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Integrative Physiology/Experimental Medicine

Rimonabant, a Selective Cannabinoid CB1 Receptor Antagonist, Inhibits Atherosclerosis in LDL Receptor–Deficient Mice

Frédérique Dol-Gleizes, Réjane Paumelle, Virgile Visentin, Anne-Marie Marés, Perrine Desitter, Nathalie Hennuyer, Andries Gilde, Bart Staels, Paul Schaeffer, Françoise Bono
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https://doi.org/10.1161/ATVBAHA.108.168757
Arteriosclerosis, Thrombosis, and Vascular Biology. 2009;29:12-18
Originally published December 17, 2008
Frédérique Dol-Gleizes
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Réjane Paumelle
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Virgile Visentin
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Anne-Marie Marés
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Perrine Desitter
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Nathalie Hennuyer
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Andries Gilde
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Bart Staels
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Paul Schaeffer
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Françoise Bono
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Abstract

Objective— The objective of this study was to determine whether the potent selective cannabinoid receptor-1 antagonist rimonabant has antiatherosclerotic properties.

Methods and Results— Rimonabant (50 mg/kg/d in the diet) significantly reduced food intake (from 3.35±.04 to 2.80±0.03 g/d), weight gain (from 14.6±0.7 g to −0.6±0.3 g), serum total cholesterol (from 8.39±0.54 to 5.32±0.18 g/L), and atherosclerotic lesion development in the aorta (from 1.7±0.22 to 0.21±0.037 mm2) and aortic sinus (from 101 000±7800 to 27 000±2900 μm2) of LDLR−/− mice fed a Western-type diet for 3 months. Rimonabant also reduced plasma levels of the proinflammatory cytokines MCP-1 and IL12 by 85% (P<0.05) and 76% (P<0.05), respectively. Pair-fed animals had reduced weight gain (6.2±0.6 g gain), but developed atherosclerotic lesions which were as large as those of untreated animals, showing that the antiatherosclerotic effect of rimonabant is not related to reduced food intake. Interestingly, rimonabant at a lower dose (30 mg/kg/d in the diet) reduced atherosclerosis development in the aortic sinus (from 121 000±20 000 to 62 000±11 000 μm2, 49% reduction, P<0.05), without affecting serum total cholesterol (7.8±0.7 g/L versus 8.1±1.3 g/L in the control group). Rimonabant decreased lipopolysaccharide (LPS)- and IL1β-induced proinflammatory gene expression in mouse peritoneal macrophages in vitro as well as thioglycollate-induced recruitment of macrophages in vivo (10 mg/kg, po bolus).

Conclusions— These results show that rimonabant has antiatherosclerotic effects in LDLR−/− mice. These effects are partly unrelated to serum cholesterol modulation and could be related to an antiinflammatory effect.

  • atherosclerosis
  • rimonabant
  • obesity
  • LDLR-deficient mice

Rimonabant is the first cannabinoid receptor type 1 (CB1) antagonist to reach advanced development for the treatment of obesity and the associated cardio-metabolic disorders in humans.1,2 The role of the endocannabinoid system in different physiological and pathological mechanisms is now being more and more recognized.3 The main endocannabinoids anandamide (arachidonoylethanolamide, AEA) and 2-arachidonoylglycerol (2-AG) are produced during cellular activation and activate G protein–coupled receptors, two of which, CB1 and CB2 receptors, have been cloned and characterized biochemically. Whereas CB1 receptors are mainly expressed in the brain, CB2 receptors have been localized essentially to immune cells, but both receptors are also present in cardiovascular tissues, albeit at lower levels.3 CB2 receptor activation has been shown to attenuate tumor necrosis factor (TNF)α-induced smooth muscle4 and endothelial cell5 activation and leads to decreased atherosclerosis in ApoE-deficient mice,6 clearly demonstrating the role of CB2 receptors in the progression of atherosclerosis. The role of CB1 receptors in atherosclerosis is less clear, although the protective effects of CB1 receptor inhibition on doxorubicin-induced cardiotoxicity7 and contractile dysfunction in cirrhotic animals8 point to a potential interest of CB1 receptor blockade in cardiovascular disorders.

See accompanying article on page 7

However, obesity by itself is now generally recognized as a significant risk factor for atherosclerosis and cardiovascular disease,9,10 and leads to increased levels of circulating endocannabinoids11 and dysregulation of the adipose tissue endocannabinoid system.12 Recent studies have shown that fat tissue is not a simple energy storage organ but exerts important endocrine and immune functions. These are achieved predominantly through release of adipocytokines like leptin or adiponectin, as well as some more classical cytokines including MCP-1 and IL12. These cytokines are probably released by inflammatory cells infiltrating fat, eg, macrophages, and might aggravate the preexisting proinflammatory condition in atherosclerotic lesions.10 Although the significant weight loss induced by rimonabant is therefore of potential interest in the field of atherosclerosis, rimonabant has also been shown to produce an improvement in other cardiovascular risk factors, eg, a decrease of the LDL/HDL cholesterol ratio and triglycerides in overweight patients,13 and also C reactive protein in dyslipidemic obese patients.14 The effects of rimonabant on these secondary end points could only partly be explained by weight loss, suggesting that rimonabant has additional metabolic or protective effects leading to a further decrease in risk factor levels.

