Fertility preservation is a common concern for male cancer survivors of reproductive age. However, except for testicular tissue cryopreservation, which is not very effective, there is no feasible and precise therapy capable of protecting spermatogenesis for prepubertal boys before or during gonadotoxic treatment. This study aims to investigate the effects of inhibiting necroptosis of spermatogonial stem cell (SSC) in fertility preservation. Male mice 12 weeks of age were used to establish gonadotoxicity with two intraperitoneal injections of busulfan at a total dose of 40 mg kg−1. The mouse model and the primary cultured mouse SSCs were used to characterize the relationship between necroptosis of SSC and gonadotoxicity. Meanwhile, the effects of an inhibitor of necroptosis pathway, RIPA-56, were observed on day 36 in the mouse model of busulfan-induced gonadotoxicity. We found that the number of SSCs was decreased, but the level of necroptosis was upregulated on day 18 after busulfan treatment in testes from gonadotoxic mice. Further experiments in primary cultured cells showed that the necroptosis caused cell death in busulfan-treated SSCs and could be inhibited by RIPA-56. After suppressing the necroptosis of SSCs, the busulfan-induced mice had a decreased loss of spermatogenic cells as shown by histology and an increased Johnsen's score. Moreover, the quantities of SSCs and epididymal spermatozoa were restored after intervention with RIPA-56, indicating a series of beneficial effects by targeting the necroptosis of SSCs in mice undergoing busulfan treatment. In conclusion, our findings reveal that the necroptosis of SSCs plays a critical role in busulfan-induced gonadotoxicity and may be a potential target for male fertility preservation.
Get full access to this article
View all access options for this article.
References
1.
BrayF, FerlayJ, SoerjomataramI, SiegelRL, TorreLA and JemalA. (2018). Global cancer statistics 2018: GLOBOCAN estimates of incidence and mortality worldwide for 36 cancers in 185 countries. CA Cancer J Clin, 68:394–424.
2.
ZhaoH, JinL, LiY, ZhangC, WangR, LiY, HuangW, CuiC, ZhangH, et al (2019). Oncofertility: what can we do from bench to bedside?. Cancer Lett, 442:148–160.
3.
MossJL, ChoiAW, Fitzgerald KeeterMK and BranniganRE. (2016). Male adolescent fertility preservation. Fertil Steril, 105:267–273.
4.
FrydmanR and GrynbergM. (2016). Introduction: male fertility preservation: innovations and questions. Fertil Steril, 105:247–248.
5.
OktayK, HarveyBE, PartridgeAH, QuinnGP, ReineckeJ, TaylorHS, WallaceWH, WangET and LorenAW. (2018). Fertility preservation in patients with cancer: ASCO Clinical Practice Guideline Update. J Clin Oncol, 36:1994–2001.
6.
KenneyLB, AntalZ, GinsbergJP, HoppeBS, BoberSL, YuRN, ConstineLS, van SantenHM, SkinnerR and GreenDM. (2018). Improving male reproductive health after childhood, adolescent, and young adult cancer: progress and future directions for survivorship research. J Clin Oncol, 36:2160–2168.
7.
WynsC, CurabaM, VanabelleB, Van LangendoncktA and DonnezJ. (2010). Options for fertility preservation in prepubertal boys. Hum Reprod Update, 16:312–328.
8.
ShettyG and MeistrichML. (2005). Hormonal approaches to preservation and restoration of male fertility after cancer treatment. J Natl Cancer Inst Monogr. 36–39.
9.
SuomalainenL, HakalaJK, PentikainenV, OtalaM, ErkkilaK, PentikainenMO and DunkelL. (2003). Sphingosine-1-phosphate in inhibition of male germ cell apoptosis in the human testis. J Clin Endocrinol Metab, 88:5572–5579.
KotzurT, Benavides-GarciaR, MecklenburgJ, SanchezJR, ReillyM and HermannBP. (2017). Granulocyte colony-stimulating factor (G-CSF) promotes spermatogenic regeneration from surviving spermatogonia after high-dose alkylating chemotherapy. Reprod Biol Endocrinol, 15:7.
12.
