NSUN2 promotes lung adenocarcinoma progression through stabilizing PIK3R2 mRNA in an m5C-dependent manner.

It is well known that 5-methylcytosine (m5C) is involved in variety of crucial biological processes in cancers. However, its biological roles in lung adenocarcinoma (LAUD) remain to be determined. The LUAD samples were used to assess the clinical value of NOP2/Sun RNA Methyltransferase 2 (NSUN2). Dot blot was used to determine global m5C levels. ChIP and dual-luciferase assays were performed to investigate the MYC-associated zinc finger protein (MAZ)-binding sites in NSUN2 promoter. RNA-seq was used to explore the downstream molecular mechanisms of NSUN2. Dual luciferase reporter assay, m5C-RIP-qPCR, and mRNA stability assay were conducted to explore the effect of NSUN2-depletion on target genes. Cell viability, transwell, and xenograft mouse model were designed to demonstrate the characteristic of NSUN2 in promoting LUAD progression. The m5C methyltransferase NSUN2 was highly expressed and caused elevated m5C methylation in LUAD samples. Mechanistically, MAZ positively regulated the transcription of NSUN2 and was related to poor survival of LUAD patients. Silencing NSUN2 decreased the global m5C levels, suppressed proliferation, migration and invasion, and inhibited activation of PI3K-AKT signaling in A549 and SPAC-1 cells. Phosphoinositide-3-Kinase Regulatory Subunit 2 (PIK3R2) was upregulated by NSUN2-mediated m5C methylation by enhancing its mRNA stabilization and activated the phosphorylation of the PI3K-AKT signaling. The present study explored the underlying mechanism and biological function of NSUN2-meditated m5C RNA methylation in LUAD. NSUN2 was discovered to facilitate the malignancy progression of LUAD through regulating m5C modifications to stabilize PIK3R2 activating the PI3K-AKT signaling, suggesting that NSUN2 could be a novel biomarker and promising therapeutic target for LUAD patients.

TXNIP-NLRP3-GSDMD axis-mediated inflammation and pyroptosis of islet ß-cells is involved in cigarette smoke-induced hyperglycemia, which is alleviated by andrographolide.

Epidemiologic surveys have indicated that cigarette smoking is an important risk factor for diabetes, but its mechanisms remain unclear. Andrographolide, an herb traditionally utilized in medicine, provides anti-inflammatory benefits for various diseases. In the present work, 265 patients with Type 2 diabetes (T2D) were investigated, and male C57BL/6 mice were exposed to cigareete smoke (CS) and/or to intraperitoneally injected andrographolide for 3 months. To elucidate the mechanism of CS-induced hyperglycemia and the protective mechanism of andrographolide, MIN6 cells were exposed to cigarette smoke extract (CSE) and/or to andrographolide. Our data from 265 patients with T2D showed that urinary creatinine and serum inflammatory cytokines (interleukin 6 (IL-6), IL-8, IL-1ß, and tumor necrosis factor α (TNF-α)) increased with smoking pack-years. In a mouse model, CS induced hyperglycemia, decreased insulin secretion, and elevated inflammation and pyroptosis in ß-cells of mice. Treatment of mice with andrographolide preserved pancreatic function by reducing the expression of inflammatory cytokines; the expression of TXNIP, NLRP3, cleaved caspase 1, IL-1ß; and the N-terminal of gasdermin D (GSDMD) protein. For MIN6 cells, CSE caused increasing secretion of the inflammatory cytokines IL-6 and IL-1ß, and the expression of TXNIP and pyroptosis-related proteins; however, andrographolide alleviated these changes. Furthermore, silencing of TXNIP showed that the blocking effect of andrographolide may be mediated by TXNIP. In sum, our results indicate that CS induces hyperglycemia through TXNIP-NLRP3-GSDMD axis-mediated inflammation and pyroptosis of islet ß-cells and that andrographolide is a potential therapeutic agent for CS-induced hyperglycemia.

CircRNA_0026344 via miR-21 is involved in cigarette smoke-induced autophagy and apoptosis of alveolar epithelial cells in emphysema.

