AngII infusion causes persistent glomerular desmin expression.

<p>Ang II infusion increased desmin protein levels (A). At sacrifice there was a significant increase of desmin protein in former AngII-infused rats and surprisingly in control rats as well. Compared to control rats, glomerular desmin expression of AngII-infused rats was significantly higher after stopping AngII infusion. Representative photomicrographs of desmin stained renal sections (B). (## p<0.01, ### p<0.001 <i>vs</i>. control at time of biopsy; *p<0.05 <i>vs</i>. AngII at time of biopsy; $ p<0.05 <i>vs</i>. control at time of sacrifice).</p

Regression of lymph vessels after Ang II withdrawal, no effect on peritubular capillaries.

<p>Ang II infusion increased mRNA expression of PDPN, however this was not significant (A). Ang II-induced lymphangiogenesis was evidenced by an increase in lymph vessel formation (B), with spontaneous regression to control levels eight weeks after withdrawal of Ang II. Ang II infusion had no effects on peritublar capillaries (C). Representative photomicrographs of Podoplanin stained renal sections (D). (##p<0.01 vs. control at time of biopsy; ***p<0.001 <i>vs</i>. Ang II at time of biopsy)</p

Increased expression of TNF-α and MCP-1 in double mutant mice, lacking both collagen XV and XVIII compared to WT.

<p><i>A–B:</i> Immunofluorescent staining of MCP-1 (red) in WT (<i>A</i>) and double mutant mice (<i>B</i>) at day 5. Double mutant mice showed an increased expression of MCP-1 in peri-tubular capillaries. Scale bars 20 µm. <i>C:</i> mRNA expression of MCP-1 in renal tissue of <i>Col15a1^−/−</i>, <i>Col18a1^−/− and Col15a1^−/−×Col18a1^−/−</i> double compound compared to WT mice at day 5 after I/R (***: p<0.001 double mutant mice compared to WT). <i>D:</i> mRNA expression of TNF-α in renal tissue of <i>Col15a1^−/−</i>, <i>Col18a1^−/− and Col15a1^−/−×Col18a1^−/−</i> double compound compared to WT mice at day 5 after I/R (**: p<0.01 double mutant mice compared to WT).</p

Effects of Ang II on OPN expression.

<p>Spp1 mRNA (A) and OPN protein (B) levels were increased by Ang II infusion. After stopping Ang II infusion Spp1 mRNA and OPN protein levels returned to control levels. In time, OPN protein expression of control rats significantly increased. Representative photomicrographs of OPN stained renal sections (C). (## p<0.01, ### p<0.001 <i>vs</i>. control at time of biopsy; ***p<0.001 <i>vs</i>. Ang II at time of biopsy).</p

Normalization of mRNA expression of fibrotic markers after cessation of Ang II infusion.

<p>mRNA expression of the fibrotic markers TGF-beta, Col4a1 and Col5a1 was upregulated under Ang II infusion (A, B, C). Eight weeks after stopping Ang II infusion, mRNA levels of all markers returned to control levels again. (#p<0.05, ##p<0.01 <i>vs</i>. control at time of biopsy; *p<0.05, **p<0.01 <i>vs</i>. Ang II at time of biopsy).</p

Decrease in Col1a1 mRNA expression after stopping Ang II infusion, ongoing deposition of Collagen I proteins.

<p>Ang II infusion increased Col1a1 mRNA (A) expression. After withdrawal of Ang II, Col1a1 mRNA levels returned to control values. Protein expression of Collagen I, as measured by dot blot analysis, was unaffected after infusion with Ang II. However, eight weeks after cessation of Ang II protein levels of Collagen I increased (B). Collagen I protein expression of control rats also significantly increased at sacrifice compared to their own biopsy levels (B). At time of sacrifice, former AngII-infused rats had significantly higher Collagen I protein levels when compared to control (B). Representative photomicrographs of Collagen I stained renal sections (C). Representative examples of immunoblot analysis of Collagen I of two tissue samples in triplicate (D). (#p<0.05, ###p<0.001 vs. control at time of biopsy; *p<0.05, ***p<0.001 vs. AngII at time of biopsy, $<0.05 vs. control at time of sacrifice).</p

Complete reversion of Ang II-induced pre-fibrotic changes.

<p>Ang II infusion increased α-SMA mRNA (A) and protein (B) expression. At time of sacrifice, mRNA levels of Ang II-infused rats returned to control values. The same pattern holds through for α-SMA protein expression, with a complete reversion of pre-fibrotic changes after cessation of Ang II. Representative photomicrographs of α-SMA stained renal sections (C). (##p<0.01, ###p<0.001 vs. control at time of biopsy; *p<0.05, ***p<0.001 vs. Ang II at time of biopsy).</p

Effects of Ang II infusion and withdrawal on tubular damage.

<p>Havcr1 mRNA (A) and protein (B) levels were increased by Ang II infusion. After stopping Ang II infusion, recovery of proximal tubular damage was established, evidenced by control levels of Havcr1 mRNA and Kim-1 protein levels in Ang II infused rats. Representative photomicrographs of Kim-1 stained renal sections (C). (### p<0.001 vs. control at time of biopsy; ***p<0.001 vs. Ang II at time of biopsy).</p

Ang II induces hypertension, renal function loss and proteinuria with spontaneous recovery after Ang II withdrawal.

<p>Ang II infusion increased systolic blood pressure (A) and urinary protein levels (B). Furthermore Ang II infusion attenuated renal function as evidenced by a decrease in creatinine clearance (C) and an increase in plasma creatinine (D). Ang II infusion also caused an increase in plasma urea levels (E). When Ang II infusion was stopped, functional renal parameters of all rats turned to control values again. (##p<0.01, ###p<0.001 vs. control).</p

No focal segmental glomerular sclerosis after AngII infusion.

<p>Representative photomicrographs of PAS stained renal sections (A).</p