We found no impairment of the endothelial function, measured as ED-FMD, among patients with newly diagnosed RA compared with controls. There have been, to the best of our knowledge, only two previous studies on ED-FMD in newly diagnosed RA patients [19
]. In those studies, patients with RA had a blunted endothelium-dependent vasodilatation. However, our study comprises four to eight times more patients with RA than those studies, which might in part explain the different results.
There was no significant difference in IMT in patients with early RA compared with controls. There are few studies evaluating atherosclerosis in newly diagnosed RA patients. In two recent studies, RA patients had an increased IMT compared with healthy controls [36
]. Those studies comprised older patients with higher inflammatory activity than those in the present study. Moreover, there were several other methodological differences compared with the present study that might explain the different results. Also, in another recent study, evaluating coronary calcification in early RA, there were no difference in atherosclerosis compared with controls [38
]. However, in a tentative follow-up after 18 months we found a significant increase in the IMT among patients with RA while there was no equivalent significant change in the control group. This increase was significant although all patients with RA were treated with DMARDs and had a low inflammatory activity. In longstanding RA we have previously demonstrated an increased IMT [7
], a finding later confirmed by other investigators [6
]. Also in patients with longstanding RA an increase in IMT has been shown even though efficient treatment with anti-TNF therapy [39
]. One recent epidemiological study showed an increased incidence of CVD, as measured by myocardial infarction, in patients with RA during the two years prior to diagnosis [40
]. The authors concluded that their results could imply the presence of inflammatory-triggered atherosclerosis before overt symptoms and diagnosis of RA.
We also wanted to identify endothelial biomarkers reflecting the early atherosclerotic process in RA. Thus, we evaluated the involvement of adhesion molecules, MCP-1 and endothelially produced haemostatic factors in endothelial dysfunction and very early atherosclerosis, as measured by ED-FMD and IMT, respectively. Simple regression analyses revealed that decreased sL-selectin was strongly related to a lower ED-FMD in RA and associated with an increased IMT in both RA and controls. A lower level of sL-selectin has been associated with CVD in the general population [24
]. L-selectin is expressed on leucocytes and is also known to be involved in the initial leukocyte rolling on activated endothelium [24
]. L-selectin has been shown to be downregulated during chronic inflammation. We found a strong inverse relation between sL-selectin and DAS28, which reflects the current inflammatory status in RA. In multiple linear regression models, evaluating the impact of endothelial biomarkers on endothelial function and adjusting for inflammation, sL-selectin was still strongly related to ED-FMD and also close to significantly related to IMT. This implies a durable relation between sL-selectin and endothelial function in early RA, a relation that does not seem to be mediated simply via inflammatory disease activity. This is a new finding that warrants further investigation of pathophysiological mechanisms beyond the inflammatory pathway.
Regarding the other adhesion molecules, sVCAM-1 and sICAM-1 showed higher levels in RA than in controls; however, we did not find any independent relations with any of the ultrasound measurements among our relatively young patients with early RA. Although, in agreement with the findings of others, some of the adhesion molecules showed relations to measures of inflammation [27
]. In previous studies on older patients with long-standing RA, several of these markers correlated with IMT [26
]. The inflammatory status of the present patient cohort was, however, low and of short duration, leading to less stress on the endothelium.
MCP-1 was found to be a strong and independent predictor of an increased IMT. This chemoattractant for monocytes is secreted by activated endothelial cells and has been suggested to be a marker of endothelial inflammation, the first step in atherosclerosis [23
]. Moreover, MCP-1 has been shown to promote plaque growth, to be increased in acute CVD in the general population [23
] and to be correlated with IMT in uraemic patients [22
]. In the present study MCP-1 was related to tPA-mass, which in turn was associated with markers of inflammation both here and in other studies [41
]. However, when adjusting for inflammation and other endothelial biomarkers, the impact of MCP-1 on IMT was even more significant. We have previously found increased MCP-1 to precede the diagnosis of RA in anti-CCP or RF-positive individuals [42
]. Previously, a more atherogenic lipid profile, suggested to modify the inflammatory reaction, has been shown to antedate the diagnosis of RA by at least 10 years [43
]. Taken together, these findings are suggestive of a very early endothelial activation in RA, possibly already present before diagnosis.
