Changes of Src-suppressed C kinase substrate expression in cytokine induced reactive C6 glioma cells.
OBJECTIVE
To investigate effect of tumor necrosis factor-alpha (TNF-alpha) on the Src-suppressed C kinase substrate (SSeCKS) in C6 glioma cells.


METHODS
Cultured C6 glioma cells were randomly divided into two groups. In time-dependent group, cells were cultured with TNF-alpha (2 ng/mL) for 0 h, 1 h, 3 h, 6 h, 12 or 24 h, respectively; in dose-dependent group, cells were cultured with TNF-alpha (0 ng/mL, 0.02 ng/mL, 0.2 ng/mL, or 2 ng/mL) for 6 h. The expression of SSeCKS was detected by Realtime PCR and Western blot analysis, and immunocytochemistry was used to investigate SSeCKS's subcellular localization.


RESULTS
TNF-alpha induced rapid phosphorylations of protein kinase C (PKC) substrates in C6 glioma cells, and upregulated SSeCKS expression in a time and concentration dependent manner. Immunocytochemistry suggested that SSeCKS was localized in the cyroplasm and the leading end of podosomal extensions in control groups, while TNF-alpha induced translocation of SSeCKS perinuclear. This effect could be partly reversed by PKC inhibitor Ro-31-8220.


CONCLUSIONS
TNF-alpha activates PKC and upregulates SSeCKS expression in C6 glioma cells. These effects are associated with PKC activity, suggesting that SSeCKS plays a role in response to glia activation in PKC mediated pathway.
Sun
Lin-Lin
LL
Key Laboratory of Neuroregeneration of Jiangsu Province, Nantong University, Nantong 226001, China.
Cheng
Chun
C
Liu
Hai-Ou
HO
Xiao
Feng
F
Qin
Jing
J
Shao
Xiao-Yi
XY
Shen
Ai-Guo
AG
D054758:A Kinase Anchor Proteins; D000818:Animals; D001253:Astrocytes / Q000378:metabolism; D001932:Brain Neoplasms / Q000378:metabolism; D018797:Cell Cycle Proteins / Q000378:metabolism*; D004305:Dose-Response Relationship, Drug; D004789:Enzyme Activation; D005786:Gene Expression Regulation; D005910:Glioma / Q000378:metabolism; D007150:Immunohistochemistry; D011493:Protein Kinase C / Q000378:metabolism*; D021381:Protein Transport / Q000502:physiology; D011897:Random Allocation; D051381:Rats; D015398:Signal Transduction / Q000502:physiology*; D014407:Tumor Cells, Cultured; D014409:Tumor Necrosis Factor-alpha / Q000008:administration & dosage / Q000502:physiology*
D054758:A Kinase Anchor Proteins; C102361:Akap12 protein, rat; D018797:Cell Cycle Proteins; D014409:Tumor Necrosis Factor-alpha; D011493:Protein Kinase C