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Proliferative potential and phenotypic analysis of long-term cultivated human granulosa cells initiated by addition of follicular fluid

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The aim of this study was to develop and optimize a strategy for long-term cultivation of luteinizing human granulosa cells (GCs).


GCs were cultivated in DMEM/F12 medium supplemented with 2% fetal calf serum. In vitro proliferation of GCs was supported by follicular fluid as well as FSH and growth factors.


The cultured GCs were maintained for 45 days with a doubling time of 159 ± 24 h. GCs initiated by the addition of follicular fluid and cultivated under low serum conditions reached 10 ± 0.7 population doublings. GCs maintain the typical phenotypic expression and the telomere length according to specific culture conditions.


Our present study has demonstrated that GCs can be maintained in vitro for at least 45 days and this cell model can be beneficial when studying hormonal regulation associated with follicular maturation and preparation of oocytes for fertilization.

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basic fibroblast growth factor


cell line derived from a human granulosa cell tumor (Sigma-Aldrich)


dulbecco’s modified eagle’s medium


doubling time


epidermal growth factor


fetal calf serum


follicular fluid


follicle stimulating hormone


luteinizing human granulosa cells


one of the isolated GCs from patient with different characters


in vitro fertilization


luteinizing hormone


the number of population doublings


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The authors wish to acknowledge MSMT 0021620820, MSMT 0021627502 and IGA MZ NS/9781-3 for financial support of research program. The authors would like to acknowledge Ms. Rückerova for help in the tissue cultures laboratory.

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Correspondence to Tomas Soukup.

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This paper is especially focused on the improvement of the cultivation protocol enabling granulosa cells to proliferate and maintain the typical phenotypic expression.

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Bruckova, L., Soukup, T., Visek, B. et al. Proliferative potential and phenotypic analysis of long-term cultivated human granulosa cells initiated by addition of follicular fluid. J Assist Reprod Genet 28, 939–950 (2011).

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