Journal: Investigative Ophthalmology & Visual Science

Article Title: The miR-320a/PRDX3 Axis Alleviates the Oxidative Stress and Fibrotic Alterations in Fibroblasts in Thyroid Eye Disease

doi: 10.1167/iovs.66.9.41

Figure Lengend Snippet: Inhibition of miR-320a inhibited TGF-β1-induced fibroblast activation . ( A ) The miR-320a inhibition was achieved in TED OFs by transfecting miR-320a inhibitor and verified using qRT-PCR. Then, the TED OFs were transfected with miR-320a inhibitor or inhibitor negative control (NC), exposed to 10 ng/mL TGF-β1 for 48 hours, and conducted a series of experiments. ( B ) Cell viability was detected by CCK-8 assay. ( C ) DNA synthesis was determined using EdU assay. ( D ) The mRNA expression of fibronectin and vimentin were detected using qRT-PCR. ( E ) The protein levels of vimentin and fibronectin were measured using immunoblotting. ( F ) The collagen I content was ascertained using ELISA. ** P < 0.01 versus the blank group; ## P < 0.01 versus the TGF-β1 + inhibitor NC group.

Article Snippet: The secreted Collagen I levels of OFs in culture medium were determined by the Human Collagen Type I ELISA Kit (CSB-E08082h; CUSABIO).

Techniques: Inhibition, Activation Assay, Quantitative RT-PCR, Transfection, Negative Control, CCK-8 Assay, DNA Synthesis, EdU Assay, Expressing, Western Blot, Enzyme-linked Immunosorbent Assay

Journal: Investigative Ophthalmology & Visual Science

Article Title: The miR-320a/PRDX3 Axis Alleviates the Oxidative Stress and Fibrotic Alterations in Fibroblasts in Thyroid Eye Disease

doi: 10.1167/iovs.66.9.41

Figure Lengend Snippet: Inhibition of miR-320a improves the progression of TED in mice . ( A , B ) TED model was established in mice as described; the model was verified using H&E staining examining mice thyroid tissues and ELISA examining serum T4 and TSAb levels. Next, miR-320a inhibition was achieved in mice by orbital injection in TED model mice. ( C ) The photographs of the eyeballs of the model mice were showed. The orbital inflammation in TED mouse model was quantified by a semi-quantitative murine orbital activity score. ( D , E ) histopathological alterations in mice orbital tissues were evaluated using H&E ( D ) and Masson staining ( E ). The collagen volume fraction (CVF) of mice orbital tissues was calculated with ImageJ software. ( F ) The expression levels of miR-320a, vimentin, and fibronectin were determined using qRT-PCR. ( G ) The protein levels of vimentin and fibronectin were determined using immunoblotting. ( H ) The level of collagen I in orbital tissues was determined using ELISA. ** P < 0.01 versus the control group; ## P < 0.01 versus the TED + antagomir-NC group.

Article Snippet: The secreted Collagen I levels of OFs in culture medium were determined by the Human Collagen Type I ELISA Kit (CSB-E08082h; CUSABIO).

Techniques: Inhibition, Staining, Enzyme-linked Immunosorbent Assay, Injection, Activity Assay, Software, Expressing, Quantitative RT-PCR, Western Blot, Control

Journal: Investigative Ophthalmology & Visual Science

Article Title: The miR-320a/PRDX3 Axis Alleviates the Oxidative Stress and Fibrotic Alterations in Fibroblasts in Thyroid Eye Disease

doi: 10.1167/iovs.66.9.41

Figure Lengend Snippet: Inhibition of PRDX3 partially reversed the inhibitory effects of miR-320a inhibition on fibroblast activation . ( A ) PRDX3 knockdown was achieved in TED OFs by transfecting small interfering RNA (siRNA) against PRDX3 (si-PRDX3#1/#2) and verified using qRT-PCR. Next, TED OFs were co-transfected with si-PRDX3 and miR-320a inhibitor, exposed to 10 ng/mL TGF-β1 for 48 hours, and conducted a series of experiments. ( B ) Cell viability was measured using CCK-8 assay. ( C ) DNA synthesis was detected using EdU assay. ( D ) The mRNA expression levels of fibronectin and vimentin were determined using qRT-PCR. ( E ) The protein levels of vimentin and fibronectin were detected using immunoblotting. ( F ) The collagen I content in culture medium was measured using ELISA. * P < 0.05, ** P < 0.01, *** P < 0.001 versus the inhibitor NC + si-NC group; ## P < 0.01 versus the inhibitor NC + si-PRDX3 group; && P < 0.01 versus the miR-320a inhibitor + si-NC group.

Article Snippet: The secreted Collagen I levels of OFs in culture medium were determined by the Human Collagen Type I ELISA Kit (CSB-E08082h; CUSABIO).

Techniques: Inhibition, Activation Assay, Knockdown, Small Interfering RNA, Quantitative RT-PCR, Transfection, CCK-8 Assay, DNA Synthesis, EdU Assay, Expressing, Western Blot, Enzyme-linked Immunosorbent Assay