Cigarette smoke (CS), a main source of indoor air pollution, is a primary risk factor for emphysema, and aberrant cellular autophagy is related to the pathogenesis of emphysema. Circular RNAs (circRNAs) affect the expression of mRNAs via acting as microRNA (miRNA) sponges, but their role in emphysema progression is not established. In the present investigation, CS, acting on alveolar epithelial cells, caused higher levels of miR-21, p-ERK, and cleaved-caspase 3 and led to lower levels of circRNA_0026344 and PTEN, which induced autophagy and apoptosis. miR-21 suppressed the expression of PTEN, which was involved in the regulation of autophagy and apoptosis. Further, in alveolar epithelial cells, overexpression of circRNA_0026344 blocked cigarette smoke extract (CSE)-induced autophagy and apoptosis, but this blockage was reversed by upregulation of miR-21 with a mimic. These results demonstrated that, in alveolar epithelial cells, CS decreases circRNA_0026344 levels, which sponge miR-21 to inhibit the miR-21 target, PTEN, which, in turn, activates ERK and thereby promotes autophagy and apoptosis, leading to emphysema. Thus, for emphysema, circRNA_0026344 regulates the PTEN/ERK axis by sponging miR-21, which is associated with the CS-induced autophagy and apoptosis of alveolar epithelial cells. In sum, the present investigation identifies a novel mechanism for CS-induced emphysema and provides information useful for the diagnosis and treatment of CS-induced emphysema.

Long non-coding RNA LINC01480 is activated by Foxo3a and promotes hydroquinone-induced TK6 cell apoptosis by inhibiting the PI3K/AKT pathway.

Long non-coding RNAs (lncRNAs) have been shown to play a critical role in the damage caused to the body by environmental exogenous chemicals; however, few studies have explored their effects during exposure to benzene and its metabolite, hydroquinone (HQ). An emerging lncRNA, LINC01480, was found to be associated with the immune microenvironment of some cancers, but its specific function remains unknown. Therefore, this study aimed to investigate the role of LINC01480 in HQ-induced apoptosis. The biological function of LINC01480 was investigated through gain-of-function and loss-of-function experiments. Mechanically, nuclear-cytoplasmic fractionation experiment, chromatin immunoprecipitation (ChIP), dual-luciferase reporter assay, and rescue experiments were performed. In this study, when TK6 cells were treated with HQ (0, 5, 10, and 20 µM) for 12, 24, 48, and 72 h, the expression of LINC01480 was increased in a dose-dependent manner. Meanwhile, the phosphorylation levels of PI3K and AKT decreased, and apoptosis increased. As compared to the control group, HQ-induced apoptosis was significantly reduced, and the relative survival rate of TK6 cells increased after silencing LINC01480, while overexpression of LINC01480 further sensitized TK6 cells to HQ-induced apoptotic cell death. LINC01480 negatively regulated the PI3K/AKT pathway in TK6 cells, and the apoptosis-inhibiting effect of LINC01480 silencing was reversed after inhibition of the PI3K/AKT pathway. In addition, ChIP and the dual-luciferase reporter assays showed that the transcription factor Foxo3a promoted LINC01480 transcription by directly binding to the promoter regions - 149 to - 138 of LINC01480. Moreover, short-term HQ exposure promoted the expression of Foxo3a. From these findings, we can conclude that LINC01480 is activated by Foxo3a, and promotes HQ-induced apoptosis by inhibiting the PI3K/AKT pathway, suggesting that LINC01480 might become a possible target for therapeutic intervention of HQ-induced toxicity.

The aberrant cross-talk of epithelium-macrophages via METTL3-regulated extracellular vesicle miR-93 in smoking-induced emphysema.

