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Exposure through arsenic-contaminated air and food caused by the burning of coal is a major environmental public health concern in Guizhou Province of China. Previous studies have shown that immunological dysfunction is involved in the pathogenesis and carcinogenesis of arsenic; however, knowledge regarding effective prevention measures have not been fully examined. The effect of Ginkgo biloba extract (EGb761) on arsenic-induced skin damage of human immortalized keratinocyte cells (HaCaT) was first evaluated in this study. The results showed that 200 μg/mL EGb761 can reduce the expression of miR-155-5p, and the indicators reflecting arsenic-induced skin damage (Krt1, Krt6c and Krt10) in arsenic-exposed cells ( P < 0.05), the expression levels of NF-AT1; the indicators reflecting arsenic-induced immunological dysfunction (IL-2, IFN-γ) in cells; and the levels of secreted IL-2 and IFN-γ in cell supernatants were significantly increased ( P < 0.05). Further randomized controlled double-blind experiments showed that compared to the placebo control group, the expression level of miR-155-5p in the plasma of the Ginkgo biloba intervention group, the indicators in the serum reflecting arsenic-induced skin damage (Krt1, Krt6c, and Krt10) and the epithelial-mesenchymal transformation (EMT) vimentin were significantly reduced ( P < 0.05), but the levels of NF-AT1 and the indicators reflecting arsenic-induced immunological dysfunction (IL-2, IFN-γ) and EMT (E-cadherin) in serum were significantly increased ( P < 0.05). Our study provides some limited evidence that Ginkgo biloba L. can increase the expression of NF-AT1 by downregulating the level of miR-155-5p, alleviating immunological dysfunction, and decreasing the expression of EMT biomarkers, thus indirectly improving arsenic-induced skin damage.

Increasing evidence supports the role of arsenic in dysregulated immune and inflammation responses, while, safe and effective treatments have not been fully examined.

Arsenic is a prevalent environmental pollutant that targets the nervous system of living beings. Recent studies indicated that microglial injury could contribute to neuroinflammation and is associated with neuronal damage. Nevertheless, the neurotoxic mechanism underlying the arsenic-induced microglial injury requires additional research. This study explores whether cathepsin B promotes microglia cell damage caused by NaAsO2. Through CCK-8 assay and Annexin V-FITC and PI staining, we discovered that NaAsO2 induced apoptosis in BV2 cells (a microglia cell line). NaAsO2 was verified to increase mitochondrial membrane permeabilization (MMP) and promote the generation of reactive oxygen species (ROS) through JC-1 staining and DCFDA assay, respectively. Mechanically, NaAsO2 was indicated to increase the expression of cathepsin B, which could stimulate pro-apoptotic molecule Bid into the activated form, tBid, and increase lysosomal membrane permeabilization by Immunofluorescence and Western blot assessment. Subsequently, apoptotic signaling downstream of increased mitochondrial membrane permeabilization was activated, promoting caspase activation and microglial apoptosis. Cathepsin B inhibitor CA074-Me could mitigate the damage of microglial. In general, we found that NaAsO2 induced microglia apoptosis and depended on the role of the cathepsin B-mediated lysosomal-mitochondrial apoptosis pathway. Our findings provided new insight into NaAsO2-induced neurological damage.

Arsenic is an environmental toxicant. Its overdose can cause liver damage. Autophagy has been reported to be involved in arsenite (iAs3+ ) cytotoxicity and plays a dual role in cell proliferation and cell death. However, the effect and molecular regulative mechanisms of iAs3+ on autophagy in hepatocytes remains largely unknown. Here, we found that iAs3+ exposure lead to hepatotoxicity by inducing autophagosome and autolysosome accumulation. On the one hand, iAs3+ promoted autophagosome synthesis by inhibiting E2F1/mTOR pathway in L-02 human hepatocytes. On the other, iAs3+ blocked autophagosome degradation partially via suppressing the expression of INPP5E and Rab7 as well as impairing lysosomal activity. More importantly, autophagosome and autolysosome accumulation induced by iAs3+ increased the protein level of E2F7a, which could further inhibit cell viability and induce apoptosis of L-02 cells. The treatment of Ginkgo biloba extract (GBE) effectively reduced autophagosome and autolysosome accumulation and thus alleviated iAs3+ -induced hepatotoxicity. Moreover, GBE could also protect lysosomal activity, promote the phosphorylation level of E2F1 (Ser364 and Thr433) and Rb (Ser780) as well as suppress the protein level of E2F7a in iAs3+ -treated L-02 cells. Taken together, our data suggested that autophagosome and autophagolysosome accumulation play a critical role for iAs3+ -induced hepatotoxicity, and GBE is a promising candidate for intervening iAs3+ induced liver damage by regulating E2F1-autophagy-E2F7a pathway and restoring lysosomal activity.

