Publications by authors named "Huifeng Pi"

34 Publications

Rapamycin antagonizes cadmium-induced breast cancer cell proliferation and metastasis through directly modulating ACSS2.

Ecotoxicol Environ Saf 2021 Aug 16;224:112626. Epub 2021 Aug 16.

School of Medicine, Guangxi University, Nanning, Guangxi Zhuang Autonomous Region, China; Department of Environmental Medicine, School of Public Health, and Department of Emergency Medicine, First Affiliated Hospital, School of Medicine, Zhejiang University, Hangzhou, Zhejiang, China. Electronic address:

Cadmium (Cd) is a carcinogen that stimulates breast cancer (BC) progression. Rapamycin is a macrolide antibiotic produced by Streptomyces hygroscopicus that possesses a wide array of pharmacological activities, including anti-BC activity. However, the effects of rapamycin on Cd-increased BC progression and the underlying mechanism have not been fully elucidated. Here, we hypothesize that rapamycin antagonizes Cd-induced BC cell proliferation and metastasis by directly modulating ACSS2. In this study, we found that rapamycin efficiently inhibited Cd-induced proliferation, invasion and migration in MCF-7 and T47-D cells. Moreover, a surface plasmon resonance (SPR) assay confirmed that rapamycin directly binds to the ACSS2 protein with a calculated equilibrium dissociation constant (KD) of 18.3 μM. Molecular docking showed that there are three binding sites in the ACSS2 protein and that rapamycin binds at the coenzyme A (COA) binding site with a docking score of - 12.26 and a binding free energy of - 26.34 kcal/mol. More importantly, rapamycin suppresses Cd-induced BC progression by activating ACSS2. After cells were cotreated with an ACSS2 inhibitor, the effects of rapamycin were abolished. In conclusion, our findings suggest that rapamycin suppresses Cd-augmented BC progression by upregulating ACSS2, and ACSS2 may serve as a direct target of rapamycin for inhibiting xenobiotic (e.g., Cd)-mediated BC progression.
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http://dx.doi.org/10.1016/j.ecoenv.2021.112626DOI Listing
August 2021

NAC antagonizes arsenic-induced neurotoxicity through TMEM179 by inhibiting oxidative stress in Oli-neu cells.

Ecotoxicol Environ Saf 2021 Oct 28;223:112554. Epub 2021 Jul 28.

Department of Occupational Health, Army Medical University, 400038, Chongqing, People's Republic of China. Electronic address:

Arsenic is one of the most common environmental pollutants. Neurotoxicity induced by arsenic has become a major public health concern. However, the effects of arsenic-induced neurotoxicity in the brain and the underlying molecular mechanisms are not well understood. N-acetyl-cysteine (NAC) is a thiol-based antioxidant that can antagonize heavy metal-induced neurotoxicity by scavenging reactive oxygen species (ROS). Here, we used the mouse oligodendrocyte precursor cell (OPC) line Oli-neu to explore the neurotoxic effects of arsenic and the protective effects of NAC. We found that arsenic exposure decreased cell viability, increased oxidative stress, caused mitochondrial dysfunction, and led to apoptosis of Oli-neu cells. Furthermore, we revealed that NAC treatment reversed these neurotoxic effects of arsenic. TMEM179, a key membrane protein, was found highly expressed in OPCs and to be an important factor in maintaining mitochondrial functions. We found that TMEM179 played a critical role in mediating the neurotoxic effects of arsenic and the protective role of NAC. PKCβ is a downstream factor through which TMEM179 regulates the expression of apoptosis-related proteins. This study improves our understanding of the neurotoxic effects and mechanisms of arsenic exposure and the protective effects of NAC. It also identifies a potential molecular target, TMEM179, for the treatment of arsenic-induced neurotoxicity.
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http://dx.doi.org/10.1016/j.ecoenv.2021.112554DOI Listing
October 2021

Histone hypoacetylation contributes to neurotoxicity induced by chronic nickel exposure in vivo and in vitro.

Sci Total Environ 2021 Aug 10;783:147014. Epub 2021 Apr 10.

Department of Occupational Health, Army Medical University, 400038 Chongqing, People's Republic of China. Electronic address:

Nickel (Ni) is a heavy metal that is both an environmental pollutant and a threat to human health. However, the effects of Ni on the central nervous system in susceptible populations have not been well established. In the present study, the neurotoxicity of Ni and its underlying mechanism were investigated in vivo and in vitro. Ni exposure through drinking water (10 mg Ni/L, 12 weeks) caused learning and memory impairment in mice. Reduced dendrite complexity was observed in both Ni-exposed mouse hippocampi and Ni-treated (200 μM, 72 h) primary cultured hippocampal neurons. The levels of histone acetylation, especially at histone H3 lysine 9 (H3K9ac), were reduced in Ni-exposed mouse hippocampi and cultured neurons. RNA sequencing and chromatin immunoprecipitation (ChIP) sequencing analyses revealed that H3K9ac-modulated gene expression were downregulated. Treatment with sodium butyrate, a histone deacetylase inhibitor, attenuated Ni-induced H3K9 hypoacetylation, neural gene downregulation and dendrite complexity reduction in cultured neurons. Sodium butyrate also restored Ni-induced memory impairment in mice. These results indicate that Ni-induced H3K9 hypoacetylation may be a contributor to the neurotoxicity of Ni. The finding that Ni disturbs histone acetylation in the nervous system may provide new insight into the health risk of chronic Ni exposure.
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http://dx.doi.org/10.1016/j.scitotenv.2021.147014DOI Listing
August 2021

8-Formylophiopogonanone B antagonizes doxorubicin-induced cardiotoxicity by suppressing heme oxygenase-1-dependent myocardial inflammation and fibrosis.

Biomed Pharmacother 2021 Aug 29;140:111779. Epub 2021 May 29.

Department of Cardiology, Affiliated Hospital of North Sichuan Medical College, No. 63, Wenhua Road, Shunqing District, Nanchong, Sichuan 637000, China; Academician Workstation, Affiliated Hospital of North Sichuan Medical College, Nanchong, Sichuan 637000, China. Electronic address:

Doxorubicin (DOX) is a widely used antitumor drug that causes severe cardiotoxicity in patients; no effective strategy yet exists to address this problem. We previously reported that 8-formylophiopogonanone B (8-FOB), a natural isoflavone in Ophiopogon japonicas, antagonizes paraquat-induced hepatotoxicity. Here, we explored the mechanisms underlying DOX-induced cardiotoxicity as well as whether 8-FOB can alleviate DOX-induced cardiotoxicity. Acute cardiotoxicity was established by injecting C57BL/6J mice with a single dose of DOX (20 mg/kg, intraperitoneal). To elucidate the mechanisms underlying DOX-induced cardiotoxicity, differentially expressed genes between hearts from DOX-treated and control mice were identified from the Gene Expression Omnibus (GEO) database via GEO2R. Using the Cytoscape software plugin cytoHubba, five hub genes associated with DOX-induced cardiotoxicity were identified: CD68, PTEN, SERPINE1, AIF1, and HMOX1. However, of these, only HMOX1 protein expression levels were significantly increased after DOX treatment. We also confirmed that HMOX1-dependent myocardial inflammation and fibrosis were closely associated with DOX-induced cardiotoxicity. More importantly, 8-FOB protected against DOX-cardiotoxicity by ameliorating cardiac injury and dysfunction, reducing cardiac fibrosis and inflammatory cytokine release, and inhibiting HMOX1 expression. In conclusion, our results suggest that inhibition of HMOX1-dependent myocardial inflammatory insults and fibrosis is essential for 8-FOB to ameliorate DOX-caused cardiotoxicity.
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http://dx.doi.org/10.1016/j.biopha.2021.111779DOI Listing
August 2021

1800 MHz Radiofrequency Electromagnetic Field Impairs Neurite Outgrowth Through Inhibiting EPHA5 Signaling.

Front Cell Dev Biol 2021 12;9:657623. Epub 2021 Apr 12.

