Examination of the immunohistochemical localization and gene expression by RT-PCR of the oxytocin receptor in diabetic and non-diabetic mouse testis

Ayşe Aydoğan, Seyit Ali Bingöl, Ayşe Aydoğan, Seyit Ali Bingöl

Abstract

Objectives: The aim of this study was to determine Oxytocin receptor (OTR) gene expression and localization in diabetic and non-diabetic mouse testes by RT-PCR and immunohistochemistry, respectively.

Materials and methods: In this study, 18 male BALB/c mice (8-12 weeks old) were used and divided into three groups: diabetic, sham, and control. Streptozotocin (STZ) was applied to the diabetic group and sodium citrate was administered to the sham group in the same way, however, the control group was left untouched. The testicular tissues were removed on the thirtieth day of testing; the right testis tissues were passed through a routine histologic process and sections were stained with H&E and PAS staining techniques. The avidin-biotin-peroxidase method was applied to determine OTR immunoreactivity, while the left testis tissues were used for RT-PCR.

Results: It was found that the body weight had decreased in the diabetic group and the diameter of the seminiferous tubules in the said group was shorter than those of the other groups. There were no obvious differences with regard to the histologic appearance between the groups. The immunohistochemical examination showed that the OTR immunoreactivity was strong in the control and sham groups but weak in the diabetic group, and the immunoreactivity was only seen in the Leydig cells. In addition, the OTR gene expression was lower in the diabetic group than in the other groups.

Conclusion: We concluded that diabetes reduces the OTR expression in the testis. It is suggested that OTR protection should be researched in diabetes for healthy reproduction and sexuality.

Keywords: Diabetes; Immunohistochemistry Oxytocin receptor; RT-PCR Testis.

Conflict of interest statement

We report no conflicts of interest as authors of the present study.

Figures

Figure 1
Figure 1
The comparison of the diameters of tubulus seminiferous contortus (TSC) between groups. The average diameter in the diabetic group of BALB/c mice was smaller than those of the others (*P<0.05)
Figure 2
Figure 2
The comparison of the blood glucose levels between groups of BALB/c mice. The glucose level in the diabetic group on the 30th day was higher than the glucose levels in the sham and control groups on the 30th day and the glucose levels in all groups on the 0th day. *P<0.05
Figure 3
Figure 3
The normal histologic appearance of the testis in the control (A) and diabetic (B) groups. H&E stain. Arrow: Interstitial area. TSC: Tubulus seminiferous contortus. Bar: 100 μm
Figure 4
Figure 4
The histological structure of testis in the control (A) and diabetic (B) groups. The basal membrane surrounded the tubulus seminiferus contortus. PAS stain. Arrow: Basal membrane of tubulus seminiferous contortus (TSC). Bar: 50 μm
Figure 5
Figure 5
OTR immunoreactivity in the testis of the control group. OTR immunoreactivity was seen only in the interstitial area. Arrows: Interstitial area. TSC: Tubulus seminiferous contortus. Bar: 50 μm
Figure 6
Figure 6
OTR immunoreactivity in the testis of the sham group. The mice were injected with sodium citrate. OTR immunoreactivity was seen only in the interstitial area. Arrow: Interstitial area. Bar: 100 μm
Figure 7
Figure 7
OTR immunoreactivity in the testis of the diabetic group. The mice were injected with STZ (100 mg/kg, IP). There was weak OTR immunoreactivity only in the interstitial area. Arrow: Interstitial area. Bar: 100 μm
Figure 8
Figure 8
The view of PCR gel for OTR gene in the testes of control, diabetic, and sham groups of BALB/c mice
Figure 9
Figure 9
OTR Gene expression level in the testes. OTR expression in the testes of the diabetic group of BALB/c mice was lower than in the sham and control groups. *P<0.05

