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Antoni J. Duleba - One of the best experts on this subject based on the ideXlab platform.
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simvastatin reduces steroidogenesis by inhibiting cyp17a1 gene expression in rat ovarian theca interstitial cells
Biology of Reproduction, 2012Co-Authors: Israel Ortega, Donna H. Wong, Amanda B. Cress, Jesus A. Villanueva, Anna Sokalska, Benjamin C Moeller, Scott D Stanley, Antoni J. DulebaAbstract:Polycystic ovary syndrome (PCOS) is characterized by ovarian enlargement, theca-interstitial hyperplasia, and increased androgen production by theca cells. Previously, our group has demonstrated that statins (competitive inhibitors of 3-hydroxy-3-methyl-glutaryl-coenzyme A reductase, a rate-limiting step of the mevalonate pathway) reduce proliferation of theca-interstitial cells in vitro and decrease serum androgen levels in women with PCOS. The present study evaluated the effect of simvastatin on rat ovarian theca-interstitial cell steroidogenesis. Because actions of statins may be due to reduced cholesterol availability and/or isoprenylation of proteins, the present study also investigated whether steroidogenesis was affected by cell- and mitochondrion-permeable 22-Hydroxycholesterol, isoprenylation substrates (farnesyl-pyrophosphate [FPP] and geranylgeranyl-pyrophosphate [GGPP]), as well as selective inhibitors of farnesyltransferase (FTI) and geranylgeranyltransferase (GGTI). Theca-interstitial cells were cultured for 12, 24, and 48 h with or without simvastatin, GGPP, FPP, FTI, GGTI, and/or 22-Hydroxycholesterol. Simvastatin decreased androgen levels in a time- and concentration-dependent fashion. This inhibitory effect correlated with a decrease in mRNA levels of Cyp17a1, the gene encoding the key enzyme regulating androgen biosynthesis. After 48 h, GGPP alone and FPP alone had no effect on Cyp17a1 mRNA expression; however, the inhibitory action of simvastatin was partly abrogated by both GGPP and FPP. The present findings indicate that statin-induced reduction of androgen levels is likely due, at least in part, to the inhibition of isoprenylation, resulting in decreased expression of CYP17A1.
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Statins Inhibit Growth of Human Theca-Interstitial Cells in PCOS and Non-PCOS Tissues Independently of Cholesterol Availability
The Journal of clinical endocrinology and metabolism, 2010Co-Authors: Anna Sokalska, Izabela J. Rzepczynska, P. Piotrowski, Amanda B. Cress, Antoni J. DulebaAbstract:Context: Polycystic ovary syndrome (PCOS) is associated with ovarian enlargement, prominent theca-interstitial hyperplasia, and excessive androgen production. Recent clinical trials have demonstrated that statins, 3-hydroxy-3-methyl-glutaryl-CoA reductase inhibitors, decrease androgen levels in women with PCOS. Objective: The present study evaluated the effect of statins on proliferation of human ovarian theca-interstitial cells. Design and Settings: In vitro experiments were performed in the university research laboratory. Patients: Human theca-interstitial cells were isolated from ovaries of PCOS (n = 4) and non-PCOS (n = 4) patients. Main Outcome Measures: The cells were incubated for 48 h without additives (control) or with simvastatin (3–30 μm), mevastatin (3–30 μm), and/or the cell- and mitochondrion-permeable form of cholesterol (22-Hydroxycholesterol; 10 μm). To determine whether the effects of statins could be affected by leukocytes, the experiment was carried out on cells not purified of leukocy...
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Role of Isoprenylation in Simvastatin-Induced Inhibition of Ovarian Theca-Interstitial Growth in the Rat
Biology of Reproduction, 2009Co-Authors: Izabela J. Rzepczynska, P. Piotrowski, Donna H. Wong, Amanda B. Cress, Jesus A. Villanueva, Antoni J. DulebaAbstract:Statins are competitive inhibitors of 3-hydroxy-3-methylglutaryl-coenzyme A reductase, a rate-limiting step of the mevalonate pathway. The pleiotropic effects of statins may be due to inhibition of cholesterol synthesis, as well as decreased availability of several biologically important intermediate components of the mevalonate pathway, including two substrates for isoprenylation (farnesyl pyrophosphate [FPP] and geranylgeranyl pyrophosphate [GGPP]). Recently, we demonstrated statin-induced inhibition of ovarian theca-interstitial cell proliferation in vitro, as well as reduction of testosterone levels in women with polycystic ovary syndrome (PCOS). This study evaluates the relative contribution of inhibition of isoprenylation and/or cholesterol availability to the modulation of theca-interstitial proliferation. Rat theca-interstitial cells were cultured in chemically defined media with or without simvastatin, FPP, GGPP, squalene, and/or two membrane-permeable forms of cholesterol (25-Hydroxycholesterol and 22-Hydroxycholesterol). Simvastatin inhibited DNA synthesis and the count of viable cells. The effects of simvastatin were partly abrogated by FPP and GGPP but not by squalene or cholesterol. Inhibition of farnesyl transferase and geranylgeranyl transferase reduced cell proliferation. The present findings indicate that simvastatin inhibits proliferation of theca-interstitial cells, at least in part, by reduction of isoprenylation. These observations provide likely mechanisms explaining clinically observed improvement of ovarian hyperandrogenism in women with PCOS.
