牛卵泡颗粒细胞PRSS35表达模式及功能研究

doi:10.11843/j.issn.0366-6964.2018.11.010

畜牧兽医学报 ›› 2018, Vol. 49 ›› Issue (11): 2394-2401.doi: 10.11843/j.issn.0366-6964.2018.11.010

牛卵泡颗粒细胞PRSS35表达模式及功能研究

毕锡麟¹, 王锴¹, 李鹏飞², 景炅婕¹, 韩琦¹, 吕丽华^1*

1. 山西农业大学动物科技学院, 太谷 030801;
2. 山西农业大学生命科学学院, 太谷 030801

收稿日期:2018-07-16 出版日期:2018-11-23 发布日期:2018-11-23
通讯作者: 吕丽华,E-mail:lihualvsxau@126.com
作者简介:毕锡麟(1991-),男,山西太原人,硕士生,主要从事动物生殖生理研究,E-mail:342864159@qq.com
基金资助:
山西省回国留学人员科研资助项目（2014-重点5）；山西省国际科技合作项目（201603D421006）；山西省青年三晋学者项目；山西省重点研发计划（一般）农业项目（201703D221020-1）；山西农业大学创新基金项目（zdpy201403/201503）

Expression Profile and Function Analysis of PRSS35 in Bovine Follicle Granulosa Cells

BI Xi-lin¹, WANG Kai¹, LI Peng-fei², JING Jiong-jie¹, HAN Qi¹, LÜ Li-hua^1*

1. College of Animal Science and Veterinary Medicine, Shanxi Agricultural University, Taigu 030801, China;
2. College of Life Sciences, Shanxi Agricultural University, Taigu 030801, China

Received:2018-07-16 Online:2018-11-23 Published:2018-11-23

摘要/Abstract

摘要：

旨在探究丝氨酸蛋白酶35（serine protease 35，PRSS35）在牛卵泡颗粒细胞（granulosa cells，GCs）中的表达及功能。采用B超超声波检测确定牛优势卵泡（dominant follicle，DF）与从属卵泡（subordinate follicles，SF），分离优势卵泡与从属卵泡颗粒细胞；利用qRT-PCR和Western blotting技术检测PRSS35在DF与SF的表达情况；免疫组化技术对PRSS35进行定位研究；设计PRSS35 siRNA基因沉默序列，经脂质体转染GCs，测定培养液雌激素（estrogen，E₂）浓度和GCs凋亡情况，研究PRSS35对牛卵泡颗粒细胞生长发育的影响。结果表明：1）PRSS35 mRNA在DF中的表达量极显著高于SF （P<0.001）；2） PRSS35在牛DF和SF颗粒层与膜层均有表达；3） PRSS35在DF中的表达量极显著高于SF （P<0.01）；4）PRSS35基因沉默后，GCs凋亡细胞百分率极显著增高（P<0.001），培养液雌激素浓度极显著降低（P<0.01）。综上表明，PRSS35在牛卵泡颗粒层和膜层均有表达，且在DF表达量极显著高于SF；PRSS35基因沉默后，通过抑制GCs增殖，降低E₂分泌浓度，从而抑制牛卵泡的生长发育。该研究为深入探讨PRSS35与牛卵泡发育的关系奠定基础。

Abstract:

