Assessment of Gene Expression on the Gap Junction Connexin of Cumulus Cells on Infertile Women With Polycystic Ovary Syndrome and Poor Ovarian Response: The Novel Role of Propranolol
,
Abstract
Objective: Building upon prior research, our investigation focused on examining changes in gene expression of Connexins 37 and 43 (Cx) influenced by β2-adrenergic agents in cumulus cell cultures from women with polycystic ovarian syndrome (PCOS) and poor ovarian response (POR), all of whom were candidates for in vitro fertilization (IVF).
Materials and methods: This experimental study was conducted between April 2021 and November 2023, involving three groups: a control group (donated eggs) and two study groups (POR and PCO). All three groups received ovulation stimulation drugs. Following oocyte puncture, cumulus cells (CCs) were isolated and placed in a culture medium. After three passages, CCs were exposed to the ADR-β2 agonist isoproterenol and its antagonist propranolol (100nM for both drugs). RNA extraction was performed, and cDNA was synthesized. Real-time PCR was used to determine gene expression, and protein levels were measured through the Western blotting method.
Results: The gene expression of Cx 37/43 was significantly reduced in all three groups (P<0.001). For women with PCO and POR, Isop notably decreased expressions (P<0.001), while Prop increased them (P<0.001). Western Blot results confirmed these findings.
Conclusion: The findings of this in-vitro study suggest that the beta-2 adrenergic antagonist propranolol could upregulate gene expression of Cx37/43 in the cellular connections of CCs among infertile women. Consequently, propranolol may enhance communication between CCs and oocytes, facilitating the transfer of signalling messengers and other essential agents required for oocyte development. This novel discovery could have significant implications for oocyte growth and maturation, offering valuable perspectives on drug treatment and assisted reproductive technology. This novel discovery could have significant implications for oocyte growth and maturation, offering valuable perspectives on drug treatment and assisted reproductive technology.
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3. Liu S, Lan Y, Zhao Y, Zhang Q, Lin T, Lin K,et al. Expression of connexin 43 protein in cardiomyocytes of heart failure mouse model. Front Cardiovasc Med. 2022;9:1028558.
4. Orellana JA, Retamal MA, Moraga-Amaro R, Stehberg J. Role of Astroglial Hemichannels and Pannexons in Memory and Neurodegenerative Diseases. Front Integr Neurosci. 2016; 10:26.
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6. Matzuk MM, Burns KH, Viveiros MM, Eppig JJ. Intercellular communication in the mammalian ovary: Oocytes carry the conversation. Science. 2002; 296: 2178–2180.
7. Alam MH, Miyano T. Interaction between growing oocytes and granulosa cells in vitro. Reprod Med Biol. 2020; 19: 13–23.
8. Zhang Y, Wang Y, Feng X, Zhang S, Xu X, Li L, et al. Oocyte-derived microvilli control female fertility by optimizing ovarian follicle selection in mice. Nat Commun. 2021;12(1):2523.
9. Gupta S, Pandey S, Parmar MS, Somal A, Paul A, Panda BSK, et al. Impact of oocyte-secreted factors on its developmental competence in buffalo. Zygote. 2017; 25: 313–320.
10. Xie J, Xu X, Liu S. Intercellular communication in the cumulus–oocyte complex during folliculogenesis: A review. Front Cell Dev Biol. 2023; 11: 1087612.
11. Decrock E, Vinken M, De Vuyst E, Krysko DV, D’Herde K, Vanhaecke T, et al. Connexin-related signaling in cell death: to live or let die? Cell Death Differ. 2009; 524-536.
12. Wang N, De Vuyst E, Ponsaerts R, Boengler K, Palacios-Prado N, Wauman J, et al. Connexins and their channels in inflammation. Crit Rev Biochem Mol Biol. 2016; 51: 413-439.
13. Mugisho OO, Green CR, Kho DT, Zhang J, Graham ES, Acosta ML, et al. The inflammasome pathway is amplified and perpetuated in an autocrine manner through connexin43 hemichannel mediated ATP release. Biochim Biophys Acta Gen Subj. 2018;1862(3):385-393.
