Effects and potential mechanism of Ca<sup>2+</sup>/calmodulin‑dependent protein kinase II pathway inhibitor KN93 on the development of ovarian follicle

Yu,Jianjie; Xie,Xianguo; Ma,Yabo; Yang,Yi; Wang,Chao; Xia,Guoliang; Ding,Xiangbin; Liu,Xinfeng

doi:10.3892/ijmm.2022.5177

Effects and potential mechanism of Ca²⁺/calmodulin‑dependent protein kinase II pathway inhibitor KN93 on the development of ovarian follicle

Authors:
- Jianjie Yu
- Xianguo Xie
- Yabo Ma
- Yi Yang
- Chao Wang
- Guoliang Xia
- Xiangbin Ding
- Xinfeng Liu
View Affiliations

Affiliations: Key Laboratory of Ministry of Education for Conservation and Utilization of Special Biological Resources in Western China, College of Life Science, Ningxia University, Yinchuan, Ningxia Hui Autonomous Region 750021, P.R. China, Key Laboratory of Ministry of Education for Conservation and Utilization of Special Biological Resources in Western China, College of Life Science, Ningxia University, Yinchuan, Ningxia Hui Autonomous Region 750021, P.R.China, Tianjin Key Laboratory of Agricultural Animal Breeding and Healthy Husbandry, College of Animal Science and Veterinary Medicine, Tianjin Agricultural University, Tianjin 300384, P.R. China
Published online on: August 4, 2022 https://doi.org/10.3892/ijmm.2022.5177
Article Number: 121
Copyright: © Yu et al. This is an open access article distributed under the terms of Creative Commons Attribution License.

Metrics: Total Views: 0 (Spandidos Publications: | PMC Statistics: )

Metrics: Total PDF Downloads: 0 (Spandidos Publications: | PMC Statistics: )

Cited By (CrossRef): 0 citations Loading Articles...

Abstract

Adequate regulation of the speed of follicular development has been reported to prolong the reproductive life of the ovary. The aim of the present study was to assess the potential effects and mechanism of the Ca²⁺/calmodulin‑dependent protein kinase II (CaMKII) pathway on the development of ovarian follicle. In the present study, the expression of CaMKII was measured in the ovary of mice at different developmental stages by immunofluorescence, confirming that CaMKII has a role in follicular development. Subsequently, the 17.5 days post‑coitus (dpc) embryonic ovaries were collected and cultured with KN93 for 4 days in vitro. It was revealed that KN93 inhibited the development of follicles, where it reduced the expression levels of oocyte and granulosa cell markers DEAD‑box helicase 4 (DDX4) and forkhead box L2 (FOXL2). These results suggested that KN93 could delay follicular development. Proteomics technology was then used to find that 262 proteins of KN93 treated 17.5 dpc embryonic ovaries were significantly altered after in vitro culture. Bioinformatics analysis was used to analyze these altered proteins. In total, four important Kyoto Encyclopedia of Genes and Genome pathways, namely steroid biosynthesis, p53 signaling pathway and retinol metabolism and metabolic pathways, were particularly enriched. Further analysis revealed that the upregulated proteins NADP‑dependent steroid dehydrogenase‑like (Nsdhl), lanosterol synthase (Lss), farnesyl‑diphosphate farnesyltransferase 1 (Fdft1), cytochrome P450 family 51 family A member 1 (Cyp51a1), hydroxymethylglutaryl‑CoA synthase 1 (Hmgcs1), fatty acid synthase (Fasn) and dimethylallyltranstransferase (Fdps) were directly interacting with each other in the four enriched pathways. In summary, the potential mechanism of KN93 in slowing down follicular development most likely lies in its inhibitory effects on CaMKII, which upregulated the expression of Nsdhl, Lss, Fdft1, Cyp51a1, Hmgcs1, Fasn and Fdps. This downregulated the expression of oocyte and granulosa cell markers DDX4 and FOXL2 in the follicles, thereby delaying follicular development. Overall, these results provide novel insight into the potential mechanism by which KN93 and CaMKII can delay follicular development.

