TY - JOUR
T1 - Characterization of serum-free buffalo granulosa cell culture and analysis of genes involved in terminal differentiation from FSH- to LH-responsive phenotype
AU - Monga, R.
AU - Sharma, I.
AU - Datta, T. K.
AU - Singh, D.
N1 - Funding Information:
We thank the Director, National Dairy Research Institute, Karnal, for providing necessary facilities for this work. We also thank Dr. Ravinder Malhotra, Principal Scientist, Dairy Economics, Statistics and Management, for helping in statistical data analysis. This work was financially supported by NAIP (grant C4/C1056 ) of the Indian Council of Agriculture Research (ICAR), India.
PY - 2011/11
Y1 - 2011/11
N2 - In the present study, buffalo granulosa cells were cultured under serum-free conditions and characterized to study the changes in gene expression associated with the transition of granulosa cells from estrogen- to progesterone-secreting phenotype. The cells were cultured in vitro under completely serum-free conditions for 8 d. Gene expression and hormone analysis showed that on day 4 granulosa cells exhibit FSH responsiveness with preovulatory phenotype having highest CYP19 gene expression and 17β-estradiol production, whereas a significant increase in transcript abundance of STAR, CYP11, and HSD3B genes accompanied with an increase in progesterone production was observed on day 8. Cells treated with LH on day 4 followed by gene expression analysis at 1, 2, 4, 6, 18, and 24 h showed significant increase in transcripts of LH-responsive genes. In conclusion, culture condition used in the present study showed that granulosa cells were FSH responsive and attained attributes of granulosa cells of dominant follicles at day 4 with highest CYP19 and LHR gene expression beyond which they acquired the ability to luteinize and thus were more LH responsive. In addition, after LH treatment, analysis of early LH-responsive genes (EGR2, RUNX1, and NR4A1) on day 4 showed that granulosa cells at this stage in culture exhibits phenotype similar to that of preovulatory follicles before LH surge in vivo and corresponds to the in vivo transition of well-orchestrated gene expression profile after LH surge. The characterized culture conditions represent a suitable in vitro model for analysis of genes involved in terminal differentiation of granulosa cells from FSH- to LH-responsive phenotype during folliculogenesis in buffalo.
AB - In the present study, buffalo granulosa cells were cultured under serum-free conditions and characterized to study the changes in gene expression associated with the transition of granulosa cells from estrogen- to progesterone-secreting phenotype. The cells were cultured in vitro under completely serum-free conditions for 8 d. Gene expression and hormone analysis showed that on day 4 granulosa cells exhibit FSH responsiveness with preovulatory phenotype having highest CYP19 gene expression and 17β-estradiol production, whereas a significant increase in transcript abundance of STAR, CYP11, and HSD3B genes accompanied with an increase in progesterone production was observed on day 8. Cells treated with LH on day 4 followed by gene expression analysis at 1, 2, 4, 6, 18, and 24 h showed significant increase in transcripts of LH-responsive genes. In conclusion, culture condition used in the present study showed that granulosa cells were FSH responsive and attained attributes of granulosa cells of dominant follicles at day 4 with highest CYP19 and LHR gene expression beyond which they acquired the ability to luteinize and thus were more LH responsive. In addition, after LH treatment, analysis of early LH-responsive genes (EGR2, RUNX1, and NR4A1) on day 4 showed that granulosa cells at this stage in culture exhibits phenotype similar to that of preovulatory follicles before LH surge in vivo and corresponds to the in vivo transition of well-orchestrated gene expression profile after LH surge. The characterized culture conditions represent a suitable in vitro model for analysis of genes involved in terminal differentiation of granulosa cells from FSH- to LH-responsive phenotype during folliculogenesis in buffalo.
KW - 17β-Estradiol
KW - Gonadotropins
KW - Progesterone
KW - QPCR
KW - Transcript abundance
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U2 - 10.1016/j.domaniend.2011.07.001
DO - 10.1016/j.domaniend.2011.07.001
M3 - Article
C2 - 21885231
AN - SCOPUS:80053582963
SN - 0739-7240
VL - 41
SP - 195
EP - 206
JO - Domestic Animal Endocrinology
JF - Domestic Animal Endocrinology
IS - 4
ER -