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Research Article Open access CC BY 3.0

Comparison of Medium Supplementation on Proliferation and Hormone Production of Bovine Granulosa Cells in a Defined Culture System

L. H. Montrezor, C. A. Piccinato, C. V. A. Collares, A. A. Vireque, A. A. M. Rosa e Silva

Journal of Scientific Research and Reports · pp. 645–659 · Published 14 Jan 2013

10.9734/JSRR/2014/7168

Abstract

Aims: The objective of this study was to develop a serum-free bovine granulosa cell (GC) culture system in which estradiol (E2) production could be maintained in a defined media, with polyvinyl alcohol (PVA), insulin and insulin-like growth factor-1 (IGF-1) and without FSH and compared the effects of two different macromolecules (PVA vs, BSA) on steroids output and cell proliferation during in vitro culture. Study Design: Bovine granulosa cell culture. Place and Duration of Study: Department of Physiology, School of Medicine of Ribeirão Preto, University of São Paulo, Brazil, 2011. Methodology: Bovine ovaries were collected from adult cows at a local abattoir and were transported in warm saline solution. Small follicles were dissected according to their vascularization and follicular fluid conditions. GC were cultured in α-MEM containing IGF-I, insulin, androstenedione, 0.1% PVA or 0.1% BSA, without FSH. After 48, 96 and 144 h of culture were analyzed GC morphology and secretion of E2 and P4. The relationship among cell shape, cell proliferation, and GC time course of steroidogenesis in vitro was further explored. Results: In the presence of PVA and BSA, E2 production reached its highest production at 144 h. There was a significant increase on P4 production on the medium containing BSA at 48 and 96 h. The changes in E2 production/P4 production ratio in PVA-cultures indicate that there was a larger increase in E2 production by these cells at 48 and 96 hrs than by cells from medium with BSA. The results of the cellular proliferation demonstrated significant tritiated thymidine incorporation in both PVA and BSA cells cultured at 144 hours. Conclusion: These results demonstrate the development of a relevant culture system for bovine GC under defined conditions with PVA. This chemically defined culture system will enable us to study the factors that regulate the physiological control of GC proliferation, differentiation and steroidogenic characteristics.

Steroidogenesis granulosa cells bovine polyvinyl alcohol reproduction

References (40)