D T Armstrong - One of the best experts on this subject based on the ideXlab platform.
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differential modulation of porcine theca granulosa and luteal cell steroidogenesis in vitro by tumor necrosis factor
Biology of Reproduction, 1993Co-Authors: Francis R Tekpetey, Heidi Engelhardt, D T ArmstrongAbstract:The role of tumor necrosis factor (TNFoa) in ovarian function was investigated using in vitro culture of theca and granulosa cells isolated from gilt follicles (4-6 mm) and small (SLC) and large (LLC) luteal cells from mid-cycle corpora lutea. TNFa did not affect basal accumulation of progesterone (P) by theca cells after 72 h of culture. However, TNFa (0.1-100 ng/ml) caused a marked dose-dependent noncytotoxic inhibition (p < 0.05) of LH or LH + insulin (I)-stimulated P accumulation by theca cells after 72 h. Maximal inhibitions averaged 87 + 6% at 5 ng/ml TNFa for LH-stimulated P and 69 ± 4% at 50 ng/ml TNFa for LH + I-stimulated P. The inhibitory effect of TNFa, evident by 24 h after culture, progressively increased on Days 2 and 3 of culture. The effect of TNFao on theca cells was mediated by cAMP generation as evidenced by TNFa inhibition of LH-induced cAMP accumulation and P accumulation in response to LH and forskolin but not dibutyryl cAMP. Consistent with this, TNFca had no effect on increased P accumulation by theca cells in the presence of 22-Hydroxycholesterol or pregnenolone alone, but inhibited further increases in P accumulation stimulated by LH plus sterol substrates. Unlike that in theca cells, FSH-induced P accumulation in granulosa cell cultures was slightly enhanced (p < 0.05) by low doses of TNFa (0.1, 0.5, and 1.0 ng/ml) after 72 h, while higher doses (5-50 ng/ml) did not alter P accumulation. Furthermore, in experiments with luteal cells, TNFot had no effect on P accumulation in cultures of SLC or LLC after 36 h. The effect of TNFa on porcine theca cells may indicate a role in preventing premature follicular luteinization; together with previous evidence, this suggests a complex and possibly evolving physiological role for TNF during follicular development.
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Regulation of steroid production in cultured porcine thecal cells by transforming growth factor-beta.
Molecular and cellular endocrinology, 1992Co-Authors: H Engelhardt, Robert E. Gore-langton, F R Tekpetey, D T ArmstrongAbstract:Evidence that transforming growth factor-beta (TGF beta) is produced by porcine thecal cells and acts upon porcine granulosa cells suggests that this peptide may be a local regulator of follicular function in this species. The objective of the present study was to investigate the effects of TGF beta on steroidogenesis in thecal cells from 4-6 mm follicles of prepubertal gilts. In this culture system, cells undergo functional luteinization such that production of androstenedione, the major steroid product in 24 h incubations, declines, and in the presence of luteinizing hormone (LH) (250 ng/ml) and insulin (1 micrograms/ml), progesterone production increases over a 3-day culture period. TGF beta (0.1-10 ng/ml) had no effect on production of androstenedione from endogenous precursors in the presence or absence of LH, although there was a slight inhibition of androstenedione production in the presence of exogenous progesterone (up to 23%). As the cells luteinized in culture, the increase in progesterone production in response to LH increased (day 1, 4.4-fold; day 3, 13-fold). TGF beta at concentrations as low as 0.1 ng/ml caused marked (up to 90%) inhibition of LH-stimulated progesterone production in day 3 cultures. In the presence of TGF beta (10 ng/ml), the response to LH was completely abolished, and the response to dibutyryl cAMP was considerably attenuated (25% of controls). Since the primary site of action of TGF beta appeared to be distal to cAMP formation, the effect of TGF beta on conversion of exogenous 22-hydroxy-cholesterol and pregnenolone to progesterone was determined in day 3 cultures. 22-Hydroxycholesterol and pregnenolone restored progesterone production to at least 80% and 89% of controls, respectively. While the primary inhibitory action of TGF beta appears to be exerted distal to cAMP formation, neither cholesterol sidechain cleavage nor the 3 beta-hydroxysteroid dehydrogenase: delta 5-delta 4 isomerase reactions are primary targets of this factor. Together with evidence of thecal production of TGF beta, the results of this study indicate that this peptide may be an autocrine regulator of thecal steroidogenesis.
