Smooth muscle AKG/OXGR1 signaling regulates epididymal fluid acid–base balance and sperm maturation

Abstract

Infertility is a global concern attributed to genetic defects, lifestyle, nutrition, and any other factors that affect the local metabolism and niche microenvironment of the reproductive system. 2-Oxoglutarate receptor 1 (OXGR1) is abundantly expressed in the testis; however, its cellular distribution and biological function of OXGR1 in the male reproductive system remain unclear. In the current study, we demonstrated that OXGR1 is primarily expressed in epididymal smooth muscle cells (SMCs). Aging and heat stress significantly reduced OXGR1 expression in the epididymis. Using OXGR1 global knockout and epididymal-specific OXGR1 knockdown models, we revealed that OXGR1 is essential for epididymal sperm maturation and fluid acid–base balance. Supplementation of α-ketoglutaric acid (AKG), the endogenous ligand of OXGR1, effectively reversed epididymal sperm maturation disorders caused by aging and heat stress. Furthermore, in vitro studies showed that AKG markedly stimulated the release of instantaneous intracellular calcium from epididymal SMCs and substantially reduced the pH_i value in the epididymal SMCs via OXGR1. Mechanistically, we discovered that AKG/OXGR1 considerably increased the expression of Na⁺/HCO₃⁻ cotransporter (NBCe1) mRNA in the epididymal SMCs, mediated by intracellular calcium signaling. The local AKG/OXGR1 system changed the epididymal fluid pH value and HCO₃⁻ concentration, thereby regulating sperm maturation via intracellular calcium signaling and NBCe1 mRNA expression. This study for the first time reveals the crucial role of OXGR1 in male fertility and sheds light on the applicability of metabolic intermediates in the nutritional intervention of reproduction.

Keywords

AKG / OXGR1 / epididymis / acid-base balance / sperm maturation

References

[1] Gunes S, Hekim GNT, Arslan MA,, et al. Effects of aging on the male reproductive system. J Assist Reprod Genet 2016; 33: 441- 54.

[2] Zhou Y, Wu F, Zhang M,, et al. EMC10 governs male fertility via maintaining sperm ion balance. J Mol Cell Biol 2018; 10: 503- 14.

[3] Pal P, Biswas S, Mukhopadhyay PK. Molecular perspective concerning fluoride and arsenic mediated disorders on epididyma maturation of spermatozoa: A concise review. Hum Exp Toxicol 2021; 40: 2025- 38.

[4] Aitken RJ, Nixon B, Lin M,, et al. Proteomic changes in mammalian spermatozoa during epididymal maturation. Asian J Androl 2007; 9: 554- 64.

[5] Zitzmann M. Effects of age on male fertility. Best Pract Res Clin Endocrinol Metab 2013; 27: 617- 28.

[6] Bisht S, Faiq M, Tolahunase M,, et al. Oxidative stress and male infertility. Nat Rev Urol 2017; 14: 470- 85.

[7] Salas-Huetos A, Bulló M, Salas-Salvadó J. Dietary patterns, foods and nutrients in male fertility parameters and fecundability: a systematic review of observational studies. Hum Reprod Update 2017; 23: 371- 89.

[8] Ilacqua A, Izzo G, Emerenziani GP,, et al. Lifestyle and fertility: the influence of stress and quality of life on male fertility. Reprod Biol Endocrinol 2018; 16: 115.

[9] Diehl J, Gries B, Pfeil U,, et al. Expression and localization of GPR91 and GPR99 in murine organs. Cell Tissue Res 2016; 364: 245- 62.

[10] Tokonami N, Morla L, Centeno G,, et al. α-Ketoglutarate regulates acid-base balance through an intrarenal paracrine mechanism. J Clin Invest 2013; 123: 3166- 71.

[11] Guerrero A, Visniauskas B, Cárdenas P,, et al. α-Ketoglutarate upregulates collecting duct (pro)renin receptor expression, tubular angiotensin II formation, and Na⁺ reabsorption during high glucose conditions. Front Cardiovasc Med 2021; 8: 644797.

[12] Yuan Y, Xu P, Jiang Q,, et al. Exercise-induced α-ketoglutaric acid stimulates muscle hypertrophy and fat loss through OXGR1- dependent adrenal activation. EMBO J 2020; 39: e103304.

