Receptor-Mediated Endocytosis of α-Galactosidase A in Human Podocytes in Fabry Disease

Prabakaran, Thaneas; Nielsen, Rikke; Larsen, Jakob Vejby; Sørensen, Søren Schwartz; Rasmussen, Ulla Feldt; Saleem, Moin A.; Petersen, Claus Munck; Verroust, P; Christensen, Erik I.

doi:10.1371/journal.pone.0025065

Cited by 111 publications

(108 citation statements)

References 67 publications

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“…Gentamicin uptake was also significantly reduced in the presence of the megalin inhibitors RAP and EDTA suggesting that megalin is likely involved in the uptake process. Our data are consistent with those from previous investigations, which demonstrated a 50% reduction in the amount of gentamicin accumulation in RAP-deficient kidneys (24) and an increase in gentamicin excretion in rat perfused proximal tubules in the presence of RAP (25). Sodium maleate, a megalin-shedding agent, also caused a significant decrease in gentamicin uptake (30-45%) providing further evidence that megalin is involved in the uptake process.…”

Section: Discussionsupporting

confidence: 93%

“…This suggests that mechanisms other than megalin-mediated endocytosis may be involved in hepatic AG uptake. Since the liver is not a physiologic site of AG-induced toxicity or pharmacologic action, however, the uptake of AGs by hepatocytes has not been extensively studied and data are limited to a small number of studies in animal models which also show limited hepatic uptake after intravenous administration (25)(26)(27). Hepatocytes may therefore be a model that can be used to investigate non-megalin-mediated mechanisms of AG transport such as passive diffusion and organic cationic transport.…”

Section: Discussionmentioning

confidence: 99%

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The Role of Megalin in the Transport of Gentamicin Across BeWo Cells, an In Vitro Model of the Human Placenta

Akour

Kennedy

Gerk

2015

AAPS J

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Abstract. Aminoglycosides (AG) are known to readily cross the placenta, although the mechanisms responsible for placental transport have not been characterized. Megalin is expressed in human placenta, and it is reasonable to speculate, given its role in renal AG uptake, that it is similarly involved in placental transport. However, the role of megalin in placental AG uptake has not been established. An in vitro model to study megalin-mediated placental transport has also not been previously described. The objectives of this study, therefore, were to evaluate the human choriocarcinoma (BeWo) cell line as a model to study megalin-mediated placental transport and to assess the uptake kinetics of gentamicin, an AG antibiotic, using this in vitro model. BeWo cells were grown on Transwell® plates, and megalin expression and functional activity were assessed. Uptake of 3 H-gentamicin was also evaluated in the presence and absence of megalin inhibitors. Expression of megalin protein and mRNA in BeWo cells were confirmed via immunoblot and qPCR analysis. Uptake of fluorescein isothiocyanate (FITC)-labeled bovine serum albumin (BSA) (a megalin substrate) was time-, concentration-, and temperature-dependent consistent with a transporter-mediated process. FITC-BSA uptake was also significantly reduced in the presence of unlabeled gentamicin (a megalin substrate) and sodium maleate (to induce megalin shedding) suggesting that megalin is functionally active in BeWo cells. Gentamicin uptake exhibited time and temperature dependence, saturability and Michaelis-Menten kinetics, all of which suggest a transporter-mediated process. Gentamicin uptake was also significantly reduced in the presence of the megalin inhibitors RAP and EDTA suggesting that megalin is likely involved in gentamicin uptake.

