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Podocyte-secreted angiopoietin-like-4 mediates proteinuria in glucocorticoid-sensitive nephrotic syndrome.

Clement LC, Avila-Casado C, Macé C, Soria E, Bakker WW, Kersten S, Chugh SS - Nat. Med. (2010)

Bottom Line: In this study, we show that the glomerular expression of angiopoietin-like-4 (Angptl4), a secreted glycoprotein, is glucocorticoid sensitive and is highly upregulated in the serum and in podocytes in experimental models of MCD and in the human disease.Podocyte-specific transgenic overexpression of Angptl4 (NPHS2-Angptl4) in rats induced nephrotic-range, and selective, proteinuria (over 500-fold increase in albuminuria), loss of glomerular basement membrane (GBM) charge and foot process effacement, whereas transgenic expression specifically in the adipose tissue (aP2-Angptl4) resulted in increased circulating Angptl4, but no proteinuria.When we fed the sialic acid precursor N-acetyl-D-mannosamine (ManNAc) to NPHS2-Angptl4 transgenic rats it increased the sialylation of Angptl4 and decreased albuminuria by more than 40%.

View Article: PubMed Central - PubMed

Affiliation: Glomerular Disease Therapeutics Laboratory, and Nephrology Research and Training Center, University of Alabama at Birmingham, Birmingham, Alabama, USA.

ABSTRACT
The main manifestations of nephrotic syndrome include proteinuria, hypoalbuminemia, edema, hyperlipidemia and lipiduria. Common causes of nephrotic syndrome are diabetic nephropathy, minimal change disease (MCD), focal and segmental glomerulosclerosis (FSGS) and membranous nephropathy. Among the primary glomerular diseases, MCD is usually sensitive to glucocorticoid treatment, whereas the other diseases show variable responses. Despite the identification of key structural proteins in the glomerular capillary loop which may contribute to defects in ultrafiltration, many of the disease mechanisms of nephrotic syndrome remain unresolved. In this study, we show that the glomerular expression of angiopoietin-like-4 (Angptl4), a secreted glycoprotein, is glucocorticoid sensitive and is highly upregulated in the serum and in podocytes in experimental models of MCD and in the human disease. Podocyte-specific transgenic overexpression of Angptl4 (NPHS2-Angptl4) in rats induced nephrotic-range, and selective, proteinuria (over 500-fold increase in albuminuria), loss of glomerular basement membrane (GBM) charge and foot process effacement, whereas transgenic expression specifically in the adipose tissue (aP2-Angptl4) resulted in increased circulating Angptl4, but no proteinuria. Angptl4(-/-) mice that were injected with lipopolysaccharide (LPS) or nephritogenic antisera developed markedly less proteinuria than did control mice. Angptl4 secreted from podocytes in some forms of nephrotic syndrome lacks normal sialylation. When we fed the sialic acid precursor N-acetyl-D-mannosamine (ManNAc) to NPHS2-Angptl4 transgenic rats it increased the sialylation of Angptl4 and decreased albuminuria by more than 40%. These results suggest that podocyte-secreted Angptl4 has a key role in nephrotic syndrome.

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Relationship of Angptl4 overexpression with proteinuria. (a) Albuminuria in female NPHS2-Angptl4 transgenic (TG) rats. (b) Albuminuria in male heterozygous NPHS2-Angptl4 transgenic rats (c) Albuminuria in male homozygous NPHS2-Angptl4 transgenic rats. (d) GelCode blue stained SDS PAGE of urinary protein from transgenic rats, rats with PAN, and individuals with minimal change disease (MCD) and membranous nephropathy (MN). Arrow points towards prominent 70 kDa intact albumin band. Mean percentage densitometry of intact albumin is shown for each lane (details in Supplementary Fig. 3d). (e) Proteinuria following induction of low dose PAN in heterozygous male NPHS2-Angptl4 transgenic and wild type littermates. (f) Proteinuria in Wistar rats treated with glucocorticoids (PAN-S) or PBS (PAN) on alternate days starting 1 day after induction of PAN. (g) Glomerular Angptl4 mRNA expression in PAN rats described in panel f. * P<0.05; ** P < 0.01, *** P<0.001.
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Figure 3: Relationship of Angptl4 overexpression with proteinuria. (a) Albuminuria in female NPHS2-Angptl4 transgenic (TG) rats. (b) Albuminuria in male heterozygous NPHS2-Angptl4 transgenic rats (c) Albuminuria in male homozygous NPHS2-Angptl4 transgenic rats. (d) GelCode blue stained SDS PAGE of urinary protein from transgenic rats, rats with PAN, and individuals with minimal change disease (MCD) and membranous nephropathy (MN). Arrow points towards prominent 70 kDa intact albumin band. Mean percentage densitometry of intact albumin is shown for each lane (details in Supplementary Fig. 3d). (e) Proteinuria following induction of low dose PAN in heterozygous male NPHS2-Angptl4 transgenic and wild type littermates. (f) Proteinuria in Wistar rats treated with glucocorticoids (PAN-S) or PBS (PAN) on alternate days starting 1 day after induction of PAN. (g) Glomerular Angptl4 mRNA expression in PAN rats described in panel f. * P<0.05; ** P < 0.01, *** P<0.001.

