Transplanted endothelial cells repopulate the liver endothelium and correct the phenotype of hemophilia A mice

Follenzi, Antonia; Benten, Daniel; Novikoff, Phyllis M.; Faulkner, L.; Raut, S.; Gupta, Sanjeev

doi:10.1172/jci32748

Cited by 114 publications

(164 citation statements)

References 61 publications

(69 reference statements)

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“…In our study, LYVE-1-positive endothelial cells in EBs treated with AM and SB431542 expressed stabilin-2, and some exhibited fenestrae-like structures. In addition, transcription of the LSEC markers F8, Consistent with that idea, it was recently shown that transplanted sinusoidal endothelial cells can repopulate the liver endothelium and correct the phenotype of hemophilia A mice [7,15], and several reports have shown that endothelial progenitor cell transplantation ameliorates acute liver injury and liver cirrhosis in rats [18,23,31,32] . 1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 22 23 24 25 26 27 28 29 30 31 32 33 34 35 36 37 38 39 40 41 42 43 44 45 46 47 48 49 50 51 52 53 54 55 56 57 58 59 60 61 62 63 64 65 23 Our second major finding is that AM-RAMP2 system is the potentital target for the induction of LSECs.…”

Section: Discussionmentioning

confidence: 56%

Induction of LYVE-1/stabilin-2-positive liver sinusoidal endothelial-like cells from embryoid bodies by modulation of adrenomedullin-RAMP2 signaling

et al. 2011

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Embryonic stem cells (ESCs) are a useful source for various cell lineages. So far, however, progress toward reconstitution of mature liver morphology and function has been limited. We have shown that knockout mice deficient in adrenomedullin (AM), a multifunctional endogenous peptide, or its receptor-activity modifying protein (RAMP2) die in utero due to poor vascular development and hemorrhage within the liver. In this study, using embryoid bodies (EBs)-culture system, we successfully induced liver sinusoidal endothelial-like cells by modulation of AM-RAMP2. In an EB differentiation system, we found that co-administration of AM and SB431542, an inhibitor of transforming growth factor (TGF ) receptor type 1, markedly enhanced differentiation of lymphatic vessel endothelial hyaluronan receptor-1 (LYVE-1)/stabilin-2-positive endothelial cells. These cells showed robust endocytosis of acetylated low-density lipoprotein (Ac-LDL) and upregulated expression of liver sinusoidal endothelial cells (LSECs)-specific markers, including factor 8 (F8), Fc-γ receptor 2b (Fcgr2b), and mannose receptor C type 1 (Mrc1), and also possess fenestrae-like structure, a key morphological feature of LSECs. In RAMP2-null liver, by contrast, LYVE-1 was downregulated in LSECs, and the sinusoidal structure was disrupted. Our findings highlight the importance of AM-RAMP2 signaling for

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Section: Discussionmentioning

confidence: 56%

Induction of LYVE-1/stabilin-2-positive liver sinusoidal endothelial-like cells from embryoid bodies by modulation of adrenomedullin-RAMP2 signaling

et al. 2011

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“…11,21,22 The cells were cultured on collagen-coated plastic tissue culture dishes in endothelial cell media (ScienCell, Carlsbad, CA) containing 5% fetal bovine serum, 1% penicillin-streptomycin, and 1% ECGS supplement (ScienCell). Primary human hepatic sinusoidal endothelial cells (HHSECs; ScienCell), or transformed sinusoidal endothelial cells (TSECs), an SV40-immortalized mouse cell line derived from sinusoidal endothelial cells, 23 were grown on uncoated plastic dishes in the same media.…”

Section: Cell Culturementioning

confidence: 99%

Aquaporin-1 Promotes Angiogenesis, Fibrosis, and Portal Hypertension Through Mechanisms Dependent on Osmotically Sensitive MicroRNAs

