Research Paper

Figure 7. Rescue experiments proved that hucMSCs treatment reduced mechanical stiffness by regulating circANKRD42–miR-136-5p–YAP1 signal pathway in vivo and in vitro. (A) AFM images depicted that BLM caused the roughness of the lung surface and the thickening of the lung tissues. hucMSCs treatment improved the lung tissue morphology, but this therapeutic effect was reversed by overexpressed circANKRD42. The effect of circANKRD42 mutation was weaker than that of overexpressed circANKRD42. (B) In the TGF-β1 group, the cells became slender and flat. Stress fibers arranged in parallel along the main axis of cells were observed in the group. In the hucMSCs-treated group, the cell shape significantly retracted and the corresponding fibers were reduced. Overexpressed circANKRD42 reversed the effect of hucMSCs treatment on cells. (C) Young’s modulus measurement showed that hucMSCs treatment decreased the force compared with that in the BLM group, but overexpressed circANKRD42 reversed the effect of hucMSCs treatment on the force. The effect of circANKRD42 mutation on the force was weaker than that of overexpressed circANKRD42. The measured average Young’s modulus was 1291.667 ± 82.735 MPa in the BLM group, 563.833 ± 31.822 MPa in the BLM+hucMSCs group, 520 ± 20.460 MPa in the BLM+hucMSCs+circANKRD42 BP group, 942.05 ± 28.203 MPa in the BLM+hucMSCs+circANKRD42 RP group, and 604.233 ± 29.259 MPa in BLM+hucMSCs+circANKRD42 Mut. (D) Young’s modulus measurement revealed that hucMSCs treatment decreased the force compared with that in the TGF-β1 group, but overexpressed circANKRD42 reversed the effect of hucMSCs treatment on the force. The measured average Young's modulus was 641.167 KPa in the TGF-β1 group, 389.467 KPa in the TGF-β1+hucMSCs group, 382.767 in the TGF-β1+hucMSCs+circANKRD42 BP group, and 495.433 KPa in TGF-β1+hucMSCs+circANKRD42 RP. (E) Young’s modulus measurement showed that hucMSCs treatment decreased the reaction forces of cells compared with that in the TGF-β1 group, but overexpressed circANKRD42 reversed the effect of hucMSCs treatment on the force. The measured average Young's modulus was 8.858 ± 0.526 KPa in the TGF-β1 group, 4.756 ± 0.306 KPa in the TGF-β1+hucMSCs group, 4.504 ± 0.160 KPa in the TGF-β1+hucMSCs+circANKRD42 BP group, and 6.092±0.240 KPa in TGF-β1+hucMSCs+circANKRD42 RP. (F) Cytoskeleton staining with FITC-phalloidin depicted that cytoskeleton tension was weakened by hucMSCs treatment but enhanced by circANKRD42 overexpression. (G) hucMSCs treatment inhibited the expression levels of YAP1, Myo1c, and F-actin and increased p-YAP1 expression in vivo and in vitro. circANKRD42 overexpression increased the expression of YAP1, Myo1c, and F-actin and decreased p-YAP1 expression, thus reversing the effect of hucMSCs treatment in vivo and in vitro. The effect of circANKRD42 mutation was weaker than that of overexpressed circANKRD42. Each bar represents the mean ± SD; n = 6; ^*p < 0.05.