Figure 4.

MiR-223-3p overexpression and FOXO3 knockdown reversed the inhibitory effect of Que on cell proliferation and the expressions of proliferation-related and autophagy-related genes in ISO-reduced RCFs. (a). The expression of miR-223-3p after transfection with miR-223-3p mimic was detected by RT-qPCR, U6 was used as an internal control. (b).The expression of FOXO3 protein in RCFs after transfection with siFOXO3 were detected by Western blot, β-actin was used as internal control. (c).The expression of FOXO3 mRNA in RCFs after transfection with siFOXO3 were detected by RT-qPCR, β-actin was used as internal control. (d). The expression of miR-223-3p in ISO-induced RCFs after treatment with Que and transfection with miR-223-3p mimic or siFOXO3 was detected by RT-qPCR, U6 was used as an internal control. (e). The cell proliferation of ISO-induced RCFs after treatment with Que and transfection with miR-223-3p mimic or siFOXO3 was detected by MTT assay. (f). The mRNA expressions of Cyclin D1, Fibronectin, and Col1A1 of ISO-induced RCFs after treatment with Que and transfection with miR-223-3p mimic or siFOXO3 was detected by RT-qPCR, β-actin was used as an internal control. (g-h) The protein expressions of Cyclin D1, Fibronectin, and Col1A1 of ISO-induced RCFs after treatment with Que and transfection with miR-223-3p mimic or siFOXO3 was detected by Western blot, β-actin was used as an internal control. (i-l) The protein expressions of FOXO3, ATG7, LC3B-I, LC3B-II, and p62/SQSTM1 of ISO-induced RCFs after treatment with Que and transfection with miR-223-3p mimic or siFOXO3 was detected by Western blot, β-actin was used as an internal control. All experiments were conducted three times. (***P < 0.001, vs. ISO, ^{^^}P < 0.01, ^{^^^}P < 0.001, vs. ISO+Que-H). (Que: quercetin, ISO: isoprenaline, M: miR-223-3p mimic, MC: mimic control, siFOXO3: small interfering RNA directed against FOXO3, siNC: siRNA negative control)