Testicular Injury Attenuated by Rapamycin Through Induction of Autophagy and Inhibition of Endoplasmic Reticulum Stress in Streptozotocin- Induced Diabetic Rats

Author(s): Wenjiao Shi, Zhixin Guo*, Ruixia Yuan

Journal Name: Endocrine, Metabolic & Immune Disorders - Drug Targets
Formerly Current Drug Targets - Immune, Endocrine & Metabolic Disorders

Volume 19 , Issue 5 , 2019

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Graphical Abstract:


Background and Objective: This study investigated whether rapamycin has a protective effect on the testis of diabetic rats by regulating autophagy, endoplasmic reticulum stress, and oxidative stress.

Methods: Thirty male Sprague-Dawley rats were randomly divided into three groups: control, diabetic, and diabetic treated with rapamycin, which received gavage of rapamycin (2mg.kg-1.d-1) after induction of diabetes. Diabetic rats were induced by intraperitoneal injection of streptozotocin (STZ, 65mg.Kg-1). All rats were sacrificed at the termination after 8 weeks of rapamycin treatment. The testicular pathological changes were determined by hematoxylin and eosin staining. The protein or mRNA expression of autophagy-related proteins (Beclin1, microtubule-associated protein light chain 3 (LC3), p62), ER stress marked proteins (CCAAT/enhancer-binding protein (C/EBP) homologous protein (CHOP), caspase-12), oxidative stress-related proteins (p22phox, nuclear factor erythroid2-related factor 2 (Nrf2)) and apoptosis-related proteins (Bax, B cell lymphoma-2 (Bcl-2)) were assayed by western blot or real-time fluorescence quantitative PCR.

Results: There were significant pathological changes in the testes of diabetic rats. The expression of Beclin1, LC3, Nrf2, Bcl-2 were significantly decreased and p62, CHOP, caspase12, p22phox, and Bax were notably increased in the testis of diabetic rats (P <0.05). However, rapamycin treatment for 8 weeks significantly reversed the above changes in the testis of diabetic rats (P <0.05).

Conclusion: Rapamycin appears to produce a protective effect on the testes of diabetic rats by inducing the expression of autophagy and inhibiting the expression of ER-stress, oxidative stress, and apoptosis.

Keywords: Rapamycin, autophagy, endoplasmic reticulum stress, oxidative stress, testis, type 1 diabetes.

