The renin inhibitor aliskiren attenuates high-glucose induced extracellular matrix synthesis and prevents apoptosis in cultured podocytes - PubMed (original) (raw)

The renin inhibitor aliskiren attenuates high-glucose induced extracellular matrix synthesis and prevents apoptosis in cultured podocytes

Lynetta M Phillips et al. Nephron Exp Nephrol. 2011.

Abstract

Background/aims: Altered extracellular matrix (ECM) remodeling and podocyte apoptosis are characteristic features of diabetic nephropathy (DN). Aliskiren (ALI) inhibits the renin-catalyzed conversion of angiotensinogen to angiotensin I. This study tested ALI's effect on podocyte ECM accretion and survival in a high-glucose environment in vitro.

Methods: Conditionally immortalized mouse podocytes were incubated in normal glucose (NG; 5.5 mM) or high glucose (HG; 40 mM) for 24-48 h with and without ALI (20 nM). Real-time RT-PCR was performed for fibronectin (FN), collagen α5(type IV) (Cola5IV), matrix metalloproteinases 2 and 9 (MMP2 and MMP9), and tissue inhibitor of metalloproteinases 1 and 2 (TIMP1 and TIMP2). Western blots were performed for FN, Cola5IV, MMP2, MMP9, TIMP1 and cleaved (activated) caspase-3.

Results: ALI significantly reduced the mRNA and protein levels of FN, Cola5IV and TIMP1, and the mRNA of TIMP2 and cleaved caspase-3. ALI had no effect on MMP2 mRNA or protein or MMP9 mRNA tested under HG conditions. Under NG conditions, ALI had no effect on FN, Cola5IV, MMP2, MMP9 and activated caspase-3 proteins. ALI decreased the activated caspase-3 protein and evidence of apoptosis by TUNEL staining observed in podocytes cultured under HG conditions.

Conclusion: These results show for the first time that renin inhibition with ALI mitigates the profibrotic and apoptotic effects of HG in cultured podocytes. These data strengthen the therapeutic rationale for renin inhibition with ALI beyond its hemodynamic effects.

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Conflict of interest statement

Novartis Institutes for Biomedical Research provided the ALI and paid for the supplies utilized in these experiments. Novartis did not pay the salaries of authors L.M.P., Y.W., T.D., J.L. or S.G.A. D.L.F. is an employee of Novartis. This work was supported in part by a grant from Novartis Pharmaceuticals.

Figures

Fig. 1

FN mRNA and protein under NG and HG conditions with and without ALI in podocytes. a FN mRNA in NG (5.5 m_M_ glucose), HG (40 m_M_ glucose) and HG + ALI (40 m_M_ glucose + 20 m_M_ ALI), n = 3. b FN protein in NG (5.5 m_M_ glucose), HG (40 m_M_ glucose) and HG + ALI (40 m_M_ glucose + 20 m_M_ ALI), n = 6. c In NG, ALI did not increase FN protein, nor was FN protein changed by isotonic mannitol (5.5 m_M_ glucose + 34.5 m_M_ mannitol, n = 3 in each group). * p ≤ 0.05 vs. HG; ** p < 0.05 vs. NG.

Fig. 2

Cola5IV in podocytes. a Cola5IV mRNA in NG (5.5 m_M_ glucose), HG (40 m_M_ glucose) and HG + ALI (40 m_M_ glucose + 20 m_M_ ALI), n = 3. b Cola5IV protein in NG (5.5 m_M_ glucose), HG (40 m_M_ glucose) and HG + ALI (40 m_M_ glucose + 20 m_M_ ALI), n = 6. c In NG, ALI did not increase Cola5IV protein, nor was Cola5IV protein changed by isotonic mannitol (5.5 m_M_ glucose + 34.5 m_M_ mannitol, n = 3 in each group). ** p < 0.05 vs. NG, * p < 0.05 vs. HG.

Fig. 3

24-hour MMP2 in podocytes. a MMP2 mRNA in NG (5.5 m_M_ glucose), HG (40 m_M_ glucose) and HG + ALI (40 m_M_ glucose + 20 m_M_ ALI), n = 3. b Activated MMP2 protein in NG (5.5 m_M_ glucose), HG (40 m_M_ glucose) and HG + ALI (40 m_M_ glucose + 20 m_M_ ALI), n = 3. c In NG, ALI did not reduce MMP2 protein, nor was MMP2 protein changed by isotonic mannitol (5.5 m_M_ glucose + 34.5 m_M_ mannitol, n = 3 in each group).

Fig. 4

24-hour TIMP2 in podocytes. TIMP2 mRNA in NG (5.5 m_M_ glucose), HG (40 m_M_ glucose) and HG + ALI (40 m_M_ glucose + 20 m_M_ ALI), n = 3. * p < 0.05 vs. HG.

Fig. 5

24-hour MMP9 in podocytes. a MMP9 mRNA in NG (5.5 m_M_ glucose), HG (40 m_M_ glucose) and HG + ALI (40 m_M_ glucose + 20 m_M_ ALI), n = 3. b MMP9 protein in NG (5.5 m_M_ glucose), HG (40 m_M_ glucose) and HG + ALI (40 m_M_ glucose + 20 m_M_ ALI), n = 3. c In NG, ALI did not reduce MMP9 protein, nor was MMP9 protein changed by isotonic mannitol (5.5 m_M_ glucose + 34.5 m_M_ mannitol, n = 3 in each group).

Fig. 6

24-hour TIMP1 in podocytes. a TIMP1 mRNA in NG (5.5 m_M_ glucose), HG (40 m_M_ glucose) and HG + ALI (40 m_M_ glucose + 20 m_M_ ALI), n = 3. b HG (40 m_M_) induced no significant change in TIMP1 protein vs. NG (5.5 m_M_), but ALI significantly lowered TIMP1 protein under HG culture conditions (40 m_M_ glucose + 20 m_M_ ALI), n = 3. * p < 0.05 vs. HG.

Fig. 7

48-hour cleaved caspase-3 in podocytes. a Cleaved caspase-3 protein in NG (5.5 m_M_ glucose), HG (40 m_M_ glucose) and HG + ALI (40 m_M_ glucose + 20 m_M_ ALI), n = 3. b In NG, ALI did not reduce cleaved caspase-3 protein, nor was cleaved caspase-3 protein changed by isotonic mannitol (5.5 m_M_ glucose + 34.5 m_M_ mannitol, n = 3 in each group). c TUNEL staining showed a significant increment in apoptotic podocytes in HG vs. NG, which was significantly attenuated by ALI. * p < 0.05 vs. HG; ** p < 0.05 vs. NG.

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The renin inhibitor aliskiren attenuates high-glucose induced extracellular matrix synthesis and prevents apoptosis in cultured podocytes - PubMed (original) (raw)