Methotrexate and anti-tumor necrosis factor treatment improves endothelial function in patients with inflammatory arthritis

Inflammatory arthritis (IA), including rheumatoid arthritis (RA), ankylosing spondylitis (AS), and psoriatic arthritis (PsA), has increased cardiovascular (CV) morbidity and mortality, probably due to cardiovascular disease (CVD) caused by atherosclerosis [1‐5]. The first step in the development of atherosclerosis is endothelial dysfunction (ED) which is initially a reversible process [6]. Thus, improving endothelial function (EF) might be of great importance in preventing atherosclerosis. The endothelium has several vital homeostatic functions, including regulation of vascular tone and growth, thrombogenesis and thrombolysis, and interactions between platelets and leukocytes and the vessel wall. The endothelium secretes vasorelaxing (e.g., nitric oxide) and vasoconstricting (e.g., endothelin-1) substances in response to mechanical stress [6, 7]. ED is characterized by impaired ability of the artery to dilate in response to physical and chemical stimuli [8, 9]. Assessment of EF has been used to estimate the CV risk in IA patients [10, 11].

Clinical studies indicate that antirheumatic treatment, including methotrexate (MTX) and antitumor necrosis factor (anti-TNF) treatment, not only ameliorates disease activity but also reduces CV morbidity and mortality in RA patients [12, 13]. There is also evidence that anti-TNF treatment improves EF in RA, and reduces arterial stiffness and intima-media thickness in patients with RA, PsA, and AS [14, 15].

However, information on the effect of antirheumatic drugs on EF in AS and PsA patients is still limited. Therefore, the aim of this study was to compare EF in RA, AS, and PsA patients, and to examine the effect of antirheumatic treatment (MTX and/or anti-TNF) on EF in these patient groups.

To our knowledge, this is the first study to compare the effect of MTX monotherapy and anti-TNF ± MTX treatment on EF in IA patients, and to compare levels of RHI between RA, PsA, and AS patients with active disease.

In IA patients with ED, antirheumatic treatment was associated with improvement in EF both at 6 weeks and 6 months of follow-up compared to baseline. However, after 6 weeks, EF continued to improve only in the MTX group.

Because MTX monotherapy was initiated only in MTX-naive patients, and the combination therapy only in patients who had previously used MTX without sufficient effect, our findings may indicate that MTX treatment in MTX-naive patients has a greater and more sustained vasculoprotective effect than anti-TNF monotherapy, or anti-TNF added to MTX treatment in MTX nonresponders. It is likely that, in MTX nonresponders, MTX also exhibited a poor response on disease activity after the addition of anti-TNF (MTX in this group was provided first of all to reduce side-effects of anti-TNF therapy). One might speculate that the poor response of MTX on inflammation is associated also with a poor effect on EF.

The exact mechanism behind the protective effect of antirheumatic treatment on ED is not known [25]. Theoretically, it might be mediated by inhibition of systemic inflammatory factors and the corresponding metabolic abnormalities. However, this explanation is not supported by our findings since the improvement in RHI was not related to systemic markers of disease activity, such as ESR and CRP. Moreover, we did not find any significant relationships between RHI levels and inflammatory markers at baseline.

Another explanation might be that the examined drugs might have a direct beneficial effect on the vessel walls, including the endothelium. It has been shown that MTX and anti-TNF treatments are associated with improvements in reverse cholesterol transport by various mechanisms [26, 27]. For example, MTX increases high-density lipoprotein (HDL) capacity to promote cholesterol efflux from cells [28]. Anti-TNF agents counteract the deleterious effects of TNF on the expression of genes involved in cholesterol efflux and reduce cell cholesterol accumulation through amelioration of serum lipoprotein functions and through reverse signaling following direct interaction with cell membrane-bound TNF [26].

Although most focus has been on the importance of impaired cell cholesterol efflux in the development of foam cells from macrophages in atherosclerotic plaques, the same mechanism may also underlie disturbances in endothelial cells, with reduction of their vasodilating and anti-inflammatory functions [29]. In fact, increased cholesterol efflux through the membrane transporters ATP-binding cassette A1 and G1 and Scavenger Receptor class B type I in endothelial cells is associated with promotion of eNOS expression and PGI2 production [30‐32]. Moreover, serum HDL capacity to promote cell cholesterol efflux is directly correlated to flow mediated dilation [33]. Thus, the improved cell cholesterol efflux due to antirheumatic treatment might both protect from atheroma formation and from ED.

IA patients have been reported to have more inflammation, involving overexpression of TNF, in their vascular media and adventitia compared to non-IA patients with CVD [34, 35]. It might even be that inflammation located in deep vascular layers might affect the luminal part of the artery, including the phenotype of the endothelial cells [36]. Thus, in theory, antirheumatic treatment, such as anti-TNF, could also ameliorate EF by inhibition of vascular inflammation.

ED occurs when the endothelium is activated and is characterized by cytokine production, loss of vascular integrity, and expression of adhesion molecules [37]. Adhesion molecules such as intercellular adhesion molecule (ICAM)-1, E-selectin, and vascular cell adhesion molecule (VCAM)-1 make the endothelium surface more adhesive to leukocytes and facilitate their migration into the vessel wall (including atherosclerotic lesions) [37, 38].

In keeping with our results, both MTX and anti-TNF have been previously reported to downregulate expression of adhesion molecules on endothelial cells, i.e., circulating markers of ED [39‐43]. Also, a recent review and meta-analysis article concluded that anti-TNF treatment might improve EF in RA patients [44].

