Testicular Ischemia-reperfusion Research Articles

TRapamycin reduces testicular ischemia-reperfusion injury by enhancing autophagy

Objectives To confirm the effects of autophagy on testicular ischemia-reperfusion (I/R) injury. Methods Forty rats were divided into sham group, I/R group, I/R+Rap (rapamycin, autophagy activator) group and I/R+ 3-MA (3-methyl adenine, autophagy inhibitor) group. Before inducing ischemia, rapamycin and 3-MA were intraperitoneally injected into I/R+Rap and I/R+ 3-ma groups, respectively. Subsequently, we then assessed testicular tissue damage. Immunohistochemistry was used to detect Beclin-1 and Caspase-3, while Western blot and qRT-PCR detected LC-II, Beclin-1 and p62. TUNEL and transmission electron microscopy were used to observe apoptosis and autophagosome. Results I/R activated autophagy in rat testicles. Rapamycin significantly improved testicular function after I/R by enhancing autophagy, reducing spermatogenic cell apoptosis, and decreasing testicular tissue damage scores. Conclusions Enhancing autophagy has a protective effect in ischemic-reperfusion injury by reducing apoptosis of rat testicular sperm cells.

Integrative analysis of transcriptomics and metabolomics reveals the protective effect and mechanism of salidroside on testicular ischemia-reperfusion injury

Testicular torsion is a critical urologic condition for which testicular detorsion surgery is considered irreplaceable as well as the golden method of reversal. However, the surgical treatment is equivalent to a blood reperfusion process, and no specific drugs are available to treat blood reperfusion injuries. Salidroside (SAL) is one of the main effective substances in rhodiola, which has been shown to have antioxidant and antiapoptosis activities. This study was designed to determine whether SAL exerted a protective effect on testicular ischemia-reperfusion (I/R) injury. In this study, the I/R injury model of the testes and reoxygenation (OGD/R) model were used for verification, and SAL was administered at doses of 100 mg/kg and 0.05 mmol/L, respectively. After the experiments, the testicular tissue and TM4 Sertoli cells were collected for histopathologic and biochemical analyses. The results revealed that SAL improves the structure of testicular tissue and regulates the oxidation–antioxidation system. To further understand the molecular mechanisms of SAL in treating testicular I/R injuries, transcriptomics and metabonomics analyses were integrated. The results show that the Nfr2/HO-1/GPX4/ferroptosis signaling pathway is enriched significantly, indicating that it may be the main regulatory pathway for SAL in the treatment of testicular I/R injuries. Thereafter, transfection with Nrf2 plasmid–liposome was used to reverse verify that the Nfr2/HO-1/GPX4/ferroptosis signaling pathway was the main pathway for SAL anti-testicular I/R injury treatment. Thus, it is suggested that SAL can protect against testicular I/R injuries by regulating the Nfr2/HO-1/GPX4 signaling pathway to inhibit ferroptosis and that SAL may be a potential drug for the treatment of testicular I/R injuries.

Protective effect of allicin on ischemia-reperfusion injury caused by testicular torsion-detorsion in rats.

Testicular ischemia-reperfusion induced by testicular torsion-detorsion increases the level of reactive oxygen species, leading to testicular damage. Allicin, one of the most active ingredients in garlic, is a significant exogenous antioxidant. In the research, the efficacy of allicin in treating testicular ischemia-reperfusion injury was assessed. The study included sixty Sprague-Dawley male rats. Three groups with 20 rats per group were created as follows: control group, testicular ischemia/reperfusion-induced group, and testicular ischemia-reperfusion plus treatment with allicin group. The control group underwent a sham operation of the left testis without other interventions. In the testicular ischemia/reperfusion-induced group, rat left testis was subjected to 720° torsion for two hours and then detorsion. In the allicin-treated group, in addition to testicular ischemia-reperfusion, 50 mg/kg of allicin was injected intraperitoneally, starting immediately following detorsion. Testicular tissue samples were obtained to measure the protein expression of xanthine oxidase, which is a major source of reactive oxygen species formation, malondialdehyde level (a reliable marker of reactive oxygen species), and testicular spermatogenic function. Testicular ischemia-reperfusion significantly increased the expression of xanthine oxidase and malondialdehyde levels in ipsilateral testes while reducing testicular spermatogenic function. The expression of xanthine oxidase and malondialdehyde levels were significantly lower in ipsilateral testes, whereas testicular spermatogenic function in the allicin-treated group was significantly higher compared with those in the testicular ischemia-reperfusion group. Our findings indicate that allicin administration improves ischemia/reperfusion-induced testicular damage by limiting reactive oxygen species generation via inhibition of xanthine oxidase expression.

