Fig. 4

Gucy1α1 co-labels Pdgfrβ-, αSma- and Vim-positive fractions of the baseline and activated kidney fibroblasts in the medullary areas. (A) Schematic of the kidney anatomy with respect to medullary area (highlighted with a black frame). (B) Representative IF images of the control, UIR/UUO Day 1, 4, 7, 14 and 28 kidneys, Gucy1α1 (magenta), Pdgfrβ (white), αSma (cyan), Vim (green), DAPI, blue. The upper panels show all combined signals, the middle panels show individual channels, and three bottom panels demonstrate Gucy1α1 along with Pdgfrβ, αSma or Vim, respectively. Original magnification, × 60, maximal intensity projection from a Z-stack, 0.09 μm/px Nyquist zoom, scale 15 μm. (C) Violin plot showing quantitative IF analysis of medullary patterns of kidney fibrosis markers expression. Note near-total overlap between Gucy1α1- and Pdgfrβ-positive areas in the control kidneys and at all stages of both injuries (average percentage of Pdgfrβ co-expression in Gucy1α1-positive areas for control: 92%; UIR Day 1: 97.75%, Day 4: 98.25%, Day 7: 97.75%, Day 14: 93%, Day 28: 91.75%; UUO Day 1: 97%, Day 4: 97.75%, Day 7: 93.25%, Day 14: 95.5%, Day 28: 95.25%). Also note that medullary Gucy1α1-expressing fibroblasts acquired αSma co-expression more abruptly and robustly than the cortical ones, and retained it at higher percentages (average double positivity for control: 16.5%; UIR Day 1: 85.75%, Day 4: 92%, Day 7: 85%, Day 14: 86.75%, Day 28: 69.25%; UUO Day 1: 76%, Day 4: 96.25%, Day 7: 87.5%, Day 14: 96.5%, Day 28: 88.25%). Similar to the cortex, most medullary Gucy1α1-positive areas exhibited Vim co-expression. N = 4 animals per group.