TLR4 knockdown by miRNA-140-5p improves tendinopathy: an in vitro study
Tu Jun 1
Xu Bin 1
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Department of Sports Medicine and Arthroscopic Surgery, The First Affiliated Hospital of Anhui Medical University, Hefei City, Anhui Province, China
Submission date: 2019-10-24
Final revision date: 2019-11-07
Acceptance date: 2019-11-24
Online publication date: 2020-02-24
Arch Med Sci 2024;20(2)
The purpose of this study was to determine whether TLR4 knockdown induced by miRNA-140-5p improves tendinopathy in an in vitro experiment.

Material and methods:
Extraction of tendon-derived stem cells (TDSCs) from SD rats was performed using TGF-1 to develop a tendinopathy cell model. In the first step, we knocked down TLR4 by si-TLR4 to investigate TLR4 in tendinopathy development, and the next we used miRNA-140-5p to investigate miRNA-140-5p in tendinopathy development. The inflammatory factors and Hyp concentration were evaluated by ELISA assay; the cell viability was measured by MTT assay; the cell apoptosis was evaluated by TUNEL and/or flow cytometry. The relative mRNA was measured by RT-qPCR assay; the relative proteins expression was evaluated by cellular immunofluorescence and/or WB assay. The correlation between miRNA-140-5p and TLR4 was analyzed by Luciferase reporter assay.

With miRNA-140-5p overexpression or TLR4 knockdown, the cell viability was significantly increased with cell apoptosis depressing compared with the Model group (p < 0.05, respectively). Meanwhile, the inflammatory factors TNF-α, IL-1β and IL-6 and Hyp concentration were significantly improved (p < 0.05, respectively), whereas the TLR4, MyD88 and NF-κB(p65) protein expression levels were significantly depressed with TLR4 knockdown by si-TLR4 or miRNA-140-5p which target TLR4.

The present results showed that TLR4 knockdown induced by miRNA-140-5p or si-TLR4 improved tendinopathy in an in vitro cell experiment.