Skip to main content

Advertisement

Springer Nature Link
Log in

Role of src-suppressed C kinase substrate in rat pulmonary microvascular endothelial hyperpermeability stimulated by inflammatory cytokines

  • Original Research Paper
  • Published:
Inflammation Research Aims and scope Submit manuscript

Abstract

Objective

The aim of the study was to investigate the role of src-suppressed C kinase substrate (SSeCKS) in the modulation of rat pulmonary microvascular endothelial cells (RPMVEC) permeability elicited by interleukin (IL)-1β and tumor necrosis factor (TNF)-α.

Methods

The gene expression of SSeCKS was analyzed by reverse transcription-polymerase chain reaction. Immunoblotting was used to determine the SSeCKS protein expression and the activation of the protein kinase C (PKC) signaling pathway. A RPMVEC monolayer was constructed to determine changes of transendothelial electrical resistance (TER) and FITC-dextran flux (P d) across the monolayer. SSeCKS-specific small interfering RNA was transfected into RPMVEC.

Results

IL-1β and TNF-α activated the PKC signaling pathway in RPMVEC, and up-regulated the gene and protein expression of SSeCKS. Depletion of endogenous SSeCKS in RPMVEC significantly attenuated cytokine-induced decrease in TER and increase in P d, but not to the basal levels. PKC inhibitors also significantly decreased cytokine-induced hyperpermeability and SSeCKS expression.

Conclusions

SSeCKS is involved in the endothelial hyperpermeability induced by IL-1β and TNF-α in inflammatory process.

This is a preview of subscription content, log in via an institution to check access.

Access this article

Log in via an institution

Subscribe and save

Springer+
from $39.99 /Month
  • Starting from 10 chapters or articles per month
  • Access and download chapters and articles from more than 300k books and 2,500 journals
  • Cancel anytime
View plans

Buy Now

Price excludes VAT (USA)
Tax calculation will be finalised during checkout.

Instant access to the full article PDF.

Institutional subscriptions

Fig. 1
Fig. 2
Fig. 3
Fig. 4
Fig. 5
Fig. 6
Fig. 7

Similar content being viewed by others

Explore related subjects

Discover the latest articles and news from researchers in related subjects, suggested using machine learning.

References

  1. Sedgwick JB, Menon I, Gern JE, Busse WW. Effects of inflammatory cytokines on the permeability of human lung microvascular endothelial cell monolayers and differential eosinophil transmigration. J Allergy Clin Immunol. 2002;110:752–6.

    Article  CAS  PubMed  Google Scholar 

  2. Angelini DJ, Hyun SW, Grigoryev DN, Garg P, Gong P, Singh IS, et al. TNF-alpha increases tyrosine phosphorylation of vascular endothelial cadherin and opens the paracellular pathway through fyn activation in human lung endothelia. Am J Physiol Lung Cell Mol Physiol. 2006;291:L1232–45.

    Article  CAS  PubMed  Google Scholar 

  3. Koss M, Pfeiffer GR 2nd, Wang Y, Thomas ST, Yerukhimovich M, Gaarde WA, et al. Ezrin/radixin/moesin proteins are phosphorylated by TNF-alpha and modulate permeability increases in human pulmonary microvascular endothelial cells. J Immunol. 2006;176:1218–27.

    CAS  PubMed  Google Scholar 

  4. Ferro T, Neumann P, Gertzberg N, Clements R, Johnson A. Protein kinase C-alpha mediates endothelial barrier dysfunction induced by TNF-alpha. Am J Physiol Lung Cell Mol Physiol. 2000;278:L1107–17.

    CAS  PubMed  Google Scholar 

  5. Cheng C, Liu H, Ge H, Qian J, Qin J, Sun L, et al. Lipopolysaccharide induces expression of SSeCKS in rat lung microvascular endothelial cell. Mol Cell Biochem. 2007;305:1–8.

    Article  CAS  PubMed  Google Scholar 

  6. Gelman IH. The role of SSeCKS/gravin/AKAP12 scaffolding proteins in the spaciotemporal control of signaling pathways in oncogenesis and development. Front Biosci. 2002;7:d1782–97.

    Article  CAS  PubMed  Google Scholar 

  7. Lin X, Tombler E, Nelson PJ, Ross M, Gelman IH. A novel src- and ras-suppressed protein kinase C substrate associated with cytoskeletal architecture. J Biol Chem. 1996;271:28430–8.

    Article  CAS  PubMed  Google Scholar 

  8. Lee SW, Kim WJ, Choi YK, Song HS, Son MJ, Gelman IH, et al. SSeCKS regulates angiogenesis and tight junction formation in blood-brain barrier. Nat Med. 2003;9:900–6.