Considering the beneficial effect of rimonabant on serum lipid parameters in patients as well as animal models,15 rimonabant would be expected to decrease atherosclerotic lesion development. However, atherosclerosis is a multifactorial disease, and besides dyslipidemia, inflammation is now recognized as one of the major factors influencing the course of the disease. Interestingly, rimonabant has been shown to reduce the levels of the proinflammatory cytokine TNFα in a model of obesity-associated hepatic steatosis,16 and is able to inhibit LPS-induced TNFα increase17 and adjuvant-induced arthritis in rats,18 suggesting that it might be able to directly or indirectly decrease the inflammatory component of atherosclerotic plaque progression.

To assess the potential effect of rimonabant on atherosclerosis, we have now evaluated the effect of this compound in several in vitro and in vivo models related to lesion formation. As a first step, the effect of rimonabant on lesion development was studied in LDLR−/− mice on high-fat diet. This mouse model of atherosclerotic lesion development is particularly interesting, because it has been shown previously that male LDLR−/− mice are more sensitive than apoE-deficient mice (ApoE−/−) to diet-induced obesity,19 and rimonabant is essentially targeted to obese and diabetic patients. To clarify the mechanism of action of rimonabant, we also determined whether rimonabant is able to modulate inflammatory gene expression in macrophages, which are one of the major cell types involved in atherosclerotic lesion progression. The effect of rimonabant on macrophage recruitment in vivo was also assessed.

Materials and Methods

For expanded Materials and Methods, please see the online data supplement, available online at http://atvb.ahajournals.org.

Mice

Male LDLR-deficient mice (LDLR−/−) on a C57BL/6 background were obtained from Charles River Company, L’Arbresle, France, at the age of 3 months. Western diet (0.21% cholesterol; 20% fat) with or without rimonabant was from Research Diets. Mice were weighed every week and food ingestion was measured every day to restrict the food intake of the pair fed group. All procedures and care of animals were approved by the Animal Care and Use Committee of sanofi-aventis R & D.

Determination of Atherosclerosis, Serum Lipoproteins, and Cytokines

For atherosclerotic area determination, thoracic and abdominal segments of the aorta were opened longitudinally, pinned onto paraffin, and stained with Sudan black B. Serum lipoprotein and cytokine levels were measured by commercially available enzymatic or ELISA kits.

Peritoneal Macrophage Isolation and Cell Culture

Murine thioglycollate-elicited peritoneal macrophages were isolated as described (Bellosta et al, 1998). Macrophage mRNA levels were measured by quantitative polymerase chain reaction (PCR).

Peritonitis Model of Macrophage Recruitment

Female CB1R+/+ and CB1R−/− mice on the C57Bl/6 background between 8 and 14 weeks of age were used (12 to 14 per group). Peritonitis was induced by a single intraperitoneal injection of 1 mL 4% thioglycollate.

Statistical Analysis

Statistical analysis of the in vitro experiment was performed using multivariate ANOVA tests followed by Scheffe post hoc test. In vivo experiments were analyzed using the nonparametric Kruskal-Wallis test or multivariate ANOVA tests followed by either Scheffe post hoc test or Dunnet post hoc test. Values of P<0.05 were considered as significant.

Results

Effect of Rimonabant on Atherosclerosis in LDLR−/− Mice Fed a Western-Type Diet as Compared to Pair-Fed Mice

As rimonabant would be expected to reduce food intake, a first experiment was designed to allow discrimination between effects of rimonabant resulting from a reduction of food intake and other effects of rimonabant. Therefore, 4 different groups of LDLR−/− mice were used: one group of mice was fed a standard diet (3% fat, no cholesterol), one group was fed a Western diet (0.21% cholesterol and 20% fat) ad libitum, a third group was fed a Western diet containing rimonabant (0.45 g/kg of food), and finally one group was fed Western diet but with access to food restricted to the mean amount of food consumed by the rimonabant-fed group of mice (pair fed group).

As shown in Figure 1A, during a 3 month period, the weight of LDLR-KO mice fed a Western diet increased significantly more than the weight of the standard diet group, even though the Western diet group ate less food than the standard diet group (Figure 1B). Actually, when food intake was expressed on energy (J) and not on a weight (g) basis, the daily calorie uptake was slightly higher for mice fed Western diet as compared to chow diet (65.9±0.8 versus 63.0±0.7 kJ/d, respectively).

Figure1
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Figure 1. Effect of rimonabant on body weight (A) and food intake (B) in LDLR−/− mice fed either standard chow diet (▾), Western-type diet (○), or Western-type diet containing rimonabant (50 mg/kg/d, •) or pair fed Western type diet (▿). mean±SEM, n=10 to 17.

