Is ferroptosis involved in ROS-induced testicular lesions in a varicocele rat model?

BACKGROUND: Ferroptosis is an iron-dependent cell death that is distinct from apoptosis. Based on excessive amounts of iron and reactive oxygen species in varicocele (VCL) rats, we hypothesize that ferroptosis might be involved in VCL. In addition, since alpha-lipoic acid (ALA) was shown to have both antioxidant and anti-ferroptotic activity we assessed in the present work the status of ferroptosis in our varicocele model and the protective effect of ALA. To this end, 70 male Wistar rats were divided into 7 groups: control, sham and varicocele groups which were initially sacrificed 2 months after the operation to verify the induction of varicocele. A second batch of the same 3 groups were sacrificed 4 months after varicocele induction to evaluate the effect of ALA supplementation. The parameters measured were chromatin integrity (aniline blue and acridine orange staining), lipid peroxidation (BODIPY staining), testicular morphometry and iron content. In addition, redox (GSH and NADPH) and ferroptosis (Nrf2, Slc7a11, P53 and p-Jnk) markers were evaluated at 2 and 4 months post-operation. RESULT: The alteration of the spermatic parameters made it possible to verify the induction of the varicocele. Iron accumulated well in the testicles during varicocele and decreased significantly following ALA treatment. Ferroptotic molecular markers at the mRNA and protein levels were not significantly altered. ALA supplementation did not alter NADPH values, but increased GSH levels. CONCLUSION: Despite the increased accumulation of iron in the testes 2 and 4 months after surgical induction of varicocele, molecular evidence did not demonstrate the involvement of ferroptosis. This could be explained by the mosaic nature of the varicocele affecting some seminiferous tubules and not others which could mask variations in molecular markers. In parallel, our study confirms that ALA stimulates the NRF2 pathway.

RéSUMé: CONTEXTE: La ferroptose est une mort cellulaire dépendante du fer qui est distincte de l'apoptose. Sur la base de quantités excessives de fer et d'espèces réactives de l'oxygène chez les rats varicocèles (VCL), nous avons fait l'hypothèse que la ferroptose pourrait être impliquée dans la VCL. Comme l'acide alpha-lipoïque (ALA) s'est avéré avoir des activités antioxydante et anti-ferroptotique nous avons dans ce travail testé le statut ferroptose dans notre modèle de rats VCL et évalué l'effet protecteur de ALA. Dans ce but, 70 rats mâles Wistar ont été divisés en plusieurs groupes: témoin, fictif et groupes varicocèle qui ont été initialement sacrifiés deux mois après l'opération pour vérifier l'induction de la varicocèle. Un deuxième lot des 3 mêmes groupes a été sacrifié 4 mois après l'induction de la varicocèle pour évaluer l'effet de la supplémentation en ALA. Les paramètres mesurés étaient l'intégrité de la chromatine (via les tests au bleu d'aniline et à l'orange acridine), la peroxydation lipidique (via le test BODIPY), la morphométrie testiculaire et la teneur en fer. De plus, les marqueurs redox (GSH et NADPH) et ferroptotique (Nrf2, Slc7a11, P53 et p-Jnk) ont été évalués 2 et 4 mois après l'opération. RéSULTATS: L'altération des paramètres spermatiques a permis de vérifier l'induction de la VCL. Le fer s'est bien accumulé dans les testicules pendant la VCL et a diminué de manière significative après le traitement ALA. Les marqueurs moléculaires ferroptotiques n'ont pas été modifiés de manière significative que ce soit en quantité d'ARNm et de protéines. La supplémentation en ALA n'a pas modifié la teneur en NADPH, mais a augmenté les niveaux de GSH. CONCLUSIONS: Malgré l'accumulation accrue de fer dans les testicules 2 et 4 mois après l'induction chirurgicale de la VCL, les investigations au niveau moléculaire n'ont pas clairement démontré l'implication de la ferroptose. Cela pourrait s'expliquer par la nature mosaïque de la VCL qui affecte certains tubules séminifères et pas d'autres, ce qui pourrait atténuer les évaluations réalisées sur organe entier. En parallèle, notre étude confirme que l'ALA stimule la voie NRF2 stimulant la réponse anti-oxydante.

