The Role and Mechanism of Ambra1-Mediated Mitophagy in TDCPP-Exposed Mouse Hippocampal Neurons.

Tri(1,3-dichloro-2-propyl)phosphate (TDCPP) is one of the most widely used organophosphorus flame retardants in consumer products. TDCPP has been confirmed to be neurotoxic, but its mechanism has not been clarified and may be related to mitophagy. AMBRA1 can promote neurological autophagy, but whether AMBRA1 is involved in the mechanism of TDCPP-induced neurotoxicity has not been elucidated. In this study, the optimal neuronal damage model was established by exposing mice hippocampal neurons to TDCPP. Furthermore, on the basis of this model, siRNA was used to knock down AMBRA1. Combined with qRT-PCR and Western blot techniques, we identified AMBRA1-mediated mitophagy-induced neuronal damage in vitro mechanism. The experimental results indicated that TDCPP treatment for 24 h led to a decrease in the cell viability of mouse hippocampal neurons, causing neuronal damage. Meanwhile, TDCPP exposure increased autophagy marker proteins p62 and LC3B, and down-regulated mitochondrial DNA ND1 damage and TOMM20 protein, suggesting that TDCPP exposure promoted mitophagy. In addition, TDCPP exposure led to changes in the expression of AMBRA1 and the key factors of mitophagy, FUNDC1, PINK1, and PARKIN, whereas mitophagy was inhibited after knockdown of AMBRA1. The research results indicated that exposure to TDCPP induced neuronal damage and promoted mitophagy. The mechanism may be that AMBRA1 promoted mitophagy in neuronal cells through the PARKIN-dependent/non-dependent pathway. This study revealed the toxic effects of TDCPP on the nervous system and its potential molecular mechanisms, which provided important clues for further understanding the mechanism of action of AMBAR1-mediated mitophagy.

Paraquat-induced oxidative stress regulates N6-methyladenosine (m6A) modification of long noncoding RNAs in Neuro-2a cells.

Paraquat (PQ) is a ubiquitously applied herbicide. Long-term PQ exposure with low dose has been reported to induce abnormal expression of long non-coding RNAs (lncRNAs) in brain nerve cells, which could further lead to Parkinson's disease (PD). N6-methyladenosine (m6A) modification has recently been identified as having an important role in regulating the function of lncRNAs. However, how m6A modification regulates lncRNAs following PQ exposure remains largely unknown. Herein, this study reported m6A modification of lncRNAs in mouse neuroblastoma cells (Neuro-2a) following PQ induced reactive oxide species (ROS). M6A sequencing was performed to explore the m6A modificated pattern of lncRNAs in Neuro-2a cells which were treated with 200 µM PQ for 3 h. It was found that PQ hypermethylated total RNA and changed the expression of m6A methyltransferase and demethylase proteins, which leading to the alteration of m6A modification of lncRNAs. Furthermore, the functional analysis further revealed that N-acetyl-L-cysteine (NAC)，a ROS scavengers, partly reversed PQ-induced distinct m6A modificated pattern of lncRNAs. In addition, tow specific m6A modified lncRNAs were identified: cell division cycle 5-like (lncRNA CDC5L) and signal transducer and activator of transcription 3 (lncRNA STAT3), which could influence downstream autophagy related biological function. In summary, this work could potentially contribute to the new insight of lncRNAs m6A modification mechanism in the field of environmental toxicology.

Reactive oxygen species regulate miR-17-5p expression via DNA methylation in paraquat-induced nerve cell damage.

There is emerging evidence suggesting that oxidative stress and DNA methylation can alter miRNA expression. However, little is known on the mechanism of miR-17-5p expression changes in paraquat (PQ)-induced nerve cell damage. In the present study, neuro-2a cells were pretreated with antioxidant N-acetylcysteine (NAC) or DNA methylation inhibitor decitabine (DAC), then exposed to different concentrations of PQ, while the expression levels of miR-17-5p were detected by qRT-PCR. Here, it is showed that PQ downregulated the expression of miR-17-5p dose-dependently in neuro-2a cells. The DNA methylation level was upregulated after PQ exposure, while downregulated with the pretreatment of NAC in the above content, detected by 5-mC immunofluorescence technique. The interaction effect of NAC and PQ in alternating DNA methylation level was further confirmed by flow cytometry. NAC and DAC individually had an interaction effect in PQ-induced nerve cell damage. After using NAC, PQ-induced ROS elevation and DNA methylation are reduced, thereby preventing the proapoptotic effect of miR-17-5p. Above all, PQ can induce DNA methylation variations through ROS production, leading to the downregulation of miR-17-5p expression in PQ-induced nerve cell damage.

