Behavior Of Human Umbilical Vein Endothelial Cells Research Articles

Fabrication of chitin-fibrin hydrogels to construct the 3D artificial extracellular matrix scaffold for vascular regeneration and cardiac tissue engineering.

As the cornerstone of tissue engineering and regeneration medicine research, developing a cost-effective and bionic extracellular matrix (ECM) that can precisely modulate cellular behavior and form functional tissue remains challenging. An artificial ECM combining polysaccharides and fibrillar proteins to mimic the structure and composition of natural ECM provides a promising solution for cardiac tissue regeneration. In this study, we developed a bionic hydrogel scaffold by combining a quaternized β-chitin derivative (QC) and fibrin-matrigel (FM) in different ratios to mimic a natural ECM. We evaluated the stiffness of those composite hydrogels with different mixing ratios and their effects on the growth of human umbilical vein endothelial cells (HUVECs). The optimal hydrogels, QCFM1 hydrogels were further applied to load HUVECs into nude mice for in vivo angiogenesis. Besides, we encapsulated human pluripotent stem cell-derived cardiomyocytes (hPSC-CMs) into QCFM hydrogels and employed 3D bioprinting to achieve batch fabrication of human-engineered heart tissue (hEHT). Finally, the myocardial structure and electrophysiological function of hEHT were evaluated by immunofluorescence and optical mapping. Designed artificial ECM has a tunable modulus (220-1380 Pa), which determines the different cellular behavior of HUVECs when encapsulated in these. QCFM1 composite hydrogels with optimal stiffness (800 Pa) and porous architecture were finally identified, which could adapt for in vitro cell spreading and in vivo angiogenesis of HUVECs. Moreover, QCFM1 hydrogels were applied in 3D bioprinting successfully to achieve batch fabrication of both ring-shaped and patch-shaped hEHT. These QCFM1 hydrogels-based hEHTs possess organized sarcomeres and advanced function characteristics comparable to reported hEHTs. The chitin-derived hydrogels are first used for cardiac tissue engineering and achieve the batch fabrication of functionalized artificial myocardium. Specifically, these novel QCFM1 hydrogels provided a reliable and economical choice serving as ideal ECM for application in tissue engineering and regeneration medicine.

Comparison of the Behavior of 3D-Printed Endothelial Cells in Different Bioinks.

Biomaterials with characteristics similar to extracellular matrix and with suitable bioprinting properties are essential for vascular tissue engineering. In search for suitable biomaterials, this study investigated the three hydrogels alginate/hyaluronic acid/gelatin (Alg/HA/Gel), pre-crosslinked alginate di-aldehyde with gelatin (ADA-GEL), and gelatin methacryloyl (GelMA) with respect to their mechanical properties and to the survival, migration, and proliferation of human umbilical vein endothelial cells (HUVECs). In addition, the behavior of HUVECs was compared with their behavior in Matrigel. For this purpose, HUVECs were mixed with the inks both as single cells and as cell spheroids and printed using extrusion-based bioprinting. Good printability with shape fidelity was determined for all inks. The rheological measurements demonstrated the gelling consistency of the inks and shear-thinning behavior. Different Young's moduli of the hydrogels were determined. However, all measured values where within the range defined in the literature, leading to migration and sprouting, as well as reconciling migration with adhesion. Cell survival and proliferation in ADA-GEL and GelMA hydrogels were demonstrated for 14 days. In the Alg/HA/Gel bioink, cell death occurred within 7 days for single cells. Sprouting and migration of the HUVEC spheroids were observed in ADA-GEL and GelMA. Similar behavior of the spheroids was seen in Matrigel. In contrast, the spheroids in the Alg/HA/Gel ink died over the time studied. It has been shown that Alg/HA/Gel does not provide a good environment for long-term survival of HUVECs. In conclusion, ADA-GEL and GelMA are promising inks for vascular tissue engineering.

Human Umbilical Vein Endothelial Cells Form a Network on a Hyaluronic Acid/Gelatin Composite Hydrogel Moderately Crosslinked and Degraded by Hydrogen Peroxide.

