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Effect of mitogen-activated protein kinases on chemokine synthesis induced by substance P in mouse pancreatic acinar cells.

Ramnath RD, Sun J, Adhikari S, Bhatia M - J. Cell. Mol. Med. (2007 Nov-Dec)

Bottom Line: Moreover, substance P-induced ERK 1/2, JNK, NFkappaB and AP-1 activation as well as chemokine synthesis were blocked by pre-treatment with either extracellular signal-regulated protein kinase kinase 1 (MEK1) inhibitor or JNK inhibitor.In addition, substance P-induced activation of ERK 112, JNK, NFkappaB and AP-1-driven chemokine production were attenuated by CP96345, a selective NK1 R antagonist, in pancreatic acinar cells.Taken together, these results suggest that substance P-NK1 R induced chemokine production depends on the activation of MAPKs-mediated NFkappaB and AP-1 signalling pathways in mouse pancreatic acini.

View Article: PubMed Central - PubMed

Affiliation: Department of Pharmacology, National University of Singapore, Singapore.

ABSTRACT
Substance P, acting via its neurokinin 1 receptor (NK1 R), plays an important role in mediating a variety of inflammatory processes. Its interaction with chemokines is known to play a crucial role in the pathogenesis of acute pancreatitis. In pancreatic acinar cells, substance P stimulates the release of NFkappaB-driven chemokines. However, the signal transduction pathways by which substance P-NK1 R interaction induces chemokine production are still unclear. To that end, we went on to examine the participation of mitogen-activated protein kinases (MAPKs) in substance P-induced synthesis of pro-inflammatory chemokines, monocyte chemoanractant protein-1 (MCP-I), macrophage inflammatory protein-lalpha (MIP-lalpha) and macrophage inflammatory protein-2 (MIP-2), in pancreatic acini. In this study, we observed a time-dependent activation of ERK1/2, c-Jun N-terminal kinase (JNK), NFkappaB and activator protein-1 (AP-1) when pancreatic acini were stimulated with substance P. Moreover, substance P-induced ERK 1/2, JNK, NFkappaB and AP-1 activation as well as chemokine synthesis were blocked by pre-treatment with either extracellular signal-regulated protein kinase kinase 1 (MEK1) inhibitor or JNK inhibitor. In addition, substance P-induced activation of ERK 112, JNK, NFkappaB and AP-1-driven chemokine production were attenuated by CP96345, a selective NK1 R antagonist, in pancreatic acinar cells. Taken together, these results suggest that substance P-NK1 R induced chemokine production depends on the activation of MAPKs-mediated NFkappaB and AP-1 signalling pathways in mouse pancreatic acini.

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NK1R is involved in substance P-induced NFκ B and AP-1 activation as well as MCP-1, MIP-1α and MIP-2 production. Freshly isolated pancreatic acini, obtained from three mice, were pre-incubated with 1 μgM CP96345 for half an hour followed by stimulation with 1 μM substance P for 45 min. Acini were separated from incubation medium by centrifugation. The pellet was used for (A) NFκB and (B) AP-1 extraction and NFκB (p65) and AP-1 (c-Jun) DN-Abinding assays were carried out. The supernatant was used to measure (C) MCP-1, (D) MIP-1α and (E) MIP-2 levels by ELISA. The results are representative of three independent experiments. Results shown are the means + SE. * P≤0.05 when compared to control, + P≤0.05 when compared to substance P (SP).
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fig09: NK1R is involved in substance P-induced NFκ B and AP-1 activation as well as MCP-1, MIP-1α and MIP-2 production. Freshly isolated pancreatic acini, obtained from three mice, were pre-incubated with 1 μgM CP96345 for half an hour followed by stimulation with 1 μM substance P for 45 min. Acini were separated from incubation medium by centrifugation. The pellet was used for (A) NFκB and (B) AP-1 extraction and NFκB (p65) and AP-1 (c-Jun) DN-Abinding assays were carried out. The supernatant was used to measure (C) MCP-1, (D) MIP-1α and (E) MIP-2 levels by ELISA. The results are representative of three independent experiments. Results shown are the means + SE. * P≤0.05 when compared to control, + P≤0.05 when compared to substance P (SP).

Mentions: The role of NK1R in substance P-induced NFκB and AP-1 activation and chemokine production was confirmed by pre-treating the cells with 1 μM of CP96345 followed by stimulation with 1 μM of substance P for 45 min. The cells were used for nuclear extraction to determine NFκB and AP-1 activation, whereas the supernatant obtained was used for detection of chemokines MCP-1, MIP-1α and MIP-2 by ELISA Our results, in Figure 9, show that pre-treatment with selective antagonist CP96345 significantly inhibited substance P-induced NFκB and AP-1 activation in pancreatic acinar cells when compared to substance P-only treated cells. As shown in Figure 9, CP96345 significantly attenuated substance P-induced MCP-1, MIP-1α, and MIP-2 synthesis when compared to substance P-only treated cells.


Effect of mitogen-activated protein kinases on chemokine synthesis induced by substance P in mouse pancreatic acinar cells.