14. Naus CC & Giaume C. Bridging the gap to therapeutic strategies based on connexin/pannexin biology.
J Transl Med. 2016; 14: 330.
15. Jacobowitz D, Wallach EE. Histochemical and chemical studies of the autonomic innervation of the ovary. Endocrinology. 1967; 81: 1132–1139.
16. Lara HE, Ferruz JL, Luza S, Bustamante DA, Borges Y, Ojeda SR. Activation of ovarian sympathetic nerves in polycystic ovary syndrome. Endocrinology. 1993; 133: 2690-5.
17. Stener-Victorin E, Ploj K, Larsson BM, Holmäng A. Rats with steroid-induced polycystic ovaries develop hypertension and increased sympathetic nervous system activity. Reprod Biol Endocrinol. 2005; 3:44.
18. Barria A, Leyton V, Ojeda S R, Lara HE. Ovarian steroidal response to gonadotropins and beta-adrenergic stimulation is enhanced in polycystic ovary syndrome: role of sympathetic innervation. Endocrinology. 1993; 133: 2696-2703.
19. Lara HE, Dorfman M, Venegas M, Luza SM, Luna SL, Mayerhofer A, et al. Changes in sympathetic nerve activity of the mammalian ovary during a normal estrous cycle and in polycystic ovary syndrome: Studies on norepinephrine release. Microsc Res Tech. 2002; 59: 495-502.
20. Hanyaloglu AC, von Zastrow M. Regulation of GPCRs by endocytic membrane trafficking and its potential implications. Annu Rev Pharmacol Toxicol. 2008; 48: 537–568.
21. Wallukat G. The beta-adrenergic receptors. Herz. 2002;27(7):683-90.
22. Li TY, Colley D, Barr KJ, Yee SP, Kidder GM. Rescue of oogenesis in Cx37-null mutant mice by oocyte-specific replacement with Cx43. J Cell Sci. 2007; 120: 4117–4125.
23. Nesbit CB, Huang J, Bhuchitra S, Singh B, Maher JY, Pastore ML. et al. New perspectives on the genetic causes of diminished ovarian reserve and opportunities for genetic screening: systematic review and meta-analysis. F&S Rev. 2020; 1: 1–15.
24. Cruz G, Fernandois D, Paredes AH. Ovarian function and reproductive senescence in the rat: role of ovarian sympathetic innervation. Reproduction. 2017; 153: R59–R68.
25. Ramírez Hernández DA, Vieyra Valdez E, Rosas Gavilán G, Linares Culebro R, Espinoza Moreno JA, Chaparro Ortega A, et al. Role of the superior ovarian nerve in the regulation of follicular development and steroidogenesis in the morning of diestrus 1. J Assist Reprod Genet. 2020; 37: 1477–88.
26. Garrido MP, Fernandois D, Venegas M, Paredes AH. Effects of sympathectomy on ovarian follicular development and steroid secretion. Reproduction. 2018; 155: 173–81.
27. Chen L, Wang H, Zhou H, Bai H, Wang T, Shi W, et al. Follicular Output Rate and Follicle-to-Oocyte Index of Low Prognosis Patients According to POSEIDON Criteria: A Retrospective Cohort Study of 32,128 Treatment Cycles. Front Endocrinol (Lausanne). 2020; 11:181.
28. Rotterdam ESHRE/ASRM-Sponsored PCOS Consensus Workshop Group Revised 2003 consensus on diagnostic criteria and long-term health risks related to polycystic ovary syndrome. Fertil Steril. 2004; 81: 19-25.
29. Bahr J, Kao L, Nalbandov AV. The role of catecholamines and nerves in ovulation. Biol Reprod. 1974; 10: 273-90.
30. Ferruz J, Ahmed CE, Ojeda SR, Lara HE. Norepinephrine release in the immature ovary is regulated by autoreceptors and neuropeptide-Y. Endocrinology. 1992; 130:1345–1351.
31. Pan BO, Li J. The art of oocyte meiotic arrest regulation. Reprod Biol Endocrinol. 2019; 17: 8.
32. Richani D, Gilchrist BR. The epidermal growth factor network: role in oocyte growth, maturation and developmental competence. Hum Reprod Update. 2018; 24:1-14.
