Molecular and Cellular Biochemistry

, Volume 440, Issue 1–2, pp 199–208 | Cite as

The significance of aryl acylamidase activity of acetylcholinesterase in osteoblast differentiation and mineralization

  • Raj Kumar Chinnadurai
  • Ponne Saravanaraman
  • Rathanam Boopathy
Article

Abstract

Osteoblast differentiation is an essential event in the developmental process, which is favoured by the production of extra cellular matrix proteins and various enzymes including discrete ones like acetylcholinesterase (AChE). Despite the fact that AChE facilitates osteoblast differentiation, the significance of its catalytic functions [esterase and aryl acylamidase (AAA) activities] in the process is yet to be ascertained. In this context, SaOS-2 cell line was used in the present study to implicate the catalytic activities of AChE in process of osteoblast differentiation and mineralization. During differentiation, it was found that the activity of both esterase and AAA increased 1.13 and 1.46 folds respectively, signifying the involvement of catalytic activities of AChE in the process. Inhibition of both the catalytic activities of AChE with edrophonium significantly reduced the amount of mineralization by decreasing the alkaline phosphatase (ALP) activity and expression of differentiation-related genes such as RUNX-2, COL1A, ALP, OC, and OP significantly (p < 0.05). Inhibition of esterase activity without altering the AAA activity using gallamine significantly increased the level ALP activity and expression of differentiation-associated genes (p < 0.05), thus favouring mineralization. Therefore, this study concludes and confirms that the AAA activity of AChE is actively involved in the process of osteoblast differentiation and mineralization.

Keywords

Osteoblast Aryl acylamidase Esterase Acetylcholinesterase Differentiation Mineralization 

Notes

Acknowledgements

We thank Dr. Paul G. Layer for encouragements, valuable comments, helpful discussions and counsel of the manuscript. PS acknowledges the research fellowship (DST-INSPIRE) provided by the Department of Science and Technology, Government of India.

Compliance with ethical standards

Conflict of interest

The authors of the manuscript declare no conflict of interest.

Supplementary material

11010_2017_3167_MOESM1_ESM.doc (9.6 mb)
Supplementary material 1 (DOC 9881 kb)

