Long non-coding RNAs mediate the association between short-term PM2.5 exposure and circulating biomarkers of systemic inflammation.

Although short-term fine particulate matter (PM2.5) exposure is associated with systemic inflammation, the effect of lncRNA on these association remains unknown. This study aims to investigate whether the plasma lncRNA mediate the effect of short-term PM2.5 exposure on systemic inflammation. In this cross-sectional study, plasma Clara cell protein 16 (CC16), interleukin 6 (IL-6), IL-8, tumor necrosis factor-α (TNF-α) and lncRNA expression levels were measured in 161 adults between March and April in 2018 in Shijiazhuang, China. PM2.5 concentrations were estimated 0-3 days prior to the examination date and the moving averages were calculated. Multiple linear regressions were used to evaluate the associations between PM2.5, the four biomarkers and lncRNA expression levels. Mediation analyses were performed to explore the potential roles of lncRNA expression in these associations. The median concentration of PM2.5 ranged from 39.65 to 60.91 mg/m3 across different lag days. The most significant effects on IL-6 and TNF-α per interquartile range increase in PM2.5 were observed at lag 0-3 days, with increases of 0.70 pg/mL (95% CI: 0.33, 1.07) and 0.21 pg/mL (95% CI: 0.06, 0.36), respectively. While the associations between PM2.5 and IL-8 (0.68 pg/mL, 95% CI: 0.34, 1.02) and CC16 (3.86 ng/mL, 95% CI: 1.60, 6.13) were stronger at lag 0 day. Interestingly, a negative association between PM2.5 and the expression of four novel lncRNAs (lnc-ACAD11-1:1, lnc-PRICKLE1-4:1, lnc-GPR39-7:2, and lnc-MTRNR2L12-3:6) were observed at each lag days. Furthermore, these lncRNAs mediated the effects of PM2.5 on the four biomarkers, with proportions of mediation ranged from 2.27% (95% CI: 1.19%, 9.82%) for CC16 to 35.60% (95% CI: 17.16%, 175.45%) for IL-6. Our findings suggested that plasma lncRNA expression mediat the acute effects of PM2.5 exposure on systematic inflammation. These highlight a need to consider circulating lncRNA expression as biomarkers to reduce health risks associated with PM2.5.

Upregulation of Circ_0035266 Contributes to the Malignant Progression of Inflammation-Associated Malignant Transformed Cells Induced by Tobacco-Specific Carcinogen NNK.

Cigarette smoking-induced chronic inflammation has been considered a vital driver of lung tumorigenesis. The compounds 4-(methylnitrosamino)-1-(3-pyridyl)-1-butanone (NNK), a tobacco-specific carcinogen, and lipopolysaccharide (LPS), an inflammatory inducer, are important components of tobacco smoke which have been implicated in inflammation-driven carcinogenesis. However, the biological effects and underlying mechanisms of LPS-mediated inflammation on NNK-induced tumorigenesis are still unclear. In this study, BEAS-2B human bronchial epithelial cells were exposed to NNK, LPS or both, for short- or long-term periods. We found that acute LPS exposure promoted the secretion of granulocyte-macrophage colony stimulating factor (GM-CSF) and interleukin (IL)-6 in NNK-treated BEAS-2B cells. In addition, chronic LPS exposure facilitated the NNK-induced malignant transformation process by promoting cell proliferation, cell cycle alteration, migration, and clonal formation. Previously, we determined that circular RNA circ_0035266 enhanced cellular inflammation in response to NNK + LPS by sponging miR-181d-5p and regulating expression of its downstream target DEAD-Box Helicase 3 X-Linked (DDX3X). Here, we found that knockdown of circ_0035266 or DDX3X led to a remarkable inhibition of the proliferation, cell cycle progression, and migration of NNK + LPS-transformed BEAS-2B cells, whereas overexpression of these genes produced the opposite effects, indicating the oncogenic roles of circ_0035266 and DDX3X in the malignant progression of chronic inflammation-driven malignant transformed cells. Moreover, the regulatory relationships among circ_0035266, miR-181d-5p, and DDX3X were further confirmed using a group of lung cancer tissues. Conclusively, our findings provide novel insights into our understanding of inflammation-driven tumorigenesis using a cellular malignant transformation model, and indicate a novel tumor-promoting role for circ_0035266 in chemical carcinogenesis.

