Inactivation of Src family kinases inhibits angiogenesis in vivo: implications for a mechanism involving organization of the actin cytoskeleton

Inhibition of angiogenesis could be a treatment strategy for diseases such as cancer, rheumatoid arthritis, and diabetic retinopathy. PP2 is a pharmacological inhibitor of Src family kinases and was found to inhibit FGF-2 induced angiogenesis in vivo. Experiments in vitro showed that PP2 inhibited invasive growth and sprouting of both endothelial and vascular smooth muscle cells into a fibrin matrix. PP2 inhibited the formation of lamellopodia and expression of kinase inactive c-Src reduced phosphorylation of cortactin and paxillin, suggesting a model in which Src kinases are involved in organization of the actin cytoskeleton. Consequently, endothelial cells expressing kinase inactive c-Src failed to spread and form cord-like structures on a collagen matrix. These data suggest that pharmacological inactivation of Src family kinases inhibits FGF-2 stimulated angiogenesis by interference with organization of the actin cytoskeleton in both endothelial and vascular smooth muscle cells, which affects cell migration.     

Interaction of plasminogen-related protein B with endothelial and smooth muscle cells in vitro

Plasminogen-related protein B (PRP-B) closely resembles the N-terminal plasminogen activation peptide, which is released from plasminogen during conversion to plasmin. We have previously demonstrated that the steady-state level of mRNA encoding PRP-B is increased within tumor tissues, and that recombinant PRP-B antagonizes neoplastic growth when administered systemically to mice harboring tumors, but no insights into the cell targets of PRP-B have been presented. Employing serum-free medium optimized for culturing human endothelial or smooth muscle cells, we show that recombinant PRP-B inhibits basic fibroblast growth factor-dependent cell migration for both cell types, as well as tube formation of endothelial cells. Comparison with the angiogenesis inhibitors angiostatin and endostatin revealed similar results. Recombinant PRP-B is effective in promoting cell attachment of endothelial and smooth muscle cells, and antibody interference experiments reveal that the interaction of recombinant PRP-B with endothelial cells is mediated at least in part by alpha(v)-containing integrins. Inhibition of angiogenesis in vivo by PRP-B was demonstrated in the chicken chorioallantoic membrane assay. PRP-B and other antiangiogenic molecules may elicit metabolic perturbations in endothelial cells as well as perivascular mesenchymal cells such as smooth muscle cells and pericytes.     

Neovascular responses induced by cultured aortic endothelial cells

Neovascularization was studied in the chorioallantoic membrane of the chick embryo after implantation of bovine aortic endothelial and smooth muscle cells, Swiss and BALB/c 3T3 cells and human diploid fibroblasts cultured separately on microcarrier beads. Quantitative analysis of neovascularization indicated a 3 1/2-fold increase in the number of blood vessels responding to endothelial cells while smooth muscle cells induced a twofold increase when compared to the response of beads without cells. Skin fibroblasts and Swiss 3T3 cells did not elicit a comparable response. The marked angiogenic response induced by endothelial cells was characterized by a 137% increase in total vessel length and a 35% increase in average vessel area when compared to controls. Two of the properties required for an angiogenesis factor--stimulation of cellular migration and proliferation--can also be demonstrated using endothelial cell-conditioned medium in cell culture systems. Medium from cultured bovine aortic endothelium stimulates DNA synthesis, proliferation, and migration of smooth muscle cells. In addition, conditioned media from both endothelial cells and smooth muscle cells produced an angiogenic response in the chorioallantoic membrane assay, which was comparable to that produced by intact cells growing on microcarrier beads. Similar responses were not evident with medium conditioned by other cell types. These results indicate the potential importance of endothelial cells and endothelial cell products in regulating blood vessel growth.     

Hepatoma-derived growth factor is a pulmonary endothelial cell-expressed angiogenic factor