References

  1. 1.
    Ollis DL, Cheah E, Cygler M, Dijkstra B, Frolow F, Franken SM, Harel M, Remington SJ, Silman I, Schrag J et al (1992) The alpha/beta hydrolase fold. Protein Eng 5(3):197–211CrossRefPubMedGoogle Scholar
  2. 2.
    Bigbee JW, Sharma KV, Chan EL, Bogler O (2000) Evidence for the direct role of acetylcholinesterase in neurite outgrowth in primary dorsal root ganglion neurons. Brain Res 861(2):354–362CrossRefPubMedGoogle Scholar
  3. 3.
    Grifman M, Galyam N, Seidman S, Soreq H (1998) Functional redundancy of acetylcholinesterase and neuroligin in mammalian neuritogenesis. Proc Natl Acad Sci 95(23):13935–13940CrossRefPubMedPubMedCentralGoogle Scholar
  4. 4.
    Koenigsberger C, Chiappa S, Brimijoin S (1997) Neurite differentiation is modulated in neuroblastoma cells engineered for altered acetylcholinesterase expression. J Neurochem 69(4):1389–1397CrossRefPubMedGoogle Scholar
  5. 5.
    Layer PG, Weikert T, Alber R (1993) Cholinesterases regulate neurite growth of chick nerve cells in vitro by means of a non-enzymatic mechanism. Cell Tissue Res 273(2):219–226CrossRefPubMedGoogle Scholar
  6. 6.
    Conscience JF, Meier W (1980) Coordinate expression of erythroid marker enzymes during dimethylsulfoxide-induced differentiation of Friend erythroleukemia cells. Exp Cell Res 125(1):111–119CrossRefPubMedGoogle Scholar
  7. 7.
    Lev-Lehman E, Ginzberg D, Hornreich G, Ehrlich G, Meshorer A, Eckstein F, Soreq H, Zakut H (1994) Antisense inhibition of acetylcholinesterase gene expression causes transient hematopoietic alterations in vivo. Gene Ther 1(2):127–135PubMedGoogle Scholar
  8. 8.
    Genever PG, Birch MA, Brown E, Skerry TM (1999) Osteoblast-derived acetylcholinesterase: a novel mediator of cell-matrix interactions in bone? Bone 24(4):297–303CrossRefPubMedGoogle Scholar
  9. 9.
    Inkson CA, Brabbs AC, Grewal TS, Skerry TM, Genever PG (2004) Characterization of acetylcholinesterase expression and secretion during osteoblast differentiation. Bone 35(4):819–827. doi: 10.1016/j.bone.2004.05.026 CrossRefPubMedGoogle Scholar
  10. 10.
    Fujimoto D (1976) Serotonin-sensitive aryl acylamidase activity of acetylcholinesterase. FEBS Lett 72(1):121–123CrossRefPubMedGoogle Scholar
  11. 11.
    Grisaru D, Lev-Lehman E, Shapira M, Chaikin E, Lessing JB, Eldor A, Eckstein F, Soreq H (1999) Human osteogenesis involves differentiation-dependent increases in the morphogenically active 3′ alternative splicing variant of acetylcholinesterase. Mol Cell Biol 19(1):788–795CrossRefPubMedPubMedCentralGoogle Scholar
  12. 12.
    Costagli C, Galli A (1998) Inhibition of cholinesterase-associated aryl acylamidase activity by anticholinesterase agents: focus on drugs potentially effective in Alzheimer’s disease. Biochem Pharmacol 55(10):1733–1737CrossRefPubMedGoogle Scholar
  13. 13.
    Akincioglu A, Akincioglu H, Gulcin I, Durdagi S, Supuran CT, Goksu S (2015) Discovery of potent carbonic anhydrase and acetylcholine esterase inhibitors: novel sulfamoylcarbamates and sulfamides derived from acetophenones. Bioorg Med Chem 23(13):3592–3602. doi: 10.1016/j.bmc.2015.04.019 CrossRefPubMedGoogle Scholar
  14. 14.
    Gocer H, Akincioglu A, Oztaskin N, Goksu S, Gulcin I (2013) Synthesis, antioxidant and antiacetylcholinesterase activities of sulfonamide derivatives of dopamine-related compounds. Arch Pharm 346(11):783–792. doi: 10.1002/ardp.201300228 CrossRefGoogle Scholar
  15. 15.
    Chinnadurai RK, Saravanaraman P, Boopathy R (2015) Understanding the molecular mechanism of aryl acylamidase activity of acetylcholinesterase—an in silico study. Arch Biochem Biophys 580:1–13. doi: 10.1016/j.abb.2015.06.002 CrossRefPubMedGoogle Scholar