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Section: Discussionsupporting

confidence: 93%

Section: Discussionmentioning

confidence: 99%

The Role of Megalin in the Transport of Gentamicin Across BeWo Cells, an In Vitro Model of the Human Placenta

Akour

Kennedy

Gerk

2015

AAPS J

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“…The expression of megalin and cubilin has been localized in rat and human podocytes, [31][32][33] and subepithelial immune deposits were observed in rats infused with antimegalin IgG. 16 Our immunohistochemical data are consistent with these studies and confirm the expression of megalin in human and rat podocytes.…”

Section: Discussionsupporting

confidence: 85%

Intravital Imaging Reveals Angiotensin II–Induced Transcytosis of Albumin by Podocytes

Schießl¹,

Hammer²,

Kattler³

et al. 2016

Journal of the American Society of Nephrology

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Albuminuria is a hallmark of kidney disease of various etiologies and usually caused by deterioration of glomerular filtration barrier integrity. We recently showed that angiotensin II (Ang II) acutely increases albumin filtration in the healthy kidney. Here, we used intravital microscopy to assess the effects of Ang II on podocyte function in rats. Acute infusion of 30, 60, or 80 ng/kg per minute Ang II enhanced the endocytosis of albumin by activation of the type 1 Ang II receptor and resulted in an average (6SEM) of 3.762.2, 72.3618.6 (P,0.001), and 239.4634.6 mm 3 (P,0.001) albumin-containing vesicles per glomerulus, respectively, compared with none at baseline or 10 ng/kg per minute Ang II. Immunostaining of Ang II-infused kidneys confirmed the presence of albumin-containing vesicles, which colocalized with megalin, in podocin-positive cells. Furthermore, podocyte endocytosis of albumin was markedly reduced in the presence of gentamicin, a competitive inhibitor of megalin-dependent endocytosis. Ang II infusion increased the concentration of albumin in the subpodocyte space, a potential source for endocytic protein uptake, and gentamicin further increased this concentration. Some endocytic vesicles were acidified and colocalized with LysoTracker. Most vesicles migrated from the capillary to the apical aspect of the podocyte and were eventually released into the urinary space. This transcytosis accounted for approximately 10% of total albumin filtration. In summary, the transcellular transport of proteins across the podocyte constitutes a new pathway of glomerular protein filtration. Ang II enhances the endocytosis and transcytosis of plasma albumin by podocytes, which may eventually impair podocyte function.

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“…20 Furthermore, new receptors involved in dosedependent agalsidase uptake in human podocytes have recently been described. 21 Taken together, these observations indicate a pivotal role of podocytes in early progression of nephropathy.…”

Section: Discussionmentioning

confidence: 82%

Agalsidase Benefits Renal Histology in Young Patients with Fabry Disease

Tøndel

Bostad

Larsen

et al. 2013

Journal of the American Society of Nephrology

223

212

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The effect of early-onset enzyme replacement therapy on renal morphologic features in Fabry disease is largely unknown. Here, we evaluated the effect of 5 years of treatment with agalsidase alfa or agalsidase beta in 12 consecutive patients age 7-33 years (median age, 16.5 years). We performed renal biopsies at baseline and after 5 years of enzyme replacement therapy; 7 patients had additional biopsies after 1 and 3 years. After a median of 65 months, biopsy findings from all patients showed total clearance of glomerular endothelial and mesangial cell inclusions, and findings from 2 patients showed complete clearance of inclusions from epithelial cells of the distal tubule. The 4 patients who received the highest dose of agalsidase exhibited substantial clearance of podocyte inclusions, and the youngest patient had nearly complete clearance of these inclusions. Linear regression analysis showed a highly significant correlation between podocyte globotriaocylceramide clearance and cumulative agalsidase dose (r50.804; P50.002). Microalbuminuria normalized in five patients. In summary, long-term enzyme replacement therapy in young patients can result in complete globotriaocylceramide clearance of mesangial and glomerular endothelial cells across all dosage regimens, and clearance of podocyte inclusions is dosedependent.

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Receptor-Mediated Endocytosis of α-Galactosidase A in Human Podocytes in Fabry Disease

Cited by 111 publications

References 67 publications

The Role of Megalin in the Transport of Gentamicin Across BeWo Cells, an In Vitro Model of the Human Placenta

The Role of Megalin in the Transport of Gentamicin Across BeWo Cells, an In Vitro Model of the Human Placenta

Intravital Imaging Reveals Angiotensin II–Induced Transcytosis of Albumin by Podocytes

Agalsidase Benefits Renal Histology in Young Patients with Fabry Disease

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