Rimonabant had a strong effect on the body weight of treated mice. Indeed, their weight did not increase over time, and stayed below the weight of the standard diet group. As shown in Figure 1B, the food intake of the rimonabant-treated group was decreased in comparison to the Western diet group. The average food intake of the rimonabant group was 2.8 g per day, giving a calculated dosage of approximately 50 mg/kg. This effect of rimonabant on food intake was expected because it has already been reported in prior studies.20,21 Nevertheless, the weight loss in the rimonabant-treated group was more pronounced than in the pair fed group, showing that the effect on weight loss is not only attributable to a restriction in food intake.

LDLR-KO mice maintained on a standard chow diet had a higher level of cholesterol (total or LDL fraction) than control C57BL/6 mice, but their level of triglycerides was close to the value in control C57BL/6 mice (supplemental Table I). The serum lipid levels were dramatically increased after a 3-month period of Western diet. Surprisingly, the pair-fed group which had a restricted access to food had lipid levels which were comparable to the levels of the Western diet group. Rimonabant (50 mg/kg) reduced lipid levels by 37% for total cholesterol, 50% for LDL cholesterol and 41% for triglycerides (supplemental Table I). On a reduced number of animals, (n=5), the results on total and LDL cholesterol were confirmed by high-performance liquid chromatography (HPLC) analysis and showed an increase in HDL cholesterol (48%) for the rimonabant treated group (not shown). Whatever the diet, LDLR−/− mice after a 3-month period had normal glycemia and insulinemia. They did not develop any diabetes.

LDLR−/− mice did not develop significant atherosclerotic lesions when fed a standard diet for 3 months, although older animals are known to develop spontaneous lesions under these conditions. Conversely, fatty streak formation in the aorta was dramatically increased after a 3 month period of Western diet (supplemental Figure IA and Figure 2A). Similarly, the atherosclerotic plaque size was dramatically increased in the aortic sinus in the Western diet group compared with the standard diet group (supplemental Figure IC and Figure 2B). More interestingly, the levels of atherosclerotic lesions were identical in the aorta or in the aortic sinus whatever the Western diet group, fed ad libitum or pair-fed to the rimonabant group. A 3-month treatment with rimonabant nearly abolished the atherosclerotic development in the aorta (90% reduction compared with pair-fed group, P<0.001) or in the aortic sinus (70% reduction, P<0.01).

Figure2
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Figure 2. Effect of rimonabant (50 mg/kg/d) on atherosclerotic lesion size in LDLR−/− mice. Top, Effect of rimonabant (50 mg/kg/d) as compared to pair-feeding in the aorta (A) and aortic sinus (B). Bottom, Effect of increasing doses of rimonabant (10 mg/kg/d and 30 mg/kg/d) in the aorta (C) and aortic sinus (D; mean±SEM).

To clarify the mechanism of action of rimonabant, the serum levels of some adipokines and chemokines were also measured. Serum leptin increased after a 3-month period of Western diet from 8.7±1.1 ng/mL in the standard diet group to 45.8±6.5 ng/mL in the Western diet group fed ad libitum (supplemental Table II). In the group with a restricted access to food (pair-fed group) leptin levels were lower at 16.8±2.5 ng/mL. In the group treated with rimonabant, circulating leptin levels were dramatically reduced reaching a value of 0.53 ng/mL, much lower than the value obtained in the standard diet group.

In contrast, adiponectin levels were identical in untreated animals, whatever the composition of the diet or the food intake (supplemental Table II). Interestingly, in the rimonabant-treated group, the levels of adiponectin were significantly increased in comparison to the pair fed group, from 6.9±0.5 μg/mL to 8.1±0.5 μg/mL (P<0.05).

To assess the inflammatory state of the animals, MCP1, IL12p40/p70, and vascular cell adhesion molecule (VCAM)-1 levels were also determined. For these 3 proteins, the levels increased during the 3 month-period of Western diet, showing that this diet was able to induce an inflammatory state. The restriction of food intake in the pair-fed group had no influence on any of the 3 markers. Conversely, rimonabant treated animals had significantly reduced MCP-1, IL12p40/p70, and VCAM-1 levels (supplemental Table II).

Dose-Effect of Rimonabant on Atherosclerosis in LDLR−/− Mice Fed a Western-Type Diet

Considering the impressive effects of rimonabant, a second study was designed to test lower doses of rimonabant. Two groups of animals treated with different doses of Rimonabant (average dose of 10 mg/kg and 30 mg/kg in the diet) were compared to a Western diet group.

Although doses were lower, rimonabant again induced a decrease in body weight as compared to the untreated animals: final weights at the end of the 3-month treatment were reduced by 15% (P<0.001) and 20% (P<0.001) in the 10 mg/kg– and 30 mg/kg–treated animals, respectively (supplemental Table III). Again, triglyceride levels were also decreased; they were respectively 27% (ns) and 49% (P<0.05) lower in the 10 mg/kg group and 30 mg/kg-treated groups as compared to the control group (supplemental Table III). In this study, triglyceride levels in the control group were higher than in the previous study because blood was withdrawn without fasting the animals.

Interestingly, total cholesterol and LDL cholesterol levels were not significantly different between the control group and the rimonabant 10 mg/kg–treated group. In the rimonabant 30 mg/kg–treated group, total cholesterol was decreased by 4% (ns) and LDL cholesterol by 14% (ns, supplemental Table III). As in the previous study, no changes in glycemia or insulinemia were observed between the groups.