Cigarette smoke (CS), a complex chemical indoor air pollutant, induces degradation of elastin, resulting in emphysema. Aberrant cross-talk between macrophages and bronchial epithelial cells is essential for the degradation of elastin that contributes to emphysema, in which extracellular vesicles (EVs) play a critical role. The formation of N6-methyladenosine (m6A) is a modification in miRNA processing, but its role in the development of emphysema remains unclear. Here, we established that production of excess mature microRNA-93 (miR-93) in bronchial epithelial cells via enhanced m6A modification was mediated by overexpressed methyltransferase-like 3 (METTL3) induced by CS. Mature miR-93 was transferred from bronchial epithelial cells into macrophages by EVs. In macrophages, miR-93 activated the JNK pathway by targeting dual-specificity phosphatase 2 (DUSP2), which elevated the levels of matrix metalloproteinase 9 (MMP9) and matrix metalloproteinase 12 (MMP12) and induced elastin degradation, leading to emphysema. These results demonstrate that METTL3-mediated formation of EV miR-93, facilitated by m6A, is implicated in the aberrant cross-talk of epithelium-macrophages, indicating that this process is involved in the smoking-related emphysema. EV miR-93 may use as a novel risk biomarker for CS-induced emphysema.

miRNA-21, which disrupts metabolic reprogramming to facilitate CD4+ T cell polarization toward the Th2 phenotype, accelerates arsenite-induced hepatic fibrosis.

Elevated levels of arsenic may be present in groundwater, and long-term exposure to arsenic increases hepatic fibrosis. T helper 2 (Th2) cells are involved in the fibrotic cascade, and cell metabolism is a regulatory factor participating in CD4+ T cell differentiation and function. However, the mechanism for Th2 cell regulation of arsenite-induced hepatic fibrosis is not fully understood. In present study, for arsenite-fed mice, activated hepatic stellate cells may be involved in the infiltration of CD4+ T cells, accompanied by up-regulation of GATA3, a transcription factor, and IL-13, the major Th2 cytokine. Exposed to arsenite, Jurkat cells had increased aerobic glycolysis to promote the cell cycle and cell proliferation. Further, this process elevated levels of marker molecules, including those of the Th2 paradigm characterized by GATA3, IL-4, and IL-13. LX-2 cells were activated when treated with culture medium from Jurkat cells exposed to arsenite. miR-21 may be a therapeutic target for arsenite-induced hepatic fibrosis. In vitro, miR-21 knock-down caused inhibition of the PTEN/PI3K/AKT pathway induced by arsenite. It also reversed the elevated glycolysis and the accelerated cell cycle and cell proliferation. Indeed, this alteration led to diminished expression of GATA3, IL-4, and IL-13 in T cells differentiated under Th2 conditions, which inhibits activation of LX-2 cells. Consistent with the results in vitro, miR-21 knock-out in mice reversed hepatic fibrosis and attenuated the levels of GATA3 and IL-13 induced by arsenite. These findings indicate that miR-21 regulates the glycolysis of CD4+ T cells through the PTEN/PI3K/AKT pathway to accelerate the cell cycle, thereby facilitating CD4+ T cell polarization toward Th2 and releasing the fibrogenic factor IL-13, which participates in arsenite-associated hepatic fibrosis. Inhibition of Th2 polarization of CD4+T cells or miR-21 could be a therapeutic strategy to combat hepatic fibrosis caused by exposure to arsenic.

LncRNA H19 via miR-29a-3p is involved in lung inflammation and pulmonary fibrosis induced by neodymium oxide.

The occupational and environmental health safety of rare earths has attracted considerable attention. In China, the rare earth neodymium oxide (Nd2O3) is extensively refined and utilized. However, the mechanisms of Nd2O3-induced lung injury are elusive. In the present study, we found that exposure of mice to Nd2O3 caused an inflammatory reaction and fibrosis in lung tissues, which was in relation to the Nd2O3-induced higher levels of the lncRNA H19 (H19), tumor necrosis factor receptor 1 (TNFRSF1A), p-p65, and p-IKKß and lower levels of miR-29a-3p. Further, in mouse monocyte macrophage leukemia cells (RAW264.7), Nd2O3 induced an inflammatory reaction, increases of H19 and TNFRSF1A levels, decreases of miR-29a-3p levels, and activation of the nuclear factor (NF)-κB signaling pathway. Further, we established that miR-29a-3p regulates TNFRSF1A expression. Up-regulation of miR-29a-3p and down-regulation of H19 blocked the Nd2O3-induced secretion of TNF-α, MIP-1α, and IL-6; the increases of TNFRSF1A levels; and activation of the NF-κB signaling pathway in RAW264.7 cells. Further, in Nd2O3-treated RAW26.4 cells, H19 inhibited the expression of miR-29a-3p, which targets TNFRSF1A, and activated the NF-κB signaling pathway to enhance the expression of TNF-α, MIP-1α, and IL-6. Moreover, for mice, up-regulation of miR-29a-3p reversed lung tissue inflammation, pulmonary fibrosis, and activation of the NF-κB signaling pathway induced by Nd2O3. In sum, the present investigation shows that H19 via miR-29a-3p is involved in lung inflammation and pulmonary fibrosis induced by Nd2O3, which is a mechanism for the Nd2O3-induced lung inflammatory response and pulmonary fibrosis. This information is useful for development of a biomarker of Nd2O3-induced lung injury.