Hepatoma-derived growth factor (HDGF) was previously identified as a developmentally regulated cardiovascular and renal gene that is mitogenic for vascular smooth muscle and aortic endothelial cells. As reciprocal interactions of smooth muscle and endothelial cells are necessary for vascular formation, we examined whether HDGF plays a role in angiogenesis. According to immunohistochemistry, HDGF was highly expressed in endothelial cells of nonmuscularized, forming blood vessels of the fetal lung. HDGF was also expressed in endothelial cells of small (20 microm) mature arteries and veins. By Western immunoblotting, HDGF was highly expressed by human pulmonary microvascular endothelial cells in vitro. Adenoviral overexpression of HDGF was mitogenic for human pulmonary microvascular endothelial cells in serum-free medium, stimulating a 1.75-fold increase in bromodeoxyuridine (BrdU) uptake and a twofold increase in cell migration. With the chick chorioallantoic membrane (CAM), a biologic assay for angiogenesis, exogenous recombinant HDGF significantly stimulated blood vessel formation and a dose-dependent reorganization of cells within the CAM into a more compact, linear alignment reminiscent of tube formation. According to double immunostaining for endothelial cells with a transforming growth factor-betaII receptor antibody and BrdU as a marker of cell proliferation, exogenous HDGF selectively stimulated endothelial cell BrdU uptake. HDGF also activated specific ERK1/2 signaling and did not overlap with VEGF SAPK/JNK, Akt-mediated pathways. We conclude that HDGF is a highly expressed vascular endothelial cell protein in vivo and is a potent endothelial mitogen and regulator of endothelial cell migration by mechanisms distinct from VEGF.     

Synthesis and structure-activity relationship of diarylamide derivatives as selective inhibitors of the proliferation of human endothelial cells

A series of diarylamide urea derivatives were synthesized and evaluated for their inhibitory activities against human coronary artery endothelial cells (ECs) and human coronary artery smooth muscle cells (SMCs). Compound was superior to Tranilast, in terms of both cell selectivity and the potency of its inhibitory activity toward the proliferation and angiogenesis of ECs.     

Quiescence, cell viability, apoptosis and necrosis of smooth muscle cells using different growth inhibitors

Smooth muscle cells and endothelial cells play an important role in cardiovascular diseases and may therefore be a potential target for gene therapy. Most in vitro experiments are performed using proliferating cell cultures. Nevertheless, non-dividing cells would represent more realistic in vivo conditions for gene therapy. Therefore, a simple method to achieve physiologically quiescence in cell cultures is needed for experiments. Growth to confluence is sufficient for endothelial cells to reach quiescence, in contrast to smooth muscle cells. Alternative techniques were investigated to achieve quiescence for smooth muscle cells. N-acetyl-cysteine, heparin, aphidicolin and serum-free medium are known inhibitors of smooth muscle cell proliferation and were tested for cell viability, necrosis and apoptosis. The inhibition status was evaluated counting cells in a cell counter. Toxicity, necrosis and apoptosis were determined using FACS analysis. Then, smooth muscle cells and endothelial cells were transfected with plasmid containing the beta-galactosidase gene using liposomes. Analysis of gene expression in transfected cells included a quantitative beta-galactosidase assay and X-gal staining. Growth inhibition was achieved with all agents tested. Using N-acetyl-cysteine, only slightly reduced growth rates were observed. Aphidicolin stopped cell growth almost immediately, but demonstrated enhanced toxicity. The amount of apoptotic and necrotic cells was lowest using heparin in the presence of foetal calf serum. Transfection experiments using stationary cultures of smooth muscle cells using heparin or aphidicolin demonstrated 5-10-fold lower transfection rates compared to transfected proliferating cell cultures serving as controls. Transfection experiments using stationary cultures of endothelial cells using growth inhibition through confluence demonstrated 40-fold lower transfection rates than transfected proliferating cell cultures. Transfer efficiency was much lower in endothelial cells compared to smooth muscle cells. In conclusion, quiescent cells simulate more realistically the in vivo situation and may therefore represent a better model for future in vivo experiments based on in vitro findings.     

Two-chain high molecular weight kininogen induces endothelial cell apoptosis and inhibits angiogenesis: partial activity within domain 5.

We previously reported that the binding of two-chain high molecular weight kininogen (HKa) to endothelial cells may occur through interactions with endothelial urokinase receptors. Since the binding of urokinase to urokinase receptors activates signaling responses and may stimulate mitogenesis, we assessed the effect of HKa binding on endothelial cell proliferation. Unexpectedly, HKa inhibited proliferation in response to several growth factors, with 50% inhibition caused by approximately 10 nM HKa. This activity was Zn(2+) dependent and not shared by either single-chain high molecular weight kininogen (HK) or low molecular weight kininogen. HKa selectively inhibited the proliferation of human umbilical vein and dermal microvascular endothelial cells, but did not affect that of umbilical vein or human aortic smooth muscle cells, trophoblasts, fibroblasts, or carcinoma cells. Inhibition of endothelial proliferation by HKa was associated with endothelial cell apoptosis and unaffected by antibodies that block the binding of HK or HKa to any of their known endothelial receptors. Recombinant HK domain 5 displayed activity similar to that of HKa. In vivo, HKa inhibited neovascularization of subcutaneously implanted Matrigel plugs, as well as rat corneal angiogenesis. These results demonstrate that HKa is a novel inhibitor of angiogenesis, whose activity is dependent on the unique conformation of the two-chain molecule.     