The study of the capillary-like network formation of human umbilical vein endothelial cells (HUVECs) in vitro is important for understanding the factors that promote or inhibit angiogenesis. Here, we report the behavior of HUVECs on the composite hydrogels containing hyaluronic acid (HA) and gelatin with different degrees of degradation, inducing the different physicochemical properties of the hydrogels. The hydrogels were obtained through horseradish peroxidase (HRP)-catalyzed hydrogelation consuming hydrogen peroxide (H2O2, 16 ppm) supplied from the air, and the degradation degree was tuned by altering the exposure time to the air. The HUVECs on the composite hydrogel with intermediate stiffness (1.2 kPa) obtained through 120 min of the exposure were more elongated than those on the soft (0.4 kPa) and the stiff (2.4 kPa) composite hydrogels obtained through 15 min and 60 min of the exposure, respectively. In addition, HUVECs formed a capillary-like network only on the stiff composite hydrogel although those on the hydrogels with comparable stiffness but containing gelatin alone or alginate instead of HA did not form the network. These results show that the HA/gelatin composite hydrogels obtained through the H2O2-mediated crosslinking and degradation could be a tool for studies using HUVECs to understand the promotion and inhibition of angiogenesis.

Behavior of Human Umbilical Vein Endothelial Cells on Titanium Surfaces Functionalized with VE-Cadherin Extracellular 1-4 Domains.

Angiogenesis is essential for the optimal functioning of orthopedic medical implants. Protein functionalization of implant surfaces can improve tissue integration through proper vascularization and prevent implant failure in patients lacking sufficient angiogenesis. The aim of this study was to evaluate the angiogenic activity of titanium surfaces functionalized with recombinant VE-cadherin extracelluar1-4 (VE-CADEC1-4) protein in human umbilical vein endothelial cells (HUVECs). After titanium discs were coated with recombinant VE-CADEC1-4 protein at appropriate concentrations, the behavior of HUVECs on the VE-CADEC1-4-functionalized titanium discs were evaluated by cell adhesion assay, proliferation assay, and real-time RT-PCR. Recombinant VE-CADEC1-4-functionalized titanium surfaces improved the adhesion of HUVECs by 1.8-fold at the optimal concentration, and the proliferative activity was 1.3-fold higher than the control at 14 days. In addition, when angiogenesis markers were confirmed by real-time RT-PCR, PECAM-1 increased approximately 1.2-fold, TEK approximately 1.4-fold, KDR approximately 1.6-fold, and Tie-1 approximately 2.1-fold compared to the control. Recombinant VE-CADEC1-4-functionalized titanium surfaces improved cell adhesion, proliferation, and angiogenic differentiation of HUVECs, suggesting that the VE-CADEC1-4-functionalization of titanium surfaces can offer angiogenic surfaces with the potential to improve bone healing in orthopedic applications.

MiR-195 alleviates oxygen–glucose deprivation/reperfusion-induced cell apoptosis via inhibition of IKKα-mediated NF-κB pathway

Objectives Increasing evidence confirmed that miRNA plays a critical role in the occurrence and development of ischemic stroke. Here, the aim of this study was to examine the function and mechanisms of miR-195 in vascular endothelial cell apoptosis induced by oxygen–glucose deprivation (OGD). Methods This study intended to use OGD to simulate ischemia in vitro. The mRNA expression of miR-195, IKKα and NF-κB in human umbilical vein endothelial cells (HUVECs) were detected by RT-qPCR. The proliferation and apoptosis ability of HUVECs were evaluated using MTT assay, colony formation assay and flow cytometry, respectively. Western blot was applied to examine related protein expression. The interaction between miR-195 and IKKα was verified by dual-luciferase reporter gene assay. Results OGD significantly inhibited cell viability and induced cell apoptosis in HUVECs. Meanwhile, OGD treatment notably decreased the expression of miR-195, as well as enhanced NF-κB expression. Moreover, miR-195 directly interacted with IKKα and suppressed its expression. Mechanically, overexpression of miR-195 exhibited pro-proliferation and anti-apoptotic effect on HUVECs treated with OGD through targeting IKKα-mediated NF-κB pathway. At the molecular level, through suppressing IKKα/NF-κB pathway, miR-195 inhibited the expression of pro-apoptotic protein Bax and active caspase-3, but increased the expression of anti-apoptotic Bcl-2 in HUVECs. Conclusions Our finding uncovers the protective effect of miR-195 on the biological behavior of HUVECs via suppression of the NF-κB pathway induced by IKKα, which may provide a new potential strategy for ischemic stroke clinical treatment.

