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Upregulation of RGS4 expression by IL-1beta in colonic smooth muscle is enhanced by ERK1/2 and p38 MAPK and inhibited by the PI3K/Akt/GSK3beta pathway.

ABSTRACT: Initial Ca(2+)-dependent contraction of intestinal smooth muscle is inhibited upon IL-1beta treatment. The decrease in contraction reflects the upregulation of regulator of G protein signaling-4 (RGS4) via the canonical inhibitor of NF-kappaB kinase-2 (IKK2)/IkappaB-alpha/NF-kappaB pathway. Here, we show that the activation of various protein kinases, including ERK1/2, p38 MAPK, and phosphoinositide 3-kinase (PI3K), differentially modulates IL-1beta-induced upregulation of RGS4 in rabbit colonic muscle cells. IL-1beta treatment caused a transient phosphorylation of ERK1/2 and p38 MAPK. It also caused the phosphorylation of Akt and glycogen synthase kinase-3beta (GSK3beta), sequential downstream effectors of PI3K. Pretreatment with PD-98059 (an ERK inhibitor) and SB-203580 (a p38 MAPK inhibitor) significantly inhibited IL-1beta-induced RGS4 expression. In contrast, LY-294002 (a PI3K inhibitor) augmented, whereas GSK3beta inhibitors inhibited, IL-1beta-induced RGS4 expression. PD-98059 blocked IL-1beta-induced phosphorylation of IKK2, degradation of IkappaB-alpha, and phosphorylation and nuclear translocation of NF-kappaB subunit p65, whereas SB-203580 had a marginal effect, implying that the effect of ERK1/2 is exerted on the canonical IKK2/IkappaB-alpha/p65 pathway of NF-kappaB activation but that the effect of p38 MAPK may not predominantly involve NF-kappaB signaling. The increase in RGS4 expression enhanced by LY-294002 was accompanied by an increase in the phosphorylation of IKK2/IkappaB-alpha/p65 and blocked by pretreatment with inhibitors of IKK2 (IKK2-IV) and IkappaB-alpha (MG-132). Inhibition of GSK3beta abolished IL-1beta-induced phosphorylation of IKK2/p65. These findings suggest that ERK1/2 and p38 MAPK enhance IL-1beta-induced upregulation of RGS4; the effect of ERK1/2 reflects its ability to promote IKK2 phosphorylation and increase NF-kappaB activity. GSK3beta acts normally to augment the activation of the canonical NF-kappaB signaling. The PI3K/Akt/GSK3beta pathway attenuates IL-1beta-induced upregulation of RGS4 expression by inhibiting NF-kappaB activation.

SUBMITTER: Hu W

PROVIDER: S-EPMC2692422 | biostudies-literature | 2009 Jun

REPOSITORIES: biostudies-literature

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Upregulation of RGS4 expression by IL-1beta in colonic smooth muscle is enhanced by ERK1/2 and p38 MAPK and inhibited by the PI3K/Akt/GSK3beta pathway.

Hu Wenhui W Li Fang F Mahavadi Sunila S Murthy Karnam S KS

American journal of physiology. Cell physiology 20090415 6

Initial Ca(2+)-dependent contraction of intestinal smooth muscle is inhibited upon IL-1beta treatment. The decrease in contraction reflects the upregulation of regulator of G protein signaling-4 (RGS4) via the canonical inhibitor of NF-kappaB kinase-2 (IKK2)/IkappaB-alpha/NF-kappaB pathway. Here, we show that the activation of various protein kinases, including ERK1/2, p38 MAPK, and phosphoinositide 3-kinase (PI3K), differentially modulates IL-1beta-induced upregulation of RGS4 in rabbit colonic ...[more]