33. He M, Zhang T, Yang Y, Wang C. Mechanisms of Oocyte Maturation and Related Epigenetic Regulation. Front Cell Dev Biol. 2021; 9:654028.
34. Pei Z, Deng K, Xu C, Zhang S. The molecular regulatory mechanisms of meiotic arrest and resumption in Oocyte development and maturation. Reprod Biol Endocrinol. 2023;21(1):90.
35. Zhang M, Xia G. Hormonal control of mammalian oocyte meiosis at diplotene stage. Cell Mol Life Sc. 2012; 69: 1279-1288.
36. Zangeneh FZ, Bagheri M, Shoushtari MS, Naghizadeh MM. Expression of ADR-α1, 2 and ADR-β2 in cumulus cell culture of infertile women with polycystic ovary syndrome and poor responder who are a candidate for IVF: the novel strategic role of clonidine in this expression. J Recept Signal Transduct Res. 2021; 41: 263-272.
37. Venegas B, De León Gordillo LY, Rosas G, Espinoza JA, Morán C, Domínguez R, et al. In rats with estradiol valerate-induced polycystic ovary syndrome, the acute blockade of ovarian β-adrenoreceptors improve ovulation. Reprod Biol Endocrinol. 2019;17(1):95.
38. Hatefi A, Zare Shahneh A, Ansari Pirsaraie Z, Alizadeh AM, Atashnak MP, Masoudi R, et al. The stimulation and inhibition of beta-2 adrenergic receptor on the inflammatory responses of ovary and immune system in the aged laying hens. BMC Vet Res. 2021;17(1):195.
39. Anitole-Misleh KG, Brown KM. Developmental regulation of catecholamine levels during sea urchin embryo morphogenesis. Comp Biochem Physiol A Mol Integr Physiol. 2004; 137:39–50.
40. Shu-Rong Z, Bremme K, Eneroth P, Nordberg A. The regulation in vitro of placental release of human chorionic gonadotropin, placental lactogen, and prolactin: effects of an adrenergic beta-receptor agonist and antagonist. Am J Obstet Gynecol. 1982;143(4):444-50.
41. Eppig JJ. Reproduction: Oocytes call, granulosa cells connect. Curr Biol. 2018; 28: R354–R356.
42. Turathum B, Gao E-M, Chian RC. The Function
of cumulus cells in oocyte growth and maturation and in subsequent ovulation and fertilization. Cells. 2021; 10: 2292.
2. Kordowitzki P, Sokołowska G, Wasielak-Politowska M, Skowronski MT. Pannexins and Connexins: Their relevance for oocyte developmental competence. Int
J Mol Sci. 2021; 22: 5918.
3. Liu S, Lan Y, Zhao Y, Zhang Q, Lin T, Lin K,et al. Expression of connexin 43 protein in cardiomyocytes of heart failure mouse model. Front Cardiovasc Med. 2022;9:1028558.
4. Orellana JA, Retamal MA, Moraga-Amaro R, Stehberg J. Role of Astroglial Hemichannels and Pannexons in Memory and Neurodegenerative Diseases. Front Integr Neurosci. 2016; 10:26.
5. Contreras JE, Sáez JC, Bukauskas FF, Bennett MV. Gating and regulation of connexin 43 (Cx43) hemichannels. Proc Natl Acad Sci U S A. 2003;100(20):11388-93.
6. Matzuk MM, Burns KH, Viveiros MM, Eppig JJ. Intercellular communication in the mammalian ovary: Oocytes carry the conversation. Science. 2002; 296: 2178–2180.
7. Alam MH, Miyano T. Interaction between growing oocytes and granulosa cells in vitro. Reprod Med Biol. 2020; 19: 13–23.
8. Zhang Y, Wang Y, Feng X, Zhang S, Xu X, Li L, et al. Oocyte-derived microvilli control female fertility by optimizing ovarian follicle selection in mice. Nat Commun. 2021;12(1):2523.
9. Gupta S, Pandey S, Parmar MS, Somal A, Paul A, Panda BSK, et al. Impact of oocyte-secreted factors on its developmental competence in buffalo. Zygote. 2017; 25: 313–320.
