spacer gif spacer gif spacer gif spacer gif spacer gif
QUICK SEARCH:   [advanced]


spacer gif
     Home     Help     Feedback     Subscriptions     Archive     Search     Table of Contents    

First published online 8 June 2005
doi: 10.1242/dev.01902

Development 132, 3293-3303 (2005)
Published by The Company of Biologists 2005
This Article
Right arrow Summary Freely available
Right arrow Full Text
Right arrow Full Text (PDF)
Right arrow Alert me when this article is cited
Right arrow Alert me if a correction is posted
Services
Right arrow Email this article to a friend
Right arrow Similar articles in this journal
Right arrow Similar articles in PubMed
Right arrow Alert me to new issues of the journal
Right arrow Download to citation manager
Right arrow reprints & permissions
Citing Articles
Right arrow Citing Articles via HighWire
Right arrow Citing Articles via Google Scholar
Google Scholar
Right arrow Articles by Carpenter, B.
Right arrow Articles by Wang, R.
Right arrow Search for Related Content
PubMed
Right arrow PubMed Citation
Right arrow Articles by Carpenter, B.
Right arrow Articles by Wang, R.

VEGF is crucial for the hepatic vascular development required for lipoprotein uptake

Brian Carpenter1,*,{dagger}, Yuankai Lin1,*, Stephanie Stoll1, Robert L. Raffai1,2, Robert McCuskey3 and Rong Wang1,{ddagger}

1 Pacific Vascular Research Laboratory, Division of Vascular Surgery, Department of Surgery, University of California, San Francisco, CA 94143-0507, USA
2 Veterans Administration Medical Research Center, Surgical Services (112G), San Francisco, CA 94121, USA
3 Department of Cell Biology and Anatomy, University of Arizona College of Medicine, Tucson, AZ 85724, USA



View larger version (45K):

[in a new window]
  		Fig. 1. Dox-controlled VEGFR2T expression in hepatocytes. (A) Schematic diagram of the transgene TRE-VEGFR2T. (B) Western blotting of newborn liver using an anti-HA antibody. Lanes 1-3, TRE-VEGFR2T sub-line 4377.5; lanes 4-6, sub-line 3647.5. (C,D) Expression of tTA by lacZ reporter assay in (C) E10.5 and (D) E11.5 embryos. (E,F) Immunostaining for the extracellular domain of VEGFR2 on LAP-tTA control liver (E) and LAP-tTA/TRE-VEGFR2T mutant liver (F). Nuclei are stained blue with DAPI. (G) VEGF levels in liver by ELISA. Error bars represent s.d. *P<0.01.

 


View larger version (36K):

[in a new window]
  		Fig. 2. Gross morphological and histological phenotype of LAP-tTA/TRE-VEGFR2T newborn mice. (A) A dark-red appearance distinguishes the LAP-tTA/TRE-VEGFR2T mutant livers from controls. Dox suppresses this phenotype. (B,C) Body weight and liver/body weight ratios of LAP-tTA/TRE-VEGFR2T newborns are comparable to LAP-tTA controls. (D) Western blotting with an anti-albumin antibody. (E) Insignificant reduction in hepatocyte proliferation in mutant livers by PCNA staining. (F) No significant difference in apoptosis. The number of proliferating cells (E) or apoptotic cells (F) is per 40 x microscopic field. Error bars represent the s.d.

 


View larger version (137K):

[in a new window]
  		Fig. 3. The vascular sinusoidal network is disorganized in mutant livers. (A,C,E,G) Control livers; (B,D,F,H) mutant livers. (A-D) H&E staining of unperfused (A,B) and perfused (C,D) newborn livers shows that red blood cells accumulate in the livers of TRE-VEGFR2T/LAP-tTA transgenic newborns and cannot be flushed out (D). (E,F) Anti-CD31 staining (brown) shows that the microvascular networks in TRE-VEGFR2T/LAP-tTA livers fail to fully develop when compared with controls, with some sinusoids appearing to be collapsed (arrow in F). (G,H) Anti-LYVE1 staining (brown) shows a similar expression pattern and level.

 


View larger version (131K):

[in a new window]
  		Fig. 4. Lipid accumulation in hepatocytes is impaired in LAP-tTA/TRE-VEGFR2T mice. (A,B) Oil Red O staining of liver sections reveals decreased hepatocellular lipids in mutant livers (B) when compared with littermate control livers (A). Samples are counterstained with Hematoxylin. (C,D) Infusion experiment with DiI-labeled remnant lipoproteins demonstrates the quick and efficient uptake and transportation of serum lipid by hepatocytes in control livers (C) but not in mutant livers (D). (E,F) Anti-APOE staining shows continuous APOE accumulation in the space of Disse of control livers (E), whereas, in the mutant livers, an area with little or disrupted APOE distribution is evident (F).

 


View larger version (149K):

[in a new window]
  		Fig. 5. Electron microscopy of liver sections shows defects in SEC and hepatocyte morphology. There is an abundance of lipid droplets (white arrowheads) in the control (A), but not in mutant (B), livers. SEM of livers shows fewer sinusoidal channels (white arrows) in the mutant (B). TEM of control (C,E) and mutant (D,F) livers reveals abnormal hepatocyte morphology, reduced hepatocyte microvilli projections (black arrows), and disrupted space of Disse (black arrowheads) in the mutant livers. SEM shows that mutant SECs lack fenestrae (H), whereas SECs in control livers are highly fenestrated (white arrowheads in G). EC, endothelial cell; L, lipid; H, hepatocyte; N, neutrophil; Er, erythrocyte; S, sinusoid.

 


View larger version (93K):

[in a new window]
  		Fig. 6. Vascular development is defective in livers that express VEGFR2T during organogenesis. The developing liver vasculature was observed by ß-galactosidase staining of ECs with the VEGFR2lacZ allele (blue). The vasculature of control (A,C,E,G) and mutant (B,D,F,H) livers was visualized at E11.5 (A,B), E12.5 (C,D) and E13.5 (E,F), and in newborn pups (G,H). Arrows denote major blood vessels; arrowheads highlight the microvasculature (E,F). (I,J) The sinusoidal network is equally disrupted in newborn mutant livers (J) when Tet was removed at E12.5.

 




© The Company of Biologists Ltd 2005