壳聚糖通过miR-29b经TGF-β1/Smad3信号通路抑制跟腱成纤维细胞生长。
Chitosan inhibits fibroblasts growth in Achilles tendon via TGF-β1/Smad3 pathway by miR-29b.
作者信息
Chen Qiang, Lu Hui, Yang Hu
机构信息
Department of Hand Surgery, The First Affiliated Hospital, College of Medicine, Zhejiang University China.
出版信息
Int J Clin Exp Pathol. 2014 Dec 1;7(12):8462-70. eCollection 2014.
BACKGROUND AND AIM
Chitosan, is a natural polymer, plays an important role in prevention of tendon adhesion in tendon healing process. However, the molecular mechanisms underlying the prevention effect is unclear. Here we investigated the effects of chitosan on Achilles tendon injury rats and fibroblasts.
METHODS
Eight weeks after surgery, gliding excursion and the content of collagen fibers in Achilles tendon injury rats were determined to evaluate the chitosan effect on tendon healing. Fibroblasts isolated from scar tissue of repaired tendon were treated with different concentration of chitosan, and then cell inhibition, apoptosis and cell cycle were measured using MTT and Flow Cytometry respectively. The expression of microRNAs (miRNAs) was quantified by real-time PCR and protein expression of TGF-β1, Smad3 and P21 were quantified by western blotting. MiR-29b inhibitor was transfected in cells to evaluate the mechanism underlying the effects of chitosan on tendon fibroblasts.
RESULTS
The gliding excursion of repaired tendon was increased and the content of collagen fibers was decreased by chitosan in rats. Chitosan inhibited the fibroblasts growth and arrested cells in G1 phase. Chitosan also elevated the expression of miR-29b and P21 while reduced the levels of TGFβ1 and Smad3 in both repaired tendon and fibroblasts. In addition, miR-29b inhibitor revered the effects of chitosan on fibroblasts.
CONCLUSIONS
The current study demonstrated that chitosan improving the condition of tendon healing after surgery, which is reduced by the high expression of miR-29b and its down-regulation of TGF-β1/Smad3 level and inhibition of fibroblasts growth.
背景与目的
壳聚糖是一种天然聚合物，在肌腱愈合过程中对预防肌腱粘连起着重要作用。然而，其预防作用的分子机制尚不清楚。在此，我们研究了壳聚糖对跟腱损伤大鼠和成纤维细胞的影响。
方法
术后8周，测定跟腱损伤大鼠的滑动幅度和胶原纤维含量，以评估壳聚糖对肌腱愈合的影响。用不同浓度的壳聚糖处理从修复肌腱的瘢痕组织中分离出的成纤维细胞，然后分别使用MTT法和流式细胞术检测细胞抑制、凋亡和细胞周期。通过实时PCR定量检测微小RNA（miRNA）的表达，通过蛋白质免疫印迹法定量检测TGF-β1、Smad3和P21的蛋白表达。将miR-29b抑制剂转染到细胞中，以评估壳聚糖对肌腱成纤维细胞作用的机制。
结果
壳聚糖可增加大鼠修复肌腱的滑动幅度，减少胶原纤维含量。壳聚糖抑制成纤维细胞生长并使细胞停滞于G1期。壳聚糖还提高了修复肌腱和成纤维细胞中miR-29b和P21的表达，同时降低了TGFβ1和Smad3的水平。此外，miR-29b抑制剂可逆转壳聚糖对成纤维细胞的作用。
结论
本研究表明，壳聚糖可改善术后肌腱愈合状况，这是通过miR-29b的高表达及其对TGF-β1/Smad3水平的下调和对成纤维细胞生长的抑制来实现的。
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