Upregulation Of VEGF Expression Research Articles

The Developing Bladder is Hypoxic in Vivo and Bladder Angiogenesis is Controlled Through VEGF by Tissue Oxygenisation

Purpose During embryogenesis the fetus is subjected to a relative hypoxic condition. Tissue hypoxia in the neural tube and heart is considered to enhance embryonic angiogenesis, partly by upregulating VEGF expression. We hypothesized that the developing bladder is hypoxic in vivo, and that oxygen tensions modulate VEGF expression in theembryonic bladder. Material and methods Pimonidazole, a hypoxia-marker, was administered to pregnant mice at embryonic days (E)14,16 and 18, spanning bladder inception to a stage when the detrusor and lamina propria become well-defined and detected by immunohistochemistry (IHC) . Western blot and IHC were used to detect VEGF and CD31 (endothelial marker). Explanted E14 bladders were cultured in 20%O2 (representing normoxia for postnatal animals) or 3%O2 (representing a relatively hypoxic state, perhaps more equivalent to the embryo milieu). Percentages of CD31 expressing endothelia were measured by flowcytometry and capillary density was assessed using IHC Results At E14, both VEGF and hypoxic pimonidazole-adducts immunocolocalized to urothelia, whereas, at E16-18, both were more prominently detected in detrusor. At E14, endothelia (CD31) were visualized directly beneath the urothelium and, at later embryonic stages; capillaries were prominent in lamina propria and also detrusor layers. VEGF protein levels within explanted bladders themselves, and also in media conditioned by rudiments, was increased in 3%O2 versus 20%O2. Similarly, the amount and density of endothelia in explants increased after culture in hypoxic versus 20%O2 atmosphere Conclusions In vivo, the developing bladder contains hypoxic regions rich in VEGF and close to nascent capillary beds. Invitro, when oxygen tensions are experimentally altered, expression of VEGF protein positively correlates with endothelial mass and density.

C-myc Amplification Is Frequent in Esophageal Adenocarcinoma and Correlated with the Upregulation of VEGF-A Expression

BACKGROUND: Deregulation of c-myc plays a major role in the carcinogenesis of human malignancies. We investigated the amplification of the c-myc gene in a surgical series of Barrett cancers. METHODS: Primary resected esophageal (Barrett) adenocarcinomas (n = 84) were investigated for c-myc amplification using chromogene in situ hybridization. Tumor samples were assembled in a tissue microarray. c-myc gene dosage was correlated with clinicopathologic parameters, including the survival and gene expression of cyclooxygenases (COX-1 and COX-2) and proangiogenic growth factors (VEGF-A and VEGF-C). RESULTS: The majority (70 of 84; 83.3%) exhibited amplification of the c-myc gene. There were low-level amplifications in 63 (75.0%) cases and high-level amplifications in 7 (8.3%) cases. No amplification was found in 14 (16.7%) cases. Tumors without c-myc amplification had lower VEGF-A, VEGF-C, and COX-2 expression levels than tumors with low-level and high-level c-myc amplification (statistically significant for VEGF-A; P = .0348). c-myc amplification was not correlated with clinicopathological parameters or survival. Only diffuse and mixed-type tumors, according to Lauren classification, exhibited c-myc amplifications more frequently (P = .0466). CONCLUSIONS: Amplifications of the c-myc gene are frequent in Barrett cancer. c-myc may be involved in the regulation of angiogenesis.

Thrombopoietin and the Hematopoietic Stem Cell

The molecular mechanisms that underlie the favorable effects of thrombopoietin on stem cell survival, self-renewal, and expansion are unknown. On the basis of known effects of HoxB4, HoxA9, and vascular endothelial cell growth factor on stem cells, we explored whether TPO might affect these pathways. We found that TPO enhances the transcription of HoxB4, induces the nuclear localization of HoxA9, and upregulates VEGF expression by reducing the ubiquination of hypoxia inducible factor.