Effect of lipopolysaccharide on the adherence of human umbilical vein endothelial cells (HUVEC) on a natural substrate.

Polymers are often contaminated with lipopolysaccharides also known as endotoxins. Even small amounts of endotoxins can have strong effects on endothelial cell function so that the endothelialisation of cardiovascular implants might be hampered. An open question is how endothelial cells seeded on a body foreign substrate respond to shear load after adding Lipid A (LPA), the domain, which is responsible for much of the toxicity of gram-negative bacteria, and whether morphological changes of endothelial cells occur.LPA supplementation to the culture medium in increasing concentrations (5, 25 and 50μg/ml) resulted in progressive reductions of the density of adherent HUVEC after shear load (p < 0.001). 48% of the HUVEC in control cultures (0μg/ml LPA) were still adherent after 2 hours of shearing at 6 dyne/cm2, while 80 minutes after addition of 50μg/ml LPA, 88% of the HUVEC had already detached from the substrate and after 100 minutes no more HUVEC were attached.The results demonstrate that endotoxins are of extreme importance for the behavior of HUVEC and that in vivo pathologies can be increasingly simulated in vitro.

Immobilization of Heparin on Bacterial Nanocellulose Hydrogels Induces Tubulogenesis of Human Endothelial Cells

Models that mimic the angiogenesis initial processes, as adhesion, migration, proliferation and tubulogenesis, are extremely valuable for investigating the action of new anti-cancer drugs. There still is a need for an angiogenesis model that reflects in vivo environment. To address this challenge, we developed a 3D matrix well-defined based on covalently immobilization of heparin (HEP) on bacterial nanocellulose (BNC) hydrogels. Successful immobilization was confirmed by qualitative and quantification analysis. Human umbilical vein endothelial cells (HUVECs) were seeded on bottom and top surfaces of BNC and BNC-HEP hydrogels and cells behavior were analyzed. The bottom surfaces of BNC-HEP hydrogels were able to support cell adhesion and promote proliferation and tubulogenesis formation. Results here presented indicate that the tubulogenesis process could be controlled by physico-chemical properties of the developed hydrogel. The interaction between the bioactive molecule, heparin, and the particular microstructure of BNC induced tubulogenenic behavior of HUVECs in vitro.

Strontium-Doped Bioactive Glass Nanoparticles in Osteogenic Commitment.

The present work has explored bioactive glass nanoparticles (BGNPs) and developed strontium-doped nanoparticles (BGNPsSr), envisioning orthopedic strategies compatible with vascularization. The nanoparticles were synthesized by the sol-gel method, achieving a diameter of 55 nm for BGNPs and 75 nm for BGNPsSr, and the inclusion of strontium caused no structural alteration. The nanoparticles exhibited high cytocompatibility for human umbilical vein endothelial cells (HUVECs) and SaOS-2. Additionally, the incorporation of strontium emphasized the tubule networking behavior of HUVECs. Our results demonstrate that the nanoparticle dissolution products encouraged the osteogenic differentiation of human adipose stem cells as it favored the expression of key genes and proteins associated with osteogenic lineage. This effect was markedly enhanced for BGNPsSr, which could prompt stem cell osteogenic differentiation without the typical osteogenic inducers. This study not only supports the hypothesis that BGNPs might play a significant role in osteogenic commitment but also highlights that the designed BGNPsSr is a valuable tool for stem cell "tune-up" in bone tissue engineering applications.

Gadd45a knockdown alleviates oxidative stress through suppressing the p38 MAPK signaling pathway in the pathogenesis of preeclampsia

