Up-regulation of the vascular cell adhesion molecule-1 (VCAM-1) induced by Theiler's murine encephalomyelitis virus infection of murine brain astrocytes.
The present article reports the up-regulation of the expression of the vascular cell adhesion molecule-1 (VCAM-1) by SJL/J mouse brain astrocytes infected with Theiler's murine encephalomyelitis virus (TMEV). Complementary RNA (cRNA) from mock- and TMEV-infected cells was hybridized to the Affymetrix whole murine genome U74v2 DNA microarray. Hybridization data analysis revealed background expression in untreated cells and the up-regulation of three sequences coding for VCAM-1, as described by the SCOP (Structural Classification Of Proteins) database. The authors further studied its regulation, confirming and validating their mRNA increase by reverse transcriptase-polymerase chain reaction (RT-PCR) and quantitative real-time RT-PCR. The presence of the 100-kDa VCAM-1 protein in mock- and TMEV-infected cells was demonstrated in the cell membrane by a specific cell-based enzyme-linked immunosorbent assay (ELISA), in addition to flow cytometry and confocal immunohistochemistry. Further, Western blots were used to quantify the amount of VCAM-1 molecules in cell extracts. All these data demonstrated a mean 75% increase in the expression of VCAM-1 on the surface of TMEV-infected cells. Three inflammatory cytokines, interleukin-1alpha (IL-1alpha), interferon gamma (IFNgumma), and specially tumor necrosis factor alpha (TNF-α), some of which are also induced by TMEV in astrocytes (IL-1alpha and TNF-alpha), were potent inducers of VCAM-1 expression. To demonstrate whether the VCAM-1 molecules were biologically active, mediating adhesion to other cells as the integrin alpha4-expressing CD4+ T lymphocytes, the authors used a cell adhesion test. It was also demonstrated by immunohistochemistry that in vivo VCAM-1 expression is enhanced after TMEV intracraneal infection. The present data show a small but statistically significant overexpression of VCAM-1 after astrocyte infection with TMEV that could play a significant role in vivo.
Rubio
Nazario
N
Instituto Cajal, C.S.I.C., Madrid, Spain. nazario@cajal.csic.es
Sanz-Rodriguez
Francisco
F
Arevalo
Maria-Angeles
MA
D000818:Animals; D000831:Animals, Newborn; D001253:Astrocytes / Q000378:metabolism* / Q000821:virology*; D001921:Brain / Q000166:cytology* / Q000821:virology; D002478:Cells, Cultured; D004797:Enzyme-Linked Immunosorbent Assay; D005434:Flow Cytometry; D020869:Gene Expression Profiling; D007120:Immunochemistry; D051379:Mice; D008815:Mice, Inbred Strains; D012333:RNA, Messenger / Q000235:genetics / Q000378:metabolism; D020133:Reverse Transcriptase Polymerase Chain Reaction; D017945:Theilovirus / Q000502:physiology*; D015854:Up-Regulation; D019010:Vascular Cell Adhesion Molecule-1 / Q000096:biosynthesis* / Q000235:genetics*
D012333:RNA, Messenger; D019010:Vascular Cell Adhesion Molecule-1