Metformin treatment may increase omentin-1 levels in women with polycystic ovary syndrome

Abstract

Objective: Polycystic ovary syndrome (PCOS) is associated with the metabolic syndrome. Decreased omentin-1 levels are associated with obesity and diabetes. To study the effects of metformin treatment on omentin-1 levels in PCOS subjects and effects of omentin-1 on in vitro migration and angiogenesis.

Research design and methods: Serum omentin-1 was measured by ELISA. Angiogenesis was assessed by studying capillary tube formation in human microvascular endothelial cells (HMEC-1) on growth factor reduced Matrigel. Endothelial cell migration assay was performed in a modified Boyden chamber. Nuclear factor-κB (NF-κB) was studied by stably transfecting HMEC-1 cells with a cis-reporter plasmid containing luciferase reporter gene linked to five repeats of NF-κB binding sites. Akt phosphorylation was assessed by Western blotting.

Results: Serum omentin-1 was significantly lower in PCOS women (P < 0.05). After 6 months of metformin treatment, there was a significant increase in serum omentin-1 (P < 0.01). Importantly, changes in hs-CRP were significantly negatively correlated with changes in serum omentin-1 (P = 0.036). In vitro migration and angiogenesis were significantly increased in serum from PCOS women (P < 0.01) compared with matched control subjects; these effects were significantly attenuated by metformin treatment (P < 0.01) plausibly through the regulation of omentin-1 levels via NF-κB and Akt pathways. CRP and VEGF induced in vitro migration, and angiogenesis was significantly decreased by omentin-1.

Conclusions: Increases in omentin-1 levels may play a role but are not sufficient to explain the decreased inflammatory and angiogenic effects of sera from metformin-treated PCOS women.

FIG. 1.

In vitro tube formation by human serum at 24 h from normal control subjects (n = 39) with or without omentin-1 (200 ng/ml), PCOS women (n = 21) with or without omentin-1 (200 ng/ml) or PI3K/Akt inhibitor (LY294002; 10 μmol/l) or CRP immunodepletion (CRP_IP), PCOS women after 6 months of metformin treatment (n = 21) with or without omentin-1 immunodepletion (Omentin-1_IP), CRP (1ug/ml) with or without addition of omentin-1 (200 ng/ml) (n = 6), and VEGF (10 ng/ml) with or without addition of omentin-1 (200 ng/ml) (n = 6), respectively. Data are expressed as % difference of median of normal control subjects [medians (interquartile range)]. Each experiment was performed in three replicates. Group comparison by Kruskal–Wallis ANOVA (post hoc analysis: Dunn's test) and Mann–Whitney U test, respectively. ^#P < 0.05, ^##P < 0.01; *P < 0.05, **P < 0.01.

FIG. 2.

Endothelial cell migration by human serum at 24 h from normal control subjects (n = 39) with or without omentin-1 (200 ng/ml), PCOS women (n = 21) with or without omentin-1 (200 ng/ml) or PI3K/Akt inhibitor (LY294002; 10 μmol/l) or CRP immunodepletion (CRP_IP), PCOS women after 6 months of metformin treatment (n = 21) with or without omentin-1 immunodepletion (Omentin-1_IP), CRP (1ug/ml) with or without addition of omentin-1 (200 ng/ml) (n = 6), and VEGF (10 ng/ml) with or without addition of omentin-1 (200 ng/ml) (n = 6), respectively. Data are expressed as % difference of median of normal control subjects [medians (interquartile range)]. Each experiment was performed in three replicates. Group comparison by Kruskal–Wallis ANOVA (post hoc analysis: Dunn's test) and Mann-Whitney U test, respectively. ^#P < 0.05, ^##P < 0.01; *P < 0.05, **P < 0.01.

FIG. 3.

Effects of serum from normal control subjects (n = 39) with or without omentin-1 (200 ng/ml), PCOS women (n = 21) with or without omentin-1 (200 ng/ml) or CRP immunodepletion (CRP_IP), PCOS women after 6 months of metformin treatment (n = 21) with or without omentin-1 immunodepletion (Omentin-1_IP), CRP (1ug/ml) with or without addition of omentin-1 (200 ng/ml) (n = 6), and TNF-α (10 ng/ml) with or without addition of omentin-1 (200 ng/ml) (n = 6), respectively, on NF-κB activation in HMEC-1 cells stably transfected with pNF-κB-luciferase at 2 h. Cells were lysed, and luciferase activities (RLU) were measured. Data are expressed as % difference of median of basal [medians (interquartile range)]. Each experiment was performed in three replicates. Group comparison by Kruskal–Wallis ANOVA (post hoc analysis: Dunn's test) and Mann–Whitney U test, respectively. ^#P < 0.05, ^##P < 0.01; *P < 0.05, **P < 0.01.

FIG. 4.

Effects of serum from normal control subjects (n = 39), PCOS women (n = 21), and PCOS women after 6 months of metformin treatment with or without addition of omentin-1 (200 ng/ml) (n = 21) on Erk1/2 and Akt phosphorylation in HMEC-1 cells at 2 and 5 min, respectively. Data are expressed as % difference of median of basal [medians (interquartile range)]. Each experiment was performed in three replicates. Group comparison by Kruskal–Wallis ANOVA (post hoc analysis: Dunn's test) and Mann–Whitney U test, respectively. ^#P < 0.05, ^##P < 0.01; *P < 0.05, **P < 0.01.