[13] Covarrubias AJ, Perrone R, Grozio A,, et al. NAD metabolism and its roles in cellular processes during ageing. Nat Rev Mol Cell Biol 2021; 22: 119- 41.

[14] Harrison AP, Pierzynowski SG. Biological effects of 2-oxoglutarate with particular emphasis on the regulation of protein, mineral and lipid absorption/metabolism, muscle performance, kidney function, bone formation and cancerogenesis, all viewed from a healthy ageing perspective state of the art--review article. J Physiol Pharmacol 2008; 59: 91- 106.

[15] Gonzalez-Rivas PA, Chauhan SS, Ha M,, et al. Effects of heat stress on animal physiology, metabolism, and meat quality: a review. Meat Sci 2020; 162: 108025.

[16] Purandhar K, Jena PK, Prajapati B,, et al. Understanding the role of heat shock protein isoforms in male fertility, aging and apoptosis. World J Mens Health 2014; 32: 123- 32.

[17] Sullivan R, Mieusset R. The human epididymis: its function in sperm maturation. Hum Reprod Update 2016; 22: 574- 87.

[18] Visconti PE, Bailey JL, Moore GD,, et al. Capacitation of mouse spermatozoa. I. Correlation between the capacitation state and protein tyrosine phosphorylation. Development 1995; 121: 1129- 37.

[19] Hu J, Merriner DJ, O’connor AE,, et al. Epididymal cysteine-rich secretory proteins are required for epididymal sperm maturation and optimal sperm function. Mol Hum Reprod 2018; 24: 111- 22.

[20] Su Y, Wang T, Wu N,, et al. Alpha-ketoglutarate extends lifespan by inhibiting mTOR and activating AMPK. Aging (Albany NY) 2019; 11: 4183- 97.

[21] Li S, Fu C, Zhao Y,, et al. Intervention with -ketoglutarate ameliorates colitis-related colorectal carcinoma via modulation of the gut microbiome. Biomed Res Int 2019; 2019: 8020785.

[22] Liu PS, Wang H, Li X,, et al. α-Ketoglutarate orchestrates macrophage activation through metabolic and epigenetic reprogramming. Nat Immunol 2017; 18: 985- 94.

[23] Liu M, Chen Y, Wang S,, et al. α-Ketoglutarate modulates macrophage polarization through regulation of PPARγ transcription and mTORC1/p70S6K pathway to ameliorate ALI/ARDS. Shock 2020; 53: 103- 13.

[24] Gao DD, Xu JW, Qin WB,, et al. Cellular mechanism underlying hydrogen sulfide mediated epithelial K secretion in rat epididymis. Front Physiol 2018; 9: 1886.

[25] Pastor-Soler N, Piétrement C, Breton S. Role of acid/base transporters in the male reproductive tract and potential consequences of their malfunction. Physiology (Bethesda) 2005; 20: 417- 28.

[26] Althoff TF, Offermanns S. G-protein-mediated signaling in vascular smooth muscle cells—implications for vascular disease. J Mol Med (Berl) 2015; 93: 973- 81.

[27] Morgado M, Cairrão E, Santos-Silva AJ,, et al. Cyclic nucleotide-dependent relaxation pathways in vascular smooth muscle. Cell Mol Life Sci 2012; 69: 247- 66.

[28] Kanaoka Y, Maekawa A, Austen KF. Identification of GPR99 protein as a potential third cysteinyl leukotriene receptor with a preference for leukotriene E4 ligand. J Biol Chem 2013; 288: 10967- 72.

[29] Shirasaki H, Kanaizumi E, Himi T. Expression and localization of GPR99 in human nasal mucosa. Auris Nasus Larynx 2017; 44: 162- 7.

[30] Bize I, Santander G. Epinephrine decreases the potassium requirements of hamster sperm capacitation: furosemide blocks the effect of epinephrine. J Exp Zool 1985; 235: 261- 7.

[31] Rafiee B, Bagher Tabei SM. The effect of N-acetyl cysteine consumption on men with abnormal sperm parameters due to positive history of COVID-19 in the last three months. Arch Ital Urol Androl 2021; 93: 465- 7.

[32] Ericsson RJ, Buthala DA, Norland JF. Fertilization of rabbit ova in vitro by sperm with adsorbed Sendai virus. Science 1971; 173: 54- 5.

