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Local uterine Ang-(1-7) infusion augments the expression of cannabinoid receptors and differentially alters endocannabinoid metabolizing enzymes in the decidualized uterus of pseudopregnant rats.

Brosnihan KB, Pulgar VM, Gallagher PE, Neves LA, Yamaleyeva LM - Reprod. Biol. Endocrinol. (2015)

Bottom Line: CB1R mRNA was reduced by decidualization (2.7-fold, p < 0.001), but increased by Ang-(1-7) (1.9-fold, p < 0.05).CB2R mRNA was increased by decidualization (4-fold, p < 0.05) and by Ang-(1-7) (2.4-fold, p < 0.001).The enzyme metabolizing AEA, fatty acid amide hydrolase (FAAH), was reduced by decidualization (7.8 fold, p < 0.001) and unchanged by Ang-(1-7) (p > 0.05), whereas the enzyme metabolizing 2-arachidonoylglycerol, monoacyl glycerol lipase (MAGL), was unchanged by decidualization (p > 0.05) and increased by Ang-(1-7) (1.7 fold, p < 0.001).

View Article: PubMed Central - PubMed

Affiliation: Hypertension and Vascular Research Center, Wake Forest School of Medicine, Winston-Salem, NC, USA. bbrosnih@wakehealth.edu.

ABSTRACT

Background: Endocannabinoids (ECs) are important contributors to implantation and decidualization and are suppressed in early pregnancy. Elevated levels of anandamide (AEA), the endogenous ligand for the CB1 and CB2 receptors (R), interfere with receptivity of the blastocyst. Ang-(1-7) is down-regulated in the implantation site (IS) in normal pregnancy at day 7 of gestation. We determined the effects of intra-uterine angiotensin-(1-7) [Ang-(1-7)] (24 microg/kg/h) or vehicle given into the left uterine horn on the ECs in the decidualized uterus.

Methods: Ovariectomized rats were sensitized for the decidual cell reaction by steroid treatment and decidualization was induced by a bolus of oil injected into the left horn; the right horn served as a control.

Results: Decidualization increased endometrial permeability (3.1+/-0.2 vs. 7.1+/-0.5 uterus/muscle of cpm of (125)I-BSA, p < 0.0001). VEGF mRNA was increased by the decidualization (1.4-fold, p < 0.05) and by Ang-(1-7) (2.0-fold, p < 0.001). CB1R mRNA was reduced by decidualization (2.7-fold, p < 0.001), but increased by Ang-(1-7) (1.9-fold, p < 0.05). CB2R mRNA was increased by decidualization (4-fold, p < 0.05) and by Ang-(1-7) (2.4-fold, p < 0.001). The enzyme metabolizing AEA, fatty acid amide hydrolase (FAAH), was reduced by decidualization (7.8 fold, p < 0.001) and unchanged by Ang-(1-7) (p > 0.05), whereas the enzyme metabolizing 2-arachidonoylglycerol, monoacyl glycerol lipase (MAGL), was unchanged by decidualization (p > 0.05) and increased by Ang-(1-7) (1.7 fold, p < 0.001).

Conclusions: These findings report for the first time that Ang-(1-7) augments the expression of CB1R, CB2R and MAGL in the decidualized uterus and thus may interfere with the early events of decidualization.

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Effects of local intra-uterine infusion of Ang-(1–7) in the decidualized horn on uterine weight (A), permeability (B), and VEGF relative gene expression (C) in non-decidualized vs decidualized uterus. Data are expressed as mean +/− SEM. n = 9–10. *p < 0.05, ***p < 0.001 vs. non-decidualized control; ###p < 0.001 vs. decidualized control; τττp < 0.001 vs. non-decidualized in the Ang-(1–7) treated animal.
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Fig2: Effects of local intra-uterine infusion of Ang-(1–7) in the decidualized horn on uterine weight (A), permeability (B), and VEGF relative gene expression (C) in non-decidualized vs decidualized uterus. Data are expressed as mean +/− SEM. n = 9–10. *p < 0.05, ***p < 0.001 vs. non-decidualized control; ###p < 0.001 vs. decidualized control; τττp < 0.001 vs. non-decidualized in the Ang-(1–7) treated animal.