View Figures

View References

1	Shah JS, Sabouni R, Vaught KCC, Owen CM, Albertini DF and Segars JH: Biomechanics and mechanical signaling in the ovary: A systematic review. J Assist Reprod Genet. 35:1135–1148. 2018. View Article : Google Scholar : PubMed/NCBI
2	Green LJ and Shikanov A: In vitro culture methods of preantral follicles. Theriogenology. 86:229–238. 2016. View Article : Google Scholar : PubMed/NCBI
3	West ER, Shea LD and Woodruff TK: Engineering the follicle microenvironment. Semin Reprod Med. 25:287–299. 2007. View Article : Google Scholar : PubMed/NCBI
4	Simon LE, Kumar TR and Duncan FE: In vitro ovarian follicle growth: A comprehensive analysis of key protocol variables. Biol Reprod. 103:455–470. 2020. View Article : Google Scholar : PubMed/NCBI
5	Zhang T, He M, Zhao L, Qin S, Zhu Z, Du X, Zhou B, Yang Y, Liu X, Xia G, et al: HDAC6 regulates primordial follicle activation through mTOR signaling pathway. Cell Death Dis. 12:5592021. View Article : Google Scholar : PubMed/NCBI
6	Guo R and Pankhurst MW: Accelerated ovarian reserve depletion in female anti Müllerian hormone knockout mice has no effect on lifetime fertility†. Biol Reprod. 102:915–922. 2020. View Article : Google Scholar
7	Chen J, Liu W, Lee KF, Liu K, Wong BPC and Yeung WSB: Overexpression of Lin28a induces a primary ovarian insufficiency phenotype via facilitation of primordial follicle activation in mice. Mole Cell Endocrinol. 539:1114602022. View Article : Google Scholar
8	Zhang J, Yan L, Wang Y, Zhang S, Xu X, Dai Y, Zhao S, Li Z, Zhang Y, Xia G, et al: In vivo and in vitro activation of dormant primordial follicles by EGF treatment in mouse and human. Clin Transl Med. 10:e1822020. View Article : Google Scholar : PubMed/NCBI
9	Xu J and Gridley T: Notch2 is required in somatic cells for break-down of ovarian germ-cell nests and formation of primordial follicles. BMC Biol. 11:132013. View Article : Google Scholar
10	Zhang H and Liu K: Cellular and molecular regulation of the activation of mammalian primordial follicles: Somatic cells initiate follicle activation in adulthood. Hum Reprod Update. 21:779–786. 2015. View Article : Google Scholar : PubMed/NCBI
11	Huang K, Wang Y, Zhang T, He M, Sun G, Wen J, Yan H, Cai H, Yong C, Xia G and Wang C: JAK signaling regulates germline cyst breakdown and primordial follicle formation in mice. Biol Open. 7:bio0294702018.
12	Wang ZP, Mu XY, Guo M, Wang YJ, Teng Z, Mao GP, Niu WB, Feng LZ, Zhao LH and Xia GL: Transforming growth factor-β signaling participates in the maintenance of the primordial follicle pool in the mouse ovary. J Biol Chem. 289:8299–8311. 2014. View Article : Google Scholar : PubMed/NCBI
13	Komatsu K and Masubuchi S: Increased supply from blood vessels promotes the activation of dormant primordial follicles in mouse ovaries. J Reprod Dev. 66:105–113. 2020. View Article : Google Scholar : PubMed/NCBI
14	Hea Q, Chengb J and Wang Y: Chronic CaMKII inhibition reverses cardiac function and cardiac reserve in HF mice. Life Sci. 219:122–128. 2019. View Article : Google Scholar
15	Zhu Q, Hao L, Shen Q, Pan J, Liu W, Gong W, Hu L, Xiao W, Wang M, Liu X, et al: CaMK II Inhibition attenuates ROS dependent necroptosis in acinar cells and protects against acute pancreatitis in mice. Oxid Med Cell Longev. 17:41873982021.