The involvement of copper, circular RNAs, and inflammatory cytokines in chronic respiratory disease.

Exposure to high concentrations of copper is associated with pulmonary inflammation and chronic respiratory disease (CRD). Epigenetic modulation of noncoding RNAs contributes to the development of several CRDs. It is unknown whether epigenetic modulation is involved in copper mediated pulmonary inflammation and CRD. We conducted a case-control study of 101 CRD cases and 161 control subjects in Shijiazhuang, China, and evaluated circRNAs and cytokine levels (IL-6 and IL-8) by qPCR and ELISA. Urinary copper concentration was determined by inductively coupled plasma mass spectrometry. Linear mixed models and generalized linear mixed models were used to assess the associations of circRNAs with CRD, urinary copper, and cytokines. We exposed the human bronchial epithelial cell line, 16HBE, to copper and assessed the functional role of a circRNA, circ_0008882, by RNA overexpression. Cellular location of circ_0008882 was assessed by separation of nuclear and cytoplasmic RNAs. Nine circRNAs were associated with an increased risk for CRDs, while the relative expression of circ_0008882 was decreased after copper exposure in vitro and in vivo. Copper exposure stimulated 16HBE cells to release proinflammatory IL-6 and IL-8. The release of the cytokines was inhibited by overexpression of circ_0008882. These results suggest a role for circ_0008882 in the regulation of CRD associated inflammation following copper exposure.

Circular RNA circNIPBL promotes NNK-induced DNA damage in bronchial epithelial cells via the base excision repair pathway.

Environmental chemical exposure often causes DNA damage, which leads to cellular dysfunction and the development of diseases. 4-(Methylnitrosamino)-1-(3-pyridyl)-1-butanone (NNK), a tobacco-specific carcinogen that is known to cause DNA damage, while remains unknown about the underlying mechanism. In this study, simulated doses of NNK exposure in smokers, ranging from 50 to 300 µM, were used to detect the DNA damage effects of NNK in two human bronchial epithelial cells, 16HBE and BEAS-2B. The comet assay revealed increased DNA damage in response to NNK treatment, as measured by increased Olive tail moment (OTM). NNK treatment also led to elevated foci formation and protein expression of γ-H2AX, a DNA damage sensor. Dysregulation of proliferation, cell cycle arrest and apoptosis, was also observed in NNK-treated cells. Furthermore, the most effective dose of NNK (300 µM) was used in subsequent mechanistic studies. A circular RNA circNIPBL was identified to be significantly up-regulated in NNK-treated cells, circNIPBL knockdown successfully alleviated NNK-induced DNA damage and reversed the cellular dysregulation, while circNIPBL overexpression had the opposite effect. Mechanistically, we identified an interaction between circNIPBL and PARP1, a critical enzyme of the base excision repair (BER) pathway. CircNIPBL silencing successfully alleviated the NNK-induced inhibition of BER pathway proteins, including PARP1, XRCC1, PCNA and FEN1, while overexpression of circNIPBL had the opposite effect. In summary, our study shows for the first time that circNIPBL promotes NNK-induced DNA damage and cellular dysfunction through the BER pathway. In addition, our findings reveal the crucial role of epigenetic regulation in carcinogen-induced genetic lesions and further our understanding of environmental carcinogenesis.

Circular RNA circ_Cabin1 promotes DNA damage in multiple mouse organs via inhibition of non-homologous end-joining repair upon PM2.5 exposure.

