Effect of bacterial endotoxin lipopolysaccharide treatment on duck Leydig cells

Yongcong Lao; Hongjia Ouyang; Xuebing Huang; Yunmao Huang

doi:10.21451/1984-3143-AR2019-0002

Original Article

Effect of bacterial endotoxin lipopolysaccharide treatment on duck Leydig cells

Yongcong Lao; Hongjia Ouyang; Xuebing Huang; Yunmao Huang

http://dx.doi.org/10.21451/1984-3143-AR2019-0002 Anim Reprod, vol.16, n4, p.871-879, 2019

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Abstract

Abstract: This study aimed to investigate the effects of bacterial endotoxin lipopolysaccharide (LPS) on hormone production and gene expression in duck Leydig cells and its underlying mechanisms. Leydig cells were collected from 200-day-old mallard ducks and divided into five treatment groups (0, 50, 100, 200, and 400 ng/mL LPS). After treatment with LPS for 6, 12, 24, and 48 h, testosterone, activin, and inhibin levels in the cell supernatants were determined using enzyme-linked immunosorbent assay (ELISA) kits. The expression levels of testosterone synthesis-related genes, including steroidogenic acute regulatory protein (StAR), 3-beta-hydroxysteroid dehydrogenase (3β-HSD), and cytochrome P450 aromatase (P450arom), and reproductive-related genes, including gonadotropin-inhibitory hormone receptor (GnIHR), follicle stimulating hormone receptor (FSHR), and luteinizing hormone receptor (LHR) were detected using quantitative real-time polymerase chain reaction (qRT-PCR). We successfully isolated and cultured duck Leydig cells with cell purity above 90%. Compared with the control group, the levels of testosterone, activin, and inhibin secreted in Leydig cells decreased gradually with increasing LPS concentration. After treatment with LPS, the expression of StAR and 3β-HSD genes in Leydig cells was upregulated at 12 h, and that of GnIHR was upregulated at 24 h; whereas the expression of FSHR and LHR was reduced at 24 h. This study indicates that LPS can inhibit the secretion of hormones and regulate the expression of related genes in duck Leydig cells.

Keywords

duck, Leydig cells, LPS, testosterone, gene expression

References

Anthoney N, Foldi I, Hidalgo A. Toll and Toll-like receptor signalling in development. Development. 2018;145(9):dev156018. http://dx.doi.org/10.1242/dev.156018. PMid:29695493.

Bosmann HB, Hales KH, Li X, Liu Z, Stocco DM, Hales DB. Acute in vivo inhibition of testosterone by endotoxin parallels loss of steroidogenic acute regulatory (StAR) protein in Leydig cells. Endocrinology. 1996;137(10):4522-5. http://dx.doi.org/10.1210/endo.137.10.8828518. PMid:8828518.

Brecchia G, Cardinali R, Mourvaki E, Collodel G, Moretti E, Dal Bosco A, Castellini C. Short- and long-term effects of lipopolysaccharide-induced inflammation on rabbit sperm quality. Anim Reprod Sci. 2010;118(2-4):310-6. http://dx.doi.org/10.1016/j.anireprosci.2009.06.016. PMid:19625146.

Carmody RJ, Chen YH. Nuclear factor-kappaB: activation and regulation during toll-like receptor signaling. Cell Mol Immunol. 2007;4(1):31-41. PMid:17349209.

Chen H, Cangello D, Benson S, Folmer J, Zhu H, Trush MA, Zirkin BR. Age-related increase in mitochondrial superoxide generation in the testosterone-producing cells of Brown Norway rat testes: relationship to reduced steroidogenic function? Exp Gerontol. 2001;36(8):1361-73. http://dx.doi.org/10.1016/S0531-5565(01)00118-8. PMid:11602210.

Collodel G, Castellini C, Del Vecchio MT, Cardinali R, Geminiani M, Rossi B, Spreafico A, Moretti E. Effect of a bacterial lipopolysaccharide treatment on rabbit testis and ejaculated sperm. Reprod Domest Anim. 2012;47(3):372-8.

Drosdowsky M, Menon KM, Forchielli E, Dorfman RI. Requirements of the cholesterol side-chain-cleaving enzyme system of rat-testis mitochondria. Biochim Biophys Acta. 1965;104(1):229-36. http://dx.doi.org/10.1016/0304-4165(65)90240-0. PMid:4378799.

Grzywacz FW, Chen H, Allegretti J, Zirkin BR. Does age-associated reduced Leydig cell testosterone production in Brown Norway rats result from under-stimulation by luteinizing hormone? J Androl. 1998;19(5):625-30. PMid:9796624.

Hales DB, Xiong Y, Tur-Kaspa I. The role of cytokines in the regulation of Leydig cell P450c17 gene expression. J Steroid Biochem Mol Biol. 1992;43(8):907-14. http://dx.doi.org/10.1016/0960-0760(92)90318-D. PMid:22217835.

