[Phosphorylated Focal Adhesion Kinase, Phosphinositides 3 Kinase and Aggrecan Genes and Proteins in Cartilage Cells Are Probably Involved in Needle Knife Intervention Induced Improvement of Knee Osteoarthritis in Rabbits].

ABSTRACT

OBJECTIVE: We have demonstrated that needle knife (acupotomy) treatment can improve knee osteoarthritis (KOA) in rabbits. The present study was designed to examine its effect on expression of phosphorylated focal adhesion kinase (p-FAK), phosphinositides 3 kinase (p-PI 3 K) and Aggrecan genes and proteins in the knee-joint cartilage tissues of KOA rabbits, so as to explore its partial molecular mechanism underlying improvement of KOA. METHODS: Forty-nine New Zealand male rabbits were randomly divided into normal control, model, model +inhibitor, needle knife, needle knife+ inhibitor, electroacupuncture (EA), EA +inhibitor groups (n=7 in each). The KOA model was established by modified Videman method (left hindlimb extension immobilization and ankle dorsal flexion 60°). Acupotomy relaxing manipulation was applied to the lateral collateral ligament and patellar ligament of the left knee-joint, two times a week for 4 weeks, and EA (2 Hz/100 Hz, 3 mA) was applied to the left "Liangmen" (ST 21), "Xuehai" (SP 10), "Neixiyan " (EX-LE 4) and "Dubi" (ST 35) for 20 min, three times a week, for 4 weeks. About 2 h before every needle-knife or EA treatment or at the corresponding time-point, intra-articular cavity injection of PF-562271(a specific antagonist of FAK, 200 µmol/L, 0.5 mL)was performed in the three inhibitor groups. The expression levels of p-FAK, p-PI 3 K, Aggrecan genes and proteins in the cartilage tissues were measured with quantitative Real-time PCR and Western blot, separately. RESULTS: After modeling, the expression levels of p-FAK and p-PI 3 K genes and proteins were significantly up-regulated (P<0.05, P<0.01), while those of Aggrecan protein and mRNA considerably down-regulated in the model group in comparison with the normal group (P<0.01). Following 4 weeks' needle-knife or EA treatment, the expression levels of p-FAK and p-PI 3 K and Aggrecan proteins in both EA and needle-knife groups, and Aggrecan mRNA in the needle knife group were significantly up-regulated (P<0.05, P<0.01). After administration of p-FAK antagonist, modeling-induced upregulation of expression of p-FAK mRNA and protein and p-PI 3 K protein, as well as modeling-induced down-regulation of Aggrecan mRNA and protein were significantly suppressed in the model+inhibitor group (P<0.05, P<0.01), and needle knife-induced and EA-induced up-regulation of expression of p-FAK, p-PI 3 K and Aggrecan mRNAs and proteins was notably suppressed respectively in comparison with the needle knife and EA groups (P<0.05, P<0.01). The effect of needle knife was significantly superior to that of EA in up-regulating p-PI 3 K and Aggrecan mRNAs as well as p-FAK, p-PI 3 K and Aggrecan proteins (P<0.05, P<0.01). CONCLUSION: Needle knife intervention can up-regulate the expression levels of p-FAK, p-PI 3 K and Aggrecan proteins and mRNAs in the cartilage tissue of the knee-joint in KOA rabbits, suggesting an involvement of FAK-PI 3 K signaling in the needle knife-induced improvement of KOA.