In this study, the fatty streak lesions in the thoracic aorta covered a surface of 1.1±0.37 mm2, which is comparable to the area obtained for the untreated Western diet–fed group of the first study (1.60±0.13 mm2; Figure 2C). A 67% to 68% reduction of lesion size was observed in atherosclerosis development for both the 10 mg/kg– and 30 mg/kg–treated groups (0.34±0.12 mm2 (P=0.065) and 0.35±0.09 mm2 (P=0.068), respectively. At the aortic sinus level, the cumulative plaque surface was 121 000±20 000 μm2 in the control animals, again in the same range as the untreated group of the first study. In the rimonabant 10 mg/kg-treated group, the aortic sinus plaque area was reduced by 9% (110 000±15 000 μm2, ns), whereas, in the rimonabant 30 mg/kg-treated group, the plaque area was 49% smaller than in control animals (62 000±11 000, P<0.05; Figure 2D).

As could be expected from the effect on animal weight, rimonabant significantly decreased leptin levels both at 10 mg/kg and 30 mg/kg (by 57% [P<0.05] and 74% [P<0.001] respectively, supplemental Table III). Adiponectin levels were slightly, but not significantly, increased at the 30 mg/kg dose, whereas MCP-1 levels were not significantly affected (supplemental Table III).

Effect of Rimonabant on Inflammatory Gene Expression in Mouse Peritoneal Macrophages

Taking into consideration the effects of rimonabant on inflammatory marker levels in vivo, we also determined whether rimonabant is able to reduce inflammatory markers in macrophages, one of the main cell types involved in atherosclerosis-related inflammation. As expected, LPS (Figure 3A) and IL1β (Figure 3B) induced a strong increase in IL6, TNFα, and MCP1 expression in mouse peritoneal macrophages. This effect was identical in macrophages from male and female animals (relative IL6, TNFα, and MCP1 levels were 4860±840 versus 4670±1500, 98±37 versus 78±17, and 430±84 versus 401±41 in male versus female animals, respectively). Rimonabant (0.3 to 1 μmol/L) significantly reduced LPS as well as IL1β-induced IL6 and TNFα mRNA expression, but did not significantly affect MCP1 mRNA levels in these cells. Similar effects were also observed in RAW 264,7 murine macrophages (supplemental Figure II). To determine whether these effects of rimonabant are attribtuable to CB1 receptor blockade, the same experiments were performed in parallel in peritoneal macrophages from CB1-receptor wild type (CB1R+/+) and CB1-receptor deficient (CB1R−/−) mice. As shown in supplemental Figure III, rimonabant (1 μmol/L) reduced LPS-induced IL6 and TNFα mRNA expression to a similar extent in CB1R+/+ and CB1R−/− mice, suggesting that this effect of rimonabant was not related to CB1-receptor antagonism.

Figure3
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Figure 3. Effect of rimonabant on inflammatory gene expression induced by LPS and interleukin (IL)-1β in peritoneal macrophages. Macrophages were pretreated with rimonabant for 2 hours followed by 3 hours of LPS (100 ng/mL; A) or IL-1β (10 ng/mL; B).

Effect of Rimonabant on Macrophage Recruitment in a Mouse Model of Thioglycollate-Induced Peritonitis

Resistance to thioglycollate injection induced macrophage recruitment into the peritoneal space has been correlated with atherosclerosis resistance in different mouse strains.22 Although the effect of thioglycollate was not different in CB1R+/+ and CB1R−/− mice (Figure 4), rimonabant (10 mg/kg, po bolus) significantly reduced thioglycollate-induced macrophage recruitment in CB1R+/+ mice (37% reduction, P<0.01), but was inactive in CB1R−/− mice.

Figure4
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Figure 4. Effect of rimonabant on thioglycollate-induced macrophage recruitment in CB1R+/+ and CB1R−/− mice. Mice were pretreated with rimonabant (10 mg/kg) or vehicle (methylcellulose) for 1 hour and injected with thioglycollate 4%. Mice were then treated with rimonabant or vehicle once a day during 3 days (mean±SEM; n=18).

Discussion

The beneficial effects of rimonabant on weight, serum HDL/LDL cholesterol ratio, triglyceride levels, and serum inflammatory markers in clinical studies suggest that this compound might reduce atherosclerotic lesion development in obese patients. Western diet–fed LDLR−/− mice barely develop lesions on normal diet, but gain weight and develop extensive lesions on a high-fat Western-type diet,19,23 and would therefore be expected to at least partially mimic the patient population used in the clinical studies of rimonabant.

Interestingly, a 3-month treatment of LDLR−/− mice with rimonabant (10 to 50 mg/kg/d) incorporated into Western diet resulted in a dose-dependent inhibition of atherosclerosis development in the aorta and aortic sinus, together with a decrease in body weight, which was even stronger than the effects reported in previous studies in high-fat fed mice.15,21 However, the lesion size in pair-fed animals did not differ significantly from the lesion size in Western diet–fed untreated animals, demonstrating that the antiatherosclerotic effect of rimonabant was not related to inhibition of food intake.

