OBJECTIVETo investigate the value of application of recombinant human VEGF to accelerate flap viability in a rat model of non-ischemic prefabricated flap.

METHODSPrefabricated Flaps were created in 48 SD rats. An autologous tail artery loop was anastomosed to the femoral artery and vein, and implanted subcutaneously in the lower abdomen. Flaps were divided into four groups of 12 each. At the time of loop implantation, the control groups received 0.9% NaCl (Control 1) and 16% (V/W) polyvinyl alcohol (PVA) solution (Control 2). The treatment groups received VEGF in 0.9% NaCl (treatment 1) and VEGF in PVA (treatment 2). In each group, a 3 cm x 4 cm flap nurtured by the tail artery pedicle was elevated and resutured into place after 3, 4 and 5 weeks. The percentage of surviving skin of each flap was determined by planimetry 7 days after flap elevation.

RESULTSMean skin survival areas at 3, 4, and 5 weeks were 1%, 0%, 10% in control; 0%, 16%, 25% in control 2; 3.57%, 39.13%, 75.00% in treatment 1; 8.13%, 41.98%, 58.41% in treatment 2. VEGF significantly improved flap survival by 5 weeks (P < 0.05).

CONCLUSIONThese results suggest VEGF can accelerate maturation of prefabricated flaps.

Animals ; Endothelial Growth Factors ; pharmacology ; Female ; Lymphokines ; pharmacology ; Rats ; Rats, Sprague-Dawley ; Recombinant Proteins ; pharmacology ; Surgical Flaps ; physiology ; Vascular Endothelial Growth Factor A ; Vascular Endothelial Growth Factors

OBJECTIVETo screen for the inhibitor of vascular endothelial growth factor (VEGF) 165 from random peptide library.

METHODSPositive phage clones were rescued after two rounds of panning and competitive elution. Its affinity activity to KDR was monitored through ELISA, immunohistochemical method, Chicken CAM assay and MTT.

RESULTSFive specific binding positive target molecule phage clones were obtained which were able to bind to cells whose surface had high KDR, among which, clone 3 and 13 could effectively block the vascularization of the chorioallantoic membrane of chick embryo, but they were not inhibitive on the proliferation of high KDR expression cells.

CONCLUSIONThe peptides, being the inhibitors of VEGF, may be useful in the treatment of cancers.

Animals ; Binding Sites ; Endothelial Growth Factors ; antagonists & inhibitors ; metabolism ; Enzyme-Linked Immunosorbent Assay ; Humans ; Intercellular Signaling Peptides and Proteins ; metabolism ; Lymphokines ; antagonists & inhibitors ; metabolism ; Peptide Library ; Peptides ; pharmacology ; Vascular Endothelial Growth Factor A ; Vascular Endothelial Growth Factors

BACKGROUNDPigment epithelium-derived factor (PEDF) is expressed in several normal organs and identified as an inhibitor of neovascularization. In the present study, we investigated the effect of PEDF in an in vitro model of ocular choroidal neovascularization.

METHODSMicrodissection was used to isolate the human choroidal endothelial cells (CECs), followed by the use of superparamagnetic beads (Dynabeads) coated with the CD31 antibody, which selectively binds to the endothelial cell surface. The mitogenic and motogenic effects of vascular endothelial growth factor (VEGF) on cultured choroidal capillary endothelial cells were examined in the presence or absence of PEDF (1, 10, 100, and 1000 ng/ml) using cell counts and migration assays.

RESULTSCells bound to the beads were isolated using a magnetic particle concentrator and they were successfully cultured and characterized to be endothelial cells that possessed greater than 95% immunoreactivity to von Willebrand factor. PEDF suppressed the proliferation and migration of VEGF-induced choroidal capillary endothelial cells. However, the concentration of PEDF which we used has little effect on normal CECs.

CONCLUSIONSPEDF played an important role on the growth and migration of VEGF-stimulated choroidal endothelial cell. These findings suggest that PEDF may be an effective approach to the treatment of choroidal neovascular disorders.

Cell Movement ; drug effects ; Cell Proliferation ; drug effects ; Cells, Cultured ; Choroid ; blood supply ; drug effects ; Choroidal Neovascularization ; drug therapy ; Endothelial Cells ; cytology ; drug effects ; Eye Proteins ; pharmacology ; Humans ; Nerve Growth Factors ; pharmacology ; Serpins ; pharmacology ; Vascular Endothelial Growth Factor A ; pharmacology

OBJECTIVETo observe the effect of human vascular endothelial growth factor (VEGF) fragment (3 approximately 4 exon) in E. coli on anti-angiogenesis.

METHODSThrough RT-PCR amplification, endonuclease cut and DNA sequence analysis identification, hVEGF fragment cDNA was inserted into E. coli expression vector pTrcHis2A. The prokaryotic expression plasmid pTrcHis2A/VEGF(3 approximately 4) was constructed and transformed into TOP10F.

