Attenuation Effect of Salvianolic Acid B on Testicular Ischemia-Reperfusion Injury in Rats

During testicular ischemia-reperfusion, overproduction of reactive oxygen species is associated with testicular injury. We injected hydrogen peroxide (a representative of reactive oxygen species) into normal testis via the testicular artery. The experiment demonstrates that reactive oxygen species can cause spermatogenic injury. Salvianolic acid B, the most abundant bioactive component in Salvia miltiorrhiza Bunge, has been reported to possess a potent antioxidant activity. This study was conducted to evaluate the effect of salvianolic acid B on testicular ischemia-reperfusion injury in a rat testicular torsion-detorsion model. Rats were randomly separated into three groups, including 20 rats in each group: control group with sham operation, testicular ischemia-reperfusion group, and testicular ischemia-reperfusion + salvianolic acid B-treated group. In the testicular ischemia-reperfusion group, left testicular torsion of 720° for 2 hours was induced, and then testicular detorsion was carried out. Rats in the salvianolic acid B-treated group additionally had salvianolic acid B administered intravenously at detorsion. At 4 hours after detorsion, testes of 10 rats from each group were collected to analyze the protein expression of xanthine oxidase which catalyzes generation of reactive oxygen species and malondialdehyde concentration (an indirect indicator of reactive oxygen species). At 3 months after detorsion, testes of the remaining 10 rats from each group were collected to analyze spermatogenesis. Compared with the control group, xanthine oxidase protein expression and malondialdehyde concentration in ipsilateral testes of testicular ischemia-reperfusion group increased significantly, while spermatogenesis decreased significantly. In the salvianolic acid B-treated group, xanthine oxidase protein expression and malondialdehyde concentration in ipsilateral testes decreased significantly, while spermatogenesis increased significantly, compared with the testicular ischemia-reperfusion group. These results suggest that salvianolic acid B can attenuate testicular torsion/detorsion-induced ischemia/reperfusion injury by downregulating the xanthine oxidase protein expression to inhibit reactive oxygen species formation.

Curcumin Along With Fe3O4 Nanoparticles Improved Sperm Parameters In Rats With Testicular Ischemia

Background: Ischemic/reperfusion (I/R) in testicular tissue is one reason for the worldwide increase in male infertility. In the present study, we assessed the effects of curcumin and Fe3O4 nanoparticles (NPs) on sperm parameters in rats with I/R damage. Materials and Methods: Forty-eight adult male rats were divided into two groups (n=24 per group): control and torsion/detorsion. The control and torsion/detorsion groups were divided into four subgroups include sham, Fe3O4 NPs, curcumin, and Fe3O4 NPs+curcumin. After the rats were sacrificed, semen was collected from their epididymal tissues to assess sperm viability, motility, concentration, and morphology. Results: Curcumin significantly improved viability, motility, and normal sperm morphology in rats with I/R damage compared to the control group; however, it did not have a significant effect on sperm concentration (P<0.001). Fe3O4 NPs alone decreased all sperm parameters in the control and I/R rats (P<0.001). However, concomitant administration of Fe3O4 nanoparticles with curcumin significantly improved sperm parameters in rats with I/R damage (P<0.001). Conclusion: The increase in all semen parameters in the experimental groups with concomitant use of Fe3O4 NPs plus curcumin indicated that green synthesis of NPs could be recommended for future clinical studies.

Impact of NiO2 Nanoparticles and Curcumin on Testis Torsion/Detorsion Injury: Role of miR-34 and circRNA 0001518

Background: Testicular torsion is one cause of infertility without proper treatment. In this study, we investigate the effects of NiO2 nanoparticles (NPs) and curcumin on sperm parameters in rats and the expressions of genes involved in the apoptotic pathway, as well as expressions of miR-34 and circRNA 0001518. Materials and Methods: Forty-eight rats were randomly divided into eight groups: control (healthy rats), control rats that received NiO2-NPs, healthy rats that received curcumin, rats that received simultaneous NiO2-NPs and curcumin, untreated testicular ischemia/reperfusion (I/R) rats, testicular I/R rats that received NiO2-NPs, testicular I/R rats that received curcumin, and testicular I/R rats that received NiO2-NPs and curcumin. Then, sperms were extracted from the rats’ epididymides to analyze concentration, viability, morphology, and motility. The cellular apoptosis level was studied using flow cytometry. Also, Bad and Bcl-X gene expressions, as well as miR-34 and circRNA 0001518 levels were measured. Results: We observed improved sperm parameters in the testicular I/R) rats that received curcumin and NiO2-NPs. Administration of NiO2-NPs to healthy rats increased both apoptosis and the Bad/Bcl-X expression ratio. However, its administration to testicular I/R rats alone or in combination with curcumin decreased apoptosis and the Bad/Bcl-X expression ratio and increased expressions of miR-34 and circRNA 0001518. Conclusion: Administration of NiO2-NPs and curcumin, alone or in combination, can have therapeutic effects in testicular I/R conditions by altering the expressions of genes in the mitochondrial apoptotic pathway and their regulatory elements.

