OBJECTIVE: To evaluate and compare protein expression profiles of synovial fibroblasts using proteome analysis in swine knee injuries with joint instability, during early post-traumatic arthritis (PTA) development. METHOD: Experimental PTA was induced by transection of the anterior cruciate ligament (ACL) in swine left knee joints. After sacrifice at 8 weeks, cartilage and synovium obtained from both knee joints were prepared for histopathologic examination. Cultured synovial fibroblasts were processed for 2-dimensional electrophoresis and mass spectrometric analysis. Histopathologic examination showed overt arthritic changes that supported the development of early PTA. RESULTS: Proteome analyses led to the identification of more than 1,500 protein spots and of 11 differently expressed protein spots. Of those, six proteins were down-regulated (cytoskeletal beta actin, cofilin-1, destrin, Rho GDP dissociation inhibitor alpha, and unnamed protein product), and five proteins were up-regulated (alpha-B crystallin, smooth muscle protein 22-alpha, and cytoskeletal beta actin) in ACL-transected synovial fibroblasts. That is, proteins related to cellular organization and signal transduction are down-regulated, and those related to cell rescue, defence, and stress are up-regulated. CONCLUSION: These results may suggest that joint instability contributes to the development of PTA and is one of the major etiologic factors of PTA. In addition, this suggests that the proteome analysis of synovial fibroblasts is a useful approach in examining a joint after an injury and can be used to understand the pathogenesis of PTA.
Actins ; Anterior Cruciate Ligament ; Arthritis* ; Cartilage ; Crystallins ; Destrin ; Electrophoresis ; Fibroblasts* ; Guanine Nucleotide Dissociation Inhibitors ; Joint Instability ; Joints ; Knee Injuries ; Knee Joint ; Muscle, Smooth ; Proteome ; Signal Transduction ; Swine ; Synovial Membrane

OBJECTIVETo identify two differentiation-associated proteins induced by rhIL-6 in M1 mouse myeloid leukemia cells.

METHODSProtein spots were excised from 2-D gels and digested in-gel with trypsin. The trypsin lysis products were first analyzed by matrix-assisted laser desorption/ionization-time of flight-mass spectrometry (MALDI-TOF-MS) through peptide mass fingerprinting and then performed peptide sequencing by nano-electrospray ionization mass spectrometry/mass spectrometry (nano-ESI-MS/MS). The database search was finished with the Mascot search engine (http://www.matrixscience.co.uk) using the data processed through MaxEnt3 and MasSeq.

RESULTSThe two proteins were not revealed by peptide mass fingerprint using MALDI-TOF-MS, while they were respectively identified as Destrin and Putative protein after the sequence of their trypic peptides were obtained by the nano-ESI-MS/MS techniques.

CONCLUSIONNano-ESI-MS/MS technique can successfully identify the two differentiation-associated proteins induced by rhIL-6 and has great advantage in protein analysis.

Actin Depolymerizing Factors ; Amino Acid Sequence ; Animals ; Apoptosis ; Cell Transformation, Neoplastic ; drug effects ; Destrin ; Interleukin-6 ; analysis ; pharmacology ; Leukemia, Myeloid, Acute ; metabolism ; pathology ; Mice ; Microfilament Proteins ; analysis ; Molecular Sequence Data ; Nanotechnology ; Recombinant Proteins ; pharmacology ; Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization ; methods ; Tumor Suppressor Proteins ; analysis