Amanda B. Cress - One of the best experts on this subject based on the ideXlab platform.
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simvastatin reduces steroidogenesis by inhibiting cyp17a1 gene expression in rat ovarian theca interstitial cells
Biology of Reproduction, 2012Co-Authors: Israel Ortega, Donna H. Wong, Amanda B. Cress, Jesus A. Villanueva, Anna Sokalska, Benjamin C Moeller, Scott D Stanley, Antoni J. DulebaAbstract:Polycystic ovary syndrome (PCOS) is characterized by ovarian enlargement, theca-interstitial hyperplasia, and increased androgen production by theca cells. Previously, our group has demonstrated that statins (competitive inhibitors of 3-hydroxy-3-methyl-glutaryl-coenzyme A reductase, a rate-limiting step of the mevalonate pathway) reduce proliferation of theca-interstitial cells in vitro and decrease serum androgen levels in women with PCOS. The present study evaluated the effect of simvastatin on rat ovarian theca-interstitial cell steroidogenesis. Because actions of statins may be due to reduced cholesterol availability and/or isoprenylation of proteins, the present study also investigated whether steroidogenesis was affected by cell- and mitochondrion-permeable 22-Hydroxycholesterol, isoprenylation substrates (farnesyl-pyrophosphate [FPP] and geranylgeranyl-pyrophosphate [GGPP]), as well as selective inhibitors of farnesyltransferase (FTI) and geranylgeranyltransferase (GGTI). Theca-interstitial cells were cultured for 12, 24, and 48 h with or without simvastatin, GGPP, FPP, FTI, GGTI, and/or 22-Hydroxycholesterol. Simvastatin decreased androgen levels in a time- and concentration-dependent fashion. This inhibitory effect correlated with a decrease in mRNA levels of Cyp17a1, the gene encoding the key enzyme regulating androgen biosynthesis. After 48 h, GGPP alone and FPP alone had no effect on Cyp17a1 mRNA expression; however, the inhibitory action of simvastatin was partly abrogated by both GGPP and FPP. The present findings indicate that statin-induced reduction of androgen levels is likely due, at least in part, to the inhibition of isoprenylation, resulting in decreased expression of CYP17A1.
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Statins Inhibit Growth of Human Theca-Interstitial Cells in PCOS and Non-PCOS Tissues Independently of Cholesterol Availability
The Journal of clinical endocrinology and metabolism, 2010Co-Authors: Anna Sokalska, Izabela J. Rzepczynska, P. Piotrowski, Amanda B. Cress, Antoni J. DulebaAbstract:Context: Polycystic ovary syndrome (PCOS) is associated with ovarian enlargement, prominent theca-interstitial hyperplasia, and excessive androgen production. Recent clinical trials have demonstrated that statins, 3-hydroxy-3-methyl-glutaryl-CoA reductase inhibitors, decrease androgen levels in women with PCOS. Objective: The present study evaluated the effect of statins on proliferation of human ovarian theca-interstitial cells. Design and Settings: In vitro experiments were performed in the university research laboratory. Patients: Human theca-interstitial cells were isolated from ovaries of PCOS (n = 4) and non-PCOS (n = 4) patients. Main Outcome Measures: The cells were incubated for 48 h without additives (control) or with simvastatin (3–30 μm), mevastatin (3–30 μm), and/or the cell- and mitochondrion-permeable form of cholesterol (22-Hydroxycholesterol; 10 μm). To determine whether the effects of statins could be affected by leukocytes, the experiment was carried out on cells not purified of leukocy...