Vasculogenic and angiogenic potential of adipose stromal vascular fraction cell populations in vitro

Adipose-derived stromal vascular fraction (SVF) is a heterogeneous cell source that contains endothelial cells, pericytes, smooth muscle cells, stem cells, and other accessory immune and stromal cells. The SVF cell population has been shown to support vasculogenesis in vitro as well vascular maturation in vivo. Matrigel, an extracellular matrix (ECM) mixture has been utilized in vitro to evaluate tube formation of purified endothelial cell systems. We have developed an in vitro system that utilizes freshly isolated SVF and ECM molecules both in pure form (fibrin, laminin, collagen) as well as premixed form (Matrigel) to evaluate endothelial tip cell formation, endothelial stalk elongation, and early stages of branching and inosculation. Freshly isolated SVF rat demonstrate cell aggregation and clustering (presumptive vasculogenesis) on Matrigel ECM within the first 36 h of seeding followed by tip cell formation, stalk cell formation, branching, and inosculation (presumptive angiogenesis) during the subsequent 4 days of culture. Purified ECM molecules (laminin, fibrin, and collagen) promote cell proliferation but do not recapitulate events seen on Matrigel. We have created an in vitro system that provides a functional assay to study the mechanisms of vasculogenesis and angiogenesis in freshly isolated SVF to characterize SVF’s blood vessel forming potential prior to clinical implantation.     

Cultured endothelial cells produce heparinlike inhibitor of smooth muscle cell growth

Using cultured cells from bovine and rat aortas, we have examined the possibility that endothelial cells might regulate the growth of vascular smooth muscle cells. Conditioned medium from confluent bovine aortic endothelial cells inhibited the proliferation of growth-arrested smooth muscle cells. Conditioned medium from exponential endothelial cells, and from exponential or confluent smooth muscle cells and fibroblasts, did not inhibit smooth muscle cell growth. Conditioned medium from confluent endothelial cells did not inhibit the growth of endothelial cells or fibroblasts. In addition to the apparent specificity of both the producer and target cell, the inhibitory activity was heat stable and not affected by proteases. It was sensitive flavobacterium heparinase but not to hyaluronidase or chondroitin sulfate ABC lyase. It thus appears to be a heparinlike substance. Two other lines of evidence support this conclusion. First, a crude isolate of glycosaminoglycans (TCA-soluble, ethanol-precipitable material) from endothelial cell-conditioned medium reconstituted in 20 percent serum inhibited smooth muscle cell growth; glycosaminoglycans isolated from unconditioned medium (i.e., 0.4 percent serum) had no effect on smooth muscle cell growth. No inhibition was seen if the glycosaminoglycan preparation was treated with heparinase. Second, exogenous heparin, heparin sulfate, chondroitin sulfate B (dermatan sulfate), chondroitin sulfate ABC, and hyaluronic acid were added to 20 percent serum and tested for their ability to inhibit smooth muscle cell growth. Heparin inhibited growth at concentrations as low as 10 ng/ml. Other glycosaminoglycans had no effect at doses up to 10 μg/ml. Anticoagulant and non- anticoagulant heparin were equally effective at inhibiting smooth muscle cell growth, as they were in vivo following endothelial injury (Clowes and Karnovsk. Nature (Lond.). 265:625-626, 1977; Guyton et al. Circ. Res. 46:625-634, 1980), and in vitro following exposure of smooth muscle cells to platelet extract (Hoover et al. Circ. Res. 47:578-583, 1980). We suggest that vascular endothelial cells may secrete a heparinlike substance in vivo which may regulate the growth of underlying smooth muscle cells.     

