title: Blockade of NF-κB and MAPK pathways by ulinastatin attenuates wear particle-stimulated osteoclast differentiation in vitro and in vivo.
abstract: Ulinastatin, a urinary trypsin inhibitor (UTI), is widely used to clinically treat lipopolysaccharide (LPS)-related inflammatory disorders recently. Adherent pathogen-associated molecular patterns (PAMPs), of which LPS is the best-studied and classical endotoxin produced by Gram-negative bacteria, act to increase the biological activity of osteopedic wear particles such as polymethyl-methacrylate (PMMA) and titanium particles in cell culture and animal models of implant loosening. The present study was designed to explore the inhibitory effect of UTI on osteoclastogenesis and inflammatory osteolysis in LPS/PMMA-mediated Raw264.7 cells and murine osteolysis models, and investigate the potential mechanism. The in vitro study was divided into the control group, LPS-induced group, PMMA-stimulated group and UTI-pretreated group. UTI (500 or 5000 units/ml) pretreatment was followed by PMMA (0.5 mg/ml) with adherent LPS. The levels of inflammatory mediators including tumour necrosis factor-α (TNF-α), matrixmetallo-proteinases-9 (MMP-9) and interleukin-6 (IL-6), receptor activation of nuclear factor NF-κB (RANK), and cathepsin K were examined and the amounts of phosphorylated I-κB, MEK, JNK and p38 were measured. In vivo study, murine osteolysis models were divided into the control group, PMMA-induced group and UTI-treated group. UTI (500 or 5000 units/kg per day) was injected intraperitoneally followed by PMMA suspension with adherent LPS (2×108particles/25 μl) in the UTI-treated group. The thickness of interfacial membrane and the number of infiltrated inflammatory cells around the implants were assessed, and bone mineral density (BMD), trabecular number (Tb.N.), trabecular thickness (Tb.Th.), trabecular separation (Tb.Sp.), relative bone volume over total volume (BV/TV) of distal femur around the implants were calculated. Our results showed that UTI pretreatment suppressed the secretion of proinflammatory cytokines including MMP-9, IL-6, TNF-α, RANK and cathepsin K through down-regulating the activity of nuclear factor kappa B (NF-κB) and MAPKs partly in LPS/PMMA-mediated Raw264.7 cells. Finally, UTI treatment decreased the inflammatory osteolysis reaction in PMMA-induced murine osteolysis models. In conclusion, these results confirm the anti-inflammatory potential of UTI in the prevention of particle disease.
authors.affiliation: Department of Orthopedics, The First People's Hospital of Yangzhou City, Second Clinical School of Yangzhou University, Yangzhou 225000, China.
authors.forename: Jiang-Ying
authors.initials: JY
authors.lastname: Ru
authors.affiliation: Department of Orthopedics, Jinling Hospital, School of Medicine, Nanjing University, Nanjing 210002, China.
authors.forename: Hai-Dong
authors.initials: HD
authors.lastname: Xu
authors.affiliation: Department of Orthopedics, The First People's Hospital of Yangzhou City, Second Clinical School of Yangzhou University, Yangzhou 225000, China.
authors.forename: Dai
authors.initials: D
authors.lastname: Shi
authors.affiliation: Department of Orthopedics, The First People's Hospital of Yangzhou City, Second Clinical School of Yangzhou University, Yangzhou 225000, China.
authors.forename: Jun-Bo
authors.initials: JB
authors.lastname: Pan
authors.affiliation: Department of Orthopedics, The First People's Hospital of Yangzhou City, Second Clinical School of Yangzhou University, Yangzhou 225000, China.
authors.forename: Xiao-Jin
authors.initials: XJ
authors.lastname: Pan
authors.affiliation: Department of Pathology, The First People's Hospital of Yangzhou City, Second Clinical School of Yangzhou University, Yangzhou 225000, China wangyanfen315@163.com.
authors.forename: Yan-Fen
authors.initials: YF
authors.lastname: Wang
mesh_terms: D000818:Animals; D002454:Cell Differentiation / Q000187:drug effects; D006023:Glycoproteins / Q000008:administration & dosage*; D007249:Inflammation / Q000139:chemically induced / Q000188:drug therapy* / Q000473:pathology; D008070:Lipopolysaccharides / Q000633:toxicity; D051379:Mice; D020929:Mitogen-Activated Protein Kinase Kinases / Q000096:biosynthesis; D016328:NF-kappa B / Q000096:biosynthesis; D010010:Osteoclasts / Q000187:drug effects*; D010012:Osteogenesis / Q000187:drug effects*; D010014:Osteolysis / Q000139:chemically induced / Q000188:drug therapy* / Q000473:pathology; D000069452:Pathogen-Associated Molecular Pattern Molecules; D000067996:RAW 264.7 Cells; D015398:Signal Transduction / Q000187:drug effects; D014409:Tumor Necrosis Factor-alpha / Q000096:biosynthesis
chemical_list: D006023:Glycoproteins; D008070:Lipopolysaccharides; D016328:NF-kappa B; D000069452:Pathogen-Associated Molecular Pattern Molecules; D014409:Tumor Necrosis Factor-alpha; D020929:Mitogen-Activated Protein Kinase Kinases; C028665:urinastatin
keywords: aseptic loosening (AL); lipopolysaccharide (LPS); mitogen activated protein kinase (MAPK); nuclear factor kappa B (NF-κB); osteoclast; osteolysis; polymethyl-methacrylate (PMMA); ulinastatin