Department of Occupational Health, Third Military Medical University, Chongqing, China.

The increasing intensity of environmental radiofrequency electromagnetic fields (RF-EMF) has increased public concern about its health effects. Of particular concern are the influences of RF-EMF exposure on the development of the brain. The mechanisms of how RF-EMF acts on the developing brain are not fully understood. Here, based on high-throughput RNA sequencing techniques, we revealed that transcripts related to neurite development were significantly influenced by 1800 MHz RF-EMF exposure during neuronal differentiation. Exposure to RF-EMF remarkably decreased the total length of neurite and the number of branch points in neural stem cells-derived neurons and retinoic acid-induced Neuro-2A cells. The expression of Eph receptors 5 (EPHA5), which is required for neurite outgrowth, was inhibited remarkably after RF-EMF exposure. Enhancing EPHA5 signaling rescued the inhibitory effects of RF-EMF on neurite outgrowth. Besides, we identified that cAMP-response element-binding protein (CREB) and RhoA were critical downstream factors of EPHA5 signaling in mediating the inhibitory effects of RF-EMF on neurite outgrowth. Together, our finding revealed that RF-EMF exposure impaired neurite outgrowth through EPHA5 signaling. This finding explored the effects and key mechanisms of how RF-EMF exposure impaired neurite outgrowth and also provided a new clue to understanding the influences of RF-EMF on brain development.
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http://dx.doi.org/10.3389/fcell.2021.657623DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8075058PMC
April 2021

Guidelines for the use and interpretation of assays for monitoring autophagy (4th edition).

Autophagy 2021 Jan 8;17(1):1-382. Epub 2021 Feb 8.

University of Crete, School of Medicine, Laboratory of Clinical Microbiology and Microbial Pathogenesis, Voutes, Heraklion, Crete, Greece; Foundation for Research and Technology, Institute of Molecular Biology and Biotechnology (IMBB), Heraklion, Crete, Greece.

In 2008, we published the first set of guidelines for standardizing research in autophagy. Since then, this topic has received increasing attention, and many scientists have entered the field. Our knowledge base and relevant new technologies have also been expanding. Thus, it is important to formulate on a regular basis updated guidelines for monitoring autophagy in different organisms. Despite numerous reviews, there continues to be confusion regarding acceptable methods to evaluate autophagy, especially in multicellular eukaryotes. Here, we present a set of guidelines for investigators to select and interpret methods to examine autophagy and related processes, and for reviewers to provide realistic and reasonable critiques of reports that are focused on these processes. These guidelines are not meant to be a dogmatic set of rules, because the appropriateness of any assay largely depends on the question being asked and the system being used. Moreover, no individual assay is perfect for every situation, calling for the use of multiple techniques to properly monitor autophagy in each experimental setting. Finally, several core components of the autophagy machinery have been implicated in distinct autophagic processes (canonical and noncanonical autophagy), implying that genetic approaches to block autophagy should rely on targeting two or more autophagy-related genes that ideally participate in distinct steps of the pathway. Along similar lines, because multiple proteins involved in autophagy also regulate other cellular pathways including apoptosis, not all of them can be used as a specific marker for autophagic responses. Here, we critically discuss current methods of assessing autophagy and the information they can, or cannot, provide. Our ultimate goal is to encourage intellectual and technical innovation in the field.
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http://dx.doi.org/10.1080/15548627.2020.1797280DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7996087PMC
January 2021

Cadmium exposure impairs pancreatic β-cell function and exaggerates diabetes by disrupting lipid metabolism.

Environ Int 2021 04 25;149:106406. Epub 2021 Jan 25.

Department of Environmental Medicine, and Department of Emergency Medicine of First Affiliated Hospital, Zhejiang University School of Medicine, Hangzhou, China. Electronic address:

Cadmium is known as an environmental pollutant that contributes to pancreatic damage and the pathogenesis of diabetes. However, less attention has been devoted to elucidating the mechanisms underlying Cd-induced pancreatic β-cell dysfunction and the role of Cd toxicity in the development of diabetes. In this study, we demonstrated that exposure to Cd caused remarkable pancreatic β-cell dysfunction and death, both in vitro and in vivo. Lipidomic analysis of Cd-exposed pancreatic β-cells using high-resolution mass spectrometry revealed that Cd exposure altered the profile and abundance of lipids. Cd exposure induced intracellular lipid accumulation, promoted lipid biogenesis, elevated pro-inflammatory lipid contents and inhibited lipid degradation. Furthermore, Cd exposure upregulated the expression levels of TNF-α, IL-1β and IL-6 in pancreatic β-cells and elevated the TNF-α, IL1-β and IL-6 levels in the serum and pancreas. Taken together, the results of our study demonstrated that environmental relevant Cd exposure causes pro-inflammatory lipids elevation and insulin secretion dysfunction in β-cells and hence exaggerates diabetes development. Combined exposure to environmental hazardous chemicals might markedly increase the probability of developing diabetes in humans. This study provides new metabolic and pharmacological targets for antagonizing Cd toxicity.
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http://dx.doi.org/10.1016/j.envint.2021.106406DOI Listing
April 2021

Cadmium promotes breast cancer cell proliferation, migration and invasion by inhibiting ACSS2/ATG5-mediated autophagy.

Environ Pollut 2021 Jan 15;273:116504. Epub 2021 Jan 15.

School of Medicine, Guangxi University, Nanning, Guangxi Zhuang Autonomous Region, China; Department of Environmental Medicine, School of Public Health, And Department of Emergency Medicine, First Affiliated Hospital, School of Medicine, Zhejiang University, Hangzhou, Zhejiang, China. Electronic address:

Cadmium (Cd), which is considered a carcinogenic metal, promotes breast cancer (BC) progression, but the precise mechanism remains unclear. Herein, MCF-7 and T47-D cells were treated with 0.1, 1, and 10 μM cadmium chloride (CdCl) for 24, 48 and 72 h. In our study, Cd exposure significantly accelerated the proliferation, migration and invasion of MCF-7 and T47-D cells. Notably, Cd inhibited autophagic flux by suppressing ATG5-dependent autophagosome formation but had no significant effect on autophagosome-lysosome fusion and lysosomal function. The genetic enhancement of autophagy through ATG5 overexpression suppressed the Cd-mediated increases in proliferation, migration and invasion, which indicated a carcinogenic role of autophagy impairment in Cd-exposed BC cells. GSEA and GeneMANIA were utilized to demonstrate that the Cd-induced decrease in ACSS2 expression mechanistically inhibited ATG5-dependent autophagy in BC cells. Importantly, ACSS2 overexpression increased the level of H3K27 acetylation in the promoter region of ATG5, and this result maintained autophagic flux and abolished the Cd-induced increases in proliferation, migration and invasion. We also verified that the expression of ACSS2 in BC tissues was low and positively related to ATG5 expression. These findings indicated that the promoting effect of Cd on BC cell proliferation, migration and invasion through the impairment of ACSS2/ATG5-dependent autophagic flux suggests a new mechanism for BC cell proliferation and metastasis stimulated by Cd.
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http://dx.doi.org/10.1016/j.envpol.2021.116504DOI Listing
January 2021

BIX-01294, a G9a inhibitor, suppresses cell proliferation by inhibiting autophagic flux in nasopharyngeal carcinoma cells.

