Biomedical subjects

Yin-Shan Ng

Publications and source records attributed to Yin-Shan Ng.

11 recordsLinked to original sources

PubMed2006

An in vitro assay reveals a role for the diaphragm protein PV-1 in endothelial fenestra morphogenesis.

Fenestrae are small pores in the endothelium of renal glomerular, gastrointestinal, and endocrine gland capillaries and are involved in the bidirectional exchange of molecules between blood and tissues. Although decades of studies have characterized fenestrae at the ultrastructural level, little is known on the mechanisms by which fenestrae form. We present the development of an in vitro assay in which rapid and abundant fenestra induction enables a detailed study of their biogenesis. Through the use of agents that stabilize or disassemble actin microfilaments, we show that actin microfilament remodeling is part of fenestra biogenesis in this model. Furthermore, by using a loss-of-function approach, we show that the diaphragm protein PV-1 is necessary for fenestral pore architecture and the ordered arrangement of fenestrae in sieve plates. Together, these data provide insight into the cell biology of fenestra formation and open up the future study of the fenestra to a combined morphological and biochemical analysis.

Actin Cytoskeleton↗

PubMed2005

A therapeutic aptamer inhibits angiogenesis by specifically targeting the heparin binding domain of VEGF165.

Aptamers recognize their targets with extraordinary affinity and specificity. The aptamer-based therapeutic, Macugen, is derived from a modified 2'fluoro pyrimidine RNA inhibitor to vascular endothelial growth factor (VEGF) and is now being used to treat the wet form of age-related macular degeneration. This VEGF(165) aptamer binds specifically to the VEGF(165) isoform, a dimeric protein with a receptor-binding domain and a heparin-binding domain (HBD). To understand the molecular recognition between VEGF and this aptamer, binding experiments were used to show that the HBD contributes the majority of binding energy in the VEGF(165)-aptamer complex. A tissue culture-based competition assay demonstrated that the HBD effectively competes with VEGF(165) for aptamer binding in vivo. Comparison of NMR spectra revealed that structural features of the smaller HBD-aptamer complex are present in the full-length VEGF(164)-aptamer complex. These data show that the HBD provides the binding site for the aptamer and is the primary determinant for the affinity and specificity in the VEGF(165)-aptamer complex.

Aptamers, Nucleotide↗

PubMed2005

VEGF function in vascular pathogenesis.

Vascular endothelial growth factor (VEGF) has proven to be the most critical angiogenic factor identified to date; its discovery has transformed the field of angiogenesis research. After two decades of intense studies by numerous laboratories, the complex biology of VEGF is slowly being elucidated. VEGF is a collection of several different isoforms with overlapping but also unique functions. Furthermore, there are at least four different VEGF receptors, adding to the complexity of the VEGF/receptor system. VEGF exerts its effects on endothelial cells, inflammatory cells and neuronal cells, and its expression may contribute to the pathologies of diseases ranging from retinopathies to cancer. This review focuses on the biochemistry and cell biology of VEGF, with particular emphasis on the different isoforms and receptors, thereby illustrating the multifunctional nature of this growth factor. Lastly, critical considerations based on the complex biology of VEGF for developing specific and more targeted anti-VEGF therapeutic agents will be discussed.

Angiogenesis Inhibitors↗

PubMed2004

VEGF expression is downregulated in nitrofen-induced congenital diaphragmatic hernia.

BACKGROUND: Vascular endothelial growth factor (VEGF) is upregulated in pulmonary alveolarization. However, developmental expression of pulmonary VEGF and its possible role in the pathogenesis of CDH are not well described. METHODS: Timed-pregnant VEGF-LacZ mice, possessing a beta-galactosidase reporter introduced into the 3' region of the VEGF gene, were used to examine fetal lung gene expression in a model of nitrofen-induced CDH. RESULTS: VEGF gene expression increased from embryonic day 13 until its peak at embryonic day 16 and then decreased until term in all groups. This pattern was most apparent in the periphery with smaller differences noted in central lung locations. Expression of VEGF/beta-gal in the lungs of nitrofen-treated mice was less than controls at all time-points (P <.0001) The type-II pneumocyte population did not significantly differ between the groups. Study concentrations of nitrofen showed no effect on vascular endothelial proliferation in vitro. CONCLUSIONS: Nitrofen downregulates the production of VEGF during gestation and attenuates the peak seen at the onset of the canalicular stage, despite preservation of type-II pneumocytes. This effect was most pronounced in peripheral lung tissue. The authors speculate that altered VEGF expression may have a pivotal role in the pathogenesis of nitrofen-induced CDH.