Dahlquist, G.; Källén, B. Mortality in childhood-onset type 1 diabetes: A population-based study. Diabetes Care, 2005, 28(10), 2384-2387.
Mayerdavis, E.J.; Beyer, J.; Bell, R.A.; Dabelea, D.; Jr, D.A.R.; Imperatore, G.; Lawrence, J.M.; Liese, A.D.; Liu, L.; Marcovina, S. Diabetes in african american youth: Prevalence, incidence, and clinical characteristics: The SEARCH for Diabetes in Youth Study Diabetes Care, 2009, 32 Suppl 2(3), S112-122.
Amaral, S.; Oliveira, P.J.; Ramalhosantos, J. Diabetes and the impairment of reproductive function: Possible role of mitochondria and reactive oxygen species. Curr. Diabetes Rev., 2008, 4(1), 46-54.
Scarano, W.R.; Messias, A.G.; Oliva, S.U.; Klinefelter, G.R.; Kempinas, W.G. Sexual behaviour, sperm quantity and quality after short-term streptozotocin-induced hyperglycaemia in rats. Int. J. Androl., 2006, 29(4), 482-488.
Brody, H. Diabetes. Nature, 2012, 485(7398), S1.
Yang, C.; Kaushal, V.; Shah, S.V.; Kaushal, G.P. Autophagy is associated with apoptosis in cisplatin injury to renal tubular epithelial cells. Am. J. Physiol. Renal Physiol., 2008, 294(4), F777-F787.
Levine, B.; Kroemer, G. Autophagy in the pathogenesis of disease. Cell, 2008, 132(1), 27-42.
Levine, B.; Klionsky, D.J. Development by self-digestion: Molecular mechanisms and biological functions of autophagy. Dev. Cell, 2004, 6(4), 463-477.
Levine, B. Cell biology: Autophagy and cancer. Nature, 2007, 446(7137), 745-747.
Levine, B.; Deretic, V. Unveiling the roles of autophagy in innate and adaptive immunity. Nat. Rev. Immunol., 2007, 7(10), 767-777.
Meijer, A.J.; Codogno, P. Signalling and autophagy regulation in health, aging and disease. Mol. Aspects Med., 2006, 27(5-6), 411-425.
Williams, A.; Jahreiss, L.; Sarkar, S.; Saiki, S.; Menzies, F.M.; Ravikumar, B.; Rubinsztein, D.C. Aggregate-prone proteins are cleared from the cytosol by autophagy: Therapeutic implications. Curr. Top. Dev. Biol., 2006, 76, 89-101.
Rubinsztein, D.C.; Gestwicki, J.E.; Murphy, L.O.; Klionsky, D.J. Potential therapeutic applications of autophagy. Nat. Rev. Drug Discov., 2007, 6(4), 304-312.
Terman, A.; Brunk, U.T. Autophagy in cardiac myocyte homeostasis, aging, and pathology. Cardiovasc. Res., 2005, 68(3), 355-365.
Ebato, C.; Uchida, T.; Arakawa, M.; Komatsu, M.; Ueno, T.; Komiya, K.; Azuma, K.; Hirose, T.; Tanaka, K.; Kominami, E. Autophagy is important in islet homeostasis and compensatory increase of beta cell mass in response to high-fat diet. Cell Metab., 2008, 8(4), 325-332.
Masini, M.; Bugliani, M.; Lupi, R.; Guerra, S.D.; Boggi, U.; Filipponi, F.; Marselli, L.; Masiello, P.; Marchetti, P. Autophagy in human type 2 diabetes pancreatic beta cells. Islets, 2009, 52(6), 1083-1086.
Xie, Z.; Kai, L.; Eby, B.; Lozano, P.; He, C.; Pennington, B.; Li, H.; Rathi, S.; Dong, Y.; Tian, R. Improvement of cardiac functions by chronic metformin treatment is associated with enhanced cardiac autophagy in diabetic OVE26 mice. Diabetes, 2011, 60(6), 1770-1778.
Das, A.; Durrant, D.; Koka, S.; Salloum, F.N.; Lei, X.; Kukreja, R.C. Mammalian target of rapamycin (mTOR) inhibition with rapamycin improves cardiac function in type 2 diabetic mice. J. Biol. Chem., 2014, 289(7), 4145-41460.
Lin, W.; Shen, M.; Guo, X.; Bo, W.; Xia, Y.; Ning, W.; Qian, Z.; Jia, L.; Wang, X. Volume-sensitive outwardly rectifying chloride channel blockers protect against high glucose-induced apoptosis of cardiomyocytes via autophagy activation. Sci. Rep., 2017, 7, 44265.