Stem Leydig cells support macrophage immunological homeostasis through mitochondrial transfer in mice

As testicular mesenchymal stromal cells, stem Leydig cells (SLCs) show great promise in the treatment of male hypogonadism. The therapeutic functions of mesenchymal stromal cells are largely determined by their reciprocal regulation by immune responses. However, the immunoregulatory properties of SLCs remain unclear. Here, we observe that SLCs transplantation restore male fertility and testosterone production in an ischemia‒reperfusion injury mouse model. SLCs prevent inflammatory cascades through mitochondrial transfer to macrophages. Reactive oxygen species (ROS) released from activated macrophages inducing mitochondrial transfer from SLCs to macrophages in a transient receptor potential cation channel subfamily member 7 (TRPM7)-mediated manner. Notably, knockdown of TRPM7 in transplanted SLCs compromised therapeutic outcomes in both testicular ischemia‒reperfusion and testicular aging mouse models. These findings reveal a new mechanism of SLCs transplantation that may contribute to preserve testis function in male patients with hypogonadism related to immune disorders.

The potential prophylactic and therapeutic impacts of niacin on ischemia/reperfusion injury of testis

The testicular ischemia-reperfusion (I/R) injury is characterized by the excessive aggregation of un-scavenged reactive oxygen species, leading to the heightened levels of oxidative stress. This phenomenon plays a pivotal role in the pathophysiology of testicular torsion damage. The current study aimed to detect the prophylactic and therapeutic effects of niacin on testicular I/R injury. Twenty-four healthy adult male Sprague Dawley rats were randomly allocated into three groups as follows: (1) sham group, (2) torsion/detorsion (T/D) group, and (3) treatment group which received 200mg/kg niacin along with testicular T/D. Torsion/detorsion was induced by 2h of torsion followed by 10 days of reperfusion period. In the treatment group, niacin was injected 30min before the reperfusion period intraperitoneally and continued for 10 days by oral gavage. T/D was associated with marked decreases in terms of sperm count, viability, and kinematic parameters versus the sham group (P<0.05), which niacin significantly reverted the kinematic parameters (P<0.05). I/R injury caused a significant increase in the number of abnormal epididymal sperms compared to the sham group (P<0.05). Niacin decreased the epididymal sperm abnormality significantly compared to the T/D group (P<0.05). Tissue abnormalities in T/D group, such as edema, hyperemia, inflammation, and necrosis were completely visible histopathologically, while the histological changes in the niacin-treated group were better than those in the T/D group. Regarding the pathological parametric evaluations, I/R injury significantly reduced the mean testicular biopsy score (MTBS), germinal epithelial cell thickness (GECT), and mean seminiferous tubular diameter (MSTD), and increased the tubular hypoplasia/atrophy (THA) compared to the sham group (P<0.05), which niacin treatment significantly improved the MTBS and GECT compared to the T/D group (P<0.05). T/D significantly increased the oxidative stress index (OSI) and lipid peroxidation (MDA) (P<0.05). Niacin significantly reduced the OSI and MDA levels compared to the T/D group (P<0.05). The current study found that niacin has preventive/therapeutic effects against the elevation of oxidative stress markers and depletion of antioxidants during I/R injury. Following administration of niacin, a reduction in histologic injury was observed in rats. In our study, we showed the antioxidant properties of niacin and its capacity to protect against I/R damage. The findings of the present investigation revealed that niacin, as an antioxidant agent, can suppress the oxidative stress induced by testicular I/R injury, and can be used as a supplementary agent in the treatment of those undergoing testicular torsion surgery.