RESULTSAfter 8hr isopropy-beta-D-thiogalactoside (IPTG) induction, VEGF fragment was expressed in 15% of total proteins through sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE). The expressed protein was highly antigenic and specific. The VEGF fragment was further purified by affinity, which could inhibit HUVEC proliferation and neovascularization of the chick chorioallantoic membrane.

CONCLUSIONVEGF fragment is anti-angiogenetic, which may potentially be used in oncologico-biological targeting therapy.

Angiogenesis Inhibitors ; isolation & purification ; pharmacology ; Cloning, Molecular ; Endothelial Growth Factors ; genetics ; isolation & purification ; pharmacology ; Escherichia coli ; genetics ; Gene Expression ; drug effects ; Genetic Vectors ; Humans ; Intercellular Signaling Peptides and Proteins ; genetics ; isolation & purification ; pharmacology ; Isopropyl Thiogalactoside ; pharmacology ; Lymphokines ; genetics ; isolation & purification ; pharmacology ; Peptide Fragments ; genetics ; isolation & purification ; pharmacology ; Plasmids ; genetics ; Polymerase Chain Reaction ; Vascular Endothelial Growth Factor A ; Vascular Endothelial Growth Factors

To induce endothelial cell, canine bone marrow-derived mesenchymal stem cells (MSCs) were separated from bone marrow by density gradient centrifugation. The isolated MSCs were induced to form endothelial-like cell in the presence of vascular endothelial growth factor (VEGF), endothelial growth factor (EGF) and so on. These results showed that the cells uniformly took on a cobblestone morphology under the light microscope, and cell nucleolus was in the middle of the cells. The cells displayed Weibel-Palade bodies under the transmission electron microscope. vWF, a specific marker of endothelial cell was positive in the cells. The above results demonstrate that MSCs may be differentiated into endothelial cells in vitro.
Animals ; Bone Marrow Cells ; cytology ; Cell Culture Techniques ; Cell Differentiation ; Cells, Cultured ; Dogs ; Endothelial Cells ; cytology ; Endothelial Growth Factors ; pharmacology ; Mesenchymal Stromal Cells ; cytology ; Vascular Endothelial Growth Factor A ; pharmacology

OBJECTIVETo investigate the effect of local application of bFGF combined with sucralfate on the cell proliferation during continuous tissue expansion (CTE).

METHODSNine white pigs were selected to undergo the continuous tissue expansion in this study and treated with bFGF and sucralfate, respectively as the following groups: group 1 with both bFGF and sucralfate, group 2 only with bFGF, group 3 with only sucralfate, and group 4 with saline as control. Fifteen samples were taken in each pig for immunohistochemistry analysis 1-14 days and 6 weeks after the operation.

RESULTSIn the group with both bFGF and sucralfate, the epidermic basal cells proliferated significantly after the operation and reached top level in 3 days, which was statistical higher than the control group, but the multiplication of basal cell was the lowest 14 days after the operation, still more than the control group. In dermal layer, proliferation of fibroblasts, vessel endothelial cells, hair follicles epidermic cells and sweat gland epicytes was also significant higher in the group with both bFGF and sucralfate than that the control group and reached top level 7 day after the operation, but the proliferation of cells decreased obviously 14 days after the operation, still higher than the control group. The mitotic activity of cells returned to the basal level in 42 days. There were no significant differences among the group 2, group 3 and group 4.

CONCLUSIONLocal application of both bFGF and sucralfate could be more effect to induce cells multiplication during early skin expansion to facilitate the growth of neoformed skin soft tissue.

Animals ; Cell Proliferation ; drug effects ; Endothelial Cells ; cytology ; drug effects ; Fibroblast Growth Factors ; pharmacology ; Fibroblasts ; cytology ; drug effects ; Sucralfate ; pharmacology ; Swine ; Time Factors ; Tissue Expansion ; Tissue Expansion Devices

Tumor angiogenesis is a complicated process in which VEGF and the members of Ang family have been proposed to play an important role. Ang family is the only vascular growth factor family including activators and inhibitors. They can all bind with specific tyrosine kinase receptor Tie-2 and participate in angiogenesis. Vascular endothelial growth factor (VEGF) is secreted by tumor cells and coordinates with Ang to promote angiogenesis. Further research on the structure and functional mechanism of VEGF and Ang will contribute to develop a new type of antiangiogenesis medicine, which might have good perspect in clinical practice. In this paper, the structure and action mechanism of Ang family and its receptor Tie-2, the application of Ang family and Tie in tumor therapy, and the synergic mechanism between Ang and VEGF were summarized.
Angiogenesis Inhibitors ; pharmacology ; Angiopoietin-2 ; pharmacology ; Animals ; Drug Synergism ; Humans ; Neoplasms ; blood supply ; Neovascularization, Pathologic ; prevention & control ; Vascular Endothelial Growth Factors ; pharmacology