Surprisingly, the relationship between atherosclerosis reduction and reduction of serum cholesterol and inflammation markers appeared to be nonlinear. Actually, serum triglycerides and leptin levels were already decreased at low doses (30 and 10 mg/kg/d, respectively), whereas serum cholesterol, IL12, and MCP-1 were only reduced, and adiponectin was only increased, at the highest dose of rimonabant (50 mg/kg/d). This clearly suggests a complex mechanism of action for the antiatherosclerotic effect of rimonabant, with some mechanisms, like decreased leptin and perhaps triglyceride lowering, being present at low doses of rimonabant and others, like serum cholesterol changes and adiponectin, additionally playing a role at higher doses. However, the serum cholesterol decrease at a high dose of rimonabant was much stronger than the decrease observed in humans,13 suggesting that the importance of this effect might be exacerbated in mice as compared to patients.

Weight loss was already present at the lowest dose of rimonabant and must therefore be taken into consideration as one of the potential determinants of the antiatherosclerotic effect of rimonabant. Actually, although the link between obesity and cardiovascular disease is still unclear, emerging evidence indicates that factors produced by adipose tissue may directly influence vascular disease. Leptin, one of these factors, is produced primarily by adipocytes, and leptin levels are correlated to body mass index in rodents and humans.24 Recently, an association between elevated leptin levels and cardiovascular events has been demonstrated in patients,25 exogenous leptin has been shown to increase atherosclerosis in apoE−/− mice,26 and the adipokine has been shown to modulate the development of atherosclerosis in LDLR−/− mice through the immune response.27 As rimonabant at 30 mg/kg/d normalized leptin levels of Western diet–fed mice to the levels found in mice on normal chow diet, the effect of rimonabant on leptin levels could clearly contribute to the antiatherosclerotic effect of the lower doses of the compound. Interestingly, the reductions in leptin levels induced by rimonabant were more pronounced than expected from the weight reduction alone, especially at the higher dose. As inflammatory cytokines are known to induce leptin expression,28 and rimonabant at the highest dose reduced inflammatory cytokine levels, one possible mechanism for this additional effect of rimonabant could be related to a decrease of leptin expression after inhibition of inflammation in the animals.

Low doses of rimonabant have actually been shown to exert antiinflammatory effects in several different inflammation-related animal models,16–18 and this also suggested the idea to check whether the antiatherosclerotic effects of rimonabant could be consistent with an antiinflammatory effect at lower doses of the compound. The lower dose of rimonabant, which remained antiatherosclerotic (30 mg/kg/d), reduced circulating MCP1 levels by more than 20%, but this effect did not reach statistical significance. However, MCP1 levels in atherosclerotic animals were quite fluctuating, and we therefore used a direct induction of the stronger inflammation induced by LPS to study the potential antiinflammatory effects of rimonabant. The in vitro effects of rimonabant on LPS or IL1β-induced expression of IL6 and TNFα, but not MCP1, suggest that rimonabant, without being an outright antiinflammatory compound, might modulate inflammation through the expression of some proinflammatory factors. In this respect, TNFα is particularly interesting, because local loss of TNF receptor-1 in the arterial wall has recently been shown to be sufficient to decrease atherosclerosis progression,29 and the antiatherosclerotic effect of systemic TNFα inhibition is now well established.30,31 In addition to its proinflammatory effects on cells in the vascular wall, TNFα might also induce a proatherogenic serum lipid profile,32 suggesting that the inhibition of TNFα production might play a role in the antiatherosclerotic effects of rimonabant. The fact that the effect of rimonabant on IL6 and TNFα expression persisted in macrophages from CB1R−/− mice clearly demonstrates that this effect is unrelated to CB1 receptor blockade, in contrast to the effect of rimonabant on LPS-induced TNFα induction in vivo, but in a similar way to the effect of rimonabant on indomethacin-induced intestinal ulcers.17 These data therefore strengthen the case for a modulation of inflammatory pathways by rimonabant, although the exact biochemical mechanism and in vivo relevance of this effect remain to be determined.

To determine whether these antiinflammatory effects of rimonabant could translate to in vivo effects at a similar dose, the effect of rimonabant on thioglycollate-induced macrophage recruitment was also assessed in normal and CB1 receptor–deficient mice. Interestingly, whereas rimonabant was able to significantly decrease macrophage recruitment, this effect was absent in CB1R−/− mice, showing that this effect is related to CB1 receptor blockade by rimonabant. The level of thioglycollate-induced macrophage recruitment is correlated with atherosclerosis resistance in different mouse strains,22 and the inhibitory effects of rimonabant therefore suggest that part of the effects of rimonabant might be related to inhibition of macrophage recruitment to atherosclerotic lesions.