Zhang, M.Z.; Wang, Y.; Paueksakon, P.; Harris, R.C. Epidermal growth factor receptor inhibition slows progression of diabetic nephropathy in association with a decrease in endoplasmic reticulum stress and an increase in autophagy. Diabetes, 2014, 63(6), 2063-2072.
Towns, R.; Guo, C.; Shangguan, Y.; Hong, S.; Wiley, J.W. Type 2 diabetes with neuropathy: Autoantibody stimulation of autophagy via Fas. Neuroreport, 2008, 19(3), 265-269.
Smith, M.H.; Ploegh, H.L.; Weissman, J.S. Road to ruin: Targeting proteins for degradation in the endoplasmic reticulum. Science, 2011, 334(6059), 1086-1090.
Ron, D.; Walter, P. Signal integration in the endoplasmic reticulum unfolded protein response. Nat. Rev. Mol. Cell Biol., 2007, 8(7), 519-529.
Hampton, R.Y. ER stress response: Getting the UPR hand on misfolded proteins. Cur. Biol., 2000, 10(14), R518-R521.
Jing, G.; Wang, J.J.; Zhang, S.X. ER Stress and apoptosis: A new mechanism for retinal cell death. J. Diabetes Res., 2013, 2012(1)589589
Balasubramanyam, M.; Singh, L.P.; Rangasamy, S.; Rangasamy, S. Molecular intricacies and the role of er stress in diabetes. Exp. Diabetes Res., 2012, 2012(2012), 958169.
O’Sullivanmurphy, B.; Urano, F. ER stress as a trigger for β-cell dysfunction and autoimmunity in type 1 diabetes. Diabetes, 2012, 61(4), 780-781.
Su, J.; Zhou, L.; Kong, X.; Yang, X.; Xiang, X.; Zhang, Y.; Li, X.; Sun, L. Endoplasmic reticulum is at the crossroads of autophagy, inflammation, and apoptosis signaling pathways and participates in the pathogenesis of diabetes mellitus. J. Diabetes Res., 2013, 2013(3)193461
Li, M.; Liu, Z.; Zhuan, L.; Wang, T.; Guo, S.; Wang, S.; Liu, J.; Ye, Z. Effects of apocynin on oxidative stress and expression of apoptosis-related genes in testes of diabetic rats. Mol. Med. Rep., 2013, 7(1), 47-52.
Maiese, K.; Morhan, S.D.; Zhao, Z.C. Oxidative stress biology and cell injury during type 1 and type 2 diabetes mellitus. Curr. Neurovasc. Res., 2007, 4(1), 63-71.
Demirtas, L.; Guclu, A.; Erdur, F.M.; Akbas, E.M.; Ozcicek, A.; Onk, D.; Turkmen, K. Apoptosis, autophagy & endoplasmic reticulum stress in diabetes mellitus. Indian J. Med. Res., 2016, 144(4), 515-524.
Tang, X.Y.; Zhang, Q.; Dai, D.Z.; Ying, H.J.; Wang, Q.J.; Dai, Y. Effects of strontium fructose 1,6-diphosphate on expression of apoptosis-related genes and oxidative stress in testes of diabetic rats. Int. J. Urol., 2008, 15(3), 251-256.
Martin, D.E.; Hall, M.N. The expanding TOR signaling network. Curr. Opin. Cell Biol., 2005, 17(2), 158-166.
Maiese, K. Cutting through the complexities of mTOR for the treatment of stroke. Curr. Neurovasc. Res., 2014, 11(2), 177-186.
Tang, Z.; Baykal, A.T.; Gao, H.; Quezada, H.C.; Zhang, H.; Bereczki, E.; Serhatli, M.; Baykal, B.; Acioglu, C.; Wang, S. mTor is a signaling hub in cell survival: A mass-spectrometry-based proteomics investigation. J. Proteome Res., 2014, 13(5), 2433-2444.
Bacharwikstrom, E.; Wikstrom, J.D.; Ariav, Y.; Tirosh, B.; Kaiser, N.; Cerasi, E.; Leibowitz, G. Stimulation of autophagy improves endoplasmic reticulum stress-induced diabetes. Diabetes, 2013, 62(4), 1227-1237.
Maiese, K. mTOR: Driving apoptosis and autophagy for neurocardiac complications of diabetes mellitus. World J. Diabetes, 2015, 6(2), 217-224.