[33] Mietens A, Tasch S, Stammler A,, et al. Time-lapse imaging as a tool to investigate contractility of the epididymal duct—effects of cGMP signaling. PLoS One 2014; 9: e92603.

[34] Borges CS, Kiguti LRA, Pupo AS,, et al. In vitro contraction of isolated cauda epididymal duct smooth muscle as a complimentary approach to physiological, pathological, toxicological, and pharmacological studies on epididymal function. Methods Mol Biol 2021; 2240: 65- 76.

[35] Mueller A, Kiguti LRA, Silva E JR, et al. Contractile effects of serotonin (5-HT) in the rat cauda epididymis: expression and functional characterization of 5-HT receptors. J Pharmacol Exp Ther 2019; 369: 98- 106.

[36] Hiroshige T, Uemura KI, Hirashima S,, et al. Morphological analysis of interstitial cells in murine epididymis using light microscopy and transmission electron microscopy. Acta Histochem 2021; 123: 151761.

[37] Liu Y, Wang DK, Chen LM. The physiology of bicarbonate transporters in mammalian reproduction. Biol Reprod 2012; 86: 99.

[38] Zoukhri D, Hodges RR, Sergheraert C,, et al. Cholinergic-induced Ca2+ elevation in rat lacrimal gland acini is negatively modulated by PKCdelta and PKCepsilon. Invest Ophthalmol Vis Sci 2000; 41: 386- 92.

[39] Perry C, Quissell DO, Reyland ME,, et al. Electrogenic NBCe1 (SLC4A4), but not electroneutral NBCn1 (SLC4A7), cotransporter undergoes cholinergic-stimulated endocytosis in salivary ParC5 cells. Am J Physiol Cell Physiol 2008; 295: C1385- 98.

[40] Handlogten ME, Osis G, Lee HW, et al. NBCe1 expression is required for normal renal ammonia metabolism. Am J Physiol Renal Physiol 2015; 309: F658- 66.

[41] Almeida S, Rato L, Sousa M, et al. Fertility and sperm quality in the aging male. Curr Pharm Des 2017; 23: 4429- 37.

[42] Qin DZ, Cai H, He C,, et al. Melatonin relieves heat-induced spermatocyte apoptosis in mouse testes by inhibition of ATF6 and PERK signaling pathways. Zool Res 2021; 42: 514- 24.

[43] Demidenko O, Barardo D, Budovskii V,, et al. Rejuvant®, a potential life-extending compound formulation with alpha-ketoglutarate and vitamins, conferred an average 8 year reduction in biological aging, after an average of 7 months of use, in the TruAge DNA methylation test. Aging (Albany NY) 2021; 13: 24485- 99.

[44] Asadi Shahmirzadi A, Edgar D, Liao C-Y,, et al. Alphaketoglutarate, an endogenous metabolite, extends lifespan and compresses morbidity in aging mice. Cell Metab 2020; 32: 447- 56.

[45] Gomes AP, Ilter D, Low V, et al. Age-induced accumulation of methylmalonic acid promotes tumour progression. Nature 2020; 585: 283- 7.

[46] Li Y, Chen L, Zhao W,, et al. Food reward depends on TLR4 activation in dopaminergic neurons. Pharmacol Res 2021; 169: 105659.

[47] Yin Q, Shen J, Wan X,, et al. Impaired sperm maturation in conditional Lcn6 knockout mice. Biol Reprod 2018; 98: 28- 41.

[48] Wang T, Yin Q, Ma X,, et al. Ccdc87 is critical for sperm function and male fertility. Biol Reprod 2018; 99: 817- 27.

[49] Godínez-Solís Y, Solís-Heredia MDJ, Roa-Espitia A,, et al. Low concentrations of lead decrease the sperm fertilization ability by altering the acrosome reaction in mice. Toxicol Appl Pharmacol 2019; 380: 114694.

[50] Xu WM, Shi QX, Chen WY,, et al. Cystic fibrosis transmembrane conductance regulator is vital to sperm fertilizing capacity and male fertility. Proc Natl Acad Sci USA 2007; 104: 9816- 21.

[51] Demarco IA, Espinosa F, Edwards J,, et al. Involvement of a Na⁺/HCO ₃⁻ cotransporter in mouse sperm capacitation. J Biol Chem 2003; 278: 7001- 9.

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2022 The Author(s) 2022. Published by Oxford University Press on behalf of Higher Education Press.