title: Lipopolysaccharide enhances Wnt5a expression through toll-like receptor 4, myeloid differentiating factor 88, phosphatidylinositol 3-OH kinase/AKT and nuclear factor kappa B pathways in human dental pulp stem cells.
abstract: BACKGROUND
  Lipopolysaccharide (LPS) has been implicated in mesenchymal stem cell differentiation processes. Wnt5a, one of the "non-canonical" Wnt family members, is important in signaling stem cell differentiation and in the inflammatory responses of immune cells. Here we studied whether LPS can regulate the expression of Wnt5a in human dental pulp stem cells (hDPSCs) and investigated the intracellular signaling pathways activated by LPS.
  
  
  METHODS
  Wnt5a mRNA and protein expression changes in hDPSCs were investigated by real-time polymerase chain reaction analysis and enzyme-linked immunosorbent assay. In addition, real-time polymerase chain reaction, enzyme-linked immunosorbent assay, and luciferase activity assays were used to determine whether toll-like receptor 4 (TLR4), myeloid differentiating factor 88 (MyD88), nuclear factor kappa B (NF-kB), or the phosphatidylinositol 3-OH kinase (PI3K)/AKT pathways are involved in LPS-induced Wnt5a expression. The activation of PI3K and AKT in hDPSCs was measured by Western blot analysis.
  
  
  RESULTS
  Wnt5a mRNA and protein expression was rapidly increased in response to LPS in a time- and dose-dependent manner. LPS-induced Wnt5a expression was effectively attenuated by administration of a TLR4 neutralizing antibody, MyD88 inhibitory peptide, PI3-kinase inhibitors (LY294002 and wortmannin), an AKT inhibitor, or NF-κB inhibitor (pyrrolidine dithiocarbamate), IκBa phosphorylation inhibitor (Bay 117082), or IκB protease inhibitor (L-1-tosylamido-2-phenylethyl chloromethyl ketone). Treatment of hDPSCs with LPS activated PI3-kinase (p85) and AKT signaling in a time-dependent manner. Moreover, LPS-mediated increases in κB-luciferase activity were diminished by the overexpression of dominant negative mutants of TLR4, MyD88, p85, AKT, and IκBa.
  
  
  CONCLUSIONS
  These results demonstrated that LPS-induced Wnt5a expression was mediated through the TLR4/MyD88/PI3-kinase/AKT pathway, which then initiated NF-κB activation in hDPSCs.
authors: Department of Operative Dentistry, School of Dentistry, The Fourth Military Medical University, Xi'an, PR China. Electronic address: hewenxi@fmmu.edu.cn.
authors: Wenxi
authors: W
authors: He
authors: Department of Operative Dentistry, School of Dentistry, The Fourth Military Medical University, Xi'an, PR China.
authors: Zhihua
authors: Z
authors: Wang
authors: Department of Operative Dentistry, School of Dentistry, The Fourth Military Medical University, Xi'an, PR China.
authors: Zeyuan
authors: Z
authors: Zhou
authors: Department of Operative Dentistry, School of Dentistry, The Fourth Military Medical University, Xi'an, PR China.
authors: Yaqing
authors: Y
authors: Zhang
authors: Department of Operative Dentistry, School of Dentistry, The Fourth Military Medical University, Xi'an, PR China.
authors: Qinglin
authors: Q
authors: Zhu
authors: Department of Operative Dentistry, School of Dentistry, The Fourth Military Medical University, Xi'an, PR China.
authors: Kewen
authors: K
authors: Wei
authors: Department of Operative Dentistry, School of Dentistry, The Fourth Military Medical University, Xi'an, PR China.
authors: Yuan
authors: Y
authors: Lin
authors: Department of Oral Biology, School of Dentistry, University of Birmingham, Birmingham, United Kingdom.
authors: Paul R
authors: PR
authors: Cooper
authors: Department of Oral Biology, School of Dentistry, University of Birmingham, Birmingham, United Kingdom.
authors: Anthony J
authors: AJ
authors: Smith
authors: Department of Operative Dentistry, School of Dentistry, The Fourth Military Medical University, Xi'an, PR China. Electronic address: hefmmu@aliyun.com.
authors: Qing
authors: Q
authors: Yu
mesh_terms: D000293:Adolescent; D000730:Androstadienes / Q000494:pharmacology; D018929:Cell Culture Techniques; D002478:Cells, Cultured; D002867:Chromones / Q000494:pharmacology; D003782:Dental Pulp / Q000166:cytology*; D003979:Diazonium Compounds / Q000494:pharmacology; D004305:Dose-Response Relationship, Drug; D004926:Escherichia coli; D006801:Humans; D020672:I-kappa B Proteins / Q000037:antagonists & inhibitors; D008070:Lipopolysaccharides / Q000494:pharmacology*; D009025:Morpholines / Q000494:pharmacology; D009183:Mycotoxins / Q000494:pharmacology; D053594:Myeloid Differentiation Factor 88 / Q000187:drug effects*; D000072000:NF-KappaB Inhibitor alpha; D016328:NF-kappa B / Q000037:antagonists & inhibitors / Q000187:drug effects*; D009570:Nitriles / Q000494:pharmacology; D019869:Phosphatidylinositol 3-Kinases / Q000187:drug effects*; D000081082:Phosphoinositide-3 Kinase Inhibitors; D011518:Proto-Oncogene Proteins / Q000187:drug effects*; D051057:Proto-Oncogene Proteins c-akt / Q000187:drug effects*; D011759:Pyrrolidines / Q000494:pharmacology; D015398:Signal Transduction / Q000187:drug effects; D013234:Stem Cells / Q000187:drug effects*; D013450:Sulfones / Q000494:pharmacology; D013859:Thiocarbamates / Q000494:pharmacology; D013997:Time Factors; D051197:Toll-Like Receptor 4 / Q000187:drug effects*; D051153:Wnt Proteins / Q000187:drug effects*; D060449:Wnt Signaling Pathway / Q000187:drug effects*; D000071818:Wnt-5a Protein; D000077191:Wortmannin; D055815:Young Adult; D000096926:Benzenesulfonamides
chemical_list: D000730:Androstadienes; D002867:Chromones; D003979:Diazonium Compounds; D020672:I-kappa B Proteins; D008070:Lipopolysaccharides; D009025:Morpholines; D009183:Mycotoxins; D053594:Myeloid Differentiation Factor 88; D000072000:NF-KappaB Inhibitor alpha; D016328:NF-kappa B; D009570:Nitriles; D019869:Phosphatidylinositol 3-Kinases; D000081082:Phosphoinositide-3 Kinase Inhibitors; D011518:Proto-Oncogene Proteins; D051057:Proto-Oncogene Proteins c-akt; D011759:Pyrrolidines; D013450:Sulfones; D013859:Thiocarbamates; D051197:Toll-Like Receptor 4; D051153:Wnt Proteins; D000071818:Wnt-5a Protein; D000077191:Wortmannin; C434003:3-(4-methylphenylsulfonyl)-2-propenenitrile; C507779:MYD88 protein, human; C000606204:NFKBIA protein, human; C495345:TLR4 protein, human; C085389:WNT5A protein, human; C009506:1-tosylamido-2-phenylethyldiazomethylketone; D000096926:Benzenesulfonamides; C020972:pyrrolidine dithiocarbamic acid; C085911:2-(4-morpholinyl)-8-phenyl-4H-1-benzopyran-4-one
keywords: Dental pulp stem cells; LPS; NF-kappaB; PI3K/AKT; TLR4; Wnt5a