    Article  CAS  PubMed  Google Scholar 

  9. Chapline C, Cottom J, Tobin H, Hulmes J, Crabb J, Jaken S. A major, transformation-sensitive PKC-binding protein is also a PKC substrate involved in cytoskeletal remodeling. J Biol Chem. 1998;273:19482–9.

    Article  CAS  PubMed  Google Scholar 

  10. Gelman IH, Lee K, Tombler E, Gordon R, Lin X. Control of cytoskeletal architecture by the src-suppressed C kinase substrate, SSeCKS. Cell Motil Cytoskeleton. 1998;41:1–17.

    Article  CAS  PubMed  Google Scholar 

  11. Kitamura H, Okita K, Fujikura D, Mori K, Iwanaga T, Saito M. Induction of Src-suppressed C kinase substrate (SSeCKS) in vascular endothelial cells by bacterial lipopolysaccharide. J Histochem Cytochem. 2002;50:245–55.

    CAS  PubMed  Google Scholar 

  12. You QH, Sun GY, Wang N, Shen JL, Wang Y. Interleukin-17F-induced pulmonary microvascular endothelial monolayer hyperpermeability via the protein kinase C pathway. J Surg Res. 2009 (in press).

  13. Irwin DC, Tissot van Patot MC, Tucker A, Bowen R. Direct ANP inhibition of hypoxia-induced inflammatory pathways in pulmonary microvascular and macrovascular endothelial monolayers. Am J Physiol Lung Cell Mol Physiol. 2005;288:L849–59.

    Article  CAS  PubMed  Google Scholar 

  14. Liu T, Guevara OE, Warburton RR, Hill NS, Gaestel M, Kayyali US. Modulation of HSP27 alters hypoxia-induced endothelial permeability and related signaling pathways. J Cell Physiol. 2009;220:600–10.

    Article  CAS  PubMed  Google Scholar 

  15. Van der Poll T. Immunotherapy of sepsis. Lancet Infect Dis. 2001;1:165–74.

    Article  PubMed  Google Scholar 

  16. Said FA, Werts C, Elalamy I, Couetil JP, Jacquemin C, Hatmi M. TNF-alpha, inefficient by itself, potentiates IL-1beta-induced PGHS-2 expression in human pulmonary microvascular endothelial cells: requirement of NF-kappaB and p38 MAPK pathways. Br J Pharmacol. 2002;136:1005–14.

    Article  CAS  PubMed  Google Scholar 

  17. Zulueta JJ, Sawhney R, Kayyali U, Fogel M, Donaldson C, Huang H, et al. Modulation of inducible nitric oxide synthase by hypoxia in pulmonary artery endothelial cells. Am J Respir Cell Mol Biol. 2002;26:22–30.

    CAS  PubMed  Google Scholar 

  18. Chen SF, Fei X, Li SH. A new simple method for isolation of microvascular endothelial cells avoiding both chemical and mechanical injuries. Microvasc Res. 1995;50:119–28.

    Article  PubMed  Google Scholar 

  19. Laitinen L. Griffonia simplicifolia lectins bind specifically to endothelial cells and some epithelial cells in mouse tissues. Histochem J. 1987;19:225–34.

    Article  CAS  PubMed  Google Scholar 

  20. Ichikawa N, Naora K, Hirano H, Hashimoto M, Masumura S, Iwamoto K. Isolation and primary culture of rat cerebral microvascular endothelial cells for studying drug transport in vitro. J Pharmacol Toxicol Methods. 1996;36:45–52.

    Article  CAS  PubMed  Google Scholar 

  21. Kelly JJ, Moore TM, Babal P, Diwan AH, Stevens T, Thompson WJ. Pulmonary microvascular and macrovascular endothelial cells: differential regulation of Ca2+ and permeability. Am J Physiol. 1998;274:L810–9.

    CAS  PubMed  Google Scholar 

  22. Tinsley JH, Breslin JW, Teasdale NR, Yuan SY. PKC-dependent, burn-induced adherens junction reorganization and barrier dysfunction in pulmonary microvascular endothelial cells. Am J Physiol Lung Cell Mol Physiol. 2005;289:L217–23.

    Article  CAS  PubMed  Google Scholar 

  23. Yan M, Cheng C, Jiang J, Liu Y, Gao Y, Guo Z, et al. Essential role of SRC suppressed C kinase substrates in schwann cells adhesion, spreading and migration. Neurochem Res. 2009;34:1002–10.

    Article  CAS  PubMed  Google Scholar 

  24. Tinsley JH, Hunter FA, Childs EW. PKC and MLCK-dependent, cytokine-induced rat coronary endothelial dysfunction. J Surg Res. 2009;152:76–83.

    Article  CAS  PubMed  Google Scholar 

  25. Tamaru M, Narumi S. E-selectin gene expression is induced synergistically with the coexistence of activated classic protein kinase C and signalings elicited by interleukin-1beta but not tumor necrosis factor-alpha. J Biol Chem. 1999;274:3753–63.