16	Altobelli GG, Van Noorden S, Cimini D, Illario M, Sorriento1 D and Cimini V: Calcium/calmodulin-dependent kinases can regulate the TSH expression in the rat pituitary. J Endocrinol Invest. 44:2387–2394. 2021. View Article : Google Scholar : PubMed/NCBI
17	Hoch B, Haase H, Schulze W, Haqemann D, Morano I, Krause EG and Karczewski P: Differentiation-dependent expression of cardiac delta-CaMKII isoforms. J Cell Biochem. 68:259–268. 1998. View Article : Google Scholar : PubMed/NCBI
18	Medvedev S, Stein P and Schultz RM: Specificity of calcium/calmodulin-dependent protein kinases in mouse egg activation. Cell Cycle. 13:1482–1488. 2014. View Article : Google Scholar : PubMed/NCBI
19	Jin XL and O'Neill C: The presence and activation of two essential transcription factors (cAMP response element-binding protein and cAMP-dependent transcription factor ATF1) in the two-cell mouse embryo. Biol Reprod. 82:459–468. 2010. View Article : Google Scholar
20	Wang J, Xu X, Jia W, Zhao D, Boczek T, Gao Q, Wang Q, Fu Y, He M, Shi R, et al: Calcium-/calmodulin-dependent protein kinase II (CaMKII) inhibition induces learning and memory impairment and apoptosis. Oxid Med Cell Longev. 2021:46350542021. View Article : Google Scholar :
21	Johnson CN, Pattanayek R, Potet F, Rebbeck RT, Blackwell DJ, Nikolaienko R, Sequeira V, Meur RL, Radwański PB, Davis JP, et al: The CaMKII inhibitor KN93-calmodulin interaction and implications for calmodulin tuning of NaV1.5 and RyR2 function. Cell Calcium. 82:1020632019. View Article : Google Scholar :
22	Yang X, Wu N, Song L and Liu Z: Intrastriatal injections of KN-93 ameliorates levodopa-induced dyskinesia in a rat model of Parkinson's disease. Neuropsychiatr Dis Treat. 9:1213–1220. 2013.PubMed/NCBI
23	Liu Y, Liang Y, Hou B, Liu M, Yang X, Liu C, Zhang J, Zhang W, Ma Z and Gu X: The inhibitor of calcium/calmodulin-dependent protein kinase II KN93 attenuates bone cancer pain via inhibition of KIF17/NR2B traffiffifficking in mice. Pharmacol Biochem Behav. 124:19–26. 2014. View Article : Google Scholar : PubMed/NCBI
24	Edvinsson L, Povlsen GK, Ahnstedt H and Waldsee R: CaMKII inhibition with KN93 attenuates endothelin and serotonin receptor-mediated vasoconstriction and prevents subarachnoid hemorrhage-induced deficits in sensorimotor function. J Neuroinflammation. 11:2072014. View Article : Google Scholar : PubMed/NCBI
25	Chen W, An P, Quan XZ, Zhang J, Zhou ZY, Zou LP and Luo HS: Ca²⁺/calmodulin-dependent protein kinase II regulates colon cancer proliferation and migration via ERK1/2 and p38 pathways. World J Gastroenterol. 23:6111–6118. 2017. View Article : Google Scholar : PubMed/NCBI
26	Shin H, Han C, Labuz ML, Kim J, Kim J, Cho S, Gho YS, Takayama S and Parka J: High-yield isolation of extracellular vesicles using aqueous two-phase system. Sci Rep. 5:131032015. View Article : Google Scholar : PubMed/NCBI
27	He Y, Chen Q, Dai J, Cui Y, Zhang C, Wen X, Li J, Xiao Y, Peng X, Liu M, et al: Single-cell RNA-Seq reveals a highly coordinated transcriptional program in mouse germ cells during primordial follicle formation. Aging Cell. 20:e134242021. View Article : Google Scholar : PubMed/NCBI
28	Bazot M, Robert Y, Mestdagh P, Boudghène F and Rocourt N: Ovarian functional disorders. J Radiol. 81:1801–1818. 2000.In French.
29	Fortune JE, Rivera GM and Yang MY: Follicular development: The role of the follicular microenvironment in selection of the dominant follicle. Anim Reprod Sci. 82:109–126. 2004. View Article : Google Scholar : PubMed/NCBI