  1. 1 Initial and Cyclic Recruitment of Ovarian Follicles [DOI]
  2. 2 Expression of Follicle-Stimulating Hormone and Luteinizing Hormone Receptor Messenger Ribonucleic Acids in Bovine Follicles during the First Follicular Wave1 [DOI]
  3. 3 Follicle Selection in Monovular Species [DOI]
  4. 4 Effects of insulin, insulin-like growth factor I, and gonadotropins on bovine granulosa cell proliferation, progesterone production, estradiol production, and(or) insulin-like growth factor I production in vitro1 [DOI]
  5. 5 Development of a Long-Term Bovine Granulosa Cell Culture System: Induction and Maintenance of Estradiol Production, Response to Follicle-Stimulating Hormone, and Morphological Characteristics1 [DOI]
  6. 6 Differentiation of Dominant Versus Subordinate Follicles in Cattle1 [DOI]
  7. 7 Changes in Messenger Ribonucleic Acid Encoding Luteinizing Hormone Receptor, Cytochrome P450-Side Chain Cleavage, and Aromatase are Associated with Recruitment and Selection of Bovine Ovarian Follicles1 [DOI]
  8. 8 Alterations in the relative abundance of gene transcripts in preimplantation bovine embryos cultured in medium supplemented with either serum or PVA [DOI]
  9. 9 Impact of Bovine Oocyte Maturation Media on Oocyte Transcript Levels, Blastocyst Development, Cell Number, and Apoptosis1 [DOI]
  10. 10 Development of a culture system for bovine granulosa cells: effects of growth hormone, estradiol, and gonadotropins on cell proliferation, steroidogenesis, and protein synthesis. [DOI]
  11. 11 Induction and maintenance of oestradiol and immunoreactive inhibin production with FSH by ovine granulosa cells cultured in serum-free media [DOI]
  12. 12 Effect of the Absence or Presence of Various Protein Supplements on Further Development of Bovine Oocytes During In Vitro Maturation1 [DOI]
  13. 13 In Vitro Growth and Development of Bovine Oocyte-Granulosa Cell Complexes on the Flat Substratum: Effects of High Polyvinylpyrrolidone Concentration in Culture Medium1 [DOI]
  14. 14 Follicle Selection in Cattle: Dynamics of Follicular Fluid Factors During Development of Follicle Dominance1 [DOI]
  15. 15 Interactions Between Follicle-Stimulating Hormone and Growth Factors in Modulating Secretion of Steroids and Inhibin-Related Peptides by Nonluteinized Bovine Granulosa Cells1 [DOI]
  16. 16 Effects of oocyte maturation media on development of pig embryos produced by in vitro fertilization [DOI]
  17. 17 Developmental potential of bovine oocytes cultured in different maturation and culture conditions [DOI]
  18. 18 Regulation of Steroidogenic Genes by Insulin-Like Growth Factor-1 and Follicle-Stimulating Hormone: Differential Responses of Cytochrome P450 Side-Chain Cleavage, Steroidogenic Acute Regulatory Protein, and 3β-Hydroxysteroid Dehydrogenase/Isomerase in Rat Granulosa Cells1 [DOI]
  19. 19 In vitro and in vivo regulation of follicular formation and activation in cattle [DOI]
  20. 20 Simultaneous radioimmunoassay of progestins, androgens and estrogens in rat testis [DOI]
  21. 21 Effects of recombinant bovine somatotrophin, insulin-like growth factor-I and insulin on bovine granulosa cell steroidogenesis in vitro [DOI]
  22. 22 Patterns of [3H] thymidine Incorporation Differ in Immature Rats and Mature, Cycling Rats1 [DOI]
  23. 23 Steroidogenesis in cumulus cells of bovine cumulus–oocyte-complexes matured in vitro with BSA and different concentrations of steroids [DOI]
  24. 24 Ovarian Follicular Steroidogenic Acute Regulatory Protein, Low-Density Lipoprotein Receptor, and Cytochrome P450 Side-Chain Cleavage Messenger Ribonucleic Acids in Cattle Undergoing Superovulation1 [DOI]
  25. 25 Novel function of ovarian growth factors: combined studies by DNA microarray, biochemical and physiological approaches [DOI]
  26. 26 Cholesterol Side-Chain Cleavage Cytochrome P450 and 3β-Hydroxysteroid Dehydrogenase Expression and the Concentrations of Steroid Hormones in the Follicular Fluids of Different Phenotypes of Healthy and Atretic Bovine Ovarian Follicles1 [DOI]
  27. 27 Preimplantation development and expression of Hsp-70 and Bax genes in bovine blastocysts derived from oocytes matured in alpha-MEM supplemented with growth factors and synthetic macromolecules [DOI]
  28. 28 Effect of polyvinylpyrrolidone on bovine oocyte maturation in vitro and subsequent fertilization and embryonic development [DOI]
  29. 29 Secretion and gene expression of inhibin, oxytocin and steroid hormones during the in vitro differentiation of bovine granulosa cells [DOI]
  30. 30 Levels of messenger ribonucleic acid for cholesterol side-chain cleavage cytochrome P-450 and 3 beta-hydroxysteroid dehydrogenase in bovine preovulatory follicles decrease after the luteinizing hormone surge [DOI]
  31. 31 The Effects of Insulin on 3β-Hydroxysteroid Dehydrogenase Expression in Human Luteinized Granulosa Cells [DOI]
  32. 32 In Vitro Production of Estradiol by Bovine Granulosa Cells: Evaluation of Culture Condition, Stage of Follicular Development, and Location of Cells Within Follicles1 [DOI]
  33. 33 Norepinephrine stimulates progesterone production in highly estrogenic bovine granulosa cells cultured under serum-free, chemically defined conditions [DOI]
  34. 34 Differentiation of bovine preovulatory follicles during the follicular phase is associated with increases in messenger ribonucleic acid for cytochrome P450 side-chain cleavage, 3 beta-hydroxysteroid dehydrogenase, and P450 17 alpha-hydroxylase, but not P450 aromatase [DOI]
  35. 35 Expression of Messenger Ribonucleic Acid (mRNA) Encoding 3β-Hydroxysteroid Dehydrogenase Δ 4,Δ 5 Isomerase (3β-HSD) during Recruitment and Selection of Bovine Ovarian Follicles: Identification of Dominant Follicles by Expression of 3β-HSD mRNA within the Granulosa Cell Layer1 [DOI]
  36. 36 Culture of bovine ovarian follicle wall sections maintained the highly estrogenic profile under basal and chemically defined conditions [DOI]
  37. 37 Polyvinyl alcohol is effective in a defined media long-term bovine granulosa cell culture in the maintaince of 17 beta-estradiol production.
  38. 38 The meiotic response of cumulus cell‐enclosed mouse oocytes to follicle‐stimulating hormone in the presence of different macromolecules [DOI]
  39. 39 The effect of maturation time on the subsequent in vitro development of bovine oocytes [DOI]
  40. 40 The role of pregnenolone-metabolizing enzymes in the regulation of oestradiol biosynthesis during development of the first wave dominant follicle in the cow [DOI]

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