Ramnath RD, Sun J, Adhikari S, Bhatia M - J. Cell. Mol. Med. (2007 Nov-Dec)

NK1R is involved in substance P-induced NFκ B and AP-1 activation as well as MCP-1, MIP-1α and MIP-2 production. Freshly isolated pancreatic acini, obtained from three mice, were pre-incubated with 1 μgM CP96345 for half an hour followed by stimulation with 1 μM substance P for 45 min. Acini were separated from incubation medium by centrifugation. The pellet was used for (A) NFκB and (B) AP-1 extraction and NFκB (p65) and AP-1 (c-Jun) DN-Abinding assays were carried out. The supernatant was used to measure (C) MCP-1, (D) MIP-1α and (E) MIP-2 levels by ELISA. The results are representative of three independent experiments. Results shown are the means + SE. * P≤0.05 when compared to control, + P≤0.05 when compared to substance P (SP).
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Related In: Results  -  Collection

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getmorefigures.php?uid=PMC4401295&req=5

fig09: NK1R is involved in substance P-induced NFκ B and AP-1 activation as well as MCP-1, MIP-1α and MIP-2 production. Freshly isolated pancreatic acini, obtained from three mice, were pre-incubated with 1 μgM CP96345 for half an hour followed by stimulation with 1 μM substance P for 45 min. Acini were separated from incubation medium by centrifugation. The pellet was used for (A) NFκB and (B) AP-1 extraction and NFκB (p65) and AP-1 (c-Jun) DN-Abinding assays were carried out. The supernatant was used to measure (C) MCP-1, (D) MIP-1α and (E) MIP-2 levels by ELISA. The results are representative of three independent experiments. Results shown are the means + SE. * P≤0.05 when compared to control, + P≤0.05 when compared to substance P (SP).
Mentions: The role of NK1R in substance P-induced NFκB and AP-1 activation and chemokine production was confirmed by pre-treating the cells with 1 μM of CP96345 followed by stimulation with 1 μM of substance P for 45 min. The cells were used for nuclear extraction to determine NFκB and AP-1 activation, whereas the supernatant obtained was used for detection of chemokines MCP-1, MIP-1α and MIP-2 by ELISA Our results, in Figure 9, show that pre-treatment with selective antagonist CP96345 significantly inhibited substance P-induced NFκB and AP-1 activation in pancreatic acinar cells when compared to substance P-only treated cells. As shown in Figure 9, CP96345 significantly attenuated substance P-induced MCP-1, MIP-1α, and MIP-2 synthesis when compared to substance P-only treated cells.

Bottom Line: Moreover, substance P-induced ERK 1/2, JNK, NFkappaB and AP-1 activation as well as chemokine synthesis were blocked by pre-treatment with either extracellular signal-regulated protein kinase kinase 1 (MEK1) inhibitor or JNK inhibitor.In addition, substance P-induced activation of ERK 112, JNK, NFkappaB and AP-1-driven chemokine production were attenuated by CP96345, a selective NK1 R antagonist, in pancreatic acinar cells.Taken together, these results suggest that substance P-NK1 R induced chemokine production depends on the activation of MAPKs-mediated NFkappaB and AP-1 signalling pathways in mouse pancreatic acini.

View Article: PubMed Central - PubMed

Affiliation: Department of Pharmacology, National University of Singapore, Singapore.

ABSTRACT
Substance P, acting via its neurokinin 1 receptor (NK1 R), plays an important role in mediating a variety of inflammatory processes. Its interaction with chemokines is known to play a crucial role in the pathogenesis of acute pancreatitis. In pancreatic acinar cells, substance P stimulates the release of NFkappaB-driven chemokines. However, the signal transduction pathways by which substance P-NK1 R interaction induces chemokine production are still unclear. To that end, we went on to examine the participation of mitogen-activated protein kinases (MAPKs) in substance P-induced synthesis of pro-inflammatory chemokines, monocyte chemoanractant protein-1 (MCP-I), macrophage inflammatory protein-lalpha (MIP-lalpha) and macrophage inflammatory protein-2 (MIP-2), in pancreatic acini. In this study, we observed a time-dependent activation of ERK1/2, c-Jun N-terminal kinase (JNK), NFkappaB and activator protein-1 (AP-1) when pancreatic acini were stimulated with substance P. Moreover, substance P-induced ERK 1/2, JNK, NFkappaB and AP-1 activation as well as chemokine synthesis were blocked by pre-treatment with either extracellular signal-regulated protein kinase kinase 1 (MEK1) inhibitor or JNK inhibitor. In addition, substance P-induced activation of ERK 112, JNK, NFkappaB and AP-1-driven chemokine production were attenuated by CP96345, a selective NK1 R antagonist, in pancreatic acinar cells. Taken together, these results suggest that substance P-NK1 R induced chemokine production depends on the activation of MAPKs-mediated NFkappaB and AP-1 signalling pathways in mouse pancreatic acini.

Show MeSH
Related in: MedlinePlus