Cadmium induces ovarian granulosa cell damage by activating PERK-eIF2α-ATF4 through endoplasmic reticulum stress.

This study aimed to investigate whether cadmium induces ovarian granulosa cell damage by activating protein kinase R-like endoplasmic reticulum kinase (PERK)-eIF2α-ATF4 through endoplasmic reticulum (ER) stress and to elucidate the underlying regulation mechanism. Two models of cadmium exposure were established. In one model, ovarian granulosa cells isolated from 21-day-old female Sprague Dawley rats were cultured in vitro for 36 h and exposed to CdCl2 (0, 5, 10, and 20 µM), and in another model, a human ovarian granulosa tumor cell line (COV434) was used to construct the binding immunoglobulin protein (BIP)-knockdown cell line sh-BIP and exposed to 0 and 20 µM CdCl2. After exposure to cadmium for 12 h, the expression mRNA and protein levels of BIP, p-PERK, and p-eIF2α were determined in the two models. miRNAs related to BIP were also detected in granulosa cells after cadmium exposure. We found that mRNA and protein levels of all factors were upregulated in each cadmium-dose group, except for BIP mRNA expression in the 5 µM Cd group. The BIP gene was knocked down in COV434 cells before exposure to cadmium. All factors were upregulated in COV434 cells exposed to Cd, and the expression of the p-eIF2α protein was downregulated in sh-BIP cells exposed to Cd. In addition, no differences in BIP-related miRNAs were detected in cadmium-exposed rat ovarian granulosa cells versus the control group. Cadmium induces ovarian granulosa cell damage by inducing ER stress.

LncRNA NR_030777 promotes mitophagy by targeting CDK1-related mitochondrial fission and ATG12 to attenuate paraquat-induced Parkinson's disease.

With the evidence emerging that abnormal expression of long noncoding RNAs (lncRNAs) are involved in onset of Parkinson's disease (PD), the role of NR_030777 contributing to this disease is of great interest. We recently found that a novel lncRNA "NR_030777" demonstrates protective effects on PQ-induced neurodegeneration. However, the underlying molecular mechanisms of NR_030777 in the regulation of mitochondrial fission and mitophagy involved in PQ-induced neuronal damage remain to be explored. NR_030777 brain conditional overexpressing mice as well as in vitro primary neuronal cells from cerebral cortex and Neuro2a cells were adopted. Immunofluorescence, Immunohistochemistry, qRT-PCR and Western blotting were used to evaluate the expression levels of RNA and proteins. RNA immunoprecipitation and RNA pulldown experiment were used to evaluate the interaction of NR_030777 with its target proteins. NR_030777 and mitophagy were increased, and tyrosine hydroxylase (TH) levels recovered after NR_030777 overexpression upon PQ treatment. The overexpression and knockdown of NR_030777 unveiled that NR_030777 positively regulated mitophagy such as the upregulation of LC3B-II:I, ATG12-ATG5, p62 and NBR1. Moreover, the application of mdivi-1, a DRP-1 inhibitor, in combination with NR_030777 genetic modified cells unveiled that NR_030777 promoted DRP1-mediated mitochondrial fission and mitophagy. Furthermore, NR_030777 were directly bound to CDK1 to increase p-DRP1 levels at the Ser616 site, leading to mitochondrial fission and mitophagy. On the other hand, NR_030777 acted directly on ATG12 within the ATG12-ATG5 complex in the 800-1400 nt region to modulate the membrane formation. Accordingly, NR_030777 deficiency in neuron cells compromised cell mitophagy. Finally, the above findings were confirmed using NR_030777-overexpressing mice. NR_030777 exerted a protective effect on PQ-exposed mice by enhancing mitophagy. Our data provide the first scientific evidence for the precise invention of PQ-induced PD. Our findings further propose a breakthrough for understanding the regulatory relationship between NR_030777, CDK1, ATG12 and mitophagy in PQ-induced PD.