Abnormalities, Drug-Induced↗

PubMed2004

Identification of genes involved in VEGF-mediated vascular morphogenesis using embryonic stem cell-derived cystic embryoid bodies.

The vasculature forms during development via two processes, vasculogenesis and angiogenesis, in which vessels form de novo from angioblast precursors or as sprouts from pre-existing vessels, respectively. A common and critical aspect of both processes is vascular morphogenesis, which includes branching of endothelial cell cords and lumen formation. Although ample evidence support the central role of vascular endothelial growth factor (VEGF) in both vasculogenesis and angiogenesis, the role of VEGF in vascular morphogenesis is unclear and little is known about the regulation of vascular morphogenesis, in general. We have used the in vitro vessel differentiation system of embryonic stem (ES) cell-derived cystic embryonic bodies (CEB) as a model for studying VEGF-mediated vessel formation. Whereas CEB formed from wild-type ES cells make well-formed vessel-like structures, CEB derived from VEGF-null ES cells contain PECAM-1-positive endothelial cells, but these cells do not participate in vascular morphogenesis. Using gene expression microarray analysis to compare gene expression in these two systems, we have been able to identify many genes and novel ESTs that are downstream of VEGF function, and which may be involved in VEGF-mediated vascular morphogenesis including caveolin-1 and HEY-1. These results support using the CEB model, in combination with gene knockout ES cells, for studying vascular morphogenesis.

Animals↗

PubMed2003

VEGF164-mediated inflammation is required for pathological, but not physiological, ischemia-induced retinal neovascularization.

Hypoxia-induced VEGF governs both physiological retinal vascular development and pathological retinal neovascularization. In the current paper, the mechanisms of physiological and pathological neovascularization are compared and contrasted. During pathological neovascularization, both the absolute and relative expression levels for VEGF164 increased to a greater degree than during physiological neovascularization. Furthermore, extensive leukocyte adhesion was observed at the leading edge of pathological, but not physiological, neovascularization. When a VEGF164-specific neutralizing aptamer was administered, it potently suppressed the leukocyte adhesion and pathological neovascularization, whereas it had little or no effect on physiological neovascularization. In parallel experiments, genetically altered VEGF164-deficient (VEGF120/188) mice exhibited no difference in physiological neovascularization when compared with wild-type (VEGF+/+) controls. In contrast, administration of a VEGFR-1/Fc fusion protein, which blocks all VEGF isoforms, led to significant suppression of both pathological and physiological neovascularization. In addition, the targeted inactivation of monocyte lineage cells with clodronate-liposomes led to the suppression of pathological neovascularization. Conversely, the blockade of T lymphocyte-mediated immune responses with an anti-CD2 antibody exacerbated pathological neovascularization. These data highlight important molecular and cellular differences between physiological and pathological retinal neovascularization. During pathological neovascularization, VEGF164 selectively induces inflammation and cellular immunity. These processes provide positive and negative angiogenic regulation, respectively. Together, new therapeutic approaches for selectively targeting pathological, but not physiological, retinal neovascularization are outlined.

Animals↗

PubMed2002

Won't you be my neighbor? Local induction of arteriogenesis.

Two recent papers in Cell and Developmental Cell provide evidence that VEGF directs arterial differentiation. Evidence that sensory nerves direct arteriogenesis and that the membrane-spanning Notch signaling system is downstream of VEGF emphasize the paracrine regulation of vessel formation.

Animals↗

PubMed2002

Defective pulmonary development in the absence of heparin-binding vascular endothelial growth factor isoforms.