30	Sun L, Chen L, Jiang Y, Zhao Y, Wang F, Zheng X, Li C and Zhou X: Metabolomic profifiling of ovary in mice treated with FSH using ultra performance liquid chromatography/mass spectrometry. Biosci Rep. 38:BSR201809652018. View Article : Google Scholar
31	Hirshfifield AN: Development of follicles in the mammalian ovary. Int Rev Cytol. 124:43–101. 1991. View Article : Google Scholar
32	Li T, Liu X, Gong X, Qiukai E and Zhang X and Zhang X: microRNA 92b-3p regulate primordial follicle assembly by targeting TSC1 in neonatal mouse ovaries. Cell Cycle. 18:824–833. 2019. View Article : Google Scholar : PubMed/NCBI
33	Wang J, Ge W, Zhai QY, Liu JC, Sun XW, Liu WX, Li L, Lei CZ, Dyce PW, De Felici M and Shen W: Single-cell transcriptome landscape of ovarian cells during primordial follicle assembly in mice. PLoS Biol. 18:e30010252020. View Article : Google Scholar : PubMed/NCBI
34	Bristol-Gould SK, Kreeger PK, Selkirk CG, Kilen SM, Cook RW, Kipp JL, Shea LD, Mayo KE and Woodruff TE: Postnatal regulation of germ cells by activin: The establishment of the initial follicle pool. Dev Biol. 298:132–148. 2006. View Article : Google Scholar : PubMed/NCBI
35	Yin H, Kristensen SG, Jiang H, Rasmussen A and Andersen CY: Survival and growth of isolated pre-antral follicles from human ovarian medulla tissue during long-term 3D culture. Hum Reprod. 31:1531–1539. 2016. View Article : Google Scholar : PubMed/NCBI
36	Zhai J, Zhang J, Zhang L, Liu X, Deng W, Wang H, Zhang Z, Liu W, Chen B, Wu C, et al: Autotransplantation of the ovarian cortex after in vitro activation for infertility treatment: A shortened procedure. Hum Reprod. 36:2134–2147. 2021. View Article : Google Scholar : PubMed/NCBI
37	Song K, Ma W, Huang C, Ding J, Cui D and Zhang M: expression pattern of mouse vasa homologue (MVH) in the ovaries of C57BL/6 female mice. Med Sci Monit. 22:2656–2663. 2016. View Article : Google Scholar : PubMed/NCBI
38	Georges A, Auguste A, Bessière L, Vanet A, Todeschini AL and Veitia RA: FOXL2: A central transcription factor of the ovary. J Mol Endocrinol. 52:R17–R33. 2013. View Article : Google Scholar : PubMed/NCBI
39	Su Y, Sugiura K, Wigglesworth K, O'Brien MJ, Affourtit JP, Pangas SA, Matzuk MM and Eppig JJ: Oocyte regulation of metabolic cooperativity between mouse cumulus cells and oocytes: BMP15 and GDF9 control cholesterol biosynthesis in cumulus cells. Development. 135:111–121. 2008. View Article : Google Scholar
40	Ershov P, Kaluzhskiy L, Mezentsev Y, Yablokov E, Gnedenko O and Ivanov A: Enzymes in the cholesterol synthesis pathway: Interactomics in the cancer context. Biomedicines. 9:8952021. View Article : Google Scholar : PubMed/NCBI
41	Nakamura T, Iwase A, Bayasula B, Nagatomo Y, Kondo M, Nakahara T, Takikawa S, Goto M, Kotani T, Kiyono T and Kikkawa F: CYP51A1 induced by growth differentiation factor 9 and follicle-stimulating hormone in granulosa cells is a possible predictor for unfertilization. Reprod Sci. 22:377–384. 2015. View Article : Google Scholar :
42	Jin Y, Chen Z, Dong J, Wang B, Fan S, Yang X and Cui M: SREBP1/FASN/cholesterol axis facilitates radioresistance in colorectal cancer. FEBS Open Bio. 11:1343–1352. 2021. View Article : Google Scholar : PubMed/NCBI
43	Yang X, Zhao Z, Fan Q, Li H, Liu LZC and Liang X: Cholesterol metabolism is decreased in patients with diminished ovarian reserve. Reprod Biomed Online. 44:185–192. 2022. View Article : Google Scholar