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. 2022 Jun 7;12(1):9400.
doi: 10.1038/s41598-022-13488-4.

Gene expression of the endocannabinoid system in endometrium through menstrual cycle

Keisuke Tanaka  1   2 , Akwasi A Amoako  3   4 , Sally Mortlock  5 , Peter A W Rogers  6   7 , Sarah J Holdsworth-Carson  6   7   8 , Jacqueline F Donoghue  6   7 , Wan Tinn Teh  6 , Grant W Montgomery  5 , Brett McKinnon  5   9
Affiliations

Gene expression of the endocannabinoid system in endometrium through menstrual cycle

Keisuke Tanaka et al. Sci Rep. .

Abstract

Endocannabinoids mediate cellular functions and their activity is controlled by a complex system of enzymes, membrane receptors and transport molecules. Endocannabinoids are present in endometrium, a cyclical regenerative tissue requiring tightly regulated cellular mechanisms for maturation. The objective of this study was to investigate the gene expression of key elements involved in the endocannabinoid system across the menstrual cycle. RNA was isolated from endometrial tissue and genome-wide gene expression datasets were generated using RNA-sequencing. An a priori set of 70 genes associated with endocannabinoid system were selected from published literature. Gene expression across the menstrual cycle was analyzed using a moderated t test, corrected for multiple testing with Bonferroni's method. A total of 40 of the 70 genes were present in > 90% of the samples, and significant differential gene expression identified for 29 genes. We identified 4 distinct regulation patterns for synthesizing enzymes, as well as a distinct regulation pattern for degradations and transporting enzymes. This study charts the expression of endometrial endocannabinoid system genes across the menstrual cycle. Altered expression of genes that control endocannabinoid may allow fine control over endocannabinoid concentrations and their influence on cellular function, maturation and differentiation as the endometrium matures through the menstrual cycle.

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Conflict of interest statement

The authors declare no competing interests.

Figures

Figure 1
Figure 1
Gene expression of ECS genes in endometrium across the menstrual cycle. (A) Using 206 endometrial samples we found that of the 70 genes identified through the literature to function as part of ECS, 40 were expressed in at least 90% of the samples. The level of expression of each gene varied significantly between individuals and mean expression varied between genes. (B) A heat map analysis identifying the relative expression (cpm) of each gene across the menstrual cycle. The stage variation identified for some of these genes could be explained in part by differences that occurred in expression across the menstrual cycle. M = Menstrual, EP = Early proliferative, MP = Mid proliferative, LP = Late proliferative, ES = Early secretory, MS = Mid secretory, LS = Late secretory.
Figure 2
Figure 2
The expression profile of the four distinct groups of endocannabinoid synthesizing enzymes. (A) Group 1 showed high gene expression (≈ 50–150 CPM) that increased across the proliferative stage and peaked during the early secretory to mid secretory stage. (B) The second group was characterised by median gene expression (≈ 20–80 CPM) that generally increased from the menstrual to early proliferative stage and then showed a gradual decline. (C) This group was characterised by a low- medium expression (≈ 10–80 CPM) that decreased in the early proliferative stage and subsequently increased in the secretory stage. (D) The fourth group was characterised by low gene expression (≈ 1–30 CPM) that showed only small changes across the stages. M = Menstrual, EP = Early proliferative, MP = Mid proliferative, LP = Late proliferative, ES = Early secretory, MS = Mid secretory, LS = Late secretory.
Figure 3
Figure 3
The expression profile of genes encoding ECS transport and degrading proteins. Transcription expression that showed a significant difference at any stage in the menstrual cycle were plotted against cycle stage. (A) For the transporting proteins moderate expression was observed for FABP5 and FABP3 that was significantly increased across the menstrual cycle. (B) ECS degrading enzymes had a moderate to low expression that were most commonly altered between the menstrual and early proliferative stage, or during the period from the late proliferative to mid secretory stage. Two genes ABHD12 and NAAA showed a consistent increase over the proliferative stage, after which there was decrease in expression. M = Menstrual, EP = Early proliferative, MP = Mid proliferative, LP = Late proliferative, ES = Early secretory, MS = Mid secretory, LS = Late secretory.
Figure 4
Figure 4
Logistic regression analysis of endocannabinoid receptor. Four of the five endocannabinoid receptor did not show consistent expression above > 90% in all samples. A logistic regression analysis examining the percentage of samples displaying a positive expression showed significant variation across the menstrual cycle for CNR1, CNR2 and GPR55. Only a small number of samples expressed TRPA1 and this was limited to the period between the mid proliferative to mid secretory stage. M = Menstrual, EP = Early proliferative, MP = Mid proliferative, LP = Late proliferative, ES = Early secretory, MS = Mid secretory, LS = Late secretory.
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