Unknown,Transcriptomics,Genomics,Proteomics

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Global gene expression profile of RAW264.7 cells with or without SWCNT exposure

ABSTRACT: Carbon nanotubes (CNTs) are one of the most attractive nanomaterials that have many potential applications in industrial, medical, and environmental sectors. To investigate the toxic effect of single-walled CNTs (SWCNTs) on the immune system, we examined the influences of SWCNTs (depleted of metal impurities) on viability and global gene expression in murine macrophages. RAW264.7 Cells were exposed to various concentrations of SWCNTs, and cell viability was analyzed at various time points using WST-1 assay. SWCNT cytotoxicity in macrophages was concentration and time dependent, with significant viability loss occurring within 4 hr at 50Î¼g/ml SWCNT exposure. As low as 1Î¼g/ml SWCNTs reduced cell viability at 12 hr, whereas 50Î¼g/ml SWCNT exposure completely abolished cell proliferation. Laser confocal microscopy indicated that SWCNTs could enter and aggregate in cell cytoplasm and nuclear areas. Microarray, real-time reverse-transcriptase polymerase chain reaction and enzyme-linked immunosorbent assay revealed that SWCNT altered gene and protein expression pattern. The most differentially expressed genes were related to proinflammatroy cytokines and chemokine, such as CCL-3 and CCL-4, apoptosis (e.g., Caspase-1, and -3), protective response such as Hmox-1and GSTM3) and cell surface molecules such as ICAM-1, Itgb2, indicating a possible involvement of SWCNT in inflammation and pulmonary granuloma formation, carcinogenesis and Th2 polarization of T cells. Furthermore, a substantial amount of down-regulated genes were related to ribosomal assembly and mitochondrial respiration chain, implicating a possible mechanism of SWCNT-induced oxidative stress and gene expression inhibition. The results are a first step in identifying molecular targets for the toxicity of SWCNTs and the elucidating of the molecular mechanisms for the immune modulation ability of SWCNTs. Cells were exposure to 50ug/ml of acid-functionzalized SWCNTs for 24 hours before gene expression profiling, cells without SWCNT exposure were used as control.

ORGANISM(S): Mus musculus

SUBMITTER: Bin Wan

PROVIDER: E-GEOD-16948 | biostudies-arrayexpress |

REPOSITORIES: biostudies-arrayexpress

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Project description:Single-walled carbon nanotubes (SWCNTs) are being explored and used for a wide range of applications in industrial and medical sectors, and the increasing exposure of SWCNTs necessitate the studies of their potential environmental and health effects. Considerable efforts have been made to improve the dispersion of SWCNTs by chemical modifications. The present study was designed to determine if the acid functionalization of SWCNTs enhanced the toxicity and more efforts were made to understand the molecular mechanisms. RAW264.7 cells were exposed to 0-100μg/mL of SWCNTs and AF-SWCNTs for 24 hours and cell proliferation, viability, and gene expression profiles were assessed and compared. Results showed that the magnitude of AF-SWCNT-induced cell proliferation inhibition was higher than that of SWCNTs. 10μg/mL of AF-SWCNTs was determined as non-toxic to cells by viability analysis. AF-SWCNTs could enter and aggregate in cell cytoplasm and nuclear areas. Microarray, real-time reverse-transcriptase polymerase chain reaction (RT-PCR) and enzyme-linked immunosorbent assays (ELISA) demonstrated that AF-SWCNTs (1, 10, and 50μg/mL) altered gene and protein expression patterns. The most differentially expressed genes were related to pro-inflammatory cytokines and chemokines (such as CCL-3 and CCL-4), apoptosis (e.g., Caspase-1, -4, and Dffa), protective response (such as Hmox-1and GSTM3), and cell surface molecules such as ICAM-1, Itgb2, indicating a possible involvement of SWCNT in inflammation, anti-apoptotic and DNA fragmentation, carcinogenesis and biased T cell polarization. Furthermore, a substantial amount of down-regulated genes were found related to ribosomal assembly and mitochondrial respiration chain, implicating a possible mechanism of SWCNT-induced oxidative stress and gene expression inhibition. RAW264.7 gene expression after exposure to 1-50ug/ml AF-SWCNT-24 hours. 0ug/ml of AF-SWCNT was used as control sample. The data were obtained from three biological replicates.

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Global gene expression profile of RAW264.7 cells with or without SWCNT exposure

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