Our previous article based on the same patient sample demonstrated that MTX and anti-TNF ± MTX treatment significantly reduced inflammatory activity (determined by ESR, CRP, WBC count, PGA, and PtGA) both at 6 weeks and at 6 months compared to baseline [16]. This may indicate that both treatment regimens have a longstanding effect on inflammation, but only MTX (in patients potentially responding to it) has a prolonged beneficial effect on the endothelial cells.

We cannot definitely rule out the possibility that the observed differences in the effect of the antirheumatic treatments on EF might be based on differences in patient populations or other factors. For example, it might be that patients with longer and more therapy-resistant IA (i.e., features typical for the anti-TNF ± MTX group; Table 1) had a higher CV risk and were less likely to improve their EF by antirheumatic treatment than the remaining IA patients (Table 2). Nevertheless, the differences in RHI change between baseline and 6 months in the two treatment groups were independent of rheumatic disease duration, IA diagnosis, and age. Moreover, there were no statistically significant differences in the examined traditional CV risk factors, the occurrences of clinical CVD and ED, and median RHI values at baseline between the two treatment groups (Table 2).

As different immune and other mechanisms are involved in the pathogenesis of RA, PsA, and AS, it might be that ED in these diseases is also mediated partly by different factors. Consequently, the effect of different antirheumatic drugs on ED in these particular diseases might also be different.

When comparing the three diagnostic groups, the AS group were the most likely to have ED and CV comorbidity (Table 2), in spite of being younger, having a similar disease duration as the RA and PsA groups, and being less likely to use systemic glucocorticoids than the RA group (Table 1). It is possible that the increased occurrence of certain CV risk factors, such as a high proportion of men and smokers, could partly explain the impaired EF in the AS group [45].

The improvements in RHI from baseline to 6 weeks and 6 months were apparent in all diagnostic groups, but it was statistically significant only in the total IA and in the RA group. The lack of statistically significant differences in RHI improvement in the other groups might be due to their relatively low sample size. Indeed, other studies indicate that antirheumatic treatment (anti-TNF) also improves EF in patients with PsA and AS [46, 47]. We cannot exclude the possibility that AS patients experienced less protection from antirheumatic treatment because they were treated only with anti-TNF and not MTX.

The cause of the observed decreased effect of anti-TNF on EF at 6 months is unclear. Among other factors, it might be caused by the well-known secondary nonresponse effect due to the development of antidrug antibodies [48].

Interestingly, women had statistically greater RHI improvement after 6 months of treatment than men (Table 2). Thus, our results may indicate that women have a better ability to reverse ED than men, independently of IA diagnosis, when treated with MTX or anti-TNF ± MTX. We do not know the molecular mechanism behind this phenomenon.

Rheumatic disease duration showed a stable negative association with RHI change from baseline to 6 months in several multiple regression models. It seems more difficult to achieve an EF improvement in patients with longer rheumatic disease duration, and this applies for both treatment regimens. Thus, these data support the notion that early antirheumatic treatment is important not only for prevention of joint damage, but also for protection from atherosclerosis. However, our results have to be confirmed in larger studies.

As in most studies, ours has several limitations. First, due to ethical reasons (to avoid prescribing MTX to patients in need of anti-TNF, and to avoid overtreatment in patients that might be sufficiently treated with MTX monotherapy) we conducted an observational study instead of a randomized controlled trial (RCT). Thus, we could not secure the same level of similarity between study groups at baseline as in a RCT, nor conduct double-blinded evaluation. On the other hand, observational studies have other advantages, e.g., they can more accurately reflect real life, and therefore have increasingly been called for over the last years. To compensate for baseline differences between the groups, we adjusted for several baseline characteristics in multiple regression models.

As MTX is the drug of choice in most patients with peripheral chronic arthritis, patients with these conditions who receive anti-TNF treatment are likely to have a longer and more severe IA. Nevertheless, the anti-TNF group did not differ from the MTX group when comparing several disease activity markers. In fact, the MTX group had statistically significantly higher PGA scores than the anti-TNF ± MTX group (Table 1).

Second, we were not able to evaluate differences in monotherapies with MTX and anti-TNF as most of the patients using anti-TNF also used MTX comedication.

Third, we evaluated RHI change only in patients with ED because we could not expect substantial RHI improvement in patients with normal EF. Therefore, regression to the mean might be questioned. However, in contrast to patients with low RHI, RHI mean values in patients with normal EF did not change towards the RHI mean at any of the control points of time. Taken together, these observations diminish the suspicion that the observed RHI differences in the ED group could be explained by regression to the mean only.

Finally, owing to a relatively small sample size, the apparent lack of some differences and associations may be due to type II errors and insufficient statistical power. Still, as this is to our knowledge the first study comparing the effect of MTX and anti-TNF regimens in IAs on EF, and comparing EF in RA, PsA, and AS, it brings new important insights into CVD in IAs, and indicates the need for further research.

An advantage of our study is a well-characterized study sample, and a design that makes it possible to examine the effect of two of the main antirheumatic treatment regimens on EF in three common IAs.

In conclusion, treatment with MTX and anti-TNF ± MTX appears to improve EF relatively quickly in IA patients with ED. After 6 months, the EF improvement was more pronounced in the MTX users than in the anti-TNF ± MTX users. Among other factors, this might be due to a more sustained beneficial effect of MTX on the vasculature.

Because the EF improvement was independent of improvement in rheumatic disease activity, modes of action other than the anti-inflammatory effect might play a role.

Among patients with active RA, AS, and PsA, those with AS had the worst endothelial function (the difference was statistically significantly different compared to those with PsA), although they were the youngest.