ObjectivePreeclampsia (PE) is a hypertensive disease associated with vascular oxidative stress (OS). Besides, cell response to OS triggers growth arrest and DNA damage inducible alpha (Gadd45a). Herein, we investigated the effect of Gadd45a on OS in PE. MethodsUmbilical cord tissues and peripheral blood samples were collected from PE patients and normal pregnant women. Immunohistochemistry was applied to identify Gadd45a level and extent of p38 phosphorylation. To verify the effect of Gadd45a on CRL1730 human umbilical vein endothelial cell (HUVEC) behavior, HUVECs were treated with hypoxia/reoxygenation (H/R) and Gadd45a-siRNA or P79350. OS products were detected using thiobarbituric acid-reactive-substance assay, immune enzyme assay, enzyme-linked immunosorbent assay (ELISA) and super oxide dismutase (SOD) kit. HUVEC behaviors were evaluated using flow cytometry, wound-healing test, and Transwell assay. The tube formation ability was assessed using in vitro tube formation assay. ResultsPE umbilical cord tissues exhibited increased Gadd45a expression, extent of p38 phosphorylation, 8-isoprostane and oxidatively modified low density lipoprotein (Ox-LDL) and decreased SOD levels. HUVECs treated with H/R or agonist + H/R showed similar expression tendencies of related genes, enhanced apoptosis and inhibited migration, invasion, and blood vessel formation. Gadd45a-siRNA conferred alleviation to OS with decreased apoptosis, greater cell migration, invasion and blood vessel, which appeared to be weakened by treatment of Gadd45a siRNA + P79350. ConclusionOver-expression of Gadd45a and activation of the p38 MAPK signaling pathway are associated with OS in PE. Furthermore, Gadd45a knockdown promotes cell migration and invasion, and the tube formation, thus alleviating OS in PE.

Influence of Micropatterned Silk Fibroin Films on Human Umbilical Endothelial Cell Behaviors

To design functional vascular grafts, it is essential to explore the interactions between endothelial cells and the surface of substrate biomaterials. The present study thus aimed at evaluating the influence of micropatterned silk fibroin films on human umbilical vein endothelial cell (HUVEC) behaviors including adhesion, migration, proliferation, and protein expressions. Silk fibroin films with different grafting micropatterns (10 μm × 10 μm, 20 μm × 20 μm, 20 μm × 30 μm, 40 μm × 40 μm) were prepared by soft-lithographic technique. HUVECs were shown to attach and grow well on the silk fibroin films. The results indicated that the 10 × 10 and 40 × 40 gratings induced higher proliferation rates as compared to the 20 × 20 and 20 × 30 groups. However, the increased orientation and alignment degree and stronger protein expressions (CD31, VWF and Vinculin) were observed on the 20 × 20 and 20 × 30 groups. Overall, this study suggested that the HUVEC behaviors could be regulated in vitro by different micropattern topographies. The micropatterned silk fibroin films may offer a potential improvement for the development of functional vascular grafts.

Engineering a Dual-Layer Chitosan–Lactide Hydrogel To Create Endothelial Cell Aggregate-Induced Microvascular Networks In Vitro and Increase Blood Perfusion In Vivo

Here, we report the use of chemically cross-linked and photo-cross-linked hydrogels to engineer human umbilical vein endothelial cell (HUVEC) aggregate-induced microvascular networks to increase blood perfusion in vivo. First, we studied the effect of chemically cross-linked and photo-cross-linked chitosan-lactide hydrogels on stiffness, degradation rates, and HUVEC behaviors. The photo-cross-linked hydrogel was relatively stiff (E = ∼15 kPa) and possessed more compact networks, denser surface texture, and lower enzymatic degradation rates than the relatively soft, chemically cross-linked hydrogel (E = ∼2 kPa). While both hydrogels exhibited nontoxicity, the soft chemically cross-linked hydrogels expedited the formation of cell aggregates compared to the photo-cross-linked hydrogels. Cells on the less stiff, chemically cross-linked hydrogels expressed more matrix metalloproteinase (MMP) activity than the stiffer, photo-cross-linked hydrogel. This difference in MMP activity resulted in a more dramatic decrease in mechanical stiffness after 3 days of incubation for the chemically cross-linked hydrogel, as compared to the photo-cross-linked one. After determining the physical and biological properties of each hydrogel, we accordingly engineered a dual-layer hydrogel construct consisting of the relatively soft, chemically cross-linked hydrogel layer for HUVEC encapsulation, and the relatively stiff, acellular, photo-cross-linked hydrogel for retention of cell-laden microvasculature above. This dual-layer hydrogel construct enabled a lasting HUVEC aggregate-induced microvascular network due to the combination of stable substrate, enriched cell adhesion molecules, and extracellular matrix proteins. We tested the dual-layer hydrogel construct in a mouse model of hind-limb ischemia, where the HUVEC aggregate-induced microvascular networks significantly enhanced blood perfusion rate to ischemic legs and decreased tissue necrosis compared with both no treatment and nonaggregated HUVEC-loaded hydrogels within 2 weeks. This study suggests an effective means for regulating hydrogel properties to facilitate a stable, HUVEC aggregate-induced microvascular network for a variety of vascularized tissue applications.

