kovacs A et al. (2007) Human C-reactive protein slows atherosclerosis development in a mouse model with human-like hypercholesterolemia. Proc Natl Acad Sci USA 104: 13768–13773

11 Pepys MB et al. (2006) Targeting C-reactive protein for the treatment of cardiovascular disease. Nature 440: 1217–1221

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