Invest New Drugs 2021 Jun 2;39(3):686-696. Epub 2021 Jan 2.

Department of Otolaryngology Head and Neck Surgery, Xinqiao Hospital, Army Medical University (Third Military Medical University), Chongqing, China.

G9a, a histone methyltransferase, has been found to be upregulated in a range of tumor tissues, and contributes to tumor growth and metastasis. However, the impact of G9a inhibition as a potential therapeutic target in nasopharyngeal carcinoma (NPC) is unclear. In the present study we aimed to investigate the anti-proliferative effect of G9a inhibition in the NPC cell lines CNE1 and CNE2, and to further elucidate the molecular mechanisms underlying these effects. The expression of G9a in NPC tumor tissues was significantly higher than that in normal nasopharyngeal tissues. The pharmacological inhibition of G9a by BIX-01294 (BIX) inhibited proliferation and induced caspase-independent apoptosis in NPC cells in vitro. Treatment with BIX induced autophagosome accumulation, which exacerbated the cytotoxic activity of BIX in NPC cells. Mechanistic studies have found that BIX impairs autophagosomes by initiating autophagy in a Beclin-1-independent way, and impairs autophagic degradation by inhibiting lysosomal cathepsin D activation, leading to lysosomal dysfunction. BIX was able to suppress tumor growth, possibly by inhibiting autophagic flux; it might therefore constitute a promising candidate for NPC therapy.
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http://dx.doi.org/10.1007/s10637-020-01053-7DOI Listing
June 2021

Bisphenol A promotes breast cancer cell proliferation by driving miR-381-3p-PTTG1-dependent cell cycle progression.

Chemosphere 2021 Apr 15;268:129221. Epub 2020 Dec 15.

Department of Environmental Medicine, School of Public Health, and Department of Emergency Medicine, First Affiliated Hospital, School of Medicine, Zhejiang University, China. Electronic address:

Bisphenol A (BPA) is a high-production-volume industrial chemical that facilitates the development of breast cancer. However, the molecular mechanism associated with BPA-induced breast cancer cell proliferation and migration remains elusive. In our study, we exposed MCF-7 cells to different concentrations of BPA (0.1, 1 and 10 μM) for 24, 48, or 72 h. We found that BPA exposure significantly promoted MCF-7 cell proliferation and migration but not invasion. To elucidate the mechanisms, the differentially expressed genes between the BPA and control groups were investigated with the Gene Expression Omnibus (GEO) database through GEO2R. Kyoto Encyclopedia of Genes and Genomes (KEGG) and pathway action network analyses demonstrated the important role of the cell cycle pathway in the effects of BPA exposure on MCF-7 cells. Importantly, analysis with the cytoHubba plugin of Cytoscape software coupled with analysis of enriched genes in the cell cycle pathway identified PTTG1 and CDC20 (two hub genes) as key targets associated with BPA-induced MCF-7 cell proliferation and migration. Interestingly, BPA significantly increased the protein expression levels of PTTG1 but not CDC20. Knockdown of PTTG1 inhibited the BPA-induced increase in proliferation and maintained cell cycle progression. In addition, we confirmed that the increased expression of PTTG1 upon BPA exposure was caused by miR-381-3p inhibition. Moreover, we verified that miR-381-3p expression was low and inversely correlated with PTTG1 expression in breast cancer tissues. Together, these findings demonstrate that BPA promotes high PTTG1 expression and alters the cell cycle to enhance MCF-7 cell proliferation by inhibiting miR-381-3p expression.
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http://dx.doi.org/10.1016/j.chemosphere.2020.129221DOI Listing
April 2021

Long-term bisphenol A exposure exacerbates diet-induced prediabetes via TLR4-dependent hypothalamic inflammation.

J Hazard Mater 2021 01 14;402:123926. Epub 2020 Sep 14.

Department of Environmental Medicine, and Department of Emergency Medicine of First Affiliated Hospital, Zhejiang University School of Medicine, Hangzhou, People's Republic of China. Electronic address:

Bisphenol A (BPA), an environmental endocrine-disrupting compound, has been revealed associated with metabolic disorders such as obesity, prediabetes, and type 2 diabetes (T2D). However, its underlying mechanisms are still not fully understood. Here, we provide new evidence that BPA is a risk factor for T2D from a case-control study. To explore the detailed mechanisms, we used two types of diet models, standard diet (SD) and high-fat diet (HFD), to study the effects of long-term BPA exposure on prediabetes in 4-week-old mice. We found that BPA exposure for 12 weeks exacerbated HFD-induced prediabetic symptoms. Female mice showed increased body mass, serum insulin level, and impaired glucose tolerance, while male mice only exhibited impaired glucose tolerance. No change was found in SD-fed mice. Besides, BPA exposure enhanced astrocyte-dependent hypothalamic inflammation in both male and female mice, which impaired proopiomelanocortin (POMC) neuron functions. Moreover, eliminating inflammation by toll-like receptor 4 (TLR4) knockout significantly abolished the effects of BPA on the hypothalamus and diet-induced prediabetes. Taken together, our data establish a key role for TLR4-dependent hypothalamic inflammation in regulating the effects of BPA on prediabetes.
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http://dx.doi.org/10.1016/j.jhazmat.2020.123926DOI Listing
January 2021

KIF5A-dependent axonal transport deficiency disrupts autophagic flux in trimethyltin chloride-induced neurotoxicity.

Autophagy 2021 04 30;17(4):903-924. Epub 2020 Mar 30.

Department of Environmental Medicine, and Department of Emergency Medicine of the First Affiliated Hospital, Zhejiang University School of Medicine, Hangzhou, China.