    Article  CAS  PubMed  Google Scholar 

  26. Tinsley JH, Teasdale NR, Yuan SY. Involvement of PKCdelta and PKD in pulmonary microvascular endothelial cell hyperpermeability. Am J Physiol Cell Physiol. 2004;286:C105–11.

    Article  CAS  PubMed  Google Scholar 

  27. Nelson PJ, Moissoglu K, Vargas J Jr, Klotman PE, Gelman IH. Involvement of the protein kinase C substrate, SSeCKS, in the actin-based stellate morphology of mesangial cells. J Cell Sci. 1999;112(Pt 3):361–70.

    CAS  PubMed  Google Scholar 

  28. Rao R, Redha R, Macias-Perez I, Su Y, Hao C, Zent R, et al. Prostaglandin E2-EP4 receptor promotes endothelial cell migration via ERK activation and angiogenesis in vivo. J Biol Chem. 2007;282:16959–68.

    Article  CAS  PubMed  Google Scholar 

  29. Alba-Loureiro TC, Martins EF, Miyasaka CK, Lopes LR, Landgraf RG, Jancar S, et al. Evidence that arachidonic acid derived from neutrophils and prostaglandin E2 are associated with the induction of acute lung inflammation by lipopolysaccharide of Escherichia coli. Inflamm Res. 2004;53:658–63.

    Article  CAS  PubMed  Google Scholar 

  30. Siflinger-Birnboim A, Johnson A. Protein kinase C modulates pulmonary endothelial permeability: a paradigm for acute lung injury. Am J Physiol Lung Cell Mol Physiol. 2003;284:L435–51.

    CAS  PubMed  Google Scholar 

  31. Rung-ruangkijkrai T, Fujikura D, Kitamura H, Saito M, Iwanaga T. The expression of src-suppressed C kinase substrate (SSeCKS) and uptake of exogenous particles in endothelial and reticular cells. Arch Histol Cytol. 2004;67:135–47.

    Article  CAS  PubMed  Google Scholar 

  32. Mountain DJ, Singh M, Menon B, Singh K. Interleukin-1beta increases expression and activity of matrix metalloproteinase-2 in cardiac microvascular endothelial cells: role of PKCalpha/beta1 and MAPKs. Am J Physiol Cell Physiol. 2007;92:C867–75.

    Google Scholar 

  33. Defazio G, Nico B, Trojano M, Ribatti D, Giorelli M, Ricchiuti F, et al. Inhibition of protein kinase C counteracts TNFalpha-induced intercellular adhesion molecule 1 expression and fluid phase endocytosis on brain microvascular endothelial cells. Brain Res. 2000;863:245–8.

    Article  CAS  PubMed  Google Scholar 

Download references

Acknowledgments

This work was supported by a grant from the National Natural Science Foundation of China (No. 30670935). The authors thank Pro. Yuan Wang (Laboratory of Molecular Biology, Key Laboratory of Gene Resource Utilization for Genetic Diseases of Ministry of Education and Anhui Province) for helpful suggestions and technical support.

Conflict of interest statement

None.

Author information

Authors and Affiliations

  1. Department of Respiratory Medicine, The First Affiliated Hospital of Anhui Medical University, 218 Jixi Road, Hefei, 230022, Anhui, People’s Republic of China

    Qing-Hai You, Geng-Yun Sun, Nan Wang & Shan Chen

  2. Key Laboratory of Gene Resource Utilization for Severe Diseases of Ministry of Education, Key Laboratory of Zoonoses (Anhui Medical University), Hefei, 230022, Anhui, People’s Republic of China

    Qing-Li Luo

Authors
  1. Qing-Hai You
    View author publications

    Search author on:PubMed Google Scholar

  2. Geng-Yun Sun
    View author publications

    Search author on:PubMed Google Scholar

  3. Nan Wang
    View author publications

    Search author on:PubMed Google Scholar

  4. Shan Chen
    View author publications

    Search author on:PubMed Google Scholar

  5. Qing-Li Luo
    View author publications

    Search author on:PubMed Google Scholar

Corresponding author

Correspondence to Geng-Yun Sun.

Additional information

Responsible Editor: Makoto Katori.

Rights and permissions

Reprints and permissions

About this article

Cite this article

You, QH., Sun, GY., Wang, N. et al. Role of src-suppressed C kinase substrate in rat pulmonary microvascular endothelial hyperpermeability stimulated by inflammatory cytokines. Inflamm. Res. 59, 949–958 (2010). https://doi.org/10.1007/s00011-010-0207-3

Download citation

  • Received:

  • Revised:

  • Accepted:

  • Published:

  • Issue Date:

  • DOI: https://doi.org/10.1007/s00011-010-0207-3

Keywords