Yongmei Qi

Global DNA hypomethylation, rather than reactive oxygen species (ROS), a potential facilitator of cadmium-stimulated K562 cell proliferation

Cell proliferation plays a critical role in the process of cadmium (Cd) carcinogenesis. Although both induction of reactive oxygen species (ROS) and alteration of DNA methylation are involved in Cd-stimulated cell proliferation, the detailed mechanism of Cd-stimulated cell proliferation remains poorly understood. In this study, K562 cells pre-treated with N-acetylcysteine (NAC) or methionine (Meth) were exposed to Cd to investigate the potential contribution of ROS and global DNA methylation pathways in Cd-induced cell proliferation. The results showed that Cd-stimulated cell proliferation, increased ROS and DNA damage levels, and induced global DNA hypomethylation. The increases of ROS and DNA damage levels were attenuated by pre-treatment with NAC. Cd-stimulated cell proliferation did not appear to be suppressed through eliminating ROS by NAC. However, methionine was shown to prevent Cd-induced global DNA hypomethylation and Cd-stimulated cell proliferation. Our results suggest that global DNA hypomethylation, rather than ROS, is a potential facilitator of Cd-stimulated K562 cell proliferation.

Mediating effect of ROS on mtDNA damage and low ATP content induced by arsenic trioxide in mouse oocytes

Mitochondria provide most of the adenosine triphosphates (ATP) necessary for the maturation of oocytes. Various environmental toxicants would lead damage to mitochondrial DNA (mtDNA) and hence interfere with oocytes development. In the current study, the effect of arsenic trioxide (As2O3) on the common 3867 bp deletion and the copy number of mtDNA in mitochondria of mouse oocytes in vivo or in vitro, as well as the molecular pathway leading to the damage were investigated. PCR strategy was used to detect the damage of mtDNA. Reactive oxygen species (ROS) and ATP content in oocytes were checked to determine the influence of As2O3 on oxidative stress and activity of mitochondria. The results showed that As2O3 could obviously decrease the copy number of mtDNA and cause severe 3867 bp deletion in mitochondria together with elevated ROS level, while ATP content was decreased. Co-treatment with N-Acetyl-Cysteine (NAC) efficiently eliminated ROS induced by As2O3, lessened the mtDNA damage and enhanced ATP content in mouse oocytes both in vivo and in vitro. Taken together, the present study revealed that As2O3 could cause severe mtDNA damage and decrease ATP content by inducing excessive ROS, and this damage would then probably restrain the further development of mouse oocytes.

Cadmium induces mitophagy through ROS-mediated PINK1/Parkin pathway.

Abstract Context and objective: Recent reports have highlighted the relationship between cadmium (Cd) and autophagy, however, whether Cd can activate mitophagy remains enigmatic. This study aims to investigate the effects of Cd on mitophagy and its potential mechanism. Methods: Mice were intraperitoneally injected with Cd for 3 d. Mitochondrial membrane potential (MMP), mitophagosomes, LC3-II/LC3-I ratio, PINK1 level and mitochondrial mass were evaluated to indicate the effects of Cd on mitophagy. To elucidate the mechanism, reactive oxygen species (ROS) scavenger N-acetyl-l-cysteine (NAC) or acetyl-l-carnitine (ALC) as well as the mitophagy inhibitor cyclosporine A (CsA) were introduced to verify the role of ROS in mitophagy. Results and conclusions: The results showed that Cd significantly induced MMP collapse and typical mitophagosomes formation, increased LC3-II/LC3-I ratio and PINK1 level, and decreased mitochondrial mass, revealing that Cd could induce mitophagy. However, NAC or ALC pretreatment markedly decreased Cd-induced ROS and simultaneously rescued MMP and mitochondrial mass, suggesting ROS played a crucial role in regulating mitophagy. NAC or ALC also dramatically lessened PINK1 level and mitochondrial accumulation of Parkin, indicating that ROS were related to PINK1/Parkin pathway. Notably, CsA compromised Cd-induced mitophagy, PINK1 accumulation and Parkin translocation while failed to block ROS increase, suggesting ROS functioned as an upstream signal for PINK1/Parkin pathway. Taken together, the results indicated that Cd induced ROS-mediated mitophagy through PINK1/Parkin pathway in kidneys of mice. The present study proposes a new perspective to evaluate the nephrotoxicity and its molecular mechanism under Cd exposure in vivo.