Endemic arsenicosis is a public health problem that affects thousands of people worldwide. However, the biological mechanism involved is not well characterized, and there is no specific treatment. Exposure to arsenic may be associated with immune-related problems. In the present work, we performed an investigation to determine whether the Th17/Treg balance was abnormal in peripheral blood mononuclear cells (PBMCs) of patients with arsenicosis caused by burning coal. Furthermore, we investigated the effect of Ginkgo biloba extract (GBE) on the Th17/Treg imbalance in patients with arsenicosis. In this trial, 81 arsenicosis patients and 37 controls were enrolled. The numbers of Th17 and Treg cells, as well as related transcription factors and serum cytokines, were determined at the beginning and end of the study. Patients with arsenicosis exhibited higher levels of Th17 cells, Th17-related cytokines (IL-17A and IL-6), and the transcription factor RORγt. There were lower levels of Treg cells, a Treg-related cytokine (IL-10), and the transcription factor Foxp3 as compared with controls. There was a positive correlation between the levels of Th17 cells and IL-17A and the levels of arsenic in hair. Arsenicosis patients were randomly assigned to a GBE treatment group or a placebo group. After 3 months of follow-up, 74 patients completed the study (39 cases in the GBE group and 35 in the placebo group). Administration of GBE to patient upregulated the numbers of Treg cells and the level of IL-10 and downregulated the numbers of Th17 cells and the levels of cytokines associated with Th17 cells. The mRNA levels of Foxp3 and RORγt were increased and decreased, respectively. These results indicated that exposure to arsenic is associated with immune-related problems. The present investigation describes a previously unknown mechanism showing that an imbalance of pro- and anti-inflammatory T cells is involved in the pathogenesis of arsenicosis and that a GBE exerts effects on arsenicosis through regulation of the pro- and anti-inflammatory T cell balance.

Arsenic poisoning is a geochemical disease that seriously endangers human health. The liver is one of the important target organs for arsenic poisoning, several studies have shown that oxidative stress plays an important role in arsenic-induced liver damage. However, the specific mechanism of arsenic-induced oxidative stress has not yet been fully elucidated, and currently, there are no effective intervention measures for the prevention and treatment of arsenic-induced liver damage. In this study, the effect of the Nrf2/GPX4 signaling pathway and oxidative stress in the arsenic-induced liver damage was first evaluated. The results show that arsenic can activate the Nrf2/GPX4 signaling pathway and increase the oxidative stress, which in turn promotes arsenic-induced liver damage in MIHA cells. Moreover, when we applied the Nrf2 inhibitor, the promoting effect of arsenic on liver damage was alleviated by inhibiting the activation of the Nrf2/GPX4 signaling pathway. Subsequently, the Rosa roxburghii Tratt [Rosaceae] (RRT) intervention experiments in cells and arsenic poisoning population were designed. The results revealed that RRT can inhibit Nrf2/GPX4 signaling pathway to reduce oxidative stress, thereby alleviates arsenic-induced liver damage. This study provides some limited evidence that arsenite can activate Nrf2/GPX4 signaling pathway to induce oxidative stress, which in turn promotes arsenic-induced liver damage in MIHA cells. The second major finding was that Kaji-ichigoside F1 may be a potential bioactive compound of RRT, which can inhibit Nrf2/GPX4 signaling pathway to reduce oxidative stress, thereby alleviates arsenic-induced liver damage. Our study will contribute to a deeper understanding of the mechanisms in arsenic-induced liver damage, these findings will identify a possible natural medicinal food dual-purpose fruit, RRT, as a more effective prevention and control strategies for arsenic poisoning.