Up to now, rimonabant is approved in Europe, but not in the United States, because of the FDA’s concerns about potential neuropsychiatric side effects. Recently, the data from the first clinical trial (STRADIVARIUS) designed to determine the antiatherosclerotic effect of rimonabant in patients by intravascular ultrasound (IVUS), became available.33 Although the effect of rimonabant on the primary end point (percent atheroma volume) did not reach statistical significance during the 18-month treatment period, rimonabant significantly decreased total atheroma volume, a secondary end point in this study. These first clinical data suggest that rimonabant indeed holds promise in the treatment of atherosclerosis, although longer treatment durations may be necessary to show this effect conclusively. The results from the ongoing CRESCENDO trial (5-year duration of treatment, NCT00263042), which directly measures cardiovascular outcomes, should clarify this point.34

In summary, in LDLR−/− mice fed a Western-type diet, rimonabant shows dose-dependent antiatherosclerotic effects which are absent in pair-fed animals. Whereas high doses increase the HDL/LDL cholesterol ratio and nearly abolish atherosclerotic lesion development, lower doses significantly decrease atherosclerosis without any effect on cholesterol levels but decrease circulating leptin levels. Rimonabant decreases the LPS-induced expression of IL6 and TNFα levels in mouse macrophages in vitro and thioglycollate-induced macrophage recruitment in vivo. It can therefore be concluded that rimonabant shows potent antiatherosclerotic effects in LDLR−/− mice on Western diet and that these effects are not solely attributable to its effects on cholesterol levels, but probably related to a decrease of inflammation by rimonabant.

Acknowledgments

We thank Noëlle Boussac-Marlière for statistical analysis.

Sources of Funding

This work was funded in part by a grant from sanofi-aventis.

Disclosures

F.D., V.V., A.M.M., P.S., and F.B. are employees of sanofi-aventis and hold significant stock of the company. R.J., P.D., N.H., A.G., and B.S. declare no competing interests beyond funding of their experiments through an unrestricted grant from sanofi-aventis.

Footnotes

  • Original received April 16, 2008; final version accepted September 26, 2008.