Development of the airways, alveoli, and the pulmonary vasculature in the fetus is a process that is precisely controlled. One of the growth factors involved, vascular endothelial growth factor (VEGF), is so critical for embryonic development that in the mouse, elimination of just a single allele is lethal. In the early stages of lung development, the mouse VEGF gene expresses three isoforms (120, 164, and 188) in a distinct temporo-spatial pattern, suggesting a specific function for each. We engineered mice that express only VEGF 120, to study the role of VEGF isoforms in lung development. Lung vessel development in these mice was studied by scanning electron microscopy of Mercox casts of lung vasculature. Airway and air-blood barrier development was analyzed by light microscopy, transmission electron microscopy, immunohistochemistry, and morphometry. In all VEGF120/120 fetuses and pups, lung vascular casts were smaller and less dense compared with 120/+ and wild-type littermates. Although the generation count of pre-acinar vessels was similar in all three genotypes, the most peripheral vessels were dilated and were more widely separated in 120/120 fetuses of all ages, compared with 120/+ and wild-type littermates. In addition, 120/120 animals had fewer air-blood barriers and a decreased airspace-parenchyma ratio compared with 120/+ and wild-type littermates. We concluded that the absence of VEGF 164 and 188 isoforms impairs lung microvascular development and delays airspace maturation, indicating an essential role for heparin-binding VEGF isoforms in normal lung development.

Alternative Splicing↗

PubMed2002

Arteriolar and venular patterning in retinas of mice selectively expressing VEGF isoforms.

The murine VEGF gene is alternatively transcribed to yield the VEGF(120), VEGF(164), and VEGF(188) isoforms, which differ in their potential to bind to heparan sulfate and neuropilin-1 and to stimulate endothelial growth. Here, their role in retinal vascular development was studied in mice selectively expressing single isoforms. VEGF(164/164) mice were normal, healthy, and had normal retinal angiogenesis. In contrast, VEGF(120/120) mice exhibited severe defects in vascular outgrowth and patterning, whereas VEGF(188/188) mice displayed normal venular outgrowth but impaired arterial development. It is noteworthy that neuropilin-1, a receptor for VEGF(164), was predominantly expressed in retinal arterioles. These findings reveal distinct roles of the various VEGF isoforms in vascular patterning and arterial development in the retina.

Animals↗

PubMed2002

Skeletal defects in VEGF(120/120) mice reveal multiple roles for VEGF in skeletogenesis.

Angiogenesis is an essential component of skeletal development and VEGF signaling plays an important if not pivotal role in this process. Previous attempts to examine the roles of VEGF in vivo have been largely unsuccessful because deletion of even one VEGF allele leads to embryonic lethality before skeletal development is initiated. The availability of mice expressing only the VEGF120 isoform (which do survive to term) has offered an opportunity to explore the function of VEGF during embryonic skeletal development. Our study of these mice provides new in vivo evidence for multiple important roles of VEGF in both endochondral and intramembranous bone formation, as well as some insights into isoform-specific functions. There are two key differences in vascularization of developing bones between wild-type and VEGF(120/120) mice. VEGF(120/120) mice have not only a delayed recruitment of blood vessels into the perichondrium but also show delayed invasion of vessels into the primary ossification center, demonstrating a significant role of VEGF at both an early and late stage of cartilage vascularization. These findings are the basis for a two-step model of VEGF-controlled vascularization of the developing skeleton, a hypothesis that is supported by the new finding that VEGF is expressed robustly in the perichondrium and surrounding tissue of cartilage templates of future bones well before blood vessels appear in these regions. We also describe new in vivo evidence for a possible role of VEGF in chondrocyte maturation, and document that VEGF has a direct role in regulating osteoblastic activity based on in vivo evidence and organ culture experiments.

Animals↗

PubMed2001

Therapeutic angiogenesis for cardiovascular disease.

The identification of angiogenic growth factors, such as vascular endothelial growth factor and fibroblast growth factor, has fueled interest in using such factors to induce therapeutic angiogenesis. The results of numerous animal studies and clinical trials have offered promise for new treatment strategies for various ischemic diseases. Increased understanding of the cellular and molecular biology of vessel growth has, however, prompted investigators and clinicians alike to reconsider the complexity of therapeutic angiogenesis. The realization that formation of a stable vessel is a complex, multistep process may provide useful insights into the design of the next generation of angiogenesis therapy.

Journal Article↗