1119. Isolation of a Novel Therapeutic Angiogenic Peptide, AG-30, by High Throughput Screening Using the HVJ-E Vector

Top of pageAbstract In a large proportion of critical limb ischemia patients, the anatomic extent and distribution of arterial occlusive disease make the patients unsuitable for operative or percutaneous revascularization. Thus, novel strategies, such as therapeutic angiogenesis, are expected to play an increasing role in the treatment of these patients. The goal of the present study was to screen a human cDNA library to isolate potent and powerful angiogenic factors that may produce better patient outcomes. Our study utilizes Hemagglutinating Virus of Japan (HVJ; Sendai virus) envelope vector (HVJ-E) for high throughput screening, based on the advantage of this vector demonstrated in previous studies. The advantages of high throughput functional screening system based on HVJ-E included: 1) rapid preparation of the vector containing the DNA library; 2) effective fusion-mediated transfer of the plasmids to various cells with minimal toxicity; and 3) easy cloning of candidate genes by transformation of E. coli. These advantages result in a lower probability of damage to isolated clones and in minimization of the time needed to screen for candidate genes. In fact, the present experiment was able to isolate candidate genes within four weeks. A human heart cDNA library (approximately 1 |[times]| 107 independent clones) was amplified, and approximately 4 |[times]| 106 clones were randomly infused into HVJ-E vector, which were transferred to human aortic endothelial cells (HAEC). This assay identified three of the genes that were superior to VEGF in terms of c-fos promoter activity. One gene in particular showed marked HAEC proliferative activity and c-fos promoter activity in comparison to VEGF and was used for subsequent experiments. Further study on this clone revealed that 96 bp segment is the active site of the clone. Thus, 30-amino acid peptide (AG-30) that derived from the segment was synthesized and used for further analysis. AG-30 showed a dose-dependent increase in HAEC viability and cell migration. Further, to confirm the angiogenic ability, an in vitro tube formation assay was performed. Tube formation was greater in response to AG-30 treatment than to recombinant VEGF. The in vivo angiogenic activity of AG-30 has been evaluated by an established mouse Matrigel plug assay. Neovessels containing intact red blood cells were observed in the Matrigel plug containing AG-30. The increased presence of neovessels in the AG-30-treated group relative to the other groups was confirmed by immunostaining with anti-CD31 antibody. Real-time RT PCR revealed that AG-30 strongly and specifically upregulate the mRNA of IGF-1 of HAEC in a time-dependent manner for 72 hours. The relatively durability of the effect of AG-30 makes it potentially suitable for clinical angiogenic therapy. Secondary effects of AG-30 may include: 1) IGF-1-mediated upregulation of VEGF expression, and 2) IGF-1-mediated Akt activation, thereby further promoting angiogenesis. Since small peptides are typically not stable in vivo, development of a delivery system to establish slow release of this peptide would be required for clinical utility.

Cyclic stretch increases VEGF expression in pulmonary arterial smooth muscle cells via TGF-β1 and reactive oxygen species: a requirement for NAD(P)H oxidase

Our previous studies have indicated that transforming growth factor (TGF)-beta1 and VEGF expression are increased in the smooth muscle cell (SMC) layer of the pulmonary vessels of lambs with pulmonary hypertension secondary to increased pulmonary blood flow. Furthermore, we found that TGF-beta1 expression increased before VEGF. Because of the increased blood flow in the shunt lambs, the SMC in the pulmonary vessels are exposed to increased levels of the mechanical force, cyclic stretch. Thus, in this study, using primary cultures of pulmonary arterial SMC isolated from pulmonary arteries of 4-wk-old lambs, we investigated the role of cyclic stretch in the apparent coordinated regulation of TGF-beta1 and VEGF. Our results demonstrated that cyclic stretch induced a significant increase in VEGF expression both at the mRNA and protein levels (P < 0.05). The increased VEGF mRNA was preceded by both an increased expression and secretion of TGF-beta1 and an increase in reactive oxygen species (ROS) generation. In addition, a neutralizing antibody against TGF-beta1 abolished the cyclic stretch-dependent increases in both superoxide generation and VEGF expression. Our data also demonstrated that cyclic stretch activated an NAD(P)H oxidase that was TGF-beta1 dependent and that NAD(P)H oxidase inhibitors abolished the cyclic stretch-dependent increase in VEGF expression. Therefore, our results indicate that cyclic stretch upregulates VEGF expression via the TGF-beta1-dependent activation of NAD(P)H oxidase and increased generation of ROS.

