Figure 1. Dedifferentiated VSMCs contribute to neointima formation in AVF.

A. VSMC marker, SMA-α, was absent in arterial media in the anastomosis of AVF The anastomosis regions in the longitudinal section of AVFs were showed in frames, the black arrows point to the SMA-α negative cells (A, arterial side; V, venous side; L, lumen, scale bar = 500 μm). B. Double immunofluorescent staining of the SMMHC/SMA-α in anastomosed area of AVFs (2 weeks). The arrows point to the VSMC marker negative cells in the media of the arterial side of the anastomosis. C. Generating VSMC reporter transgenic mice. After tamoxifen induction, only VSMCs and their lineages will be labeled with GFP. D. VSMCs (SMMHC⁺) were labeled with GFP in the common carotid artery in RFP^flox/flox-GFP/SMMHC-ERCre⁺ mice (VSMC^GFP mice) after tamoxifen treatment. E. AVFs were created in VSMC^GFP mice after 14 days of last dose of tamoxifen treatment. The expression of SMMHC and GFP were determined in anastomosis (E, left panel) of the 2 week AVFs (yellow arrow points SMMHC⁺/GFP⁺ cells; white arrows point SMMHC⁻/GFP⁺ cells); the percentage of the two types of cells were counted (E, right panel). F. mRNA ratio of SMMHC and GFP were determined from common carotid artery and anastomosis of AVFs that were created in VSMC^GFP mice (representative data was shown from 3 repeated experiments, *, p < 0.05). G. AVFs were created in VSMC^GFP mice after 14 days of last dose of tamoxifen treatment. The expression of SMA-α and GFP were determined in neointima area in the venous side of the 4 week AVFs. The total GFP⁺ cells and GFP⁺/SMA-α⁻ cells were counted (right panel. n = 6). Scale bar = 50 μm in panel B, D, E, and G.