Prevalence status and associated factors of wrist postural injury in the Chinese occupational population.

Objective: This study investigated the prevalence of wrist injuries in 15 industries and different types of work in China. Study on the associated factors of wrist injuries provides a scientific basis for prevention and treatment of wrist diseases in occupational workers. Methods: A cross-sectional study of musculoskeletal symptoms of related practitioners in 15 industries, including automobile manufacturing, was conducted to retrieve worker demographic information, working wrist posture, and pain conditions. Multivariable binary logistic regression analyses were performed to identify factors associated with work-related musculoskeletal disorders (WMSDs). Results: The prevalence of wrist injuries among the study population was 13.2%. Toy manufacturing, animal husbandry, automobile manufacturing, shoe manufacturing, and biopharmaceutical manufacturing had the highest wrist injury rates at 29.1, 19.1, 14.9, 14.9, and 14.0%, respectively. Among the types of jobs, enamel workers (63.0%), butchers (43.6%), combers (32.5%), welders (31.3%), and scaffolders (26.5%) had the highest prevalence rates. Based on the final multivariate logistic regression analysis: female [odds ratios (OR) = 1.24; 95% confidence interval (CI), 1.15-1.35], 6-10 years of service (OR = 1.11; 95% CI, 1.03-1.18), >10 years of service (OR = 1.15; 95% CI, 1.06-1.25), frequent upward and downward flexion in wrist posture at work (OR = 1.81; 95% CI, 1.84-2.11), and frequent wrist placement on the edge of angular objects increased the OR of injury (OR = 1.52; 95% CI, 1.44-1.61). Need to squeeze objects tightly while working (OR = 1.72; 95% CI, 1.57-1.89), prolonged wrist flexion (OR = 1.86; 95% CI, 1.75-1.97), and work hand position above the shoulder for prolonged periods (OR = 1.11; 95% CI, 1.04-1.19) also suggested the relationship between these factors and the higher prevalence of wrist injury in the workers. The associated factor was physical activity (OR = 0.86; 95% CI, 0.80-0.94). Conclusion: This study suggested the relationship between these factors and the higher prevalence of wrist injury in the toy manufacturing, animal husbandry, automobile manufacturing, and shoe-making industries, enamel workers, butchers, and combers. And are work types that require special attention. Females, working age, physical activity, and abnormal posture of the wrist were factors significantly associated with WMSDs.

Drp1-mediated mitochondrial fission contributes to mitophagy in paraquat-induced neuronal cell damage.

Paraquat (PQ) is one of the most widely used herbicides in the world due to its excellent weed control effects. Accumulating evidence has revealed that long-term exposure to PQ can significantly increase the risk of Parkinson's disease (PD). However, the underlying molecular mechanisms are yet to be fully understood. Hence, we investigated the potential role of reactive oxygen species (ROS) and dynamin-related protein 1 (DRP1) in PQ-induced mitophagy, aiming to elaborate on possible molecular mechanisms involved in PQ-triggered neurotoxicity. Our results showed that ROS were increased, mitochondrial membrane potential was decreased at 100, 200, and 300 µM PQ concentrations, and autophagy pathways were activated at a concentration of 100 µM in neuronal cells. In addition, excessive mitophagy was observed in neurons exposed to 300 µM PQ for 24 h. Then, ROS-mediated mitochondrial fission was found to contribute to PQ-induced excessive mitophagy. Moreover, all aforementioned changes were significantly ameliorated by mdivi-1. Thus, our findings provide a novel neurotoxic mechanism and reveal the DRP1-mitochondrial fission pathway as a potential target for treatments of PQ-induced excessive mitophagy, serving as an alternative target for the prevention and treatment of Parkinson's disease. Because harmful substances are transmitted and enriched in the food chain, the toxic effect of environmental paraquat is nonnegligible, and more investigations are needed.