Benavides-GarciaR, JoachimR, PinaNA, MutojiKN, ReillyMA and HermannBP. (2015). Granulocyte colony-stimulating factor prevents loss of spermatogenesis after sterilizing busulfan chemotherapy. Fertil Steril, 103:270–280.e8.
13.
CarmelyA, MeirowD, PeretzA, AlbeckM, BartoovB and SredniB. (2009). Protective effect of the immunomodulator AS101 against cyclophosphamide-induced testicular damage in mice. Hum Reprod, 24:1322–1329.
14.
NeuhausN and SchlattS. (2019). Stem cell-based options to preserve male fertility. Science, 363:1283–1284.
15.
FayomiAP, PetersK, SukhwaniM, Valli-PulaskiH, ShettyG, MeistrichML, HouserL, RobertsonN, RobertsV, et al. (2019). Autologous grafting of cryopreserved prepubertal rhesus testis produces sperm and offspring. Science, 363:1314–1319.
16.
SkinnerR, MulderRL, KremerLC, HudsonMM, ConstineLS, BardiE, BoekhoutA, Borgmann-StaudtA, BrownMC, et al. (2017). Recommendations for gonadotoxicity surveillance in male childhood, adolescent, and young adult cancer survivors: a report from the International Late Effects of Childhood Cancer Guideline Harmonization Group in collaboration with the PanCareSurFup Consortium. Lancet Oncol, 18:e75–e90.
17.
XieY, DengCC, OuyangB, LvLY, YaoJH, ZhangC, ChenHC, LiXY, SunXZ, DengCH and LiuGH. (2019). Establishing a nonlethal and efficient mouse model of male gonadotoxicity by intraperitoneal busulfan injection. Asian J Androl. DOI: 10.4103/aja.aja_41_19
ShanB, PanH, NajafovA and YuanJ. (2018). Necroptosis in development and diseases. Genes Dev, 32:327–340.
20.
WeinlichR, OberstA, BeereHM and GreenDR. (2017). Necroptosis in development, inflammation and disease. Nat Rev Mol Cell Biol, 18:127–136.
21.
LiD, MengL, XuT, SuY, LiuX, ZhangZ and WangX. (2017). RIPK1-RIPK3-MLKL-dependent necrosis promotes the aging of mouse male reproductive system. Elife, 6. DOI: 10.7554/eLife.27692
22.
de Rooij DG. (2017). The nature and dynamics of spermatogonial stem cells. Development, 144:3022–3030.
23.
MizunoK, HayashiY, KojimaY, KurokawaS, SasakiS and KohriK. (2007). Influence for testicular development and histological peculiarity in the testes of flutamide-induced cryptorchid rat model. Int J Urol, 14:67–72.
24.
WuY, ZhouH, FanX, ZhangY, ZhangM, WangY, XieZ, BaiM, YinQ, et al. (2015). Correction of a genetic disease by CRISPR-Cas9-mediated gene editing in mouse spermatogonial stem cells. Cell Res, 25:67–79.
25.
DongL, GulM, HildorfS, PorsSE, KristensenSG, HoffmannER, CortesD, ThorupJ and AndersenCY. (2019). Xeno-free propagation of spermatogonial stem cells from infant boys. Int J Mol Sci, 20. DOI: 10.3390/ijms20215390
26.
HouJ, NiuM, LiuL, ZhuZ, WangX, SunM, YuanQ, YangS, ZengW, et al. (2015). Establishment and characterization of human germline stem cell line with unlimited proliferation potentials and no tumor formation. Sci Rep, 5:16922.
27.
BaertY, BrayeA, StruijkRB, van PeltAM and GoossensE. (2015). Cryopreservation of testicular tissue before long-term testicular cell culture does not alter in vitro cell dynamics. Fertil Steril, 104:1244–1252.e1–e4.
28.
ValliH, SukhwaniM, DoveySL, PetersKA, DonohueJ, CastroCA, ChuT, MarshallGR and OrwigKE. (2014). Fluorescence- and magnetic-activated cell sorting strategies to isolate and enrich human spermatogonial stem cells. Fertil Steril, 102:566–580.e7.
29.
RabacaA, SousaM, AlvesMG, OliveiraPF and SaR. (2015). Novel Drug Therapies for Fertility preservation in men undergoing chemotherapy: clinical relevance of protector agents. Curr Med Chem, 22:3347–3369.