Mentions: Figure 2 compares the uterine weight and permeability levels in the decidualized uterine horn in the presence or absence of Ang-(1–7) local infusion. There was a 9.6 fold increase in uterine weight in the oil-induced decidualized horn as compared to the non-decidualized horn, which was not altered by Ang-(1–7) infusion. One of the first events of decidualization is an increase in permeability. Permeability measured as the ratio of radiolabeled I(125)-albumin in the uterus relative to the radioactivity of skeletal muscle albumin increased with decidualization, but this increase was unchanged with Ang-(1–7) treatment. VEGF, also known as vascular permeability factor, was increased with decidualization; in the presence of Ang-(1–7) treatment VEGF mRNA expression was augmented 2-fold in the decidualized horn as compared to the control decidualized horn. There was no effect of decidualization or Ang-(1–7) treatment on markers of apoptosis, including caspase 3, caspase 9, and APAF mRNA (Table 1). In addition, uterine tissue was immunostained for vimentin (Figure 3A and B). There was a clear difference in intensity of vimentin staining between the anitmesometrial and mesometrial poles (Figure 3A and B). There was no difference in the antimesometrial staining between treated and non-treated uteri. Vimentin staining was evident in the lining of the vascular components of the mesometrium of the decidulized uterus and was similar in Ang-(1–7)-infused vs. PBS-infused decidualized uterine horns (PBS-infused horn: 0.33+/−0.04 vs. Ang-(1–7)-infused horn: 0.28+/−0.04 relative intensity of vimentin staining, n = 3–5, p > 0.05).Decidualization is characterized by a 2.7–fold decrease in CB1R mRNA levels, 4-fold increase in CB2R mRNA, 7.8 fold decrease in FAAH mRNA, and no significant change in MAGL mRNA levels (Figure 4A-D). The Ang-(1–7) treated decidualized horn (Figure 4A) resulted in a 1.9-fold increase in CB1R mRNA levels as compared to the control decidualized horn; the Ang-(1–7) treated horn, however, remained significantly lower than the non-decidualized horn. The Ang-(1–7) treatment resulted in an augmented 2.4-fold increase in CB2R mRNA levels over the control decidualized horn (Figure 4B); the Ang-(1–7)-treated decidualized horn remained 9-fold higher than the non-decidualized horn. Ang-(1–7) treatment increased the expression of MAGL mRNA by 1.7-fold over its non-decidualized control and by 1.7-fold over the decidualized control (Figure 4C). Although there was a significant reduction in FAAH mRNA levels with decidualization, this reduction was unchanged with Ang-(1–7) treatment in the decidualized horn (Figure 4D).Circulating levels of Ang I, Ang II, and Ang-(1–7) were measured in control and Ang-(1–7) treated pseudopregnant rats. Circulating levels of Ang I and Ang II were unchanged by the intra-uterine infusion of Ang-(1–7) (Figure 5), but there was an increase in circulating levels of Ang-(1–7) with the local uterine Ang-(1–7) infusion.Figure 2


Local uterine Ang-(1-7) infusion augments the expression of cannabinoid receptors and differentially alters endocannabinoid metabolizing enzymes in the decidualized uterus of pseudopregnant rats.

Brosnihan KB, Pulgar VM, Gallagher PE, Neves LA, Yamaleyeva LM - Reprod. Biol. Endocrinol. (2015)

Effects of local intra-uterine infusion of Ang-(1–7) in the decidualized horn on uterine weight (A), permeability (B), and VEGF relative gene expression (C) in non-decidualized vs decidualized uterus. Data are expressed as mean +/− SEM. n = 9–10. *p < 0.05, ***p < 0.001 vs. non-decidualized control; ###p < 0.001 vs. decidualized control; τττp < 0.001 vs. non-decidualized in the Ang-(1–7) treated animal.
© Copyright Policy - open-access
Related In: Results  -  Collection