10. Xie J, Xu X, Liu S. Intercellular communication in the cumulus–oocyte complex during folliculogenesis: A review. Front Cell Dev Biol. 2023; 11: 1087612.
11. Decrock E, Vinken M, De Vuyst E, Krysko DV, D’Herde K, Vanhaecke T, et al. Connexin-related signaling in cell death: to live or let die? Cell Death Differ. 2009; 524-536.
12. Wang N, De Vuyst E, Ponsaerts R, Boengler K, Palacios-Prado N, Wauman J, et al. Connexins and their channels in inflammation. Crit Rev Biochem Mol Biol. 2016; 51: 413-439.
13. Mugisho OO, Green CR, Kho DT, Zhang J, Graham ES, Acosta ML, et al. The inflammasome pathway is amplified and perpetuated in an autocrine manner through connexin43 hemichannel mediated ATP release. Biochim Biophys Acta Gen Subj. 2018;1862(3):385-393.
14. Naus CC & Giaume C. Bridging the gap to therapeutic strategies based on connexin/pannexin biology.
J Transl Med. 2016; 14: 330.
15. Jacobowitz D, Wallach EE. Histochemical and chemical studies of the autonomic innervation of the ovary. Endocrinology. 1967; 81: 1132–1139.
16. Lara HE, Ferruz JL, Luza S, Bustamante DA, Borges Y, Ojeda SR. Activation of ovarian sympathetic nerves in polycystic ovary syndrome. Endocrinology. 1993; 133: 2690-5.
17. Stener-Victorin E, Ploj K, Larsson BM, Holmäng A. Rats with steroid-induced polycystic ovaries develop hypertension and increased sympathetic nervous system activity. Reprod Biol Endocrinol. 2005; 3:44.
18. Barria A, Leyton V, Ojeda S R, Lara HE. Ovarian steroidal response to gonadotropins and beta-adrenergic stimulation is enhanced in polycystic ovary syndrome: role of sympathetic innervation. Endocrinology. 1993; 133: 2696-2703.
19. Lara HE, Dorfman M, Venegas M, Luza SM, Luna SL, Mayerhofer A, et al. Changes in sympathetic nerve activity of the mammalian ovary during a normal estrous cycle and in polycystic ovary syndrome: Studies on norepinephrine release. Microsc Res Tech. 2002; 59: 495-502.
20. Hanyaloglu AC, von Zastrow M. Regulation of GPCRs by endocytic membrane trafficking and its potential implications. Annu Rev Pharmacol Toxicol. 2008; 48: 537–568.
21. Wallukat G. The beta-adrenergic receptors. Herz. 2002;27(7):683-90.
22. Li TY, Colley D, Barr KJ, Yee SP, Kidder GM. Rescue of oogenesis in Cx37-null mutant mice by oocyte-specific replacement with Cx43. J Cell Sci. 2007; 120: 4117–4125.
23. Nesbit CB, Huang J, Bhuchitra S, Singh B, Maher JY, Pastore ML. et al. New perspectives on the genetic causes of diminished ovarian reserve and opportunities for genetic screening: systematic review and meta-analysis. F&S Rev. 2020; 1: 1–15.
24. Cruz G, Fernandois D, Paredes AH. Ovarian function and reproductive senescence in the rat: role of ovarian sympathetic innervation. Reproduction. 2017; 153: R59–R68.
25. Ramírez Hernández DA, Vieyra Valdez E, Rosas Gavilán G, Linares Culebro R, Espinoza Moreno JA, Chaparro Ortega A, et al. Role of the superior ovarian nerve in the regulation of follicular development and steroidogenesis in the morning of diestrus 1. J Assist Reprod Genet. 2020; 37: 1477–88.
26. Garrido MP, Fernandois D, Venegas M, Paredes AH. Effects of sympathectomy on ovarian follicular development and steroid secretion. Reproduction. 2018; 155: 173–81.
27. Chen L, Wang H, Zhou H, Bai H, Wang T, Shi W, et al. Follicular Output Rate and Follicle-to-Oocyte Index of Low Prognosis Patients According to POSEIDON Criteria: A Retrospective Cohort Study of 32,128 Treatment Cycles. Front Endocrinol (Lausanne). 2020; 11:181.