The aim of this study was to explore the expression profile and analyze the function of serine protease 35 (PRSS35) in bovine follicular granulosa cells. Cattle dominant follicles (DF) and subordinate follicles (SF) were separated according their sizes measured using B-mode ultrasonography technology. The expression profile of PRSS35 was analyzed using qRT-PCR and Western blotting, and its location in DF and SF was detected by immunohistochemistry. The siRNA sequences of PRSS35 were designed and synthesized and PRSS35-gene-silenced GCs line was obtained. Estrogen (E₂) concentration and cell apoptosis were detected through studying the effects of PRSS35 on growth and development of bovine follicular GCs. The results showed that:1) The expression of PRSS35 mRNA in DF was significantly higher than that in SF (P<0.001). 2) The PRSS35 was expressed both in DF and SF granular layer and theca layer. 3) The expression of RSS35 protein in DF was significantly higher than that in SF (P<0.01). 4) When PRSS35 gene was silenced, the percentage of apoptotic GCs was significantly increased (P<0.001), and the concentration of E₂ in cell culture media was significantly decreased (P<0.01). The results suggested that PRSS35 was expressed both in bovine follicular granular layer and theca layer, and the PRSS35 expression was significantly higher in DF than that in SF. The proliferation of GCs was inhibited, and the concentration of E₂ was reduced when PRSS35 gene was silenced, indicating the growth and development of bovine follicles were inhibited. This study provides a basis for further studying the relationship between PRSS35 and follicular development in cattle.

中图分类号:

S823.2

毕锡麟, 王锴, 李鹏飞, 景炅婕, 韩琦, 吕丽华. 牛卵泡颗粒细胞PRSS35表达模式及功能研究[J]. 畜牧兽医学报, 2018, 49(11): 2394-2401.

BI Xi-lin, WANG Kai, LI Peng-fei, JING Jiong-jie, HAN Qi,Lü Li-hua. Expression Profile and Function Analysis of PRSS35 in Bovine Follicle Granulosa Cells[J]. ACTA VETERINARIA ET ZOOTECHNICA SINICA, 2018, 49(11): 2394-2401.