References

  1. 1.↵
    Gelfand EV, Cannon CP. Rimonabant: a selective blocker of the cannabinoid CB1 receptors for the management of obesity, smoking cessation and cardiometabolic risk factors. Expert Opin Investig Drugs. 2006; 15: 307–315.
    OpenUrlCrossRefPubMed
  2. 2.↵
    Kyrou I, Valsamakis G, Tsigos C. The endocannabinoid system as a target for the treatment of visceral obesity and metabolic syndrome. Ann N Y Acad Sci. 2006; 1083: 270–305.
    OpenUrlCrossRefPubMed
  3. 3.↵
    Pacher P, Batkai S, Kunos G. The endocannabinoid system as an emerging target of pharmacotherapy. Pharmacol Rev. 2006; 58: 389–462.
    OpenUrlAbstract/FREE Full Text
  4. 4.↵
    Rajesh M, Mukhopadhyay P, Hasko G, Huffman JW, Mackie K, Pacher P. CB2 cannabinoid receptor agonists attenuate TNF-alpha-induced human vascular smooth muscle cell proliferation and migration. Br J Pharmacol. 2008; 153: 347–357.
    OpenUrlCrossRefPubMed
  5. 5.↵
    Rajesh M, Mukhopadhyay P, Batkai S, Hasko G, Liaudet L, Huffman JW, Csiszar A, Ungvari Z, Mackie K, Chatterjee S, Pacher P. CB2-receptor stimulation attenuates TNF-alpha-induced human endothelial cell activation, transendothelial migration of monocytes, and monocyte-endothelial adhesion. Am J Physiol Heart Circ Physiol. 2007; 293: H2210–H2218.
    OpenUrlAbstract/FREE Full Text
  6. 6.↵
    Steffens S, Veillard NR, Arnaud C, Pelli G, Burger F, Staub C, Karsak M, Zimmer A, Frossard JL, Mach F. Low dose oral cannabinoid therapy reduces progression of atherosclerosis in mice. Nature. 2005; 434: 782–786.
    OpenUrlCrossRefPubMed
  7. 7.↵
    Mukhopadhyay P, Batkai S, Rajesh M, Czifra N, Harvey-White J, Hasko G, Zsengeller Z, Gerard NP, Liaudet L, Kunos G, Pacher P. Pharmacological inhibition of CB1 cannabinoid receptor protects against doxorubicin-induced cardiotoxicity. J Am Coll Cardiol. 2007; 50: 528–536.
    OpenUrlCrossRefPubMed
  8. 8.↵
    Batkai S, Mukhopadhyay P, Harvey-White J, Kechrid R, Pacher P, Kunos G. Endocannabinoids acting at CB1 receptors mediate the cardiac contractile dysfunction in vivo in cirrhotic rats. Am J Physiol Heart Circ Physiol. 2007; 293: H1689–H1695.
    OpenUrlAbstract/FREE Full Text
  9. 9.↵
    Van Gaal LF, Mertens IL, De Block CE. Mechanisms linking obesity with cardiovascular disease. Nature. 2006; 444: 875–880.
    OpenUrlCrossRefPubMed
  10. 10.↵
    Fantuzzi G, Mazzone T. Adipose tissue and atherosclerosis: exploring the connection. Arterioscler Thromb Vasc Biol. 2007; 27: 996–1003.
    OpenUrlAbstract/FREE Full Text
  11. 11.↵
    Engeli S, Bohnke J, Feldpausch M, Gorzelniak K, Janke J, Batkai S, Pacher P, Harvey-White J, Luft FC, Sharma AM, Jordan J. Activation of the peripheral endocannabinoid system in human obesity. Diabetes. 2005; 54: 2838–2843.
    OpenUrlAbstract/FREE Full Text
  12. 12.↵
    Bluher M, Engeli S, Kloting N, Berndt J, Fasshauer M, Batkai S, Pacher P, Schon MR, Jordan J, Stumvoll M. Dysregulation of the peripheral and adipose tissue endocannabinoid system in human abdominal obesity. Diabetes. 2006; 55: 3053–3060.
    OpenUrlAbstract/FREE Full Text
  13. 13.↵
    Van Gaal LF, Rissanen AM, Scheen AJ, Ziegler O, Rossner S. Effects of the cannabinoid-1 receptor blocker rimonabant on weight reduction and cardiovascular risk factors in overweight patients: 1-year experience from the RIO-Europe study. Lancet. 2005; 365: 1389–1397.
    OpenUrlCrossRefPubMed
  14. 14.↵
    Despres JP, Golay A, Sjostrom L. Effects of rimonabant on metabolic risk factors in overweight patients with dyslipidemia. N Engl J Med. 2005; 353: 2121–2134.
    OpenUrlCrossRefPubMed
  15. 15.↵
    Poirier B, Bidouard JP, Cadrouvele C, Marniquet X, Staels B, O'Connor SE, Janiak P, Herbert JM. The anti-obesity effect of rimonabant is associated with an improved serum lipid profile. Diabetes Obes Metab. 2005; 7: 65–72.
    OpenUrlCrossRefPubMed
  16. 16.↵
    Gary-Bobo M, Elachouri G, Gallas JF, Janiak P, Marini P, Ravinet-Trillou C, Chabbert M, Cruccioli N, Pfersdorff C, Roque C, Arnone M, Croci T, Soubrie P, Oury-Donat F, Maffrand JP, Scatton B, Lacheretz F, Le Fur G, Herbert JM, Bensaid M. Rimonabant reduces obesity-associated hepatic steatosis and features of metabolic syndrome in obese Zucker fa/fa rats. Hepatology. 2007; 46: 122–129.
    OpenUrlCrossRefPubMed
  17. 17.↵
    Croci T, Landi M, Galzin AM, Marini P. Role of cannabinoid CB1 receptors and tumor necrosis factor-alpha in the gut and systemic anti-inflammatory activity of SR 141716 (rimonabant) in rodents. Br J Pharmacol. 2003; 140: 115–122.
    OpenUrlCrossRefPubMed
  18. 18.↵
    Croci T, Zarini E. Effect of the cannabinoid CB1 receptor antagonist rimonabant on nociceptive responses and adjuvant-induced arthritis in obese and lean rats. Br J Pharmacol. 2007; 150: 559–566.
    OpenUrlCrossRefPubMed
  19. 19.↵
    Schreyer SA, Vick C, Lystig TC, Mystkowski P, LeBoeuf RC. LDL receptor but not apolipoprotein E deficiency increases diet-induced obesity and diabetes in mice. Am J Physiol Endocrinol Metab. 2002; 282: E207–E214.
    OpenUrlAbstract/FREE Full Text
  20. 19A.↵
    Bellosta S, Via D, Canavesi M, Pfister P, Fumagalli R, Bernini F. HMG-CoA reductase inhibitors reduce MMP-9 secretion by macrophages. Arterioscler Thromb Vasc Biol. 1998; 18: 1671–1678.
