The C-terminal domain of thrombomodulin regulates monocyte migration with interleukin-6 stimulation

Y. W. Lin, C. Y. Huang, C. M. Shih, W. L. Chang, S. K. Shyue, Y. T. Tsai, C. Y. Lin, C. Y. Lee, Y. J. Chang, N. C. Chang, Feng Yin Lin, Chien Sung Tsai

Research output: Contribution to journalArticle

3 Citations (Scopus)

Abstract

Thrombomodulin (TM) is expressed on the surface of monocyte, which is important in the regulation of cell migration, proliferation, and inflammatory responses. In a previous study, we demonstrated that TM on monocyte is negatively associated with cell migration. However, the mechanisms involved in this process are unclear, therefore, we explored the mechanisms in this study. Chemotactic assays and immunofluorescence showed that TM siRNA increased the chemotaxis of the IL-6-activated THP-1, and aggravated actin assembly relative to the IL-6-treated control. In contrast, cells overexpressing plasmids containing full-length or domain 5 of TM followed by IL-6 treatment displayed lower chemotaxis and less actin assembly. Western blot analysis showed that TM knockdown markedly increased cytoskeleton components cofilin and LIMK1 phosphorylation in IL-6-treated THP-1, whereas, transfected cells with HA-TM FL or HA-TM D5, but not HA-TM D1-3 plasmids, reversed the effects. Activation of ERK1/2 and JNK/SAPK, upstream regulators of cytoskeleton components, were also inhibited in overexpressed group. Immunoprecipitation assay demonstrated that actin interacts with TM and intersectin1 in THP-1. Decreased interaction between intersectin1 and actin in TM knockdowns suggested that the interaction is mediated by TM. Our findings indicate that TM domain 5 is a negative regulator and seems to have the ability to inhibit paxillin, cofilin, LIMK1, and actin activation. The mechanisms for the repression effect of domain 5 may be mediated by inhibition of the ERK1/2 and JNK/SAPK activation. Expression of domain 5 of TM may represent a promising approach for controlling monocyte migration, and TM may have potential applications in treatment of inflammatory diseases.

Original languageEnglish
Pages (from-to)27-39
Number of pages13
JournalEuropean Journal of Inflammation
Volume12
Issue number1
Publication statusPublished - 2014

Fingerprint

Thrombomodulin
Monocytes
Interleukin-6
Actins
Actin Depolymerizing Factors
Chemotaxis
Cytoskeleton
Cell Movement
Plasmids
Paxillin
Aptitude
Immunoprecipitation
Small Interfering RNA

Keywords

  • Cofilin
  • Intersectin1
  • LIMK1
  • Monocyte
  • Paxicillin
  • Thrombomodulin

ASJC Scopus subject areas

  • Immunology
  • Immunology and Allergy

Cite this

The C-terminal domain of thrombomodulin regulates monocyte migration with interleukin-6 stimulation. / Lin, Y. W.; Huang, C. Y.; Shih, C. M.; Chang, W. L.; Shyue, S. K.; Tsai, Y. T.; Lin, C. Y.; Lee, C. Y.; Chang, Y. J.; Chang, N. C.; Lin, Feng Yin; Tsai, Chien Sung.

In: European Journal of Inflammation, Vol. 12, No. 1, 2014, p. 27-39.

Research output: Contribution to journalArticle

Lin, Y. W. ; Huang, C. Y. ; Shih, C. M. ; Chang, W. L. ; Shyue, S. K. ; Tsai, Y. T. ; Lin, C. Y. ; Lee, C. Y. ; Chang, Y. J. ; Chang, N. C. ; Lin, Feng Yin ; Tsai, Chien Sung. / The C-terminal domain of thrombomodulin regulates monocyte migration with interleukin-6 stimulation. In: European Journal of Inflammation. 2014 ; Vol. 12, No. 1. pp. 27-39.
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abstract = "Thrombomodulin (TM) is expressed on the surface of monocyte, which is important in the regulation of cell migration, proliferation, and inflammatory responses. In a previous study, we demonstrated that TM on monocyte is negatively associated with cell migration. However, the mechanisms involved in this process are unclear, therefore, we explored the mechanisms in this study. Chemotactic assays and immunofluorescence showed that TM siRNA increased the chemotaxis of the IL-6-activated THP-1, and aggravated actin assembly relative to the IL-6-treated control. In contrast, cells overexpressing plasmids containing full-length or domain 5 of TM followed by IL-6 treatment displayed lower chemotaxis and less actin assembly. Western blot analysis showed that TM knockdown markedly increased cytoskeleton components cofilin and LIMK1 phosphorylation in IL-6-treated THP-1, whereas, transfected cells with HA-TM FL or HA-TM D5, but not HA-TM D1-3 plasmids, reversed the effects. Activation of ERK1/2 and JNK/SAPK, upstream regulators of cytoskeleton components, were also inhibited in overexpressed group. Immunoprecipitation assay demonstrated that actin interacts with TM and intersectin1 in THP-1. Decreased interaction between intersectin1 and actin in TM knockdowns suggested that the interaction is mediated by TM. Our findings indicate that TM domain 5 is a negative regulator and seems to have the ability to inhibit paxillin, cofilin, LIMK1, and actin activation. The mechanisms for the repression effect of domain 5 may be mediated by inhibition of the ERK1/2 and JNK/SAPK activation. Expression of domain 5 of TM may represent a promising approach for controlling monocyte migration, and TM may have potential applications in treatment of inflammatory diseases.",
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AU - Shyue, S. K.

