Fig. 4
AFsHPi-Exos transport miR-21-5p to VSMCs and promote the calcification of VSMCs. A Heatmap showing expression profiles of candidate miRNAs in AFsNPi-Exos and AFsHPi-Exos. B qPCR analysis showing the level of miR-21-5p in VSMCs treated with AFsNPi-Exos or AFsHPi-Exos. C The miR-21-5p levels in AFsHPi-Exos treated with RNase, proteinase K (ProtK), or Triton X-100 as measured by qPCR. D pri-miR-21 levels in NPi/HPi treated AFs, AFsNPi-Exos/AFsHPi-Exos and AFsNPi-Exos/AFsHPi-Exos treated VSMCs as measured by qPCR. E Fluorescence microscopy analysis of PKH26-labeled exosomes derived from FAM-miR-21-5p-mimics transfected AFs internalization by VSMCs. FAM-miR-21-5p signals (green) were co-localized with PKH26 signals in the perinuclear region of VSMCs (PKH26 in red and DAPI in blue). The scale bar represents 50 μm. Western blot (F) and ALP activity assay (G) showing the Runx2 and BMP2 protein expression and ALP activity in VSMCs after treatment with vehicle or exosomes derived from AFs transfected with miR-21-5p mimics/miR-21-5p inhibitor/negative controls. The levels of Runx2 and BMP2 (H) and ALP activity (I) in VSMCs after miR-21-5p inhibitor transfection under AFsHPi-Exos treatment. The VSMCs in all groups were induced by 3.5 mM Pi simultaneously. Data are the mean ± SD of three independent experiments. *p < 0.05, **p < 0.01