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. 2019 Jul 25;4:100011. doi: 10.1016/j.mbplus.2019.100011

Fig. 7.

Fig. 7

Depletion of EMILIN1 from the ECM layer reduces paxillin area and stress fibers in renal epithelial cells. (A) Quantified results showing the paxillin area in HRPTECs on siSHAM or siEMILIN1 ECM during initial binding to the ECM (cultured for 2 h). Shown is mean ± SEM; ***P < 0.001. N = 30 fluorescent images derived from N = 3 samples. (B) Representative immunofluorescence images (400× magnification) of HRPTECs after 2 h of binding on siSHAM or siEMILIN1 ECM and stained for paxillin (green), F-actin (red) and DAPI (blue). Scale bar represents 20 μm (overview image, left) and 5 μm (zoomed-in image, right). Quantified results showing the ZO-1 (C) and F-actin area (D) in HK2 cells cultured to confluency on siSHAM or siEMILIN1 ECM. Shown is mean ± SEM; ***P < 0.001. N = 20 fluorescent Z-stacks derived from N = 4 samples. (E) Representative immunofluorescence Z-stacks (630× magnification) of HK2 cells cultured to confluency (cultured for 48 h) on siSHAM or siEMILIN1 ECM and stained for ZO-1 (green), F-actin (red) and DAPI (blue). Scale bar represents 20 μm (overview image, left) and 5 μm (zoomed-in image, right). (F) Colocalization of paxillin and F-actin in HK2 cells cultured to confluency on siSHAM or siEMILIN1 ECM. Shown is mean ± SEM; *P < 0.05. N ≥ 25 fluorescent Z-stacks, from N ≥ 5 samples. (G) Representative immunofluorescence Z-stacks (630× magnification) of HK2 cells cultured to confluency on siSHAM or siEMILIN1 ECM and stained for paxillin (green), F-actin (red) and DAPI (blue). Colocalization is displayed as yellow. Scale bar represents 15 μm (overview image, left) and 5 μm (zoomed-in image, right). (H) RhoA activation levels in HK2 cells cultured for 48 h on siSHAM or siEMILIN1 ECM. Shown is mean ± SEM; *P < 0.05. N = 5 assays.