The aim of this study was to assess the impact of conjugated linoleic acids (CLAs), vitamin E, and combination of these nutrients on serum lipid profiles and blood pressure (BP) in patients with active rheumatoid arthritis (RA). In a randomized, double-blind, placebo-controlled trial, 87 patients with active RA were divided into four groups receiving one of the following daily supplements for three months: Group C: CLAs 2.5 g equivalent to 2 g mixture of cis 9-trans 11 and trans 10-cis12 CLAs in a rate of 50/50; Group E: vitamin E: 400 mg; Group CE: CLAs and vitamin E at above doses: Group P: placebo. After supplementation, SBP levels decreased significantly in the group C in comparison with groups E and P and mean arterial pressure reduced significantly in groups C and CE. There weren’t significant differences in the levels of prostaglandin E2 (PGE2), triglycerides, cholesterol, low-density lipoprotein cholesterol (LDL-C), high-density lipoprotein cholesterol (HDL-C), LDL/HDL, cholesterol/HDL, fasting blood sugar, C-reactive protein (CRP), arylestrase activity, platelet count and body mass index between groups. CRP dropped nonsignificantly in groups P, C, E and CE (19%, 24%, 55%, and 39%, respectively). Erythrocytes sedimentation rate levels decreased in groups C, E and CE (P ≤ 0.05, P ≤ 0.05, P ≤ 0.001, respectively). It is concluded that supplementation of CLAs decreased BP and vitamin E decreased CRP. Therefore cosupplementation of CLAs and vitamin E might be profitable for heart disease prevention in RA patients.

Rheumatoid arthritis (RA), as the most common inflammatory disease, is associated with excess cardiovascular morbidity and mortality that is not entirely explained by traditional risk factors of cardiovascular disease (CVD) (De Pablo et al 2007). It might be due to an increased prevalence of cardiovascular risk factors such as dyslipidemia (Nurmohamed 2007). Risk factors for CVD, including inflammatory biomarkers, low intake of antioxidants, and vitamins, may contribute to excess CVD in persons with RA (Dessein et al 2006). There is a complex relationship between inflammation, antioxidant vitamin status, and the risk of CVD. Furthermore, low intakes of antioxidant vitamins and other micronutrients, including vitamin E, have been reported to be inversely associated with CVD incidence and mortality (De Pablo et al 2007). Conjugated linoleic acids (CLAs) are a group of naturally occurring isomers of linoleic acid that differ in the position or geometry of their double bonds (Kelly 2001; Bhattacharya et al 2006). The biological activities of CLAs have received considerable attention because of its anticancer (Parodi 1999), antiatherogenic (Rudel 1999; Arbonés-Mainar et al 2006) and antidiabetic effects (Peck et al 1998). The atheroprotective properties of CLAs have been particularly well documented (Kritchevsky et al 2000; Toomey et al 2003). Inclusion of CLAs in a hypercholesterolemic diet in rabbits led to a significant reduction in serum triglyceride (TG), low-density lipoprotein cholesterol (LDL-C) and cholesterol (CHO) levels. In addition, there was a reduced incidence of atherosclerotic plaque formation in the rabbit aorta (Lee et al 1994). Similar results have also been reported for hamster hypercholesterolemia models (Nicolosi et al 1997; Wilson et al 2000). Despite these findings, some reports suggest that CLAs may possibly have proatherogenic effects (Munday et al 1999; Gavino et al 2000). There are several molecular mechanisms for explaining the hypotriacylglycerolemic effect of CLAs. The cis-9, trans-11 and the trans-10, cis-12 isomers of CLAs are potent peroxisome proliferators-activated receptor (PPAR) agonists. PPARα is a key transcription factor that regulates hepatic lipid metabolism (Moya-Camarena and Belury 1999a). Feeding CLAs-enriched diets increases the expression of PPARα-responsive genes in animals (Moya-Camarena and Belury 1999b). Therefore, the TG-lowering effect of CLAs may be partly attributed to the effect of CLAs on PPARα. CLAs is also a PPARγ ligand. In adipose tissue, PPARγ regulates the expression of the genes that determine adipogenesis, lipid metabolism and insulin sensitivity (Lowell et al 1999; Moya-Camarena et al 1999). Some other proposed mechanisms include their role on peroxisome proliferators-sterol regulatory element-binding proteins (SREBPs) (Pai et al 1998) and stearoyl-CoA desaturase (SCD) (Miyazaki et al 2000, 2001; Ntambi 1999). SREBP1c isoforms regulate fatty acid and TG synthesis (Pai et al 1998). Studies suggest that liver SREBP-1c expression is dependent on the nuclear