Volume 65, Issue 5, Pages (May 2004)

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Volume 65, Issue 5, Pages 1647-1653 (May 2004) Pioglitazone increases renal tubular cell albumin uptake but limits proinflammatory and fibrotic responses Stephen Zafiriou, Scott R. Stanners, Tania S. Polhill, Philip Poronnik, Carol A. Pollock Kidney International Volume 65, Issue 5, Pages 1647-1653 (May 2004) DOI: 10.1111/j.1523-1755.2004.00574.x Copyright © 2004 International Society of Nephrology Terms and Conditions

Figure 1 Peroxisome proliferator-activated receptor gamma (PPARγ) is expressed in opossum kidney cells. Western Blot of opossum kidney cell lysate showing that PPARγ is expressed in opossum kidney cells and has a molecular weight of ∼50 kD. Kidney International 2004 65, 1647-1653DOI: (10.1111/j.1523-1755.2004.00574.x) Copyright © 2004 International Society of Nephrology Terms and Conditions

Figure 2 Growth in opossum kidney cells exposed to pioglitazone with and without low-density lipoprotein (LDL) and/or albumin. Cell protein (A) and cell number (B) in opossum kidney cells exposed to the defined treatments for 24hours. Symbols are: (▪) 5mmol/L glucose (5Glc); 5mmol/L glucose with pioglitazone 10 μmol/L; LDL 100, LDL 100 μg/mL; albumin, albumin 1mg/mL; LDL + Alb, LDL 100 μg/mL + albumin 1mg/mL. Kidney International 2004 65, 1647-1653DOI: (10.1111/j.1523-1755.2004.00574.x) Copyright © 2004 International Society of Nephrology Terms and Conditions

Figure 3 Albumin uptake in opossum kidney cells. (A) Following 24hours' exposure to different concentrations of pioglitazone (0.1 to 10 μmol/L), opossum kidney cells were incubated with Texas Red–bovine serum albumin (BSA) (100 μg/mL) for 2hours, after which time the intracellular Texas Red–BSA uptake was measured and corrected for cell protein. Results are standardized against control conditions (opossum kidney cells in 5mmol/L glucose). (B) Albumin uptake in opossum kidney cells exposed to pioglitazone with and without low-density lipoprotein (LDL) and/or albumin. Following 24hours' exposure to the defined treatments, opossum kidney cells were incubated with Texas Red–BSA (100 μg/mL) for 2hours, after which time the intracellular Texas Red–BSA uptake was measured and corrected for cell protein. Symbols are: (▪) 5mmol/L glucose (5Glc); 5mmol/L glucose with pioglitazone 10 μmol/L; LDL 100, LDL 100 μg/mL; albumin, albumin 1mg/mL; LDL + Alb, LDL 100 μg/mL + albumin 1mg/mL. Kidney International 2004 65, 1647-1653DOI: (10.1111/j.1523-1755.2004.00574.x) Copyright © 2004 International Society of Nephrology Terms and Conditions

Figure 4 Transforming growth factor-β1 (TGF-β1) secretion in opossum kidney cells exposed to pioglitazone with and without low-density lipoprotein (LDL) and/or albumin. Active TGF-β1 levels were measured by enzyme-linked immunosorbent assay (ELISA) in media conditioned by opossum kidney cells exposed to the defined experimental treatments for 24hours. Symbols are: (▪) 5mmol/L glucose (5Glc); 5mmol/L glucose with pioglitazone 10 μmol/L; LDL 100, LDL 100 μg/mL; albumin, albumin 1mg/mL; LDL + Alb, LDL 100 μg/mL + albumin 1mg/mL. #P < 0.05 for LDL 100 μg/mL vs. LDL 100 μg/mL with pioglitazone 10 μmol/L. Kidney International 2004 65, 1647-1653DOI: (10.1111/j.1523-1755.2004.00574.x) Copyright © 2004 International Society of Nephrology Terms and Conditions

Figure 5 Monocyte chemoattractant protein-1 (MCP-1) secretion in opossum kidney cells exposed to pioglitazone with and without low-density lipoprotein (LDL) and/or albumin. MCP-1 levels were measured by enzyme-linked immunosorbent assay (ELISA) in media conditioned by opossum kidney cells exposed to the defined experimental treatments for 24hours. Symbols are: (▪) 5mmol/L glucose (5Glc); 5mmol/L glucose with pioglitazone 10 μmol/L; LDL 100, LDL 100 μg/mL; albumin, albumin 1mg/mL; LDL + Alb, LDL 100 μg/mL + albumin 1mg/mL. #P < 0.05 for LDL 100 μg/mL vs. LDL 100 μg/mL with pioglitazone 10 μmol/L. Kidney International 2004 65, 1647-1653DOI: (10.1111/j.1523-1755.2004.00574.x) Copyright © 2004 International Society of Nephrology Terms and Conditions

Figure 6 Nuclear factor-κB (NFκB)-secreted alkaline phosphatase (SEAP) activity in opossum kidney cells exposed to pioglitazone with and without low-density lipoprotein (LDL) and/or albumin. NF-κB–specific reporter gene assay. SEAP expression of opossum kidney cells exposed to the defined treatments for 24hours. Symbols are: (▪) 5mmol/L glucose (5Glc); 5mmol/L glucose with pioglitazone 10 μmol/L; LDL 100, LDL 100 μg/mL; albumin, albumin 1mg/mL; LDL + Alb, LDL 100 μg/mL + albumin 1mg/mL. Kidney International 2004 65, 1647-1653DOI: (10.1111/j.1523-1755.2004.00574.x) Copyright © 2004 International Society of Nephrology Terms and Conditions