SIRT1 regulated hexokinase-2 promoting glycolysis is involved in hydroquinone-enhanced malignant progression in human lymphoblastoid TK6 cells.

Reprogramming of cellular metabolism is a vital event during tumorigenesis. The role of glycolysis in malignant progression promoted by hydroquinone (HQ), one of the metabolic products of benzene, remains to be understood. Recently, we reported the overexpression of sirtuin 1 (SIRT1) in HQ-enhanced malignant progression of TK6 cells and hypothesized that SIRT1 might contribute to glycolysis and favor tumorigenesis. Our data showed that acute exposure of TK6 cells to HQ for 48 h inhibited glycolysis, as indicated by reduction in glucose consumption, lactate production, hexokinase activity, and the expression of SIRT1 and glycolytic enzymes, including HIF-1α, hexokinase-2 (HK-2), ENO-1, glucose transporter 1 (Glut-1), and lactic dehydrogenase A (LDHA). Knockdown of SIRT1 or inhibition of glycolysis using the glycolytic inhibitor 2-deoxy-D-glucose (2-DG) downregulated the levels of SIRT1 and glycolytic enzymes and significantly enhanced HQ-induced cell apoptosis, although knockdown of SIRT1 or 2-DG alone had little effect on apoptosis. Furthermore, immunofluorescence and Co-IP assays demonstrated that SIRT1 regulated the expression of HK-2, and HQ treatment caused a decrease in SIRT1 and HK-2 binding to mitochondria. Importantly, we found that glycolysis was promoted with increasing HQ treatment weeks. Long-term HQ exposure increased the expression of SIRT1 and several glycolytic enzymes and promoted malignant cell progression. Moreover, compared with the PBS group, glucose consumption and lactate production increased after 10 weeks of HQ exposure, and the protein levels of SIRT1 and HK-2 were increased after 15 weeks of HQ exposure, while those of Glut-1, ENO-1, and LDHA were elevated. In addition, SIRT1 knockdown HQ 19 cells exhibited decreased lactate production, glucose consumption, glycolytic enzymes expression, cell growth, and tumor formation in nude mice. Our findings identify the high expression of SIRT1 as a strong oncogenic driver that positively regulates HK-2 and promotes glycolysis in HQ-accelerated malignant progression of TK6 cells.

miR-21-regulated M2 polarization of macrophage is involved in arsenicosis-induced hepatic fibrosis through the activation of hepatic stellate cells.

