Figure 7

TNF-α or IL-1β dose-dependently up-regulated NKCC1 mRNA and protein expression in primary astrocytes. Bar graph A shows both TNF-α and IL-1β significantly elevated (#P < 0.05) NKCC1 mRNA expression. Panels B and C depicting TNF-α (10 ng/ml or 5 ng/ml) or IL-1β (10 ng/ml or 5 ng/ml) treatment protein significantly increased NKCC1 protein when compared with that in the control group (#P <0.05). The alteration of NKCC1 expression is dose dependent, that is, a higher concentration of TNF-α or IL-1β induced more NKCC1 mRNA and protein expression (A-C) (*P < 0.05, **P < 0.01). Immunofluorescence images of astrocytes showing GFAP labeled astrocytes (D, G, J, M, red), co-labeled with NKCC1 (E, H, K, N, green). The co-localized expression of GFAP and NKCC1 can be seen in panels F, I, L and O. Note NKCC1 expression is significantly up-regulated in astrocytes after treatment with a higher concentration of TNF-α (10 ng/ml) or IL-1β (10 ng/ml) as compared with the control group. ns: non-significant, P > 0.05. Scale bars: (D–O), 20 μm. GFAP, glial fibrillary acidic protein; IL-1β, interleukin-1beta; NKCC1, Na-K-Cl Cotransporter 1; TNF-α, tumor necrosis factor-alpha.