Intratumoral hypoxia is a common feature of pancreatic cancer (PC) and also plays a role in its progression. However, hypoxia-regulated signatures in PC are still not completely understood. This study aimed to identify core hypoxia-associated genes and determine their underlying molecular mechanisms in PC cells. Transformer 2 alpha homolog (TRA2A) was found to be an important hypoxia-associated gene, which was upregulated in PC tissues and in PC cells cultured under hypoxia. High TRA2A expression was associated with advanced stage, poor differentiation, and lymph node metastasis. Under normoxic and hypoxic conditions, knockdown of TRA2A both markedly suppressed PC cell proliferation and motility in vitro and in vivo, as well as activation of the AKT pathway. Hypoxia-inducible factor 1 subunit alpha (HIF1α) upregulated the transcription of TRA2A by directly binding to its promoter. TRA2A showed a co-expression relationship with HIF1α in PC tissues. Overexpression of TRA2A alleviated the pro-inhibitive functions of HIF1α-inhibition on PC cell proliferation and motility under hypoxia. In conclusion, TRA2A is a crucial downstream gene of HIF1α that accelerates the proliferation and motility of PC cells. TRA2A may be a novel and practical molecular target for investigating the hypoxic response of PC cells. Abbreviations: TRA2A, transformer 2A protein; PC, pancreatic cancer; HIF1α, hypoxia-inducible factor 1-alpha; GEO, Gene Expression Omnibus; IHC, immunohistochemical staining.

Terpinen-4-ol (T4O), a compound isolated from the seeds of turmeric, has exhibited anti-malignancy, anti-aging, and anti-inflammatory properties in previous studies. However, the specific effects and molecular mechanisms of T4O on pancreatic cancer (PC) cells remain largely unknown. In this study, we demonstrated that T4O markedly suppressed PC cell proliferation and colony formation in vitro and induced apoptosis. Similarly, T4O significantly inhibited the migration and invasion of PC cells in vitro. Through RNA sequencing, 858 differentially expressed genes (DEGs) were identified, which were enriched in the Rhodopsin (RHO)/ Ras homolog family member A (RHOA) signaling pathway. Rho-associated coiled-coil containing protein kinase 2 (ROCK2), a DEG enriched in the RHO/RHOA signaling pathway, was considered as a key target of T4O in PC cells; it was significantly reduced after T4O treatment, highly expressed in PC tissues, and negatively associated with patient outcome. Overexpression of ROCK2 significantly reduced the inhibitory effects of T4O on PC cell proliferation and mobility. Moreover, T4O inhibited cell proliferation in vivo and decreased the Ki-67, cell nuclear antigen, EMT markers, and ROCK2 expression. In conclusion, we consider that T4O can suppress the malignant biological behavior of PC by reducing the expression of ROCK2, thus contributing to PC therapy.

Previous studies have reported controversial results on levels of inflammatory cytokines in patients with arsenic exposure. This study aims to evaluate the associations between arsenic exposure and inflammatory cytokines and C-reaction protein (CRP).We searched the databases including PubMed, Embase, Web of Science, and China national knowledge infrastructure (CNKI) for studies reporting levels of cytokines and CRP in patients with arsenic exposure compared to the controls. The retrieval time was from January 2000 to September 2022.13 observational studies involving 1665 arsenic exposed and 1091 unexposed individuals were included. Among these studies, 6 from China, 4 from India, 2 from Bangladesh and 1 from Turkey. Our result showed that interleukin (IL)-6, IL-8, and IL-12 levels were significantly higher in arsenic-exposed individuals compared to the control group, IL-2 level was significantly lower, and Tumor necrosis factor-α, Interferon-γ, CRP, and IL-10 levels were not changed. After sensitivity analyses, tumor necrosis factor-α and Interferon-γ levels were significantly higher in arsenic-exposed individuals compared to the control group. High heterogeneity was detected in most studies.Many cytokines (such as IL-6, IL-8, and IL-12) have altered in individuals with arsenic exposure, this indicates arsenic exposure could trigger the cell-mediated inflammatory response. Regular examining immune function (such as inflammatory cytokines) in individuals with the risk of arsenic exposure is important to human health.

