OBJECTIVE: To determine if initial cell plating density alters steroidogenesis and the E(2):P ratio in granulosa cells in long-term serum-free culture. DESIGN: Experimental study. SETTING: Academic institution. ANIMAL(S): Cattle of slaughterhouse origin. INTERVENTION(S): Culture of granulosa cells in vitro at different cell plating density. MAIN OUTCOME MEASURE(S): Steroid secretion was measured by RIA, mRNA levels were measured by real-time polymerase chain reaction, and cell death was assessed by flow cytometry. RESULT(S): Low plating density favored E(2) secretion and mRNA encoding estrogenic enzymes, whereas higher density inhibited E(2) secretion and enhanced P secretion and levels of mRNA encoding progestagenic enzymes. Increasing plating density decreased the E(2):P ratio and cell health. CONCLUSION(S): Lower cell density favors an estrogenic granulosa cell phenotype, whereas higher density favors luteinization. Serum-free culture systems should be optimized with this in mind. Copyright 2010 American Society for Reproductive Medicine. Published by Elsevier Inc. All rights reserved.
OBJECTIVE: To determine if initial cell plating density alters steroidogenesis and the E(2):P ratio in granulosa cells in long-term serum-free culture. DESIGN: Experimental study. SETTING: Academic institution. ANIMAL(S): Cattle of slaughterhouse origin. INTERVENTION(S): Culture of granulosa cells in vitro at different cell plating density. MAIN OUTCOME MEASURE(S): Steroid secretion was measured by RIA, mRNA levels were measured by real-time polymerase chain reaction, and cell death was assessed by flow cytometry. RESULT(S): Low plating density favored E(2) secretion and mRNA encoding estrogenic enzymes, whereas higher density inhibited E(2) secretion and enhanced P secretion and levels of mRNA encoding progestagenic enzymes. Increasing plating density decreased the E(2):P ratio and cell health. CONCLUSION(S): Lower cell density favors an estrogenic granulosa cell phenotype, whereas higher density favors luteinization. Serum-free culture systems should be optimized with this in mind. Copyright 2010 American Society for Reproductive Medicine. Published by Elsevier Inc. All rights reserved.
Authors: Pedro Guillén-García; Elena Rodríguez-Iñigo; Isabel Guillén-Vicente; Rosa Caballero-Santos; Marta Guillén-Vicente; Stephen Abelow; Guillermo Giménez-Gallego; Juan Manuel López-Alcorocho Journal: Cartilage Date: 2014-04 Impact factor: 4.634