Abstracts

Abstracts

Vascular endothelial growth factor expression in myometrial resistance arteries in women with preeclampsia

It has recently been demonstrated that induced endothelial damage upregulates VEGF expression in medial smooth muscle cells of the arterial wall. The purpose of this study is to determine whether there is difference in VEGF expression in placentae and myometrial resistance arteries from patients with preeclampsia and those from controls to clarify the source of increased systemic level of VEGF in preeclampsia. : Placentae were collected immediately after cesarean section and a small piece of full-thickness uterine myometrium was excised from the upper margin of cesarean section incision sites from normal pregnant women (n = 10) and preeclampsia patients (n = 7). For VEGF immunohistochemistry (IHC), sections were incubated with a polyclonal goat anti-human VEGF, and the remaining procedure was performed using the ABC kit. VEGF mRNA in situ hybridization (ISH) was performed with paraffin-embedded tissue blocks using VEGF probe. For detection, the sections were covered with streptavidin-AP and incubated for 20 minutes and washed in TBS for 5 minutes and then covered with BCIP/NBT and incubated in the dark for 60 minutes and washed in water for 5 minutes. In placental tissue, VEGF was identified primarily in syncytiotrophoblasts in IHC and mRNA ISH, and there were no differences in staining between placentae from preeclampsia and those from control group. We also identified VEGF staining in smooth muscle cell in myometrium and perivascular smooth muscle in myometrial resistance arteries. However, there were no differences in VEGF staining between preeclampsia and control groups. This study suggests placenta and perivascular smooth muscle are not the origin of the increased circulating VEGF in preeclampsia. To understand the role of VEGF in preeclampsia clearly, further studies are required to find out the sites of increased VEGF synthesis and changes in VEGF metabolism in preeclampsia.

Late-Breaking Science

### H11 Kinase Promotes or Inhibits Cardiac Cell Growth and Survival, Depending on its Subcellular Distribution Christophe Depre, Anna Zajac, Nadia Hedhli, Chull Hong, Jing Liu, Guiping Yang, Li Wang, Huacheng Dai, Univ. Medicine New Jersey, Newark, NJ; Thomas Wagner, Greenville Hosp System, Greenville, SC; Stephen F Vatner; Univ. Medicine New Jersey, Newark, NJ H11 kinase is a protein of unknown function mainly expressed in heart and skeletal muscle. Its expression is markedly increased in the ischemic heart together with an array of cytoprotective and anti-apoptotic genes, which supposes that H11 kinase promotes cell survival. Paradoxically, overexpression of H11 kinase in other cell types induces apoptosis. Therefore, the goal of the present study was to determine how H11 kinase affects cardiac cell survival and death, using both a cardiac-specific transgenic (TG) mouse model and cardiac cell culture. After 45 min coronary occlusion in vivo, infarct size in hearts from TG mice (7-fold overexpression) was reduced by 80% compared to wild type (P<0.01). This protection was related to a significant (P<0.05) increase in the expression of proteins promoting cell survival, including heat-shock proteins (HSP70, mmDJA4), phospho-Akt, the mTOR targets p70S6K and 4EBP-1, and the glucose transporter GLUT1. Surprisingly, although overexpression of H11 kinase is protective in the TG animal, ischemia reduced H11 kinase expression by 60% in wild type mice (P<0.01 versus sham). To reconcile this paradox, we performed a subcellular fractionation, which revealed that H11 kinase is expressed in both the cytosol and the nucleus. In wild type mice, the cytosolic expression of H11 kinase during ischemia was reduced by more than 80%, whereas the nuclear expression was unaffected. In the TG mouse, overexpression of H11 kinase accumulated as a nuclear protein, which therefore mediates the survival effect. To understand the role of the cytosolic form of the protein, we transfected isolated cardiac myocytes with increasing doses of adenovirus (3 MOI to 100 MOI) harboring the …

Upregulated expression of VEGF and Ca IX correlate with the vascular metastasis pattern of intestinal type carcinoma

Upregulated expression of VEGF and Ca IX correlate with the vascular metastasis pattern of intestinal type carcinoma

Revascularization of myocardial scar tissue following prostaglandin E1-therapy in patients with ischemic heart disease.