Protective Effects of Diospyros Kaki Leaves Extract against Oxidative Damage and Inflammation in the Testicular Ischemia-Reperfusion Model in Rats

Protective Effects of Diospyros Kaki Leaves Extract against Oxidative Damage and Inflammation in the Testicular Ischemia-Reperfusion Model in Rats

Oxidative stress, apoptosis, inflammation, and proliferation modulator function of visnagin provide gonadoprotective activity in testicular ischemia-reperfusion injury.

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The Regulatory Effects of JAK2/STAT3 on Spermatogenesis and the Redox Keap1/Nrf2 Axis in an Animal Model of Testicular Ischemia Reperfusion Injury.

The male reproductive system requires the pleiotropic activity of JAK/STAT to maintain its function, especially spermatogenesis. The study aims to investigate the effect of JAK2 signaling on the expression of the Keap1/Nrf2 axis, spermatogenesis, and the Sertoli cells (Sc) junctions in an animal model of testicular ischemia reperfusion injury (tIRI). Testes subjected to tIRI exhibited increased JAK2/STAT3 activity associated with spermatogenic arrest and reduced expression of the Sc junctions. In addition, there was an increased protein expression of Keap1 and decreased Nrf2., which was coupled with the downregulation of gene expression of antioxidant enzymes. Reduced SOD and CAT activities were accompanied by increased lipid peroxidation and protein carbonylation during tIRI. Increased caspase 9 activity and Bax/Bcl2 ratio indicated initiation of apoptosis. Inhibition of JAK2 activity by AG490 maintained the integrity of spermatogenesis and SC junctions, normalized the expression of the Keap1/Nrf2 axis and its downstream antioxidant enzymes, and prevented germ cell apoptosis. The results further emphasized the regulatory role of JAK2/STAT3 on spermatogenesis, Keap1/Nrf2 signaling, and maintenance of the testicular redox balance to combat testicular dysfunction and male infertility.

Expression of concern: Involvement of mitogen-activated protein kinases (MAPKs) during testicular ischemia–reperfusion injury in nuclear factor-κB knock-out mice [Life Sci. 81 (2007) 413–422

Expression of concern: Involvement of mitogen-activated protein kinases (MAPKs) during testicular ischemia–reperfusion injury in nuclear factor-κB knock-out mice [Life Sci. 81 (2007) 413–422

Dimethyl Fumarate Protects Rats against Testicular Ischemia–Reperfusion Injury

Ischemia–reperfusion injury (IRI) after testicular torsion is linked to significant damage in testicular tissue. Dimethyl fumarate (DMF) activates the nuclear factor erythroid 2-related factor 2 (Nrf2) signaling pathway, thereby inducing antioxidant and anti-inflammatory effects. We analyzed the usefulness of DMF in preventing IRI following testicular torsion/detorsion in Sprague-Dawley rats ( n = 32 ). The animals were classified into control (sham), DMF (200 mg/kg/day), IRI, and IRI+DMF (IRI with 200 mg/kg/day DMF) groups. Testicular IRI was induced by detorsion after 1.5 h of torsion. DMF was administered via oral gavage daily from 1 h before testicular detorsion until day 7, when orchiectomy was performed. The testis-to-body weight ratio was calculated. Histopathological evaluation was performed using the Johnsen and Cosentino scores for seminiferous tubules. Malondialdehyde, superoxide dismutase, and total glutathione levels were determined in testicular tissues. Moreover, Nrf2, heme oxygenase 1 (HO-1), NAD(P)H quinone dehydrogenase 1 (NQO1), nuclear factor kappa B (NF-κB), and inflammatory cytokine (interleukin 1b (IL1b), IL6, and tumor necrosis factor alpha (TNF-α)) levels were determined through quantitative polymerase chain reaction. Nuclear Nrf2 and cytoplasmic HO-1 and NQO1 protein levels were also evaluated. DMF significantly improved the testis-to-body weight ratio and reduced histopathological damage in the testes. Moreover, it significantly improved the concentration of malondialdehyde, superoxide dismutase, and total glutathione. Furthermore, it inhibited NF-κB and inflammatory cytokine mRNA expression compared with the findings obtained in untreated rats with IRI (all p &lt; 0.05 ). Nrf2, HO-1, and NQO1 expressions (mRNA and protein) were markedly elevated following DMF treatment in rats with IRI (all p &lt; 0.05 ). DMF administration activated the Nrf2 signaling pathway and induced antioxidant and anti-inflammatory effects, thereby improving IRI-induced testicular damage. Thus, DMF may prevent IRI following testicular torsion.