hormone liver X receptor (LXR). C9 t11 isomer was shown to down-regulate mRNA expression of LXR-α and SREBP-1c, whereas the t10 c12 isomer had no effect (Roche et al 2002). The results suggested that c9t11 isomer positively influences lipid metabolism by reduced synthesis and cleavage of hepatic SREBP-1, which in turn is regulated by LXRα expression. SREBP-1c enhances the transcription of the genes required for fatty acid synthesis and fatty acid elongation including fatty acids synthetase (FAS) and SCD (Pai et al 1998; König et al 2008). Hypertension is also a common pathological state associated with an increased risk of cardiovascular diseases. Some studies have shown that CLAs (50:50) or the t10c12 isomer decreases blood pressure (BP) and hypertension in various rat models prone to develop obesity, diabetes and hypertension (Nagao et al 2003a, 2003b; Alice et al 2005). One possible mechanism is through inhibition of cyclooxygenases, both of them have been implicated in the development of atherosclerosis (Belton et al 2003). c9, t11- and t10, c12-CLAs isomers are not substrates for cyclooxygenases (COX) but compete with arachidonic acid for access to the enzymes. In one study, it was seen that feeding rabbits with an atherogenic diet rich in CLAs reduced atherosclerotic lesion sizes (Valetille et al 2005). A number of studies in rabbits have considered the potential for the lipophilic antioxidant vitamin E to inhibit atherosclerosis in vitro by decreasing lipid peroxidation of LDL (Schwenke et al 1998). Some epidemiological studies have shown an association between high dietary intake and high serum concentrations of vitamin E and lower rates of ischemic heart disease (Gey et al 1990; Rimm et al 1993; Stampfer et al 1993) and myocardial infarction (Stephens et al 1996). Here, we investigated the effect of 50:50 blend of c9, t11: t10, c12-CLAs on BP and lipid profiles in active RA patients.

In RA disease, chronic systemic inflammation may contribute to the higher incidence of CVD. (Hannawi et al 2007; Metosios et al 2007). To our knowledge; this is the first study which investigated the effects of CLAs and its combination with vitamin E in human with RA disease. In the present study, we didn’t find any significant changes in the plasma levels of TG and CHO in any of the studied groups. The magnitude of the change in plasma TG concentrations in our study is comparable with that achieved following 50:50 CLAs supplementation in Benito and colleagues’ (2001) study. They did not find any effect on plasma CHO, lipoprotein, or TG. Noone and colleagues’ (2002) study did not show any modification of plasma CHO and lipoprotein concentrations in healthy subjects receiving the 50:50 CLAs mix, however they showed a decrease in the TG concentration. Some other studies showed that trans-10, cis-12 CLAs isomer may be the effective hypotriglycerolemic isomer (Gavino et al 2000; Lin et al 2001). The TG-lowering effect of CLAs may be partly attributed to the effect of CLAs on PPARα. Furthermore, CLAs is a PPARγ ligand. In adipose tissue, PPARγ regulates the expression of the genes that determine adiposeness, lipid metabolism, and insulin sensitivity (Houseknecht et al 1998; Lowell 1999). In our study, there was a nonsignificant decrease in CHO in CE group. We used CLAs capsules with equal proportions of the cis-9, trans-11 and trans-10, cis-12 CLAs in triglyceride form. Recent vitro and animal studies suggest that the individual CLAs isomers may have different effects on lipid metabolism. Human studies regarding CHO-related parameters were not in line with the animal ones. The inconsistency of results may arise from the effects of particular CLAs consists the usual CLAs mix. Moreover, the dose of CLAs mix administered to humans, on a weight basis, was much lower than that for animal models and may not reach the effective level (Smedman and Vessby 2001). However, the effects of the CLAs-enriched dairy products on blood lipids were more complex. In our study similar to Desroches and colleagues’ (2005) study after the consumption of the CLAs supplements, LDL-C, and HDL-C concentrations and the ratio of total to HDL-C and LDL-C to HDL-C were not significantly changed. Singhal and colleagues (2001) study results in patients with coronary heart disease were similar to our study and plasma TG concentration didn’t change significantly in the vitamin E-supplemented group. A large base of epidemiological evidence suggests that a 1 mg/dl increment in HDL-C would be associated with a 2%–3% decrement in CVD risk (Gordon et al 1989). Our data show that