|Year : 2019 | Volume
| Issue : 1 | Page : 125
Testosterone and zinc supplementations on renal ischemia-reperfusion injury in orchiectomized rats
Fatemeh Moslemi1, Farzan Piudeh1, Mohammad-Reza Hajian1, Amir Khodarahmi1, Mehdi Nematbakhsh2
1 Water and Electrolytes Research Center, Isfahan University of Medical Sciences, Isfahan, Iran
2 Water and Electrolytes Research Center; Department of Physiology, Isfahan University of Medical Sciences; Isfahan MN Institute of Basic and Applied Sciences Research, Isfahan, Iran
|Date of Submission||25-Feb-2018|
|Date of Acceptance||30-Apr-2018|
|Date of Web Publication||19-Jul-2019|
Water and Electrolytes Research Center, Isfahan University of Medical Sciences, Isfahan
Source of Support: None, Conflict of Interest: None
Background: Renal ischemia-reperfusion (IR) injury has numerous deleterious effects on the kidney function. An experimental investigation was conducted to determine the possible protective role of testosterone (TES) and zinc (Zn) supplementations on the kidney function after IR injury in orchiectomized rats. Methods: Orchiectomized rats (n = 32) were divided into the five groups as sham operated (Group 1), IR (Group 2), IR pretreatment with TES (IR + TES, Group 3), Zn (IR + Zn, Group 4), and TES + Zn (IR + TES + Zn, Group 5). Twenty-four hours' post-IR injury, the animals were sacrificed and the required parameters were measured. Results: The results revealed that there were not any significant difference in serum levels of creatinine (Cr), nitrite and malondialdehyde (MDA), Cr clearance (ClCr), renal sodium (Na) load, and percentage of Na excretion (ENa%) between sham and IR groups. The pretreatment with TES and Zn either alone or combine did not alter the serum levels of Cr, nitrite and MDA, and ClCr, Na load, and ENa%. However, pretreatment with Zn, TES, or combined altered kidney weight, kidney tissue levels of nitrite and MDA, and urine flow in IR groups. Conclusions: The orchiectomy itself performed protective effect against renal IR injury. However, pretreatment with Zn or TES may not alter kidney function against renal IR in orchiectomized rats.
Keywords: Orchiectomy, renal ischemia, testosterone, zinc
|How to cite this article:|
Moslemi F, Piudeh F, Hajian MR, Khodarahmi A, Nematbakhsh M. Testosterone and zinc supplementations on renal ischemia-reperfusion injury in orchiectomized rats. Int J Prev Med 2019;10:125
|How to cite this URL:|
Moslemi F, Piudeh F, Hajian MR, Khodarahmi A, Nematbakhsh M. Testosterone and zinc supplementations on renal ischemia-reperfusion injury in orchiectomized rats. Int J Prev Med [serial online] 2019 [cited 2020 Jul 12];10:125. Available from: http://www.ijpvmjournal.net/text.asp?2019/10/1/125/263086
| Introduction|| |
The restriction of blood flow to vital organs followed by reperfusion is characterized as ischemia-reperfusion(IR) injury. The renal IR injury usually is accompanied with formation or activation of many substances such as reactive oxygen species, cytokines, and chemokines. In clinic, acute kidney injury and chronic kidney disease usually are resulted in renal IR injury. Gender also plays a pivotal role in the outcome of kidney IR injury, and the sex hormones are highlighted while males compared with females are considered to be more susceptible to renal IR injury., Some controversial studies indicated that orchiectomy has a protective role in renal IR injury and testosterone(TES) supplementation could reverse it while others' experimental studies proved otherwise.,,
On the other hand, the antioxidant therapy by zinc(Zn) supplementation may protect the kidney against IR injury. Zn inhibits the apoptosis process after renal IR injury, and the protective role of Zn is reported to act in a dose-dependent manner., Previously, we reported that Zn could protect the kidney against renal IR injury gender dependently. In the current study, the roles of TES and Zn supplementations were considered in orchiectomized rats.
| Methods|| |
Thirty-two adult male(200±20g) Wistar rats (Animal Center, Isfahan University of Medical Sciences, Isfahan, Iran) were housed under standard conditions. The protocol of this study was confirmed to be in accordance by Ethics Committee of the Isfahan University of Medical Sciences.