  16. 16.
    Ellman GL, Courtney KD, Andres V Jr, Feather-Stone RM (1961) A new and rapid colorimetric determination of acetylcholinesterase activity. Biochem Pharmacol 7:88–95CrossRefPubMedGoogle Scholar
  17. 17.
    Rajesh RV, Chitra L, Layer PG, Boopathy R (2009) The aryl acylamidase activity is much more sensitive to Alzheimer drugs than the esterase activity of acetylcholinesterase in chicken embryonic brain. Biochimie 91(9):1087–1094. doi: 10.1016/j.biochi.2009.07.004 CrossRefPubMedGoogle Scholar
  18. 18.
    Bessey OA, Lowry OH, Brock MJ (1946) A method for the rapid determination of alkaline phosphates with five cubic millimeters of serum. J Biol Chem 164:321–329PubMedGoogle Scholar
  19. 19.
    Lowry OH, Rosebrough NJ, Farr AL, Randall RJ (1951) Protein measurement with the Folin phenol reagent. J Biol Chem 193(1):265–275PubMedGoogle Scholar
  20. 20.
    Golub EE, Boesze-Battaglia K (2007) The role of alkaline phosphatase in mineralization. Curr Opin Orthop 18(5):444–448. doi: 10.1097/BCO.0b013e3282630851 CrossRefGoogle Scholar
  21. 21.
    Mornet E, Stura E, Lia-Baldini AS, Stigbrand T, Menez A, Le Du MH (2001) Structural evidence for a functional role of human tissue nonspecific alkaline phosphatase in bone mineralization. J Biol Chem 276(33):31171–31178. doi: 10.1074/jbc.M102788200 CrossRefPubMedGoogle Scholar
  22. 22.
    Anderson HC, Sipe JB, Hessle L, Dhanyamraju R, Atti E, Camacho NP, Millan JL (2004) Impaired calcification around matrix vesicles of growth plate and bone in alkaline phosphatase-deficient mice. Am J Pathol 164(3):841–847CrossRefPubMedPubMedCentralGoogle Scholar
  23. 23.
    Weiss MJ, Cole DE, Ray K, Whyte MP, Lafferty MA, Mulivor RA, Harris H (1988) A missense mutation in the human liver/bone/kidney alkaline phosphatase gene causing a lethal form of hypophosphatasia. Proc Natl Acad Sci 85(20):7666–7669CrossRefPubMedPubMedCentralGoogle Scholar
  24. 24.
    Wennberg C, Hessle L, Lundberg P, Mauro S, Narisawa S, Lerner UH, Millan JL (2000) Functional characterization of osteoblasts and osteoclasts from alkaline phosphatase knockout mice. J Bone Miner Res 15(10):1879–1888. doi: 10.1359/jbmr.2000.15.10.1879 CrossRefPubMedGoogle Scholar
  25. 25.
    Yadav MC, Simao AM, Narisawa S, Huesa C, McKee MD, Farquharson C, Millan JL (2011) Loss of skeletal mineralization by the simultaneous ablation of PHOSPHO1 and alkaline phosphatase function: a unified model of the mechanisms of initiation of skeletal calcification. J Bone Miner Res 26(2):286–297. doi: 10.1002/jbmr.195 CrossRefPubMedGoogle Scholar
  26. 26.
    Yoon K, Golub E, Rodan GA (1989) Alkaline phosphatase cDNA transfected cells promote calcium and phosphate deposition. Connect Tissue Res 22(1–4):17–25PubMedGoogle Scholar
  27. 27.
    Komori T (2010) Regulation of osteoblast differentiation by Runx2. Adv Exp Med Biol 658:43–49. doi: 10.1007/978-1-4419-1050-9_5 CrossRefPubMedGoogle Scholar
  28. 28.
    Fujita T, Azuma Y, Fukuyama R, Hattori Y, Yoshida C, Koida M, Ogita K, Komori T (2004) Runx2 induces osteoblast and chondrocyte differentiation and enhances their migration by coupling with PI3K-Akt signaling. J Cell Biol 166(1):85–95. doi: 10.1083/jcb.200401138 CrossRefPubMedPubMedCentralGoogle Scholar
  29. 29.
    Zhang X, Yang M, Lin L, Chen P, Ma KT, Zhou CY, Ao YF (2006) Runx2 overexpression enhances osteoblastic differentiation and mineralization in adipose—derived stem cells in vitro and in vivo. Calcif Tissue Int 79(3):169–178. doi: 10.1007/s00223-006-0083-6 CrossRefPubMedGoogle Scholar
  30. 30.
    Komori T (2005) Regulation of skeletal development by the Runx family of transcription factors. J Cell Biochem 95(3):445–453. doi: 10.1002/jcb.20420 CrossRefPubMedGoogle Scholar
