Abstract
Recent evidence supports a neuroprotective role of Src homology 2-containing protein tyrosine phosphatase 2 (SHP-2) against ischemic brain injury. However, the molecular mechanisms of SHP-2 activation and those governing how SHP-2 exerts its function under oxidative stress conditions are not well understood. Recently we have reported that reactive oxygen species (ROS)-mediated oxidative stress promotes the phosphorylation of endogenous SHP-2 through lipid rafts, and that this phosphorylation strongly occurs in astrocytes, but not in microglia. To investigate the molecules involved in events leading to phosphorylation of SHP-2, raft proteins were analyzed using astrocytes and microglia. Interestingly, caveolin-1 and -2 were detected only in astrocytes but not in microglia, whereas flotillin-1 was expressed in both cell types. To examine whether the H2O2-dependent phosphorylation of SHP-2 is mediated by caveolin-1, we used specific small interfering RNA (siRNA) to downregulate caveolin- 1 expression. In the presence of caveolin-1 siRNA, the level of SHP-2 phosphorylation induced by H2O2 was significantly decreased, compared with in the presence of control siRNA. Overexpression of caveolin- 1 effectively increased H2O2-induced SHP-2 phosphorylation in microglia. Lastly, H2O2 induced extracellular signal-regulated kinase (ERK) activation in astrocytes through caveolin-1. Our results suggest that caveolin-1 is involved in astrocyte-specific intracellular responses linked to the SHP-2-mediated signaling cascade following ROS-induced oxidative stress.
Highlights
Src homology 2-containing protein tyrosine phosphatase 2 (SHP-2), a member of a subfamily of protein tyrosine phosphatases (PTPs), is highly expressed in specific regions of the rat brain, including the cortex, cerebellum, and hippocampus (Suzuki et al, 1995)
Primary cells were cultured as described in the Materials and Methods section, and cultures were confirmed as being enriched with astrocytes and microglia by immunoblotting with anti-glial fibrillary acidic protein (GFAP) and anti-ionized calcium-binding adaptor molecule 1 (Iba-1) antibodies, markers for astrocytes and microglia, respectively
We demonstrated that H2O2mediated oxidative stress strongly induces SHP-2 phosphorylation, in astrocytes, and lipid rats are involved in this events (Park et al, 2009)
Summary
Src homology 2-containing protein tyrosine phosphatase 2 (SHP-2), a member of a subfamily of protein tyrosine phosphatases (PTPs), is highly expressed in specific regions of the rat brain, including the cortex, cerebellum, and hippocampus (Suzuki et al, 1995). Earlier studies show that SHP-2 is involved in neuroprotection in response to ischemic brain injury (Aoki et al, 2000; Chong et al, 2003; Gee and Mansuy, 2005). Overexpression of a catalytically inactive mutant of SHP-2 increases susceptibility to focal cerebral ischemia/reperfusion injury in the mouse adult brain (Aoki et al, 2000). Recent studies in our laboratory have suggested that H2O2-mediated oxidative stress induces SHP-2 phosphorylation and activation through lipid rafts, by H2O2 in rat primary astrocytes, but barely detectable in microglia. The goal of the present study is to elucidate the means by which SHP-2 phosphorylation and modification occurs, in astrocytes in the presence of H2O2. We examined the expression pattern of raft proteins, in astrocytes and microglia, and showed for the first time that caveolin-1 and -2 are expressed in astrocytes and that the presence of caveolin-1 in astrocytes contributes to enhanced SHP-2/ERK signaling in response to H2O2
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