To explore the feasibility of mesenchymal stem cells (MSCs) acting as seed cells in tissue engineering, we isolated human bone marrow MSCs and differentiated them into vascular endothelial-like cells (ELCs) in vitro. Bone marrow mononuclear cells (BMSCs) were isolated by the method of percoll density centrifugation, and seeded in Dulbecco Modified Eagle Medium supplemented with 10% fetal bovine serum. MSCs were purified through multiple adherent cultures, and differentiated into ELCs induced by endothelial cell growth medium-2 (EBM-2) medium containing vascular endothelial growth factor (VEGF), human fibroblast growth factor (hFGF), insulin like growth factors 1 (IGF-1), and human epidermal growth factor (hEGF). The relative biologic characteristics of ELCs including cell morphology and phenotype were studied by inverted microscope and flow cytometry. The induced cells were identified by immunofluorescence with CD31 and Von Willebrand factor (vWF). The results showed that the morphology of MSCs was long-spindle and vortex-like growth. After induction of differentiation, the cells were round, and similar to vascular endothelial cells (ECs). Flow cytometric analysis revealed that ELCs expressed ECs specific surface markers of CD31 and vascular endothelial cadherin (VE-cadherin), but not CD133. Immunofluorescence results also confirmed that ELCs expressed CD31 and vWF. The results suggested that ELCs possed similar cell biological characteristics with ECs. In one word, human MSCs derived from bone marrow have the potential to differentiate into ECs in vitro, and show clinical feasibility acting as ideal donor cells of vascular tissue engineering.
Antigens, CD ; metabolism ; Bone Marrow Cells ; Cadherins ; metabolism ; Cell Culture Techniques ; Cell Differentiation ; Cells, Cultured ; Culture Media ; chemistry ; Endothelial Cells ; cytology ; Epidermal Growth Factor ; pharmacology ; Fibroblast Growth Factors ; pharmacology ; Flow Cytometry ; Humans ; Insulin-Like Growth Factor I ; pharmacology ; Mesenchymal Stromal Cells ; cytology ; Platelet Endothelial Cell Adhesion Molecule-1 ; metabolism ; Tissue Engineering ; Vascular Endothelial Growth Factor A ; pharmacology ; von Willebrand Factor ; metabolism

After ischemic stroke, there is neovascularization around the infarcted area, which is called penumbra. Angiogenesis and arteriogenesis are responsible for the new vessel formation. Until recently, vasculogenesis has been proved to involve mechanisms in postischemic neovascularization, which was thought to be restricted to embryonic development. New blood vessels' formation is a complex pathologic process after ischemic stroke, in which many factors are properly involved. There are factors stimulating neovascularization, such as vascular endothelial growth factor, platelet-derived growth factor, basic fibroblast growth factor and angiopoietin; there are also factors inhibiting neovascularization, such as thrombospondin. Functional recovery was found after stroke, which may contribute to angiogensis in the periinfarct tissue. Thus, therapeutic angiogenesis has been initially studied in animal models, but there is still a long way to go for therapeutic angiogenesis to be used in the treatment of stroke patient.
Angiogenesis Inducing Agents ; pharmacology ; Angiopoietin-1 ; metabolism ; Brain ; blood supply ; Brain Infarction ; metabolism ; physiopathology ; Humans ; Neovascularization, Physiologic ; Platelet-Derived Growth Factor ; metabolism ; Vascular Endothelial Growth Factors ; metabolism

The objective of this study was to explore the effects of BMP-4 and VEGF on the development of primary hematopoietic stem cells during the differentiation of embryonic stem cells (ESCs) into embryoid body (EB). Murine E14 ESCs were seeded into semisolid methylcellulose-based medium for EB formation. According to added or not cytokines, experiments were divided into: (1) group of spontaneous differentiation without cytokine as control; (2) group of BMP-4 in different concentrations (0, 5, 15, 25 and 50 ng/ml); (3) group of BMP-4 combined with VEGF; (4) group of VEGF alone. EBs were collected on days 3, 6, 9, 12, 15, and the proportion of Flk-1(+) cells were assayed by flow cytometry. The results showed that in the different BMP-4 concentration groups, the proportions of Flk-1(+) cells were significantly different, and it reached the peak values in 25 ng/ml BMP-4 group as 6.51 +/- 1.02% at day 3 and 7.70 +/- 1.12% at day 6 respectively, which were statistically higher than those in control group without-BMP-4 and in 5 ng/ml BMP-4 group (p < 0.05). When BMP-4 was used in combination with VEGF, Flk-1(+) cells went to peak proportion value at day 9 as 27.53 +/- 8.14%, which was statistically higher than that in spontaneous differentiation group as 8.77 +/- 2.35% (p < 0.05) and VEGF treatment group as 11.21 +/- 2.23% (p < 0.05). It is concluded that BMP-4 in combination with VEGF can promote Flk-1(+) cells genesis during EB formation in vitro, which provides experimental evidence for researches on directed differentiation of ESCs into hematopoietic stem cells simulating the microenvironment in vivo.
Animals ; Bone Morphogenetic Protein 4 ; pharmacology ; Cell Differentiation ; drug effects ; Cells, Cultured ; Embryonic Stem Cells ; cytology ; drug effects ; Hematopoietic Stem Cells ; cytology ; Mice ; Vascular Endothelial Growth Factors ; pharmacology