Hedger MP, Eddy EM. Monoclonal antibodies against rat Leydig cell surface antigens. Biol Reprod. 1986;35(5):1309-19. http://dx.doi.org/10.1095/biolreprod35.5.1309. PMid:3548835.

Hedger MP, Eddy EM. The heterogeneity of isolated adult rat Leydig cells separated on Percoll density gradients: an immunological, cytochemical, and functional analysis. Endocrinology. 1987;121(5):1824-38. http://dx.doi.org/10.1210/endo-121-5-1824. PMid:2822376.

Kajihara T, Okagaki R, Ishihara O. LPS-induced transient testicular dysfunction accompanied by apoptosis of testicular germ cells in mice. Med Mol Morphol. 2006;39(4):203-8. http://dx.doi.org/10.1007/s00795-006-0334-7. PMid:17187183.

Kakar SS, Musgrove LC, Devor DC, Sellers JC, Neill JD. Cloning, sequencing, and expression of human gonadotropin releasing hormone (GnRH) receptor. Biochem Biophys Res Commun. 1992;189(1):289-95. http://dx.doi.org/10.1016/0006-291X(92)91556-6. PMid:1333190.

Keeney DS, Mendis-Handagama SM, Zirkin BR, Ewing LL. Effect of long term deprivation of luteinizing hormone on Leydig cell volume, Leydig cell number, and steroidogenic capacity of the rat testis. Endocrinology. 1988;123(6):2906-15. http://dx.doi.org/10.1210/endo-123-6-2906. PMid:3197648.

Lee MS, Kim YJ. Signaling pathways downstream of pattern-recognition receptors and their cross talk. Annu Rev Biochem. 2007;76(1):447-80. http://dx.doi.org/10.1146/annurev.biochem.76.060605.122847. PMid:17328678.

Livak KJ, Schmittgen TD. Analysis of relative gene expression data using real-time quantitative PCR and the 2(-Delta Delta C(T)) Method. Methods. 2001;25(4):402-8. http://dx.doi.org/10.1006/meth.2001.1262. PMid:11846609.

McLachlan RI, Robertson DM, De Kretser DM, Burger HG. Advances in the physiology of inhibin and inhibin-related peptides. Clin Endocrinol (Oxf). 1988;29(1):77-112. http://dx.doi.org/10.1111/j.1365-2265.1988.tb00251.x. PMid:3073881.

Metukuri MR, Reddy CMT, Reddy PRK, Reddanna P. Bacterial LPS mediated acute inflammation-induced spermatogenic failure in rats: role of stress response proteins and mitochondrial dysfunction. Inflammation. 2010;33(4):235-43. http://dx.doi.org/10.1007/s10753-009-9177-4. PMid:20087639.

Molenaar R, Rommerts FF, van der Molen HJ. Non-specific esterase: a specific and useful marker enzyme for Leydig cells from mature rats. J Endocrinol. 1986;108(3):329-34, NP. http://dx.doi.org/10.1677/joe.0.1080329. PMid:3457892.

Morizane S, Kajita A, Mizuno K, Takiguchi T, Iwatsuki K. Toll-like receptor signalling induces the expression of serum amyloid A in epidermal keratinocytes and dermal fibroblasts. Clin Exp Dermatol. 2019;44(1):40-6. http://dx.doi.org/10.1111/ced.13604. PMid:29770468.

Nagata Y, Homma H, Matsumoto M, Imai K. Stimulation of steroidogenic acute regulatory protein (StAR) gene expression by D-aspartate in rat Leydig cells. FEBS Lett. 1999;454(3):317-20. http://dx.doi.org/10.1016/S0014-5793(99)00840-6. PMid:10431830.

Ni Y, Zhou Y, Lu L, Grossmann R, Zhao R. Developmental changes of FSH-R, LH-R, ER-beta and GnRH-I expression in the ovary of prepubertal ducks (Anas platyrhynchos). Anim Reprod Sci. 2007;100(3-4):318-28. http://dx.doi.org/10.1016/j.anireprosci.2006.08.012. PMid:16989964.

O’Bryan MK, Schlatt S, Phillips DJ, de Kretser DM, Hedger MP. Bacterial lipopolysaccharide-induced inflammation compromises testicular function at multiple levels in vivo. Endocrinology. 2000;141(1):238-46. http://dx.doi.org/10.1210/endo.141.1.7240. PMid:10614644.

O’Neill LA, Golenbock D, Bowie AG. The history of Toll-like receptors - redefining innate immunity. Nat Rev Immunol. 2013;13(6):453-60. http://dx.doi.org/10.1038/nri3446. PMid:23681101.

O’Shaughnessy PJ. Steroidogenic enzyme activity in the hypogonadal (hpg) mouse testis and effect of treatment with luteinizing hormone. J Steroid Biochem Mol Biol. 1991;39(6):921-8. http://dx.doi.org/10.1016/0960-0760(91)90350-E. PMid:1751391.