Mentions: We noted significant albuminuria in both NPHS2-Angptl4 TG rat founder lines. Female homozygous and male heterozygous rats developed albuminuria at age 1 month (Fig. 3a–3c, Supplementary Fig. 3c). Homozygous females developed up to 100-fold, heterozygous males up to 20-fold and homozygous males over 500-fold increase in albuminuria over time. Heterozygous females were not albuminuric. Over 90% of the urinary protein comprises of intact albumin (Fig. 3d, Supplementary Fig. 3d), thereby making these rats the first model of selective proteinuria. Blood pressure was significantly lower in proteinuric heterozygous NPHS2-Angptl4 transgenic rats compared to wild type controls (Supplementary Fig. 3e). By contrast, aP2-Angptl4 transgenic rats did not develop albuminuria (Fig. 3d), despite high circulating Angptl4 levels (Supplementary Fig. 4a–4d). Despite upregulation of Angptl4 in pancreas in NPHS2-Angptl4 transgenic rats, circulating Angptl4 levels were not elevated, and were in fact marginally reduced (Supplementary Fig. 4a–4d) due to presence of proteinuria in these rats. In keeping with the proteinuric effects of podocyte secreted Angptl4, NPHS2-Angptl4 transgenic rats developed more proteinuria (Fig. 3e) than wild type littermates after induction of PAN. Serum electrolytes, creatinine and glucose levels were indistinguishable from wild type rats.


Podocyte-secreted angiopoietin-like-4 mediates proteinuria in glucocorticoid-sensitive nephrotic syndrome.

Clement LC, Avila-Casado C, Macé C, Soria E, Bakker WW, Kersten S, Chugh SS - Nat. Med. (2010)

Relationship of Angptl4 overexpression with proteinuria. (a) Albuminuria in female NPHS2-Angptl4 transgenic (TG) rats. (b) Albuminuria in male heterozygous NPHS2-Angptl4 transgenic rats (c) Albuminuria in male homozygous NPHS2-Angptl4 transgenic rats. (d) GelCode blue stained SDS PAGE of urinary protein from transgenic rats, rats with PAN, and individuals with minimal change disease (MCD) and membranous nephropathy (MN). Arrow points towards prominent 70 kDa intact albumin band. Mean percentage densitometry of intact albumin is shown for each lane (details in Supplementary Fig. 3d). (e) Proteinuria following induction of low dose PAN in heterozygous male NPHS2-Angptl4 transgenic and wild type littermates. (f) Proteinuria in Wistar rats treated with glucocorticoids (PAN-S) or PBS (PAN) on alternate days starting 1 day after induction of PAN. (g) Glomerular Angptl4 mRNA expression in PAN rats described in panel f. * P<0.05; ** P < 0.01, *** P<0.001.
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Figure 3: Relationship of Angptl4 overexpression with proteinuria. (a) Albuminuria in female NPHS2-Angptl4 transgenic (TG) rats. (b) Albuminuria in male heterozygous NPHS2-Angptl4 transgenic rats (c) Albuminuria in male homozygous NPHS2-Angptl4 transgenic rats. (d) GelCode blue stained SDS PAGE of urinary protein from transgenic rats, rats with PAN, and individuals with minimal change disease (MCD) and membranous nephropathy (MN). Arrow points towards prominent 70 kDa intact albumin band. Mean percentage densitometry of intact albumin is shown for each lane (details in Supplementary Fig. 3d). (e) Proteinuria following induction of low dose PAN in heterozygous male NPHS2-Angptl4 transgenic and wild type littermates. (f) Proteinuria in Wistar rats treated with glucocorticoids (PAN-S) or PBS (PAN) on alternate days starting 1 day after induction of PAN. (g) Glomerular Angptl4 mRNA expression in PAN rats described in panel f. * P<0.05; ** P < 0.01, *** P<0.001.
Mentions: We noted significant albuminuria in both NPHS2-Angptl4 TG rat founder lines. Female homozygous and male heterozygous rats developed albuminuria at age 1 month (Fig. 3a–3c, Supplementary Fig. 3c). Homozygous females developed up to 100-fold, heterozygous males up to 20-fold and homozygous males over 500-fold increase in albuminuria over time. Heterozygous females were not albuminuric. Over 90% of the urinary protein comprises of intact albumin (Fig. 3d, Supplementary Fig. 3d), thereby making these rats the first model of selective proteinuria. Blood pressure was significantly lower in proteinuric heterozygous NPHS2-Angptl4 transgenic rats compared to wild type controls (Supplementary Fig. 3e). By contrast, aP2-Angptl4 transgenic rats did not develop albuminuria (Fig. 3d), despite high circulating Angptl4 levels (Supplementary Fig. 4a–4d). Despite upregulation of Angptl4 in pancreas in NPHS2-Angptl4 transgenic rats, circulating Angptl4 levels were not elevated, and were in fact marginally reduced (Supplementary Fig. 4a–4d) due to presence of proteinuria in these rats. In keeping with the proteinuric effects of podocyte secreted Angptl4, NPHS2-Angptl4 transgenic rats developed more proteinuria (Fig. 3e) than wild type littermates after induction of PAN. Serum electrolytes, creatinine and glucose levels were indistinguishable from wild type rats.

