title: The role of phospholipase D in osteoblast response to titanium surface microstructure.
abstract: Biomaterial surface properties such as microtopography and energy can change cellular responses at the cell-implant interface. Phospholipase D (PLD) is required for the differentiation of osteoblast-like MG63 cells on machined and grit-blasted titanium surfaces. Here, we determined if PLD is also required on microstructured/high-energy substrates and the mechanism involved. shRNAs for human PLD1 and PLD2 were used to silence MG63 cells. Wild-type and PLD1 or PLD1/2 silenced cells were cultured on smooth-pretreatment surfaces (PT); grit-blasted, acid-etched surfaces (SLA); and SLA surfaces modified to have higher surface energy (modSLA). PLD was inhibited with ethanol or activated with 24,25-dihydroxyvitamin-D(3) [24R,25(OH)(2)D(3)]. As surface roughness/energy increased, PLD mRNA and activity increased, cell number decreased, osteocalcin and osteoprotegerin increased, and protein kinase C (PKC) and alkaline phosphatase specific activities increased. Ethanol inhibited PLD and reduced surface effects on these parameters. There was no effect on these parameters after knockdown of PLD1, but PLD1/2 double knockdown had effects comparableto PLD inhibition. 24R,25(OH)(2)D(3) increased PLD activity and the production of osteocalcin and osteoprotegerin, but decreased cell number on the rough/high-energy surfaces. These results confirm that surface roughness/energy-induced PLD activity is required for osteoblast differentiation and that PLD2 is the main isoform involved in this pathway. PLD is activated by 24R,25(OH)(2)D(3) in a surface-dependent manner and inhibition of PLD reduces the effects of surface microstructure/energy on PKC, suggesting that PLD mediates the stimulatory effect of microstructured/high-energy surfaces via PKC-dependent signaling.
authors.affiliation: Department of Biomedical Engineering, Georgia Institute of Technology, Atlanta, Georgia, USA.
authors.forename: Mimi
authors.initials: M
authors.lastname: Fang
authors.forename: Rene
authors.initials: R
authors.lastname: Olivares-Navarrete
authors.forename: Marco
authors.initials: M
authors.lastname: Wieland
authors.forename: David L
authors.initials: DL
authors.lastname: Cochran
authors.forename: Barbara D
authors.initials: BD
authors.lastname: Boyan
authors.forename: Zvi
authors.initials: Z
authors.lastname: Schwartz
mesh_terms: D001483:Base Sequence; D002460:Cell Line; D002999:Clone Cells; D004789:Enzyme Activation / Q000187:drug effects; D055785:Gene Knockdown Techniques; D020868:Gene Silencing / Q000187:drug effects; D006801:Humans; D007527:Isoenzymes / Q000378:metabolism; D008969:Molecular Sequence Data; D010006:Osteoblasts / Q000187:drug effects* / Q000201:enzymology*; D010739:Phospholipase D / Q000037:antagonists & inhibitors / Q000378:metabolism*; D011493:Protein Kinase C / Q000378:metabolism; D034741:RNA, Small Interfering / Q000235:genetics / Q000378:metabolism; D013379:Substrate Specificity / Q000187:drug effects; D013499:Surface Properties / Q000187:drug effects; D014025:Titanium / Q000737:chemistry* / Q000494:pharmacology*; D014807:Vitamin D / Q000031:analogs & derivatives / Q000378:metabolism
chemical_list: D007527:Isoenzymes; D034741:RNA, Small Interfering; C118756:dihydroxy-vitamin D3; D014807:Vitamin D; D014025:Titanium; D011493:Protein Kinase C; D010739:Phospholipase D
references.citation: Biochem Biophys Res Commun. 2006 Oct 6;348(4):1398-402
references.pmid: 16919239
references.citation: Clin Oral Implants Res. 2004 Dec;15(6):683-92
references.pmid: 15533129
references.citation: J Biomed Mater Res. 1999 Nov;47(2):139-51
references.pmid: 10449625
references.citation: Steroids. 2001 Sep;66(9):683-94
references.pmid: 11546556
references.citation: J Cell Biochem. 2009 May 1;107(1):155-62
references.pmid: 19288498
references.citation: Crit Rev Oral Biol Med. 1996;7(4):329-45
references.pmid: 8986395
references.citation: J Biomed Mater Res A. 2005 Jul 1;74(1):49-58
references.pmid: 15924300
references.citation: J Biomed Mater Res. 1995 Mar;29(3):389-401
references.pmid: 7542245
references.citation: Connect Tissue Res. 2003;44 Suppl 1:130-5