Male rats were anesthetized with chloral hydrate (450 mg/kg, ip), and a midline abdominal incision was made and the testicles were removed and the skin was sutured.
One week after orchiectomy, the animals were randomly assigned into five experimental groups, and they received one of the following treatments:
- Group 1 (n = 6, sham operated): Rats received sesame oil alone intramuscularly once a week for 3 weeks and 1 week later underwent renal IR injury procedure without clamping renal vessels
- Group 2 (n = 5, IR group): Rats received sesame oil alone intramuscularly once a week for 3 weeks and 1 week later underwent renal IR injury surgery
- Group 3 (n = 6, IR + TES group): Rats received TES (10 mg/kg dissolved in sesame oil) intramuscularly once a week for 3 weeks and 1 week later underwent renal IR surgery
- Groups 4 (n = 7, IR + Zn group): Rats received sesame oil alone intramuscularly once a week for 3 weeks and 1 week later underwent renal IR surgery, but 5 consecutive days before surgery, they received Zn supplement (10 mg/kg/day, ip)
- Group 5 (n = 8, IR + TES + Zn group): Rats received TES once a week for 3 weeks and 1 week later underwent renal IR surgery, but 5 consecutive days before surgery, they received Zn supplement. The summary of the group's treatments is shown in [Table 1].
|Table 1: The summary of the groups' treatments. The experiment was designed in five steps|
Click here to view
Renal ischemia-reperfusion injury
The animals were anesthetized by chloral hydrate (450 mg/kg, ip). Two small incisions were made on the flanks, and the renal artery and vein on both sides were clamped. After 45 min, the clamps were removed and the kidneys were allowed to reperfuse. Eighteen hours later, the animals were placed in metabolic cages to collect the urine for the next 6 h. Twenty-four hours' postrenal IR injury, blood samples were obtained through heart puncture. Finally, all animals were sacrificed and kidneys were removed and weighed immediately. The right kidney was homogenized and centrifuged. The Cr clearance (ClCr) was calculated based on clearance formula as ClCr = urine flow (UF) × Ucr/Pcr, where UF, Ucr, and Pcr stand for UF, urine Cr level, and serum Cr level.
The levels of Cr in serum and urine were determined using quantitative diagnostic kits (Pars Azmoon, Iran). Assessments of malondialdehyde (MDA) level in the serum and kidney tissue were performed by the manual method. The serum and kidney levels of nitrite (stable nitric oxide [NO] metabolite) were measured using Griess method. The levels of sodium (Na) in serum and urine were measured using flame photometer assay.
Data were reported as mean ± standard error of mean. The exact statistic method (Kruskal–Wallis H-test and Mann–Whitney U-test) was applied to compare the parameters between the groups. P < 0.05 was considered statistically significant.
| Results|| |
The serum levels of Cr, nitrite and MDA, ClCr, filtrate Na load, and percentage of Na excretion (ENa%) were not significantly different between the groups [Figure 1]. The total kidney weight (KW) per 100 g body weight in IR, IR + Zn, and IR + TES + Zn groups was decreased significantly when compared with sham group [P < 0.05, [Figure 1]]. In addition, KW in IR + TES group was increased statistically when compared with IR group (P < 0.05). The tissue level of nitrite in IR + TES, IR + Zn, and IR + TES + Zn groups was greater than sham group (P < 0.05) while this marker in IR + TES + Zn group was increased when compared with IR groups significantly [P < 0.05, [Figure 1]]. The tissue MDA level in IR + Zn and IR + TES + Zn groups were lower than IR group statistically [P < 0.05, [Figure 1]]. In the all IR group (Groups 2–5), the mean levels of UF were increased; however, statistically, a higher UF was detected in IR + TES, IR + Zn, and IR + TES + Zn groups when compared with sham group [P < 0.05, [Figure 1].