Arsenicosis induced by chronic exposure to arsenic is recognized as one of the main damaging effects on public health. Exposure to arsenic can cause hepatic fibrosis, but the molecular mechanisms by which this occurs are complex and elusive. It is not known if miRNAs are involved in arsenic-induced liver fibrosis. We found that in the livers of mice exposed to arsenite, there were elevated levels of microRNA-21 (miR-21), phosphorylated mammalian target of rapamycin (p-mTOR), and arginase 1 (Arg1); low levels of phosphatase and tensin homolog (PTEN); and more extensive liver fibrosis. For cultured cells, arsenite-induced miR-21, p-mTOR, and Arg1; decreased PTEN; and promoted M2 polarization of macrophages derived from THP-1 monocytes (THP-M), which caused secretion of fibrogenic cytokines, including transforming growth factor-ß1. Coculture of arsenite-treated, THP-M with LX-2 cells induced α-SMA and collagen I in the LX-2 cells and resulted in the activation of these cells. Downregulation of miR-21 in THP-M inhibited arsenite-induced M2 polarization and activation of LX-2 cells, but cotransfection with PTEN siRNA or a miR-21 inhibitor reversed this inhibition. Moreover, knockout of miR-21 in mice attenuated liver fibrosis and M2 polarization compared with WT mice exposed to arsenite. Additionally, LN, PCIII, and HA levels were higher in patients with higher hair arsenic levels, and levels of miR-21 were higher than controls and positively correlated with PCIII, LN, and HA levels. Thus, arsenite induces the M2 polarization of macrophages via miR-21 regulation of PTEN, which is involved in the activation of hepatic stellate cells and hepatic fibrosis. The results establish a previously unknown mechanism for arsenicosis-induced fibrosis.

MicroRNA-191 blocking the translocation of GLUT4 is involved in arsenite-induced hepatic insulin resistance through inhibiting the IRS1/AKT pathway.

Environmental exposure to arsenic can cause a variety of health problems. Epidemiological and experimental studies have established a diabetogenic role for arsenic, but the mechanisms responsible for arsenic-induced impairment of insulin action are unclear. MicroRNAs (miRNAs) are involved in various metabolic disorders, particularly in the development of insulin resistance. The present study investigated whether arsenite, an active form of arsenic, induces hepatic insulin resistance and the mechanisms underlying it. After male C57BL/6J mice were exposed to arsenite (0 or 20 ppm) in drinking water for 12 months, intraperitoneal glucose tolerance tests (IPGTTs) and insulin tolerance tests (ITTs) revealed an arsenite-induced glucose metabolism disorder. Hepatic glycogen levels were lower in arsenite-exposed mice. Further, for livers of mice exposed to arsenite, miR-191 levels were higher, and protein levels of insulin receptor substrate 1 (IRS1), p-IRS1, and phospho-protein kinase B (p-AKT) were lower. Further, glucose transporter 4 (GLUT4) had lower levels on the plasma membrane. For insulin-treated L-02 cells, arsenite decreased glucose consumption and glycogen levels, increased miR-191 levels, and inhibited the IRS1/AKT pathway and the translocation of GLUT4 from the cytoplasm to the plasma membrane. For insulin-treated L-02 cells, the decreases of glucose consumption, glycogen levels, GLUT4 on the plasma membrane, and p-AKT levels induced by arsenite were reversed by SC79 (agonist of AKT) and an miR-191 inhibitor; these effects caused by miR-191 inhibitor were restored by IRS1 siRNA. In insulin-treated L-02 cells, miR-191, via IRS1, was involved in the arsenite-induced decreases of glucose consumption and glycogen levels and in inhibition of the translocation of GLUT4. Thus, miR-191 blocking the translocation of GLUT4 was involved in arsenite-induced hepatic insulin resistance through inhibiting the IRS1/AKT pathway. Our study reveals a mechanism for arsenite-induced hepatic insulin resistance, which provides clues for discovering biomarkers for the development of type 2 diabetes and for prevention and treatment of arsenic poisoning.

The ubiquitination and acetylation of histones are associated with male reproductive disorders induced by chronic exposure to arsenite.