Trimethyltin chloride (TMT) is widely used as a constituent of fungicides and plastic stabilizers in the industrial and agricultural fields, and is generally acknowledged to have potent neurotoxicity, especially in the hippocampus; however, the mechanism of induction of neurotoxicity by TMT remains elusive. Herein, we exposed Neuro-2a cells to different concentrations of TMT (2, 4, and 8 μM) for 24 h. Proteomic analysis, coupled with bioinformatics analysis, revealed the important role of macroautophagy/autophagy-lysosome machinery in TMT-induced neurotoxicity. Further analysis indicated significant impairment of autophagic flux by TMT via suppressed lysosomal function, such as by inhibiting lysosomal proteolysis and changing the lysosomal pH, thereby contributing to defects in autophagic clearance and subsequently leading to nerve cell death. Mechanistically, molecular interaction networks of Ingenuity Pathway Analysis identified a downregulated molecule, KIF5A (kinesin family member 5A), as a key target in TMT-impaired autophagic flux. TMT decreased KIF5A protein expression, disrupted the interaction between KIF5A and lysosome, and impaired lysosomal axonal transport. Moreover, overexpression restored axonal transport, increased lysosomal dysfunction, and antagonized TMT-induced neurotoxicity . Importantly, in TMT-administered mice with seizure symptoms and histomorphological injury in the hippocampus, TMT inhibited KIF5A expression in the hippocampus. Gene transfer of enhanced autophagic clearance in the hippocampus and alleviated TMT-induced neurotoxicity . Our results are the first to demonstrate KIF5A-dependent axonal transport deficiency to cause autophagic flux impairment via disturbance of lysosomal function in TMT-induced neurotoxicity; manipulation of KIF5A may be a therapeutic approach for antagonizing TMT-induced neurotoxicity.: 3-MA: 3-methyladenine; AAV: adeno-associated virus; ACTB: actin beta; AGC: automatic gain control; ATG: autophagy-related; ATP6V0D1: ATPase H transporting lysosomal V0 subunit D1; ATP6V1E1: ATPase H transporting lysosomal V1 subunit E1; CA: cornu ammonis; CQ: chloroquine; CTSB: cathepsin B; CTSD: cathepsin D; DCTN1: dynactin subunit 1; DG: dentate gyrus; DYNLL1: dynein light chain LC8-type 1; FBS: fetal bovine serum; GABARAP: GABA type A receptor-associated protein; GABARAPL1: GABA type A receptor associated protein like 1; GABARAPL2: GABA type A receptor associated protein like 2; GAPDH: glyceraldehyde-3-phosphate dehydrogenase; IPA: Ingenuity Pathway Analysis; KEGG: Kyoto Encyclopedia of Genes and Genomes; KIF5A: kinesin family member 5A; LAMP: lysosomal-associated membrane protein; MAP1LC3B/LC3B: microtubule-associated protein 1 light chain 3 beta; NBR1: NBR1 autophagy cargo receptor; OPTN: optineurin; PBS: phosphate-buffered saline; PFA: paraformaldehyde; PIK3C3/VPS34: phosphatidylinositol 3-kinase catalytic subunit type 3; PRM: parallel reaction monitoring; siRNA: small interfering RNA; SQSTM1/p62: sequestosome 1; SYP: synaptophysin; TAX1BP1: Tax1 binding protein 1; TMT: trimethyltin chloride; TUB: tubulin.
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http://dx.doi.org/10.1080/15548627.2020.1739444DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8078766PMC
April 2021

Long-term exercise prevents hepatic steatosis: a novel role of FABP1 in regulation of autophagy-lysosomal machinery.

FASEB J 2019 11 31;33(11):11870-11883. Epub 2019 Jul 31.

School of Aerospace Medicine, Fourth Military Medical University, Xi'an, China.

Nonalcoholic fatty liver disease (NAFLD) is one of the most prevalent diseases worldwide. Exercise is a first-line therapy and an important preventive measure for patients with NAFLD, but the underlying mechanisms are not clear. C57BL/6 mice were fed a high-fat diet (HFD) and subjected to 12 wk swimming exercise. Exercise protected against hepatic lipid accumulation and alleviated hepatocyte damage in HFD mice. Tandem mass tag-based quantitative proteomic analyses and ingenuity pathway analysis revealed that exercise down-regulated fatty acid-binding protein (FABP)1 signaling pathway, which was most closely associated with lipid metabolism. Moreover, exercise significantly decreased FABP1 expression, and liver-specific overexpression of FABP1 abolished the protective effect of exercise in NAFLD mice. Specifically, exercise significantly increased autophagic flux restoring lysosomal function, including lysosomal proteolysis and lysosomal acidification maintenance, contributing to enhancement in autophagic clearance and subsequently alleviation of hepatic steatosis. Conversely, overexpression in the mouse liver blocked the protective effect of exercise inhibiting autophagy flux. The present study identified FABP1 inhibition-mediated replenishment of the autophagy-lysosomal machinery as a novel endogenous mechanism whereby long-term exercise improves lipid homeostasis and ameliorates hepatic steatosis in NAFLD.-Pi, H., Liu, M., Xi, Y., Chen, M., Tian, L., Xie, J., Chen, M., Wang, Z., Yang, M., Yu, Z., Zhou, Z., Gao, F. Long-term exercise prevents hepatic steatosis: a novel role of FABP1 in regulation of autophagy-lysosomal machinery.
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http://dx.doi.org/10.1096/fj.201900812RDOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6902714PMC
November 2019

Inhibition of SERPINA3N-dependent neuroinflammation is essential for melatonin to ameliorate trimethyltin chloride-induced neurotoxicity.

J Pineal Res 2019 Oct 6;67(3):e12596. Epub 2019 Aug 6.

Department of Environmental Medicine, and Department of Emergency Medicine of First Affiliated Hospital, Zhejiang University School of Medicine, Hangzhou, China.

Trimethyltin chloride (TMT) is a potent neurotoxin that causes neuroinflammation and neuronal cell death. Melatonin is a well-known anti-inflammatory agent with significant neuroprotective activity. Male C57BL/6J mice were intraperitoneally injected with a single dose of melatonin (10 mg/kg) before exposure to TMT (2.8 mg/kg, ip). Thereafter, the mice received melatonin (10 mg/kg, ip) once a day for another three consecutive days. Melatonin dramatically alleviated TMT-induced neurotoxicity in mice by attenuating hippocampal neuron loss, inhibiting epilepsy-like seizures, and ameliorating memory deficits. Moreover, melatonin markedly suppressed TMT-induced neuroinflammatory responses and astrocyte activation, as shown by a decrease in inflammatory cytokine production as well as the downregulation of neurotoxic reactive astrocyte phenotype markers. Mechanistically, serine peptidase inhibitor clade A member 3N (SERPINA3N) was identified as playing a central role in the protective effects of melatonin based on quantitative proteome and bioinformatics analysis. Most importantly, melatonin significantly suppressed TMT-induced SERPINA3N upregulation at both the mRNA and protein levels. The overexpression of Serpina3n in the mouse hippocampus abolished the protective effects of melatonin on TMT-induced neuroinflammation and neurotoxicity. Melatonin protected cells against TMT-induced neurotoxicity by inhibiting SERPINA3N-mediated neuroinflammation. Melatonin may be a promising and practical agent for reducing TMT-induced neurotoxicity in clinical practice.
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http://dx.doi.org/10.1111/jpi.12596DOI Listing
October 2019

SCD1 activation impedes foam cell formation by inducing lipophagy in oxLDL-treated human vascular smooth muscle cells.

J Cell Mol Med 2019 08 22;23(8):5259-5269. Epub 2019 May 22.

School of Aerospace Medicine, Fourth Military Medical University, Xi'an, China.

The formation of fat-laden foam cells, which contributes to the fatty streaks in the plaques of atheromas, is an important process in atherosclerosis. Vascular smooth muscle cells (VSMCs) are a critical origin of foam cells. However, the mechanisms that underlie VSMC foam cell formation are not yet completely understood. Here, we demonstrated that oxidized low-density lipoprotein (oxLDL) inhibited lipophagy by suppressing lipid droplet (LD)-lysosome fusion and increased VSMC foam cell formation. Moreover, although oxLDL treatment inhibited lysosomal biogenesis, it had no significant effect on lysosomal proteolysis and lysosomal pH. Notably, through TMT-based quantitative proteomic analysis and database searching, 94 differentially expressed proteins were identified, of which 54 were increased and 40 were decreased in the oxLDL group compared with those in the control group. Subsequently, SCD1, a protein of interest, was further investigated. SCD1 levels in the VSMCs were down-regulated by exposure to oxLDL in a time-dependent manner and the interaction between SCD1 and LDs was also disrupted by oxLDL. Importantly, SCD1 overexpression enhanced LD-lysosome fusion, increased lysosomal biogenesis and inhibited VSMC foam cell formation by activating TFEB nuclear translocation and its reporter activity. Modulation of the SCD1/TFEB-mediated lipophagy machinery may offer novel therapeutic approaches for the treatment of atherosclerosis.
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http://dx.doi.org/10.1111/jcmm.14401DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6652860PMC
August 2019

AKT inhibition-mediated dephosphorylation of TFE3 promotes overactive autophagy independent of MTORC1 in cadmium-exposed bone mesenchymal stem cells.

