Bisphenol A exposure stimulates prostatic fibrosis via exosome-triggered epithelium changes.

Fibrosis is the pathological basis for the clinical progression of benign prostatic hyperplasia (BPH). Prostatic fibrosis is an important risk factor in patients with BPH who experience lower urinary tract symptoms. Bisphenol A (BPA) is an environmental endocrine disruptor (EED) that causes prostate defects. The effects of BPA on the prostate were investigated in this study using mouse and human prostate cell models. BPA-induced mouse prostatic fibrosis is characterized by collagen deposition and an increase in hydroxyproline concentration. Furthermore, BPA-exposed prostatic stromal fibroblasts exosomes promote the epithelial-mesenchymal transition of epithelial cells. High-throughput RNA sequencing and functional enrichment analyses show that substantially altered mRNAs, lncRNAs and circRNAs play roles in cellular interactions and the hypoxia-inducible factor-1 signaling pathway. The results showed that exosomes participated in the pro-fibrogenic effects of BPA on the prostate by mediating communication between stromal and epithelial cells and triggering epithelial changes.

Mediation of PM2.5-induced cytotoxicity: the role of P2X7 receptor in NR8383 cells.

Ambient fine particulate matter (PM2.5) is a threat to public health. The P2 X 7purinergic receptor (P2X7R) is a modulator that responds to inflammation. Yet the role of P2X7R in the mediation of PM2.5-induced pulmonary cytotoxicity is rarely investigated. In this study, the expression of P2X7R and its effect on cell viability, oxidative damage, apoptosis, mitochondrial dysfunction and underlying mechanism following PM2.5 treatment in rat alveolar macrophages (NR8383) were analyzed. The outcome indicated that PM2.5 exposure significantly increased the expression of P2X7R, while P2X7R antagonist oATP markedly alleviate the production of reactive oxygen species (ROS), Nitrite Oxidation (NO), mitochondrial membrane potential, apoptosis rate, and release of inflammatory cytokines. In contrast, P2X7 agonist BzATP showed opposite effect in PM2.5-treated NR8383 cells. Therefore, these results demonstrated that P2X7R participated in PM2.5-induced pulmonary toxicity, while the blockade of P2X7R is a promising therapeutic approach of treating PM2.5-induced lung diseases.

Circ_0027470 promotes cadmium exposure-induced prostatic fibrosis via sponging miRNA-1236-3p and stimulating SHH signaling pathway.

Cadmium (Cd) is a toxic heavy metal pollutant and serves as an important environmental endocrine-disrupting chemical. Cd exposure is believed to can enhance the risks of age-related disorders including benign prostatic hyperplasia (BPH). This study was to investigate the harms of Cd exposure on mice prostate and human nonmalignant prostate epithelial RWPE-1 cells. Mice prostate fibrosis was evaluated by visualizing the prostatic collagen deposition via Masson and Sirius red staining, and detecting the content of hydroxyproline. Additionally, the epithelial-mesenchymal transition (EMT), primary ciliogenesis and SHH signaling pathways in both mice prostate and RWPE-1 cells were evaluated. It was found that Cd exposure stimulated prostatic collagen deposition, EMT and primary ciliogenesis, as well as enhanced the circ_0027470 level and reduced the miRNA-1236-3p level. Circ_0027470 functioned as a sponge of miRNA-1236-3p, which had the inhibiting target of SHH. The whole results showed that circ_0027470 promoted Cd exposure-induced prostatic fibrosis via sponging miRNA-1236-3p and subsequently stimulating SHH signaling pathway. This study shed a light on a novel molecular mechanism involved in circRNA for Cd exposure-induced prostate deficits.

Alcohol Exposure Induces Depressive and Anxiety-like Behaviors via Activating Ferroptosis in Mice.

Alcohol use disorder (AUD) is a global public health problem and is frequently comorbid with mental disorders, including anxiety and depression. Ferroptosis is an iron-dependent cell death, which is involved in the pathological process of various diseases such as neurodegenerative diseases, but the role of ferroptosis in the mediation of AUD and its induced mental disorders is unclear. In this study, we aimed to investigate whether ferroptosis was involved in alcohol-induced depressive and anxiety-like behaviors in mice. Following an 8-week period of intermittent alcohol exposure, the alcohol group showed noticeable depressive and anxiety-like behaviors. In addition, nissl staining revealed that alcohol exposure induced neuron damage in the hippocampus (Hip) and prefrontal cortex (PFC) of mice. The levels of synapse-related proteins were significantly reduced in the alcohol group. Iron staining demonstrated that alcohol increased the number of iron-positive staining cells. The protein expression of the transferrin receptor (TFRC) was increased, and the expression of glutathione peroxidase 4 (GPX4) was decreased, respectively, in the alcohol group. Furthermore, the ferroptosis inhibitor ferrostatin-1 significantly prevented alcohol-induced neuron damage and enhanced the expression of N-methyl-d-aspartic acid (NMDA) receptor 2B (NR2B), α-amino-3-hydroxy-5-methyl-4-isoxazole-propionic acid (AMPA) receptor 1 (GluA1) and GPX4 in vitro. These results indicated that alcohol exposure could induce depressive and anxiety-like behaviors, and that this effect may occur via activating ferroptosis.

