Intra-articular injection of interleukin-4 decreases nitric oxide production by chondrocytes and ameliorates subsequent destruction of cartilage in instability-induced osteoarthritis in rat knee joints.
OBJECTIVE
To investigate the in vitro and in vivo effects of interleukin (IL)-4 on mechanical stress-induced nitric oxide (NO) expression by chondrocytes, and destruction of cartilage and NO production in an instability-induced osteoarthritis (OA) model in rat knee joints, respectively.


METHODS
Cyclic tensile stress (CTS; 0.5Hz and 7% elongation) was applied to cultured normal rat chondrocytes with or without pre-incubation with recombinant rat IL-4 (rrIL-4). Inducible NO synthase (iNOS) mRNA expression and NO production were examined with real-time polymerase chain reaction and the Griess reaction, respectively. OA was induced in rat knee joints by transection of the anterior cruciate and medial collateral ligaments and resection of the medial meniscus. rrIL-4 (10, 50, and 100 ng/joint/day) was injected intra-articularly, and knee joint samples were collected 2, 4, and 6 weeks after surgery. Cartilage destruction was evaluated by the modified Mankin score and Osteoarthritis Research Society International scoring system on paraffin-embedded sections stained with safranin O. Cleavage of aggrecan and NO production were examined by immunohistochemistry for aggrecan neoepitope (NITEGE) and of nitrotyrosine (NT), respectively.


RESULTS
rrIL-4 down-regulated CTS-induced iNOS mRNA expression and NO production by chondrocytes. The intra-articular injection of rrIL-4 gave rise to a limited, but significant amelioration of cartilage destruction, prevention of loss of aggrecan, and decrease in the number of NT-positive chondrocytes, an effect that was not dose-dependent.


CONCLUSIONS
The present study suggests that IL-4 may exert chondroprotective properties against mechanical stress-induced cartilage destruction, at least in part, by inhibiting NO production by chondrocytes.
Yorimitsu
M
M
Department of Orthopaedic Surgery, Okayama University Graduate School of Medicine, Dentistry and Pharmaceutical Sciences, 2-5-1 Shikata-cho, Okayama City, Okayama 700-8558, Japan.
Nishida
K
K
Shimizu
A
A
Doi
H
H
Miyazawa
S
S
Komiyama
T
T
Nasu
Y
Y
Yoshida
A
A
Watanabe
S
S
Ozaki
T
T
D053674:Aggrecans / Q000378:metabolism; D000818:Animals; D001169:Arthritis, Experimental / Q000378:metabolism / Q000473:pathology / Q000517:prevention & control*; D002478:Cells, Cultured; D019902:Chondrocytes / Q000187:drug effects* / Q000378:metabolism; D004305:Dose-Response Relationship, Drug; D005269:Femur / Q000473:pathology; D015971:Gene Expression Regulation, Enzymologic / Q000187:drug effects; D007270:Injections, Intra-Articular; D015847:Interleukin-4 / Q000494:pharmacology* / Q000627:therapeutic use; D040542:Mechanotransduction, Cellular / Q000187:drug effects; D009569:Nitric Oxide / Q000096:biosynthesis*; D052247:Nitric Oxide Synthase Type II / Q000096:biosynthesis / Q000235:genetics; D010003:Osteoarthritis / Q000517:prevention & control*; D012333:RNA, Messenger / Q000235:genetics; D051381:Rats; D017208:Rats, Wistar; D011994:Recombinant Proteins / Q000494:pharmacology / Q000627:therapeutic use; D020133:Reverse Transcriptase Polymerase Chain Reaction / Q000379:methods; D013314:Stress, Mechanical; D014443:Tyrosine / Q000031:analogs & derivatives / Q000378:metabolism
D053674:Aggrecans; D012333:RNA, Messenger; D011994:Recombinant Proteins; D015847:Interleukin-4; D009569:Nitric Oxide; C002744:3-nitrotyrosine; D014443:Tyrosine; D052247:Nitric Oxide Synthase Type II