Huebert

Jagavelu

Hendrickson

et al. 2011

The American Journal of Pathology

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Changes in hepatic vasculature accompany fibrogenesis, and targeting angiogenic molecules often attenuates fibrosis in animals. Aquaporin-1 (AQP1) is a water channel, overexpressed in cirrhosis, that promotes angiogenesis by enhancing endothelial invasion. The effect of AQP1 on fibrogenesis in vivo and the mechanisms driving AQP1 expression during cirrhosis remain unclear. The purpose of this study was to test the effect of AQP1 deletion in cirrhosis and explore mechanisms regulating AQP1. After bile duct ligation, wild-type mice overexpress AQP1 that colocalizes with vascular markers and sites of robust angiogenesis. AQP1 knockout mice demonstrated reduced angiogenesis compared with wild-type mice, as evidenced by immunostaining and endothelial invasion/proliferation in vitro. Fibrosis and portal hypertension were attenuated based on immunostaining, portal pressure, and spleen/body weight ratio. AQP1 protein, but not mRNA, was induced by hyperosmolality in vitro, suggesting post-transcriptional regulation. Endothelial cells from normal or cirrhotic mice were screened for microRNA (miR) expression using an array and a quantitative PCR. miR-666 and miR-708 targeted AQP1 mRNA and were decreased in cirrhosis and in cells exposed to hyperosmolality, suggesting that these miRs mediate osmolar changes via AQP1. Binding of the miRs to the untranslated region of AQP1 was assessed using luciferase assays. In conclusion, AQP1 promotes angiogenesis, fibrosis, and portal hypertension after bile duct ligation and is regulated by osmotically sensitive miRs.

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“…A representative fractionation is shown in Figure 6. The first peak (fractions 4-10) corresponds to the high-density WPB-containing fractions, whereas the second peak (fractions [20][21][22][23][24][25] contains the subcellular fractions derived of organelles of the secretory pathway (endoplasmatic reticulum (ER), Golgi, trans-Golgi network) and constitutively released vesicles. 37 The amount of VWF stored in WPBs was similar for non-transduced (12-15%; Figure 6A) and FVIII-GFP transduced (15-21%; Figure 6B) BOECs.…”

Section: Quantitative Determination Of the Amount Of Von Willebrand Fmentioning

confidence: 99%

“…Studies in hemophilia A mice have demonstrated that transplantation of liver sinusoidal endothelial cells can correct the hemophilic phenotype. 5,21 In addition, transplantation of genetically modified BOECs intravenously 10,22 or implanted subcutaneously in a Matrigel TM scaffold 10 results in long-term therapeutic levels of FVIII.…”

Section: Introductionmentioning

confidence: 99%

Storage and regulated secretion of factor VIII in blood outgrowth endothelial cells

Biggelaar¹,

Bouwens²,

Kootstra³

et al. 2009

Haematologica

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BackgroundGene therapy provides an attractive alternative for protein replacement therapy in hemophilia A patients. Recent studies have shown the potential benefit of directing factor (F)VIII gene delivery to cells that also express its natural carrier protein von Willebrand factor (VWF). In this study, we explored the feasibility of blood outgrowth endothelial cells as a cellular FVIII delivery device with particular reference to long-term production levels, intracellular storage in Weibel-Palade bodies and agonist-induced regulated secretion. Design and MethodsHuman blood outgrowth endothelial cells were isolated from peripheral blood collected from healthy donors, transduced at passage 5 using a lentiviral vector encoding human Bdomain deleted FVIII-GFP and characterized by flow cytometry and confocal microscopy. ResultsBlood outgrowth endothelial cells displayed typical endothelial morphology and expressed the endothelial-specific marker VWF. Following transduction with a lentivirus encoding FVIII-GFP, 80% of transduced blood outgrowth endothelial cells expressed FVIII-GFP. Levels of FVIII-GFP positive cells declined slowly upon prolonged culturing. Transduced blood outgrowth endothelial cells expressed 1.6±1.0 pmol/1×10 6 cells/24h FVIII. Morphological analysis demonstrated that FVIII-GFP was stored in Weibel-Palade bodies together with VWF and P-selectin. FVIII levels were only slightly increased following agonist-induced stimulation, whereas a 6-to 8-fold increase of VWF levels was observed. Subcellular fractionation revealed that 15-22% of FVIII antigen was present within the dense fraction containing Weibel-Palade bodies. ConclusionsWe conclude that blood outgrowth endothelial cells, by virtue of their ability to store a significant portion of synthesized FVIII-GFP in Weibel-Palade bodies, provide an attractive cellular on-demand delivery device for gene therapy of hemophilia A.

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Transplanted endothelial cells repopulate the liver endothelium and correct the phenotype of hemophilia A mice

Cited by 114 publications

References 61 publications

Induction of LYVE-1/stabilin-2-positive liver sinusoidal endothelial-like cells from embryoid bodies by modulation of adrenomedullin-RAMP2 signaling

Induction of LYVE-1/stabilin-2-positive liver sinusoidal endothelial-like cells from embryoid bodies by modulation of adrenomedullin-RAMP2 signaling

Aquaporin-1 Promotes Angiogenesis, Fibrosis, and Portal Hypertension Through Mechanisms Dependent on Osmotically Sensitive MicroRNAs

Storage and regulated secretion of factor VIII in blood outgrowth endothelial cells

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