Granulosa cells with stem cell properties in the rat ovary

doi:10.3969/j.issn.2095-4344.2015.01.014

Abstract

Abstract:

BACKGROUND: Human and rat ovarian granulosa cells in dominant follicles have the phenomenon of expressing stem cell characteristics.
OBJECTIVE: To investigate the expression of stem cell-related factors in rat ovarian granulosa cells.
METHODS: After the paraffin sections of rat ovarian tissue, immunohistochemical method was used to detect CD34, CD133, ABCG2/Bcrp1, Pou5f1/Oct-4 expressions. Granulosa cells cultured in vitro were harvested by follicular puncture method, and then the immunohistochemical method was used to detect the expression of FSHR receptor in order to identify the purity of granule cells. In the cultured granulosa cells, CD44 and C-Kit expressions were detected immunohistochemically, RT-PCR was used to detect ABCG2/Bcrp1, Pou5f1/Oct-4, Nanog gene expressions in ovarian tissue and granulosa cells.
RESULTS AND CONCLUSION: Immunohistochemistry detection on paraffin sections showed that a part of ovarian granulosa cells expressed CD34, CD133, ABCG2/Bcrp1 and Pou5f1/Oct-4, and the expression of Pou5f1/Oct-4 protein gradually increased in the development of ovarian follicles, significantly enhanced during the luteal phase, and then disappeared after the formation of corpus albicans, displaying a periodic expression characteristics. FSHR receptor positive identification rate of primary cells harvested by the folliclar puncture method was more than 95%. Granulosa cells cultured in vitro were mainly long spindle-shaped or diamond, and some cells presented with aggregation growth and expressed CD44 and C-Kit. RT-PCR test results showed that there were no Nanog in the ovarian tissue and cultured granulosa cells, low expression of Pou5f1/Oct-4 in the ovarian tissue, strong expression of ABCG2/Bcrp1 in the ovarian tissue, weak expression of Pou5f1/Oct-4 in the cultured granulosa cells, and strong expression of ABCG2/Bcrp1 in the cultured granulosa cells. These findings suggest that a part of granulosa cells in the rat ovarian have the characteristics of stem cells.

中国组织工程研究杂志出版内容重点：干细胞；骨髓干细胞；造血干细胞；脂肪干细胞；肿瘤干细胞；胚胎干细胞；脐带脐血干细胞；干细胞诱导；干细胞分化；组织工程

全文链接：

Key words: Stem Cells, Ovary, Granulosa Cells

CLC Number:

R394.2

Liu Hui-ping, Lin Zheng-hua, Zhang Guo-min, Yu Rong, Liu Wen-e, Li Ling, Gu Xu-yu. Granulosa cells with stem cell properties in the rat ovary[J]. Chinese Journal of Tissue Engineering Research, 2015, 19(1): 78-84.

