Abstract
BackgroundJoint contracture is a fibrous disease characterized as joint capsule fibrosis that results in joint dysfunction and disability. The purpose of this study was to analyze the biological activities of chaperonin containing T-complex polypeptide (CCT) subunits and to determine the role of CCT chaperone in joint contracture in a rat model.MethodsIn this study, the rat model of joint contracture was established by immobilizing the rat knee for 8 weeks. Then, fibroblasts were isolated from the posterior joint capsule and were cultured for functional analysis such as qRT-PCR, Western blot, transwell assay, and collagen assay. The effect of CCT subunit was determined by employing a lentivirus containing target gene and transfecting it into fibroblasts.ResultsResults of qRT-PCR and Western blot showed that among all CCT subunits, CCT6b significantly decreased in the fibroblasts from contractive joints compared to cells from normal joints (p < 0.05). Overexpression of CCT6b by transfection of lentivirus containing CCT6b gene to active fibroblasts significantly inhibited fibrous marker (α-SMA, COL-1) expressions, fibroblast migration, and collagen synthesis (all p < 0.05). Moreover, fibrosis-related chaperone CCT7 expression was decreased with CCT6b overexpression (p < 0.05).ConclusionThe biological activities of CCT subunits in fibroblasts from the joint contracture rat model were analyzed in this study. CCT6b significantly decreased in the active fibroblasts, and overexpression of CCT6b significantly inhibited fibroblast functions. These findings indicate that CCT6b appears to be a potential molecular biomarker and therapeutic target for the novel therapies of joint contracture.
Highlights
Joint contracture, characterized as losing active and passive range of motion (ROM) on the affected joints, is a severe complication resulting in irreversible disability after joint surgery or longtime immobilization [1]
Results of the Western blot indicated that among all containing T-complex polypeptide (CCT) subunits, the expressions of CCT6b and CCT7 were significantly different in the Fibroblasts derived from the immobilized knees (IF) group compared to the Fibroblasts derived from the contralateral knees (CF) group (p < 0.05), while other CCT subunits showed no difference between the two groups (Fig. 3a)
Comparing to the CF group, CCT6b expression was significantly decreased in fibroblasts from the IF group, while an increased expression of CCT7 was observed (p < 0.05). Quantitative real-time polymerase chain reaction (qRT-PCR) demonstrated the similar trends of CCT6b and CCT7 mRNA expressions (p < 0.05) in the IF group when compared to the CF group (Fig. 3b)
Summary
Joint contracture, characterized as losing active and passive range of motion (ROM) on the affected joints, is a severe complication resulting in irreversible disability after joint surgery or longtime immobilization [1]. We have previously confirmed that chaperonin containing T-complex polypeptide (CCT) subunit eta (CCT7) was a potential marker in joint contracture in the rat model [8]. The eight subunits may contribute to the proliferation and activation of fibroblasts since they cooperate in many biological functions [18]. The biological functions of CCT subunits have not yet been systematically analyzed in joint contracture. Joint contracture is a fibrous disease characterized as joint capsule fibrosis that results in joint dysfunction and disability. The purpose of this study was to analyze the biological activities of chaperonin containing T-complex polypeptide (CCT) subunits and to determine the role of CCT chaperone in joint contracture in a rat model
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