| Objective: The purpose of this study is to explore the efficacy and mechanism of low intensity pulsed ultrasound in improving the arthrogenic contracture of knee.Methods:(1)Experimental groups:(1)6 New Zealand white rabbits without any treatment: control group(C).(2)Thirty New Zealand white rabbits were received left hind limb knee extension immobilization,then they were randomly divided into the following 5 groups(n=6): 6 weeks immobilization group(IM-6w);1 week of remobilization group(RM-1w)and LIPUS intervention group(LIPUS-1W)after 6weeks of immobilization;2 week of remobilization group(RM-2w)and LIPUS intervention group(LIPUS-2W)after 6 weeks of immobilization.(2)Evaluation method of experimental results:(1)Evaluation of arthrogenic contracture:measurement with range of motion measuring instrument.(2)To evaluate the degree of fibrosis of the joint capsule: The percentage of collagen deposition.The protein expression level of type I collagen,type III collagen and α-SMA.(3)Evaluate the level of reactive oxygen species(ROS)in the joint capsule: The content of malondialdehyde and superoxide dismutase was detected.(4)Evaluate the activation of TGF-β1/Smad signal pathway in joint capsule: the expression level of TGF-β1,p-Smad2,Smad2,p-Smad3,Smad3 and Smad4 protein.Number of p-Smad2/3 positive cells.Results:(1)Compared with group C,after 6 weeks of immobilization,the percentage of arthrogenic contracture and collagen deposition in group IM-6w was increased(p<0.05),the protein expression level of type I collagen,type III collagen and α-SMA was increased(p<0.05).Malondialdehyde activity was increased(p<0.05),correspondingly,superoxide dismutase content was decreased(p<0.05).The protein level of TGF-β1,p-Smad2/Smad2,p-Smad3/Smad3 and Smad4 were increased(p<0.05).It is showing that TGF-β1/Smad signaling pathway was significantly activated.(2)Arthrogenic contracture:(1)Compared with the IM-6w group,the arthrogenic contracture in the RM-1w group was worse(p<0.05),but the arthrogenic contracture in the RM-2w group was less(p<0.05).(2)Compared with IM-6w group,the degree of arthrogenic contracture decreased with the increase of LIPUS intervention time(p<0.05).(3)At each time point,LIPUS group showed reduced arthrogenic contractures compared to RM group(p<0.05).(3)Joint capsule fibrosis,ROS level,the activation of TGF-β1/Smad signaling pathway in knee joint capsule:(1)Compared with IM-6w group,the percentage of collagen deposition in RM group was significantly increased(p<0.05),and the protein expression levels of type I collagen,type III collagen and α-SMA were elevated(p<0.05);Similarly,malondialdehyde activity was increased(p<0.05),while superoxide dismutase content was decreased(p<0.05).Besides,the protein expression levels of TGF-β1,p-Smad2/Smad2,p-Smad3/Smad3 and Smad4 were elevated(p<0.05).The number of P-Smad2/3positive cells were increased(p<0.05).However,there was no statistical difference in these indexes between RM-1w group and RM-2w group(p>0.05).(2)Compared with IM-6w group,fibrosis indicators mentioned above in LIPUS group were decreased gradually with the progression of intervention time(p<0.05).Correspondingly,malondialdehyde activity decreased gradually(p<0.05),while superoxide dismutase content increased gradually(p<0.05).Related indicators of TGF-β1/Smad signaling pathways in LIPUS group were also decreased gradually(p<0.05).(3)At each time point,fibrosis level in LIPUS group was decreased,and ROS production and TGF-β1/Smad signaling pathway were inhibited in LIPUS group compared to RM group(p<0.05).Conclusions: High ROS levels and overactivity of TGF-β1/Smad signaling pathway may be one of the causes of immobilization-induced knee capsule fibrosis.LIPUS reduces the degree of immobilization-induced knee capsule fibrosis by inhibiting ROS production and activation of TGF-β1/Smad signaling pathway. |
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