Raucci F, D’Aniello A, Di Fiore MM. Stimulation of androgen production by D-aspartate through the enhancement of StAR, P450scc and 3beta-HSD mRNA levels in vivo rat testis and in culture of immature rat Leydig cells. Steroids. 2014;84:103-10. http://dx.doi.org/10.1016/j.steroids.2014.03.016. PMid:24713504.

Reddy MM, Mahipal SVK, Subhashini J, Reddy MC, Roy KR, Reddy GV, Reddy PRK, Reddanna P. Bacterial lipopolysaccharide-induced oxidative stress in the impairment of steroidogenesis and spermatogenesis in rats. Reprod Toxicol. 2006;22(3):493-500. http://dx.doi.org/10.1016/j.reprotox.2006.03.003. PMid:16644180.

Sadasivam M, Ramatchandirin B, Ayyanar A, Prahalathan C. Bacterial lipopolysaccharide differently modulates steroidogenic enzymes gene expressions in the brain and testis in rats. Neurosci Res. 2014;83:81-8. http://dx.doi.org/10.1016/j.neures.2014.02.011. PMid:24594480.

Sharova VS, Izvol Skaya MS, Tillet Y, Voronova SN, Zakharova LA. The morphogenetic effect of bacterial endotoxin lipopolysaccharide on the functioning of the reproductive system in rats. Dokl Biol Sci. 2014;455(1):79-82. http://dx.doi.org/10.1134/S0012496614020100. PMid:24795176.

Shi L, Song R, Yao X, Ren Y. Effects of selenium on the proliferation, apoptosis and testosterone production of sheep Leydig cells in vitro. Theriogenology. 2017;93:24-32. http://dx.doi.org/10.1016/j.theriogenology.2017.01.022. PMid:28257863.

Shimizu T, Ishizawa S, Magata F, Kobayashi M, Fricke PM, Miyamoto A. Involvement of lipopolysaccharide in ovarian cystic follicles in dairy cow: expressions of LPS receptors and steroidogenesis-related genes in follicular cells of cystic follicles. Anim Reprod Sci. 2018;195:89-95. http://dx.doi.org/10.1016/j.anireprosci.2018.05.010. PMid:29843942.

Toocheck C, Clister T, Shupe J, Crum C, Ravindranathan P, Lee TK, Ahn JM, Raj GV, Sukhwani M, Orwig KE, Walker WH. Mouse Spermatogenesis Requires Classical and Nonclassical Testosterone Signaling. Biol Reprod. 2016;94(1):11. http://dx.doi.org/10.1095/biolreprod.115.132068. PMid:26607719.

Topo E, Soricelli A, D’Aniello A, Ronsini S, D’Aniello G. The role and molecular mechanism of D-aspartic acid in the release and synthesis of LH and testosterone in humans and rats. Reprod Biol Endocrinol. 2009;7(1):120. http://dx.doi.org/10.1186/1477-7827-7-120. PMid:19860889.

Vale W, Rivier C, Hsueh A, Campen C, Meunier H, Bicsak T, Vaughan J, Corrigan A, Bardin W, Sawchenko P, Et A. Chemical and biological characterization of the inhibin family of protein hormones. Recent Prog Horm Res. 1988;44:1-34. PMid:3064204.

Vijay K. Toll-like receptors in immunity and inflammatory diseases: Past, present, and future. Int Immunopharmacol. 2018;59:391-412. http://dx.doi.org/10.1016/j.intimp.2018.03.002. PMid:29730580.

Wang C, Sinha Hikim AP, Lue YH, Leung A, Baravarian S, Swerdloff RS. Reproductive aging in the Brown Norway rat is characterized by accelerated germ cell apoptosis and is not altered by luteinizing hormone replacement. J Androl. 1999;20(4):509-18. PMid:10452595.

Wu C, Su Z, Lin M, Ou J, Zhao W, Cui J, Wang RF. NLRP11 attenuates Toll-like receptor signalling by targeting TRAF6 for degradation via the ubiquitin ligase RNF19A. Nat Commun. 2017;8(1):1977. http://dx.doi.org/10.1038/s41467-017-02073-3. PMid:29215004.

Yang JY, Zhang YF, Li YX, Guan GP, Kong XF, Liang AM, Ma KW, Da Li G, Bai XF. Effects of T-2 toxin on the regulation of steroidogenesis in mouse Leydig cells. Toxicol Ind Health. 2016;32(10):1801-7. http://dx.doi.org/10.1177/0748233715590516. PMid:26085520.

Ying SY. Inhibins, activins, and follistatins: gonadal proteins modulating the secretion of follicle-stimulating hormone. Endocr Rev. 1988;9(2):267-93. http://dx.doi.org/10.1210/edrv-9-2-267. PMid:3136011.

Effect of bacterial endotoxin lipopolysaccharide treatment on duck Leydig cells

Yongcong Lao; Hongjia Ouyang; Xuebing Huang; Yunmao Huang

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