Caralluma fimbriata Extract Improves Vascular Dysfunction in Obese Mice Fed a High-Fat Diet

Abstract

Background: Obesity is a risk factor for developing cardiovascular diseases (CVDs) by impairing normal vascular function. Natural products are gaining momentum in the clinical setting due to their high efficacy and low toxicity. Caralluma fimbriata extract (CFE) has been shown to control appetite and promote weight loss; however, its effect on vascular function remains poorly understood. This study aimed to determine the effect that CFE had on weight loss and vascular function in mice fed a high-fat diet (HFD) to induce obesity, comparing this effect to that of lorcaserin (LOR) (an anti-obesity pharmaceutical) treatment.

Methods: C57BL/6J male mice (n = 80) were fed a 16-week HFD to induce obesity prior to being treated with CFE and LOR as standalone treatments or in conjunction. Body composition data, such as weight gain and fat mass content were measured, isometric tension analyses were performed on isolated abdominal aortic rings to determine relaxation responses to acetylcholine, and immunohistochemistry studies were utilized to determine the expression profiles on endothelial nitric oxide synthase (eNOS) and cell stress markers (nitrotyrosine (NT) and 78 kDa glucose-regulated protein (GRP78)) in the endothelial, medial and adventitial layers of aortic rings.

Results: The results demonstrated that CFE and CFE + LOR treatments significantly reduced weight gain (17%; 24%) and fat mass deposition (14%; 16%). A HFD markedly reduced acetylcholine-mediated relaxation (p < 0.05, p < 0.0001) and eNOS expression (p < 0.0001, p < 0.01) and significantly increased NT (p < 0.05, p < 0.0001) and GRP78 (p < 0.05, p < 0.01, p < 0.001). Obese mice treated with CFE exhibited significantly improved ACh-induced relaxation responses, increased eNOS (p < 0.05, p < 0.01) and reduced NT (p < 0.01) and GRP78 (p < 0.05, p < 0.01) expression.

Conclusions: Thus, CFE alone or in combination with LOR could serve as an alternative strategy for preventing obesity-related cardiovascular diseases.

Keywords: Caralluma fimbriata extract; bio-active compounds; high-fat diet; lorcaserin; natural product; obesity; vascular dysfunction.

Figure 1

Body weights during the 16-week study. Mice body weights are shown each week during HFD induction from week 0 to week 8. The body weights from week 9 to week 16 represent the treatment period with CFE and/or LOR. All the data are derived from n = 16/group and values plotted are presented as the mean ± SEM. The statistical significance * p ≤ 0.05 represents the comparison between the control and the HFD groups; ‘#’ represents HFD with CFE + LOR, and ‘$’ represents LOR with CFE + LOR. Abbreviations: CFE, Caralluma fimbriata extract; HFD, high-fat diet; LOR, lorcaserin; SEM, standard error of the mean.

Figure 2

Change in weight gain during the 16-week study. The weight gain of all the groups during the HFD induction and treatment periods are represented individually. Each group had an n = 16, and the values plotted are the mean ± SEM. Significance is set at p ≤ 0.05. Abbreviations: CFE, Caralluma fimbriata extract; HFD, high-fat diet; LOR, lorcaserin; SEM, standard error of the mean.

Figure 3

Change in fat content during the treatment period (8 weeks): EchoMRI-based fat mass measurement difference between weeks 8 and 16. Each group consisted of n = 16 mice, and the values plotted are represented as the mean ± SEM. Significance is set at p ≤ 0.05. Abbreviations: CFE, Caralluma fimbriata extract; HFD, high-fat diet; LOR, lorcaserin; SEM, standard error of the mean.

Figure 4

Fat and lean mass percentage with respect to bodyweight. Lean mass percentage (A) and fat mass percentage (B) during the 8th and 16th weeks with respect to bodyweights. Each group included n = 16 mice and the values plotted are the mean ± SEM. Significance was set at p ≤ 0.05. Abbreviations: CFE, Caralluma fimbriata extract; HFD, high-fat diet; LOR, lorcaserin; SEM, standard error of the mean; Wk, weeks.

Figure 5

Relaxation responses of abdominal aortic rings to ACh dose–response. (A) The aortic rings from the mice fed a 16-week HFD had significantly reduced ability to relax to ACh when compared to the mice fed a control diet (mean ± SEM, p < 0.05, p < 0.0001). (B) Treatment with LOR was unable to improve relaxation to ACh in mice fed a HFD (mean ± SEM). (C) CFE was able to significantly enhance relaxation responses to ACh in mice fed a HFD (mean ± SEM, p < 0.05, p < 0.01, p < 0.001). (D) The combination treatment of CFE + LOR was unable to significantly increase ACh-mediated relaxation (mean ± SEM). Abbreviations: ACh, acetylcholine; CFE, Caralluma fimbriata; HFD, high-fat diet; LOR, lorcaserin; SEM, standard error of the mean. Key: * = p < 0.05, ** = p < 0.01 and **** = p < 0.0001.