大鼠主动脉内皮细胞和平滑肌细胞的原代培养及生物学特性比较

目的培养大鼠主动脉平滑肌细胞和内皮细胞,细胞纯化与鉴定,比较生物学特性的差异。方法采用血管环贴壁法培养动脉内皮细胞,组织块贴壁法培养动脉平滑肌细胞,并采用有限稀释法挑选内皮细胞单克隆,免疫细胞荧光鉴定二者的特异性标志,相差显微镜观察二者单个细胞及细胞群体在形态上的差异性,CCK-8试剂盒检测细胞的增殖,比较二者对胰酶消化,粘附,冻存后复苏的情况。结果血管环贴壁法成功培养血管内皮细胞,组织块培养法成功培养出血管平滑肌细胞,内皮细胞能够形成单克隆集落,培养的细胞均表达相应的特异性标志,内皮细胞增殖速度和平滑肌细胞有差异,内皮细胞对胰酶的耐受性较差,内皮细胞粘附所需时间短,对冻存后的耐受性较好。结论组织块贴壁法适合内皮细胞和平滑肌细胞的培养,有限稀释法能够纯化原代培养的内皮细胞,大鼠主动脉平滑肌细胞和内皮细胞在细胞形态、增殖、粘附、对胰酶的反应、冻存后复苏均存在差异。     

Functional roles of angiogenic factors and receptors on non‐endothelial cells in the oropharyngeal cavity of the chukar partridge (Alectoris chukar)

The vascular endothelial growth factor (VEGF) family belong to the platelet‐derived growth factor supergene family and is involved in angiogenesis and mitogenesis. The VEGF–VEGFR system regulates endothelial cell proliferation, migration, vascular permeability, secretion and other non‐endothelial cells functions. To clarify the possible role of endothelial and non‐endothelial cells, VEGF and its receptors, vascular endothelial cell growth inhibitor (VEGI) were immunohistochemically examined in oropharyngeal organs. Ten adult partridges were used in this study and the pharynx and larynx were dissected together with the palate and tongue. VEGI, VEGF and its receptor were highly expressed in luminal epithelial and stromal cells, when compared to glandular epithelial and muscle cells (P < 0.05). Moreover, VEGF, its receptors and VEGI were expressed rather strongly in the endothelial cells of the blood capillaries and in both the endothelial and smooth muscle cells of the large and small blood vessels. In conclusion, VEGF and its receptors (flt1/fms, flk1/KDR and flt4) and VEGI were expressed by various cell groups at varying intensity in the oropharyngeal organs. This demonstrates that they play a critical role in the regulation and maintenance of the functions in cells different from endothelial ones as well as in cell proliferation, differentiation, apoptosis and angiogenesis.     

Comparative endocrinology-paracrinology-autocrinology of human adult large vessel endothelial and smooth muscle cells

Summary Endothelial and smooth muscle cells were isolated from human adult large blood vessels to compare their proliferative response to hormones and growth factors. Neural extracts and the medium from differentiated hepatoma cells were used as concentrated sources of required hormones and growth factors that supported both cell types. Active hormones and growth factors were identified from the neural extracts and hepatoma medium by substitution or direct isolation and biochemical characterization. Epidermal growth factor, lipoproteins, and heparin-binding growth factors elicited growth-stimulatory effects on both endothelial and smooth muscle cells. Both types of human vascular cells displayed 7600 to 8600 specific heparin-binding growth factor receptors per cell with a similar apparent dissociation constant (Kd) of 200 to 250 pM. Heparin modified the response of both endothelial and smooth muscle cells to heparin-binding growth factors dependent on the type of heparin-binding growth factor and amount of heparinlike material present. In addition, heparin exerted a growth factor-independent inhibition of smooth muscle cell proliferation. Platelet-derived growth factor, insulinlike growth factors, and glucocorticoid specifically supported proliferation of smooth muscle cells with no apparent effect on endothelial cell proliferation. Growth-factorlike proteinase inhibitors had an impact specifically on endothelial cell proliferation. Transforming growth factor beta was a specific inhibitor of endothelial cells, but had a positive effect on smooth muscle cell proliferation. The results provide a framework for differential control of the two vascular cell types at normal and atherosclerotic blood vessel sites by the balance among positive and negative effectors of endocrine, paracrine and autocrine origin. This research was supported by NIH grants CA37589, HL33847, and AM35310 from the National Institutes of Health, Bethesda, MD; grant 1718 from the Council for Tobacco Research; and a grant from RJR/Nabisco, Inc.     