Exposure to arsenic, which occurs via various routes, can cause reproductive toxicity. However, the mechanism for arsenic-induced reproductive disorders in male mice has not been extensively investigated. Here, 6-week-old male mice were dosed to 0, 5, 10, or 20 ppm sodium arsenite (NaAsO2), an active form of arsenic, in drinking water for six months. For male mice exposed to arsenite, fertility was lower compared to control mice. Moreover, for exposed mice, there were lower sperm counts, lower sperm motility, and higher sperm malformation ratios. Further, the mRNA and protein levels of the gonadotropin-regulated testicular RNA helicase (DDX25) and chromosome region maintenance-1 protein (CRM1), along with proteins associated with high mobility group box 2 (HMGB2), phosphoglycerate kinase 2 (PGK2), and testicular angiotensin-converting enzyme (tACE) were lower. Furthermore, chronic exposure to arsenite led to lower H2A ubiquitination (ubH2A); histone H3 acetylation K18 (H3AcK18); and histone H4 acetylations K5, K8, K12, and K16 (H4tetraAck) in haploid spermatids from testicular tissues. These alterations disrupted deposition of protamine 1 (Prm1) in testes. Overall, the present results indicate that the ubiquitination and acetylation of histones is involved in the spermiogenesis disorders caused by chronic exposure to arsenite, which points to a previously unknown connection between the modification of histones and arsenite-induced male reproductive toxicity.

Pre-miR-27a rs895819 polymorphism and risk of diffuse large B-cell lymphoma.

BACKGROUND: Recently, several studies have investigated the relationship between Pre-miR-27a rs895819 polymorphism and risk of various cancers. However, the relationship between rs895819 and diffuse large B-cell lymphoma (DLBCL) has not been well known. METHODS: In this study, we conducted a case-control study to explore the role of Pre-miR-27a rs895819 in risk of DLBCL. The PCR-TaqMan and luciferase assays and in vitro experiments were used to evaluate polymorphism function. RESULTS: As a result, we found subjects carrying with rs895819 AG/GG genotype had a significantly decreased risk when compared with those carrying the AA genotype. Further qPCR assay showed that the DLBCL patients carrying AG/GG genotypes showed a lower level of mature miR-27a when compared with patients carrying AA genotype. Moreover, miR-27a levels were upregulated in DLBCL tissues compared with normal lymphoid tissues. Further in vitro experiments showed that miR-27a might function as an oncogene through target TGFBR1. In addition, TGFBR1 overexpression rescues effects of miR-27a inhibitor on DLBCL cells phenotypes. CONCLUSIONS: In conclusion, these findings indicate that rs895819 A > G might reduce the expression of mature miR-27a, and leading a higher level of TGFBR1, ultimately inhibiting the development of DLBCL.

NF-κB-regulation of miR-155, via SOCS1/STAT3, is involved in the PM2.5-accelerated cell cycle and proliferation of human bronchial epithelial cells.

Air pollution, especially fine particulate matter (PM2.5, particles <2.5 µm in size), induces adverse health effects on the respiratory system. Uncontrolled proliferation of bronchial epithelial cells, resulting from deregulated cell cycle progression, contributes to pulmonary homeostatic imbalance. Although dysregulation of miRNAs is involved in a variety of pathophysiologic processes, the role of miRNAs in lung injury caused by PM2.5 is unclear. In the present study, we found that different concentrations of PM2.5 caused a biphasic effect on proliferation of human bronchial epithelial (HBE) cells. PM2.5 induced an aberrant cell cycle and proliferation of HBE cells, and up-regulated miR-155 levels with a concentration-dependent manner. High miR-155 expression, mediated by NF-κB activation, produced an accelerated G1/S phase and cell proliferation though the STAT3 pathway, which targeted SOCS1. These findings indicate that NF-κB-mediated miR-155 induces an altered cell cycle through epigenetic modulation of the SOCS1/STAT3 signaling pathway and provide a mechanism for the biphasic effect of different concentrations of PM2.5 in inducing respiratory injury.

Role of B-Cell Lymphoma 2 Ovarian Killer (BOK) in Acute Toxicity of Human Lung Epithelial Cells Caused by Cadmium Chloride.