Prostaglandin E1 (PGE-1) treatment has proved to stimulate angiogenesis in vital non-infarcted myocardium of patients with ischemic cardiomyopathy (ICMP). We investigated infarcted myocardial tissue for a possible angiogenic response to PGE-1. Neovascularization was investigated in infarcted areas of 12 hearts explanted from patients with ICMP who had been treated with PGE-1 before heart transplantation (HTX). In transmural sections containing myocardial scar tissue, CD34 and VEGF were immunohistochemically quantified to estimate capillary density and the extent of angiogenesis. To investigate a possible effect of PGE-1 on collagen turnover, the collagen content was determined in myocardial scar tissue by assessing the intensity of the area positively stained with sirius red. PGE-1-treated patients had significantly more CD34- and VEGF-positive cells in infarcted areas, and showed a significant reduction in collagen content as compared with the non-PGE-1 group (CD34: 120.3 +/- 6.1 vs. 47.7 +/- 6.1 capillary profiles/mm2; VEGF: 52.8 +/- 5.6 vs. 24.0 +/- 4.8 capillary profiles/mm2, and collagen content: 2.18 +/- 0.4 eU vs. 3.59 +/- 0.38 eU). Our data demonstrate that PGE-1 stimulates angiogenesis by upregulating VEGF expression, and reduces fibrosis in cardiac scar tissue of ischemic origin. The induction of therapeutic angiogenesis in vital and at sites of putative dead myocardial scar tissue, along with the hemodynamic improvement in patients with severe ICMP, might explain the favorable clinical outcome in PGE-1-treated patients before HTX.

Coordinating Etk/Bmx activation and VEGF upregulation to promote cell survival and proliferation.

Etk/Bmx, a member of the Tec family of non-receptor tyrosine kinase, is characterized by an N-terminal PH domain and has recently been shown to be involved in the regulation of various cellular processes, including proliferation, differentiation, motility and apoptosis. Since VEGF and the activation of its signaling pathway have been implicated in modulating a variety of biological responses, we characterized the role of Etk-dependent signaling pathways involved in the upregulation of VEGF expression, and explored the functional implications of this enhancement in sustaining cell proliferation and survival. Using Northern and Western analyses, transient transfections, and pharmacological agents, we demonstrate that Etk activation alone is sufficient to transcriptionally induce VEGF expression, independent of the previously identified hypoxia response element (HRE), in both Pa-4 epithelial and TR-BBB endothelial cells under normoxia. In addition, Etk utilizes both MEK/ERK and PI3-K/Pak1 signaling pathways in concert to activate VEGF transcription. Functionally, Etk activation elicits a profound stimulatory effect on TR-BBB cell proliferation and formation of capillary-like networks in Matrigel containing reduced levels of growth factors. Finally, antisense oligonucleotides against either endogenous VEGF or Etk abrogate the proliferation of Etk-activated TR-BBB cells, and exogenous VEGF treatment stimulates endogenous Etk tyrosine phosphorylation in HUVECs. Taken together, these results indicate that VEGF is both an Etk downstream target gene and an Etk upstream activator, constituting a reciprocal Etk-VEGF autoregulatory loop. These findings, to our knowledge, are the first delineation of a network of positive feedforward signaling pathways that converge on the Etk-VEGF axis, causally associating Etk-mediation of VEGF induction with enhanced cellular processes in both epithelial and endothelial cells.

Clinical and experimental evidence of prostaglandin E1-induced angiogenesis in the myocardium of patients with ischemic heart disease.

Prostaglandin E1 (PGE-1) is a potent vasodilative agent which has been used to bridge patients with chronic heart failure listed for heart transplantation (HTX). In various experimental settings PGE-1 appears to stimulate angiogenesis by inducing vascular endothelial growth factor expression. This observational clinical study sought to investigate the angiogenic effects of PGE-1 in the failing human heart. Neovascularization was investigated in 14 explanted hearts from patients with ischemic cardiomyopathy (ICMP) who had been bridged to HTX with PGE-1 (8+/-1 mg/kg/min, 97+/-75.6 days) and compared with 14 hearts who did not receive PGE-1 prior to HTX. In three sectional areas obtained from the left ventricular wall CD34, von Willebrand factor (vWf), nuclear Ki67 (MIB-1), and VEGF were quantified by immunohistochemistry to estimate capillary density and endothelial cell proliferation. Additionally, to investigate a possible angiogenic effect of PGE-1 in vitro, cultured human coronary artery smooth muscle cells (HCASMCs) were treated with PGE-1. PGE-1-treated patients had significantly more CD34- and vWf-positive cells in the subepicardium (both P<0.01), myocardium (both P<0.0001) and subendocardium (P<0.01 and P<0.001) as compared to the nonPGE-1 group. Proliferative endothelial activity expressed by the presence of MIB-1- and VEGF-positive cells (both P<0.0001 in all layers) was increased more than twofold. Addition of PGE-1 to HCASMCs in cell culture resulted in a significant increase in VEGF production (164.0+/-19.7 pg/10(5) cells/24 h, P<0.005) as compared to the control cell line (66.6+/-8.7 pg/10(5) cells/24 h, P<0.005). Our data demonstrate that PGE-1 is a potent stimulator of angiogenesis via upregulation of VEGF expression. The induction of therapeutic angiogenesis in patients with severe ICMP might explain the favorable clinical outcome in PGE-1 treated patients until HTX.

