title: Multi-walled carbon nanotube directed gene and protein expression in cultured human aortic endothelial cells is influenced by suspension medium.
abstract: The use and production of multi-walled carbon nanotubes (MWCNTs) have significantly increased over the last decade due to their versatility in numerous applications. Their unique physical and chemical properties make them desirable for various biomedical applications, but the same properties also raise concerns about their safety to human health, particularly at the cellular level. The vascular endothelium could be exposed to nanomaterials either by direct intravenous administration in nanomedicine or by translocation following inhalational exposure in an occupational setting. We hypothesized that direct exposure to MWCNTs will increase the expression of inflammatory markers in human aortic endothelial cells (HAEC). We also investigated the effect of the route of exposure on activation by changing the suspension medium of the MWCNTs. HAEC were treated in vitro with MWCNTs (1 or 10 μg/cm(2)) suspended in either cell culture medium [(M)-MWCNTs] or 10% clinical grade pulmonary surfactant [(S)-MWCNTs]. The zeta potential of the (S)-MWCNTs was significantly more negative than the (M)-MWCNTs suggesting a more stable suspension. Treatment of HAEC with (S)-MWCNTs; as compared to (M)-MWCNTs resulted in a significantly higher up-regulation of mRNA transcripts for cell adhesion molecules VCAM1, SELE, ICAM1 and the chemokine CCL2. Time dependent changes in VCAM1 and CCL2 protein levels were confirmed by immunofluorescence, flow cytometry and ELISA. A label free quantitative mass spectrometry proteomic analysis was utilized to compare protein expression patterns between the two suspensions of MWCNTs. We identified significant expression changes in >200 unique proteins in MWCNT treated HAEC. However, the two suspensions of MWCNTs resulted in different protein expression patterns with the eIF2 pathway as the only common pathway identified between the two suspensions. These data suggest that direct exposure to MWCNTs induces acute inflammatory and protein expression changes in HAEC, which is influenced by the type of media used for suspension of MWCNTs and their resulting zeta potential.
authors.affiliation: Department of Physiology, Brody School of Medicine, East Carolina University, Greenville, NC 27834, USA.
authors.forename: Achini K
authors.initials: AK
authors.lastname: Vidanapathirana
authors.forename: Xianyin
authors.initials: X
authors.lastname: Lai
authors.forename: Susana C
authors.initials: SC
authors.lastname: Hilderbrand
authors.forename: Josh E
authors.initials: JE
authors.lastname: Pitzer
authors.forename: Ramakrishna
authors.initials: R
authors.lastname: Podila
authors.forename: Susan J
authors.initials: SJ
authors.lastname: Sumner
authors.forename: Timothy R
authors.initials: TR
authors.lastname: Fennell
authors.forename: Christopher J
authors.initials: CJ
authors.lastname: Wingard
authors.forename: Frank A
authors.initials: FA
authors.lastname: Witzmann
authors.forename: Jared M
authors.initials: JM
authors.lastname: Brown
mesh_terms: D001011:Aorta / Q000166:cytology / Q000187:drug effects / Q000473:pathology; D002448:Cell Adhesion / Q000187:drug effects; D002460:Cell Line; D002470:Cell Survival / Q000187:drug effects; D002478:Cells, Cultured; D018932:Chemokine CCL2 / Q000235:genetics / Q000378:metabolism; D003470:Culture Media / Q000737:chemistry*; D042783:Endothelial Cells / Q000187:drug effects* / Q000473:pathology; D004730:Endothelium, Vascular / Q000166:cytology* / Q000187:drug effects / Q000473:pathology; D004797:Enzyme-Linked Immunosorbent Assay; D005455:Fluorescent Antibody Technique; D006801:Humans; D019570:Inhalation Exposure; D037742:Nanotubes, Carbon / Q000737:chemistry* / Q000633:toxicity; D040901:Proteomics; D012333:RNA, Messenger / Q000235:genetics / Q000378:metabolism; D015854:Up-Regulation
chemical_list: C111835:CCL2 protein, human; D018932:Chemokine CCL2; D003470:Culture Media; D037742:Nanotubes, Carbon; D012333:RNA, Messenger