Paraquat-induced oxidative stress regulates N6-methyladenosine (m6A) modification of circular RNAs.

Paraquat (PQ), a widely used herbicide and well-known oxidative stress inducer, has been linked to numerous neurodegenerative diseases, but the underlying mechanism(s) remains unknown. Circular RNAs (circRNAs) have recently been reported to be associated with oxidative stress in Parkinson's disease. Herein, we performed methylated RNA immunoprecipitation and RNA sequencing assays for mouse neuroblastoma (Neuro-2a) cells and successfully established a positive link between the alteration of circRNAs driven by m6A modification and PQ-induced oxidative stress. We observed oxidative stress and antioxidative stress present distinct m6A modification pattern of circRNAs as well as biological effect. Gene ontology and pathway analysis predicted that differentially m6A-methylated and expressed circRNAs are highly clustered in pathways associated with function and development of nervous system, including axon cargo transport, nervous system development, long-term potentiation, and neurotrophic signaling pathways. Moreover, we demonstrated that the alteration of m6A-methylated circRNAs upon PQ exposure could be partially reversed by N-acetylcysteine pretreatment. The mechanistic analysis further demonstrated that N-acetylcysteine pretreatment attenuated the decreased expression of target genes (UBC and PPP2CA) induced by PQ. These findings revealed distinct patterns of differentially m6A-modified circRNAs, indicating that m6A could participate in a specific regulatory network of circRNAs to modulate the expression of downstream genes in response to PQ-induced oxidative stress. In conclusion, our work established a link between m6A modification of circRNAs and PQ-induced oxidative stress, and further studies are required to explore the underlying molecular mechanisms associated with PQ-induced neurotoxicity.

LncRNA NR_030777 Alleviates Paraquat-Induced Neurotoxicity by Regulating Zfp326 and Cpne5.

Paraquat (PQ) is herbicide widely used in agricultural production. It is identified as an environmental toxicant that could lead to neurodegeneration damage. Parkinson's disease (PD) is a central nervous system degenerative disease that occurs in the elderly. Main risk factors for PD include genetic and environmental variables, but its specific mechanism is still not well understood. Emerging evidence suggests that long noncoding RNAs (lncRNAs) play an important role in PD. LncRNA NR_030777 has a full length of 2208 bp and is highly conserved among species. RNA profiling showed a significant alteration in lncRNA NR_030777 expression upon PQ-induced neurotoxicity. However, little is known on the functional relevance of lncRNA NR_030777 in the development of PQ. In this study, we discovered a vital protective role of lncRNA NR_030777 in PQ-induced neurotoxicity. The expression of NR_030777 correlates with elevated level of reactive oxygen species induced by PQ. In addition, activated expression of NR_030777 alleviates neurotoxicity by regulating the expression of Zfp326 and Copine 5. We report that lncRNA NR_030777 has a vital protective role in neurotoxicity induced by environmental toxicants such as PQ. This study could serve as an exemplary case for lncRNAs to be considered as a potential target for the prevention and treatment of PQ-induced neurodegenerative disorders such as PD.

Anti-Müllerian hormone participates in ovarian granulosa cell damage due to cadmium exposure by negatively regulating stem cell factor.

Cadmium is involved in female gonadal toxicity. Although many studies concur in suggesting a significant influence on female reproduction, the mechanisms of action are still needed to study. This study was designed to examine the characteristics of 0, 5, 10, and 20 µM cadmium toxicity in granulosa cells culture for 12 h, and the subsequent effect of cadmium exposure on AMH production and formation. Here we demonstrate that the viability of granulosa cells cadmium exposed was decreased, and the apoptosis was increased. Intriguingly, After exposure to cadmium the SCF was decreased and the AMH was increased in granulosa cells. This study may demonstrate that AMH signaling is involved in Cd action through SCF activation. The signaling pathways may play a role in the reproductive effects caused by environmental and occupational exposures to Cd. These results further confirm the functional significance of AMH signaling in mediating the intraovarian actions of Cd.