Endotoxin-Induced Chemokine Expression in Murine Peritoneal Mesothelial Cells: The Role of Toll-Like Receptor 4

Sawako Kato, Yukio Yuzawa, Naotake Tsuboi, Shoichi Maruyama, Yoshiki Morita, Tetsuya Matsuguchi, Seiichi Matsuo

Research output: Contribution to journalArticle

72 Citations (Scopus)

Abstract

Acute peritonitis, in which peritoneal mesothelial cells are directly exposed to bacterial components, is a major cause of peritoneal dysfunction in continuous ambulatory peritoneal dialysis patients. We have previously shown that Toll-like receptors (TLR) are expressed in kidney cells, and LPS induces TLR4-dependent chemokine production in tubular epithelial cells. The present work was designed to investigate the involvement of TLR, especially TLR4, in the lipid A-mediated chemokine production by murine peritoneal mesothelial cells (MPMC). A primary cell culture of MPMC from C3H/HeN mice (wild-type mice; LPS sensitive) and from C3H/HeJ mice (containing a point mutation of TLR4; LPS hyposensitive) was established. The expression profile of the TLR family and their accessory molecules, CD14 and MD-2, which are requisite for the LPS signaling pathway, was examined by RT-PCR, Northern blot test, and immunohistochemical staining. Synthetic lipid A-mediated chemokine production by MPMC was studied. The involvement of MAP kinase family (ERK, JNK, and p38 mitogen-activated protein kinase) and nuclear factor (NF)-κB in these processes was also studied. MPMC constitutively express TLR4, CD14, and MD-2. A prominent induction of monocyte chemotactic protein-1 (MCP-1) and macrophage inflammatory protein (MIP)-2 by MPMC was detected after lipid A stimulation and was strictly dependent on TLR4. Furthermore, TLR4-dependent chemokine production followed by leukocyte influx into the peritoneal cavity was also confirmed in vivo after stimulation with LPS. mRNA expression of MCP-1 was abolished by NF-κB inhibition, but were not affected by the inhibition of ERK, JNK, or p38. As compared with MCP-1, MIP-2 mRNA expression was inhibited by a high dose of curcumin but not by NF-κB decoy oligodeoxynucleotide and individual inhibitions of MAP kinase, suggesting that the additional signaling pathway with NF-κB might be involved in mRNA expression of MIP-2. These show that TLR4 is directly involved in the production of MCP-1 and MIP-2 by MPMC in a NF-κB-dependent manner, but the process does not require any MAP kinase activation. The results provide a candidate molecular target in prevention of it.

Original languageEnglish
Pages (from-to)1289-1299
Number of pages11
JournalJournal of the American Society of Nephrology
Volume15
Issue number5
Publication statusPublished - 01-05-2004
Externally publishedYes

Fingerprint

Toll-Like Receptor 4
Chemokines
Endotoxins
Chemokine CCL2
Lipid A
Toll-Like Receptors
Chemokine CXCL2
Phosphotransferases
Inbred C3H Mouse
Messenger RNA
Curcumin
Primary Cell Culture
Continuous Ambulatory Peritoneal Dialysis
JNK Mitogen-Activated Protein Kinases
Oligodeoxyribonucleotides
Peritoneal Cavity
p38 Mitogen-Activated Protein Kinases
Peritonitis
Point Mutation
Northern Blotting

All Science Journal Classification (ASJC) codes

  • Nephrology

Cite this

Kato, Sawako ; Yuzawa, Yukio ; Tsuboi, Naotake ; Maruyama, Shoichi ; Morita, Yoshiki ; Matsuguchi, Tetsuya ; Matsuo, Seiichi. / Endotoxin-Induced Chemokine Expression in Murine Peritoneal Mesothelial Cells : The Role of Toll-Like Receptor 4. In: Journal of the American Society of Nephrology. 2004 ; Vol. 15, No. 5. pp. 1289-1299.
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Kato, S, Yuzawa, Y, Tsuboi, N, Maruyama, S, Morita, Y, Matsuguchi, T & Matsuo, S 2004, 'Endotoxin-Induced Chemokine Expression in Murine Peritoneal Mesothelial Cells: The Role of Toll-Like Receptor 4', Journal of the American Society of Nephrology, vol. 15, no. 5, pp. 1289-1299.

Endotoxin-Induced Chemokine Expression in Murine Peritoneal Mesothelial Cells : The Role of Toll-Like Receptor 4. / Kato, Sawako; Yuzawa, Yukio; Tsuboi, Naotake; Maruyama, Shoichi; Morita, Yoshiki; Matsuguchi, Tetsuya; Matsuo, Seiichi.

In: Journal of the American Society of Nephrology, Vol. 15, No. 5, 01.05.2004, p. 1289-1299.

