title: PEP-1-GRX-1 Modulates Matrix Metalloproteinase-13 and Nitric Oxide Expression of Human Articular Chondrocytes.
abstract: BACKGROUND
  The protein transduction domain (PTD) enables therapeutic proteins to directly penetrate the membranes of cells and tissues, and has been increasingly utilized. Glutaredoxin-1 (GRX-1) is an endogenous antioxidant enzyme involved in the cellular redox homeostasis system. In this study, we investigated whether PEP-1-GRX-1, a fusion protein of GRX-1 and PEP-1 peptide, a PTD, could suppress catabolic responses in primary human articular chondrocytes and a mouse carrageenan-induced paw edema model.
  
  
  METHODS
  Human articular chondrocytes were isolated enzymatically from articular cartilage and cultured in a monolayer. The transduction efficiency of PEP-1-GRX-1 into articular chondrocytes was measured by western blot and immunohistochemistry. The effects of PEP-1-GRX-1 on matrix metalloproteinases (MMPs) and catabolic factor expression in interleukin (IL)-1β- and lipopolysaccharide (LPS)-treated chondrocytes were analyzed by real-time quantitative reverse transcription-polymerase chain reaction and western blot. The effect of PEP-1-GRX1 on the mitogen-activated protein kinase (MAPK) and nuclear factor kappa-light chain-enhancer of activated B cells (NF-κB) signaling pathway were also analyzed by western blot. Finally, the inhibitory effect of PEP-1-GRX-1 on MMP-13 production was measured in vivo in a mouse carrageenan-induced paw edema model.
  
  
  RESULTS
  PEP-1-GRX-1 significantly penetrated into human chondrocytes and mouse cartilage, whereas GRX-1 did not. PEP-1-GRX-1 significantly suppressed MMP-13 expression and nitric oxide (NO) production in LPS-stimulated chondrocytes, and NO production in IL-1β-stimulated chondrocytes, compared with GRX-1. In addition, PEP-1-GRX-1 decreased IL-1β- and LPS-induced activation of MAPK and NF-κB. In the mouse model of carrageenan-induced paw edema, PEP-1-GRX-1 significantly suppressed carrageenan-induced MMP-13 production as well as paw edema.
  
  
  CONCLUSIONS
  These results demonstrate that PEP-1-GRX-1 can be transduced efficiently in vitro and in vivo into human chondrocytes and mouse cartilage tissue and downregulate catabolic responses in chondrocytes by inhibiting the MAPK and NF-κB pathway. PEP-1-GRX-1 thus has the potential to reduce catabolic responses in chondrocytes and cartilage.
authors.forename: Hyun Sook
authors.initials: HS
authors.lastname: Hwang
authors.forename: In Young
authors.initials: IY
authors.lastname: Park
authors.forename: Hyun Ah
authors.initials: HA
authors.lastname: Kim
authors.forename: Soo Young
authors.initials: SY
authors.lastname: Choi
mesh_terms: D000818:Animals; D002351:Carrageenan / Q000633:toxicity; D002358:Cartilage, Articular / Q000166:cytology / Q000187:drug effects / Q000378:metabolism*; D002478:Cells, Cultured; D003543:Cysteamine / Q000031:analogs & derivatives / Q000378:metabolism; D004195:Disease Models, Animal; D015536:Down-Regulation / Q000187:drug effects; D004487:Edema / Q000139:chemically induced / Q000378:metabolism / Q000473:pathology; D054477:Glutaredoxins / Q000235:genetics / Q000378:metabolism; D006801:Humans; D007150:Immunohistochemistry; D053583:Interleukin-1beta / Q000494:pharmacology; D008070:Lipopolysaccharides / Q000633:toxicity; D008297:Male; D053509:Matrix Metalloproteinase 13 / Q000235:genetics / Q000378:metabolism*; D051379:Mice; D008810:Mice, Inbred C57BL; D020928:Mitogen-Activated Protein Kinases / Q000378:metabolism; D016328:NF-kappa B / Q000378:metabolism; D009569:Nitric Oxide / Q000378:metabolism*; D010455:Peptides / Q000235:genetics / Q000378:metabolism; D011993:Recombinant Fusion Proteins / Q000096:biosynthesis / Q000302:isolation & purification / Q000494:pharmacology; D015398:Signal Transduction / Q000187:drug effects
chemical_list: D054477:Glutaredoxins; D053583:Interleukin-1beta; D008070:Lipopolysaccharides; D016328:NF-kappa B; C503924:Pep-1 peptide; D010455:Peptides; D011993:Recombinant Fusion Proteins; D009569:Nitric Oxide; D003543:Cysteamine; D002351:Carrageenan; D020928:Mitogen-Activated Protein Kinases; D053509:Matrix Metalloproteinase 13
keywords: Chondrocyte; Glutaredoxin-1; Matrix metalloproteinase; Osteoarthritis.; Protein transduction domain