Figure 2. Proximal tubular-specific ADAM17 KO confers protection against fibrosis after IRI.

Control (SLC34a1^GCE/+ ADAM17^WT/WT, Cre WT/WT) and ADAM17 PTC-KO (SLC34a1^GCE/+ ADAM17^fl/fl, Cre fl/fl) mice were subjected to bilateral ischemia for 29 minutes followed by reperfusion. (A) KO specificity was examined in healthy kidneys (left: control, right: PTC-KO mouse) by immunostaining of ADAM17 (red color) together with LTL staining (green color) as a proximal tubular marker (thick arrows, LTL-positive tubules; thin arrows, LTL-negative tubules; scale bar: 50 μm). (B–D) Time-course of BUN, urinary NGAL, and urinary KIM1 (n = 6–15). (E) The induction of fibronectin and αSMA was examined in kidney cortex by immunostaining at day 21 (left: representative images, right: quantification, n = 10; scale bar: 50 μm). (F) Fibronectin protein expression at day 21 after injury was examined by Western blot (top: sample blots; bottom: quantification; GAPDH was used as loading control; n = 4). (G and H) Masson’s trichrome staining in kidney cortex at day 21 after injury (G: representative images; H: quantification; n = 6; scale bar: 100 μm). (I) Total interstitial area (n = 6) and (J) dilated tubules (n = 6) were quantified in kidney cortex at day 21 after injury. *P < 0.05; **P < 0.01; ***P < 0.001 as determined by an unpaired 2-tailed Student’s t test. Diamond symbol denotes position of kidney capsule. ADAM17, a disintegrin and metalloprotease 17; BUN, blood urea nitrogen; IRI, ischemia reperfusion injury; KIM1, kidney injury molecule 1; LTL, Lotus Tetragonolobus lectin; MT, Masson’s trichrome; NGAL, neutrophil gelatinase–associated lipocalin; PTC-KO, proximal tubule cell KO; αSMA, alpha smooth muscle actin; WB, Western blot.