Research output: Contribution to journalArticle

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T1 - Endotoxin-Induced Chemokine Expression in Murine Peritoneal Mesothelial Cells

T2 - The Role of Toll-Like Receptor 4

AU - Kato, Sawako

AU - Yuzawa, Yukio

AU - Tsuboi, Naotake

AU - Maruyama, Shoichi

AU - Morita, Yoshiki

AU - Matsuguchi, Tetsuya

AU - Matsuo, Seiichi

PY - 2004/5/1

Y1 - 2004/5/1

N2 - Acute peritonitis, in which peritoneal mesothelial cells are directly exposed to bacterial components, is a major cause of peritoneal dysfunction in continuous ambulatory peritoneal dialysis patients. We have previously shown that Toll-like receptors (TLR) are expressed in kidney cells, and LPS induces TLR4-dependent chemokine production in tubular epithelial cells. The present work was designed to investigate the involvement of TLR, especially TLR4, in the lipid A-mediated chemokine production by murine peritoneal mesothelial cells (MPMC). A primary cell culture of MPMC from C3H/HeN mice (wild-type mice; LPS sensitive) and from C3H/HeJ mice (containing a point mutation of TLR4; LPS hyposensitive) was established. The expression profile of the TLR family and their accessory molecules, CD14 and MD-2, which are requisite for the LPS signaling pathway, was examined by RT-PCR, Northern blot test, and immunohistochemical staining. Synthetic lipid A-mediated chemokine production by MPMC was studied. The involvement of MAP kinase family (ERK, JNK, and p38 mitogen-activated protein kinase) and nuclear factor (NF)-κB in these processes was also studied. MPMC constitutively express TLR4, CD14, and MD-2. A prominent induction of monocyte chemotactic protein-1 (MCP-1) and macrophage inflammatory protein (MIP)-2 by MPMC was detected after lipid A stimulation and was strictly dependent on TLR4. Furthermore, TLR4-dependent chemokine production followed by leukocyte influx into the peritoneal cavity was also confirmed in vivo after stimulation with LPS. mRNA expression of MCP-1 was abolished by NF-κB inhibition, but were not affected by the inhibition of ERK, JNK, or p38. As compared with MCP-1, MIP-2 mRNA expression was inhibited by a high dose of curcumin but not by NF-κB decoy oligodeoxynucleotide and individual inhibitions of MAP kinase, suggesting that the additional signaling pathway with NF-κB might be involved in mRNA expression of MIP-2. These show that TLR4 is directly involved in the production of MCP-1 and MIP-2 by MPMC in a NF-κB-dependent manner, but the process does not require any MAP kinase activation. The results provide a candidate molecular target in prevention of it.

AB - Acute peritonitis, in which peritoneal mesothelial cells are directly exposed to bacterial components, is a major cause of peritoneal dysfunction in continuous ambulatory peritoneal dialysis patients. We have previously shown that Toll-like receptors (TLR) are expressed in kidney cells, and LPS induces TLR4-dependent chemokine production in tubular epithelial cells. The present work was designed to investigate the involvement of TLR, especially TLR4, in the lipid A-mediated chemokine production by murine peritoneal mesothelial cells (MPMC). A primary cell culture of MPMC from C3H/HeN mice (wild-type mice; LPS sensitive) and from C3H/HeJ mice (containing a point mutation of TLR4; LPS hyposensitive) was established. The expression profile of the TLR family and their accessory molecules, CD14 and MD-2, which are requisite for the LPS signaling pathway, was examined by RT-PCR, Northern blot test, and immunohistochemical staining. Synthetic lipid A-mediated chemokine production by MPMC was studied. The involvement of MAP kinase family (ERK, JNK, and p38 mitogen-activated protein kinase) and nuclear factor (NF)-κB in these processes was also studied. MPMC constitutively express TLR4, CD14, and MD-2. A prominent induction of monocyte chemotactic protein-1 (MCP-1) and macrophage inflammatory protein (MIP)-2 by MPMC was detected after lipid A stimulation and was strictly dependent on TLR4. Furthermore, TLR4-dependent chemokine production followed by leukocyte influx into the peritoneal cavity was also confirmed in vivo after stimulation with LPS. mRNA expression of MCP-1 was abolished by NF-κB inhibition, but were not affected by the inhibition of ERK, JNK, or p38. As compared with MCP-1, MIP-2 mRNA expression was inhibited by a high dose of curcumin but not by NF-κB decoy oligodeoxynucleotide and individual inhibitions of MAP kinase, suggesting that the additional signaling pathway with NF-κB might be involved in mRNA expression of MIP-2. These show that TLR4 is directly involved in the production of MCP-1 and MIP-2 by MPMC in a NF-κB-dependent manner, but the process does not require any MAP kinase activation. The results provide a candidate molecular target in prevention of it.

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