Autophagy 2019 04 20;15(4):565-582. Epub 2018 Oct 20.

a Department of Environmental Medicine, and Department of Critical Care Medicine of the First Affiliated Hospital , Zhejiang University School of Medicine , Hangzhou , China.

Cadmium (Cd) is a toxic metal that is widely found in numerous environmental matrices and induces serious adverse effects in various organs and tissues. Bone tissue seems to be a crucial target of Cd contamination. Macroautophagy/autophagy has been proposed to play a pivotal role in Cd-mediated bone toxicity. However, the mechanisms that underlie Cd-induced autophagy are not yet completely understood. We demonstrated that Cd treatment increased autophagic flux and inhibition of the autophagic process using Atg7 gene silencing blocked the Cd-induced mesenchymal stem cell death. Mechanistically, Cd activated nuclear translocation of TFE3 but not that of TFEB or MITF, which contributed to the expression of autophagy-related genes and lysosomal biogenesis. Specifically, Cd decreased expression of phospho-AKT (Ser473). The reduction in AKT activity led to dephosphorylation of cytosolic TFE3 at Ser565 and promoted TFE3 nuclear translocation independently of MTORC1. Notably, Cd treatment increased the activity of PPP3/calcineurin, and pharmacological inhibition of PPP3/calcineurin with FK506 suppressed AKT dephosphorylation and TFE3 activity. These results suggest that PPP3/calcineurin negatively regulates AKT phosphorylation and is involved in Cd-induced TFE3-dependent autophagy. Modulation of the PPP3/calcineurin-AKT-TFE3 autophagic-lysosomal machinery may offer novel therapeutic approaches for the treatment of Cd-induced bone damage. Abbreviations: ACTB: actin: beta; AKT: thymoma viral proto-oncogene; AMPK: AMP-activated protein kinase; ATG: autophagy related; Baf A1: bafilomycin A; Cd: cadmium; FOXO3: forkhead box O3; MAP1LC3/LC3: microtubule-associated protein 1 light chain 3; MITF: melanogenesis associated transcription factor; MSC: mesenchymal stem sell; MTORC1: mechanistic target of rapamycin kinase complex 1; RPS6KB1: ribosomal protein S6 kinase: polypeptide 1; SGK1: serum/glucocorticoid regulated kinase 1; SQSTM1/p62: sequestosome 1;TFE3: transcription factor E3; TFEB: transcription factor EB; TFEC: transcription factor EC.
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http://dx.doi.org/10.1080/15548627.2018.1531198DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6526814PMC
April 2019

Transcription factor E3 protects against cadmium-induced apoptosis by maintaining the lysosomal-mitochondrial axis but not autophagic flux in Neuro-2a cells.

Toxicol Lett 2018 Oct 17;295:335-350. Epub 2018 Jul 17.

Department of Environmental Medicine, and Department of Critical Care Medicine of the First Affiliated Hospital, Zhejiang University School of Medicine, Hangzhou, China. Electronic address:

Cadmium (Cd), is a well-known environmental and occupational hazard with a potent neurotoxic action. However, the mechanism underlying cadmium-induced neurotoxicity remains unclear. Herein, we exposed Neuro-2a cells to different concentrations of cadmium chloride (CdCl) (12.5, 25 and 50 μM) for 24 h and found that Cd significantly induced lysosomal membrane permeabilization (LMP) with the release of cathepsin B (CTSB) to the cytosol, which in turn caused the release of mitochondrial cytochrome c (Cyt c) and eventually triggered caspase-dependent apoptosis. Interestingly, Cd decreased TFE3 expression but induced the nuclear translocation of TFE3 and TFE3 target-gene expression, which might be associated with lysosomal stress mediated by Cd. Notably, Tfe3 overexpression protected against Cd-induced neurotoxicity by maintaining the lysosomal-mitochondrial axis, and the protective effect of TFE3 is not dependent on the restoration of autophagic flux. In conclusion, our study demonstrated for the first time that lysosomal-mitochondrial axis dependent apoptosis, a neglected mechanism, may be the most important reason for Cd-induced neurotoxicity and that manipulation of TFE3 signaling may be a potential therapeutic approach for treatment of Cd-induced neurotoxicity.
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http://dx.doi.org/10.1016/j.toxlet.2018.07.015DOI Listing
October 2018

Inhibiting MT2-TFE3-dependent autophagy enhances melatonin-induced apoptosis in tongue squamous cell carcinoma.

J Pineal Res 2018 Mar 11;64(2). Epub 2018 Jan 11.

Department of Occupational and Environmental Medicine, School of Public Health, Zhejiang University, Hangzhou, China.

Autophagy modulation is a potential therapeutic strategy for tongue squamous cell carcinoma (TSCC). Melatonin possesses significant anticarcinogenic activity. However, whether melatonin induces autophagy and its roles in cell death in TSCC are unclear. Herein, we show that melatonin induced significant apoptosis in the TSCC cell line Cal27. Apart from the induction of apoptosis, we demonstrated that melatonin-induced autophagic flux in Cal27 cells as evidenced by the formation of GFP-LC3 puncta, and the upregulation of LC3-II and downregulation of SQSTM1/P62. Moreover, pharmacological or genetic blockage of autophagy enhanced melatonin-induced apoptosis, indicating a cytoprotective role of autophagy in melatonin-treated Cal27 cells. Mechanistically, melatonin induced TFE3 dephosphorylation, subsequently activated TFE3 nuclear translocation, and increased TFE3 reporter activity, which contributed to the expression of autophagy-related genes and lysosomal biogenesis. Luzindole, a melatonin membrane receptor blocker, or MT2-siRNA partially blocked the ability of melatonin to promote mTORC1/TFE3 signaling. Furthermore, we verified in a xenograft mouse model that melatonin with hydroxychloroquine or TFE3-siRNA exerted a synergistic antitumor effect by inhibiting autophagy. Importantly, TFE3 expression positively correlated with TSCC development and poor prognosis in patients. Collectively, we demonstrated that the melatonin-induced increase in TFE3-dependent autophagy is mediated through the melatonin membrane receptor in TSCC. These data also suggest that blocking melatonin membrane receptor-TFE3-dependent autophagy to enhance the activity of melatonin warrants further attention as a treatment strategy for TSCC.
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http://dx.doi.org/10.1111/jpi.12457DOI Listing
March 2018

Enhancing lysosomal biogenesis and autophagic flux by activating the transcription factor EB protects against cadmium-induced neurotoxicity.

Sci Rep 2017 02 27;7:43466. Epub 2017 Feb 27.

Department of Occupational Health, Third Military Medical University, Chongqing 400038, People's Republic of China.

