Transgenic human CRP is not pro-atherogenic, pro-atherothrombotic or pro-inflammatory in apoE^−/− mice

Abstract

The pathogenic significance, if any, of the epidemiological association between baseline C-reactive protein (CRP) values and future atherothrombotic events is not known. We therefore investigated spontaneous atherosclerosis and atherothrombosis, and systemic markers of inflammation (acute phase proteins), in aged, normal diet-fed, male apolipoprotein E deficient (apoE^−/−) mice with and without transgenic expression of human CRP. At 18 months of age, aortic atherosclerosis was extensive but with no significant difference in plaque size between C57BL/6apoE^−/− mice with (apoE^−/−-hCRP⁺) and without transgenic human CRP (apoE^−/−). Atherosclerotic lesions in brachiocephalic arteries were typically complex and layered, with extensive fibrotic-cholesterol deposits, calcification and occasional recent intraplaque haemorrhage and thrombus, but with no significant overall differences between apoE^−/− and apoE^−/−-hCRP⁺ animals. Concentrations of mouse serum amyloid P component (SAP) were essentially normal throughout and did not differ between apoE^−/− and apoE^−/−-hCRP⁺ mice, or between wild-type (apoE^+/+) and apoE^−/− mice, regardless of human CRP expression. Mouse serum amyloid A protein (SAA), and human CRP concentrations were modestly but significantly higher in apoE^−/−-hCRP⁺ than in apoE^+/+-hCRP⁺ animals, but mouse SAA values were unaffected by transgenic expression of human CRP in either background. Thus, there was no evidence in this 18 month study of apoE^−/−, and control apoE^+/+ mice, that transgenic human CRP was pro-atherogenic, pro-inflammatory or pro-atherothrombotic.

Introduction

There is currently intense interest in the relationship between the classical human acute phase protein, C-reactive protein (CRP), and cardiovascular disease. The epidemiological and experimental observations and their implications for pathogenesis, risk assessment and clinical management are controversial [1]. It is therefore critical to distinguish clearly the very different pathological processes involved in cardiovascular disease, which include: (1) the slow lifelong development of atherosclerosis; (2) the catastrophic acute atherothrombotic event that causes arterial occlusion; and (3) the consequences of tissue ischaemia after arterial occlusion. Equally important is recognition that human CRP is produced non-specifically in response to almost all forms of tissue damage and inflammation [2], [3], and that its swift and enormous dynamic range encompasses up to 10,000-fold difference in circulating concentration between baseline and the acute phase response, with correspondingly very different potential functional effects [2].

There is a significant epidemiological association between baseline CRP values and future coronary atherothrombotic events in general population cohorts [4], but recent large scale studies show no relationship between CRP and the necessarily indirect measures of atherosclerosis burden [5], [6]. Furthermore, neither epidemiological studies nor experimental work in vitro and in vivo support the idea that CRP plays a causal role in atherosclerosis [1]. We previously reported that transgenic expression of human CRP did not affect development of spontaneous aortic atherosclerosis of standard diet-fed apolipoprotein E deficient (apoE^−/−) mice up to the age of 12 months and was not associated with increased production of mouse serum amyloid P component (SAP), an exquisitely sensitive acute phase reactant [7]. In the present study, in order to evaluate the more critical question of any association with atherothrombosis rather than just atheroma, we followed a cohort of apoE^−/− mice up to 18 months of age when they develop advanced, complex, thrombosis-prone atherosclerotic lesions in the brachiocephalic artery [8], [9]. We also monitored SAP and serum amyloid A protein (SAA), the most sensitive murine acute phase proteins, to determine whether transgenic expression of human CRP has any pro-inflammatory effects in mice in vivo, and whether development of extensive severe atherosclerosis in this model is associated with increased concentrations of systemic markers of inflammation.