30.
PuyoS, MontaudonD and PourquierP. (2014). From old alkylating agents to new minor groove binders. Crit Rev Oncol Hematol, 89:43–61.
31.
BucciLR and MeistrichML. (1987). Effects of busulfan on murine spermatogenesis: cytotoxicity, sterility, sperm abnormalities, and dominant lethal mutations. Mutat Res, 176:259–268.
32.
LeeSB, KimJJ, HanSA, FanY, GuoLS, AzizK, NowsheenS, KimSS, ParkSY, et al. (2019). The AMPK-Parkin axis negatively regulates necroptosis and tumorigenesis by inhibiting the necrosome. Nat Cell Biol, 21:940–951.
33.
DondelingerY, HulpiauP, SaeysY, BertrandMJM and VandenabeeleP. (2016). An evolutionary perspective on the necroptotic pathway. Trends Cell Biol, 26:721–732.
34.
ZhouW and YuanJ. (2014). Necroptosis in health and diseases. Semin Cell Dev Biol, 35:14–23.
35.
XieY, LvL, YaoJ, ZhangC, ChenH, ChenW, LiangX, SunX, DengC and LiuG. (2019). Phosphorylated mixed lineage kinase domain-like protein in human seminal plasma: a potential novel biomarker of spermatogenic function. Andrologia, 51:e13310.
36.
XuD, JinT, ZhuH, ChenH, OfengeimD, ZouC, MifflinL, PanL, AminP, et al. (2018). TBK1 suppresses RIPK1-driven apoptosis and inflammation during development and in aging. Cell, 174:1477–1491.e19.
37.
CaccamoA, BrancaC, PirasIS, FerreiraE, HuentelmanMJ, LiangWS, ReadheadB, DudleyJT, SpangenbergEE, et al. (2017). Necroptosis activation in Alzheimer's disease. Nat Neurosci, 20:1236–1246.
38.
RenY, SuY, SunL, HeS, MengL, LiaoD, LiuX, MaY, LiuC, et al. (2017). Discovery of a highly potent, selective, and metabolically stable inhibitor of receptor-interacting protein 1 (RIP1) for the treatment of systemic inflammatory response syndrome. J Med Chem, 60:972–986.
39.
Kanatsu-ShinoharaM and ShinoharaT. (2013). Spermatogonial stem cell self-renewal and development. Annu Rev Cell Dev Biol, 29:163–187.
40.
GoossensE, Van SaenD and TournayeH. (2013). Spermatogonial stem cell preservation and transplantation: from research to clinic. Hum Reprod, 28:897–907.
41.
MenonS, RivesN, Mousset-SimeonN, SibertL, VannierJP, MazurierS, MasseL, DuchesneV and MaceB. (2009). Fertility preservation in adolescent males: experience over 22 years at Rouen University Hospital. Hum Reprod, 24:37–44.
42.
WallaceWH, AndersonRA and IrvineDS. (2005). Fertility preservation for young patients with cancer: who is at risk and what can be offered?. Lancet Oncol, 6:209–218.
43.
YangRF, LiuTH, ZhaoK and XiongCL. (2014). Enhancement of mouse germ cell-associated genes expression by injection of human umbilical cord mesenchymal stem cells into the testis of chemical-induced azoospermic mice. Asian J Androl, 16:698–704.
44.
OfengeimD, MazzitelliS, ItoY, DeWittJP, MifflinL, ZouC, DasS, AdiconisX, ChenH, et al. (2017). RIPK1 mediates a disease-associated microglial response in Alzheimer's disease. Proc Natl Acad Sci U S A, 114:E8788–E8797.
45.
OfengeimD, ItoY, NajafovA, ZhangY, ShanB, DeWittJP, YeJ, ZhangX, ChangA, et al. (2015). Activation of necroptosis in multiple sclerosis. Cell Rep, 10:1836–1849.
Supplementary Material
Please find the following supplemental material available below.
For Open Access articles published under a Creative Commons License, all supplemental material carries the same license as the article it is associated with.
For non-Open Access articles published, all supplemental material carries a non-exclusive license, and permission requests for re-use of supplemental material or any part of supplemental material shall be sent directly to the copyright owner as specified in the copyright notice associated with the article.