Regarding the haemostatic factors, increased tPA-mass was related to a higher IMT. There was also a tendency towards a significant relation between increased PAI-1-mass and impaired ED-FMD. Increased values for PAI-1 and tPA mass have been shown to be associated with an increased risk of CVD in the general population [21
] as well as in patients with RA [26
]. tPA is produced endothelially and an increase in this biomarker might be another reflection of an endothelial activation [21
]. In this study, there was a relation between tPA-mass and DAS28 in patients with RA. Increasing PAI-1-mass was also related to markers of inflammation. PAI-1 is an acute-phase reactant and this finding is consistent with our previous study in which PAI-1 was increased in RA patients compared with controls [7
]. Both tPA and PAI-1 have also been shown to be associated with inflammation in patients with diabetes mellitus [41
]. An increase in these biomarkers leads to a prothrombotic state [21
]. Furthermore, our finding that the level of VWF, another endothelially produced haemostatic factor, was independently related to the IMT, is in accordance with other reports [8
]. This relation also remained when adjusted for ESR. Taken together, our findings implicate a procoagulant state among our RA patients with newly diagnosed RA.
It appears, as a result of this study, that sL-selectin, being involved in the initial leukocyte rolling process, is a strong predictor of the early endothelial activation in RA, as measured by ED-FMD, whereas MCP-1 and VWF are indicators of increasing IMT, a later phase of the early atherosclerotic process. Corresponding results were not found in the control group.
In the RA group, the IMT was higher among men than females, and IMT also increased with age, higher systolic blood pressure, higher cholesterol and a longer period of smoking or using oral snuff. Also, there was a decrease in ED-FMD in men compared with women, ED-FMD decreased with increasing age and increasing systolic blood pressure. All of these findings are in line with studies on the general population [11
] or patients with established RA [4
Our ongoing study is one of the first to investigate the ED-FMD and IMT prospectively among newly diagnosed patients with RA. A strength of the study is that in northern Sweden nearly all patients with newly diagnosed RA are included in a structured follow up. All patients aged 60 years or younger were asked to participate in this study within 12 months of their diagnosis. As our main interest is to evaluate the impact of inflammation and endothelial activation on the early atherosclerotic process prospectively over a long time, a young cohort comprising patients with very early RA will be less affected by obscuring generalised atherosclerosis [42
] and by other comorbidities and mortality in the coming decades. Moreover, as inflammation seems to be relatively more important for the atherosclerotic progression in younger patients [4
] this cohort should, however, be ideal for unmasking any difference prospectively. Although it may have rendered it more difficult to determine baseline differences in RA vs. controls, the design allows repeated analyses in a comparably large cohort. The first follow-up only 18 months after the initial ultrasound measurement could be accomplished in only a subgroup. Nevertheless, this provided a unique opportunity to investigate rapid changes in the measurements of atherosclerosis among patients with RA. A complete follow-up in all RA patients and controls five years after the first ultrasound investigation is in progress. Another strength of this study was that the same person (EL) undertook all of the ultrasound measurements, thereby eliminating interpersonal variation. A limitation is that there was only one control per two RA patients, which could decrease the statistical power in the control group. Both the control group and, in particular, the RA-group are, however, larger than in most previous studies on RA using this technique. Furthermore, our main focus regarding associations between physiological measures and biomarkers was in the larger RA-group. Another limitation could be that all individuals were already on and continued their medications, including DMARDs, corticosteroids, non-steroidal anti-inflammatory drugs and coxibs throughout the ultrasound measurement, due to the fact that this work is part of a prospective observational study. Medications at the first ultrasound investigation, or up until the first follow-up, had however no statistically significant influence on the results.
Taken together, our results point towards an ongoing endothelial activation among patients with early RA. This endothelial activation was not translated into endothelial dysfunction, as measured by ED-FMD. Changes of ED-FMD can be rapid and could have reflected a high inflammatory activity if present. The variability of IMT is complex and therefore more inert to temporary changes. A more sensitive physiological method would possibly have been able to uncover a relation. However, there were differences in the levels of biomarkers, indicating endothelial activation in early RA to be related to the disease process, yet independent of acute-phase reaction. Also, after 18 months this had been translated into a significant increase of atherosclerosis as measured by IMT, not evident among the controls. Furthermore, more biomarkers were related to the physiological measurements within the RA patient cohort than among the controls, while the controls exhibited more relations between the physiological measurements and traditional CVD risk factors. This supports the hypothesis that the disease process in RA is involved in early endothelial activation and atherosclerosis, in a way distinguished from the general population. This possibly indicates that the endothelial activating process also has a different course compared with that leading to joint destruction in RA. In previous studies of patients with long-standing RA, in whom endothelial activation had continued for several years, the IMT was consistently shown to be increased compared with controls [6