Cadmium (Cd), a highly ubiquitous heavy metal, is a well-known inducer of neurotoxicity. However, the mechanism underlying cadmium-induced neurotoxicity remains unclear. In this study, we found that Cd inhibits autophagosome-lysosome fusion and impairs lysosomal function by reducing the levels of lysosomal-associated membrane proteins, inhibiting lysosomal proteolysis and altering lysosomal pH, contributing to defects in autophagic clearance and subsequently leading to nerve cell death. In addition, Cd decreases transcription factor EB (TFEB) expression at both the mRNA and protein levels. Furthermore, Cd induces the nuclear translocation of TFEB and TFEB target-gene expression, associated with compromised lysosomal function or a compensatory effect after the impairment of the autophagic flux. Notably, restoration of the levels of lysosomal-associated membrane protein, lysosomal proteolysis, lysosomal pH and autophagic flux through Tfeb overexpression protects against Cd-induced neurotoxicity, and this protective effect is incompletely dependent on TFEB nuclear translocation. Moreover, gene transfer of the master autophagy regulator TFEB results in the clearance of toxic proteins and the correction of Cd-induced neurotoxicity in vivo. Our study is the first to demonstrate that Cd disrupts lysosomal function and autophagic flux and manipulation of TFEB signalling may be a therapeutic approach for antagonizing Cd-induced neurotoxicity.
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http://dx.doi.org/10.1038/srep43466DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5327481PMC
February 2017

Dihydromyricetin protects against liver ischemia/reperfusion induced apoptosis via activation of FOXO3a-mediated autophagy.

Oncotarget 2016 11;7(47):76508-76522

Department of Hepatobiliary Surgery, Fuzhou General Hospital of PLA, Fuzhou, Fujian, China.

Liver ischemia and reperfusion (I/R) injury is characterized by defective liver autophagy accompanied by alterations to the endogenous defense system. Dihydromyricetin (DHM) is a natural flavonoid that demonstrates a wide range of physiological functions, and has been implicated as a regulator of autophagy. This study investigates the protective effects of DHM pretreatment on liver injury caused by ischemia/reperfusion (I/R) and elucidates the potential mechanism of DHM-mediated protection. Mice were subjected to 60 minutes of ischemia followed by 5 hours of reperfusion. DHM (100 mg/kg bw/day) or the vehicle was administered daily by gavage 7 days before ischemia and immediately before reperfusion. In this study, DHM markedly decreased serum aminotransferase activity and inhibited liver I/R -stimulated apoptosis. Moreover, DHM exerted hepatoprotective effects by upregulating mRNA levels of various essential autophagy-related genes including ATG5, ATG12, BECN1, and LC3. Autophagy inhibitor chloroquine or Atg5 knockdown blocked DHM -mediated elevation in liver function. Specifically, DHM significantly increased FOXO3a expression, and enhanced FOXO3a nuclear translocation and Ser588 phosphorylation modification. Importantly, the inhibition of FOXO3a with FOXO3a-siRNA in mice decreased DHM-induced autophagy-related genes and diminished the protective effects of DHM against liver I/R injury. In summary, these findings identify DHM as a novel hepatoprotective small molecule by elevating FOXO3a expression and nuclear translocation, stimulating autophagy-related genes and suppressing liver I/R-induced apoptosis, suggesting FOXO3a may have therapeutic value in liver cell protection in liver I/R injury.
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http://dx.doi.org/10.18632/oncotarget.12894DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5363527PMC
November 2016

From the Cover: Autophagy Induction Contributes to Cadmium Toxicity in Mesenchymal Stem Cells via AMPK/FOXO3a/BECN1 Signaling.

Toxicol Sci 2016 11 4;154(1):101-114. Epub 2016 Aug 4.

Department of Occupational Health, Third Military Medical University, Chongqing 400038, People's Republic of China

Mesenchymal stem cells (MSCs) are a valuable in vitro model for investigating the bone toxicity of cadmium (Cd). Autophagy has been proposed to play a pivotal role in Cd-mediated toxicity. The FOXO family proteins are important transcription factors that are essential to autophagy induction. This study investigated the role of autophagy in Cd-induced skeleton damage and its potential mechanism. We exposed MSCs to different concentrations of cadmium chloride (3.5, 7, and 14 μM) for 24 h. We demonstrated that Cd treatment increased autophagic flux, and inhibition of autophagic process using BENC1 gene silencing blocked Cd-induced cell death. Cd treatment also significantly increased mRNA levels of various essential autophagy-related genes including ATG5, ATG12, BECN1, LC3, and ULK1. Specifically, Cd increased FOXO3a and FOXO1 expression at the mRNA and protein levels, and AMPK was demonstrated to enhance FOXO3a nuclear translocation and transcriptional activity by phosphorylating FOXO3a at specific serine residues (Ser588) in Cd-treated MSCs. Notably, knockdown of FOXO3a, but not FOXO1, prevented autophagy-related genes expression and autophagosome formation after Cd treatment. Taken together, our results demonstrate that Cd-induced cell death via the overactivation of FOXO3a-dependent autophagy. Modulation of the FOXO3a autophagy pathway may offer novel therapeutic approaches for the treatment of Cd-induced bone damage.
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http://dx.doi.org/10.1093/toxsci/kfw144DOI Listing
November 2016

Exposure to nickel oxide nanoparticles induces pulmonary inflammation through NLRP3 inflammasome activation in rats.

Int J Nanomedicine 2016 22;11:3331-46. Epub 2016 Jul 22.

Department of Occupational Health.

With recent advances in the manufacture and application of nickel oxide nanoparticles (NiONPs), concerns about their adverse effects on the respiratory system are increasing. However, the underlying cellular and molecular mechanisms of NiONP-induced pulmonary toxicity remain unclear. In this study, we focused on the impacts of NiONPs on pulmonary inflammation and investigated whether the NLRP3 inflammasome is involved in NiONP-induced pulmonary inflammation and injury. NiONP suspensions were administered by single intratracheal instillation to rats, and inflammatory responses were evaluated at 3 days, 7 days, or 28 days after treatment. NiONP exposure resulted in sustained pulmonary inflammation accompanied by inflammatory cell infiltration, alveolar proteinosis, and cytokine secretion. Expression of Nlrp3 was markedly upregulated by the NiONPs, which was accompanied by overexpression of the active form of caspase-1 (p20) and interleukin (IL)-1β secretion in vivo. NiONP-induced IL-1β secretion was partially prevented by co-treatment with a caspase-1 inhibitor in macrophages. Moreover, siRNA-mediated Nlrp3 knockdown completely attenuated NiONP-induced cytokine release and caspase-1 activity in macrophages in vitro. In addition, NiONP-induced NLRP3 inflammasome activation requires particle uptake and reactive oxygen species production. Collectively, our findings suggest that the NLRP3 inflammasome participates in NiONP-induced pulmonary inflammation and offer new strategies to combat the pulmonary toxicity induced by NiONPs.
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http://dx.doi.org/10.2147/IJN.S106912DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4965228PMC
February 2017

Melatonin antagonizes cadmium-induced neurotoxicity by activating the transcription factor EB-dependent autophagy-lysosome machinery in mouse neuroblastoma cells.

J Pineal Res 2016 Oct 3;61(3):353-69. Epub 2016 Aug 3.

Department of Occupational Health, Third Military Medical University, Chongqing, China.