EGCG inhibits Cd2+-induced apoptosis through scavenging ROS rather than chelating Cd2+ in HL-7702 cells

Abstract Context and objective: Epigallocatechin-3-gallat (EGCG), the major catechin in green tea, shows a potential protective effect against heavy metal toxicity to humans. Apoptosis is one of the key events in cadmium (Cd2+)-induced cytotoxicity. Nevertheless, the study of EGCG on Cd2+-induced apoptosis is rarely reported. The objective of this study was to clarify the effect and detailed mechanism of EGCG on Cd2+-induced apoptosis. Methods: Normal human liver cells (HL-7702) were treated with Cd2+ for 21 h, and then co-treated with EGCG for 3 h. Cell viability, apoptosis, intracellular reactive oxygen species (ROS), malondialdehyde (MDA), mitochondrial membrane potential (MMP) and caspase-3 activity were detected. On the other hand, the chelation of Cd2+ with EGCG was tested by UV-Vis spectroscopy analysis and Nuclear Magnetic Resonance (1H NMR) spectroscopy under neutral condition (pH 7.2). Results and conclusion: Cd2+ significantly decreased the cell viability and induced apoptosis in HL-7702 cells. Conversely, EGCG co-treatment resulted in significant inhibition of Cd2+-induced reduction of cell viability and apoptosis, implying a rescue effect of EGCG against Cd2+ poisoning. The protective effect most likely arises from scavenging ROS and maintaining redox homeostasis, as the generation of intracellular ROS and MDA is significantly reduced by EGCG, which further prevents MMP collapse and suppresses caspase-3 activity. However, no evidence is observed for the chelation of EGCG with Cd2+ under neutral condition. Therefore, a clear conclusion from this work can be made that EGCG could inhibit Cd2+-induced apoptosis by acting as a ROS scavenger rather than a metal chelating agent.

ROS act as an upstream signal to mediate cadmium-induced mitophagy in mouse brain

As a well known generator of reactive oxygen species (ROS), cadmium (Cd) is found to be an effective inducer of mitophagy in mouse kidney and liver cells. Here, we aim to elucidate whether Cd can also initiate mitophagy in mouse brain and what role ROS play in this process. Our results showed that Cd caused overproduction of ROS. Meanwhile, Cd induced mitophagy, as indicated by the collapse of mitochondrial membrane potential (MMP), formation of mitophagosomes, increases of PINK1 level and LC3-II/LC3-I ratio and decrease of mitochondrial mass. Scavenging of ROS by N-acetyl-L-cysteine (NAC) or acetyl-L-carnitine (ALC) rescued MMP and mitochondrial mass, and squelched PINK1 level, mitochondrial accumulation of Parkin and LC3-II/LC3-I ratio, suggesting that ROS were associated with Cd-induced mitophagy. Cyclosporine A (CsA), an inhibitor of mitophagy, blocked Cd-induced mitophagy and PINK1/Parkin pathway but failed to suppress ROS increase, revealing that ROS are the causes rather than the results of Cd-induced mitophagy. In conclusion, this study suggested that ROS functioned on the upstream of PINK1/Parkin pathway to mediate Cd-induced mitophagy.

2,4-Dichlorophenol induces apoptosis in primary hepatocytes of grass carp (Ctenopharyngodon idella) through mitochondrial pathway

2,4-Dichlorophenol (2,4-DCP), a major type of chlorophenols, has been widely used to produce some herbicides and pharmaceuticals, yet due to its incomplete degradation and bioaccumulation characteristics, it is toxic to aquatic organisms. Apoptosis is one of the most severe outcomes of cell poisoning and injury. So far, the potential molecular mechanism of 2,4-DCP-induced apoptosis has not been reported. This study showed that 2,4-DCP significantly induced apoptosis in primary hepatocytes of grass carp (Ctenopharyngodon idella). 2,4-DCP exposure upregulated mRNA of caspase-3, reduced the mitochondrial membrane potential (Δψm), increased intracellular reactive oxygen species (ROS) and the Bax/Bcl-2 ratio, while protection of mitochondria with acetyl-l-carnitine hydrochloride (ALC) rescued 2,4-DCP-induced apoptosis, restored the Δψm and reduced the Bax/Bcl-2 ratio. Taken together, this is the first study that has identified that 2,4-DCP exposure induced apoptosis through the mitochondria-dependent pathway in primary hepatocytes of grass carp.

Synergic effect of 3'-azido-3'-deoxythymidine and arsenic trioxide in suppressing hepatoma cells.

The aim of this study was to investigate the synergic antitumor effects of arsenic trioxide (As2O3) and 3′-azido-3′-deoxythymidine (AZT) on hepatoma cells and explore the possible molecular basis of these effects. These results showed that AZT enhanced the inhibitory effect of As2O3 on HepG2 and SMMC-7721 cell growth. The IC50 of As2O3 in combination with AZT was lower than that of As2O3 alone. A concentration-dependent synergic effect of As2O3 and AZT (CI <1) was observed in all the tested combinations of these compounds. These results also showed that the combination of As2O3 and AZT dramatically and significantly increased the number of apoptotic cells in HepG2 and SMMC-7721 cells. Studies in vivo showed that the combination of As2O3 and AZT was statistically superior to either As2O3 or AZT alone in the treatment of tumor-bearing mice. As2O3 (1 mg/kg) containing AZT (50 mg/kg) inhibits proliferation of implanted hepatoma 22 by 56.35%. These results suggest that treating hepatoma with a combinination of As2O3 and AZT offers the advantages of reduced toxic side effects and improved therapeutic efficacy. To understand the mechanism through which As2O3 and AZT suppress tumors, we studied the effects of these compounds, both separately, and in combination, on telomerase and caspase-3 activity. The results showed that the growth inhibitory and apoptotic effects of As2O3 and AZT on human hepatoma cells could be related to the inhibition of telomerase and the activation of caspase 3.