references.pmid: 12952186
references.citation: Steroids. 2001 Mar-May;66(3-5):363-74
references.pmid: 11179745
references.citation: Biomaterials. 2004 Aug;25(19):4759-66
references.pmid: 15120522
references.citation: Biomaterials. 1999 Dec;20(23-24):2311-21
references.pmid: 10614937
references.citation: J Biol Chem. 1995 Aug 18;270(33):19465-72
references.pmid: 7642630
references.citation: J Biomed Mater Res A. 2006 Feb;76(2):323-34
references.pmid: 16270344
references.citation: Biochim Biophys Acta. 2001 Jan 15;1499(3):209-21
references.pmid: 11341968
references.citation: Clin Oral Implants Res. 2000;11 Suppl 1:33-58
references.pmid: 11168256
references.citation: J Biomed Mater Res A. 2004 Sep 1;70(3):361-9
references.pmid: 15293309
references.citation: Biomaterials. 2006 Nov;27(32):5502-11
references.pmid: 16857255
references.citation: Cell Mol Life Sci. 2005 Jul;62(13):1448-61
references.pmid: 15924269
references.citation: J Bone Joint Surg Am. 2008 Nov;90(11):2485-98
references.pmid: 18978418
references.citation: Neuroscience. 2007 May 25;146(3):946-61
references.pmid: 17434680
references.citation: Biochim Biophys Acta. 1999 Jul 30;1439(2):229-44
references.pmid: 10425398
references.citation: Biomaterials. 2008 Oct;29(28):3822-35
references.pmid: 18617258
references.citation: Biomaterials. 2001 Apr;22(7):731-41
references.pmid: 11246968
references.citation: J Biol Chem. 1997 Feb 7;272(6):3860-8
references.pmid: 9013646
references.citation: Biochem J. 1996 Oct 1;319 ( Pt 1):285-91
references.pmid: 8870680
references.citation: J Biomed Mater Res A. 2004 Nov 1;71(2):217-25
references.pmid: 15386491
references.citation: J Biomed Mater Res. 1998 Jan;39(1):77-85
references.pmid: 9429099
references.citation: Biochim Biophys Acta. 1991 Jul 9;1084(2):155-8
references.pmid: 1854800
references.citation: J Biol Chem. 1995 Jun 23;270(25):14935-43
references.pmid: 7797473
references.citation: J Cell Biochem. 2006 Jun 1;98(3):590-617
references.pmid: 16440327
references.citation: J Biomed Mater Res A. 2006 Dec 15;79(4):1023-32
references.pmid: 17034031
references.citation: Biomaterials. 2006 Jul;27(20):3716-25
references.pmid: 16569430
references.citation: Cell Signal. 2005 Nov;17(11):1423-32
references.pmid: 15951158
references.citation: Clin Oral Implants Res. 2005 Dec;16(6):650-6
references.pmid: 16307571
references.citation: Chem Phys Lipids. 1999 Apr;98(1-2):127-40
references.pmid: 10358935
references.citation: Biomaterials. 1998 Dec;19(23):2219-32
references.pmid: 9884063
references.citation: FASEB J. 1998 Oct;12(13):1309-17
references.pmid: 9761774
references.citation: J Biomed Mater Res. 1991 Jul;25(7):889-902
references.pmid: 1918105
references.citation: Biochem Biophys Res Commun. 2005 Feb 25;327(4):1047-51
references.pmid: 15652502
references.citation: Crit Rev Oral Biol Med. 2002;13(2):143-54
references.pmid: 12097357
references.citation: J Leukoc Biol. 2005 Jun;77(6):999-1007
references.pmid: 15774548
references.citation: J Biomed Mater Res. 1996 Sep;32(1):55-63
references.pmid: 8864873
references.citation: FEBS Lett. 1994 Apr 4;342(2):149-53
references.pmid: 8143868
references.citation: J Biol Chem. 1995 Dec 15;270(50):29640-3
references.pmid: 8530346
references.citation: Biochim Biophys Acta. 1999 Jul 30;1439(2):135-49
references.pmid: 10425391
references.citation: Adv Dent Res. 1999 Jun;13:38-48
references.pmid: 11276745
references.citation: Biochem J. 2005 Jul 1;389(Pt 1):207-14
references.pmid: 15769249
references.citation: Biochem J. 2000 Feb 1;345 Pt 3:401-15
references.pmid: 10642495
references.citation: Nature. 1984 Apr 19-25;308(5961):693-8
references.pmid: 6232463
references.citation: Curr Biol. 1997 Mar 1;7(3):191-201
references.pmid: 9395408
references.citation: J Biomed Mater Res. 2001 Jun 5;55(3):350-9
references.pmid: 11255188
references.citation: Biomaterials. 2007 Jun;28(18):2821-9
references.pmid: 17368532
references.citation: Clin Oral Implants Res. 1997 Dec;8(6):442-7
references.pmid: 9555202
references.citation: Proc Natl Acad Sci U S A. 2008 Oct 14;105(41):15767-72
references.pmid: 18843104
references.citation: Implant Dent. 2003;12(2):175-81
references.pmid: 12861887