Trefoil Factor-3 (TFF3) Stimulates De Novo Angiogenesis in Mammary Carcinoma both Directly and Indirectly via IL-8/CXCR2.

Mammary carcinoma cells produce pro-angiogenic factors to stimulate angiogenesis and tumor growth. Trefoil factor-3 (TFF3) is an oncogene secreted from mammary carcinoma cells and associated with poor prognosis. Herein, we demonstrate that TFF3 produced in mammary carcinoma cells functions as a promoter of tumor angiogenesis. Forced expression of TFF3 in mammary carcinoma cells promoted proliferation, survival, invasion and in vitro tubule formation of human umbilical vein endothelial cells (HUVEC). MCF7-TFF3 cells with forced expression of TFF3 generated tumors with enhanced microvessel density as compared to tumors formed by vector control cells. Depletion of TFF3 in mammary carcinoma cells by siRNA concordantly decreased the angiogenic behavior of HUVEC. Forced expression of TFF3 in mammary carcinoma cells stimulated IL-8 transcription and subsequently enhanced IL-8 expression in both mammary carcinoma cells and HUVEC. Depletion of IL-8 in mammary carcinoma cells with forced expression of TFF3, or antibody inhibition of IL-8, partially abrogated mammary carcinoma cell TFF3-stimulated HUVEC angiogenic behavior in vitro, as did inhibition of the IL-8 receptor, CXCR2. Depletion of STAT3 by siRNA in MCF-7 cells with forced expression of TFF3 partially diminished the angiogenic capability of TFF3 on stimulation of cellular processes of HUVEC. Exogenous recombinant hTFF3 also directly promoted the angiogenic behavior of HUVEC. Hence, TFF3 is a potent angiogenic factor and functions as a promoter of de novo angiogenesis in mammary carcinoma, which may co-coordinate with the growth promoting and metastatic actions of TFF3 in mammary carcinoma to enhance tumor progression.

Differential effects of a soluble or immobilized VEGFR-binding peptide

Regulating endothelial cell behavior is a key step in understanding and controlling neovascularization for both pro-angiogenic and anti-angiogenic therapeutic strategies. Here, we characterized the effects of a covalently immobilized peptide mimic of vascular endothelial growth factor, herein referred to as VEGF receptor-binding peptide (VR-BP), on human umbilical vein endothelial cell (HUVEC) behavior. Self-assembled monolayer arrays presenting varied densities of covalently immobilized VR-BP and varied densities of the fibronectin-derived cell adhesion peptide Gly-Arg-Gly-Asp-Ser-Pro (GRGDSP) were used to probe for changes in HUVEC attachment, proliferation and tubulogenesis. In a soluble form, VR-BP exhibited pro-angiogenic effects in agreement with previous studies, indicated by increases in HUVEC proliferation. However, when presented to cells in an insoluble context, covalently immobilized VR-BP inhibited several pro-angiogenic HUVEC behaviors, including attachment and proliferation, and also inhibited HUVEC response to soluble recombinant VEGF protein. Furthermore, substrates with covalently immobilized VR-BP also modulated HUVEC tubulogenesis when a matrigel overlay assay was used to provide cells with a pseudo-three dimensional environment. Taken together, these results demonstrate that the context in which ligands are presented to cell surface receptors strongly influences their effects, and that the same ligand can be an agonist or an antagonist depending on the manner of presentation to the cell.

Effect of hydrogen on the behavior of cultured human umbilical vein endothelial cells (HUVEC) on titanium oxide films fabricated by plasma immersion ion implantation and deposition

In this paper, titanium oxide (Ti–O) thin films were fabricated by plasma immersion ion implantation and deposition (PIII-D), and subsequently hydrogen ions were implanted into the film surface by PIII to improve the biocompatibility of blood and vascular contacting biomaterials and prosthesis such as stents, etc. The films were characterized using X-ray diffraction (XRD), X-ray photoelectron spectroscopy (XPS), secondary ion mass spectroscopy (SIMS), and atomic force microscopy (AFM). The contact angle of Ti–O films with simulated body fluid was measured. The biological behavior of cultured human umbilical vein endothelial cells (HUVECs) on Ti–O film surfaces was evaluated by an in vitro cultured HUVEC experiment. The in vitro investigation of cultured ECs on Ti–O films revealed that the biological behavior of HUVECs, such as adherence, growth and proliferation etc., were better biologically for hydrogen-containing Ti–O films than for untreated Ti–O films. The results of our study support the hypothesis that the surface morphology and biological properties such as wettability and surface tension etc. of the implanted Ti–O films are changed to a more desirable range by the high energy H plasma immersion ion implantation and by H incorporation in the films. As a result, the H-containing Ti–O films possess good biocompatibility.