Vascular smooth muscle cell growth-promoting factor/F-spondin inhibits angiogenesis via the blockade of integrin alphavbeta3 on vascular endothelial cells

Vascular smooth muscle cell growth-promoting factor (VSGP) was originally isolated from bovine ovarian follicular fluid as a stimulator of vascular smooth muscle cell proliferation. Homology searches indicate that bovine and human VSGPs are orthologs of rat F-spondin. Here, we examined whether recombinant human VSGP/F-spondin affected the biological activities of endothelial cells. VSGP/F-spondin did not affect the proliferation of human umbilical vein endothelial cells (HUVECs); however, it did inhibit VEGF- or bFGF-stimulated HUVEC migration. To clarify the mechanism of this inhibitory effect, we examined the adhesion of HUVECs to extracellular matrix proteins. VSGP/F-spondin specifically inhibited the spreading of HUVECs on vitronectin via the functional blockade of integrin alphavbeta3. As a result, VSGP/F-spondin inhibited the tyrosine phosphorylation of focal adhesion kinase (FAK) when HUVECs were plated on vitronectin. Moreover, VSGP/F-spondin inhibited the activation of Akt when HUVECs on vitronectin were stimulated with VEGF. VSGP/F-spondin inhibited tube formation by HUVECs in vitro and neovascularization in the rat cornea in vivo. These results indicate that VSGP/F-spondin inhibits angiogenesis at least in part by the blockade of endothelial integrin alphavbeta3.     

Early treatment with fumagillin, an inhibitor of methionine aminopeptidase-2, prevents Pulmonary Hypertension in monocrotaline-injured rats

Pulmonary Hypertension (PH) is a pathophysiologic condition characterized by hypoxemia and right ventricular strain. Proliferation of fibroblasts, smooth muscle cells, and endothelial cells is central to the pathology of PH in animal models and in humans. Methionine aminopeptidase-2 (MetAP2) regulates proliferation in a variety of cell types including endothelial cells, smooth muscle cells, and fibroblasts. MetAP2 is inhibited irreversibly by the angiogenesis inhibitor fumagillin. We have previously found that inhibition of MetAP2 with fumagillin in bleomycin-injured mice decreased pulmonary fibrosis by selectively decreasing the proliferation of lung myofibroblasts. In this study, we investigated the role of fumagillin as a potential therapy in experimental PH. In vivo, treatment of rats with fumagillin early after monocrotaline injury prevented PH and right ventricular remodeling by decreasing the thickness of the medial layer of the pulmonary arteries. Treatment with fumagillin beginning two weeks after monocrotaline injury did not prevent PH but was associated with decreased right ventricular mass and decreased cardiomyocyte hypertrophy, suggesting a direct effect of fumagillin on right ventricular remodeling. Incubation of rat pulmonary artery smooth muscle cells (RPASMC) with fumagillin and MetAP2-targeting siRNA inhibited proliferation of RPASMC in vitro. Platelet-derived growth factor, a growth factor that is important in the pathogenesis of PH and stimulates proliferation of fibroblasts and smooth muscle cells, strongly increased expression of MetP2. By immunohistochemistry, we found that MetAP2 was expressed in the lesions of human pulmonary arterial hypertension. We propose that fumagillin may be an effective adjunctive therapy for treating PH in patients.     

Role of lipoproteins in growth of human adult arterial endothelial and smooth muscle cells in low lipoprotein-deficient serum

Recently improved culture conditions for human adult arterial endothelial and smooth muscle cells from a wide variety of donors have been used to study the effects of lipoproteins on proliferation of both cell types in low serum culture medium. Optimal growth of endothelial and smooth muscle cells in an optimal nutrient medium (MCDB 107) containing epidermal growth factor, a partially purified fraction from bovine brain, and 1% (v/v) lipoprotein-deficient serum was dependent on either high- or low-density lipoprotein. High- and low-density lipoprotein stimulated cell growth by three- and five-fold, respectively, over a 6-day period. Optimal stimulation of both endothelial and smooth muscle cell growth occurred between 20 and 60 micrograms/ml of high- and low-density lipoproteins, respectively. No correlation between the activation of 3-hydroxyl-3-methylglutaryl coenzyme. A reductase activity and lipoprotein-stimulated cell proliferation was observed. Lipid-free total apolipoproteins or apolipoprotein C peptides from high-density lipoprotein were partially effective and together with oleic acid effectively replaced native high-density lipoprotein for the support of endothelial cell growth. In contrast, apolipoproteins or apolipoprotein C peptides from high-density lipoprotein alone or with oleic acid had no effect on smooth muscle cell proliferation. The results suggest a functional role of high- and low-density lipoproteins and apolipoproteins in the proliferation of human adult endothelial and smooth muscle cells.     

