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Efficient uptake of blood-borne BK and JC polyomavirus-like particles in endothelial cells of liver sinusoids and renal vasa recta.

Simon-Santamaria J, Rinaldo CH, Kardas P, Li R, Malovic I, Elvevold K, McCourt P, Smedsrød B, Hirsch HH, Sørensen KK - PLoS ONE (2014)

Bottom Line: Most VLP-positive vessels in renal medulla did not express PV-1/Meca 32, suggesting location to the non-fenestrated part of vasa recta.The endothelial cells of these vessels also efficiently endocytosed a scavenger receptor ligand, formaldehyde-denatured albumin, suggesting high endocytic activity compared to other renal endothelia.We conclude that LSECs very effectively cleared a large fraction of blood-borne BK- and JC-VLPs, indicating a central role of these cells in early removal of polyomavirus from the circulation.

View Article: PubMed Central - PubMed

Affiliation: Department of Medical Biology, UiT - The Arctic University of Norway, Tromsø, Norway.

ABSTRACT
Liver sinusoidal endothelial cells (LSECs) are specialized scavenger cells that mediate high-capacity clearance of soluble waste macromolecules and colloid material, including blood-borne adenovirus. To explore if LSECs function as a sink for other viruses in blood, we studied the fate of virus-like particles (VLPs) of two ubiquitous human DNA viruses, BK and JC polyomavirus, in mice. Like complete virions, VLPs specifically bind to receptors and enter cells, but unlike complete virions, they cannot replicate. 125I-labeled VLPs were used to assess blood decay, organ-, and hepatocellular distribution of ligand, and non-labeled VLPs to examine cellular uptake by immunohisto- and -cytochemistry. BK- and JC-VLPs rapidly distributed to liver, with lesser uptake in kidney and spleen. Liver uptake was predominantly in LSECs. Blood half-life (∼1 min), and tissue distribution of JC-VLPs and two JC-VLP-mutants (L55F and S269F) that lack sialic acid binding affinity, were similar, indicating involvement of non-sialic acid receptors in cellular uptake. Liver uptake was not mediated by scavenger receptors. In spleen, the VLPs localized to the red pulp marginal zone reticuloendothelium, and in kidney to the endothelial lining of vasa recta segments, and the transitional epithelium of renal pelvis. Most VLP-positive vessels in renal medulla did not express PV-1/Meca 32, suggesting location to the non-fenestrated part of vasa recta. The endothelial cells of these vessels also efficiently endocytosed a scavenger receptor ligand, formaldehyde-denatured albumin, suggesting high endocytic activity compared to other renal endothelia. We conclude that LSECs very effectively cleared a large fraction of blood-borne BK- and JC-VLPs, indicating a central role of these cells in early removal of polyomavirus from the circulation. In addition, we report the novel finding that a subpopulation of endothelial cells in kidney, the main organ of polyomavirus persistence, showed selective and rapid uptake of VLPs, suggesting a role in viremic organ tropism.

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Electron microscopy of BK- and JC-virus like particles (VLPs).The figure show transmission electron micrographs of negatively stained BK- and JC-VLPs constructed from BK-VP1 or JC-VP1 proteins: A) BK-VP1 VLPs (Dunlop strain); B) JC-VP1-Mad-1 VLPs; C) JC-VP1-L55F VLPs; D) JC-VP1-S269F VLPs. Scale bars: 200 nm.
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pone-0111762-g001: Electron microscopy of BK- and JC-virus like particles (VLPs).The figure show transmission electron micrographs of negatively stained BK- and JC-VLPs constructed from BK-VP1 or JC-VP1 proteins: A) BK-VP1 VLPs (Dunlop strain); B) JC-VP1-Mad-1 VLPs; C) JC-VP1-L55F VLPs; D) JC-VP1-S269F VLPs. Scale bars: 200 nm.

Mentions: BK- and JC-VLPs were prepared as described previously [48]. Briefly, BK- and JC-VLPs were isolated from Sf9 insect cells infected with recombinant baculovirus encoding Dunlop BK-VP1 or Mad-1 JC-VP1 capsid protein. Recombinant baculoviruses containing PML variants of JC VP1 sequences L55F or S269F were obtained by site specific mutagenesis PCR using the following primers: L55F-Fwd (5′-ACC CAG ATG AGC ATT TTA GGG GTT TTA G-3′) and L55F-Rev (5′-CTA AAA CCC CTA AAA TGC TCA TCT GGG T-3′), S269F-Fwd (5′-TAA CAG ATC TGG TTT CCA GCA GTG GAG A-3′) and S269F-Rev (5′-TCT CCA CTG CTG GAA ACC AGA TCT GTT A-3′), respectively. Infected Sf9 cells were disrupted by sonication and glass mortar and pestle treatment. VLPs were purified from cellular lysate by CsCl gradient and stored in buffer consisting of 150 mM NaCl, 10 mM Tris-HCl pH 7.4 and 1.0 mM CaCl2. The 3-dimensional structure and integrity of VLPs was confirmed by transmission electron microscopy (Figure 1). In short, JC-VLPs or BK-VLPs in 0.9% NaCl were placed on discharged pallodium coated mesh grids for 2 min; then washed twice with double distilled water, and once with freshly made 2% uranyl acetate. Another drop of 2% uranyl acetate was added for 10 sec. The grids were gently washed on drops of double distilled water and air dried before electron microscopy. The images were recorded in a FEI CM100 TEM transmission electron microscope (FEI, OR, USA), operating at 80 kV, and equipped with Veleta digital CCD camera (Olympus, Germany).


Efficient uptake of blood-borne BK and JC polyomavirus-like particles in endothelial cells of liver sinusoids and renal vasa recta.