AU - Tsai, Y. T.

AU - Lin, C. Y.

AU - Lee, C. Y.

AU - Chang, Y. J.

AU - Chang, N. C.

AU - Lin, Feng Yin

AU - Tsai, Chien Sung

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N2 - Thrombomodulin (TM) is expressed on the surface of monocyte, which is important in the regulation of cell migration, proliferation, and inflammatory responses. In a previous study, we demonstrated that TM on monocyte is negatively associated with cell migration. However, the mechanisms involved in this process are unclear, therefore, we explored the mechanisms in this study. Chemotactic assays and immunofluorescence showed that TM siRNA increased the chemotaxis of the IL-6-activated THP-1, and aggravated actin assembly relative to the IL-6-treated control. In contrast, cells overexpressing plasmids containing full-length or domain 5 of TM followed by IL-6 treatment displayed lower chemotaxis and less actin assembly. Western blot analysis showed that TM knockdown markedly increased cytoskeleton components cofilin and LIMK1 phosphorylation in IL-6-treated THP-1, whereas, transfected cells with HA-TM FL or HA-TM D5, but not HA-TM D1-3 plasmids, reversed the effects. Activation of ERK1/2 and JNK/SAPK, upstream regulators of cytoskeleton components, were also inhibited in overexpressed group. Immunoprecipitation assay demonstrated that actin interacts with TM and intersectin1 in THP-1. Decreased interaction between intersectin1 and actin in TM knockdowns suggested that the interaction is mediated by TM. Our findings indicate that TM domain 5 is a negative regulator and seems to have the ability to inhibit paxillin, cofilin, LIMK1, and actin activation. The mechanisms for the repression effect of domain 5 may be mediated by inhibition of the ERK1/2 and JNK/SAPK activation. Expression of domain 5 of TM may represent a promising approach for controlling monocyte migration, and TM may have potential applications in treatment of inflammatory diseases.

AB - Thrombomodulin (TM) is expressed on the surface of monocyte, which is important in the regulation of cell migration, proliferation, and inflammatory responses. In a previous study, we demonstrated that TM on monocyte is negatively associated with cell migration. However, the mechanisms involved in this process are unclear, therefore, we explored the mechanisms in this study. Chemotactic assays and immunofluorescence showed that TM siRNA increased the chemotaxis of the IL-6-activated THP-1, and aggravated actin assembly relative to the IL-6-treated control. In contrast, cells overexpressing plasmids containing full-length or domain 5 of TM followed by IL-6 treatment displayed lower chemotaxis and less actin assembly. Western blot analysis showed that TM knockdown markedly increased cytoskeleton components cofilin and LIMK1 phosphorylation in IL-6-treated THP-1, whereas, transfected cells with HA-TM FL or HA-TM D5, but not HA-TM D1-3 plasmids, reversed the effects. Activation of ERK1/2 and JNK/SAPK, upstream regulators of cytoskeleton components, were also inhibited in overexpressed group. Immunoprecipitation assay demonstrated that actin interacts with TM and intersectin1 in THP-1. Decreased interaction between intersectin1 and actin in TM knockdowns suggested that the interaction is mediated by TM. Our findings indicate that TM domain 5 is a negative regulator and seems to have the ability to inhibit paxillin, cofilin, LIMK1, and actin activation. The mechanisms for the repression effect of domain 5 may be mediated by inhibition of the ERK1/2 and JNK/SAPK activation. Expression of domain 5 of TM may represent a promising approach for controlling monocyte migration, and TM may have potential applications in treatment of inflammatory diseases.

KW - Cofilin

KW - Intersectin1

KW - LIMK1

KW - Monocyte

KW - Paxicillin

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