BK‐induced cytosolic phospholipase A₂ expression via sequential PKC‐δ, p42/p44 MAPK, and NF‐κB activation in rat brain astrocytes

Abstract

Bradykinin (BK), an inflammatory mediator, has been shown to induce cytosolic phospholipase A2 (cPLA2) expression implicating in inflammatory responses in various cell types. However, the detailed mechanisms underlying BK-induced cPLA2 expression in astrocytes remain unclear. RT-PCR and Western blotting analysis showed that BK induced the expression of cPLA2 mRNA and protein, which was inhibited by Hoe140, suggesting the involvement of B2 BK receptors, confirmed by immunofluorescence staining using anti-B2 BK receptor antibody. BK-induced cPLA2 expression and phosphorylation of p42/p44 MAPK was attenuated by PD98059, indicating the involvement of MEK1/2-p42/p44 MAPK in these responses. BK-induced cPLA2 expression might be due to the translocation of NF-kappaB into nucleus which was inhibited by Hoe140, helenalin, and PD98059, implying the involvement of NF-kappaB. Moreover, BK-induced cPLA2 expression was attenuated by rottlerin, suggesting that PKC-delta might be involved in these responses. This hypothesis was supported by the transfection with a dominant negative plasmid of PKC-delta significantly attenuated BK-induced response. In addition, BK-stimulated translocation of PKC-delta from cytosol to membrane fraction was inhibited by rottlerin but not by PD98059, indicating that PKC-delta might be an upstream component of p42/p44 MAPK. Accordingly, BK-induced phosphorylation of p42/p44 MAPK was attenuated by rottlerin but not by helenalin. These results suggest that in RBA-1 cells, BK-induced cPLA2 expression was sequentially mediated through activation of PKC-delta, p42/p44 MAPK, and NF-kappaB. Understanding the regulation of cPLA2 expression induced by BK in astrocytes might provide a new therapeutic strategy of brain injury and inflammatory diseases.

Oxidized low‐density lipoprotein induces matrix metalloproteinase‐9 expression via a p42/p44 and JNK‐dependent AP‐1 pathway in brain astrocytes

“…Moreover, we also demonstrated that the PI3K may, in part, mediate regulating p42/p44 MAPK activation (Fig. 5A), consistent with our recent findings with COX-2 expression in vascular smooth muscle cells (Hsieh et al, 2006).…”

“…Initiation of GPCR signaling commonly integrates into the serine/threonine protein kinases, mitogen-activated protein kinase (MAPK) cascades including p42/p44 MAPK, c-Jun N-terminal kinases (JNK or stress-activated protein kinase), and p38 MAPK that mediate various cell responses induced by oxLDL (Anwar et al, 2005;Chien et al, 2003). In astrocytes, MAPK signals are triggered by IL1b and BK, resulting in upregulation of inflammatory proteins including MMP-9 and cPLA 2 (Hsieh et al, , 2006Wu et al, 2004). Furthermore, our previous report has indicated that PI3K/Akt mediates oxLDLinduced vascular smooth muscle cell proliferation (Chien et al, 2003) and prolongs survival of macrophage foam cells in atherosclerosic lesions (Boullier et al, 2006).…”

Protein kinase C δ is a critical regulator of CD1d‐mediated antigen presentation

“…Thus, these results suggest that PKCd regulates antigen presentation by molecules that acquire their antigens in the endocytic pathway, but not by those that present endogenous peptide ligands to CTL. It has been reported that PKCd plays a role in the control of MAPK-mediated signaling, mainly upstream of p38 and ERK1/2 [7][8][9], and we have recently demonstrated that both p38 and ERK1/2 participate in the regulation of CD1d-mediated antigen presentation [2]. Interestingly, the phosphorylation of both p38 and ERK1/2 was decreased in the DN PKCd-expressing cells as compared to the control (Fig.…”

Bradykinin induces matrix metalloproteinase‐9 expression and cell migration through a PKC‐δ‐dependent ERK/Elk‐1 pathway in astrocytes

“…Two hundred microliter of homogenization buffer A (20 mM Tris-HCl, pH 8.0, 10 mM EGTA, 2 mM EDTA, 2 mM dithiothreitol, 1 mM phenylmethylsulfonyl fluoride, 25 mg/mL aprotinin, 10 mg/mL leupeptin) was added to each dish, and the cells were scraped into a 1.5-mL tube with a rubber policeman. The cytosolic, membrane, and nuclear fractions were prepared by centrifugation as described previously (Hsieh et al, 2006). The protein concentration of each sample was determined by the BCA reagents.…”