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Mono-2-ethylhexyl phthalate (MEHP) exposure is known to induce severe testicular injury via reactive oxygen species (ROS). However, few effective treatments are available for the precise treatment of MEHP-induced germ cell damage. Epigallocatechin gallate (EGCG), one of the major polyphenols in green tea, has potential antioxidant activity and can alleviate many diseases induced by oxidative stress. This study explored whether EGCG protects germ cells from MEHP-induced oxidative stress damage. Cells were treated with 400 μM MEHP and 60 μM EGCG for 24 h. EGCG reduced MEHP-induced ROS overgeneration in the spermatogonial cell line GC-1 and spermatocyte cell line GC-2. Western blotting and immunofluorescence showed that the MEHP+EGCG group exhibited lower nuclear factor (erythroid-derived 2)-like 2 (NRF2), heme oxygenase (decycling) 1 (HO-1), and superoxide dismutase (SOD) expression than the MEHP group. Moreover, activation of the mammalian target of rapamycin (mTOR) pathway was decreased. The expression of key factors of pyroptosis was downregulated, and interleukin-10 (IL-10) expression was reduced. Additionally, apoptosis was inhibited by EGCG. The findings indicate that EGCG protects against MEHP-induced germ cell pyroptosis by scavenging ROS, suppressing the mTOR pathway, and inhibiting pyroptosis. EGCG may thus be a potential treatment for MEHP-related spermatogenic dysfunction., Competing Interests: Declaration of Competing Interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2023 Elsevier Ltd. All rights reserved.)

Most children with a neurogenic bladder (NB) have bladder fibrosis, which causes irreversible bladder dysfunction and damage to the upper urinary tract. However, the mechanism of bladder fibrosis remains unclear. This study aimed to investigate the underlying causes of bladder fibrosis. Here, the lumbar 6 (L6) and sacral 1 (S1) spinal nerves of Sprague Dawley rats were severed bilaterally to establish NB models. Using RNA-seq, we discovered that the NF-κB signaling pathway and inflammation were upregulated in spinal cord injury (SCI)-induced bladder fibrosis. Subsequent Western blotting, enzyme-linked immunosorbent assays, immunohistochemical staining, and immunofluorescence staining verified the RNA-seq findings. To further clarify whether the NF-κB signaling pathway and pyroptosis were involved in bladder fibrosis, a TGF-β1-treated urinary epithelial cell line (SV-HUC-1 cells) was used as an in vitro model. Based on the results of RNA-seq, we consistently found that the NF-κB signaling pathway and pyroptosis might play important roles in TGF-β1-treated cells. Further experiments also confirmed the RNA-seq findings in vitro. Moreover, using the NLRP3 inhibitor MCC950 rescued TGF-β1-induced fibrosis, and the NF-κB signaling pathway inhibitor BAY 11-7082 effectively rescued TGF-β1-induced pyroptosis and the deposition of extracellular matrix by SV-HUC-1 cells. In summary, our research demonstrated for the first time that the NF-κB signaling pathway inhibition rescued bladder epithelial cells pyroptosis and fibrosis in neurogenic bladders. [ABSTRACT FROM AUTHOR]

Background: Neurogenic bladder (NB) patients exhibit varying degrees of bladder fibrosis, and the thickening and hardening of the bladder wall induced by fibrosis will further affect bladder function and cause renal failure. Our study aimed to investigate the mechanism of bladder fibrosis caused by a spinal cord injury (SCI). Methods: NB rat models were created by cutting the bilateral lumbar 6 (L6) and sacral 1 (S1) spinal nerves. RNA-seq, Western blotting, immunofluorescence, cell viability and ELISA were performed to assess the inflammation and fibrosis levels. Results: The rats showed bladder dysfunction, upper urinary tract damage and bladder fibrosis after SCI. RNA-seq results indicated that hypoxia, EMT and pyroptosis might be involved in bladder fibrosis induced by SCI. Subsequent Western blot, ELISA and cell viability assays and immunofluorescence of bladder tissue confirmed the RNA-seq findings. Hypoxic exposure increased the expression of HIF-1α and induced EMT and pyroptosis in bladder epithelial cells. Furthermore, HIF-1α knockdown rescued hypoxia-induced pyroptosis, EMT and fibrosis. Conclusion: EMT and pyroptosis were involved in the development of SCI-induced bladder fibrosis via the HIF-1α pathway. Inhibition of the HIF-1α pathway may serve as a potential target to alleviate bladder fibrosis caused by SCI. [ABSTRACT FROM AUTHOR]

As the most abundantly used phthalate derivative, di-(2-ethylhexyl) phthalate (DEHP) leads to reproductive disorders, especially in males. Testicular injury can be triggered when the testis is exposed to DEHP during the immature stage. However, the potential mechanism is largely unclear. In the present study, Sprague-Dawley rats were exposed to 0, 250 and 500 mg/kg/day DEHP from postnatal day (PND) 20 to PND 30. The spermatogonia cell line GC-1 and spermatocyte cell line GC-2 were exposed to different doses of monoethylhexyl phthalate (MEHP), a metabolite of DEHP. Testicular injury was observed. Oxidative stress was evaluated both in vivo and in vitro. Our results showed that after DEHP exposure, the testicular structure was damaged and spermatogenesis was disturbed. We also found that oxidative stress was increased, as indicated by the upregulation of the important factors in the antioxidant pathway. Furthermore, the expression of autophagy-related proteins was significantly downregulated. Autophagy inhibition led to activation of the pyroptosis pathway. Nucleotide-binding and oligomerisation (NOD) domain-like receptor (NLR) family pyrin domain (PYD)-containing 3 (NLRP3), Caspase-1 and cytokine interleukin-1β (IL-1β) were significantly upregulated. Additionally, an imbalance in self-renewal and differentiation was observed in germ cells after DEHP exposure, causing the cessation of germ cell development. In summary, these data suggest that DEHP exposure enhances oxidative stress, downregulates autophagy, induces NLRP3 inflammasome activation and subsequently triggers pyroptosis in vivo and in vitro, which provides novel insight into DEHP-related injury in immature testes in the context of pyroptosis. [Display omitted] • Exposure to DEHP could induce reproductive injury in immature testes. • The ROS/mTOR/NLRP3 signaling pathway participates in testicular injury induced by DEHP exposure. • DEHP exposure also increases the level of self-renewal and differentiation of GC-1 and GC-2 cells. [ABSTRACT FROM AUTHOR]