The cannabinoid receptor CB1 affects the proliferation and apoptosis of adenomyotic human uterine smooth muscle cells of the junctional zone: a mechanism study

Sha Wang,Hua Duan,Xiao Li,Bohan Li,Fuqing Sun,Xue Shen,Zhengchen Guo

doi:10.1186/s12958-020-00690-0

Abstract

BackgroundThe denomyotic junctional zone (JZ) plays an important role in the pathogenesis of adenomyosis. Proliferating cell nuclear antigen (PCNA) is an important nuclear marker of cell proliferation. This study aimed to evaluate the effects of the cannabinoid receptor CB1 on proliferation and apoptosis in the JZ in women with and without adenomyosis.MethodsJZ smooth muscle cells (JZSMCs) of the adenomyosis and control groups were collected and cultivated. Immunohistochemistry and immunoblotting were used for protein localization and expression detection of CB1 and PCNA. Additionally, qRT-PCR was used to quantitatively analyse the mRNA expression of the two. AM251 and ACEA were used to regulate the function of CB1 receptors, and CCK-8 assay and flow cytometry assay were used to verify the proliferation and apoptosis of JZSMCs after regulation.ResultsWe demonstrated that in normal JZSMCs CB1 and PCNA messenger RNA (mRNA) and protein expression was significantly higher in the proliferative phase of the menstrual cycle than in the secretory phase. CB1 and PCNA expression in JZSMCs from women with ADS was significantly higher than that in control women and did not significantly differ across the menstrual cycle. CB1 receptor antagonist AM251 inhibited the proliferation of adenomyotic JZSMCs in a dose-dependent manner. The CB1 receptor agonist ACEA significantly promoted the proliferation of adenomyotic JZSMCs. The apoptosis rate of adenomyotic JZSMCs treated with AM251 was significantly higher than that of JZSMCs from the untreated control group. The apoptosis rate was significantly decreased in the ACEA group compared with that in the untreated control group. Furthermore, AM251 suppressed the phosphorylation of AKT and Erk1/2 in adenomyotic JZSMCs. The CB1 agonist ACEA significantly promoted the phosphorylation of AKT and Erk1/2.ConclusionsOur results indicated that the levels of CB1 and PCNA were increased in patients with adenomyosis and that cyclic changes were lost. CB1 may affect uterine JZ proliferation and apoptosis in adenomyosis by enhancing AKT and MAPK/Erk signalling.

Highlights

Adenomyosis (ADS) is a common oestrogen-dependent uterine disorder distinguished by non-malignant invasion of the bioactive endometrium into the myometrial wall, and it can result in abnormal uterine bleeding, dysmenorrhoea, and subfertility [1, 2]
We confirmed that expression of the cannabinoid receptors CB1 and CB2 was significantly higher in the ADS myometrium than in the normal myometrium, and CB1 expression in the junctional zone (JZ) was positively correlated with the severity of dysmenorrhoea in patients with ADS [13]
In the ADS and control groups, Proliferating cell nuclear antigen (PCNA) and CB1 expression was observed in endometrial glandular epithelial cells, stromal cells, vascular endothelial cells, JZ smooth muscle cells (JZSMCs) and ectopic endometrial cells (Fig. 1a)

Summary

Introduction

Adenomyosis (ADS) is a common oestrogen-dependent uterine disorder distinguished by non-malignant invasion of the bioactive endometrium into the myometrial wall, and it can result in abnormal uterine bleeding, dysmenorrhoea, and subfertility [1, 2]. The ECS has been reported to be involved in processes relevant to endometriosis, including cell migration, cell proliferation, apoptosis, and inflammation, and to interact with sex steroid hormones [9, 10]. A large number of studies have confirmed that CB1 can affect the proliferation, migration and apoptosis of many types of cells by regulating the AKT and MAPK/Erk pathways (via phosphorylation activation) [11, 12]. This study aimed to evaluate the effects of the cannabinoid receptor CB1 on proliferation and apoptosis in the JZ in women with and without adenomyosis

Objectives

Methods

Results

Conclusion