Exosomes obtained from adipose mesenchymal stem cells prevent ischemia-reperfusion injury after torsion-detorsion in rat testes.

To investigate the effect of exosomes obtainedfrom adipose-derived mesenchymal stem cells (AD-MSCs) on testicular ischemia-reperfusion (I/R) injury. AD-MSCs from rat adipose tissue were cultured. Characterization of cells was evaluated with CD44, CD90, CD34 and CD45 antibodies. Exosomes from AD-MSCs were obtained with the miRCURYexosomeisolation kit. 21 rats were divided into 3 groups. The I/R model was created as 720° torsion for 4h and reperfusion for 4h. In the Sham group (SG), only scrotal incision was made. 100µl of medium in the torsion-control group (T-CG) and 100µl of exosome in the treatment group (TG) were injected into the testicular parenchyma after detorsion. Johnsen scores of testicles were determined. Apoptosis was evaluated by the TUNEL method. It was observed that the seminiferous tubule structures were partially disrupted in T-CG, but normal in SG and TG. Johnsen scores in SG, T-CG, and TG were 8.64 ± 0.39, 7.71 ± 0.37, and 8.57 ± 0.39, respectively. Apoptotic cell distribution was 11.28 ± 5.25%, 60.58% ± 1.68% and 17.71 ± 8.34% in SG, T-CG and TG, respectively. In both parameters, the difference between SG and TG was insignificant (p > 0.05), the difference between T-CG/TG and SG/T-CG was significant (p < 0.05). Exosomes obtainedfromAD-MSCs are effective in preventing testicular I/R injury. This effect appears to occur because of suppression of apoptotic activity.

Histological study on the possible protective role of L-carnitine against testicular ischemia-reperfusion injury induced in rabbits

Histological study on the possible protective role of L-carnitine against testicular ischemia-reperfusion injury induced in rabbits

Emodin alleviates testicular ischemia-reperfusion injury through the inhibition of NLRP3-mediated pyroptosis

Ischemia-reperfusion injury (IRI) is a major cause of injury after testicular torsion and can lead to permanent impairment of spermatogenesis. Emodin (6-methyl-1,3,8-trihydroxyanthraquinone) has potent anti-inflammatory effects and may be protective against IRI in various organs. Herein, we evaluated the effects of emodin on pyroptosis in spermatogenic cells and its role in the process of testicular IRI. A testicular torsion/detorsion (TTD) mouse model and an oxygen-glucose deprivation/reperfusion (OGD/R) germ cell model were established. Hematoxylin and eosin staining was performed to evaluate the testicular ischemic injury. The expression of pyroptosis-related proteins and reactive oxygen species production in testis tissues were detected using Western blotting, quantitative real-time PCR, malondialdehyde and superoxide dismutase assay kits and immunohistochemistry. Cell viability and cytotoxicity were evaluated using Cell Counting Kit-8 and lactate dehydrogenase assay kit. Enzyme-linked immunosorbent assay, immunofluorescence and immunoblotting were performed to assess inflammatory protein levels. The results revealed that pyroptosis and inflammation levels were upregulated after testicular IRI, and emodin inhibited inflammation and pyroptosis by acting on NOD-like receptor thermal protein domain-associated protein 3 (NLRP3). Emodin exerts protective effects on testicular IRI by acting on the NLRP3 signaling pathway and inhibiting IRI-mediated pyroptosis. Emodin treatment attenuated testicular IRI and inhibited pyroptosis. Inhibitory effects of emodin on pyroptosis were attributed to the inhibition of NLRP3 inflammasomes. Thus, emodin could be an alternative treatment for testicular IRI.

Pathological analysis of spermatic dysfunction following testicular ischemia-reperfusion injury

Pathological analysis of spermatic dysfunction following testicular ischemia-reperfusion injury

NADPH Oxidase-Mediated Testicular Oxidative Imbalance Regulates the TXNIP/NLRP3 Inflammasome Axis Activation after Ischemia Reperfusion Injury.