Figures/Tables 6

References

[1] 龙海红.卵巢相关干细胞研究进展[J].现代生物医学进展,2012, 12(31): 6188-6191.
[2] Bukovsky A, Svetlikova M, Caudle MR.Oogenesis in cultures derived from adult human ovaries.Reprod Biol Endocrinol. 2005;3:17.
[3] Honda A, Hirose M, Hara K,et al.Isolation, characterization, and in vitro and in vivo differentiation of putative thecal stem cells.Proc Natl Acad Sci U S A. 2007;104(30):12389-12394.
[4] Reya T, Morrison SJ, Clarke MF,et al.Stem cells, cancer, and cancer stem cells.Nature. 2001;414(6859):105-111.
[5] Johnson J, Canning J, Kaneko T, et al.Germline stem cells and follicular renewal in the postnatal mammalian ovary. Nature. 2004;428(6979):145-150.
[6] Kossowska-Tomaszczuk K, De Geyter C, De Geyter M, et al. The multipotency of luteinizing granulosa cells collected from mature ovarian follicles.Stem Cells. 2009;27(1):210- 219.
[7] 陈毅斐,许程洁,王鑫泉,等.小鼠卵巢颗粒细胞适宜培养体系及激素干预的影响[J].中国组织工程研究,2012,16(2):282-286.
[8] Dzafic E, Stimpfel M, Virant-Klun I.Plasticity of granulosa cells: on the crossroad of stemness and transdifferentiation potential.J Assist Reprod Genet. 2013;30(10):1255-1261.
[9] Pisarska MD, Barlow G, Kuo FT.Minireview: roles of the forkhead transcription factor FOXL2 in granulosa cell biology and pathology.Endocrinology. 2011;152(4):1199-1208.
[10] Dzafic E, Stimpfel M, Novakovic S,et al.Expression of mesenchymal stem cells-related genes and plasticity of aspirated follicular cells obtained from infertile women.Biomed Res Int. 2014;2014:508216.
[11] Guo JQ, Gao X, Lin ZJ,et al.BMSCs reduce rat granulosa cell apoptosis induced by cisplatin and perimenopause.BMC Cell Biol. 2013;14:18.
[12] Wang F, Wang L, Yao X,et al.Human amniotic epithelial cells can differentiate into granulosa cells and restore folliculogenesis in a mouse model of chemotherapy-induced premature ovarian failure.Stem Cell Res Ther. 2013;4(5): 124.
[13] Chen HF, Jan PS, Kuo HC,et al.Granulosa cells and retinoic acid co-treatment enrich potential germ cells from manually selected Oct4-EGFP expressing human embryonic stem cells.Reprod Biomed Online. 2014;29(3):319-332.
[14] Bhartiya D, Singh J.FSH-FSHR3-stem cells in ovary surface epithelium: basis for adult ovarian biology, failure, aging, and cancer.Reproduction. 2015;149(1):R35-R48.
[15] Zhou S, Schuetz JD, Bunting KD,et al.The ABC transporter Bcrp1/ABCG2 is expressed in a wide variety of stem cells and is a molecular determinant of the side-population phenotype. Nat Med. 2001;7(9):1028-1034.
[16] 何志旭,林槟,黄绍良,等.体外分化胚胎干细胞为造血干细胞重建造血功能的研究[J].中国病理生理学,2008,24(8):1610-1615.
[17] 高芸,商雪莹,吕男男,等. FAT10过表达促进人乳头状甲状腺癌细胞抗凋亡及集落形成能力的研究[J].实用预防医学,2013, 20(6):647-649.
[18] Riva F, Omes C, Bassani R,et al.In-vitro culture system for mesenchymal progenitor cells derived from waste human ovarian follicular fluid.Reprod Biomed Online. 2014;29(4): 457-469.
[19] Maleki M, Ghanbarvand F, Reza Behvarz M,et al.Comparison of mesenchymal stem cell markers in multiple human adult stem cells.Int J Stem Cells. 2014;7(2):118-126.
[20] Rad SM, Bamdad T, Sadeghizadeh M,et al.Transcription factor decoy against stem cells master regulators, Nanog and Oct-4: a possible approach for differentiation therapy.Tumour Biol. 2014 Dec 3. [Epub ahead of print]
[21] Mattioli M, Gloria A, Turriani M,et al.Osteo-regenerative potential of ovarian granulosa cells: an in vitro and in vivo study.Theriogenology. 2012;77(7):1425-1437.
[22] Yuan J, Zhang D, Wang L,et al.No evidence for neo-oogenesis may link to ovarian senescence in adult monkey.Stem Cells. 2013;31(11):2538-2550.
[23] 王研,赵晓娥,杨培先,等.小鼠卵巢颗粒细胞的体外培养[J]. 西北农林科技大学学报,2007,35(8):11-14.
[24] 陈志钊,蔡凯晋. 血清干细胞因子(SCF)及其受体c-kit的研究进展[J].中外医疗,2013,32(13):186-187.
[25] Parrott JA, Skinner MK.Kit-ligand/stem cell factor induces primordial follicle development and initiates folliculogenesis.Endocrinology. 1999;140(9):4262-4271.
[26] 乔威,冉峰,刘长建.人外周血内皮祖细胞的分离、培养及鉴定[J].中国组织工程研究,2013,17(36):6508-6514.
[27] 曹青,王飞,林继先,等. 骨髓间充质干细胞对CD117+心脏干细胞分化过程中转化生长因子βⅢ型受体表达的影响[J].中国组织工程研究与临床康复,2009,13(36):7057-7062.
[28] Kossowska-Tomaszczuk K, De Geyter C.Cells with stem cell characteristics in somatic compartments of the ovary.Biomed Res Int. 2013;2013:310859.
[29] Varras M, Griva T, Kalles V,et al.Markers of stem cells in human ovarian granulosa cells: is there a clinical significance in ART.J Ovarian Res. 2012;5(1):36.