Capsaicin improves sperm quality in rats with experimental varicocele.

Capsaicin is the main capsaicinoid in chilli peppers that have numerous biological and pharmaceutical roles in the body such as antioxidant, anti-inflammatory, anticarcinogenic, analgesic, counterirritant and antiarthritic properties. Numerous studies have shown increased oxidative stress in men with varicocele that is caused by dilation of the spermatic vein and increase of testicular temperature. Therefore, we aimed to assess the effect of Capsaicin on sperm parameters in rats with experimental varicocele. At first, we induced varicocele in 30 Wistar rats and, verify varicocele model only in 10 rats by assessment of sperm parameters, oxidative stress, DNA damage and persistent histone after 2 months. Of the remaining 20 varicocelised rats, half of them were treated with 2.5 mg/kg Capsaicin for two months and the other half served as control. Then, sperm tests were assessed, and the results showed that Capsaicin can restore the mean of sperm oxidative stress (38.78 ± 3.75 versus 58.37 ± 4.34; p < .05), sperm concentration (60.14 ± 7.66 versus 34.87 ± 5.78; p < .05) and motility (62.43 ± 3.10 versus 41.22 ± 5.11; p < .05) in varicocelised rats treated with Capsaicin compared to varicocelised rats that were not treat. Therefore, Capsaicin possibly with reduction of oxidative stress level could improve mean of sperm concentration and motility in varicocele condition.

Endoplasmic Reticulum Stress (ER Stress) and Unfolded Protein Response (UPR) Occur in a Rat Varicocele Testis Model.

Using a surgically induced varicocele rat model, we show here strong evidence that the misfolded/unfolded protein response that is part of the stress response of the endoplasmic reticulum (ER) is activated in the varicocele testis (VCL), leading to the induction of apoptosis. To support this hypothesis, it is observed that the spliced variant of the X-box protein 1 (XBP1s), resulting from the activation of the inositol-requiring enzyme 1 (IRE1) membrane sensor, is significantly more represented in VCL testicular extracts. The activation of the IRE1/XBP1s pathway is also supported by the observation that the VCL testes show an increase phosphorylation of the c-Jun-kinase (JNK) known to be one intermediate of this pathway and an increased level of caspase-3, the terminal apoptotic effector, partly explaining the apoptotic status of the VCL testis.

Alpha-Lipoic Acid improves the testicular dysfunction in rats induced by varicocele.

Alpha-lipoic acid (ALA) is a disulphide compound with multifunctional antioxidant properties and is soluble in both water and lipid. Several recent studies evaluated efficacy of ALA in various diseases related to oxidative damage such as diabetes, multiple sclerosis and Alzheimer and concluded that ALA can reduce oxidative stress by quenching reactive oxygen and nitrogen species, restoring antioxidants such as glutathione, vitamins C and E, and/or improving activity of antioxidant enzymes. Varicocele, an enlargement of the veins in scrotum, is considered as the most common repairable cause of male infertility and is associated with high levels of oxidative stress. In this study, surgical varicocele was induced in 30 adult male Wistar rats with other 20 rats serving as sham-operated and nonoperated control. Varicocele caused significant worsening of sperm parameters, DNA damage and lipid peroxidation 2 and 4 months after surgery. A 2-month ALA administration after surgery was able to revert these effects. These results clearly showed that ALA can reduce the negative side effects of elevated testicular temperature and increased oxidative stress in varicocelised rats. This study warrants future clinical research to assess whether ALA is of help in the treatment of infertile men with varicocele.

A Review Study on Punica granatum L.

Punica granatum L (pomegranate) is a deciduous shrub, native to Iran. Nowadays, besides its use as a fruit, its medicinal properties have attracted the interest of researchers of many countries. Pomegranate fruit has medicinal properties such as anti-inflammatory and antibacterial activities. The pomegranate seed oil has inhibitory effect on skin and breast cancers. The pomegranate seed oil has phytoestrogenic compounds and the fruit is rich in phenolic compounds with strong antioxidant activity. Ellagic acid is one of the main components of pomegranate with phenolic structure and antioxidant activity. This review article presents the recently published findings on different aspects of this plant focusing on its medicinal properties.