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Role of Isoprenylation in Simvastatin-Induced Inhibition of Ovarian Theca-Interstitial Growth in the Rat
Biology of Reproduction, 2009Co-Authors: Izabela J. Rzepczynska, P. Piotrowski, Donna H. Wong, Amanda B. Cress, Jesus A. Villanueva, Antoni J. DulebaAbstract:Statins are competitive inhibitors of 3-hydroxy-3-methylglutaryl-coenzyme A reductase, a rate-limiting step of the mevalonate pathway. The pleiotropic effects of statins may be due to inhibition of cholesterol synthesis, as well as decreased availability of several biologically important intermediate components of the mevalonate pathway, including two substrates for isoprenylation (farnesyl pyrophosphate [FPP] and geranylgeranyl pyrophosphate [GGPP]). Recently, we demonstrated statin-induced inhibition of ovarian theca-interstitial cell proliferation in vitro, as well as reduction of testosterone levels in women with polycystic ovary syndrome (PCOS). This study evaluates the relative contribution of inhibition of isoprenylation and/or cholesterol availability to the modulation of theca-interstitial proliferation. Rat theca-interstitial cells were cultured in chemically defined media with or without simvastatin, FPP, GGPP, squalene, and/or two membrane-permeable forms of cholesterol (25-Hydroxycholesterol and 22-Hydroxycholesterol). Simvastatin inhibited DNA synthesis and the count of viable cells. The effects of simvastatin were partly abrogated by FPP and GGPP but not by squalene or cholesterol. Inhibition of farnesyl transferase and geranylgeranyl transferase reduced cell proliferation. The present findings indicate that simvastatin inhibits proliferation of theca-interstitial cells, at least in part, by reduction of isoprenylation. These observations provide likely mechanisms explaining clinically observed improvement of ovarian hyperandrogenism in women with PCOS.
Heidi Engelhardt - One of the best experts on this subject based on the ideXlab platform.
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differential modulation of porcine theca granulosa and luteal cell steroidogenesis in vitro by tumor necrosis factor
Biology of Reproduction, 1993Co-Authors: Francis R Tekpetey, Heidi Engelhardt, D T ArmstrongAbstract:The role of tumor necrosis factor (TNFoa) in ovarian function was investigated using in vitro culture of theca and granulosa cells isolated from gilt follicles (4-6 mm) and small (SLC) and large (LLC) luteal cells from mid-cycle corpora lutea. TNFa did not affect basal accumulation of progesterone (P) by theca cells after 72 h of culture. However, TNFa (0.1-100 ng/ml) caused a marked dose-dependent noncytotoxic inhibition (p < 0.05) of LH or LH + insulin (I)-stimulated P accumulation by theca cells after 72 h. Maximal inhibitions averaged 87 + 6% at 5 ng/ml TNFa for LH-stimulated P and 69 ± 4% at 50 ng/ml TNFa for LH + I-stimulated P. The inhibitory effect of TNFa, evident by 24 h after culture, progressively increased on Days 2 and 3 of culture. The effect of TNFao on theca cells was mediated by cAMP generation as evidenced by TNFa inhibition of LH-induced cAMP accumulation and P accumulation in response to LH and forskolin but not dibutyryl cAMP. Consistent with this, TNFca had no effect on increased P accumulation by theca cells in the presence of 22-Hydroxycholesterol or pregnenolone alone, but inhibited further increases in P accumulation stimulated by LH plus sterol substrates. Unlike that in theca cells, FSH-induced P accumulation in granulosa cell cultures was slightly enhanced (p < 0.05) by low doses of TNFa (0.1, 0.5, and 1.0 ng/ml) after 72 h, while higher doses (5-50 ng/ml) did not alter P accumulation. Furthermore, in experiments with luteal cells, TNFot had no effect on P accumulation in cultures of SLC or LLC after 36 h. The effect of TNFa on porcine theca cells may indicate a role in preventing premature follicular luteinization; together with previous evidence, this suggests a complex and possibly evolving physiological role for TNF during follicular development.