MT1-MMP expression promotes tumor growth and angiogenesis through an up-regulation of vascular endothelial growth factor expression.

Membrane type 1 metalloprotease (MT1-MMP) is a transmembrane metalloprotease that plays a major role in the extracellular matrix remodeling, directly by degrading several of its components and indirectly by activating pro-MMP2. We investigated the effects of MT1-MMP overexpression on in vitro and in vivo properties of human breast adenocarcinoma MCF7 cells, which do not express MT1-MMP or MMP-2. MT1-MMP and MMP-2 cDNAs were either transfected alone or cotransfected. All clones overexpressing MT1-MMP 1) were able to activate endogenous or exogenous pro-MMP-2, 2) displayed an enhanced in vitro invasiveness through matrigel-coated filters independent of MMP-2 transfection, 3) induced the rapid development of highly vascularized tumors when injected subcutaneously in nude mice, and 4) promoted blood vessels sprouting in the rat aortic ring assay. These effects were observed in all clones overexpressing MT1-MMP regardless of MMP-2 expression levels, suggesting that the production of MMP-2 by tumor cells themselves does not play a critical role in these events. The angiogenic phenotype of MT1-MMP-producing cells was associated with an up-regulation of VEGF expression. These results emphasize the importance of MT1-MMP during tumor angiogenesis and open new opportunities for the development of anti-angiogenic strategies combining inhibitors of MT1-MMP and VEGF antagonists.

FGF-4 displays an angiogenic activity through an autocrine up-regulation of VEGF expression

FGF-4 displays an angiogenic activity through an autocrine up-regulation of VEGF expression

Oxygen Regulates Vascular Endothelial Growth Factor-Mediated Vasculogenesis and Tubulogenesis

To determine whether low oxygen is a stimulus for endothelial cell differentiation and vascular development in the kidney, we examined the effect of low oxygen on rat metanephric organ culture, a model known to recapitulate nephrogenesis in the absence of vessels. After 6 days in culture in standard (20% O2) or low oxygen (1–3% O2) conditions, metanephric kidney growth and morphology were assessed by DNA measurement, and light and electron microscopy. DNA content was higher in 3% O2-treated explants (2.5 ± 0.17 μg/kidney,n= 9) than in 20% O2explants (1.5 ± 0.09 μg/kidney,n= 9),P< 0.05. Low oxygen induced proliferation of tubular epithelial cells, resulting in enhanced number of tubules of similar size. Endothelial cells forming capillaries were localized in 3% O2explants by light and electron microscopy and by immunocytochemistry using endothelial cell markers. Flt-1, Flk-1, and ACE-containing cells were detected in 3% O2-treated explants, whereas 20% O2explants were virtually negative. VEGF mRNA levels were 10-fold higher in 3% O2-treated explants than in 20% O2-treated explants. Addition of anti-VEGF antibodies to 3% O2-treated explants prevented low oxygen-induced growth and endothelial cell differentiation and proliferation. Our data indicate that low oxygen stimulates growth by cell proliferation and induces tubulogenesis, endothelial cell differentiation, and vasculogenesis in metanephric kidneys in culture. Upregulation of VEGF expression by low oxygen and prevention of low oxygen-induced tubulogenesis and vasculogenesis by anti-VEGF antibodies indicate that these changes were mediated by VEGF. These data suggest that low oxygen is the stimulus to initiate renal vascularization.

VEGF gene expression is upregulated in electrically stimulated rat skeletal muscle.