Cadmium (Cd), a highly ubiquitous heavy metal, induces neurotoxicity. Melatonin, a major secretory product of the pineal gland, protects against Cd-induced neurotoxicity. However, the mechanism that accounts for this protection remains to be elucidated. Herein, we exposed mouse neuroblastoma cells (Neuro-2a cells) to different concentrations of cadmium chloride (CdCl2 ) (12.5, 25, and 50 μ mol L(-1) ) for 24 hours. We showed that Cd inhibits autophagosome-lysosome fusion and impairs lysosomal function, subsequently leading to nerve cell death. In addition, Cd decreases the level of transcription factor EB (TFEB) but induces the nuclear translocation of TFEB, associated with compromised lysosomal function or a compensatory effect after the impairment of the autophagic flux. Moreover, compared to the 50-μ mol L(-1) Cd group, administration of 1 μ mol L(-1) melatonin increased "TFEB-responsive genes" (P<.05) and the levels of lysosomal-associated membrane protein (0.57±0.06 vs 1.00±0.11, P<.05), preserved lysosomal protease activity (0.52±0.01 vs 0.90±0.02, P<.05), maintained the lysosomal pH level (0.50±0.01 vs 0.87±0.05, P<.01), and enhanced autophagosome-lysosome fusion (0.05±0.00 vs 0.21±0.01, P<.01). Notably, melatonin enhanced TFEB expression (0.37±0.04 vs 0.72±0.07, P<.05) and nuclear translocation (2.81±0.08 vs 3.82±0.05, P<.05). Tfeb siRNA blocked the melatonin-mediated elevation in autophagy-lysosome machinery in Cd-induced neurotoxicity (P<.01). Taken together, these results uncover a potent role for TFEB-mediated autophagy in the pathogenesis of Cd-induced neurotoxicity, suggesting that control of the autophagic pathway by melatonin might provide an important clue for exploring potential targets for novel therapeutics of Cd-induced neurotoxicity.
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http://dx.doi.org/10.1111/jpi.12353DOI Listing
October 2016

SIRT1 facilitates hepatocellular carcinoma metastasis by promoting PGC-1α-mediated mitochondrial biogenesis.

Oncotarget 2016 May;7(20):29255-74

Department of Hepatobiliary Surgery, Second Affiliated Hospital of Third Military Medical University, Chongqing 400037, China.

SIRT1 is a multifaceted NAD+-dependent protein deacetylase known to act as a tumor promoter or suppressor in different cancers. Here, we describe a novel mechanism of SIRT1-induced hepatocellular carcinoma (HCC) metastasis. SIRT1 overexpression was frequently detected in human HCC specimens and was associated with microvascular invasion (P = 0.0039), advanced tumor node metastasis (TNM) stages (P = 0.0016), HCC recurrence (P = 0.021) and poor outcomes (P = 0.039). Lentivirus-mediated knockdown of SIRT1 in MHCC97H cells reduced invasion and metastasis in vitro and in vivo. SIRT1 depletion attenuated mitochondrial biogenesis and adenosine triphosphate (ATP) production but did not affect epithelial-mesenchymal transition. Elevated SIRT1 expression strongly correlated with the upregulation of PGC-1α in HCC specimens, and ectopic expression of SIRT1 increased PGC-1α levels. In cell assays and an orthotopic transplantation model, PGC-1α overexpression reversed the inhibitory effects of SIRT1 depletion on invasion and metastasis by enhancing mitochondrial biogenesis. These findings reveal the involvement of SIRT1 in HCC metastasis and provide a rationale for exploring therapeutic targets against the SIRT1/PGC-1α axis.
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http://dx.doi.org/10.18632/oncotarget.8711DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5045394PMC
May 2016

G9a-mediated histone methylation regulates cadmium-induced male fertility damage in pubertal mice.

Toxicol Lett 2016 Jun 6;252:11-21. Epub 2016 Apr 6.

Department of Occupational Health, Third Military Medical University, Chongqing 400038, People's Republic of China. Electronic address:

Increasing evidence suggests that cadmium (Cd) is associated with male fertility damage. However, the effects of histone modification on Cd-induced male fertility damage remain obscure. This study aims to evaluate the roles of histone methylation mediated by euchromatin histone methyltransferase (EHMT2/G9a) in regulating Cd-induced male fertility damage. We exposed 4-week-old male C57BL/6J mice to Cd by intraperitoneal injection at 2mg/kg for 1, 3 and 5days. Our data showed that Cd exposure decreased the numbers of impregnated females and litter sizes, which was concomitant with sperm count reduction, histological changes in the cauda epididymal ducts and seminiferous epithelium, and testicular cell apoptosis as evaluated by terminal dUTP nick-end labeling (TUNEL) assay and immunoblotting with increased levels of cleaved caspase 3, PARP and Bax and a decreased level of Bcl-2. Cd-induced male fertility damage was accompanied by enhanced G9a activity followed by increased histone H3 lysine 9 monomethylation (H3K9me1) and dimethylation (H3K9me2) in testes. Furthermore, inhibition of G9a by BIX-01294 normalized H3K9me1 and H3K9me2 to basal levels and prevented Cd-induced male fertility damage and testicular cell apoptosis. Our present study revealed that G9a-mediated histone methylation plays a critical role in Cd-induced male fertility damage and testicular cell apoptosis.
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http://dx.doi.org/10.1016/j.toxlet.2016.04.004DOI Listing
June 2016

CdSe/ZnS quantum dots induce hepatocyte pyroptosis and liver inflammation via NLRP3 inflammasome activation.

Biomaterials 2016 Jun 3;90:27-39. Epub 2016 Mar 3.

Department of Occupational Health, Third Military Medical University, Chongqing 400038, China. Electronic address:

Increased biomedical applications of quantum dots (QDs) have raised considerable concern regarding their toxicological impact. However, the toxicity of QDs is largely unknown and the underlying mechanism is still undefined. This study was conducted to examine the hepatotoxicity of CdSe/ZnS core/shell QDs and the underlying mechanism. In hepatic L02 cells, the QDs caused cytotoxicity in a dose-dependent manner. The QDs were then shown to activate the NLR pyrin domain containing 3 (NLRP3) inflammasome in hepatocytes, leading to a novel pro-inflammatory form of cell death named pyroptosis. Further experiments demonstrated that the QDs induced mitochondrial reactive oxygen species (mtROS) production, and that both a mtROS and a total ROS scavenger attenuated QDs-induced NLRP3 activation and pyroptosis. In addition, QDs increased cytoplasmic calcium (Ca(2+)) levels, while a Ca(2+) release antagonist and chelator alleviated QDs-induced mtROS, NLRP3 activation and subsequent pyroptosis in hepatocytes. In vivo, QDs administration induced liver inflammation and dysfunction. Moreover, the QDs also resulted in NLRP3 activation in liver tissue. However, QDs-induced liver inflammation and dysfunction were abolished in NLRP3 knockout mice. Also, an elevation in mtROS was observed in liver after QDs administration, and the mtROS scavenger suppressed liver NLRP3 activation, inflammation and dysfunction induced by QDs. Our data suggest that QDs induced hepatocyte pyroptosis, liver inflammation and dysfunction via NLRP3 activation, which was caused by QDs-triggered mtROS production and Ca(2+) mobilization. Our results provide novel insights into QDs-induced hepatotoxicity and the underlying mechanism, facilitating control of the side effects of QDs.
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http://dx.doi.org/10.1016/j.biomaterials.2016.03.003DOI Listing
June 2016

Extremely Low-Frequency Electromagnetic Fields Promote In Vitro Neuronal Differentiation and Neurite Outgrowth of Embryonic Neural Stem Cells via Up-Regulating TRPC1.

PLoS One 2016 7;11(3):e0150923. Epub 2016 Mar 7.

Department of Occupational Health, Faculty of Preventive Medicine, Third Military Medical University, Chongqing, China.

