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. 2020 Sep 25:7:539496.
doi: 10.3389/fvets.2020.539496. eCollection 2020.

Knockdown of CYP19A1 in Buffalo Follicular Granulosa Cells Results in Increased Progesterone Secretion and Promotes Cell Proliferation

Xingrong Lu  1 , Anqin Duan  1 , Xiaoya Ma  1 , Shasha Liang  1 , Tingxian Deng  1
Affiliations

Knockdown of CYP19A1 in Buffalo Follicular Granulosa Cells Results in Increased Progesterone Secretion and Promotes Cell Proliferation

Xingrong Lu et al. Front Vet Sci. .

Abstract

Cytochrome P450 aromatase 19A1 (CYP19A1) is a critical enzyme in estrogen synthesis. However, the effect of CYP19A1 on cell growth and hormone secretion of buffalo follicular granulosa cells (BFGCs) is poorly understood. The objective of this study was to assess the role of CYP19A1 in cell proliferation and hormone secretion of BFGCs by knocking down CYP19A1 mRNA expression. The mRNA expression level of CYP19A1 gene was knocked down in BFGCs using the siCYP19A1-296 fragment with the best interference efficiency of 72.63%, as affirmed by real-time quantitative PCR (qPCR) and cell morphology analysis. The CYP19A1 knockdown promoted the proliferation of BFGCs through upregulating the mRNA expression levels of six proliferation-related genes (CCND1, CCNE1, CCNB1, CDK2, CDKN1A, and CDKN1B). Moreover, CYP19A1 knockdown increased (P < 0.05) the concentrations of progesterone secretion (P4) in BFGCs through increasing the mRNA expression levels of three steroidogenic genes (HSD17B1, HSD17B7, and CYP17A1). Our data further found that the FSH could inhibit the mRNA expression level of CYP19A1 in BFGCs, while LH obtains the opposite effect. These findings showed that the CYP19A1 knockdown had a regulatory role in the hormone secretion and cell proliferation in BFGCs.

Keywords: CYP19A1; buffalo; cell proliferation; follicular granulosa cell; progesterone secretion.

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Figures

Figure 1
Figure 1
CYP19A1 induced cell proliferation and apoptosis of BFGCs. (A) A screen of CYP19A1 siRNA fragments in BFGCs. (B) Changes of cell morphology after CYP19A1 interference for 72 h. 200 × (bar = 100 μm). Note a: BFGCs transfected with siCYP19A1-277 for 72 h; b: BFGCs transfected with siCYP19A1-296 for 72 h; c: BFGCs transfected with siCYP19A1-375 for 72 h; d: BFGCs transfected with NC for 72 h. (C) Effect of CYP19A1 gene interference on proliferation-related genes expression. (D) Effect of siCYP19A1 on BFGCs by CCK-8 cck8 detection. (E) Effect of CYP19A1 gene interference on apoptosis-related genes expression. (F) Comparison of apoptosis rates between test group and native control group. **P < 0.01; ***P < 0.001; ****P < 0.0001; NS, no significant.
Figure 2
Figure 2
CYP19A1 regulated hormone secretion of BFGCs. (A) Effects of E2 and P4 secretion from BFGCs with CYP19A1 interference; (B) Effect of CYP19A1 knockdown on steroidogenic gene expression. (C) Effects of adding LH and FSH on CYP19A1 expression. *P < 0.05; **P < 0.01; ***P < 0.001.
Figure 3
Figure 3
CYP19A1 regulated cell function and hormone secretion of BFGCs. Red font represents test results.
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