Oxidative stress, inflammation and germ cell death are the main characteristics of testicular ischemia reperfusion injury (tIRI), which is considered as the underlying mechanism for testicular torsion and detorsion. The study aimed to examine the effect of tIRI-activated NADPH oxidase (NOX) on the expression of the NLRP3 inflammasome pathway components. Three groups of male Sprague-Dawley rats (n = 12 each) were studied: sham, unilateral tIRI only and tIRI treated with apocynin, a NOX-specific inhibitor. The tIRI rat model was subjected to 1 h of ischemia followed by 4 h of reperfusion. H&E staining, real time PCR, biochemical assays, and Western blot were utilized to evaluate spermatogenic damage, gene expression, oxidative stress markers, and NLRP3 pathway components, respectively. As a result of tIRI, decreased total antioxidant capacity and suppressed activities of superoxide dismutase and catalase were associated with spermatogenic arrest. The components of the NLRP3 inflammasome pathway (TXNIP, NLRP3, ASC, caspase-1, GSDMD, MMP-9) were upregulated transcriptionally and post-transcriptionally during tIRI. In parallel, tissue inflammation was demonstrated by a marked increase in the concentrations of myeloperoxidase, IL-1β, and IL-18. Apocynin treatment prevented testicular oxidative stress and inflammation. Thus, NOX inhibition by apocynin prevented ROS accumulation, proinflammatory cytokine overexpression and NLRP3 inflammasome activation during tIRI.

Effects of Titanium Dioxide Nanoparticles and Coenzyme Q10 on Testicular Ischemia-Reperfusion Injury: Role of the Mitochondrial Apoptosis Pathway

Background: Testicular ischemia-reperfusion (I/R) injury is a urological emergency that can lead to male infertility. So far, no suitable treatment has been found for it. Therefore, in the present study, we investigate the therapeutic effects of concomitant administration of coenzyme Q10 (CoQ10) and titanium dioxide nanoparticles (TiO2-NPs) on testicular I/R damage in rats and the expressions of genes involved in mitochondrial apoptosis, miR-21, and circRNA0001518. Materials and Methods: In this study, after induction of testicular torsion/detorsion, CoQ10 and TiO2-NPs were administered to the rats for ten days. Then, sperm extracted from the epididymides were analyzed for concentration, viability, morphology, and motility. The amount of apoptosis in testicular cells was studied by flow cytometry. Also, the expressions of the Bax and Bcl-2 genes, as well as miR-21 and circRNA0001518 levels were evaluated. Results: Sperm parameters improved in the rats’ testicular that received CoQ10. Administration of TiO2-NPs to healthy rats increased apoptosis and the Bax/Bcl-2 expression ratio. However, its administration to testicular I/R rats alone or in combination with CoQ10 caused a decrease in apoptosis, the Bax/Bcl-2 expression ratio, and an increase in miR-21 and circRNA0001518 expressions. Conclusion: Overall, individual or joint administration of TiO2-NPs or CoQ10 can have therapeutic effects on testicular I/R by altering the expressions of genes in the mitochondrial apoptotic pathway and their regulatory elements.

Role of antioxidants in preventing testicular ischemia‑reperfusion injury: a narrative review.

Testicular ischemia-reperfusion injury (TIR) is a urological emergency common among male newborns, children, and adolescents. Testicular injury and its consequences, such as altered hormone production, subfertility, and infertility, are determined by the duration and degree of testicular torsion. Early diagnosis and treatment are crucial to preserving the testes and fertility. Previous studies suggest that reactive oxygen species contribute to the pathogenesis of TIR injury, but the underlying mechanism remains unclear. Several drugs/plants reportedly exhibit antioxidative activities to protect against TIR. This review summarizes current studies on the role of antioxidants in preventing experimental TIR injury and discusses the underlying pathophysiological mechanisms.

Baicalein Alleviates Testicular Ischemia-Reperfusion Injury in a Rat Model of Testicular Torsion-Detorsion.