Endemic arsenism caused by coal burning is a unique type of biogeochemical disease that only exists in China, and it is also a disease of element imbalances. Previous studies have shown that element imbalances are involved in the pathogenesis of arsenic; however, the interaction between the various elements and effective preventive measures have not been fully studied. This study first conducted a cross-sectional study of a total of 365 participants. The results showed that arsenic exposure can increase the content of elements (Al, As, Fe, Hg, K, and Na) in the hair (

Coal-burning type of arsenism, a chronic arsenism caused by environmental arsenic pollution, found firstly at Guizhou Province of China, manifested as the disruption of pro- and anti-inflammatory T cell balance and multiple organ damage, while no specific treatment for the arsenism patients. The effect of methylation of the forkhead box P3 (Foxp3) promoter region on arsenic-induced disruption of pro- and anti-inflammatory T cell balance was first evaluated in this study, between the control and arsenism groups. The results show that arsenic can induce the hypermethylation of 6 sites in the Foxp3 promoter by upregulating the expression of recombinant DNA Methyltransferase 1 (Dnmt1) mRNA, leading to the downregulation of Foxp3 mRNA, Tregs, and interleukin 10 (IL-10, anti-inflammatory cytokine) levels, and increased the levels of interleukin 17 (IL-17, pro-inflammatory cytokine) in the peripheral blood of patients with arsenic poisoning. Further randomized controlled double-blind experiments (including the placebo control groups and the Ginkgo biloba extract (GBE) intervention groups) showed that compared to the placebo control group or before GBE intervention, the levels of Dnmt1 mRNA, Foxp3 methylation, and IL-17 in the peripheral blood of the GBE intervention group were significantly decreased after intervention (P 0.05), but the levels of regulatory T cells (Tregs) and IL-10 were significantly increased (P 0.05). Our study provides some limited evidence that GBE can attenuate the disruption of pro- and anti-inflammatory balance of peripheral blood in arsenism patients by decreasing hypermethylation of the Foxp3 promoter region. This study provides scientific basis for further understanding a possible natural medicinal plant, GBE, as a more effective measure to prevent and control the disruption of pro- and anti-inflammatory balance caused by coal-burning type of arsenism.

Exposure through arsenic-contaminated air and food caused by the burning of coal is a major environmental public health concern in Guizhou Province of China. Previous studies have shown that immunological dysfunction is involved in the pathogenesis and carcinogenesis of arsenic; however, knowledge regarding effective prevention measures have not been fully examined. The effect of

Arsenic (As) is a toxic metalloid element that causes lung cancer and multiple non-malignant respiratory diseases. The toxicity of arsenic is mediated in part by epigenetic mechanisms, such as alterations in DNA methylation. While increasing studies have highlighted the potential importance of arsenic exposure to DNA methylation patterns and the subsequent risks for arsenic toxicity, there has been little focus on DNA hydroxymethylation-a negative regulation mechanism of DNA methylation. Therefore, this study aimed to investigate the relationship between genomic DNA methylation/hydroxymethylation and lung injury in arsenicosis populations. First, an increased risk of lung injury and exacerbation of lung function impairment in the arsenicosis population was confirmed. Levels of 5-methylcytosine/deoxycytidine (5 mC/dC), 5-hydroxymethylcytosine/deoxycytidine (5 hmC/dC) and 5 hmC/5 mC in genomic DNA of peripheral blood were decreased in the arsenicosis population compared to in the control. Additionally, multivariate logistic regression models showed an increased risk of chest digital radiography (DR) abnormalities when 5 hmC/dC and 5 hmC/5 mC levels were lower (OR = 3.12 and 3.96, all P 0.001). For 3 years follow-up, regression analysis showed that a decline in 5 hmC/dC was significantly associated with the decline of lung function parameters [forced vital capacity (FVC), forced expiratory volume in 1 s (FEV1) and maximal mid-expiratory flow (MMEF); β = 0.167, 0.122 and 0.073, respectively; all P 0.05]. Using the receiver operating characteristic (ROC) curve, a combination of 5 hmC/5 dC and 5 hmC/5 mC obtained the highest value for distinguishing lung injury in all subjects (AUC = 0.82, P 0.01). In contrast, in arsenicosis subjects, 5 hmC/dC was better at distinguishing lung injury (AUC = 0.84, P 0.01). Together, the results revealed that a decrease in genomic DNA hydroxymethylation markers was associated with lung injury in coal-burning arsenicosis populations. Genomic DNA hydroxymethylation could be a novel biomarker for identifying the risk of lung injury caused by coal-burning arsenicosis.