License 1 - License 2
Show All Figures
getmorefigures.php?uid=PMC4325957&req=5

Fig2: Effects of local intra-uterine infusion of Ang-(1–7) in the decidualized horn on uterine weight (A), permeability (B), and VEGF relative gene expression (C) in non-decidualized vs decidualized uterus. Data are expressed as mean +/− SEM. n = 9–10. *p < 0.05, ***p < 0.001 vs. non-decidualized control; ###p < 0.001 vs. decidualized control; τττp < 0.001 vs. non-decidualized in the Ang-(1–7) treated animal.
Mentions: Figure 2 compares the uterine weight and permeability levels in the decidualized uterine horn in the presence or absence of Ang-(1–7) local infusion. There was a 9.6 fold increase in uterine weight in the oil-induced decidualized horn as compared to the non-decidualized horn, which was not altered by Ang-(1–7) infusion. One of the first events of decidualization is an increase in permeability. Permeability measured as the ratio of radiolabeled I(125)-albumin in the uterus relative to the radioactivity of skeletal muscle albumin increased with decidualization, but this increase was unchanged with Ang-(1–7) treatment. VEGF, also known as vascular permeability factor, was increased with decidualization; in the presence of Ang-(1–7) treatment VEGF mRNA expression was augmented 2-fold in the decidualized horn as compared to the control decidualized horn. There was no effect of decidualization or Ang-(1–7) treatment on markers of apoptosis, including caspase 3, caspase 9, and APAF mRNA (Table 1). In addition, uterine tissue was immunostained for vimentin (Figure 3A and B). There was a clear difference in intensity of vimentin staining between the anitmesometrial and mesometrial poles (Figure 3A and B). There was no difference in the antimesometrial staining between treated and non-treated uteri. Vimentin staining was evident in the lining of the vascular components of the mesometrium of the decidulized uterus and was similar in Ang-(1–7)-infused vs. PBS-infused decidualized uterine horns (PBS-infused horn: 0.33+/−0.04 vs. Ang-(1–7)-infused horn: 0.28+/−0.04 relative intensity of vimentin staining, n = 3–5, p > 0.05).Decidualization is characterized by a 2.7–fold decrease in CB1R mRNA levels, 4-fold increase in CB2R mRNA, 7.8 fold decrease in FAAH mRNA, and no significant change in MAGL mRNA levels (Figure 4A-D). The Ang-(1–7) treated decidualized horn (Figure 4A) resulted in a 1.9-fold increase in CB1R mRNA levels as compared to the control decidualized horn; the Ang-(1–7) treated horn, however, remained significantly lower than the non-decidualized horn. The Ang-(1–7) treatment resulted in an augmented 2.4-fold increase in CB2R mRNA levels over the control decidualized horn (Figure 4B); the Ang-(1–7)-treated decidualized horn remained 9-fold higher than the non-decidualized horn. Ang-(1–7) treatment increased the expression of MAGL mRNA by 1.7-fold over its non-decidualized control and by 1.7-fold over the decidualized control (Figure 4C). Although there was a significant reduction in FAAH mRNA levels with decidualization, this reduction was unchanged with Ang-(1–7) treatment in the decidualized horn (Figure 4D).Circulating levels of Ang I, Ang II, and Ang-(1–7) were measured in control and Ang-(1–7) treated pseudopregnant rats. Circulating levels of Ang I and Ang II were unchanged by the intra-uterine infusion of Ang-(1–7) (Figure 5), but there was an increase in circulating levels of Ang-(1–7) with the local uterine Ang-(1–7) infusion.Figure 2

Bottom Line: CB1R mRNA was reduced by decidualization (2.7-fold, p < 0.001), but increased by Ang-(1-7) (1.9-fold, p < 0.05).CB2R mRNA was increased by decidualization (4-fold, p < 0.05) and by Ang-(1-7) (2.4-fold, p < 0.001).The enzyme metabolizing AEA, fatty acid amide hydrolase (FAAH), was reduced by decidualization (7.8 fold, p < 0.001) and unchanged by Ang-(1-7) (p > 0.05), whereas the enzyme metabolizing 2-arachidonoylglycerol, monoacyl glycerol lipase (MAGL), was unchanged by decidualization (p > 0.05) and increased by Ang-(1-7) (1.7 fold, p < 0.001).

View Article: PubMed Central - PubMed

Affiliation: Hypertension and Vascular Research Center, Wake Forest School of Medicine, Winston-Salem, NC, USA. bbrosnih@wakehealth.edu.

ABSTRACT

Background: Endocannabinoids (ECs) are important contributors to implantation and decidualization and are suppressed in early pregnancy. Elevated levels of anandamide (AEA), the endogenous ligand for the CB1 and CB2 receptors (R), interfere with receptivity of the blastocyst. Ang-(1-7) is down-regulated in the implantation site (IS) in normal pregnancy at day 7 of gestation. We determined the effects of intra-uterine angiotensin-(1-7) [Ang-(1-7)] (24 microg/kg/h) or vehicle given into the left uterine horn on the ECs in the decidualized uterus.

Methods: Ovariectomized rats were sensitized for the decidual cell reaction by steroid treatment and decidualization was induced by a bolus of oil injected into the left horn; the right horn served as a control.

Results: Decidualization increased endometrial permeability (3.1+/-0.2 vs. 7.1+/-0.5 uterus/muscle of cpm of (125)I-BSA, p < 0.0001). VEGF mRNA was increased by the decidualization (1.4-fold, p < 0.05) and by Ang-(1-7) (2.0-fold, p < 0.001). CB1R mRNA was reduced by decidualization (2.7-fold, p < 0.001), but increased by Ang-(1-7) (1.9-fold, p < 0.05). CB2R mRNA was increased by decidualization (4-fold, p < 0.05) and by Ang-(1-7) (2.4-fold, p < 0.001). The enzyme metabolizing AEA, fatty acid amide hydrolase (FAAH), was reduced by decidualization (7.8 fold, p < 0.001) and unchanged by Ang-(1-7) (p > 0.05), whereas the enzyme metabolizing 2-arachidonoylglycerol, monoacyl glycerol lipase (MAGL), was unchanged by decidualization (p > 0.05) and increased by Ang-(1-7) (1.7 fold, p < 0.001).

Conclusions: These findings report for the first time that Ang-(1-7) augments the expression of CB1R, CB2R and MAGL in the decidualized uterus and thus may interfere with the early events of decidualization.

Show MeSH