Exposure to extremely low-frequency electromagnetic fields (ELF-EMFs) can enhance hippocampal neurogenesis in adult mice. However, little is focused on the effects of ELF-EMFs on embryonic neurogenesis. Here, we studied the potential effects of ELF-EMFs on embryonic neural stem cells (eNSCs). We exposed eNSCs to ELF-EMF (50 Hz, 1 mT) for 1, 2, and 3 days with 4 hours per day. We found that eNSC proliferation and maintenance were significantly enhanced after ELF-EMF exposure in proliferation medium. ELF-EMF exposure increased the ratio of differentiated neurons and promoted the neurite outgrowth of eNSC-derived neurons without influencing astrocyes differentiation and the cell apoptosis. In addition, the expression of the proneural genes, NeuroD and Ngn1, which are crucial for neuronal differentiation and neurite outgrowth, was increased after ELF-EMF exposure. Moreover, the expression of transient receptor potential canonical 1 (TRPC1) was significantly up-regulated accompanied by increased the peak amplitude of intracellular calcium level induced by ELF-EMF. Furthermore, silencing TRPC1 expression eliminated the up-regulation of the proneural genes and the promotion of neuronal differentiation and neurite outgrowth induced by ELF-EMF. These results suggest that ELF-EMF exposure promotes the neuronal differentiation and neurite outgrowth of eNSCs via up-regulation the expression of TRPC1 and proneural genes (NeuroD and Ngn1). These findings also provide new insights in understanding the effects of ELF-EMF exposure on embryonic brain development.
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http://journals.plos.org/plosone/article?id=10.1371/journal.pone.0150923PLOS
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4780708PMC
July 2016

Cadmium induces NLRP3 inflammasome-dependent pyroptosis in vascular endothelial cells.

Toxicol Lett 2016 Mar 22;246:7-16. Epub 2016 Jan 22.

Department of Occupational Health, Third Military Medical University, Chongqing 400038, China. Electronic address:

Cadmium (Cd) is an important and common environmental pollutant that has been linked to cardiovascular diseases, such as atherosclerosis and hypertension. Increasing evidence demonstrates that Cd impairs the cardiovascular system by targeting vascular endothelial cells, but the underlying mechanisms remain obscure. In human umbilical vein endothelial cells (HUVECs), we observed that Cd treatment led to cell death and the generation of inflammatory cytokines. The Cd-induced cell death was identified as pyroptosis, a novel pro-inflammatory form of cell death depending on caspase-1 activation. In addition, exposure of HUVECs to Cd resulted in NLRP3 inflammasome activation as evidenced by cleavage of caspase-1 and downstream interleukin (IL)-1β production. Moreover, knockdown of NLRP3 by small interfering RNA efficiently suppressed Cd-induced caspase-1 cleavage, IL-1β production and pyroptosis in HUVECs. Additional experiments demonstrated that treatment with Cd significantly increased the levels of mitochondrial reactive oxygen species (mtROS) and intracellular ROS in HUVECs. Accordingly, pre-treatment with mtROS scavenger or total ROS scavenger reduced Cd-induced activation of NLRP3 inflammasome and pyroptotic cell death. Taken together, our data suggest that NLRP3 inflammasome, activated by the generation of mtROS, mediates Cd-induced pyroptosis in HUVECs. Our results provide novel insights into Cd-induced cytotoxicity and the underlying mechanism by which Cd induces endothelial injury.
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http://dx.doi.org/10.1016/j.toxlet.2016.01.014DOI Listing
March 2016

Melatonin prevents abnormal mitochondrial dynamics resulting from the neurotoxicity of cadmium by blocking calcium-dependent translocation of Drp1 to the mitochondria.

J Pineal Res 2016 Apr 15;60(3):291-302. Epub 2016 Feb 15.

Department of Occupational Health, Third Military Medical University, Chongqing, China.

Cadmium (Cd) is a persistent environmental toxin and occupational pollutant that is considered to be a potential risk factor in the development of neurodegenerative diseases. Abnormal mitochondrial dynamics are increasingly implicated in mitochondrial damage in various neurological pathologies. The aim of this study was to investigate whether the disturbance of mitochondrial dynamics contributed to Cd-induced neurotoxicity and whether melatonin has any neuroprotective properties. After cortical neurons were exposed to 10 μM cadmium chloride (CdCl2 ) for various periods (0, 3, 6, 12, and 24 hr), the morphology of their mitochondria significantly changed from the normal tubular networks into punctuated structures within 3 hr. Following this pronounced mitochondrial fragmentation, Cd treatment led to signs of mitochondrial dysfunction, including excess reactive oxygen species (ROS) production, decreased ATP content, and mitochondrial membrane potential (▵Ψm) loss. However, 1 mM melatonin pretreatment efficiently attenuated the Cd-induced mitochondrial fragmentation, which improved the turnover of mitochondrial function. In the brain tissues of rats that were intraperitoneally given 1 mg/kg CdCl2 for 7 days, melatonin also ameliorated excessive mitochondrial fragmentation and mitochondrial damage in vivo. Melatonin's protective effects were attributed to its roles in preventing cytosolic calcium ([Ca(2+) ]i ) overload, which blocked the recruitment of Drp1 from the cytoplasm to the mitochondria. Taken together, our results are the first to demonstrate that abnormal mitochondrial dynamics is involved in cadmium-induced neurotoxicity. Melatonin has significant pharmacological potential in protecting against the neurotoxicity of Cd by blocking the disbalance of mitochondrial fusion and fission.
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http://dx.doi.org/10.1111/jpi.12310DOI Listing
April 2016

SIRT3-SOD2-mROS-dependent autophagy in cadmium-induced hepatotoxicity and salvage by melatonin.

Autophagy 2015 ;11(7):1037-51

a Department of Occupational Health; Third Military Medical University ; Chongqing , China.

Cadmium is one of the most toxic metal compounds found in the environment. It is well established that Cd induces hepatotoxicity in humans and multiple animal models. Melatonin, a major secretory product of the pineal gland, has been reported to protect against Cd-induced hepatotoxicity. However, the mechanism behind this protection remains to be elucidated. We exposed HepG2 cells to different concentrations of cadmium chloride (2.5, 5, and 10 μM) for 12 h. We found that Cd induced mitochondrial-derived superoxide anion-dependent autophagic cell death. Specifically, Cd decreased SIRT3 protein expression and activity and promoted the acetylation of SOD2, superoxide dismutase 2, mitochondrial, thus decreasing its activity, a key enzyme involved in mitochondrial ROS production, although Cd did not disrupt the interaction between SIRT3 and SOD2. These effects were ameliorated by overexpression of SIRT3. However, a catalytic mutant of SIRT3 (SIRT3(H248Y)) lacking deacetylase activity lost the capacity to suppress Cd-induced autophagy. Notably, melatonin treatment enhanced the activity but not the expression of SIRT3, decreased the acetylation of SOD2, inhibited mitochondrial-derived O2(•-) production and suppressed the autophagy induced by 10 μM Cd. Moreover, 3-(1H-1,2,3-triazol-4-yl)pyridine, a confirmed selective SIRT3 inhibitor, blocked the melatonin-mediated suppression of autophagy by inhibiting SIRT3-SOD2 signaling. Importantly, melatonin suppressed Cd-induced autophagic cell death by enhancing SIRT3 activity in vivo. These results suggest that melatonin exerts a hepatoprotective effect on mitochondrial-derived O2(•-)-stimulated autophagic cell death that is dependent on the SIRT3/SOD2 pathway.
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http://dx.doi.org/10.1080/15548627.2015.1052208DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4590599PMC
April 2016
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