PMID: 19369446

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Project description:Initial Ca2+-dependent contraction of the intestinal smooth muscle mediated by G(q)-coupled receptors is attenuated by RGS4 (regulator of G-protein signalling 4). Treatment of colonic muscle cells with IL-1beta (interleukin-1beta) inhibits acetylcholine-stimulated initial contraction through increasing the expression of RGS4. NF-kappaB (nuclear factor kappaB) signalling is the dominant pathway activated by IL-1beta. In the present study we show that RGS4 is a new target gene regulated by IL-1beta/NF-kappaB signalling. Exposure of cultured rabbit colonic muscle cells to IL-1beta induced a rapid increase in RGS4 mRNA expression, which was abolished by pretreatment with a transcription inhibitor, actinomycin D, implying a transcription-dependent mechanism. Existence of the canonical IKK2 [IkappaB (inhibitor of NF-kappaB) kinase 2]/IkappaBalpha pathway of NF-kappaB activation induced by IL-1beta in rabbit colonic muscle cells was validated with multiple approaches, including the induction of reporter luciferase activity and endogenous NF-kappaB-target gene expression, NF-kappaB-DNA binding activity, p65 nuclear translocation, IkappaBalpha degradation and the phosphorylation of IKK2 at Ser(177/181) and p65 at Ser(536). RGS4 up-regulation by IL-1beta was blocked by selective inhibitors of IKK2, IkappaBalpha or NF-kappaB activation, by effective siRNA (small interfering RNA) of IKK2, and in cells expressing either the kinase-inactive IKK2 mutant (K44A) or the phosphorylation-deficient IkappaBalpha mutant (S32A/S36A). An IKK2-specific inhibitor or effective siRNA prevented IL-1beta-induced inhibition of acetylcholine-stimulated PLC-beta (phopsholipase C-beta) activation. These results suggest that the canonical IKK2/IkappaBalpha pathway of NF-kappaB activation mediates the up-regulation of RGS4 expression in response to IL-1beta and contributes to the inhibitory effect of IL-1beta on acetylcholine-stimulated PLC-beta-dependent initial contraction in rabbit colonic smooth muscle.

Project description:BACKGROUND: Osteoporosis is a worldwide health problem predominantly affecting post-menopausal women. Therapies aimed at increasing bone mass in osteoporetic patients lag behind comparable investigation of therapeutic strategies focusing on the bone resorption process. Sesamin, a major lignan compound found in Sesamun indicum Linn., has a variety of pharmacological effects, though its activity on bone cell function is unclear. Herein we examine the effect of this lignan on osteoblast differentiation and function. METHOD: Cell cytotoxicity and proliferative in hFOB1.19 were examined by MTT and alamar blue assay up to 96 h of treatment. Gene expression of COL1, ALP, BMP-2, Runx2, OC, RANKL and OPG were detected after 24 h of sesamin treatment. ALP activity was measured at day 7, 14 and 21 of cultured. For mineralized assay, ADSCs were cultured in the presence of osteogenic media supplement with or without sesamin for 21 days and then stained with Alizarin Red S. MAPK signaling pathway activation was observed by using western blotting. RESULTS: Sesamin promoted the gene expression of COL1, ALP, OCN, BMP-2 and Runx2 in hFOB1.19. On the other hand, sesamin was able to up-regulate OPG and down-regulate RANKL gene expression. ALP activity also significantly increased after sesamin treatment. Interestingly, sesamin induced formation of mineralized nodules in adipose derived stem cells (ADSCs) as observed by Alizarin Red S staining; this implies that sesamin has anabolic effects both on progenitor and committed cell stages of osteoblasts. Western blotting data showed that sesamin activated phosphorylation of p38 and ERK1/2 in hFOB1.19. CONCLUSIONS: The data suggest that sesamin has the ability to trigger osteoblast differentiation by activation of the p38 and ERK MAPK signaling pathway and possibly indirectly regulate osteoclast development via the expression of OPG and RANKL in osteoblasts. Therefore, sesamin may be a promising phytochemical that could be developed for supplementation of osteoporotic therapy.

Dataset Information

Upregulation of RGS4 expression by IL-1beta in colonic smooth muscle is enhanced by ERK1/2 and p38 MAPK and inhibited by the PI3K/Akt/GSK3beta pathway.

Publications

Upregulation of RGS4 expression by IL-1beta in colonic smooth muscle is enhanced by ERK1/2 and p38 MAPK and inhibited by the PI3K/Akt/GSK3beta pathway.

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