Fine particulate matter (PM2.5) has been shown to induce DNA damage. Circular RNAs (circRNAs) have been implicated in various disease processes related to environmental chemical exposure. However, the role of circRNAs in the regulation of DNA damage response (DDR) after PM2.5 exposure remains unclear. In this study, male ICR mice were exposed to PM2.5 at a daily mean concentration of 382.18 µg/m3 for 3 months in an enriched-ambient PM2.5 exposure system in Shijiazhuang, China, and PM2.5 collected form Shijiazhuang was applied to RAW264.7 cells at 100 µg/mL for 48 h. The results indicated that exposure to PM2.5 induced histopathological changes and DNA damage in the lung, kidney and spleen of male ICR mice, and led to decreased cell viability, increased LDH activity and DNA damage in RAW264.7 cells. Furthermore, circ_Cabin1 expression was significantly upregulated in multiple mouse organs as well as in RAW264.7 cells upon exposure to PM2.5. PM2.5 exposure also resulted in impairment of non-homologous end joining (NHEJ) repair via the downregulation of Lig4 or Dclre1c expression in vivo and in vitro. Importantly, circ_Cabin1 promoted PM2.5-induced DNA damage via inhibiting of NHEJ repair. Moreover, the expression of circ_Cabin1 and Lig4 or Dclre1c was strongly correlated in multiple mouse organs, as well as in the blood. In summary, our study provides a new perspective on circRNAs in the regulation of DDR after environmental chemical exposure.

Tobacco-Related Exposure Upregulates Circ_0035266 to Exacerbate Inflammatory Responses in Human Bronchial Epithelial Cells.

One of the most carcinogenic chemicals found in cigarette tobacco smoke is 4-(methylnitrosamino)-1-(3-pyridyl)-1-butanone (NNK), which has been confirmed to be associated with the etiology of diverse cancers. Lipopolysaccharide (LPS), another biologically active component of cigarette smoke, is a risk factor which enhances NNK-induced lung tumorigenesis due to chronic lung inflammation. Although inflammatory responses play critical roles in the initiation of many tumors, our knowledge about the mechanisms of NNK+LPS on inflammation is currently limited. Here, we investigated the inflammatory effects of NNK+LPS in human bronchial epithelial cells (BEAS-2B) and explored the underlying mechanisms mediated by circular RNAs (circRNAs). We identified a novel circRNA, circ_0035266, which was strongly upregulated in NNK+LPS-induced BEAS-2B cells and enhanced the inflammatory responses to NNK+LPS by regulating the secretion of pro-inflammatory cytokines interleukin (IL)-6 and IL-8. Specifically, circ_0035266 knockdown alleviated NNK+LPS-induced inflammatory responses, whereas overexpression of circ_0035266 had the opposite effect. Moreover, dual-luciferase reporter and fluorescence in situ hybridization (FISH) assays verified that circ_0035266 bound to miR-181d-5p directly in the cytoplasm. qRT-PCR, dual-luciferase reporter assays, and Western blot analyses showed that DDX3X (DDX3) was the downstream target of miR-181d-5p and that DDX3X expression levels were modulated by circ_0035266. These results suggested that circ_0035266 served as a competitive endogenous RNA for miR-181d-5p to regulate DDX3X expression, which is involved in the modulation of NNK+LPS-induced inflammatory responses in BEAS-2B cells.

Circular RNA circBbs9 promotes PM2.5-induced lung inflammation in mice via NLRP3 inflammasome activation.

Fine particulate matter (PM2.5) is one of the most important components of environmental pollutants, and is associated with pulmonary injury. However, the biological mechanisms of pulmonary damage caused by PM2.5 are poorly defined, especially the molecular pathways related to inflammation. Following system exposure to PM2.5 for 3 months in normal mice and in chronic obstructive pulmonary disease (COPD) model mice, it was found that PM2.5 exposure increased the expression of IL-1ß and IL-18 in lung tissues via NLRP3 activation, and these effects were more intense in COPD model mice. Circular RNA (circRNA) sequencing showed that the expression profiles of circRNAs were changed after PM2.5 exposure, and the positive roles of circBbs9 in inflammation induced by PM2.5 were verified. The circBbs9 knockdown alleviated PM2.5-induced inflammation via NLRP3 inflammasome inactivation, as well as IL-1ß and IL-18 inhibition in RAW264.7 cells, while overexpression of circBbs9 had the opposite effect. Bioinformatics and luciferase reporter assays showed that circBbs9 bound to microRNA-30e-5p (miR-30e-5p) and co-regulated the expression of Adar, a downstream target gene of miR-30e-5p. Taken together, these results revealed that PM2.5 induced pulmonary inflammation through NLRP3 inflammasome activation regulated by the circBbs9-miR-30e-5p-Adar pathway. Our findings provide a new target, circBbs9, for the assessment of lung inflammation and COPD exacerbation induced by PM2.5 exposure.

