Volume 64, Issue 2, Pages (August 2003)

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Volume 64, Issue 2, Pages 555-564 (August 2003) Thiazide-induced hypocalciuria is accompanied by a decreased expression of Ca2+ transport proteins in kidney Tom Nijenhuis, Joost G.J. Hoenderop, Johannes Loffing, Annemiete W.C.M. van der Kemp, Carel H. van Os, René J.M. Bindels Kidney International Volume 64, Issue 2, Pages 555-564 (August 2003) DOI: 10.1046/j.1523-1755.2003.00128.x Copyright © 2003 International Society of Nephrology Terms and Conditions

Figure 1 Effect of hydrochlorothiazide (HCTZ) and extracellular volume (ECV) status on mRNA expression levels of distal convoluted tubule (DCT) transport proteins. mRNA expression levels of TRPV5 (A), calbindin-D28K (B), Na+ Cl- cotransporter (NCC) (C), and Na+/Ca2+ exchanger (NCX1) (D) in kidney cortex were determined by real-time quantitative polymerase chain reaction (PCR). Results are presented relative to controls. Controls, sham-operated animals; HCTZ, 12mg/24hours HCTZ; HCTZ/Na+-repleted, 12mg/24hours HCTZ supplemented with NaCl/KCl via the drinking water; Na+-depleted, animals fed the low-sodium diet [0.02% (wt/vol) NaCl]; and HCTZ/Na+-depleted, Na+-depleted animals receiving 12mg/24hours HCTZ. Data are presented as means ± SEM. *P < 0.05 vs. controls. Kidney International 2003 64, 555-564DOI: (10.1046/j.1523-1755.2003.00128.x) Copyright © 2003 International Society of Nephrology Terms and Conditions

Figure 2 Effect of hydrochlorothiazide (HCTZ) and extracellular volume (ECV) status on protein abundance of distal convoluted tubule (DCT) transport proteins. Protein expression of TRPV5 (A), calbindin-D28K (B), and Na+ Cl- cotransporter (NCC) (C) in kidney cortex were determined by immunohistochemistry and presented relative to controls. Controls, sham-operated animals; HCTZ, 12mg/24hours HCTZ; HCTZ/Na+-repleted, 12mg/24hours HCTZ supplemented with NaCl/KCl via the drinking water; Na+-depleted, animals fed the low-sodium diet [0.02% (wt/vol) NaCl]; and HCTZ/Na+-depleted, Na+-depleted animals receiving 12mg/24hours HCTZ. Data are presented as means ± SEM. *P < 0.05 vs. controls. Kidney International 2003 64, 555-564DOI: (10.1046/j.1523-1755.2003.00128.x) Copyright © 2003 International Society of Nephrology Terms and Conditions

Figure 3 Disturbed distribution of the Na+ Cl- cotransporter (NCC) in hydrochlorothiazide (HCTZ)-treated animals. Detail of distal convoluted tubule (DCT) showing NCC immunopositive staining of control (A) and HCTZ-treated (B) rats demonstrating that the characteristic apical NCC signal as shown in the controls is lost in a portion of tubules present in the HCTZ-treated rats. Instead, a diffuse intracellular appearance is visible (arrows). Kidney International 2003 64, 555-564DOI: (10.1046/j.1523-1755.2003.00128.x) Copyright © 2003 International Society of Nephrology Terms and Conditions

Figure 4 Hydrochlorothiazide (HCTZ) treatment reduced the amount of Na+ Cl- cotransporter (NCC) and calbindin-D28K coexpressing tubules. Number of tubules coexpressing NCC and calbindin-D28K was determined by immunohistochemistry and presented relative to controls. Controls, sham-operated animals; HCTZ, 12mg/24hours HCTZ; HCTZ/Na+-repleted, 12mg/24hours HCTZ supplemented with NaCl/KCl via the drinking water; Na+-depleted, animals fed the low-sodium diet [0.02% (wt/vol) NaCl]; and HCTZ/Na+-depleted, Na+-depleted animals receiving 12mg/24hours HCTZ. Data are presented as mean ± SEM. *P < 0.05 vs. controls. Kidney International 2003 64, 555-564DOI: (10.1046/j.1523-1755.2003.00128.x) Copyright © 2003 International Society of Nephrology Terms and Conditions

Figure 5 Hydrochlorothiazide (HCTZ) treatment reduced calbindin-D28K protein abundance as determined by immunoblotting. Immunoblots for calbindin-D28K of individual kidney samples. Controls, sham-operated animals; HCTZ, 12mg/24hours HCTZ; HCTZ/Na+-repleted, 12mg/24hours HCTZ supplemented with NaCl/KCl via the drinking water; Na+-depleted, animals fed the low-sodium diet [0.02% (wt/vol) NaCl]; and HCTZ/Na+-depleted, Na+-depleted animals receiving 12mg/24hours HCTZ. Kidney International 2003 64, 555-564DOI: (10.1046/j.1523-1755.2003.00128.x) Copyright © 2003 International Society of Nephrology Terms and Conditions