|Figure 1: The serum (S) levels of creatinine, nitrite, and malondialdehyde and kidney tissue (T) levels of malondialdehyde and nitrite and kidney weight, Cr clearance, urine flow, renal Na load, and percentage of Na excretion (ENa%) in the all experimental group. The P value in each panel was obtained by Kruskal–Wallis H-test. The data in ischemia-reperfusion + testosterone, ischemia-reperfusion + zinc, and ischemia-reperfusion + testosterone + zinc were compared with sham or ischemia-reperfusion groups using Mann–Whitney U-test, and the symbols indicate significant difference from (*) sham or (†) ischemia-reperfusion groups (P < 0.05)|
Click here to view
| Discussion|| |
The present study was designed to evaluate the effect of TES and Zn supplementations on renal IR injury in orchiectomized rat. The findings suggested that orchiectomy alone has a protective role in renal IR injury. However, although administration of TES and Zn altered some of the parameters, it seems that their certain protective roles may be failed in this model.
Gender plays a critical role in renal IR outcomes, and males are more susceptible to the renal IR and also have a delayed repair in comparison to females. Kang et al. reported that orchiectomy decreased the inflammatory response, and TES replacement enhanced renal injury. Furthermore, Park et al. showed that administration of TES in female mice increases renal inflammatory response in renal IR injury. Actually, the presence of TES rather than the absence of estrogen cause inflammation through inhibition of NO synthase activation.,
There are some studies against our findings. Soljancic et al. concluded that TES level in serum was seriously reduced after renal IR injury in normal rats, and the kidney could be protected against IR injury by the reduced TES while castration also promoted kidney injury. They also found that intravenous administration of TES (20 μg/kg/min) reduced the kidney IR injury. Other study indicated that reduction of TES level is associated with undesired outcome in coronary heart diseases. Moreover, TES exhibits a protective effect against spinal cord IR. Albayrak et al. data indicated that administration of TES protected intestinal IR injury, but such observation was not detected in the absence of testes. However, the exact mechanism by which TES protects the kidney is not documented yet. On the contrary, the TES treatment in renal IR injury did not protect the kidney while orchiectomy alone showed a protective effect. In addition and similar to our study, Park et al. found that orchiectomy itself could protect the kidney against IR injury, and the protective role of orchiectomy is related to the expression of heat-shock proteins. One possibility for these controversies is speculated that this difference may be due to the injection protocol. In this study, TES was injected once a week; however, other studies injected TES daily or more frequently.
Guo et al. found that Zn has a protective effect on renal IR by antiapoptotic and antioxidant capacity, and the protective effect of Zn on renal IR is dose dependent. Therefore, although Zn performed some protective effect against renal IR, it may fail in orchiectomized model.
| Conclusions|| |
It seems that orchiectomy itself with uncertain mechanism performed some protective effect against renal IR injury. In such condition, no protective role from TES or Zn supplementations was observed. According to other studies, the protective effect of TES or Zn against renal IR injury was reported, in normal animals. Orchidectomy disturbs many hormonal and nonhormonal pathways,, and TES also may affect Zn through glucose cycle. Therefore, exogenous administration of TES and Zn may not perform an effective role against renal IR injury in orchiectomized rats due to complex pathways disturbances.
This research was supported by Isfahan University of Medical Sciences (Grant # 296098).
Financial support and sponsorship
Conflicts of interest
There are no conflicts of interest.
| References|| |
Malek M, Nematbakhsh M. Renal ischemia/reperfusion injury; from pathophysiology to treatment. J Renal Inj Prev 2015;4:20-7.
Jang HR, Rabb H. The innate immune response in ischemic acute kidney injury. Clin Immunol 2009;130:41-50.
