title: Selected gene profiles of stressed NSC-34 cells and rat spinal cord following peripheral nerve reconstruction and minocycline treatment.
abstract: The present study was conducted to investigate the effects of minocycline on the expression of selected transcriptional and translational profiles in the rat spinal cord following sciatic nerve (SNR) transection and microsurgical coaptation. The mRNA and protein expression levels of B cell lymphoma-2 (Bcl-2), Bcl-2-associated X protein (Bax), caspase-3, major histocompatibility complex I (MHC I), tumor necrosis factor-α (TNF-α), activating transcription factor 3 (ATF3), vascular endothelial growth factor (VEGF), matrix metalloproteinase 9 (MMP9), and growth associated protein-43 (GAP-43) were monitored in the rat lumbar spinal cord following microsurgical reconstruction of the sciatic nerves and minocycline treatment. The present study used semi-quantitative reverse transcription-polymerase chain reaction (RT-PCR) and immunohistochemistry. As a PCR analysis of spinal cord tissue enabled the examination of the expression patterns of all cell types including glia, the motorneuron-like NSC-34 cell line was used to investigate expression level changes in motorneurons. As stressors, oxygen glucose deprivation (OGD) and lipopolysaccharide (LPS) treatment were performed. SNR did not induce significant degeneration of ventral horn motorneurons, whereas microglia activation and synaptic terminal retraction were detectable. All genes were constitutively expressed at the mRNA and protein levels in untreated spinal cord and control cells. SNR significantly increased the mRNA expression levels of all genes, albeit only temporarily. In all genes except MMP9 and GAP-43, the induction was seen ipsilaterally and contralaterally. The effects of minocycline were moderate. The expression levels of MMP9, TNF-α, MHC I, VEGF, and GAP-43 were reduced, whereas those of Bax and Bcl-2 were unaffected. OGD, but not LPS, was toxic for NSC-34 cells. No changes in the expression levels of Bax, caspase-3, MHC I or ATF3 were observed. These results indicated that motorneurons were not preferentially or solely responsible for SNR-mediated upregulation of these genes. MMP9, TNF-α, VEGF and Bcl-2 were stress-activated. These results suggest that a substantial participation of motorneurons in gene expression levelsin vivo. Minocycline was also shown to have inhibitory effects. The nuclear factor-κB signalling pathway may be a possible target of minocycline.
authors.affiliation: Institute of Biochemistry and Cell Biology, Otto-Von-Guericke University Magdeburg, Magdeburg D-39120, Germany.
authors.forename: Gerburg
authors.initials: G
authors.lastname: Keilhoff
authors.affiliation: Institute of Biochemistry and Cell Biology, Otto-Von-Guericke University Magdeburg, Magdeburg D-39120, Germany; Department of Trauma Surgery, Otto-Von-Guericke University Magdeburg, Magdeburg D-39120, Germany.
authors.forename: Benjamin
authors.initials: B
authors.lastname: Lucas
authors.affiliation: Institute of Biochemistry and Cell Biology, Otto-Von-Guericke University Magdeburg, Magdeburg D-39120, Germany.
authors.forename: Katja
authors.initials: K
authors.lastname: Uhde
authors.affiliation: Department of Plastic, Reconstructive and Aesthetic Surgery, Hand Surgery, Klinikum Bielefeld, Bielefeld D-33604, Germany.
authors.forename: Hisham
authors.initials: H
authors.lastname: Fansa
keywords: NSC-34 cell line; immunohistochemistry; lipopolysaccharide; oxygen glucose deprivation; reverse transcription-polymerase chain reaction