title: Lubricin/Proteoglycan 4 Binding to CD44 Receptor: A Mechanism of the Suppression of Proinflammatory Cytokine-Induced Synoviocyte Proliferation by Lubricin.
abstract: OBJECTIVE
  To evaluate the binding of recombinant human proteoglycan 4 (rhPRG4) to CD44 receptor and its consequences on cytokine-induced synoviocyte proliferation.
  
  
  METHODS
  The binding of rhPRG4 to CD44 and competition with high molecular weight (HMW) hyaluronic acid (HA) was evaluated using a direct enzyme-linked immunosorbent assay (ELISA) and surface plasmon resonance. Sialidase A and O-glycosidase digestion of rhPRG4 was performed, and CD44 binding was evaluated using ELISA. Rheumatoid arthritis (RA) fibroblast-like synoviocytes (FLS) were stimulated with interleukin-1β (IL-1β) or tumor necrosis factor α (TNFα) for 48 hours in the presence or absence of rhPRG4 or HMW HA at 20, 40, and 80 μg/ml, and cell proliferation was measured. The contribution of CD44 was assessed by coincubation with a CD44 antibody (IM7). The antiproliferative effect of rhPRG4 was investigated following treatment of PRG4(-/-) mouse synoviocytes with IL-1β or TNFα in the presence or absence of IM7.
  
  
  RESULTS
  Recombinant human PRG4 bound CD44 and interfered with the binding of HMW HA to CD44. Removal of sialic acid and O-glycosylations significantly increased CD44 binding by rhPRG4 (P < 0.001). Both rhPRG4 and HMW HA at 40 and 80 μg/ml significantly suppressed IL-1β-induced proliferation of RA FLS (P < 0.05). Recombinant human PRG4 at 20, 40, and 80 μg/ml significantly suppressed TNFα-induced RA FLS proliferation (P < 0.05). CD44 neutralization reversed the effect of rhPRG4 on IL-1β- and TNFα-stimulated RA FLS and the effect of HMW HA on IL-1β-stimulated RA FLS. Recombinant human PRG4 inhibited cytokine-induced proliferation of PRG4(-/-) synoviocytes, which could be prevented by blocking CD44.
  
  
  CONCLUSIONS
  PRG4 (lubricin) is a novel putative ligand for CD44 and may control synoviocyte overgrowth in inflammatory arthropathies via a CD44-mediated mechanism.
authors.affiliation: Massachusetts College of Pharmacy and Health Sciences University, Boston.
authors.forename: Afnan
authors.initials: A
authors.lastname: Al-Sharif
authors.affiliation: Massachusetts College of Pharmacy and Health Sciences University, Boston.
authors.forename: Maha
authors.initials: M
authors.lastname: Jamal
authors.affiliation: Rhode Island Hospital, Providence.
authors.forename: Ling X
authors.initials: LX
authors.lastname: Zhang
authors.affiliation: Brown University, Providence, Rhode Island.
authors.forename: Katherine
authors.initials: K
authors.lastname: Larson
authors.affiliation: University of Calgary, Calgary, Alberta, Canada.
authors.forename: Tannin A
authors.initials: TA
authors.lastname: Schmidt
authors.affiliation: Rhode Island Hospital and Brown University, Providence, Rhode Island.
authors.forename: Gregory D
authors.initials: GD
authors.lastname: Jay
authors.affiliation: Massachusetts College of Pharmacy and Health Sciences University, Boston.
authors.forename: Khaled A
authors.initials: KA
authors.lastname: Elsaid
mesh_terms: D000818:Animals; D001172:Arthritis, Rheumatoid / Q000378:metabolism*; D049109:Cell Proliferation / Q000187:drug effects / Q000502:physiology*; D004797:Enzyme-Linked Immunosorbent Assay; D005347:Fibroblasts / Q000187:drug effects / Q000378:metabolism*; D006801:Humans; D018960:Hyaluronan Receptors / Q000378:metabolism*; D006820:Hyaluronic Acid / Q000378:metabolism*; D053583:Interleukin-1beta / Q000494:pharmacology; D051379:Mice; D018345:Mice, Knockout; D011509:Proteoglycans / Q000235:genetics* / Q000378:metabolism*; D020349:Surface Plasmon Resonance; D013583:Synovial Membrane / Q000166:cytology*; D014409:Tumor Necrosis Factor-alpha / Q000494:pharmacology; D000099158:Proteoglycan 4
chemical_list: D018960:Hyaluronan Receptors; D006820:Hyaluronic Acid; D053583:Interleukin-1beta; D011509:Proteoglycans; D014409:Tumor Necrosis Factor-alpha; C497483:CD44 protein, human; C476399:Cd44 protein, mouse; C419197:PRG4 protein, human; D000099158:Proteoglycan 4; C498837:Prg4 protein, mouse