Exposure to arsenic-contaminated air and food caused by the burning of coal in unventilated indoor stoves is a major environmental public health concern in Guizhou Province, China. The liver is one of the main target organs for coal-fired arsenic exposure; however, there is little information about the risk assessment between cumulative arsenic exposure and the prevalence of liver damage. This study first evaluated the chronic daily intake (CDI) for two exposure pathways (inhalation and ingestion) and five environmental media (i.e., indoor and outdoor air, drinking water, rice, corn, and chili peppers) in 1998, 2006, 2014, and 2017. Then, the dose-effect and dose-response relationship between hair arsenic (HA) and cumulative arsenic (CA) levels and liver damage was analyzed. The results clearly show that the CDI in 1998 was 34.9 μg·kg-1·d-1, 22.9 μg·kg-1·d-1 in 2006, 11.7 μg·kg-1·d-1 in 2014, and 6.7 μg·kg-1·d-1 in 2017 in the arsenic exposure area. All of these values were higher than the daily baseline level of 3.0 μg·kg-1·d-1 as recommended by the Joint FAO/WHO Expert Committee on Food Additives (JECFA), and the increased HA and CA can increase the risk of coal-fired arsenic-induced liver damage. In addition, we analyzed the possible maximum acceptable CA exposure level for coal-fired arsenic-induced liver damage using the Bayesian benchmark dose. The recommended maximum acceptable CA exposure level for liver damage caused by coal-burning arsenic is 7120 mg. This study provides scientific insight into understanding the dose-response relationship of liver damage caused by coal-burning arsenic exposure and the monitoring and prevention of arsenic poisoning.

Chronic exposure to arsenic may result in the manifestation of damage in multiple organs or systems of the body. Arsenic-induced renal dysfunction has been determined, but their pathogenesis has not been fully examined. In this study, we measured the expression levels of miR-191 in plasma, the contents of pro-inflammatory (interleukin (IL)-6 and tumor necrosis factor alpha) and anti-inflammatory (IL-2 and transforming growth factor beta) cytokines, and renal dysfunction indicators (blood urea nitrogen, blood creatinine, uric acid, and cystatin C) in serum from control and arsenic poisoning populations and analyzed the relationship between the miR-191, cytokines, and renal dysfunction indicators. The results clearly show the alteration of miR-191 expression was significantly associated with arsenic-induced renal dysfunction. Overall, the association of miR-191, inflammatory response and renal dysfunction, is clearly supported by the current findings. In other words, miR-191 is involved in renal dysfunction in exposed populations by regulating inflammatory response caused by coal-burning arsenic. The study provides a scientific basis for further studies of the causes of the arsenic-induced renal dysfunction, the biological role of miR-191, and targeted prevention strategies.

Background: Currently, most arsenic (As) studies in populations are concerned with water-borne arsenicosis. However, residents in Xingren County of Guizhou Province, Southwest of China, represent a unique case of arsenicosis which is related to indoor combustion of high As-containing coal. This study aimed to assess the alterations of As levels and its risk factors in coal-borne arsenicosis residents during the past 20 years. Methods: Four follow-up investigations in Xingren County were selected from the year 1998 to 2017, a total of 245, 272, 584, and 309 residents were involved in the four investigations, respectively. Local external environmental medium (coal, soil, water, air, rice, corn and chili peppers) and biological samples (urine, hair) were collected at each time of investigation for total As analysis. Sociodemographics and lifestyles variables were extracted from the questionnaire investigation. Both univariate and multivariate unconditional logistic regression models were performed to analyze the variation of risk factors for coal-borne arsenicosis. Results: A substantial reduction of total As levels was observed both in external environmental medium and biological samples in the unique coal-borne arsenicosis region, especially since the year 2006. In addition, age, duration of consuming high As-containing coal and smoking status were found to be the most significant risk factors for coal-borne arsenicosis during the past 20 years by both two different logistic regression models. Room ventilation and grain drying modes were no longer to be risk factors since 1998 survey. Annual household income had always been an important protective factor for coal-borne arsenicosis in recent 20 years by both two different logistic regression models. Grain storage modes had become significant protective factor in 2014 and 2017 survey. A certain correlation between sex, education and coal-borne arsenicosis was observed by univariate logistic regression model but no clear links were found by multivariate logistic regression model. Conclusions: Considerable efforts to blocking As exposure from burning coal and As contaminated foods in this region are observed over the study period. Further practical health education programs may need to target individuals with long-term of As exposure, lower socioeconomic status and smoking in order to better prevent and control the occurrence and development of coal-borne arsenicosis. Keywords: Coal, Guizhou, Arsenic, Arsenicosis, Risk factors