Zuk A, Bonventre JV. Acute kidney injury. Annu Rev Med 2016;67:293-307.
Robert R, Ghazali DA, Favreau F, Mauco G, Hauet T, Goujon JM, et al.
Gender difference and sex hormone production in rodent renal ischemia reperfusion injury and repair. J Inflamm (Lond) 2011;8:14.
Lima-Posada I, Portas-Cortés C, Pérez-Villalva R, Fontana F, Rodríguez-Romo R, Prieto R, et al.
Gender differences in the acute kidney injury to chronic kidney disease transition. Sci Rep 2017;7:12270.
Kang KP, Lee JE, Lee AS, Jung YJ, Kim D, Lee S, et al.
Effect of gender differences on the regulation of renal ischemia-reperfusion-induced inflammation in mice. Mol Med Rep 2014;9:2061-8.
Soljancic A, Ruiz AL, Chandrashekar K, Maranon R, Liu R, Reckelhoff JF, et al.
Protective role of testosterone in ischemia-reperfusion-induced acute kidney injury. Am J Physiol Regul Integr Comp Physiol 2013;304:R951-8.
Park KM, Cho HJ, Bonventre JV. Orchiectomy reduces susceptibility to renal ischemic injury: A role for heat shock proteins. Biochem Biophys Res Commun 2005;328:312-7.
Rao K, Sethi K, Ischia J, Gibson L, Galea L, Xiao L, et al.
Protective effect of zinc preconditioning against renal ischemia reperfusion injury is dose dependent. PLoS One 2017;12:e0180028.
Ogawa T, Mimura Y. Antioxidant effect of zinc on acute renal failure induced by ischemia-reperfusion injury in rats. Am J Nephrol 1999;19:609-14.
Guo L, Li P, Meng C, Lu R, Yang Y, Zhou Y, et al.
Protective effect of zinc on mouse renal ischemia-reperfusion injury by anti-apoptosis and antioxidation. Curr Pharm Biotechnol 2014;15:577-82.
Moslemi F, Talebi A, Nematbakhsh M. The protective effect of zinc supplementation on renal ischemia/reperfusion injury in rat: Gender-related Difference. Int J Prev Med 2019;10:68. doi: 10.4103/ijpvm.IJPVM_279_17. eCollection 2019.
Malkin CJ, Pugh PJ, Morris PD, Asif S, Jones TH, Channer KS, et al.
Low serum testosterone and increased mortality in men with coronary heart disease. Heart 2010;96:1821-5.
Gürer B, Kertmen H, Kasim E, Yilmaz ER, Kanat BH, Sargon MF, et al.
Neuroprotective effects of testosterone on ischemia/reperfusion injury of the rabbit spinal cord. Injury 2015;46:240-8.
Albayrak Y, Halici Z, Odabasoglu F, Unal D, Keles ON, Malkoc I, et al.
The effects of testosterone on intestinal ischemia/reperfusion in rats. J Invest Surg 2011;24:283-91.
Park KM, Kim JI, Ahn Y, Bonventre AJ, Bonventre JV. Testosterone is responsible for enhanced susceptibility of males to ischemic renal injury. J Biol Chem 2004;279:52282-92.
El Majdoubi M, Ramaswamy S, Sahu A, Plant TM. Effects of orchidectomy on levels of the mRNAs encoding gonadotropin-releasing hormone and other hypothalamic peptides in the adult male rhesus monkey (Macaca mulatta
). J Neuroendocrinol 2000;12:167-76.
Fitts JM, Klein RM, Powers CA. Comparison of tamoxifen and testosterone propionate in male rats: Differential prevention of orchidectomy effects on sex organs, bone mass, growth, and the growth hormone-IGF-I axis. J Androl 2004;25:523-34.
Yousofvand N, Zarei F, Ghanbari A. Exogenous testosterone, finasteride and castration effects on testosterone, insulin, zinc and chromium in adult male rats. Iran Biomed J 2013;17:49-53.