The toxic effects of chlorophenols and associated mechanisms in fish.

Chlorophenols (CPs) are ubiquitous contaminants in the environment primarily released from agricultural and industrial wastewater. These compounds are not readily degraded naturally, and easily accumulate in organs, tissues and cells via food chains, further leading to acute and chronic toxic effects on aquatic organisms. Herein, we review the available literature regarding CP toxicity in fish, with special emphasis on the potential toxic mechanisms. CPs cause oxidative stress via generation of reactive oxygen species, induction of lipid peroxidation and/or oxidative DNA damage along with inhibition of antioxidant systems. CPs affect immune system by altering the number of mature B cells and macrophages, while suppressing phagocytosis and down-regulating the expression of immune factors. CPs also disrupt endocrine function by affecting hormone levels, or inducing abnormal gene expression and interference with hormone receptors. CPs at relatively higher concentrations induce apoptosis via mitochondria-mediated pathway, cell death receptor-mediated pathway, and/or DNA damage-mediated pathway. CPs at relatively lower concentrations promote cell proliferation, and foster cancers-prone environment by increasing the rate of point mutations and oxidative DNA lesions. These toxic effects in fish are induced directly by CPs per se or indirectly by their metabolic products. In addition, recent studies on the alteration of DNA methylation by CPs through high-throughput DNA sequencing analysis provide new insights into our understanding of the epigenetic mechanisms underlying CPs toxicity.

ATM mediates spermidine-induced mitophagy via PINK1 and Parkin regulation in human fibroblasts

The ATM (ataxia telangiectasia mutated) protein has recently been proposed to play critical roles in the response to mitochondrial dysfunction by initiating mitophagy. Here, we have used ATM-proficient GM00637 cells and ATM-deficient GM05849 cells to investigate the mitophagic effect of spermidine and to elucidate the role of ATM in spermdine-induced mitophagy. Our results indicate that spermidine induces mitophagy by eliciting mitochondrial depolarization, which triggers the formation of mitophagosomes and mitolysosomes, thereby promoting the accumulation of PINK1 and translocation of Parkin to damaged mitochondria, finally leading to the decreased mitochondrial mass in GM00637 cells. However, in GM05849 cells or GM00637 cells pretreated with the ATM kinase inhibitor KU55933, the expression of full-length PINK1 and the translocation of Parkin are blocked, and the colocalization of Parkin with either LC3 or PINK1 is disrupted. These results suggest that ATM drives the initiation of the mitophagic cascade. Our study demonstrates that spermidine induces mitophagy through ATM-dependent activation of the PINK1/Parkin pathway. These findings underscore the importance of a mitophagy regulatory network of ATM and PINK1/Parkin and elucidate a novel mechanism by which ATM influences spermidine-induced mitophagy.

AZT and emodin exhibit synergistic growth-inhibitory effects on K562/ADM cells by inducing S phase cell cycle arrest and suppressing MDR1 mRNA/p-gp protein expression.

Abstract Context: Previous studies have demonstrated that both 3′-azido-3′-deoxythymidine (AZT) and emodin, a traditional chemotherapy agent, can inhibit the growth of many types of cancer cells. Objective: This study aimed to evaluate the effect of AZT and emodin on adriamycin-resistant human chronic myelogenous leukemia (K562/ADM) cells, determine the expression of multidrug resistance 1 (MDR1) mRNA/p-glycoprotein (p-gp) protein, a protein known to induce resistance to anticancer agents, and to elucidate the underlying molecular mechanisms. Materials and methods: K562/ADM cells were treated with AZT (10–160 μM) or emodin (5–80 μM) for 24, 48 and 72 h and cell viability was measured using the MTT assay. The effect of AZT (16.5, 33 and 66 μM) and emodin (6.1, 17.6 and 33.2 μM) on K562/ADM cell cycle distribution was determined by flow cytometry, and MDR1 mRNA/p-gp protein expression was determined by real time RT-PCR and western blotting. Results: The growth suppression of emodin was dramatically enhanced by AZT in K562/ADM cells. The IC50 of AZT and emodin was lower than that of emodin alone. All examined combinations of AZT and emodin yielded a synergetic effect (CI < 1). Furthermore, AZT and emodin altered the cell cycle distribution and led to an accumulation of cells in S phase. Meanwhile, the expression of MDR1 mRNA/p-gp protein was markedly decreased. Discussion and conclusion: These results show a synergistic growth-inhibitory effect of AZT and emodin in K562/ADM cells, which is achieved through S phase arrest. MDR1 might ultimately be responsible for these phenomena.