The Effect of Oil-Based Cannabis Extracts on Metabolic Parameters and Microbiota Composition of Mice Fed a Standard and a High-Fat Diet

Abstract

The prevalence of obesity and obesity-related pathologies is lower in frequent cannabis users compared to non-users. It is well established that the endocannabinoid system has an important role in the development of obesity. We recently demonstrated that prolonged oral consumption of purified Δ-9 Tetrahydrocannabinol (THC), but not of cannabidiol (CBD), ameliorates diet-induced obesity and improves obesity-related metabolic complications in a high-fat diet mouse model. However, the effect of commercially available medical cannabis oils that contain numerous additional active molecules has not been examined. We tested herein the effects of THC- and CBD-enriched medical cannabis oils on obesity parameters and the gut microbiota composition of C57BL/6 male mice fed with either a high-fat or standard diet. We also assessed the levels of prominent endocannabinoids and endocannabinoid-like lipid mediators in the liver. THC-enriched extract prevented weight gain by a high-fat diet and attenuated diet-induced liver steatosis concomitantly with reduced levels of the lipid mediators palmitoyl ethanolamide (PEA) and docosahexaenoyl ethanolamide (DHEA) in the liver. In contrast, CBD-enriched extract had no effect on weight gain, but, on the contrary, it even exacerbated liver steatosis. An analysis of the gut microbiota revealed that mainly time but not treatment exerted a strong effect on gut microbiota alterations. From our data, we conclude that THC-enriched cannabis oil where THC is the main constituent exerts the optimal anti-obesity effects.

Keywords: cannabis; endocannabinoid system; microbiota; obesity.

Figure 1

Effect of Avidekel and Erez extracts on weight gain, caloric intake, fat pad weights, and adipocyte size. Body weight change (A), average caloric intake (B), fat pad weights (C), and quantitative analysis of adipocyte size in epididymal tissue (D) were measured and calculated at the end of the treatment regime (n = 5–7). Data are shown as mean ± SEM. * p < 0.05, ** p < 0.002, *** p < 0.001, **** p < 0.0001. Statistical significance was determined by two-way ANOVA followed by Tukey’s post hoc test.

Figure 2

Inflammation markers in adipose tissue. mRNA expression of genes involved in inflammation examined in epididymal fat: (A). Interleukin-1 beta (IL-1β). (B). Macrophage receptor F4/80. (C). Monocyte differentiation antigen CD14. (D). Interferon-γ (INF-γ). PPIA was used as a housekeeping gene. The gene expression levels were calculated as ΔΔCT and normalized to the vehicle of each diet (n = 4–6). Data are shown as the mean of fold-change ± SEM. * p < 0.05, ** p < 0.002, *** p < 0.001, **** p < 0.0001. Statistical significance was determined by two-way ANOVA followed by Tukey’s post hoc test.

Figure 3

Effect of Avidekel and Erez extract on obesity-induced steatosis. (A). Representative liver tissue sections stained with H&E (magnification ×100). (B). Morphological analysis of liver droplet density (n = 5). (C). Liver weights. (D). Levels of hepatic triglycerides (n = 3). (E). Serum cholesterol (n = 3). (F). Severity of steatosis was blindly scored as described by Liang et al. [27] (n = 3). Data are shown as mean ± SEM. * p < 0.05, ** p < 0.002, **** p < 0.0001. Statistical significance was determined by using two-way ANOVA followed by Tukey’s post hoc test.

Figure 4

Levels of phytocannabinoid metabolites in liver tissue. Levels of THC, CBD, and their metabolites were assessed at the end of the treatment regime as described in materials and methods. Data are shown as mean ± SEM (n = 4).

Figure 5

Alpha diversity and F/B ratios of gut microbiota in mice. Fecal samples were taken at various time points (T1—a month after starting HFD; T2—a day before the start of treatment; T3—one day after the start of the treatment; and T4—day of sacrifice), and gut microbiota were analyzed. The gut microbial richness was estimated by observed species; the diversity was evaluated by using the Shannon index; and Evenness was measured by using the Shannon Evenness index. Alpha diversity metrics: richness, evenness, diversity, and Firmicutes/Bacteroidetes ratio at T1 (A–D), HFD-fed mice (E–H), and STD-fed mice (I–L). The line shows the average, and the whiskers show the minimum and maximum (n = 4–7). * p < 0.05, ** p < 0.002, *** p < 0.001, **** p < 0.0001. Statistical significance was calculated by using non-parametric Mann–Whitney (A–D) and Kruskal–Wallis (E–L) tests.

Figure 6

The relative frequency (%) of microbial taxa at the order level in mice gut microbiota samples. Each vertical bar represents the average fecal sample of the respective treatment group at four different time points (T1—a month after starting HFD; T2—a day before the start of treatment; T3—one day after the start of the treatment; and T4—day of sacrifice). Data are shown as percentages (n = 4–7).