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Regulation of steroid production in cultured porcine thecal cells by transforming growth factor-β
Molecular and Cellular Endocrinology, 1992Co-Authors: Heidi Engelhardt, Francis R Tekpetey, Robert E. Gore-langton, David T. ArmstrongAbstract:Evidence that transforming growth factor-β (TGFβ) is produced by porcine thecal cells and acts upon porcine granulosa cells suggests that this peptide may be a local regulator of follicular function in this species. The objective of the present study was to investigate the effects of TGFβ on steroidogenesis in thecal cells from 4–6 mm follicles of prepubertal gilts. In this culture system, cells undergo functional luteinization such that production of androstenedione, the major steroid product in 24 h incubations, declines, and in the presence of luteinizing hormone (LH) (250 ng/ml) and insulin (1 μg/ml), progesterone production increases over a 3-day culture period. TGFβ (0.1–10 ng/ml) had no effect on production of androstenedione from endogenous precursors in the presence or absence of LH, although there was a slight inhibition of androstenedione production in the presence of exogenous progesterone (up to 23%). As the cells luteinized in culture, the increase in progesterone production in response to LH increased (day 1, 4.4-fold; day 3, 13-fold). TGFβ at concentrations as low as 0.1 ng/ml caused marked (up to 90%) inhibition of LH-stimulated progesterone production in day 3 cultures. In the presence of TGFβ (10 ng/ml), the response to LH was completely abolished, and the response to dibutyryl cAMP was considerably attenuated (25% of controls). Since the primary site of action of TGFβ appeared to be distal to cAMP formation, the effect of TGFβ on conversion of exogenous 22-hydroxy-cholesterol and pregnenolone to progesterone was determined in day 3 cultures. 22-Hydroxycholesterol and pregnenolone restored progesterone production to at least 80% and 89% of controls, respectively. While the primary inhibitory action of TGFβ appears to be exerted distal to cAMP formation, neither cholesterol sidechain cleavage nor the 3β-hydroxysteroid dehydrogenase: Δ5−Δ4 isomerase reactions are primary targets of this factor. Together with evidence of thecal production of TGFβ, the results of this study indicate that this peptide may be an autocrine regulator of thecal steroidogenesis.
Dale A. Freeman - One of the best experts on this subject based on the ideXlab platform.
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Barbiturates inhibit progesterone synthesis in cultured Leydig tumor cells and human granulosa cells
Gynecological endocrinology : the official journal of the International Society of Gynecological Endocrinology, 1999Co-Authors: Peter M. Gocze, Ildikó Szabó, Z. Porpaczy, Dale A. FreemanAbstract:Screening drugs used in obstetrical practice for effects on steroid hormone synthesis revealed that phenobarbital inhibited progesterone synthesis in MA-10 Leydig tumor cells. The inhibition was apparently a drug class effect since it could be reproduced by other barbiturates. Barbiturate blockade was reversible and could be bypassed in the MA-10 cells by using 22-Hydroxycholesterol. Human granulosa cell progesterone synthesis was also inhibited in a dose dependent fashion by phenobarbital, secobarbital and barbituric acid. Significant inhibition occurred in dose ranges that would be therapeutic for treating epilepsy. From these data we conclude that barbiturates block steroidogenesis by inhibiting cholesterol transport to the cholesterol side chain cleavage enzyme.
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Finasteride blocks progesterone synthesis in MA-10 Leydig tumor cells
Endocrinology, 1993Co-Authors: Dale A. Freeman, Peter M. Gocze, Z. PorpaczyAbstract:The 5 alpha-reductase inhibitor, finasteride, inhibits progesterone synthesis in the MA-10 Leydig tumor cells. Inhibition is dose-dependent with half maximal inhibition occurring at 10 ng/ml, a concentration significantly less than serum concentrations detected in finasteride-treated patients. Experiments to localize the site of inhibition by this compound revealed that the 3 beta-hydroxysteroid dehydrogenase delta 5-->delta 4 isomerase enzyme was not blocked by finasteride, but that cholesterol side-chain cleavage was inhibited. Thus, both dibutyryl-cAMP-stimulated and 22-Hydroxycholesterol-stimulated steroidogenesis were inhibited by finasteride. This effect of finasteride to block cholesterol side-chain cleavage may be species-specific. Inhibition is readily detected in the mouse-derived MA-10 cells; however, human granulosa cell steroidogenesis is finasteride-insensitive while rat Leydig cell steroidogenesis is only minimally effected by finasteride.
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A cholesteryl ester hydrolase inhibitor blocks cholesterol translocation into the mitochondria of MA-10 Leydig tumor cells.
Endocrinology, 1992Co-Authors: Peter M. Gocze, Dale A. FreemanAbstract:The present studies describe an unexpected action of a cholesteryl ester hydrolase inhibitor on MA-10 Leydig tumor cells. These studies were initially intended to use the inhibitor, diethylumbelliferyl phosphate, to block cholesteryl ester hydrolysis and, thus, determine the contributions of this form of cholesterol to steroidogenesis and reveal any direct hormone effects on cholesterol esterification. Although this compound acted as an effective inhibitor of the cholesteryl ester hydrolase in intact MA-10 cells, it inhibited steroidogenesis at lower concentrations and to a greater extent than could be explained by simple inhibition of the ester hydrolase enzyme. This compound proved not to be generally toxic, but blocked some process occurring between cAMP formation and cholesterol side-chain cleavage. The diethylumbelliferyl phosphate block of steroidogenesis was readily bypassed by 22-Hydroxycholesterol. These data indicated that the compound inhibited cholesterol transport. The lesion in cholesterol t...