Stability of angiogenic agents, ginsenoside Rg 1 and Re, isolated from Panax ginseng: In vitro and in vivo studies

The study was designed to investigate the stability of ginsenoside Rg 1 (Rg 1) and Re (Re), two natural herbal compounds isolated from Panax ginseng, based on their activity to promote angiogenesis in vitro and in vivo. After being treated at different temperatures, pHs, and solvent species for distinct durations, the remaining activities of Rg 1 and Re on human umbilical vein endothelial cell (HUVEC) proliferation, migration, and tube formation were examined in vitro. Additionally, the remaining activity of each treated test agent, mixed in a growth factor-reduced Matrigel, in stimulating angiogenesis was evaluated subcutaneously in a mouse model. Basic fibroblast growth factor (bFGF) was used as a control. It was found in vitro that HUVEC proliferation, migration in a Transwell plate, and tube formation on Matrigel were all significantly enhanced in the presence of bFGF, Rg 1, or Re. However, after being treated at different temperatures, pHs, or solvent species, the remaining activity of bFGF on HUVEC behaviors reduced significantly. This observation was more significant with increasing the duration of treatment. In contrast, the activities of Rg 1 and Re remained unchanged throughout the entire course of the study. The in vivo results observed on day 7 after implantation showed that the blank control (Matrigel alone) was slightly vascularized. In contrast, the density of neo-vessels in the Matrigel plug mixed with bFGF, Rg 1, or Re was significantly enhanced. However, after being treated, the density of neo-vessels was significantly reduced in the Matrigel plug mixed with bFGF, while those of Rg 1 and Re remained unchanged. The aforementioned results suggested that Rg 1 and Re could be a novel group of nonpeptide angiogenic agents with a superior stability and may be used for the management of tissue regeneration.

Antithrombogenic investigation and biological behavior of cultured human umbilical vein endothelial cells on Ti-O film

Our previous research results have shown that the Ti-O films with appropriate characteristics possess great potentials for biomaterials application. In this paper, using plasma immersion ion implantation and deposition (PIII-D), titanium oxide thin films are fabricated onto silicon wafer. The antithrombogenesis of films is evaluated in vitro through the platelet adhesion investigation. The biological behavior of human umbilical vein endothelial cells (HUVEC) on the film surface is investigated in vitro by endothelial cell (EC) culture. Our results reveal that the crystalline Ti-O films exhibit attractive blood compatibility. The in vitro HUVEC-cultured investigation of Ti-O film surface has justified that the biological behavior of HUVECs on different structure surfaces is significantly different. The adherence, growth and proliferation of HUVECs to the crystalline Ti-O film surface are in order, by forming a perfect single layer, preserving the natural original shape and displaying the cobblestone road metal rank, and obviously superior to that on the amorphous Ti-O film surface. According to our study, the crystalline Ti-O film, with proper microstructure, is helpful for seeding ECs and can be used as a functional surface for the adherence and growth of ECs.

Behavior of cultured human umbilical vein endothelial cells on titanium oxide films fabricated by plasma immersion ion implantation and deposition

Our previous research results have shown that Ti–O films with appropriate characteristics possessed good blood compatibility. In this paper, we synthesized Ti–O films using plasma immersion ion implantation and deposition (PIII-D) and investigated the characteristics and the biological behavior of cultured human umbilical vein endothelial cells (HUVEC) onto film surface. The structure and composition of the Ti–O films were investigated using X-ray diffraction (XRD), X-ray photoelectron spectroscopy (XPS), and atom force microscopy (AFM). The surface energy was measured by a contact angle method. The in vitro HUVEC-cultured investigation of Ti–O film surface justified that the biological behavior of HUVEC on different structure surfaces was significantly different. The adherence and growth of HUVEC to crystalline Ti–O film surface were much better than that of amorphous Ti–O film. The mechanism based on the relationship between the film characteristics and biological behavior of endothelial cells (ECs) is discussed.