supplementation of vitamin E and its combination with CLAs for at least three months can increase HDL-C nonsignificantly by 3.4% and 4.5%, respectively. Separate supplementation of CLAs did not improve serum HDL-C levels. The absence of such effects of CLAs supplementation should not be attributed to the interference of weight, physical activity, dietary intake or medication because these variables did not change during the study period and the patients had good compliance in taking their supplements. α-Tocopherol possibly has power to inhibit oxidation induced by α-tocopheroxyl radical (Singh et al 2005) and known synergistic action between vitamin E and CLAs (Kim et al 2005; Santoz-Zago et al 2007). Other studies have also shown that CLAs supplementation had no effect on HDL lipid composition (Nicolosi et al 1997; Gavino et al 2000; Stangl 2000). In our study, LDL-C increased in all groups and the difference between groups was not significant. Nicolosi and colleagues (2004) also showed that non-HDL-C concentrations were reduced by (9%–13%) in response to a CLAs-enriched diet. The LDL-C/HDL-C was reduced in the 80:20 CLAs group (27.2%) and the 50:50 CLAs group (22.4%), however these weren’t significant (Nicolosi et al 2004). In our investigation, CLAs plus vitamin E was more effective than CLAs in reducing LDL-C/HDL-C, CHO/HDL-C, and CRP. In the present study, FBS didn’t change significantly in four groups. Few studies have evaluated the effect of CLAs intake on glucose metabolism and insulin sensitivity in humans, but the available data suggest that glucose metabolism may be negatively affected by CLAs isomers. In Riserus and colleagues’ (2001) study, a significant increase in glucose levels was observed in both the CLAs and the control group. The increase was more pronounced in the control group (Riserus et al 2001). In another study, glucose levels increased significantly only in the group receiving the trans10 cis 12 isomer and not in the case of a mixture of different isomers. The increase was not significantly different from that which occurred in the control group (Riserus et al 2002). Smedman and Vessby (2001) also showed an increase (p = 0.054) in glycemia after the consumption of a mixture of CLAs isomers compared with the control group. They observed that insulin levels tended to increase after an intake of 2 g CLAs (Smedman and Vessby 2001). Several other studies have shown no deleterious effects on glucose or insulin levels or sensitivity to insulin attributable to the consumption of different doses of CLAs enriched with cis 9, trans 11 or trans 10, cis 12 (Tricon et al 2004; Taylor and Zahradka 2004). Results on the effect of CLAs on glucose metabolism from animal studies are controversial (Houseknecht et al 1998; Stangl 2000; Tsuboyama-Kascoke et al 2000). In our study, CLAs supplementation had no effect on body weight. This finding is in agreement with Zambell and colleagues (2000) who found no changes in body weight or body composition attributed to CLAs supplementation in human subjects. However some studies have reported that CLAs supplementation (4 g/d) had no effect on body weight, but they significantly reduced body fat (Smedman and Vessby 2001). It was shown that, 9c, 11t-CLAs, 10t, 12c-CLAs inhibited arachidonic acid and collagen-induced platelet aggregation (Truitt et al 1999). In the present study, the number of platelet decreased nonsignificantly in groups P, C, and CE. Leonard and colleagues (2007) studied the effects of vitamin E (400 IU) on cholesterol levels of hypercholesterolemic patients receiving statins. Similar to our study they didn’t find significant differences in any lipoprotein cholesterol fraction. However, unlike our study, a 6% decrease in HDL-C was detected within the vitamin E group (p < 0.05) (Leonard et al 2007). Following our study, LDL-C increased in all groups, but this increase was only significant in group E. This increase may be due to increasing the size of LDL-C or changing its fractions or by increase of its half life by decreasing LDL-C oxidation. In our study AEA, decreased nonsignificantly in groups E, P, and C, but decreased as compared with baseline in group E (P = 0.068). We had nonsignificant differences between four groups. In one study on pedigreed baboons, dietary vitamin E improved LDL quality and showed two apparently paradoxical effects on HDL metabolism: lower HDL2 and higher apo A-I. These effects have contrasting associations with CVD risk and may help account for the mixed results from clinical trials of dietary vitamin E. In our study, nonsignificant reductions