Biphasic Effect of TNF-α on the Survival and Endocrine Function of Human Ovarian Granulosa Cells

Abstract

Biphasic Effect of TNF-α on the Survival and Endocrine Function of Human Ovarian Granulosa Cells Haiyan Liang1,† , Dongkui Xu2,† , Xianhong Tong3 , Bin Ling1,4 , Wenhui Wang1,4 , Dingqing Feng1,*, Jing Liang1,* 1Department of Obstetrics and Gynecology, China-Japan Friendship Hospital, 100029 Beijing, China 2VIP Department, National Cancer Center/National Clinical Research Center for Cancer/Cancer Hospital, Chinese Academy of Medical Sciences and Peking Union Medical College, 100021 Beijing, China 3Center for Reproductive Medicine and Prenatal Diagnosis, The First Affiliated Hospital of University of Science and Technology of China, 230001 Hefei, Anhui, China 4Graduate School of Peking Union Medical College, Chinese Academy of Medical Sciences, 100730 Beijing, China *Correspondence: dingqing_feng@aliyun.com (Dingqing Feng); jacyliang@sina.com (Jing Liang) †These authors contributed equally. Academic Editor: Michael H. Dahan Submitted: 12 November 2021 Revised: 19 February 2022 Accepted: 28 February 2022 Published: 18 May 2022 Abstract Background: TNF-α has a wide range of functions in processes within the ovaries, including follicular development, ovarian hormones synthesis, ovulation, granulosa cell necroptosis, and follicular atresia. Innate immune response in the ovaries is the main source of TNF-α. The purpose of this study is to explore the dose- and time-dependent interactions between TNF-α and human granulosa cells. Methods: The follicular fluids (FF) were obtained from infertile patients undergoing IVF with PCOS (n = 20) and healthy women (n = 19). Granulosa cells were cultured and treated with varying concentration of recombinant human TNF-α. Cell viability was detected by CCK-8 method, the levels of TNF-α, estradiol, and inhibin were measured with ELISA, and cell apoptosis was detected by flow cytometry. Q-PCR and western-blot were used to evaluate the expression of TNFR1/2, P450, Inhibin α/βA, and caspase 3. Results: The level of TNF-α increased gradually with the development of follicles and was remarkably higher in the FF of PCOS patients than in that of healthy women. Treatment with TNF-α as low as 0.2 ng/mL promoted the proliferation of granulosa cells, thus upregulating P450 aromatase and inhibin A. On the other hand, a high concentration of TNF-α (0.4, 0.8, 2, and 8 ng/mL) showed dose- and time-dependent inhibitory effects on cell viability and E2 and inhibin A secretion. However, no marked differences in the expression of the two types of TNFR were observed in TNF-α-treated granulosa cells. Conclusions: The data presented here verify a biphasic effect of TNF-α on human ovarian endocrine and follicular development. Keywords: TNF-α; TNF-α receptor; granulosa cell; P450 aromatase; inhibin A