References

    1. Kimura T, Makino Y, Saji F, Takemura M, Inoue T, Kikuchi T, et al. Molecular characterization of a cloned human oxytocin receptor. Eur J Endocrinol. 1994;131:385–390.
    1. Anjum S, Anuradha A, Krishna A. A possible direct action of oxytocin on spermatogenesis and steroidogenesis in pre-pubertal mouse. Andrologia. 2018;e12958:1–11.
    1. Lockard MA, Ebert MS, Bargmann CI. Oxytocin mediated behavior in invertebrates: an evolutionary perspective. Dev Neurobiol. 2017;77:128–142.
    1. Nicholson HD, Jenkın L. Oxytocin and prostatic function. Adv Exp Med Biol. 1995;395:529–538.
    1. Roth KI. Diabetes treatment-bridging the divide. N Engl J Med. 2007;356:1499–1501.
    1. Gurka MJ, Golden SH, Musani SK, Sims M, Vishnu A, Guo Y, et al. Independent associations between a metabolic syndrome severity score and future diabetes by sex and race: the atherosclerosis risk in communities study and Jackson heart study. Diabetologia. 2017;60:1261–1270.
    1. Agbaje IM, Rogers DA, Mcvicar CM, Mcclure N, Atkinson AB, Mallidis C, et al. Insulin dependent diabetes mellitus: implications for male reproductive function. Hum Reprod. 2007;22:1871–1877.
    1. Kort HI, Massey JB, Elsner CW, Mitchell-Leef D, Shapiro DB, Witt MA, et al. Impact of body mass index values on sperm quantity and quality. J Androl. 2006;27:450–452.
    1. Bingol SA ve Kocamıs H. The gene expression profile by RT-PCR and Immunohistochemical expression pattern of catalase in the kidney tissue of both health and diabetic mice. Kafkas Univ Vet Fak Derg . 2010;16:825–834.
    1. Kanitkar M, Bhonde R. Existence of islet regenerating factors within the pancreas. The Rev Diabet Stud. 2004;1:185–192.
    1. Shu S, Ju G, Fan L. The glucose oxidase-DAB–nickel in peroxidase histochemistry of the nervous system. Neurosci Lett. 1988;85:169–171.
    1. Kasahara Y, Sato K, Takayanagi Y, Mizukami H, Ozawa K, Hidema S, et al. Oxytocin receptor in the hypothalamus is sufficient to rescue normal thermoregulatory function in male oxytocin receptor knockout mice. Endocrinology. 2013;154:4305–4315.
    1. Göçmen C, Seçilmiş A, Kumcu EK, Ertuğ PU, Önder S, Dikmen A, et al. Effects of vitamin E and sodium selenate on neurogenic and endothelial relaxation of corpus cavernosum in the diabetic mouse. Eur J Pharmacol. 2000;398:93–98.
    1. Wada J, Zhang H, Tsuchiyama Y, Hiragushi K, Hıda K, Shikata K, et al. Gene exspression profile in streptozotosin-induced diabetic mice kidneys undergoing glomerulosclerosis. Kidney Int. 2001;59:1363–1373.
    1. Elis Yildiz S, Deprem T, Karadag Sari E, Bingol SA, Koral Tasci S, Aslan S, et al. Immunohistochemical distribution of leptin in kidney tissues of melatonin treated diabetic rats. Biotech Histochem. 2015;90:270–277.
    1. Kim NK, Stankovic M, Cushman TT, Goldstein I, Munarrız R, Traish AM. Streptozotosin- induced diabetes in the rat is associated with changes in vaginal hemodynamics, morphology and biochemikal markers. BMC Physiol. 2006;6:1–9.
    1. Orman D, Vardi N, Ates B, Taslidere E, Elbe H. Aminoguanidine mitigates apoptosis, testicular seminiferous tubules damage, and oxidative stress in streptozotocin-induced diabetic rats. Tissue Cell. 2015;47:284–290.