Vascular endothelial growth factor (VEGF; also called vascular permeability factor) is a secreted mitogen with distinct target cell specificity for vascular endothelial cells. Hypoxia upregulates VEGF expression, making it a likely mediator of the angiogenesis that occurs in poorly perfused tissues. The purpose of this study was to determine whether VEGF gene expression is upregulated in chronically stimulated skeletal muscles, where hypoxia is thought to trigger the growth of blood vessels. The right anterior tibialis and extensor digitorum longus muscles of 12 rats were stimulated electrically (10 Hz, 300 microseconds pulses) for up to 21 days by way of the peroneal motor nerve. The contralateral muscles served as control. Northern analysis showed that VEGF mRNA levels increased by approximately sixfold after 4 days of stimulation and then decreased gradually over the next several days. VEGF mRNA levels were still elevated by two- to threefold after 21 days of stimulation. Higher VEGF mRNA levels in the early stages of muscle stimulation and gradually decreasing levels in later stages are consistent with a metabolic hypothesis in which tissue oxygenation controls VEGF expression. These studies support the hypothesis that VEGF has a physiological role in promoting angiogenesis in stimulated skeletal muscle.

Basic fibroblast growth factor upregulates the expression of vascular endothelial growth factor in vascular smooth muscle cells. Synergistic interaction with hypoxia.

Vascular endothelial growth factor (VEGF) is a hypoxia-inducible direct angiogenic factor. Upregulation of VEGF is thought to mediate many of the angiogenic effects of growth factors that are not direct endothelial cell mitogens. Like VEGF, basic fibroblast growth factor (bFGF) is considered to induce angiogenesis by a direct effect on endothelial cells. This study investigated the possibility that bFGF may also act indirectly by regulating VEGF expression in vascular smooth muscle cells (VSMCs). Incubation of confluent and quiescent cultures of rabbit VSMCs with bFGF caused a time- and concentration-dependent increase in steady-state levels of VEGF mRNA, as analyzed by Northern blot hybridization. Exposure of VSMCs to a threshold hypoxic stimulus (2.5% O2) caused a modest increase in VEGF mRNA levels. However, the combination of 2.5% O2 with bFGF had a marked synergistic effect. This effect was specific for VEGF as hypoxia did not enhance bFGF-induced expression of the proto-oncogene c-myc. Synergistic upregulation of VEGF mRNA expression also was observed between hypoxia and TGF-beta 1. These results suggest that bFGF may promote angiogenesis both by a direct effect on endothelial cells and also indirectly by the upregulation of VEGF in VSMCs. The synergy demonstrated between hypoxia and either bFGF or TGF-beta 1 suggests that multiple diverse stimuli may interact via the upregulation of VEGF expression in VSMCs to amplify the angiogenic response.

Indirect Angiogenic Cytokines Upregulates VEGF and bFGF Gene Expression in Vascular Smooth Muscle Cells, Whereas Hypoxia Upregulates VEGF Expression Only

Indirect Angiogenic Cytokines Upregulates VEGF and bFGF Gene Expression in Vascular Smooth Muscle Cells, Whereas Hypoxia Upregulates VEGF Expression Only

Induction of vascular endothelial growth factor expression by prostaglandin E2 and E1 in osteoblasts.

PGE1 and PGE2 are potent stimulators of bone formation. Osteogenesis is strongly dependent on angiogenesis. Vascular endothelial growth factor (VEFG), a secreted endothelial cell-specific mitogen, has been implicated in physiological and pathological angiogenesis. The aim of this study was to examine the possible role of VEGF in PG stimulation of bone formation. We found that in rat calvaria-derived osteoblast-enriched cells and in the osteoblastic RCT-3 cell line PGE2 and E1 increased VEGF mRNA and protein levels. The increased expression of VEGF mRNA produced by PGE2 was rapid (maximal at 1 h), transient (declined by 3 h), potentiated by cycloheximide, and abolished by actinomycin D. PGE2 had no effect on VEGF mRNA stability, suggesting transcriptional regulation of VEGF expression by PGF2. Rp-cAMP, a cAMP antagonist, suppressed VEGF mRNA induced by PGE2, indicating cAMP mediation. The upregulation of VEGF expression by PGE2 in the preosteoblastic RCT-1 cells was potentiated by treatment with retinoic acid, which induces the differentiation of these cells. The upregulation of VEGF mRNA by PGE2 was inhibited by dexamethasone treatment. In addition, Northern blot analysis showed that VEGF mRNA is expressed in adult rat tibia. In summary, we documented, for the first time, the expression of VEGF in osteoblasts and in bone tissue. Stimulation of VEGF expression by PGs and its suppression by glucocorticoids, which, respectively, stimulate and suppress bone formation, strongly implicate the involvement of VEGF in bone metabolism.