BackgroundThe transplantation of adipose-derived stem cells (ASCs) is a most promising treatment for diabetic erectile dysfunction (DMED). However, the effect of high glucose on the post-transplantation survival of stem cells limits the efficacy of ASCs transplantation. Prolonging the survival time of ASCs in vivo after transplantation is a key issue in the utilization of ASCs for DMED. Herein, we aimed to investigate the therapeutic effect of ASCs by downregulating NOD-, LRR-, and pyrin domain-containing protein 3 (NLRP3) as well as its mechanism of action in DMED.MethodsASCs were obtained by isolating subcutaneous fat from SD rats and were identified using lipogenic and osteogenic differentiation assays, as well as flow cytometric analysis. The shNLRP3 lentivirus with the best downregulating effect was screened, and shNLRP3 lentivirus (LV-shNLRP3) was transfected into ASCs (ASCsshNLRP3) to detect apoptosis and the reactive oxygen species (ROS) levels in each group under high glucose conditions. In DMED rats, ASCsLV-shNLRP3, ASCsLV-control, or phosphate buffered saline (PBS) were administrated via intra-cavernous injection, and normal rats served as normal controls. One week post-injection, animal imaging was performed to track the ASCs. Four weeks post-injection, erectile function was evaluated by measuring the intra-cavernosal pressure and mean arterial pressure. Corpus cavernosum pyroptosis and endothelial function were examined by western blotting and immunofluorescence.ResultsNLRP3-mediated pyroptosis might be a pathogenic mechanism of ED and DMED. ASCs were isolated successfully. Thereafter, the LV-shNLRP3 with the highest transfection efficiency was selected and used to modify ASCs successfully. LV-shNLRP3 could protect ASCs paracrine function under hyperglycemia through anti-apoptosis and anti-ROS deposition mechanisms. Furthermore, ASCsLV-shNLRP3 showed an advantage in the suppression of pyroptosis compared to ASCsLV-control. The ASCsLV-shNLRP3 group had improved cavernous endothelial function and smooth muscle injury, thus reversing erectile function, and was superior to the ASCsLV-control group.ConclusionsNLRP3 Inflammasome-mediated pyroptosis might be involved in DMED formation. Intra-cavernous injection of ASCsLV-shNLRP3 could suppress cavernosal pyroptosis, contributing to improved erectile function in DMED rats.

Abnormal amyloid-β (Aβ) abnormal accumulation and oxidative stress play important roles in Alzheimer′ disease (AD). Quercetin has been reported to possess antioxidant and anti-inflammatory properties, and thus of therapeutic interests for neurodegenerative disorders. In the present study, we aimed to characterize the mechanisms by which quercetin exerts neuroprotective effects in murine neuroblastoma N2a cells stably expressing human Swedishh mutant amyloid precursor protein (APP). Quercetin treatment exhibited low cytoxicity, attenuated APP expression and APP-induced oxidative neurotoxicity in N2a/APP cells. We found that quercetin effected via the down-regulation of phospho-extracellular signal regulated protein kinase (p-ERK1/2) pathway and up-regulation of phospho-protein kinase B (p-AKT) pathway in N2a/APP cells. In addition, quercetin ameliorated the elevated levels of reactive oxygen species using DCFH-DA flow-cytometry in N2a/APP cells, lipid peroxidation using (4-HNE), and DNA oxidation (8-OHdG assays). Quercetin ameliorated the loss of mitochondrial membrane potential using JC-1 fluorescence assay in N2a/APP cells in a dose-dependent mannor. In conclusion, we domenstrated the neuroprotective effects of quercetin against the APP expression induced oxidative neurotoxicity, impairment of mitochondrial function and oxidative stress through inactivation of the ERK1/2 signaling pathway and activation of AKT signaling pathways.