Song, Q.; Han, C.C.; Xiong, X.P.; He, F.; Gan, W.; Wei, S.H.; Liu, H.H.; Li, L.; Xu, H.Y. PI3K-Akt-mTOR signal inhibition affects expression of genes related to endoplasmic reticulum stress. Genet. Mol. Res. Gmr., 2016, 15(3)
Paschoal, V.A.; Amano, M.T.; Belchior, T.; Magdalon, J.; Chimin, P.; Andrade, M.L.; Ortizsilva, M.; Castro, É.; Yamashita, A.S.; Rosa Neto, J.C. mTORC1 inhibition with rapamycin exacerbates adipose tissue inflammation in obese mice and dissociates macrophage phenotype from function. Immunobiology, 2017, 222(2), 261-271.
Agbaje, I.M.; Rogers, D.A.; Mcvicar, C.M.; Mcclure, N.; Atkinson, A.B.; Mallidis, C.; Lewis, S.E. Insulin dependant diabetes mellitus: Implications for male reproductive function. Hum. Reprod., 2007, 22(7), 1871-1877.
Dinulovic, D.; Radonjic, G. Diabetes mellitus/male infertility. Arch. Androl., 2009, 25(3), 277-293.
Koh, P.O. Streptozotocin-induced diabetes increases apoptosis through JNK phosphorylation and Bax activation in rat testes. J. Vet. Med. Sci., 2007, 69(9), 969-971.
Yorimitsu, T.; Klionsky, D.J. Autophagy: Molecular machinery for self-eating. Cell Death Differ., 2005, 12(12)(Suppl.), 1542-1552.
Kroemer, G.; Mariño, G.; Levine, B. Autophagy and the integrated stress response. Mol. Cell, 2010, 40(2), 280-293.
Bacharwikstrom, E.; Wikstrom, J.D.; Kaiser, N.; Cerasi, E.; Leibowitz, G. Improvement of ER stress-induced diabetes by stimulating autophagy. Autophagy, 2013, 9(4), 626-628.
Schröder, M.; Kaufman, R.J. The mammalian unfolded protein response. Annu. Rev. Biochem., 2005, 74(74), 739-789.
Boyce, M.; Yuan, J. Cellular response to endoplasmic reticulum stress: A matter of life or death. Cell Death Differ., 2006, 13(3), 363-373.
Gupta, S.; Read, D.E.; Deepti, A.; Cawley, K.; Gupta, A.; Oommen, D.; Verfaillie, T.; Matus, S.; Smith, M.A.; Mott, J.L. Perk-dependent repression of miR-106b-25 cluster is required for ER stress-induced apoptosis. Cell Death Disease., 2012, 3(6)e333
Marciniak, S.J.; Yun, C.Y.; Oyadomari, S.; Novoa, I.; Zhang, Y.; Jungreis, R.; Nagata, K.; Harding, H.P.; Ron, D. CHOP induces death by promoting protein synthesis and oxidation in the stressed endoplasmic reticulum. Genes Dev., 2004, 18(24), 3066-3077.
Nakagawa, T.; Zhu, H.; Morishima, N.; Li, E.; Xu, J.; Yankner, B.A.; Yuan, J. Caspase-12 mediates endoplasmic-reticulum-specific apoptosis and cytotoxicity by amyloid-beta. Nature, 2000, 403(6765), 98-103.
Xu, M.; Dai, D.Z.; Zhang, Q.; Cheng, Y.S.; Dai, Y. Upregulated NADPH oxidase contributes to diabetic testicular complication and is relieved by strontium fructose 1,6-diphosphate. Exp. Clin. Endocrinol. Diabetes, 2010, 118(07), 459-465.
Wang, Y.; Zhang, Z.; Guo, W.; Sun, W.; Miao, X.; Wu, H.; Cong, X.; Wintergerst, K.A.; Kong, X.; Cai, L. AB196. Sulforaphane reduction of testicular apoptotic cell death in diabetic mice is associated with the up-regulation of Nrf2 expression and function. Am. J. Physiol. Endocrinol. Metab., 2014, 307(1), E14-E23.
Zhao, Y.; Tan, Y.; Dai, J.; Li, B.; Guo, L.; Cui, J.; Wang, G.; Shi, X.; Zhang, X.; Mellen, N. Exacerbation of diabetes-induced testicular apoptosis by zinc deficiency is most likely associated with oxidative stress, p38 MAPK activation, and p53 activation in mice. Toxicol. Lett., 2011, 200(1), 100-106.
Sainio-Pöllänen, S.; Henriksén, K.; Parvinen, M.; Simell, O.; Pöllänen, P. Stage-specific degeneration of germ cells in the seminiferous tubules of non-obese diabetic mice. Int. J. Androl., 1997, 20(4), 243-254.