in serum AEA of lipoprotein-associated enzymes, Paraxonase1 (PON1) in all groups except group CE was shown. The presence of PON1 on HDL particles is considered to be a major source of protection from lipoprotein oxidation. Therefore, reduction of PON1 activity may be considered to be prooxidant and pro-atherogenic (Rainwater et al 2007). Our results contradict results in rabbits and humans (Jarvik et al 2002; Jeon et al 2005). This discrepancy may reflect differences in HDL size and composition properties among the species. Alternately, vitamin E-associated reduction in oxidative stress and oxidized LDL may have lowered the metabolic requirements for these enzymes to maintain the redox balance in the circulation. Then, through feedback mechanisms, the expression of PON1 may have been downregulated (Jarvik et al 2003; Rainwater et al 2005). Furthermore the differences between groups wasn’t significant. Following of our study ESR levels decreased significantly in the groups C, E, and CE compared with baseline (P ≤ 0.05 P ≤ 0.05, P ≤ 0.001, respectively). ESR levels in group CE was significantly lower than group P (P ≤ 0.05). Vitamin E was reported to regulate transcription or activity of a number of genes involved in oxidative stress, proliferation, inflammation, and apoptosis (Dutta and Dutta 2003; Tucker and Townsend 2005; Aldred et al 2006). CRP dropped in all groups in comparison with baseline, but this decreased more in group E (P = 0.054). The reduction of WBC in group CE was significant compare with other groups (P < 0.05) (Table 3). PLT count decreased nonsignificantly in CE, C, E groups. Consequently CLAs, vitamin E, and their combination are useful in reduction of inflammation. Many studies (Igarashi and Miyazawa 2001; Kim et al 2002; Palacios et al 2003; Bergamo et al 2006, 2007; Santos-Zago et al 2007) show the antioxidant effect of CLAs, but several studies performed on healthy and obese subjects have observed that CLAs administration is associated with an increase in some lipid oxidation and peripheral inflammation parameters (Riserus et al 2002, 2004; Basu et al 2000a, 2000b; Smedman et al 2004). One of our aims was to investigate the effect of α-tocopherol and CLAs supplementation on BP. BP levels decreased significantly in the group C by 8% and 8.5% for DBP and MAP respectively, after three months of supplementation as compared with baseline. The reduction of SBP in group C was significant compared with the groups E and P (Table 2). Herrera and colleagues (2005) studied healthy primigravidas who had a family history of pre-eclampsia and diastolic notch. Calcium-CLAs supplementation reduced significantly the incidence of pregnancy-induced hypertension (PIH) in the study group (P = 0.01). Endothelial dysfunction was also significantly reduced after calcium-CLAs supplementation (Herrera et al 2005). Herrera and colleagues (2006) in another study on healthy primigravidas showed that the incidence of PIH was significantly reduced in women receiving the CLAs supplement compare with the placebo. There were no significant differences in the plasma concentrations of ionized calcium, PE2 (similar to our study), renin, angiotensin II, parathormone, and calcitonine (Herrera et al 2006). In Raff and colleagues’ (2006) investigation, a diet rich in CLAs didn’t alter SBP, DBP and pulse pressure, but this may be due to the short time for the study (five weeks). Nagao and colleagues (2003a) found that the relative mRNA expressions of angiotensinogen and leptin were suppressed by 10t, 12c-CLAs in adipose tissue. They speculate that the antihypertensive effect of 10t, 12c-CLAs can be attributed to the lowered secretion of hypertensive adipocytokines from abdominal adipose tissues (Nagao et al 2003a). In another study Nagao and colleagues (2003b) showed that CLAs suppresses the development of nonobese essential hypertension in spontaneously hypertensive rats (SHRs). They concluded that the antihypertensive effect of dietary CLAs may be due to the increase of plasma adiponectin level and is associated with the alleviation of membrane abnormality in SHRs (Nagao et al 2003b). In our study in group E, BP increased nonsignificantly, in Ward and colleagues’ (2007) study BP, pulse pressure, and heart rate increased versus placebo.

It is concluded that supplementation of 2 g CLAs 50:50 mix, and its combination with vitamin E for 12 weeks didn’t change lipid profiles and FBS significantly, but CLAs decreased BP and vitamin E decreased CRP so co-supplementation of CLAs and vitamin E could be helpful for heart disease prevention in RA patients.