Chronic exposure to arsenic remains a major environmental public health concern worldwide, affecting hundreds of millions of people. Arsenic-induced multiorgan damage and miRNA expression changes after arsenic exposure have been determined, but their associations and risks have not been fully examined. In this study, we measured the expression levels of five miRNAs in plasma from control and arsenic poisoned populations, and we analyzed the relationship between miRNAs and multiorgan damage. The results clearly show that the upregulation of miR-155 expression can increase the risk of arsenic induced skin damage (OR = 10.55; 95% CI: 6.02, 18.47); further, there is a link between the expression of miR-21 (OR = 11.84; 95% CI: 5.34, 26.28) and miR-145 (OR = 2.39; 95% CI: 1.61, 3.55) and liver damage, and miR-191 and kidney damage (OR = 3.65; 95% CI: 1.49, 8.93). In addition, we analyzed the diagnostic value of miRNAs associated with specific organ damage in arsenic-induced multiorgan damage. It was found that the miR-155 has a certain diagnostic value in arsenic-induced skin damage (AUC = 0.83), miR-21 and miR-145 have diagnostic value for liver damage (AUC = 0.80, 0.81) and miR-191 has diagnostic value for kidney damage (AUC = 0.83). This study provides the first comprehensive assessment of the association and risk of five miRNAs with arsenic-induced multiorgan damage. The study can provide a scientific basis for further understanding the causes of arsenic-induced multiorgan damage, identification of possible biological markers, and improvement of targeted prevention and control strategies.

Acinetobacter baumannii (A. baumannii), one of the major pathogens that causes severe nosocomial infections, is characterised by a high prevalence of drug resistance. It has been reported that A. baumannii triggers the NOD-like receptor 3 (NLRP3) inflammasome, but the role of its virulence-related outer membrane protein A (ompA) remains unclear. Therefore, this study aimed to explore the effects of ompA on the NLRP3 inflammasome and its underlying molecular mechanisms. Results showed that ompA enhanced inflammatory damage, which was reduced as a result of knockout of the ompA gene. Additionally, ompA-stimulated expression of NLRP3 inflammasome was significantly blocked by silencing caspase-1, but activation of NLRP3 inflammasome was not altered after silencing ASC; this indicated that ompA was dependent on the caspase-1 pathway to activate the inflammatory response. Simultaneously, the wild-type (WT) strains triggered NLRP3 inflammasome after inhibition of caspase-1 degradation by proteasome inhibitor MG-132, aggravating tissue damage. These findings indicated that ompA may be dependent on the caspase-1 pathway to enhance inflammation and exacerbate tissue damage. Taken together, these results confirmed a novel capsase-1-modulated mechanism underpinning ompA activity, which further reveals the NLRP3 inflammasome pathway as a potential immunomodulatory target against A. baumannii infections.

