Y. Figenschau et al., A SIMPLIFIED SERUM-FREE METHOD FOR PREPARATION AND CULTIVATION OF HUMAN GRANULOSA-LUTEAL CELLS, Human reproduction, 12(3), 1997, pp. 523-531
A simplified method for the preparation and long-term cultivation of g
ranulosa-luteal cells in serum-free medium is described, The cells wer
e harvested from women undergoing in-vitro fertilization, enriched by
sedimentation and dissociated by enzymatic treatment. We demonstrated,
by introducing a synthetic serum replacement (SSR2(TM)), that these p
rimary cell cultures cultivated in monolayers on an extracellular matr
ix may be used in experiments exceeding 7 days with low cell loss and
cell death, No adverse effect on progesterone production was found, Th
ere was a high diversity in progesterone production between cells from
individual patients. After several days in culture, the cells were ch
allenged with human chorionic gonadotrophin which revived the rapidly
decreasing progesterone production, We were unable to demonstrate an i
ncrease in cell number after 7 days of cultivation when the cells were
grown in medium supplemented with either serum or SSR2(TM). The mitog
ens epidermal growth factor and basic fibroblast growth factor had no
influence on proliferation, We also found that the present method prev
ents leukocyte contamination in the granulosa-luteal cell cultures, Co
mpared with the common method based on the enrichment of granulosa-lut
eal cells on a density gradient (Ficoll(TM)/Percoll(TM)), this method
saves time, labour and expense, in addition to augmenting purity.