28. Rotterdam ESHRE/ASRM-Sponsored PCOS Consensus Workshop Group Revised 2003 consensus on diagnostic criteria and long-term health risks related to polycystic ovary syndrome. Fertil Steril. 2004; 81: 19-25.
29. Bahr J, Kao L, Nalbandov AV. The role of catecholamines and nerves in ovulation. Biol Reprod. 1974; 10: 273-90.
30. Ferruz J, Ahmed CE, Ojeda SR, Lara HE. Norepinephrine release in the immature ovary is regulated by autoreceptors and neuropeptide-Y. Endocrinology. 1992; 130:1345–1351.
31. Pan BO, Li J. The art of oocyte meiotic arrest regulation. Reprod Biol Endocrinol. 2019; 17: 8.
32. Richani D, Gilchrist BR. The epidermal growth factor network: role in oocyte growth, maturation and developmental competence. Hum Reprod Update. 2018; 24:1-14.
33. He M, Zhang T, Yang Y, Wang C. Mechanisms of Oocyte Maturation and Related Epigenetic Regulation. Front Cell Dev Biol. 2021; 9:654028.
34. Pei Z, Deng K, Xu C, Zhang S. The molecular regulatory mechanisms of meiotic arrest and resumption in Oocyte development and maturation. Reprod Biol Endocrinol. 2023;21(1):90.
35. Zhang M, Xia G. Hormonal control of mammalian oocyte meiosis at diplotene stage. Cell Mol Life Sc. 2012; 69: 1279-1288.
36. Zangeneh FZ, Bagheri M, Shoushtari MS, Naghizadeh MM. Expression of ADR-α1, 2 and ADR-β2 in cumulus cell culture of infertile women with polycystic ovary syndrome and poor responder who are a candidate for IVF: the novel strategic role of clonidine in this expression. J Recept Signal Transduct Res. 2021; 41: 263-272.
37. Venegas B, De León Gordillo LY, Rosas G, Espinoza JA, Morán C, Domínguez R, et al. In rats with estradiol valerate-induced polycystic ovary syndrome, the acute blockade of ovarian β-adrenoreceptors improve ovulation. Reprod Biol Endocrinol. 2019;17(1):95.
38. Hatefi A, Zare Shahneh A, Ansari Pirsaraie Z, Alizadeh AM, Atashnak MP, Masoudi R, et al. The stimulation and inhibition of beta-2 adrenergic receptor on the inflammatory responses of ovary and immune system in the aged laying hens. BMC Vet Res. 2021;17(1):195.
39. Anitole-Misleh KG, Brown KM. Developmental regulation of catecholamine levels during sea urchin embryo morphogenesis. Comp Biochem Physiol A Mol Integr Physiol. 2004; 137:39–50.
40. Shu-Rong Z, Bremme K, Eneroth P, Nordberg A. The regulation in vitro of placental release of human chorionic gonadotropin, placental lactogen, and prolactin: effects of an adrenergic beta-receptor agonist and antagonist. Am J Obstet Gynecol. 1982;143(4):444-50.
41. Eppig JJ. Reproduction: Oocytes call, granulosa cells connect. Curr Biol. 2018; 28: R354–R356.
42. Turathum B, Gao E-M, Chian RC. The Function
of cumulus cells in oocyte growth and maturation and in subsequent ovulation and fertilization. Cells. 2021; 10: 2292.
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| Issue | Vol 19, No 2 (June 2025) | |
| Section | Original Articles | |
| DOI | https://doi.org/10.18502/jfrh.v19i2.19299 | |
| Keywords | ||
| Cumulus Cells Gap Junction Cx37/43 Expressions Polycystic Ovarian Syndrome Poor Ovarian Response Beta 2 Adrenergic Receptors Propranolol | ||
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How to Cite
1.
Zangeneh F, Naghizadeh MM, Jafarabadi M, Dehghan M, Bagheri M. Assessment of Gene Expression on the Gap Junction Connexin of Cumulus Cells on Infertile Women With Polycystic Ovary Syndrome and Poor Ovarian Response: The Novel Role of Propranolol. J Family Reprod Health. 2025;19(2):111-121.
