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Effects of high levels of glucose on the steroidogenesis and the expression of adiponectin receptors in rat ovarian cells.

Chabrolle C, Jeanpierre E, Tosca L, Ramé C, Dupont J - Reprod. Biol. Endocrinol. (2008)

Bottom Line: This was associated with substantial reductions in the amounts of 3beta HSD, p450scc, p450 aromatase and StAR proteins and MAPK ERK1/2 phosphorylation.Streptozotocin treatment did not affect adiponectin receptors in rat ovary but it increased AMPK phosphorylation without affecting MAPK ERK1/2 phosphorylation.However, the mechanism that leads to reduced ovarian steroid production seems to be different.

View Article: PubMed Central - HTML - PubMed

Affiliation: Unité de Physiologie de la Reproduction et des Comportements, Institut National de la Recherche Agronomique, 37380 Nouzilly, France. chabrolle@tours.inra.fr

ABSTRACT

Background: Reproductive dysfunction in the diabetic female rat is associated with altered folliculogenesis and steroidogenesis. However, the molecular mechanisms involved in the reduction of steroid production have not been described. Adiponectin is an adipocytokine that has insulin-sensitizing actions including stimulation of glucose uptake in muscle and suppression of glucose production in liver. Adiponectin acts via two receptor isoforms - AdipoR1 and AdipoR2 - that are regulated by hyperglycaemia and hyperinsulinaemia in liver and muscle. We have recently identified AdipoR1 and AdipoR2 in rat ovary. However, their regulation in ovaries of diabetic female rat remains to be elucidated.

Methods: We incubated rat primary granulosa cells in vitro with high concentrations of glucose (5 or 10 g/l) + or - FSH (10-8 M) or IGF-1 (10-8 M), and we studied the ovaries of streptozotocin-induced diabetic rats (STZ) in vivo. The levels of oestradiol and progesterone in culture medium and serum were measured by RIA. We used immunoblotting to assay key steroidogenesis factors (3beta HSD, p450scc, p450 aromatase, StAR), and adiponectin receptors and various elements of signalling pathways (MAPK ERK1/2 and AMPK) in vivo and in vitro. We also determined cell proliferation by [3H] thymidine incorporation.

Results: Glucose (5 or 10 g/l) impaired the in vitro production in rat granulosa cells of both progesterone and oestradiol in the basal state and in response to FSH and IGF-1 without affecting cell proliferation and viability. This was associated with substantial reductions in the amounts of 3beta HSD, p450scc, p450 aromatase and StAR proteins and MAPK ERK1/2 phosphorylation. In contrast, glucose did not affect the abundance of AdipoR1 or AdipoR2 proteins. In vivo, as expected, STZ treatment of rats caused hyperglycaemia and insulin, adiponectin and resistin deficiencies. Plasma progesterone and oestradiol levels were also reduced in STZ rats. However, the amounts of 3beta HSD and p450 aromatase were the same in STZ rat ovary and controls, and the amounts of StAR and p450scc were higher. Streptozotocin treatment did not affect adiponectin receptors in rat ovary but it increased AMPK phosphorylation without affecting MAPK ERK1/2 phosphorylation.

Conclusion: High levels of glucose decrease progesterone and oestradiol production in primary rat granulosa cells and in STZ-treated rats. However, the mechanism that leads to reduced ovarian steroid production seems to be different. Furthermore, adiponectin receptors in ovarian cells are not regulated by glucose.