Arsenic is a well established human carcinogen that causes diseases of the lung. Some studies have suggested that hypoxia-inducible factors (HIFs) and microRNAs (miRNAs) are involved in human lung cancer; however, their molecular mechanisms that causally contribute to arsenite-caused malignant transformation of cells remain unclear. To elucidate the mechanisms of angiogenesis and metastasis of lung cancer caused by arsenite, we investigated the role of HIF-2α regulation of miRNA-191 (miR-191) in the angiogenic and metastatic properties of human bronchial epithelial (HBE) cells transformed by arsenite. In HBE cells, HIF-2α binds to the hypoxia response element (HRE) in the promoter region of miR-191 and initiates transcription of miR-191. Blocking of HIF-2α with siRNA inhibited the up-regulation of miR-191, Wilms’ tumor 1 (WT1) protein, matrix metalloproteinase 9 (MMP-9), vascular endothelial growth factor (VEGF), and the down-regulation of brain acid-soluble protein 1 (BASP1). In arsenite-transformed HBE (T-HBE) cells, down-regulation of HIF-2α by siRNA blocked the process of angiogenesis and decreased their neoplastic properties and metastatic capacity, which were reversed by over-expression of miR-191 or by up-regulating WT1. Thus, HIF-2α up-regulates WT1 via miR-191, both of which are involved in the angiogenesis and metastasis of T-HBE cells. The results present a better understanding of the processes involved in lung cancer caused by arsenite exposure.

Arsenic is well established as a human carcinogen, but the molecular mechanisms leading to arsenic-induced carcinogenesis are complex and elusive. It is also not known if lncRNAs are involved in arsenic-induced liver carcinogenesis. We have found that MALAT1, a non-coding RNA, is over-expressed in the sera of people exposed to arsenite and in hepatocellular carcinomas (HCCs), and MALAT1 has a close relation with the clinicopathological characteristics of HCC. In addition, hypoxia-inducible factor (HIF)-2α is up-regulated in HCCs, and MALAT1 and HIF-2α have a positive correlation in HCC tissues. During the malignant transformation of human hepatic epithelial (L-02) cells induced by a low concentration (2.0 μM) of arsenite, MALAT1 and HIF-2α are increased. In addition, arsenite-induced MALAT1 causes disassociation of the von Hippel-Lindau (VHL) protein from HIF-2α, therefore, alleviating VHL-mediated HIF-2α ubiquitination, which causes HIF-2α accumulation. In turn, HIF-2α transcriptionally regulates MALAT1, thus forming a positive feedback loop to ensure expression of arsenite-induced MALAT1 and HIF-2α, which are involved in malignant transformation. Moreover, MALAT1 and HIF-2α promote the invasive and metastatic capacities of arsenite-induced transformed L-02 cells and in HCC-LM3 cells. The capacities of MALAT1 and HIF-2α to promote tumor growth are validated in mouse xenograft models. In mice, arsenite induces an inflammatory response, and MALAT1 and HIF-2α are over-expressed. Together, these findings suggest that the MALAT1/HIF-2α feedback loop is involved in regulation of arsenite-induced malignant transformation. Our results not only confirm a novel mechanism involving reciprocal regulation between MALAT1 and HIF-2α, but also expand the understanding of the carcinogenic potential of arsenite.

Arsenic is a known human carcinogen and the mechanisms underlying arsenic-induced tumorigenesis remain elusive. Circular RNAs (circRNAs) are involved in the development of cancers, generally acting as sponges for microRNAs (miRNAs). Here, we screened the circRNA expression profiles of HaCaT cells, which are immortalized human keratinocytes, and arsenite-transformed HaCaT cells (T-HaCaT). The presence of has_circRNA-008913 (circ008913) was confirmed in HaCaT cells. Among the circRNAs down-regulated in T-HaCaT cells, circ008913 showed the greatest decrease and was chosen for further research. In HaCaT cells, arsenite induced increases of mRNA levels of the genes for cell-surface markers (k5 and CD34) of skin stem cells, decreases of DAB2IP, and increases of ZEB1. MicroRNA (miR)-889 suppressed the expression of DAB2IP and was involved in regulation of cancer stem cells (CSCs). Moreover, overexpression of circ008913 with pLCDH-circ008913 or transfection with an miR-889 inhibitor reduced the capacity of T-HaCaT cells for colony formation, invasion, migration, and the sizes of tumors in nude mice, effects that were reversed by co-transfection with an miR-889 mimic. These results suggest that, in HaCaT cells, arsenite decreases circ008913 levels, which act as a sponge for miR-889 and down-regulate the miR-889 target, DAB2IP, which, in turn, up-regulates ZEB1, increases mRNA levels of the cell-surface markers of skin stem cells, and is involved in arsenite-induced acquisition of CSC-like properties that lead to malignant transformation. The results also indicate that circ008913 functions as a competing endogenous RNA (ceRNA) for miR-889, which is involved in the arsenite-induced acquisition of CSC-like properties by regulation of DAB2IP and elucidate a previously unknown mechanism between arsenite-induced acquisition of CSC-like properties and carcinogenesis.