PM2 .5 exposure-induced ferroptosis in neuronal cells via inhibiting ERK/CREB pathway.

PM2.5 exposure has been demonstrated to correlate with neurological disorders recently. Ferroptosis is recognized as a newly found programmed form of cell death associated with neurodegenerative diseases, while glutathione peroxidase 4 (GPX4) is a key regulator of ferroptosis. However, the relationship between PM2.5 -induced neurotoxicity and ferroptosis is still unclear. The current study aims to investigate if ferroptosis is involved in neurotoxicity post PM2.5 exposure and its underlying mechanism. The PM2.5 -treated neuronal Neuro-2a (N2A) and SH-SY5Y cells were applied to the current study. The results showed that PM2.5 significantly increased the neuronal cell death, yet the ferroptosis antagonist Ferrostain-1 (Fer-1) markedly decreased the cell death induced by PM2.5 . Western blot further confirmed that ferroptosis was triggered post PM2.5 treatment in N2A cells by decreasing expressions of GPX4 and ferritin heavy chain (FTH), as well as enhancing expressions of ferritin light chain (FTL) and transferrin receptor protein (TFRC). Meanwhile, PM2.5 treatment augmented neuronal oxidative damage and mitochondrial dysfunction. The bioinformatic analysis indicated that CREB could be the regulator of GPX4, and our results showed that ERK/CREB pathway was down-regulated in N2A cells post PM2.5 treatment. The addition of ERK1/2 agonist post PM2.5 treatment significantly inhibit ferroptosis via increasing the expression of GPX4. Taken together, the present study demonstrated that PM2.5 -induced ferroptosis via inhibiting ERK/CREB pathway, and these findings will advance our knowledge of PM2.5 -induced cytotoxicity in the nervous system.

Candidate Chinese Herbal Medicine Alleviates Methamphetamine Addiction via Regulating Dopaminergic and Serotonergic Pathways.

Methamphetamine (METH) addiction and its induced mental disorders have become a severe worldwide problem. A candidate Chinese herbal medicine (CCHM) in our lab had therapeutic effects on METH-induced locomotor sensitization, however, its chemical and pharmacological profiles remain to be elucidated. The current study aimed to investigate the effect of CCHM on conditioned place preference (CPP) induced by METH and screen the main active ingredients and key targets by using network pharmacology and molecular docking methods. Kyoto Encyclopedia of Genes and Genomes (KEGG) enrichment, Gene ontology (GO) analysis and protein-protein interaction (PPI) network were performed to discover the potential mechanisms. Results showed that CCHM could significantly inhibit METH-induced CPP behaviors in mice. A total of 123 components and 43 targets were screened. According to the network pharmacology analysis, ten hub targets including D(2) dopamine receptor (DRD2) and 5-hydroxytryptamine receptor 3A (HTR3A) were screened. GO analysis and KEGG enrichment indicated that mechanisms of CCHM treatment of METH addiction were related to multiple pathways such as dopaminergic synapse and serotoninergic synapse. Western blot results showed that the protein expressions of DRD2 in nucleus accumbens and prefrontal cortex were significantly decreased in METH group, while the protein expressions of HTR3A were significantly increased. These changes caused by METH could be prevented by CCHM pretreatment. The results of molecular docking displayed that the five active ingredients such as (S)-Scoulerine, Hyndarin, and Beta-Sitosterol had good affinities with DRD2 and HTR3A. In conclusion, this study constructed the CCHM's pharmacologic network for treating METH addiction based on the method of network analysis and experimental verification, and analyzed its major active ingredients and potential targets, indicating a new direction for further revealing its mechanisms of effect on METH addiction.

Terminalia chebula extracts ameliorate methamphetamine-induced memory deficits via activating the ERK and Nrf2 pathway.

