Expression and localization of cannabinoid receptors in human immature oocytes and unfertilized metaphase-II oocytes

Abstract

Endocannabinoid anandamide and cannabinoid receptors have been described in some organs of the female reproductive system, but little is known about the expression of these receptors in human oocytes. The aim of the study was to describe the expression of cannabinoid receptors in human oocytes and to investigate their differential distribution at various stages of meiotic resumption in human oocytes. A total of 750 human oocytes from 214 patients were analysed by Western blot, immunocytochemistry and PCR. For this study, oocytes that were not suitable for intracytoplasmic sperm injection (ICSI) (germinal-vesicle and metaphase-I stages), as well as metaphase-II oocytes that had not developed into an embryo after ICSI were used. Western blot analysis revealed the presence of CB1 and CB2 receptor proteins in human oocytes. CB1 and CB2 receptor immunostaining patterns changed during the various stages of meiotic resumption. Localization of CB1 receptor was peripheral at germinal-vesicle stage, homogeneous over the entire oocyte at metaphase I and peripheral at mature metaphase II. CB2 receptor localization was peripheral at germinal-vesicle and metaphase-I stages but homogeneous over the entire cell at metaphase II. This finding suggests a possible role for endocannabinoids, acting via receptors, in the maturation of female gametes and fertilization. The number of couples with sterility problems attending fertility programmes is rising but treatment is not always successful. Important problems associated with failure to conceive remain unresolved because many physiological aspects of human reproduction are still unknown. Endocannabinoids are endogenous chemical compounds that mimic the action of the main psychoactive component of marijuana, delta-9-tetrahydrocannabinol. An endogenous cannabinoid named anandamide has been found in human follicular fluid. Thus, in order to develop knowledge in this field, in the present study we have described the presence of the cannabinoid receptors CB1 and CB2 (the proteins required to mediate the action of the cannabinoids) in the early stages of meiotic resumption of oocytes (the stages before ovulation) and we could postulate that the endocannabinoids could act in the regulation of maturation of oocytes. Our study, together with other studies, indicates that the endocannabinoid system may play a role in human reproduction.

Dynamic of expression and localization of cannabinoid-degrading enzymes FAAH and MGLL in relation to CB1 during meiotic maturation of human oocytes

“…In human ovaries, however, only FAAH has been found localized to the theca cells of secondary and tertiary follicles, whereas CB1 immunostaining appears in oocytes from primordial to secondary follicles but not in tertiary follicles and, in contrast, intense CB2 staining has been observed in oocytes during all stages of follicle maturation (El-Talatini et al 2009). Finally, in a study focused on human oocytes separated from follicles, the presence of both CB1 and CB2 has been corroborated during the resumption of oocyte meiosis from the GV to MII stages (Peralta et al 2011). Whether the differences in the localization of CB1, CB2 and FAAH during the follicle and/or oocyte maturation observed in the aforementioned three studies reflect differences between species or are also attributable to differences in methodology (Bagavandoss and Grimshaw 2010;Peralta et al 2011) is unclear.…”

“…Finally, in a study focused on human oocytes separated from follicles, the presence of both CB1 and CB2 has been corroborated during the resumption of oocyte meiosis from the GV to MII stages (Peralta et al 2011). Whether the differences in the localization of CB1, CB2 and FAAH during the follicle and/or oocyte maturation observed in the aforementioned three studies reflect differences between species or are also attributable to differences in methodology (Bagavandoss and Grimshaw 2010;Peralta et al 2011) is unclear. In any case, taken together, all the evidence to date leads us to believe that the previous non-detection of FAAH in human oocytes (El-Talatini et al 2009) is attributable to a relatively low level of expression of FAAH rather than its absence.…”

“…Even so, the part played by this system in the ovary and in oocyte events has rarely been analyzed (El-Talatini et al 2009;Bagavandoss and Grimshaw 2010;Peralta et al 2011). The aim of the present study was to gather further data on the presence and localization of cannabinoid-degrading enzymes in relation to the CB1 cannabinoid receptor during the resumption of meiosis in human oocytes in order to form a basis for explaining a possible role of the cannabinoid system in this process.…”

“…The localization of the cannabinoid receptors and degrading enzymes in rat (Bagavandoss and Grimshaw 2010) and human ovary has led to the hypothesis that AEA plays a role in folliculogenesis, preovulatory follicle maturation oocyte maturity, and ovulation (El-Talatini et al 2009). Finally, the hypothesis with respect to the role of the cannabinoid system in oocyte maturation has been reinforced by findings concerning the localization of cannabinoid receptors during the nuclear maturation of human oocytes, in which each receptor is re-localized (Peralta et al 2011).…”

“…In view of the presence of AEA in human follicular fluid and the re-localization of cannabinoid receptors observed in the oocyte during its maturation, cannabinoid signaling can be postulated to be involved in the regulation of oocyte meiosis (Peralta et al 2011). Therefore, to shed further light on this issue, our aim is to characterize, in detail and to compare the expression and distribution of the two AEA-degrading enzymes, FAAH and MGLL and further to explore their relationship with the CB1 receptor in human oocytes during the meiotic resumption of oocytes obtained during in vitro fertilization (IVF)/intracytoplasmic sperm injection (ICSI) cycles.…”