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Effect of glucose treatment on basal and FSH- or IGF-1-stimulated progesterone and oestradiol secretions by rat granulosa cells. Granulosa cells from immature rats were cultured for 48 h in medium with serum and then in serum-free medium in the absence or in the presence of 10 g/l glucose ± 10-8 M FSH or 10-8 M IGF-1 for another 48 h (A and B) as described in Materials and Methods. The culture medium was then collected and assayed for progesterone (A) and oestradiol (B) by RIA. Results are means ± SE for three groups of granulosa cells. Each group of granulosa cells was obtained from about 30 rats. Bars with different letters are significantly different (p < 0.05). The letter "a" indicates values which are not significantly different from control (without FSH or IGF-1 and glucose).
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Figure 1: Effect of glucose treatment on basal and FSH- or IGF-1-stimulated progesterone and oestradiol secretions by rat granulosa cells. Granulosa cells from immature rats were cultured for 48 h in medium with serum and then in serum-free medium in the absence or in the presence of 10 g/l glucose ± 10-8 M FSH or 10-8 M IGF-1 for another 48 h (A and B) as described in Materials and Methods. The culture medium was then collected and assayed for progesterone (A) and oestradiol (B) by RIA. Results are means ± SE for three groups of granulosa cells. Each group of granulosa cells was obtained from about 30 rats. Bars with different letters are significantly different (p < 0.05). The letter "a" indicates values which are not significantly different from control (without FSH or IGF-1 and glucose).

Mentions: The effects of glucose treatment on steroidogenesis in rat granulosa cells were first examined in incubations containing various concentrations of glucose for 48 h. Concentrations of glucose greater than 5 g/l were found to inhibit the syntheses of progesterone and oestradiol (data not shown). We also determined whether glucose (10 g/l) affected the production of progesterone and oestradiol in response to FSH or IGF-1. In the presence of FSH (10-8M), glucose (10 g/l, 48 h) decreased the secretion of progesterone (p < 0.01) and oestradiol (p < 0.01) by a factor of about five (Fig. 1A and 1B). In the presence of IGF-1 (10-8M), glucose (10 g/l, 48 h) decreased the secretion progesterone (p < 0.01) and oestradiol (p < 0.01) by a factor of about three (Fig. 1A and 1B). Similar results were obtained with a lower dose of glucose (5 g/l, data not shown). Thus, a high glucose concentration (5 or 10 g/l) decreased both basal and FSH-or IGF-1-stimulated progesterone and oestradiol production in rat granulosa cells.


Effects of high levels of glucose on the steroidogenesis and the expression of adiponectin receptors in rat ovarian cells.

Chabrolle C, Jeanpierre E, Tosca L, Ramé C, Dupont J - Reprod. Biol. Endocrinol. (2008)

Effect of glucose treatment on basal and FSH- or IGF-1-stimulated progesterone and oestradiol secretions by rat granulosa cells. Granulosa cells from immature rats were cultured for 48 h in medium with serum and then in serum-free medium in the absence or in the presence of 10 g/l glucose ± 10-8 M FSH or 10-8 M IGF-1 for another 48 h (A and B) as described in Materials and Methods. The culture medium was then collected and assayed for progesterone (A) and oestradiol (B) by RIA. Results are means ± SE for three groups of granulosa cells. Each group of granulosa cells was obtained from about 30 rats. Bars with different letters are significantly different (p < 0.05). The letter "a" indicates values which are not significantly different from control (without FSH or IGF-1 and glucose).
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC2277425&req=5

Figure 1: Effect of glucose treatment on basal and FSH- or IGF-1-stimulated progesterone and oestradiol secretions by rat granulosa cells. Granulosa cells from immature rats were cultured for 48 h in medium with serum and then in serum-free medium in the absence or in the presence of 10 g/l glucose ± 10-8 M FSH or 10-8 M IGF-1 for another 48 h (A and B) as described in Materials and Methods. The culture medium was then collected and assayed for progesterone (A) and oestradiol (B) by RIA. Results are means ± SE for three groups of granulosa cells. Each group of granulosa cells was obtained from about 30 rats. Bars with different letters are significantly different (p < 0.05). The letter "a" indicates values which are not significantly different from control (without FSH or IGF-1 and glucose).
Mentions: The effects of glucose treatment on steroidogenesis in rat granulosa cells were first examined in incubations containing various concentrations of glucose for 48 h. Concentrations of glucose greater than 5 g/l were found to inhibit the syntheses of progesterone and oestradiol (data not shown). We also determined whether glucose (10 g/l) affected the production of progesterone and oestradiol in response to FSH or IGF-1. In the presence of FSH (10-8M), glucose (10 g/l, 48 h) decreased the secretion of progesterone (p < 0.01) and oestradiol (p < 0.01) by a factor of about five (Fig. 1A and 1B). In the presence of IGF-1 (10-8M), glucose (10 g/l, 48 h) decreased the secretion progesterone (p < 0.01) and oestradiol (p < 0.01) by a factor of about three (Fig. 1A and 1B). Similar results were obtained with a lower dose of glucose (5 g/l, data not shown). Thus, a high glucose concentration (5 or 10 g/l) decreased both basal and FSH-or IGF-1-stimulated progesterone and oestradiol production in rat granulosa cells.