As a psychoactive substance abused worldwide, methamphetamine (METH) abuse leads to multiple neurodegenerative symptoms including memory deficits. Terminalia chebula retzius extracts (TREs) isolated by our lab have great antioxidant activity and its effect on METH-induced memory deficits has not been investigated yet. The present study was designed to investigate the protective effect of TREs on METH induced cell apoptosis in vitro and memory deficits in vivo. The results showed that TREs treatment attenuated free radical release and improved cell survival of primary hippocampal neurons after METH injury. In the Morris water maze task, TREs treatment reversed METH-induced learning and memory deficits in acquisition and retention. Moreover, TREs reduced oxidative stress in the serum and hippocampus of mice. Additionally, extracellular regulated protein kinases (ERK1/2) pathway and the nuclear factor E2-related factor 2 (Nrf2) pathway were inactivated after METH treatment, and were significantly activated after TREs pretreatment. These findings suggest that TREs may exert potent neuroprotective effect via activation of both ERK and Nrf2 pathways, thus providing a basis for its potential use for ameliorating memory deficits induced by METH.

The exosome-circ_0001359 derived from cigarette smoke exposed-prostate stromal cells promotes epithelial cells collagen deposition and primary ciliogenesis.

Cigarettes consumption is continued to be popular. We found that cigarette smoke (CS) exposure promoted prostatic fibrosis. In this study, human prostate epithelial RWPE-1 cells were co-cultured with exosomes derived from CS exposed-WPMY-1 cells (CS-WPMY-1-exo). The collagen deposition, primary ciliogenesis, epithelial-mesenchymal transition (EMT) and transforming growth factor (TGF)-ß1 level of RWPE-1 were evaluated. The circRNAs profiles of WPMY-1-exo were explored by high-throughput RNA sequencing. It was found that CS-WPMY-1-exo significantly promoted RWPE-1 collagen deposition, EMT and primary ciliogenesis. There were 17 differentially expressed (DE) circRNAs (including circ_0001359) between CS-WPMY-1-exo and the negative control. Functional enrichment analyses showed that the DE circRNAs played important roles in ciliary basal body, spindle microtubule and TGF-ß signaling pathway. Circ_0001359 siRNA attenuated CS-WPMY-1 induced RWPE-1 cells collagen deposition, EMT and primary ciliogenesis, as well as inhibited the level of TGF-ß1. The whole results showed that circ_0001359 derived from CS-WPMY-1-exo contributed to prostatic fibrosis via stimulating epithelial cells phenotypes changes and collagen deposition.

Krill Oil Alleviated Methamphetamine-Induced Memory Impairment via the MAPK Signaling Pathway and Dopaminergic Synapse Pathway.

Methamphetamine (METH) abuse exerts severe harmful effects in multiple organs, especially the brain, and can induce cognitive dysfunction and memory deficits in humans. Krill oil is rich in polyunsaturated fatty acids, while its effect on METH-induced cognitive impairment and mental disorders, and the underlying mechanism remain unknown. The aim of the present study was to investigate the protective effect of krill oil on METH-induced memory deficits and to explore the molecular mechanisms by using an integrated strategy of bioinformatics analysis and experimental verification. METH-exposed mice were treated with or without krill oil. Learning and memory functions were evaluated by the Morris water maze. The drug-component-target network was constructed in combination with network pharmacology. The predicted hub genes and pathways were validated by the Western blot technique. With krill oil treatment, memory impairment induced by METH was significantly improved. 210 predicted targets constituted the drug-compound-target network by network pharmacology analysis. 20 hub genes such as DRD2, MAPK3, CREB, BDNF, and caspase-3 were filtered out as the underlying mechanisms of krill oil on improving memory deficits induced by METH. The KEGG pathway and GO enrichment analyses showed that the MAPK signaling pathway, cAMP signaling pathway, and dopaminergic synapse pathway were involved in the neuroprotective effects of krill oil. In the hippocampus, DRD2, cleaved caspase-3, and γ-H2AX expression levels were significantly increased in the METH group but decreased in the krill oil-treated group. Meanwhile, krill oil enhanced the expressions of p-PKA, p-ERK1/2, and p-CREB. Our findings suggested that krill oil improved METH-induced memory deficits, and this effect may occur via the MAPK signaling pathway and dopaminergic synapse pathways. The combination of network pharmacology approaches with experimental validation may offer a useful tool to characterize the molecular mechanism of multicomponent complexes.

The exosome-like vesicles derived from androgen exposed-prostate stromal cells promote epithelial cells proliferation and epithelial-mesenchymal transition.