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参考文献

[1] STOCK A E,FORTUNE J E.Ovarian follicular dominance in cattle:relationship between prolonged growth of the ovulatory follicle and endocrine parameters[J].Endocrinology,1993,132(3):1108-1114.
[2] FRICKE P M,AL-HASSAN M J,ROBERTS A J,et al.Effect of gonadotropin treatment on size,number,and cell proliferation of antral follicles in cows[J].Domest Anim Endocrinol,1997,14(3):171-180.
[3] SHELDON I M,NOAKES D E,RYCROFT A N,et al.Influence of uterine bacterial contamination after parturition on ovarian dominant follicle selection and follicle growth and function in cattle[J].Reproduction,2002,123(6):837-845.
[4] ARMSTRONG D G,GUTIERREZ C G,BAXTER G,et al.Expression of mRNA encoding IGF-I,IGF-Ⅱ and type 1 IGF receptor in bovine ovarian follicles[J].J Endocrinol,2000,165(1):101-113.
[5] SEN A,BETTEGOWDA A,JIMENEZ-KRASSEL F,et al.Cocaine-and amphetamine-regulated transcript regulation of follicle-stimulating hormone signal transduction in bovine granulosa cells[J].Endocrinology,2007,148(9):4400-4410.
[6] 李鹏飞,孟金柱,郝庆玲,等.PDF2和ODF1转录组测序筛选牛卵泡发育相关基因[J].畜牧兽医学报,2018,49(2):300-309.
LI P F,MENG J Z,HAO Q L,et al.Screening and analysing of genes associated with follicular development in bovine ODF1 and PDF2 transcriptome[J].Acta Veterinaria et Zootechnica Sinica,2018,49(2):300-309.(in Chinese)
[7] DIAO H L,XIAO S,LI R,et al.Distinct spatiotemporal expression of serine proteases prss23 and prss35 in periimplantation mouse uterus and dispensable function of prss35 in fertility[J].PLoS One,2013,8(2):e56757.
[8] MIYAKOSHI K,MURPHY M J,YEOMAN R R,et al.The identification of novel ovarian proteases through the use of genomic and bioinformatic methodologies[J].Biol Reprod,2006,75(6):823-835.
[9] GOES F S,HAMSHERE M L,SEIFUDDIN F,et al.Genome-wide association of mood-incongruent psychotic bipolar disorder[J].Transl Psych,2012,2(10):e180.
[10] WAHLBERG P,NYLANDER Å,AHLSKOG N,et al.Expression and localization of the serine proteases high-temperature requirement factor A1,Serine Protease 23,and Serine Protease 35 in the mouse ovary[J].Endocrinology,2008,149(10):5070-5077.
[11] LI S H,LIN M H,HWU Y M,et al.Correlation of cumulus gene expression of GJA1,PRSS35,PTX3,and SERPINE2 with oocyte maturation,fertilization,and embryo development[J].Reprod Biol Endocrinol,2015,13(1):93.
[12] LETRA A,MENEZES R,FONSECA R F,et al.Novel cleft susceptibility genes in chromosome 6p[J].J Dent Res,2010,89(9):927-932.
[13] LEBLEU V S,TENG Y Q,O'CONNELL J T,et al.Identification of human epididymis protein-4 as a fibroblast-derived mediator of fibrosis[J].Nat Med,2013,19(2):227-231.
[14] REYNOLDS A,LEAKE D,BOESE Q,et al.Rational siRNA design for RNA interference[J].Nat Biotechnol,2004,22(3):326-330.
[15] VELAZQUEZ M A,SPICER L J,WATHES D C.The role of endocrine insulin-like growth factor-I (IGF-I) in female bovine reproduction[J].Domest Anim Endocrinol,2008,35(4):325-342.
[16] DROST M.Bubaline versus bovine reproduction[J].Theriogenology,2007,68(3):447-449.
[17] SUNDERLAND S J,CROWE M A,BOLAND M P,et al.Selection,dominance and atresia of follicles during the oestrous cycle of heifers[J].J Reprod Fertil,1994,101(3):547-555.
[18] MANABE N,GOTO Y,MATSUDA-MINEHATA F,et al.Regulation mechanism of selective atresia in porcine follicles:regulation of granulosa cell apoptosis during atresia[J].J Reprod Dev,2004,50(5):493-514.
[19] GINTHER O J.Selection of the dominant follicle in cattle and horses[J].Anim Reprod Sci,2000,60-61:61-79.
[20] KOBAYASHI Y,JIMENEZ-KRASSEL F,IRELAND J J,et al.Evidence of a local negative role for cocaine and amphetamine regulated transcript (CART),inhibins and low molecular weight insulin like growth factor binding proteins in regulation of granulosa cell estradiol production during follicular waves in cattle[J].Reprod Biol Endocrinol,2006,4:22.
[21] GINTHER O J,KOT K,KULICK L J,et al.Sampling follicular fluid without altering follicular status in cattle:oestradiol concentrations early in a follicular wave[J].J Reprod Fertil,1997,109(2):181-186.
[22] SARTORI R,FRICKE P M,FERREIRA J C,et al.Follicular deviation and acquisition of ovulatory capacity in bovine follicles[J].Biol Reprod,2001,65(5):1403-1409.
[23] FORTUNE J E.Ovarian follicular growth and development in mammals[J].Biol Reprod,1994,50(2):225-232.
[24] SHIMIZU T,KOSAKA N,MURAYAMA C,et al.Apelin and APJ receptor expression in granulosa and theca cells during different stages of follicular development in the bovine ovary:involvement of apoptosis and hormonal regulation[J].Anim Reprod Sci,2009,116(1-2):28-37.
[25] 李鹏飞,孟金柱,郝庆玲,等.胰岛素和FSH对体外培养猪卵泡颗粒细胞雌激素的影响[J].畜牧兽医学报,2017,48(11):2084-2090.
LI P F,MENG J Z,HAO Q L,et al.Effects of insulin and FSH on E₂ production of pig ovarian follicular granulosa cells in vitro culture[J].Acta Veterinaria et Zootechnica Sinica,2017,48(11):2084-2090.(in Chinese)

牛卵泡颗粒细胞PRSS35表达模式及功能研究

Expression Profile and Function Analysis of PRSS35 in Bovine Follicle Granulosa Cells

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