Circular RNA 0039411 Is Involved in Neodymium Oxide-induced Inflammation and Antiproliferation in a Human Bronchial Epithelial Cell Line via Sponging miR-93-5p.

Adverse health effects induced by neodymium oxide (Nd2O3) particles have raised concern as a result of their increasing applications in various arenas. However, information on their potential cytotoxicity is currently limited. In the present study, we investigated the underlying cytotoxicity of Nd2O3 in human bronchial epithelial cells (16HBE) and the potential mechanisms mediated by circular RNAs (circRNAs). Nd2O3 exposure initiated an inflammatory response in 16HBE cells via the release of the proinflammatory cytokines interleukin (IL)-6 and IL-8. The 5-ethynyl-2'-deoxyuridine assays showed that Nd2O3 treatment inhibited 16HBE cell proliferation and caused cell cycle arrest at G0/G1 phase and cell apoptosis. Microarray analyses demonstrated that Nd2O3 treatment altered circRNA expression profiles and significantly upregulated circRNA 0039411 (circ_0039411) in 16HBE cells. Further functional studies showed that silencing circ_0039411 prevented Nd2O3-induced inflammation and reversed its antiproliferative effect by moderating the G0/G1 phase cell cycle arrest, whereas overexpression of circ_0039411 had the opposite effects. Luciferase reporter assays showed that circ_0039411 bound to miR-93-5p, whereas fluorescence in situ hybridization showed that circ_0039411 and miR-93-5p colocalized in the cytoplasm. Moreover, transfection of 16HBE cells with a miR-93-5p mimic decreased the phosphorylation of signal transducer and activator of transcription 3 (STAT3). The levels of phospho-STAT3 were decreased by circ_0039411 silencing and elevated after circ_0039411 overexpression. These results suggested that upregulation of circ_0039411 mediated Nd2O3-induced inflammation and dysfunction by sponging miR-93-5p.

The Frequency of Unhealthy Food Advertising on Mainland Chinese Television (TV) and Children and Adolescents' Risk of Exposure to Them.

OBJECTIVE: To conduct an analysis of the frequency of unhealthy food advertising on mainland Chinese television (TV) and children and adolescents' risk of exposure to them. METHODS: The frequencies of all types of advertisements (ads) on forty TV channels in mainland China, the exact ad broadcast times, and the name and brand of all snacks and western fast foods advertised were recorded from 0800 hours to 2400 hours on both a weekday and a weekend day in a week. The difference in the frequencies of the diverse types of ads over eight time intervals (each time interval was 2 hours) were compared, and the trends in ad frequencies during the time intervals were described. RESULTS: The TV channels broadcast 155 (91-183) (expressed as median [P25-P75]) food ads, 87 (38-123) snack ads, 49 (11-85) beverage ads, and 58 (25-76) ads of snacks suitable for limited consumption (SSLCs) in a day. The proportion of snack ads among food ads (SPF%) was 55.5% (40.3%-71.0%), and the proportion of SSLC ads among snack ads (LPS%) was 67.4% (55.4%-79.3%). The ad frequencies for food, snacks, SSLCs, and beverages demonstrated significant differences among the eight time intervals (all P=0.000). TV channels broadcast the most frequent ads for food, snacks, SSLCs, and beverages during the time interval from 2000 hours to 2200 hours among the eight time intervals. CONCLUSIONS: Chinese children and adolescents may be at a high risk of exposure to unhealthy food advertising on TV. Reducing the exposure risk strongly requires multisectoral cooperation.