BACKGROUND B-cell lymphoma 2 (BCL-2) ovarian killer (BOK) is a Bcl-2 family member with sequence homology to pro-apoptotic BAX and BAK, but its physiological and pathological roles remain largely unclear. Exposure of cells to cadmium may cause DNA damage, decrease DNA repair capacity, and increase genomic instability. MATERIAL AND METHODS The present study investigated the effects of BOK on the toxicity of cadmium chloride (CdCl2) to human bronchial epithelial (16HBE) cells. We constructed BOK over-expressing (16HBE-BOK) cells and BOK knockdown (16HBE-shBOK) cells using the BOK-ORF plasmid and BOK-siRNA. qRT-PCR for BOK mRNA expression. We used Trypan blue exclusion assay for cell growth, MTT colorimetric assays for cells inhibition rate, and Comet assays for detecting damaged DNA. RESULTS CdCl2, at various concentrations and exposure times, increased BOK mRNA. 16HBE-BOK cells (BOK over-expressing) proliferated more than 16HBE cells after 72 h; 16HBE-shBOK (BOK knockdown) cells proliferated less. In addition, BOK deficiency enhanced cell death induced by CdCl2. Similarly, CdCl2- and H2O2-induced DNA damage was greater in BOK-deficient cells. CONCLUSIONS These findings support a role for BOK in CdCl2-induced DNA damage and cell death.

Wnt/ß-Catenin Pathway Is Involved in Cadmium-Induced Inhibition of Osteoblast Differentiation of Bone Marrow Mesenchymal Stem Cells.

Cadmium is a common environmental pollutant that causes bone damage. However, the effects of cadmium on the osteogenic differentiation of bone marrow mesenchymal stem cells (BMMSCs) and its mechanism of action in this process are unclear. Here, we determined the effects of cadmium chloride (CdCl2) on the osteogenic differentiation of BMMSCs and the potential mechanism involved in this process. As determined in the present investigation, CdCl2, in a concentration-dependent manner, affected the viability of BMMSCs and their cytoskeletons. Exposure to 0.1 or 0.2 µM CdCl2 inhibited osteogenic differentiation of BMMSCs, which was reflected in the down-regulation of osteoblast-related genes (ALP, OCN, Runx2, OSX, and OPN); in suppression of the protein expression of alkaline phosphatase (ALP) and runt-related transcription factor 2 (Runx2); and in decreased ALP activity and capacity for mineralization. Moreover, mRNA microarray was performed to determine the roles of these factors in BMMSCs treated with CdCl2 in comparison to control BMMSCs. As determined with the microarrays, the Wingless-type (Wnt), mothers against decapentaplegic and the C. elegans gene Sam (SMAD), and Janus kinase-Signal Transducers and Activators of Transcription (JAK-STAT) signaling pathways were involved in the effects caused by CdCl2. Moreover, during differentiation, the protein levels of Wnt3a, ß-catenin, lymphoid enhancer factor 1 (LEF1), and T-cell factor 1 (TCF1) were reduced by CdCl2. The current research shows that CdCl2 suppresses the osteogenesis of BMMSCs via inhibiting the Wnt/ß-catenin pathway. The results establish a previously unknown mechanism for bone injury induced by CdCl2.

MALAT1 via microRNA-17 regulation of insulin transcription is involved in the dysfunction of pancreatic ß-cells induced by cigarette smoke extract.

Cigarettes contain various chemicals with the potential to influence metabolic health. Exposure to cigarette smoke causes a dysfunction in pancreatic ß-cells and impairs insulin production. However, the mechanisms for cigarette smoke-induced reduction of insulin remain largely unclear. Data from 558 patients with diabetes showed that, with smoking pack-years, homeostatic model assessment (HOMA)-ß (a method for assessing ß-cell function) decreased and that HOMA of insulin resistance increased. For ß-cells (MIN6), cigarette smoke extract (CSE) increased the levels of thioredoxin-interacting protein (TXNIP) and the long noncoding (lnc)RNA, metastasis-associated lung adenocarcinoma transcript 1 (MALAT1), and downregulated the levels of the transcription factor, mafA, and microRNA (miR)-17. MALAT1, one of four lncRNAs predicted to regulate miR-17, was knocked down by small interfering RNA (siRNA). For these cells, an miR-17 mimic inhibited TXNIP and enhanced the production of insulin. Knockdown of MALAT1 induced an increase in miR-17, which suppressed TXNIP and promoted the production of insulin. In the sera of patients with diabetes who smoked, there were higher MALAT1 levels and lower miR-17 levels than in the sera of nonsmokers. Thus, CSE inhibits insulin production by upregulating TXNIP via MALAT1-mediated downregulation of miR-17, which provides an understanding of the processes involved in the reduced ß-cells function caused by cigarette smoke.