  31. 31.
    Komori T (2010) Regulation of bone development and extracellular matrix protein genes by RUNX2. Cell Tissue Res 339(1):189–195. doi: 10.1007/s00441-009-0832-8 CrossRefPubMedGoogle Scholar
  32. 32.
    Mochida Y, Duarte WR, Tanzawa H, Paschalis EP, Yamauchi M (2003) Decorin modulates matrix mineralization in vitro. Biochem Biophys Res Commun 305(1):6–9CrossRefPubMedGoogle Scholar
  33. 33.
    Nair AK, Gautieri A, Chang S-W, Buehler MJ (2013) Molecular mechanics of mineralized collagen fibrils in bone. Nat Commun 4:1724CrossRefPubMedPubMedCentralGoogle Scholar
  34. 34.
    Viguet-Carrin S, Garnero P, Delmas PD (2006) The role of collagen in bone strength. Osteoporos Int 17(3):319–336. doi: 10.1007/s00198-005-2035-9 CrossRefPubMedGoogle Scholar
  35. 35.
    Wang Y, Azaïs T, Robin M, Vallée A, Catania C, Legriel P, Pehau-Arnaudet G, Babonneau F, Giraud-Guille M-M, Nassif N (2012) The predominant role of collagen in the nucleation, growth, structure and orientation of bone apatite. Nat Mater 11(8):724–733CrossRefPubMedGoogle Scholar
  36. 36.
    Bouleftour W, Bouet G, Granito RN, Thomas M, Linossier MT, Vanden-Bossche A, Aubin JE, Lafage-Proust MH, Vico L, Malaval L (2015) Blocking the expression of both bone sialoprotein (BSP) and osteopontin (OPN) impairs the anabolic action of PTH in mouse calvaria bone. J Cell Physiol 230(3):568–577. doi: 10.1002/jcp.24772 CrossRefPubMedGoogle Scholar
  37. 37.
    Reffitt DM, Ogston N, Jugdaohsingh R, Cheung HF, Evans BA, Thompson RP, Powell JJ, Hampson GN (2003) Orthosilicic acid stimulates collagen type 1 synthesis and osteoblastic differentiation in human osteoblast-like cells in vitro. Bone 32(2):127–135CrossRefPubMedGoogle Scholar
  38. 38.
    Salim A, Nacamuli RP, Morgan EF, Giaccia AJ, Longaker MT (2004) Transient changes in oxygen tension inhibit osteogenic differentiation and Runx2 expression in osteoblasts. J Biol Chem 279(38):40007–40016. doi: 10.1074/jbc.M403715200 CrossRefPubMedGoogle Scholar
  39. 39.
    Wu M, Hesse E, Morvan F, Zhang JP, Correa D, Rowe GC, Kiviranta R, Neff L, Philbrick WM, Horne WC, Baron R (2009) Zfp521 antagonizes Runx2, delays osteoblast differentiation in vitro, and promotes bone formation in vivo. Bone 44(4):528–536. doi: 10.1016/j.bone.2008.11.011 CrossRefPubMedGoogle Scholar
  40. 40.
    Boopathy R, Layer PG (2004) Aryl acylamidase activity on acetylcholinesterase is high during early chicken brain development. Protein J 23(5):325–333. doi: 10.1023/b:jopc.0000032652.99257.19 CrossRefPubMedGoogle Scholar
  41. 41.
    Dai SQ, Yu LP, Shi X, Wu H, Shao P, Yin GY, Wei YZ (2014) Serotonin regulates osteoblast proliferation and function in vitro. Braz J Med Biol Res 47(9):759–765CrossRefPubMedPubMedCentralGoogle Scholar
  42. 42.
    Spieker J, Ackermann A, Salfelder A, Vogel-Hopker A, Layer PG (2016) Acetylcholinesterase regulates skeletal in ovo development of chicken limbs by ACh-dependent and -independent mechanisms. PLoS ONE 11(8):e0161675. doi: 10.1371/journal.pone.0161675 CrossRefPubMedPubMedCentralGoogle Scholar

Copyright information

© Springer Science+Business Media, LLC 2017

Authors and Affiliations

  • Raj Kumar Chinnadurai
    • 1
    • 2
  • Ponne Saravanaraman
    • 1
    • 3
  • Rathanam Boopathy
    • 1
  1. 1.Department of BiotechnologyBharathiar UniversityCoimbatoreIndia
  2. 2.Centre for Animal Research, Training and Services (CAReTS), Central Inter-Disciplinary Research Facility (CIDRF)Mahatma Gandhi Medical College and Research Institute (MGMCRI) CampusPuducherryIndia
  3. 3.Department of BiotechnologyPondicherry Central UniversityPuducherryIndia

Personalised recommendations