Koh, P.O. Streptozotocin-induced diabetes increases the interaction of Bad/Bcl-X L and decreases the binding of pBad/14–3–3 in rat testis. Life Sci., 2007, 81(13), 1079-1084.
K. B.; Fleming, S.L.; Johnson, K.J.; Patel, S.R.; Schoenfeld, H.A., Role of sertoli cells in injury-associated testicular germ cell apoptosis. Proc. Soc. Exp. Biol. Med., 2000, 225(2), 105-115.
Fujitani, Y.; Kawamori, R.; Watada, H. The role of autophagy in pancreatic beta-cell and diabetes. Autophagy, 2009, 5(2), 280-282.
Tanaka, Y.; Kume, S.; Kitada, M.; Kanasaki, K.; Uzu, T.; Maegawa, H.; Koya, D. Autophagy as a therapeutic target in diabetic nephropathy. J. Diabetes Res., 2011, 2012(12)628978
Xu, X.; Kobayashi, S.; Chen, K.; Timm, D.; Volden, P.; Huang, Y.; Gulick, J.; Yue, Z.; Robbins, J.; Epstein, P.N. Diminished autophagy limits cardiac injury in mouse models of type 1 diabetes. J. Biol. Chem., 2013, 288(25), 18077-18092.
Zhao, Y.; Zhang, L.; Qiao, Y.; Zhou, X.; Wu, G.; Wang, L.; Peng, Y.; Dong, X.; Huang, H.; Si, L. Heme oxygenase-1 prevents cardiac dysfunction in streptozotocin-diabetic mice by reducing inflammation, oxidative stress, apoptosis and enhancing autophagy. Plos One, 2013, 8(9)e75927
Hartford, C.M.; Ratain, M.J. Rapamycin: Something old, something new, sometimes borrowed and now renewed. Clin. Pharmacol. Ther., 2007, 82(4), 381-388.
Ozcan, U.; Ozcan, L.; Yilmaz, E.; Düvel, K.; Sahin, M.; Manning, B.D.; Hotamisligil, G.S. Loss of the tuberous sclerosis complex tumor suppressors triggers the unfolded protein response to regulate insulin signaling and apoptosis. Mol. Cell, 2008, 29(5), 541-451.
Li, H.; Min, Q.; Ouyang, C.; Lee, J.; He, C.; Zou, M.H.; Xie, Z. AMPK activation prevents excess nutrient-induced hepatic lipid accumulation by inhibiting mTORC1 signaling and endoplasmic reticulum stress response. Biochim. Biophys. Acta, 2014, 1842(9), 1844-1854.
Yin, J.J.; Li, Y.B.; Wang, Y.; Liu, G.D.; Wang, J.; Zhu, X.O.; Pan, S.H. The role of autophagy in endoplasmic reticulum stress-induced pancreatic Î2 cell death. Autophagy, 2012, 8(2), 158-164.
Rubinsztein, D.C. The roles of intracellular protein-degradation pathways in neurodegeneration. Nature, 2006, 443(7113), 780-786.
Association, A.D. Diagnosis and classification of diabetes mellitus. Recenti Prog. Med., 2012, 101(7-8), 274-276.
Xiao, T.; Xu, G.; Ling, N.; Song, W.; Lei, S.; He, T.; Huang, Y.; Zhang, J.; Ke, Y.; Wang, J. Rapamycin promotes podocyte autophagy and ameliorates renal injury in diabetic mice. Mol. Cellular. Biochem., 2014, 394(1-2), 145-154.
Rovira, J.; Marcelo, A.E.; Burke, J.T.; Brault, Y.; Moyarull, D.; Bañónmaneus, E.; Ramírezbajo, M.J.; Gutiérrezdalmau, A.; Revuelta, I.; Quintana, L.F. Effect of mTOR inhibitor on body weight: From an experimental rat model to human transplant patients. Transpl. Int., 2008, 21(10), 992-998.
Yamauchi, T.; Kamon, J.; Ito, Y.; Tsuchida, A.; Yokomizo, T.; Kita, S.; Sugiyama, T.; Miyagishi, M.; Hara, K.; Tsunoda, M. Cloning of adiponectin receptors that mediate antidiabetic metabolic effects. Nature, 2007, 423(6941), 762-769.
Martin, L.J. Implications of adiponectin in linking metabolism to testicular function. Endocrine, 2014, 46(1), 16-28.

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Year: 2019
Page: [665 - 675]
Pages: 11
DOI: 10.2174/1871530319666190102112844
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