The Role of Endocannabinoids in Early Pregnancy

The endocannabinoid N-arachidonoylethanolamine (anandamide; AEA) adversely affects early pregnancy initiation and maintenance in both animals and humans. Anandamide acting through the cannabinoid receptors CB1 and CB2 affects many aspects of reproduction and is formed on demand by the hydrolysis of N-arachidonoyl-phosphatidylethanolamine (NAPE), by a specific phospholipase D (PLD) NAPE-PLD. Furthermore, AEA is catabolised by Fatty Acid Amide Hydrolase (FAAH). The aim of this thesis was to investigate how variations in the various components of the endocannabinoid system might lead to different early pregnancy outcomes. Using a newly developed UPLC-MS/MS method, plasma AEA levels were measured in women throughout pregnancy, compared to previous data and shown to be comparable. Moreover, women presenting with a viable pregnancy (on USS) with a threatened miscarriage (bleeding), who went on to miscarry and women presenting with a confirmed miscarriage (on USS) when compared with women with a viable pregnancy at the time of USS in whom there were no symptoms of miscarriage or those that went to term, all had increased plasma AEA levels. Median plasma AEA levels in the miscarriage group were 3-fold (p<0.001) higher than the levels in the live birth group. Using an AEA level of 2.0nM as cut-off for predicting subsequent miscarriage gave a sensitivity of 100% and a specificity of 94%, (negative predictive value 100%; positive predictive value 82%). Although other markers of pregnancy success (progesterone, hCG and PAPP-A) all gave the expected concentrations in on-going, failed and failing pregnancies, there was no statistically significant correlation between these factors and plasma AEA levels. Immunohistochemical evaluation of CB1, CB2, FAAH and NAPE-PLD expression in first trimester pregnancy tissues (trophoblast and decidua) showed that FAAH staining in the miscarriage/non-viable group was significantly lower than that in the surgical termination of pregnancy group. In women undergoing medical termination of pregnancy, using an anti-progesterone (mifepristone), FAAH staining was actually increased compared with that from the surgical termination of pregnancy group; whilst CB2 had increased staining and CB1 had reduced staining in the miscarriage and the medical termination groups when compared with the surgical termination of pregnancy group. There was no significant difference in NAPE-PLD staining between all three groups. These data provide new evidence that the endocannabinoid system plays a significant role in early pregnancy failure/miscarriage and that plasma AEA may be used as a future predictive marker for miscarriage. The lack of correlation between AEA and progesterone and the finding of elevated FAAH in the medical miscarriage (anti-progesterone) group, but low FAAH in spontaneous miscarriage suggest that factors other than progesterone are responsible for increased AEA levels in cases of miscarriage.