Simon-Santamaria J, Rinaldo CH, Kardas P, Li R, Malovic I, Elvevold K, McCourt P, Smedsrød B, Hirsch HH, Sørensen KK - PLoS ONE (2014)

Electron microscopy of BK- and JC-virus like particles (VLPs).The figure show transmission electron micrographs of negatively stained BK- and JC-VLPs constructed from BK-VP1 or JC-VP1 proteins: A) BK-VP1 VLPs (Dunlop strain); B) JC-VP1-Mad-1 VLPs; C) JC-VP1-L55F VLPs; D) JC-VP1-S269F VLPs. Scale bars: 200 nm.
© Copyright Policy
Related In: Results  -  Collection

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Show All Figures
getmorefigures.php?uid=PMC4222947&req=5

pone-0111762-g001: Electron microscopy of BK- and JC-virus like particles (VLPs).The figure show transmission electron micrographs of negatively stained BK- and JC-VLPs constructed from BK-VP1 or JC-VP1 proteins: A) BK-VP1 VLPs (Dunlop strain); B) JC-VP1-Mad-1 VLPs; C) JC-VP1-L55F VLPs; D) JC-VP1-S269F VLPs. Scale bars: 200 nm.
Mentions: BK- and JC-VLPs were prepared as described previously [48]. Briefly, BK- and JC-VLPs were isolated from Sf9 insect cells infected with recombinant baculovirus encoding Dunlop BK-VP1 or Mad-1 JC-VP1 capsid protein. Recombinant baculoviruses containing PML variants of JC VP1 sequences L55F or S269F were obtained by site specific mutagenesis PCR using the following primers: L55F-Fwd (5′-ACC CAG ATG AGC ATT TTA GGG GTT TTA G-3′) and L55F-Rev (5′-CTA AAA CCC CTA AAA TGC TCA TCT GGG T-3′), S269F-Fwd (5′-TAA CAG ATC TGG TTT CCA GCA GTG GAG A-3′) and S269F-Rev (5′-TCT CCA CTG CTG GAA ACC AGA TCT GTT A-3′), respectively. Infected Sf9 cells were disrupted by sonication and glass mortar and pestle treatment. VLPs were purified from cellular lysate by CsCl gradient and stored in buffer consisting of 150 mM NaCl, 10 mM Tris-HCl pH 7.4 and 1.0 mM CaCl2. The 3-dimensional structure and integrity of VLPs was confirmed by transmission electron microscopy (Figure 1). In short, JC-VLPs or BK-VLPs in 0.9% NaCl were placed on discharged pallodium coated mesh grids for 2 min; then washed twice with double distilled water, and once with freshly made 2% uranyl acetate. Another drop of 2% uranyl acetate was added for 10 sec. The grids were gently washed on drops of double distilled water and air dried before electron microscopy. The images were recorded in a FEI CM100 TEM transmission electron microscope (FEI, OR, USA), operating at 80 kV, and equipped with Veleta digital CCD camera (Olympus, Germany).

Bottom Line: Most VLP-positive vessels in renal medulla did not express PV-1/Meca 32, suggesting location to the non-fenestrated part of vasa recta.The endothelial cells of these vessels also efficiently endocytosed a scavenger receptor ligand, formaldehyde-denatured albumin, suggesting high endocytic activity compared to other renal endothelia.We conclude that LSECs very effectively cleared a large fraction of blood-borne BK- and JC-VLPs, indicating a central role of these cells in early removal of polyomavirus from the circulation.

View Article: PubMed Central - PubMed

Affiliation: Department of Medical Biology, UiT - The Arctic University of Norway, Tromsø, Norway.

ABSTRACT
Liver sinusoidal endothelial cells (LSECs) are specialized scavenger cells that mediate high-capacity clearance of soluble waste macromolecules and colloid material, including blood-borne adenovirus. To explore if LSECs function as a sink for other viruses in blood, we studied the fate of virus-like particles (VLPs) of two ubiquitous human DNA viruses, BK and JC polyomavirus, in mice. Like complete virions, VLPs specifically bind to receptors and enter cells, but unlike complete virions, they cannot replicate. 125I-labeled VLPs were used to assess blood decay, organ-, and hepatocellular distribution of ligand, and non-labeled VLPs to examine cellular uptake by immunohisto- and -cytochemistry. BK- and JC-VLPs rapidly distributed to liver, with lesser uptake in kidney and spleen. Liver uptake was predominantly in LSECs. Blood half-life (∼1 min), and tissue distribution of JC-VLPs and two JC-VLP-mutants (L55F and S269F) that lack sialic acid binding affinity, were similar, indicating involvement of non-sialic acid receptors in cellular uptake. Liver uptake was not mediated by scavenger receptors. In spleen, the VLPs localized to the red pulp marginal zone reticuloendothelium, and in kidney to the endothelial lining of vasa recta segments, and the transitional epithelium of renal pelvis. Most VLP-positive vessels in renal medulla did not express PV-1/Meca 32, suggesting location to the non-fenestrated part of vasa recta. The endothelial cells of these vessels also efficiently endocytosed a scavenger receptor ligand, formaldehyde-denatured albumin, suggesting high endocytic activity compared to other renal endothelia. We conclude that LSECs very effectively cleared a large fraction of blood-borne BK- and JC-VLPs, indicating a central role of these cells in early removal of polyomavirus from the circulation. In addition, we report the novel finding that a subpopulation of endothelial cells in kidney, the main organ of polyomavirus persistence, showed selective and rapid uptake of VLPs, suggesting a role in viremic organ tropism.

Show MeSH
Related in: MedlinePlus