Bottom Line: This was associated with substantial reductions in the amounts of 3beta HSD, p450scc, p450 aromatase and StAR proteins and MAPK ERK1/2 phosphorylation.Streptozotocin treatment did not affect adiponectin receptors in rat ovary but it increased AMPK phosphorylation without affecting MAPK ERK1/2 phosphorylation.However, the mechanism that leads to reduced ovarian steroid production seems to be different.

View Article: PubMed Central - HTML - PubMed

Affiliation: Unité de Physiologie de la Reproduction et des Comportements, Institut National de la Recherche Agronomique, 37380 Nouzilly, France. chabrolle@tours.inra.fr

ABSTRACT

Background: Reproductive dysfunction in the diabetic female rat is associated with altered folliculogenesis and steroidogenesis. However, the molecular mechanisms involved in the reduction of steroid production have not been described. Adiponectin is an adipocytokine that has insulin-sensitizing actions including stimulation of glucose uptake in muscle and suppression of glucose production in liver. Adiponectin acts via two receptor isoforms - AdipoR1 and AdipoR2 - that are regulated by hyperglycaemia and hyperinsulinaemia in liver and muscle. We have recently identified AdipoR1 and AdipoR2 in rat ovary. However, their regulation in ovaries of diabetic female rat remains to be elucidated.

Methods: We incubated rat primary granulosa cells in vitro with high concentrations of glucose (5 or 10 g/l) + or - FSH (10-8 M) or IGF-1 (10-8 M), and we studied the ovaries of streptozotocin-induced diabetic rats (STZ) in vivo. The levels of oestradiol and progesterone in culture medium and serum were measured by RIA. We used immunoblotting to assay key steroidogenesis factors (3beta HSD, p450scc, p450 aromatase, StAR), and adiponectin receptors and various elements of signalling pathways (MAPK ERK1/2 and AMPK) in vivo and in vitro. We also determined cell proliferation by [3H] thymidine incorporation.

Results: Glucose (5 or 10 g/l) impaired the in vitro production in rat granulosa cells of both progesterone and oestradiol in the basal state and in response to FSH and IGF-1 without affecting cell proliferation and viability. This was associated with substantial reductions in the amounts of 3beta HSD, p450scc, p450 aromatase and StAR proteins and MAPK ERK1/2 phosphorylation. In contrast, glucose did not affect the abundance of AdipoR1 or AdipoR2 proteins. In vivo, as expected, STZ treatment of rats caused hyperglycaemia and insulin, adiponectin and resistin deficiencies. Plasma progesterone and oestradiol levels were also reduced in STZ rats. However, the amounts of 3beta HSD and p450 aromatase were the same in STZ rat ovary and controls, and the amounts of StAR and p450scc were higher. Streptozotocin treatment did not affect adiponectin receptors in rat ovary but it increased AMPK phosphorylation without affecting MAPK ERK1/2 phosphorylation.

Conclusion: High levels of glucose decrease progesterone and oestradiol production in primary rat granulosa cells and in STZ-treated rats. However, the mechanism that leads to reduced ovarian steroid production seems to be different. Furthermore, adiponectin receptors in ovarian cells are not regulated by glucose.

Show MeSH
Related in: MedlinePlus