Molecular and Biochemical Mechanism of Cannabidiol in the Management of the Inflammatory and Oxidative Processes Associated with Endometriosis

Abstract

Endometriosis is usually associated with inflammation and chronic pelvic pain. This paper focuses the attention on the anti-inflammatory, anti-oxidant and analgesic effects of cannabidiol (CBD) and on its potential role in endometriosis. We employed an in vivo model of endometriosis and administered CBD daily by gavage. CBD administration strongly reduced lesions diameter, volume and area. In particular, it was able to modify lesion morphology, reducing epithelial glands and stroma. CBD showed anti-oxidant effects reducing lipid peroxidation, the expression of Nox-1 and Nox-4 enzymes. CBD restored the oxidative equilibrium of the endogenous cellular defense as showed by the SOD activity and the GSH levels in the lesions. CBD also showed important antifibrotic effects as showed by the Masson trichrome staining and by downregulated expression of MMP-9, iNOS and TGF-β. CBD was able to reduce inflammation both in the harvested lesions, as showed by the increased Ikb-α and reduced COX2 cytosolic expressions and reduced NFkB nuclear localization, and in the peritoneal fluids as showed by the decreased TNF-α, PGE2 and IL-1α levels. CBD has important analgesic effects as showed by the reduced mast cells recruitment in the spinal cord and the reduced release of neuro-sensitizing and pro-inflammatory mediators. In conclusion, the collected data showed that CBD has an effective and coordinated effects in endometriosis suppression.

Keywords: cannabidiol; endometriosis; inflammation.

Figure 1

CBD administration reduced lesion size endometriosis-induced: hfUS analysis: vehicle (A), CBD (B), macroscopic analysis: vehicle (C), CBD (D), lesion number (E), lesion diameter (F), lesion volume (G), lesion area (H), histological analysis: vehicle (I), CBD (J), histopathological score (K): G: glands; S: stroma; L: cyst lumen; A: adipose tissue. For each analysis, n = 5 animals were employed. For the histological analysis, n = 5 different fields from n = 5 different animals were evaluated. ** p < 0.01 vs. endometriosis.

Figure 2

CBD administration reduced pro-oxidative alteration in endometriotic lesions: malondialdehyde (MDA) levels (A), GSH levels (B), SOD activity (C), Western blot analysis of Nox-1 (D) and Nox-4 (E) expression. For each analysis, n = 5 animals were employed. ** p < 0.01 vs. endometriosis, *** p < 0.001 vs. endometriosis.

Figure 3

CBD administration reduced fibrosis endometriosis-induced: Masson trichrome staining: vehicle (A), CBD (B), aniline blue stain area (C); Western blot analysis of MMP-9 (D), iNOS (E) and TGF-β expression (F). For each analysis, n = 5 animals were employed. For the Masson trichrome staining, n = 5 different fields from n = 5 different animals were evaluated. * p < 0.05 vs. endometriosis, ** p < 0.01 vs. endometriosis, *** p < 0.001 vs. endometriosis.

Figure 4

CBD administration reduced the pro-inflammatory state endometriosis-induced: Western blot analysis of Ikb-α (A), COX2 (B), NF-kB (C), IL-1β (D), TNF-α (E) and PGE₂ (F) levels in the peritoneal fluids. For each analysis, n = 5 animals were employed. ** p < 0.01 vs. endometriosis, *** p < 0.001 vs. endometriosis, ### p < 0.001 vs. sham.

Figure 5

CBD administration reduced pain-mediators and neuroinflammatory state endometriosis-induced: Toluidine blue staining: sham (A), vehicle (B), CBD (C), mast cell number (D), Western blot analysis of c-FOS (E), and NGF (F) expression; Immunohistochemical analysis of GFAP: sham (G), vehicle (H), CBD (I), graphical quantification of GFAP expression (J), immunohistochemical analysis of Iba-1: sham (K), vehicle (L), CBD (M), graphical quantification of Iba-1 expression (N). For each analysis, n = 5 animals were employed. For the toluidine blue staining and the immunohistochemical analysis, n = 5 different fields from n = 5 different animals were evaluated. * p < 0.05 vs. endometriosis, ## p < 0.01 vs. sham, *** p < 0.001 vs. endometriosis, ### p < 0.001 vs. sham. Scale bar 100 μm.

Figure 6

CBD administration reduced pain-behavior endometriosis-induced: open field test: number of crossings (A), number of entries in central square (B), and time spent in central square (C), elevated plus maze test: number of entries in closed and open arms (D), % of open entries (E), % of time in open arms (F), acetic-acid-induced abdominal contractions (G), hot plate test (H). For each analysis, n = 5 animals were employed. ** p < 0.01 vs. endometriosis, *** p < 0.001 vs. sham, ### p < 0.001 vs. endometriosis.