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STUDY QUESTION Does fibrin introduced into the extracellular matrix affect the growth and maturation of individual primate follicles during encapsulated three-dimensional (3D) culture? SUMMARY ANSWER While not altering follicle survival, fibrin–alginate (FIBRIN) improves macaque primary, but not secondary, follicle development during encapsulated 3D culture in terms of growth, steroidogenesis, anti-Müllerian hormone (AMH)/vascular endothelial growth factor (VEGF) production and oocyte maturation. WHAT IS KNOWN ALREADY Efforts to grow non-human primate ovarian follicles from the secondary to the antral stage during encapsulated 3D culture have been successful. However, the growth and maturation of primary follicles in vitro has not been reported in primates, especially in chemically defined conditions. STUDY DESIGN, SIZE, DURATION In vitro follicle maturation was investigated using the rhesus macaque (Macaca mulatta). Ovaries (n = 7 pairs) were obtained during the early follicular phase of the menstrual cycle (cycle day 1–4). Primary (80–120 µm diameter) and secondary (125–225 µm diameter) follicles were isolated mechanically, randomly assigned to experimental groups, encapsulated into alginate (0.25% w/v) or FIBRIN (25 mg/ml fibrinogen–0.25% alginate) and cultured for 13 and 5 weeks, respectively. MATERIALS, SETTING, METHODS Individual follicles were cultured in alpha minimum essential medium supplemented with FSH. Follicle survival and growth were assessed by microscopy. Follicles that reached the antral stage were treated with recombinant hCG. Metaphase II (MII) oocytes were inseminated via ICSI. Follicle morphology was evaluated by hematoxylin and eosin (H&E) staining. Immunohistochemistry was performed for cytochrome P450 family 17 subfamily A polypeptide 1 (CYP17A1) and 19 subfamily A polypeptide 1 (CYP19A1). Culture medium was analyzed for estradiol (E2) and progesterone by chemiluminescence, androstenedione (A4) by radioimmunoassay, as well as anti-Müllerian hormone (AMH) and vascular endothelial growth factor (VEGF) by enzyme-linked immunosorbent assay. MAIN RESULTS AND THE ROLE OF CHANCE A total of 105 primary and 133 secondary follicles were collected. The presence of fibrin in the alginate matrix had no effect on either primary or secondary follicle survival. Growing primary and secondary follicles formed an antrum at Weeks 9 and 3, respectively. The percentage of growing follicles was higher (P < 0.05) for primary follicles cultured in FIBRIN than alginate at Week 13. The diameters were larger for the growing secondary follicles cultured in alginate than FIBRIN at Week 5 (P < 0.05). H&E staining revealed the typical morphology for small antral follicles. CPY17A1 immunostaining was detected in theca cells, while CYP19A1 was observed in granulosa cells. E2 increased (P < 0.05) during antrum formation in growing follicles at Week 9 for primary and Week 3 for secondary follicles. AMH levels in medium from growing primary follicles increased (P < 0.05) after Week 4 with peak levels at Weeks 9–11. AMH increased (P < 0.05) in growing secondary follicles at Weeks 3–5. VEGF levels in medium were elevated (P < 0.05) in growing primary follicles at Week 9. VEGF increased (P < 0.05) in medium from growing secondary follicles at Weeks 3–5. E2, AMH and VEGF production was higher (P < 0.05) in primary follicle culture with FIBRIN than alginate alone. One primary follicle cultured in FIBRIN (1 of 5 follicles harvested) and a secondary follicle cultured in alginate alone (1 of 15 follicles harvested) yielded an MII oocyte. The fertilized oocyte from primary follicle culture arrested without cell division after fertilization, while the oocyte from secondary follicle culture cleaved and reached the morula stage. LIMITATIONS, REASONS FOR ... [ABSTRACT FROM AUTHOR]

STUDY QUESTION Is decorin (DCN), a putative modulator of growth factor (GF) signaling, expressed in the primate ovary and does it play a role in ovarian biology? SUMMARY ANSWER DCN expression in the theca, the corpus luteum (CL), its presence in the follicular fluid (FF) and its actions revealed in human IVF-derived granulosa cells (GCs), suggest that it plays multiple roles in the ovary including folliculogenesis, ovulation and survival of the CL. WHAT IS KNOWN ALREADY DCN is a secreted proteoglycan, which has a structural role in the extracellular matrix (ECM) and also interferes with the signaling of multiple GF/GF receptors (GFRs). However, DCN expression and action in the primate ovary has yet to be determined. STUDY DESIGN, SIZE, DURATION Archival human and monkey ovarian samples were analyzed. Studies were conducted using FF and GC samples collected from IVF patients. PARTICIPANTS/MATERIALS, SETTING, METHODS Immunohistochemistry, western blotting, RT–PCR, quantitative RT–PCR (qPCR) and enzyme-linked immunosorbent assay (ELISA) studies were complemented by cellular studies, including the measurements of intracellular Ca2+, reactive oxygen species (ROS), epidermal GF receptor (EGFR) phosphorylation by DCN and caspase activity. MAIN RESULTS AND THE ROLE OF CHANCE Immunohistochemistry revealed strong DCN staining in the connective tissue and follicular thecal compartments, but not in GCs of pre-antral and antral follicles. Pre-ovulatory follicles could not be studied, but DCN was associated with connective tissue of CL samples and the cytoplasm of luteal cells. DCN expression in monkey CL doubled (P < 0.05) towards the end of the luteal lifespan. DCN was found in human FF obtained from IVF patients (mean: 12.9 ng/ml; n = 20) as determined by ELISA. DCN mRNA and/or protein were detected in freshly isolated and cultured, luteinized human GCs. In the latter, exogenous human recombinant DCN increased intracellular Ca2+ levels and induced the production of ROS in a concentration-dependent manner. DCN, like epidermal GF, phosphorylated EGFR significantly (P < 0.05) and reduced the activity of caspase 3/7 in cultured GCs. The data indicate the expression of DCN in the theca of growing follicles, in FF of ovulatory follicles and in the CL. Therefore, DCN may exert paracrine actions via GF/GFR systems in multiple ovarian compartments. LIMITATIONS, REASONS FOR CAUTION Functional studies were performed in cultures of human luteinized GCs, which are an apt model but may not fully mirror the pre-ovulatory GC compartment or the CL. Other human ovarian cells, including the thecal cells, were not available. WIDER IMPLICATIONS OF THE FINDINGS In accordance with its evolving roles in other organs, ovarian DCN is an ECM-associated component, which acts as a multifunctional regulator of GF signaling in the primate ovary. DCN may thus be involved in folliculogenesis, ovulation and the regulation of the CL survival in primates. STUDY FUNDING/COMPETING INTEREST(S) This study was supported by Deutsche Forschungsgemeinschaft (DFG) MA1080/17-3 and in part DFG MA1080/21-1 (to AM), NIH grants HD24870 (S.R.O. and R.L.S.), the Eunice Kennedy Shriver NICHD/NIH through cooperative agreement HD18185 as part of the Specialized Cooperative Centers Program in Reproduction and Infertility Research (S.R.O.) and 8P51OD011092-53 for the operation of the Oregon National Primate Research Center (G.A.D., J.D.H., S.R.O. and R.L.S). [ABSTRACT FROM PUBLISHER]