Arsenic, an established human carcinogen, causes genetic toxicity. However, the molecular mechanisms involved remain unknown. MicroRNAs (miRNAs) are regulators that participate in fundamental cellular processes. In the present investigation, we selected, as research subjects, patients with arsenic poisoning caused by burning of coal in Guizhou Province, China. For these patients, the plasma levels of miR-145 were up-regulated. In L-02 cells, arsenite, an active form of arsenic, induced up-regulation of miR-145 and down-regulation of ERCC1 and ERCC2, and caused DNA damage. For L-02 cells, transfection with an miR-145 inhibitor prevented arsenite-induced DNA damage and decreased ERCC2 levels. Luciferase reporter assays showed that miR-145 regulated ERCC2 expression by targeting the 3'-UTR of ERCC2, but not that for ERCC1. The present results demonstrate that arsenite induces the over-expression of miR-145 and inhibits DNA repair via targeting ERCC2, thus promoting DNA damage. The information provides a new mechanism for arsenic-induced liver injury.

Endemic arsenism, caused by burning coal containing high levels of arsenic, is found only in the Guizhou and Shanxi Provinces of China. Dysregulated microRNAs (miRNAs), detected in the blood, are emerging as promising biomarkers. At present, little is known about the change and clinical efficacy of circulating miRNAs in patients with endemic arsenism produced by burning of coal. Here, we determined, by using TaqMan Human miRNA Array Chips, the differential expression of plasma miRNAs between patients with arsenism caused by coal-burning and a control group. Four increased miRNAs (miR-21, miR-145, miR-155, and miR-191) were verified in a larger sample by quantitative real-time PCR. Furthermore, bioinformatics and Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway enrichment analyses were used to associate changes in plasma levels of the miRNAs with their functions and their effects on various pathways. The results of chip array assays show that the levels of miR-21, miR-141, miR-148a, miR-145, miR-155, miR-191, miR-218, and miR-491 were most prominently increased and that the levels of miR-200b, miR-200c, miR-26, and miR-34c were decreased. The qRT-PCR results confirm that the circulating levels of miR-21, miR-145, miR-155, and miR-191 are increased in patients with arsenism caused by coal-burning. KEGG analyses show that these miRNAs inhibit the target genes of pathways related to immune inflammation, oxidative stress, and DNA damage repair. Therefore, the four miRNAs may be biomarkers of endemic arsenism caused by coal-burning. Further studies with larger samples should be performed to confirm these findings and to elucidate the underlying mechanisms.

Arsenic is a well established human carcinogen that causes diseases of the lung. Some studies have suggested that hypoxia-inducible factors (HIFs) and microRNAs (miRNAs) are involved in human lung cancer; however, their molecular mechanisms that causally contribute to arsenite-caused malignant transformation of cells remain unclear. To elucidate the mechanisms of angiogenesis and metastasis of lung cancer caused by arsenite, we investigated the role of HIF-2α regulation of miRNA-191 (miR-191) in the angiogenic and metastatic properties of human bronchial epithelial (HBE) cells transformed by arsenite. In HBE cells, HIF-2α binds to the hypoxia response element (HRE) in the promoter region of miR-191 and initiates transcription of miR-191. Blocking of HIF-2α with siRNA inhibited the up-regulation of miR-191, Wilms' tumor 1 (WT1) protein, matrix metalloproteinase 9 (MMP-9), vascular endothelial growth factor (VEGF), and the down-regulation of brain acid-soluble protein 1 (BASP1). In arsenite-transformed HBE (T-HBE) cells, down-regulation of HIF-2α by siRNA blocked the process of angiogenesis and decreased their neoplastic properties and metastatic capacity, which were reversed by over-expression of miR-191 or by up-regulating WT1. Thus, HIF-2α up-regulates WT1