Testicular torsion/detorsion-induced ischemia/reperfusion injury is partly due to the overgeneration of reactive oxygen species. Baicalein, a main bioactive constituent derived from the dried root of Scutellaria baicalensis Georgi, possesses powerful antioxidative and anti-inflammatory properties. Therefore, we designed the research to explore the possible protective effect of baicalein against testicular ischemia-reperfusion injury. Sprague-Dawley rats were randomized into 4 groups, including control, testicular ischemia-reperfusion, testicular ischemia-reperfusion+vehicle injection, and testicular ischemia-reperfusion+baicalein therapy groups. The control group received surgical exposure of the left testis without torsion-detorsion. In the testicular ischemia-reperfusion group, the left testis underwent 720° counterclockwise torsion for two hours and then was allowed detorsion. Rats in the testicular ischemia-reperfusion+vehicle injection group received intraperitoneal injection of the vehicle at detorsion. In the baicalein-treated group, the intraperitoneal administration of baicalein dissolved in the vehicle was performed at detorsion. At four hours or three months following testicular detorsion, testicular tissues were removed to detect the levels of tumor necrosis factor-alpha (TNF-α) and interleukin-1beta (IL-1β) which can recruit neutrophils into the testis, myeloperoxidase activity (an index of neutrophil infiltration in the testis), protein expression of nicotinamide adenine dinucleotide phosphate (NADPH) oxidase in neutrophils which can catalyze reactive oxygen species production, malondialdehyde concentration (a common marker of reactive oxygen species), and spermatogenesis. Both testicular ischemia-reperfusion and testicular ischemia-reperfusion+vehicle injection significantly increased the TNF-α and IL-1β levels, myeloperoxidase activity, NADPH oxidase protein expression, and malondialdehyde concentration, while decreased spermatogenesis in ipsilateral testes. In contrast, baicalein administration remarkably reduced TNF-α and IL-1β levels, myeloperoxidase activity, NADPH oxidase protein expression, and malondialdehyde concentration and also elevated spermatogenesis in ipsilateral testes. The results of our experiment demonstrate that baicalein alleviates testicular ischemia-reperfusion injury by inhibiting TNF-α and IL-1β secretion, neutrophil infiltration in the testis, and NADPH oxidase protein expression in neutrophils to reduce reactive oxygen species production.

NOX-activated NLRP3 Inflammasome Pathway Triggers Pyroptosis During Testicular Ischemia Reperfusion Injury

NOX-activated NLRP3 Inflammasome Pathway Triggers Pyroptosis During Testicular Ischemia Reperfusion Injury

Targeting soluble guanylate cyclase with Riociguat has potency to alleviate testicular ischaemia reperfusion injury via regulating various cellular pathways.

Testicular ischaemia reperfusion (I/R) injury results with serious dysfunctions in testis. This study aims to explore effects of soluble guanylate cyclase (sGC) stimulator Riociguat on experimental testicular I/R injury in rats. Twenty-one male rats were divided into three groups (Control, IR and IRR). The control group was not exposed to any application. Bilateral testis from IR and IRR animals were rotated 720° in opposite directions for 3 h to induce experimental testicular ischaemia. Animals in IR and IRR groups were subjected to 3 h of reperfusion. Isotonic and Riociguat were administered to the animals 30 min prior reperfusion by oral gavage. At the end of experiment, animals were sacrificed and tissue samples were used for analyses. Riociguat treatment significantly decreased tissue malondialdehyde and Luminol levels compared to the IR group (p < 0.05). The pathological changes, pro-apoptotic proteins (Bax, Caspase 3, and Caspase 9) and apoptotic index in the IR group were down regulated in Riociguat treated animals (p < 0.05). Riociguat treatment was also significantly increased anti-apoptotic Bcl-2 expression, but alleviated tissue injury via modulating pro-inflammatory cytokine IL-1β levels and significantly (p < 0.05) down-regulating NF-κB activity. Moreover, mTOR and ERK phosphorylation increased in IR group (p < 0.05), but Riociguat treatment reduced protein phosphorylation. Our experiment indicated that targeting sGC might support surgical interventions in testicular I/R injury by modulating oxidative stress, inflammation, and apoptotic protein expression levels, but more detailed studies are required to explore the protective activity of Riociguat and underlying mechanisms in testicular I/R injury.