NF-κB-regulated miR-155, via repression of QKI, contributes to the acquisition of CSC-like phenotype during the neoplastic transformation of hepatic cells induced by arsenite.

Chronic exposure to arsenite can cause various human tumors. For the initiation and recurrence of human liver cancer, the acquisition of CSC-like properties is essential. In various cancers, microRNAs (miRNAs) act as regulators in induction of CSC-like properties. Liver cancers over-express miR-155, but the mechanism relating miR-155 and arsenite-induced liver cancer is unknown. Here, we show that long-term exposure of L-02 cells to arsenite increases miR-155 levels by activation of NF-κB and leads to the acquisition of CSC-like properties. In spheroids formed from arsenite-transformed L-02 cells, the levels of miR-155 positively relate to the levels of CD90, EpCAM, and OCT4. Inhibition of miR-155, by reduction of SOX2 and OCT4, results in suppression of spheroid formation. Luciferase reporter assays indicate that QKI is a target of miR-155. Inhibition of QKI expression by miR-155 promotes arsenite-induced acquisition of CSC-like properties, whereas QKI over-expression has the opposite effect. Collectively, the findings demonstrate that miR-155, driven by NF-κB, reduces QKI expression and is involved in acquisition of the CSC-like phenotype during neoplastic transformation of hepatic cells induced by arsenite.

Identification of interneurons required for the aversive response of Caenorhabditis elegans to graphene oxide.

BACKGROUND: So far, how the animals evade the environmental nanomaterials is still largely unclear. In this study, we employed in vivo assay system of Caenorhabditis elegans to investigate the aversive behavior of nematodes to graphene oxide (GO) and the underlying neuronal basis. RESULTS: In this assay model, we detected the significant aversive behavior of nematodes to GO at concentrations more than 50 mg/L. Loss-of-function mutation of nlg-1 encoding a neuroligin with the function in connecting pre- and post-synaptic neurons suppressed the aversive behavior of nematodes to GO. Moreover, based on the neuron-specific activity assay, we found that the NLG-1 activity in AIY or AIB interneurons was required for the regulation of aversive behavior to GO. The neuron-specific activities of NLG-1 in AIY or AIB interneurons were also required for the regulation of GO toxicity. CONCLUSIONS: Using nlg-1 mutant as a genetic tool, we identified the AIY and AIB interneurons required for the regulation of aversive behavior to GO. Our results provide an important neuronal basis for the aversive response of animals to environmental nanomaterials.

Enhanced glycolysis, regulated by HIF-1α via MCT-4, promotes inflammation in arsenite-induced carcinogenesis.

Arsenite is well established as a human carcinogen, but the molecular mechanisms leading to arsenite-induced carcinogenesis are complex and elusive. Accelerated glycolysis, a common process in tumor cells called the Warburg effect, is associated with various biological phenomena. However, the role of glycolysis induced by arsenite is unknown. We have found that, with chronic exposure to arsenite, L-02 cells undergo a metabolic shift to glycolysis. In liver cells exposed to arsenite, hypoxia inducible factor-1α (HIF-1α) and monocarboxylate transporter-4 (MCT-4) are over-expressed. MCT-4, directly mediated by HIF-1α, maintains a high level of glycolysis, and the enhanced glycolysis promotes pro-inflammatory properties, which are involved in arsenite carcinogenesis. In addition, serum lactate and cytokines are higher in arsenite-exposed human populations, and there is a positive correlation between them. Moreover, there is a positive relationship between lactate and cytokines with arsenic in hair. In sum, these findings indicate that MCT-4, mediated by HIF-1α, enhances the glycolysis induced by arsenite. Lactate, the end product of glycolysis, is released into the extracellular environment. The acidic microenvironment promotes production of pro-inflammatory cytokines, which contribute to arsenite-induced liver carcinogenesis. These results provide a link between the induction of glycolysis and inflammation in liver cells exposed to arsenite, and thus establish a previously unknown mechanism for arsenite-induced hepatotoxicity.