Advertisement

Calpain pp 209-218 | Cite as

Experimental Manipulation of Calpain Activity In Vitro

  • Seiichi Kitagawa
Protocol
Part of the Methods in Molecular Biology book series (MIMB, volume 1915)

Abstract

The calpain activity in cells can be experimentally manipulated in vitro by calpain inhibitors, and various types of calpain inhibitors such as peptide aldehydes and α-mercapto-acrylic acid derivatives are widely used as a valuable tool to elucidate the physiological and pathological roles of calpain. Here I describe the experimental procedures with calpain inhibitors, with human neutrophils being primarily used in this experiment. It should be noted that potent calpain inhibitors not only inhibit the calpain activity but also stimulate cell functions via direct activation of human formyl peptide receptors and/or other G protein-coupled receptors depending on the inhibitors used.

Key words

Calpain Calpain inhibitors Human formyl peptide receptors G protein-coupled receptors Migration Apoptosis 

References

  1. 1.
    Lokuta MA, Nuzzi PA, Huttenlocher A (2003) Calpain regulates neutrophil chemotaxis. Proc Natl Acad Sci U S A 100:4006–4011CrossRefGoogle Scholar
  2. 2.
    Katsube M, Kato T, Kitagawa M, Noma H, Fujita H, Kitagawa S (2008) Calpain-mediated regulation of the distinct signaling pathways and cell migration in human neutrophils. J Leukoc Biol 84:255–263CrossRefGoogle Scholar
  3. 3.
    Ozaki Y, Kato T, Kitagawa M, Fujita H, Kitagawa S (2008) Calpain inhibition delays neutrophil apoptosis via cyclic AMP-independent activation of protein kinase A and protein kinase A-mediated stabilization of Mcl-1 and X-linked inhibitor of apoptosis (XIAP). Arch Biochem Biophys 477:227–231CrossRefGoogle Scholar
  4. 4.
    Camins A, Verdaguer E, Folch J, Pallas M (2006) Involvement of calpain activation in neurodegenerative processes. CNS Drug Rev 12:135–148CrossRefGoogle Scholar
  5. 5.
    Mani SK, Balasubramanian S, Zavadzkas JA, Jeffords LB, Rivers WT, Zile MR, Mukherjee R, Spinale FG, Kuppuswamy D (2009) Calpain inhibition preserves myocardial structure and function following myocardial infarction. Am J Physiol Heart Circ Physiol 297:H1744–H1751CrossRefGoogle Scholar
  6. 6.
    Noma H, Kato T, Fujita H, Kitagawa M, Yamano T, Kitagawa S (2009) Calpain inhibition induces activation of the distinct signalling pathways and cell migration in human monocytes. Immunology 128:e487–e496CrossRefGoogle Scholar
  7. 7.
    Sasaki T, Kishi M, Saito M, Tanaka T, Higuchi N, Kominam E, Katunuma N, Murachi T (1990) Inhibitory effect of di- and tripeptidyl aldehydes on calpains and cathepsins. J Enzym Inhib 3:195–201CrossRefGoogle Scholar
  8. 8.
    Wang KK, Nath R, Posner A, Raser KJ, Buroker-Kilgore M, Hajimohammadreza I, Probert AW Jr, Marcoux FW, Ye Q, Takano E, Hatanaka M, Maki M, Caner H, Collins JL, Fergus A, Lee KS, Lunney EA, Hays SJ, Yuen P (1996) An alpha-mercaptoacrylic acid derivative is a selective nonpeptide cell-permeable calpain inhibitor and is neuroprotective. Proc Natl Acad Sci U S A 93:6687–6692CrossRefGoogle Scholar
  9. 9.
    Rock KL, Gramm C, Rothstein L, Clark K, Stein R, Dick L, Hwang D, Goldberg AL (1994) Inhibitors of the proteasome block the degradation of most cell proteins and the generation of peptides presented on MHC class I molecules. Cell 78:761–771CrossRefGoogle Scholar
  10. 10.
    Fujita H, Kato T, Watanabe N, Takahashi T, Kitagawa S (2011) Calpain inhibitors stimulate phagocyte functions via activation of human formyl peptide receptors. Arch Biochem Biophys 513:51–60CrossRefGoogle Scholar
  11. 11.
    Fujita H, Kato T, Watanabe N, Takahashi T, Kitagawa S (2011) Stimulation of human formyl peptide receptors by calpain inhibitors: homology modeling of receptors and ligand docking simulation. Arch Biochem Biophys 516:121–127CrossRefGoogle Scholar
  12. 12.
    Kitagawa S, Kato T, Kitagawa M, Aomatsu M, Fujita H (2013) Biological effects of calpain inhibitors on human phagocyte functions. Chapter V. In: Lashinski EM (ed) Enzymes and enzyme activity. Nova Science Publishers, New York, pp 121–144Google Scholar
  13. 13.
    Kobayashi S, Yamashita K, Takeoka T, Ohtsuki T, Suzuki Y, Takahashi R, Yamamoto K, Kaufmann SH, Uchiyama T, Sasada M, Takahashi A (2002) Calpain-mediated X-linked inhibitor of apoptosis degradation in neutrophil apoptosis and its impairment in chronic neutrophilic leukemia. J Biol Chem 277:33968–33977CrossRefGoogle Scholar
  14. 14.
    Derouet M, Thomas L, Cross A, Moots RJ, Edwards SW (2004) Granulocyte macrophage colony-stimulating factor signaling and proteasome inhibition delay neutrophil apoptosis by increasing the stability of Mcl-1. J Biol Chem 279:26915–26921CrossRefGoogle Scholar
  15. 15.
    Kato T, Kutsuna H, Oshitani N, Kitagawa S (2006) Cyclic AMP delays neutrophil apoptosis via stabilization of Mcl-1. FEBS Lett 580:4582–4458CrossRefGoogle Scholar

Copyright information

© Springer Science+Business Media, LLC, part of Springer Nature 2019

Authors and Affiliations

  1. 1.Department of Physiology, Graduate School of MedicineOsaka City UniversityOsakaJapan
  2. 2.Shitennoji Tamatsukurien ClinicOsakaJapan

Personalised recommendations