EARLY REHABILITATION OF CRITICAL TESTICULAR ISCHEMIA WITH ACUTE TORSION IN CHILDREN

Introduction. Acute testicular torsion is an urgent condition characterized by chorda spermatica turn and torsion with its vessels along vertical or horizontal axis. This condition results in testicular ischemia and loss of organ in case of lack of timely medical care. If a child is operated on within 6-12 hours their chance for complete recovery is decreased to 70 %. The timeline of 12-24 hours leaves only a 20% chance to keep a testicle. After 24 hours since the beginning of the condition there is virtually no chance to keep organ’s viability. The goal of the research is an optimization of early rehabilitation of critical testicular ischemia in children with acute testicular torsion. Material and methods. Over the period of the years 2010 – 2021 54 children with testicular torsion were observed and treated. Eight children got to a hospital as in-patients within first six hours from condition establishment, eight patients turned to a hospital within the period of 6.1-12 hours, 11 patients addressed hospital after 12.1-16 hours, 16 patients did so after 16.1-24 hours, and 11 patients addressed hospital after more than 24 hours since condition had been established. All patients with testicular torsion were admitted to a hospital in an urgent manner, their general condition was assessed as "moderate". Time before surgery was limited (up to 1 hour from the moment of hospitalization). Therefore, the examination of patients was minimized by a general blood test, a general urine test (83.3 %), determination of blood group and Rh-factor, measuring arterial pressure. A testicle was verified as viable in 12 children (22.2 %). Genital gland necrosis was diagnosed in 22 patients (40.8 %), they underwent orchophuniculectomia. Critical testicle ischemia was diagnosed in 20 boys (37 %). All patients were operated on. In all cases operation started not later than an hour after hospitalization. For 45 children operation was performed via inguinal access (83.3 %), transscrotal access was used in 9 patients (16.7 %). In all cases when a testicle was viable invasive detorsion with further orchiopexy was used. All children who underwent testiculectomy were hospitalized later than 16 hours after the onset of the disease. All boys with critical testicle ischemia underwent rehabilitation measures during operation. Results. Among children with genital gland critical ischemia 10 patients (52.6 %) recovered completely. Children who had been hospitalized during 6-12 hours after the onset of the disease were discharged from a hospital ward on the 5-7 day after operation. Two children (25 %) in this group had further testicle autolysis. Among children who addressed for healthcare support within 12–16 hours after the condition establishment, two patients (18.2 %) recovered completely. Five patients (50 %) had testicle autolysis in the postoperative period. Conclusions. 1. Critical testicular ischemia is observed in 35.2 % children with acute testicular torsion. 2. Early rehabilitation measures include a complex of conservative and operational approaches implemented in pre-operational, intra-operational and early post-operational periods. 3. Timely and full-fledged rehabilitation measures implementation allows to improve results of operational treatment and save affected genital gland with critical ischemia in 50 % of patients.

JAK Inhibition Prevents DNA Damage and Apoptosis in Testicular Ischemia-Reperfusion Injury via Modulation of the ATM/ATR/Chk Pathway

Testicular ischemia reperfusion injury (tIRI) causes oxidative stress-induced DNA damage leading to germ cell apoptosis (GCA). The aim of the study is to establish a direct link between JAK2 activation and the DNA damage response (DDR) signaling pathways and their role in tIRI-induced GCA using AG490, a JAK2 specific inhibitor. Male Sprague Dawley rats (n = 36) were divided into three groups: sham, unilateral tIRI and tIRI + AG490 (40 mg/kg). During tIRI, augmentation in the phosphorylation levels of the JAK2/STAT1/STAT3 was measured by immunohistochemistry. Observed spermatogenic arrest was explained by the presence of considerable levels of DSB, AP sites and 8OHdG and activation of caspase 9, caspase 3 and PARP, which were measured by colorimetric assays and TUNEL. The ATM/Chk2/H2AX and ATR/Chk1 pathways were also activated as judged by their increased phosphorylation using Western blot. These observations were all prevented by AG490 inhibition of JAK2 activity. Our findings demonstrate that JAK2 regulates tIRI-induced GCA, oxidative DNA damage and activation of the ATM/Chk2/H2AX and ATR/Chk1 DDR pathways, but the cell made the apoptosis decision despite DDR efforts.

Leech therapy (Hirudo medicinalis) attenuates testicular damages induced by testicular ischemia/reperfusion in an animal model

Background Testicular torsion/detorsion triggers tissue ischemia/reperfusion, leading to reactive oxygen species overgeneration and apoptosis. The saliva of leeches is full of anti-inflammatory, anticoagulants, antioxidants, and antimicrobial agents. Therefore, this study aimed to assess the protective mechanism of leech therapy on testicular ischemia/reperfusion damage. Methods 18 adult male rats were randomly divided into three groups: 1-Sham-operated group (SO). 2-Torsion/detorsion (T.D) group: two hours of testicular torsion with two hours of testicular detorsion was performed. 3-Torsion/detorsion + Leech therapy (TDL) group. Sperm parameters (motility, vitality, morphology, and concentration), oxidative stress biomarkers (MDA, CAT, GPx, and TAC), histopathological factors (Mean seminiferous tubular diameter, Germinal epithelial cell thickness, Testicular capsule thickness, Johnson’s score, and Cosentino’s score), and immunohistochemical markers for apoptosis detection (Bax, Bcl-2, and Caspase-3) were measured. Results There was a significant difference for all sperm parameters in the T. D group compared to the sham group. Leech therapy significantly increased progressive motility and normal morphology and reduced non-progressive motility. In the TDL group, MDA concentration significantly reduced, and levels of GPx, TAC, and CAT remarkably increased. All evaluated histopathological parameters in the TDL group significantly increased compared to the T. D group except for the testicular capsule thickness. T. D notably increased the expression of Bax and Caspase-3, while the treatment group slowed the rate of apoptosis compared to the control group. Bcl-2 expression in the T. D group was significantly lower than that in the sham group. Leech therapy increased the Bcl-2 expression. Conclusion Leech therapy attenuates damages to testicular tissue following torsion/detorsion due to its antioxidant, anti-inflammatory, and anti-apoptotic effects. Hence, it can be considered as an effective remedy for testicular ischemia/reperfusion. Graphical abstract