Figure 6

eNOS expression in different layers of mouse abdominal aorta. Immunohistochemical localization of eNOS in the endothelium, media and adventitia of abdominal aortae from (A) mice fed a control diet (n = 3); (B) mice fed a 16-week HFD to induce obesity (n = 4); (C) mice fed a HFD and treated with LOR (n = 4); (D) mice fed a HFD and treated with CFE (n = 4); and (E) mice fed a HFD and treated with CFE and LOR (n = 3). (F) Positive control slides were stained with the anti-CD31 antibody, while the (G) negative control slides were incubated without the primary antibody. (H) Obese mice fed a 16-week HFD had markedly reduced endothelial eNOS expression (mean ± SEM, p < 0.0001), which was significantly increased by the CFE treatment either alone (mean ± SEM, p < 0.01) or in combination with LOR (mean ± SEM, p < 0.05). (I) Media expression of eNOS in HFD-fed mice was significantly decreased (mean ± SEM, p < 0.01) and was enhanced by CFE treatment (mean ± SEM, p < 0.05). (J) Although not statistically significant, adventitial expression of eNOS was decreased by the HFD and was increased across all treatment groups. Abbreviations: CFE, Caralluma fimbriata; eNOS, endothelial nitric oxide synthase; HFD, high-fat diet; LOR, lorcaserin; PI, proportional intensity; SEM, standard error of the mean. Key: * = p < 0.05, ** = p < 0.01 and **** = p < 0.0001.

Figure 7

NT expression in different layers of mouse abdominal aorta. Immunohistochemical localization of NT in the endothelium, media and adventitia of abdominal aortae from (A) mice fed a control diet (n = 3); (B) mice fed a 16-week HFD to induce obesity (n = 4); (C) HFD-fed mice treated with LOR (n = 3); (D) HFD-fed mice treated with CFE (n = 3); and (E) HFD-fed mice treated with CFE and LOR (n = 3). Positive control slides were stained with ant-CD31 antibody (F) and negative control slides were not incubated with the primary antibody (G). (H) Obese mice fed a 16-week HFD had markedly increased endothelial NT expression (mean ± SEM, p < 0.0001), which was significantly reduced by CFE treatment, either alone (mean ± SEM, p < 0.05) or in combination with LOR (mean ± SEM, p < 0.05). (I) Media expression of NT in HFD mice was significantly elevated (mean ± SEM, p < 0.05). Although there were no significant differences, the CFE and LOR + CFE treatments appeared to reduce NT expression. (J) There were no significant differences in adventitial NT expression between groups; however, the control and the HFD-fed mice treated with CFE demonstrated lower NT levels. Abbreviations: CFE, Caralluma fimbriata; HFD, high-fat diet; LOR, lorcaserin; NT, nitrotyrosine; PI, proportional intensity; SEM, standard error of the mean. Key: * = p < 0.05 and **** = p < 0.0001.

Figure 8

GRP78 expression in different layers of mouse abdominal aorta. Immunohistochemical localization of GRP78 in the endothelium, media and adventitia of abdominal aortae from (A) mice fed a control diet (n = 3); (B) mice fed a 16-week HFD to induce obesity (n = 3); (C) HFD-fed mice treated with LOR (n = 3); (D) HFD-fed mice treated with CFE (n = 3) and (E) HFD-fed mice treated with CFE and LOR (n = 3). The positive control slides were stained with anti-CD31 antibody (F) and the negative control slides were not incubated with the primary antibody (G). (H) Obese mice fed a 16-week HFD had markedly increased endothelial GRP78 expression (mean ± SEM, p < 0.001), which was significantly reduced by LOR (mean ± SEM, p < 0.05) treatment, CFE (mean ± SEM, p < 0.01) treatment or their combination (mean ± SEM, p < 0.01). (I) Media expression of GRP78 in HFD mice was significantly elevated (mean ± SEM, p < 0.05). Although there were no statistically significant differences, the CFE, LOR and LOR + CFE treatments reduced its expression. (J) GRP78 expression in the adventitia was significantly augmented in the HFD mice (mean ± SEM, p < 0.01), and both CFE (mean ± SEM, p < 0.05) and LOR + CFE (mean ± SEM, p < 0.05) were able to markedly reduce this upregulation. Abbreviations: CFE, Caralluma fimbriata; HFD, high-fat diet; GRP78, 78 kDa glucose-regulated protein; LOR, lorcaserin; PI, proportional intensity; SEM, standard error of the mean. Key: * = p < 0.05, ** = p < 0.01 and *** = p < 0.001.