    OpenUrlAbstract/FREE Full Text
  21. 20.↵
    Carai MA, Colombo G, Maccioni P, Gessa GL. Efficacy of rimonabant and other cannabinoid CB1 receptor antagonists in reducing food intake and body weight: preclinical and clinical data. CNS Drug Rev. 2006; 12: 91–99.
    OpenUrlCrossRefPubMed
  22. 21.↵
    Ravinet-Trillou C, Arnone M, Delgorge C, Gonalons N, Keane P, Maffrand JP, Soubrie P. Anti-obesity effect of SR141716, a CB1 receptor antagonist, in diet-induced obese mice. Am J Physiol Regul Integr Comp Physiol. 2003; 284: R345–R353.
    OpenUrlAbstract/FREE Full Text
  23. 22.↵
    Stein O, Dabach Y, Ben-Naim M, Halperin G, Stein Y. Lower macrophage recruitment and atherosclerosis resistance in FVB mice. Atherosclerosis. 2006; 189: 336–341.
    OpenUrlCrossRefPubMed
  24. 23.↵
    Schreyer SA, Lystig TC, Vick CM, LeBoeuf RC. Mice deficient in apolipoprotein E but not LDL receptors are resistant to accelerated atherosclerosis associated with obesity. Atherosclerosis. 2003; 171: 49–55.
    OpenUrlCrossRefPubMed
  25. 24.↵
    Maffei M, Halaas J, Ravussin E, Pratley RE, Lee GH, Zhang Y, Fei H, Kim S, Lallone R, Ranganathan S. Leptin levels in human and rodent: measurement of plasma leptin and ob RNA in obese and weight-reduced subjects. Nat Med. 1995; 1: 1155–1161.
    OpenUrlCrossRefPubMed
  26. 25.↵
    Sierra-Johnson J, Romero-Corral A, Lopez-Jimenez F, Gami AS, Sert Kuniyoshi FH, Wolk R, Somers VK. Relation of increased leptin concentrations to history of myocardial infarction and stroke in the United States population. Am J Cardiol. 2007; 100: 234–239.
    OpenUrlCrossRefPubMed
  27. 26.↵
    Bodary PF, Gu S, Shen Y, Hasty AH, Buckler JM, Eitzman DT. Recombinant leptin promotes atherosclerosis and thrombosis in apolipoprotein E-deficient mice. Arterioscler Thromb Vasc Biol. 2005; 25: e119–e122.
    OpenUrlAbstract/FREE Full Text
  28. 27.↵
    Taleb S, Herbin O, Ait-Oufella H, Verreth W, Gourdy P, Barateau V, Merval R, Esposito B, Clement K, Holvoet P, Tedgui A, Mallat Z. Defective leptin/leptin receptor signaling improves regulatory T cell immune response and protects mice from atherosclerosis. Arterioscler Thromb Vasc Biol. 2007; 27: 2691–2698.
    OpenUrlAbstract/FREE Full Text
  29. 28.↵
    Grunfeld C, Zhao C, Fuller J, Pollack A, Moser A, Friedman J, Feingold KR. Endotoxin and cytokines induce expression of leptin, the ob gene product, in hamsters. J Clin Invest. 1996; 97: 2152–2157.
    OpenUrlCrossRefPubMed
  30. 29.↵
    Zhang L, Peppel K, Sivashanmugam P, Orman ES, Brian L, Exum ST, Freedman NJ. Expression of tumor necrosis factor receptor-1 in arterial wall cells promotes atherosclerosis. Arterioscler Thromb Vasc Biol. 2007; 27: 1087–1094.
    OpenUrlAbstract/FREE Full Text
  31. 30.↵
    Branen L, Hovgaard L, Nitulescu M, Bengtsson E, Nilsson J, Jovinge S. Inhibition of tumor necrosis factor-alpha reduces atherosclerosis in apolipoprotein E knockout mice. Arterioscler Thromb Vasc Biol. 2004; 24: 2137–2142.
    OpenUrlAbstract/FREE Full Text
  32. 31.↵
    Canault M, Peiretti F, Mueller C, Kopp F, Morange P, Rihs S, Portugal H, Juhan-Vague I, Nalbone G. Exclusive expression of transmembrane TNF-alpha in mice reduces the inflammatory response in early lipid lesions of aortic sinus. Atherosclerosis. 2004; 172: 211–218.
    OpenUrlCrossRefPubMed
  33. 32.↵
    Popa C, Netea MG, van Riel PL, van der Meer JW, Stalenhoef AF. The role of TNF-alpha in chronic inflammatory conditions, intermediary metabolism, and cardiovascular risk. J Lipid Res. 2007; 48: 751–762.
    OpenUrlAbstract/FREE Full Text
  34. 33.↵
    Nissen SE, Nicholls SJ, Wolski K, Rodes-Cabau J, Cannon CP, Deanfield JE, Despres JP, Kastelein JJ, Steinhubl SR, Kapadia S, Yasin M, Ruzyllo W, Gaudin C, Job B, Hu B, Bhatt DL, Lincoff AM, Tuzcu EM. Effect of rimonabant on progression of atherosclerosis in patients with abdominal obesity and coronary artery disease: the STRADIVARIUS randomized controlled trial. JAMA. 2008; 299: 1547–1560.
    OpenUrlCrossRefPubMed
  35. 34.↵
    Scheen AJ, Van Gaal LF. Rimonabant as an adjunct therapy in overweight/obese patients with type 2 diabetes. Eur Heart J. 2007; 28: 1401–1402.
    OpenUrlFREE Full Text
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    Rimonabant, a Selective Cannabinoid CB1 Receptor Antagonist, Inhibits Atherosclerosis in LDL Receptor–Deficient Mice
    Frédérique Dol-Gleizes, Réjane Paumelle, Virgile Visentin, Anne-Marie Marés, Perrine Desitter, Nathalie Hennuyer, Andries Gilde, Bart Staels, Paul Schaeffer and Françoise Bono
    Arteriosclerosis, Thrombosis, and Vascular Biology. 2009;29:12-18, originally published December 17, 2008
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    Rimonabant, a Selective Cannabinoid CB1 Receptor Antagonist, Inhibits Atherosclerosis in LDL Receptor–Deficient Mice
    Frédérique Dol-Gleizes, Réjane Paumelle, Virgile Visentin, Anne-Marie Marés, Perrine Desitter, Nathalie Hennuyer, Andries Gilde, Bart Staels, Paul Schaeffer and Françoise Bono
    Arteriosclerosis, Thrombosis, and Vascular Biology. 2009;29:12-18, originally published December 17, 2008
    https://doi.org/10.1161/ATVBAHA.108.168757
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