Benign prostatic hyperplasia (BPH) is an age-related disease in men. Mesenchymal /stromal and epithelial cells interactions are essential to prostate functions. In this study, human nonmalignant prostate epithelial RWPE-1 cells were cocultured with testosterone (TE) -exposed prostate stromal fibroblasts WPMY-1 cells (TE-WPMY-1). The survival rate, epithelial-mesenchymal transition (EMT) and collagen deposition of RWPE-1 were observed. The expression profiles of circRNAs, lncRNAs and mRNAs in WPMY-1-derived exosome-like vesicles (WPMY-1-exo) were explored by high-throughput RNA sequencing. Firstly, both TE-WPMY-1 and TE-WPMY-1-exo significantly promoted RWPE-1 cells proliferation. Secondly, 41 circRNAs, 132 lncRNAs and 1057 mRNAs were differentially expressed (DE) between TE-WPMY-1-exo and the control. Functional enrichment analyses, co-expression analyses and quantitative real-time PCR verification showed that the DE RNAs played important roles in cell proliferation, structure, phenotype and fibrosis. Lastly, blocking WPMY-1-exo biogenesis/release by GW4869 can attenuate TE-WPMY-1-stimulated RWPE-1 cells EMT and collagen deposition. Taken together, our results indicated that WPMY-1-exo modulated the phenotypes changes and collagen deposition of prostate epithelial cells. It provided a novel basis for understanding the underlying mechanisms of RWPE-1 cells EMT and fibrosis induced by WPMY-1 in BPH.

Lipopolysaccharide accelerates fine particulate matter-induced cell apoptosis in human lung bronchial epithelial cells.

OBJECTIVES: The aim of the study has been to investigate the effect of the Standard Reference Material of fine particulate matter (SRM 2786) on cytotoxicity and apoptosis in human lung bronchial epithelial cells (16HBE cells). Whether the lipopolysaccharide (LPS)-induced inflammation could further accelerate cell apoptosis induced by SRM 2786 stimulation has also been determined. MATERIAL AND METHODS: 16HBE cells were exposed to various doses of SRM 2786 with or without LPS. The following parameters: cytotoxicity, apoptotic rate, Bax/Bcl-2 expression, nitric oxide (NO) production, and reactive oxygen species (ROS) generation were measured. RESULTS: The results have shown that SRM 2786 induces cell damage and apoptosis of 16HBE cells as demonstrated by significant decrease in expression of Bcl-2 and increase in expression of Bax. When compared with the control cells, the apoptotic rate of cells treated by 500 µg/ml of SRM 2786 increased from 2.43±0.21% to 43.96±2.95% (p < 0.01). Further, there was an elevated production of NO and ROS post SRM 2786 treatment. The level of NO in cells treated with 500 µg/ml of SRM 2786 was 18.33±1.02 µmol/l whereas that of control cells was 1.58±0.31 µmol/l (p < 0.01). When compared with the control group, the level of intracellular ROS increased by 24% after treatment with 500 µg/ml of SRM 2786 (p < 0.05). In addition, LPS pre-treatment may accelerate cell apoptosis by increasing generation of NO and ROS followed by SRM 2786 stimulation. When compared to cells treated with 125 µg/ml of SRM 2786 alone, the levels of NO and ROS in cells pretreated with LPS increased by 28% and 11.6%, respectively (p < 0.05), and the apoptotic rate increased from 34.62±4.44% to 54.11±3.34% (p < 0.01). CONCLUSIONS: These findings have suggested that in vitro exposure to SRM 2786 could induce 16HBE cells apoptosis probably by means of the mechanism involving the generation of free radicals, while the degree of apoptosis would be further aggravated under inflammation condition. Int J Occup Med Environ Health 2018;31(2):173-183.

Combined effects of fine particulate matter and lipopolysaccharide on apoptotic responses in NR8383 macrophages.

Alveolar macrophages (AM) are the predominant lung cells responsible for both ingestion and clearance of inhaled particulate matter (PM). The aims of this study were (1) to examine effects of fine PM on rat NR8383 cell line apoptosis, and (2) to determine whether NR8383 cell functions are further affected when exposed to fine PM in the presence of inflammation induced by lipopolysaccharide (LPS). Standard Reference Material 2786 (SRM 2786) for fine PM was used to measure the following parameters: cytotoxicity, apoptotic rate, Bax/Bcl-2 expression, nitric oxide (NO) production, and reactive oxygen species (ROS) generation in NR8383 cells. Data showed that SRM 2786 alone induced damage and apoptosis in NR8383 cells in a concentration-dependent manner as demonstrated by significant decrease in expression of Bcl-2 and increase in expression of Bax, suggesting fine PM might trigger apoptosis involving a mitochondria-mediated apoptotic pathway. In addition, there was elevated production of free radicals, such as NO and ROS, suggesting oxidative stress plays a role in the observed apoptotic responses. Further, LPS pretreatment enhanced apoptosis of NR8383 cells induced by SRM 2786. Consequently, data indicate that SRM 2786 triggered cell apoptosis in NR8383 cells, probably by mechanisms involving oxidative stress, as evidenced by elevated NO and ROS levels, while the degree of apoptosis was further aggravated by inflammation.