Bottom Line: In this study, we show that the glomerular expression of angiopoietin-like-4 (Angptl4), a secreted glycoprotein, is glucocorticoid sensitive and is highly upregulated in the serum and in podocytes in experimental models of MCD and in the human disease.Podocyte-specific transgenic overexpression of Angptl4 (NPHS2-Angptl4) in rats induced nephrotic-range, and selective, proteinuria (over 500-fold increase in albuminuria), loss of glomerular basement membrane (GBM) charge and foot process effacement, whereas transgenic expression specifically in the adipose tissue (aP2-Angptl4) resulted in increased circulating Angptl4, but no proteinuria.When we fed the sialic acid precursor N-acetyl-D-mannosamine (ManNAc) to NPHS2-Angptl4 transgenic rats it increased the sialylation of Angptl4 and decreased albuminuria by more than 40%.

View Article: PubMed Central - PubMed

Affiliation: Glomerular Disease Therapeutics Laboratory, and Nephrology Research and Training Center, University of Alabama at Birmingham, Birmingham, Alabama, USA.

ABSTRACT
The main manifestations of nephrotic syndrome include proteinuria, hypoalbuminemia, edema, hyperlipidemia and lipiduria. Common causes of nephrotic syndrome are diabetic nephropathy, minimal change disease (MCD), focal and segmental glomerulosclerosis (FSGS) and membranous nephropathy. Among the primary glomerular diseases, MCD is usually sensitive to glucocorticoid treatment, whereas the other diseases show variable responses. Despite the identification of key structural proteins in the glomerular capillary loop which may contribute to defects in ultrafiltration, many of the disease mechanisms of nephrotic syndrome remain unresolved. In this study, we show that the glomerular expression of angiopoietin-like-4 (Angptl4), a secreted glycoprotein, is glucocorticoid sensitive and is highly upregulated in the serum and in podocytes in experimental models of MCD and in the human disease. Podocyte-specific transgenic overexpression of Angptl4 (NPHS2-Angptl4) in rats induced nephrotic-range, and selective, proteinuria (over 500-fold increase in albuminuria), loss of glomerular basement membrane (GBM) charge and foot process effacement, whereas transgenic expression specifically in the adipose tissue (aP2-Angptl4) resulted in increased circulating Angptl4, but no proteinuria. Angptl4(-/-) mice that were injected with lipopolysaccharide (LPS) or nephritogenic antisera developed markedly less proteinuria than did control mice. Angptl4 secreted from podocytes in some forms of nephrotic syndrome lacks normal sialylation. When we fed the sialic acid precursor N-acetyl-D-mannosamine (ManNAc) to NPHS2-Angptl4 transgenic rats it increased the sialylation of Angptl4 and decreased albuminuria by more than 40%. These results suggest that podocyte-secreted Angptl4 has a key role in nephrotic syndrome.

Show MeSH
Related in: MedlinePlus