Alzheimers disease is characterized by abnormal β-amyloid (Aβ) plaque accumulation, tau hyperphosphorylation, reactive oxidative stress, mitochondrial dysfunction and synaptic loss. Myricetin, a dietary flavonoid, has been shown to have neuroprotective effects in vitro and in vivo. Here, we aimed to elucidate the mechanism and pathways involved in myricetin protective effect on the toxicity induced by the Aβ42 oligomer. Neuronal SH-SY5Y cells were pretreated with myricetin before incubation with Aβ42 oligomer. The levels of pre- and post-synaptic proteins, mitochondrial division and fusion proteins, glycogen synthase kinase-3β (GSK-β) and extracellular regulated kinase (ERK) 1/2 were assessed by Western blotting. Flow cytometry assays for mitochondrial membrane potential (JC1) and reactive oxidative stress, as well immunofluorescence staining for lipid peroxidation (4-HNE) and DNA oxidation (8-OHdG), were performed. We found that myricetin prevented Aβ42 oligomer-induced tau phosphorylation and the reduction in pre/postsynaptic proteins. In addition, myricetin reduced reactive oxygen species generation, lipid peroxidation, and DNA oxidation induced by the Aβ42 oligomer. Moreover, myricetin prevented the Aβ42 oligomer-induced reduction in mitochondrial fusion proteins (mitofusin-1, mitofusin-2), fission protein (dynamin-related protein 1) phosphorylation, and mitochondrial membrane potential via the associated GSK-3β and ERK 1/2 signaling pathways. In conclusion, this study provides new insight into the neuroprotective mechanism of myricetin against Aβ42 oligomer-induced toxicity.

Oxidative stress is involved in the pathogenesis of Alzheimer's disease (AD), which is linked to reactive oxygen species (ROS), lipid peroxidation, and neurotoxicity. Emerging evidence suggests a role of nuclear factor (erythroid-derived 2)-like 2 (Nrf2), a major source of antioxidant response elements in AD. The molecular mechanism of oxidative stress and ferroptosis in astrocytes in AD has not yet been fully understood. Here, we aim to investigate the mechanism by which Nrf2 regulates the ferroptosis of astrocytes in AD. Postmortem frontal cortex tissues from patients with AD and nondemented controls and brain tissue from the 3xTg AD mouse model and wild-type mice (10 months old) were used. Immunofluorescence staining for Nrf2, the ROS marker NADPH oxidase 4 (NOX4), and GFAP was performed. We further induced Nrf2 deficiency in mouse astrocytes by using RNAi and assessed the changes in ROS, ferroptosis, lipid peroxidation, and mitochondrial dysfunction by using western blotting and immunofluorescence staining. We found decreased expression of Nrf2 and upregulated expression of NOX4 in the frontal cortex from patients with AD and cortex of 3xTg mice compared to control groups. We demonstrated that Nrf2 deficiency led to ferroptosis-dependent oxidative stress-induced ROS with downregulated heme oxygenase-1 and glutathione peroxidase 4 and upregulated cystine glutamate expression. Moreover, Nrf2 deficiency increased lipid peroxidation, DNA oxidation, and the mitochondrial fragmentation in mouse astrocytes. In conclusion, these results suggest that Nrf2 promotes ferroptosis of astrocytes involving oxidative stress in AD.

In patients with diffuse large B-cell lymphoma, MYC combined with Bcl2 and/or Bcl6-based protein expression is called double expression lymphoma (DEL). R-DA-EPOCH program chemotherapy is typically recommended because these patients often have a poor prognosis. Although numerous factors affect survival of patients with DEL, the roles of the tumor biomarker histone methyltransferase G9a (G9a) and the lymphocyte-to-monocyte ratio (LMR) are unknown. We performed a retrospective analysis of data from 51 patients. These patients were newly diagnosed with DEL and treated with R-DA-EPOCH at Taizhou People’ s Hospital and Northern Jiangsu People's Hospital between June 2014 and December 2019. Receiver operator characteristic curve results were used to calculate the LMR cutoff value. We used an immunohistochemical analysis to examine G9a expression in DEL tissues. The Kaplan–Meier method was used to determine progression-free survival (PFS) and overall survival (OS) characteristics. Cox proportional-hazards models were constructed for univariate and multivariate analyses to examine the prognostic values of LMRs and G9a in patients with DEL. The cutoff value for LMR was 2.18. The 5-year PFS rate was 35.3%, and the 5-year OS rate was 39.2%. Patients with DEL with lower LMRs and who were G9a-positive predicted inferior PFS and OS. Univariate analysis revealed that patients with elevated LDH levels, high National Comprehensive Cancer Network International Prognostic Index (NCCN-IPI) scores, LMRs ≤2.18, and G9a-positive results had relatively poorer PFS and OS. The multivariate analysis revealed that LMRs ≤2.18 and a G9a-positive result were independent prognostic factors for PFS and OS in patients with DEL treated with R-DA-EPOCH. The study results suggested that peripheral blood LMRs were an important marker for evaluation of prognosis in patients with DEL. High expression of G9a was associated with worse outcomes, indicating that G9a may serve as a prognostic biomarker for patients with DEL who undergo R-DA-EPOCH program chemotherapy.