Supplementary Material for: Dietary Phosphate Binding and Loading Alter Kidney Angiotensin-Converting Enzyme mRNA and Protein Content in 5/6 Nephrectomized Rats

<i>Background:</i> Vitamin D receptor activation with paricalcitol can modulate the transcription of renin-angiotensin system components in the surgical 5/6 nephrectomy rat model (5/6 NX) of chronic renal insufficiency. We tested the hypothesis whether dietary modification of phosphate influences kidney renin-angiotensin system gene expression at the mRNA level in 5/6 NX rats. <i>Methods:</i> Fifteen weeks after surgery, rats were given control diet (0.3% calcium, 0.5% phosphate), phosphate-lowering diet (3% calcium as carbonate) or high-phosphate diet (1.5%) for 12 weeks. Sham-operated rats were on control diet. <i>Results:</i> Blood pressure, plasma phosphate, parathyroid hormone, glomerulosclerosis, tubulointerstitial damage, and FGF-23 were increased in remnant kidney rats, whereas creatinine clearance was decreased. Phosphate, parathyroid hormone, glomerulosclerosis, tubulointerstitial damage, and FGF-23 were further elevated by the high-phosphate diet, but were reduced by the phosphate-lowering diet. Plasma calcium was increased with the phosphate-lowering diet and decreased with the high-phosphate diet. Remnant kidney rats on control diet showed upregulated kidney angiotensin-converting enzyme (ACE) and angiotensin (Ang) IV receptor (AT<sub>4</sub>) transcription, while ACE2, Ang II type 2 receptor and renin receptor transcription were downregulated in comparison with sham rats. Phosphate-lowering diet reduced whereas high-phosphate diet increased kidney ACE, and these effects were observed at both mRNA and protein levels. Dietary phosphate loading also resulted in lower AT<sub>1a</sub> gene transcription. <i>Conclusion:</i> Dietary phosphate loading was associated with elevated kidney ACE expression, increased tissue damage and lower AT<sub>1a</sub> transcription in 5/6 NX rats. Phosphate binding with 3% calcium carbonate had opposite effects on ACE and kidney damage.