Specific inhibition of endothelial cell proliferation by thrombospondin.

Angiogenesis is a multi-step event involving endothelial cell migration, attachment, and proliferation. A thrombospondin (TSP)-like protein has recently been described as a naturally-occurring inhibitor of angiogenesis. We now report that human platelet TSP inhibits the in vitro proliferation of endothelial cells from the rabbit corpus luteum, bovine adrenal cortex and pulmonary artery, and human umbilical vein. The antiproliferative effect of TSP was neutralized by monoclonal antibodies against TSP. The growth arrest seen with TSP was specific for endothelial cells since TSP actually stimulated the growth of vascular smooth muscle cells and human foreskin fibroblasts. These results imply that the angiogenesis-inhibiting effect of TSP is mediated through an inhibition of endothelial cell proliferation. Elucidation of the mechanism of action of TSP on endothelial cell proliferation may lead to potential therapeutic approaches for the control of neovascular diseases.     

The role of fructose-1, 6-diphosphate in cell migration and proliferation in an in vitro xenograft blood vessel model of vascular wound healing

Summary Both smooth muscle cells and endothelial cells play an important role in vascular wound healing. To elucidate the role of fructose-1, 6-diphosphate, cell proliferation and cell migration studies were performed with human endothelial cells and rat smooth muscle cells. To mimic blood vessels, endothelial and smooth muscle cells were used in 1:10, 1:5, and 1:1 concentrations, respectively, mimicking large-, mid-, and capillary-sized blood vessels. Cell migration was studied with fetal bovine serum-starved cells. For cell proliferation assay, cells were plated at 30–50% confluency and then starved. The cells were incubated for 48 h with fructose-1, 6-diphosphate at (per ml) 10 mg, 1 mg, 500 μg, 250 μg, 100 μg, and 10 μg, pulsed with tritiated-thymidine and incubated with 1 N NaOH for 30 min at room temperature, harvested, and counted. For migration assay, confluent cells were starved, wounded, and incubated for 24 h with same concentrations of fructose-1, 6-diphosphate as in proliferation assay. The cells were fixed and counted. Smooth muscle cell proliferation was inhibited by fructose-1, 6-diphosphate at 10 mg/ml. In the xenograft models of 1:10, 1:5, and 1:1 fructose-1, 6-diphosphate inhibited proliferation at 10 mg/ml. In migration studies 10 mg fructose-1, 6-diphosphate per ml was inhibitory to both cell types. In large-, mid-, and capillary-sized blood vessels, fructose-1, 6-diphosphate inhibited proliferation of both cell types at 10 mg/ml. At the individual cell level, fructose-1, 6-diphosphate is nonstimulatory to proliferation of endothelial cells while inhibiting migration, and it acts on smooth muscle cells by inhibiting both proliferation and migration.     

The role of vascular-derived perlecan in modulating cell adhesion,proliferation and growth factor signaling

Smooth muscle cell proliferation can be inhibited by heparan sulfate proteoglycans whereas the removal or digestion of heparan sulfate from perlecan promotes their proliferation. In this study we characterized the glycosaminoglycan side chains of perlecan isolated from either primary human coronary artery smooth muscle or endothelial cells and determined their roles in mediating cell adhesion and proliferation, and in fibroblast growth factor (FGF) binding and signaling. Smooth muscle cell perlecan was decorated with both heparan sulfate and chondroitin sulfate, whereas endothelial perlecan contained exclusively heparan sulfate chains. Smooth muscle cells bound to the protein core of perlecan only when the glycosaminoglycans were removed, and this binding involved a novel site in domain III as well as domain V/endorepellin and the α₂β₁ integrin. In contrast, endothelial cells adhered to the protein core of perlecan in the presence of glycosaminoglycans. Smooth muscle cell perlecan bound both FGF1 and FGF2 via its heparan sulfate chains and promoted the signaling of FGF2 but not FGF1. Also endothelial cell perlecan bound both FGF1 and FGF2 via its heparan sulfate chains, but in contrast, promoted the signaling of both growth factors. Based on this differential bioactivity, we propose that perlecan synthesized by smooth muscle cells differs from that synthesized by endothelial cells by possessing different signaling capabilities, primarily, but not exclusively, due to a differential glycanation. The end result is a differential modulation of cell adhesion, proliferation and growth factor signaling in these two key cellular constituents of blood vessels.