    1. Haan JB, Stefanovic N, Paterson DN, Scurr LL, Croft KD, Mori TA, et al. Kidney expression of glutathione peroxidase-1 is not protective against streptozotocin-induced diabetic nephropathy. Am J Physiol Renal Physiol. 2005;289:F544–F551.
    1. Jelodar G, Khaksar Z, Pourahmadi M. Endocrine profile and testicular histomorphometry in adult rat offspring of diabetic mothers. J Physiol Sci. 2009;59:377–382.
    1. Cai L, Chen S, Evans T, Deng DX, Mukherjee K, Chakrabarti S. Apoptotic germ-cell death and testicular damage in experimental diabetes: prevention by endothelin antagonism. Urol res. 2000;28:342–347.
    1. Guneli E, Tugyan K, Ozturk H, Gumustekin M, Cilaker S, Uysal N. Effect of melatonin on testicular damage in streptozotocin-induced diabetes rats. Eur Surg Res. 2008;40:354–360.
    1. Akkoc H, Kelle I, Tunik S, Erdinc M, Erdinc L, Nergiz Y. Effects of Ethyl Pyruvate on Testicular Damage in Rats with Stretozotocin-Induced Diabetes. Acta Endocrinol. 2012;8:35–45.
    1. Kianifard D, Sadrkhanlou RA, Hasanzadeh S. The ultrastructural changes of the sertoli and leydig cells following streptozotocin induced diabetes. Iran J Basic Med Sci. 2012;15:623–635.
    1. Koroglu P, Erkanli Senturk G, Yucel D, Bingol Ozakpinar O, Uras F, Arbak S. The effect of exogenous oxytocin on streptozotocin (STZ)-induced diabetic adult rat testes. Peptides. 2015;63:47–54.
    1. Erbas O, Korkmaz AH, Oltulu F, Aktug H, Yavasoglu A, Akman L, et al. Oxytocin alleviates cisplatin-induced renal damage in rats. Iran J Basic Med Sci. 2014;17:747–752.
    1. Veening JG, de Jong TR, Waldinger MD, Korte SM, Oliver B. The role of oxytocin in male and female reproductive behavior. Eur J Pharmacol. 2015;753:209–228.
    1. Gossen A, Hahn A, Westphal S, Prinz S, Schultz RT, Gründer G, et al. Oxytocin plasma concentrations after single intranasal oxytocin administration - a study in healthy men. Neuropeptides. 2012;46:211–215.
    1. Frayne J, Nıcholson HD. Localization of oxytocin receptors in the human and macaque monkey male reproductive tracts: evidence for a physiological role of oxytocin in the male. Mol Hum Reprod. 1998;4:527–532.
    1. Einspanier A, Ivell R. Oxytocin and receptor expression in reproductive tissues of the male marmoset monkey. Biol Reprod . 1997;56:416–422.
    1. Sanna F, Bratzu J, Argiolas A, Melis MR. Oxytocin induces penile erection and yawning when injected into the bed nucleus of the stria terminalis: Involment of glutamic acid, dopamine, and nitric oxide. Horm Behav. 2017;96:52–61.
    1. Li M, Wang T, Guo S, Rao K, Liu J, Ye Z. Expression of oxcytocin receptor in diabetic rat penis. Andrologia. 2012;44:716–720.
    1. Whittington K, Assinder SJ, Parkinson T, Lapwood KR, Nicholson HD. Function and localization of oxytocin receptors in the reproductive tissue of rams. Reproduction. 2001;122:317–325.
    1. Zhang XH, Filippi S, Vignozzi L, Morelli A, Mancina R, Luconi M, et al. Identification, localization and functional in vitro and in vivo activity of oxytocin receptor in the rat penis. J Endocrinol. 2005;184:567–576.
    1. Prapaiwan N, Manee-in S, Moonarmart W, Srisuwatanasagul S. The expressions in oxytocin and sex steroid receptors in the reproductive tissues of normal and unilateral cryptorchid dogs. Theriogenology. 2017;100:59–65.

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