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Effects of α-enolase gene silencing on reproductive-related hormone receptor expression and steroid hormone synthesis of primary granulosa cells from goose F1 follicles

Hong Ji
Hong Ji
, 
Chun-Yang Niu
Chun-Yang Niu
, 
Hong-Liang Zhang
Hong-Liang Zhang
, 
Jing-Ru Guo
Jing-Ru Guo
, 
Li Zhen
Li Zhen
, 
Shuai Lian
Shuai Lian
, 
Chuang Yang
Chuang Yang
, 
Huan-Min Yang
Huan-Min Yang
 y 
Jian-Fa Wang
Jian-Fa Wang
   | 31 ene 2020
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Publicado en línea: 31 ene 2020
Páginas: 141 - 149
Recibido: 06 jun 2019
Aceptado: 17 ene 2020
DOI: https://doi.org/10.2478/jvetres-2020-0008
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© 2020 H. Ji et al. published by Sciendo
This work is licensed under the Creative Commons Attribution-NonCommercial-NoDerivatives 3.0 License.

Fig. 1

Transfection of shRNA-ENO1 in granulosa cells and detection of ENO1 expression. A – transfection of shRNA-ENO1 in granulosa cells (100 ×) including granulosa cells under ordinary light and fluorescence. B – ENO1 mRNA expression by qRT-PCR in granulosa cells transfected with shRNA-ENO1 after 48 h. ENO1 mRNA expression by qRT-PCR in granulosa cells was measured by qRT-PCR. C – ENO1 expression by Western blot in granulosa cells transfected with shRNA-ENO1 after 48 h. The significance of difference for the ENO1 expression level among the different groups was determined by Student’s t-test. ** P < 0.01, and * P < 0.05 when compared with the control sample
Transfection of shRNA-ENO1 in granulosa cells and detection of ENO1 expression. A – transfection of shRNA-ENO1 in granulosa cells (100 ×) including granulosa cells under ordinary light and fluorescence. B – ENO1 mRNA expression by qRT-PCR in granulosa cells transfected with shRNA-ENO1 after 48 h. ENO1 mRNA expression by qRT-PCR in granulosa cells was measured by qRT-PCR. C – ENO1 expression by Western blot in granulosa cells transfected with shRNA-ENO1 after 48 h. The significance of difference for the ENO1 expression level among the different groups was determined by Student’s t-test. ** P < 0.01, and * P < 0.05 when compared with the control sample

Fig. 2

Quantitative RT-PCR–detected expression of FSHR, LHR, ERα, ERβ, GHR, and IGFBP-1 mRNA in the granulosa transfected with shRNA-ENO1 after 48 h detected by qRT-PCR. A – FSHR, B – LHR, C – GHR, D – IGFBP-1, E – ERα, F – ERβ. The significance of difference for the hormone receptor levels were determined by Student’s t-test. ** p < 0.01, and * p < 0.05 when compared with the control sample
Quantitative RT-PCR–detected expression of FSHR, LHR, ERα, ERβ, GHR, and IGFBP-1 mRNA in the granulosa transfected with shRNA-ENO1 after 48 h detected by qRT-PCR. A – FSHR, B – LHR, C – GHR, D – IGFBP-1, E – ERα, F – ERβ. The significance of difference for the hormone receptor levels were determined by Student’s t-test. ** p < 0.01, and * p < 0.05 when compared with the control sample

Fig. 3

ELISA-detected concentration of six hormones in the supernatant of granulosa transfected with shRNA-ENO1 after 48 h. A – oestradiol. B – activin. C – progesterone. D – testosterone. E – inhibin. F – follistatin. The significance of difference for the hormone levels was determined by Student’s t-test. ** P < 0.01, and * P < 0.05 when compared with the control sample
ELISA-detected concentration of six hormones in the supernatant of granulosa transfected with shRNA-ENO1 after 48 h. A – oestradiol. B – activin. C – progesterone. D – testosterone. E – inhibin. F – follistatin. The significance of difference for the hormone levels was determined by Student’s t-test. ** P < 0.01, and * P < 0.05 when compared with the control sample

List of primers used for quantitative RT-PCR

Target gene Primer sequence (5′–3′) Amplicon size (bp)
ENO1 F: GCGGTGCCTCAACTGGAATT 183
R: CCATGTCCAGCATCAGTTTGTC
GAPDH

F: GCTGATGCTCCCATGTTCGTGAT

R: GTGGTGCAAGAGGCATTGCTGAC		 86
LHR

F: GTAACACTGGAATAAGGGAAT

R: GAAGGCTTGACTGTGGATA		 191
ERα

F: ACCCAAACAGACCATTCAACGAA

R: CGCCAGACTAAGCCAATCATCAG		 187
ERβ

F: AAGTGGGAATGATGAAATGTGGC

R: GGACTGACCGTGCTGAGGAGAAT		 163
FSHR

F: TCCTGTGCTAACCCTTTCCTCTA

R: AACCAGTGAATAAATAGTCCCATC		 207
IGFBP-1

F: CCTTGTCAGAAGGAGCTCTA

R: CATCCAGTGAAGTTTCACACT		 145
GHR

F: GCCCCTGCTGACATTTGAGAAT

R: GGCCACTGCAGAAGATCATCAC		 115
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Life Sciences, Molecular Biology, Microbiology and Virology, other, Medicine, Veterinary Medicine
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