In Situ Endothelialization Promoted by SEMA4D and CXCL12 for Titanium-Based Biomaterials.

In situ endothelialization, aiming to create implantation surfaces capable of self-endothelialization, seems to be an extremely promising solution, particularly on those blood-contacting surfaces. In this research study, we immobilized the soluble semaphorin 4D (SEMA4D) and C-X-C motif chemokine ligand 12 (CXCL12) biomolecules together with heparin onto the metal-based biomaterial surfaces to achieve in situ endothelialization of modified samples both by stimulating the neighboring endothelial cells (ECs) migration and by capturing the circulating endothelial progenitor cells (EPCs) directly from the blood circulation. X-ray photoelectron spectroscopy data demonstrate the successful immobilization of SEMA4D and CXCL12. Due to the presence of heparin, hemocompatibility was also improved after modification. Although EC migration was mainly mediated by SEMA4D with the coordination of CXCL12, EC proliferation and haptotaxis property were also enhanced, while EC chemotaxis was slightly suppressed because the further immobilization of CXCL12 influences the release of SEMA4D. The results of the ex vivo EPC capturing assay indicated the mobilization of CXCL12 promotes EPC adhesion. In vivo implantation further demonstrated that CXCL12 cooperates with SEMA4D to promote a process of in situ endothelialization.

Constructing bio-layer of heparin and type IV collagen on titanium surface for improving its endothelialization and blood compatibility.

The modification of cardiovascular stent surface for a better micro-environment has gradually changed to multi-molecule, multi-functional designation. In this study, heparin (Hep) and type IV collagen (IVCol) were used as the functional molecule to construct a bifunctional micro-environment of anticoagulation and promoting endothelialization on titanium (Ti). The surface characterization results (AFM, Alcian Blue 8GX Staining and fluorescence staining of IVCol) indicated that the bio-layer of Hep and IVCol were successfully fabricated on the Ti surface through electrostatic self-assembly. The APTT and platelet adhesion test demonstrated that the bionic layer possessed better blood compatibility compared with Ti surface. The adhesion, proliferation, migration and apoptosis tests of endothelial cells proved that the Hep/IVCol layer was able to enhance the endothelialization of the Ti surface. The in vivo animal implantation results manifested that the bionic surface could encourage new endothelialization. This work provides an important reference for the construction of multifunction micro-environment on the cardiovascular scaffold surface.

Endothelialization strategy of implant materials surface: The newest research in recent 5 years.

In recent years, more and more metal or non-metal materials have been used in the treatment of cardiovascular diseases, but the vascular complications after transplantation are still the main factors restricting the clinical application of most grafts, such as acute thrombosis and graft restenosis. Implant materials have been extensively designed and surface optimized by researchers, but it is still too difficult to avoid complications. Natural vascular endodermis has excellent function, anti-coagulant and anti-intimal hyperplasia, and it is also the key to maintaining the homeostasis of normal vascular microenvironment. Therefore, how to promote the adhesion of endothelial cells (ECs) on the surface of cardiovascular materials to achieve endothelialization of the surface is the key to overcoming the complications after implant materialization. At present, the surface endothelialization design of materials based on materials surface science, bioactive molecules, and biological function intervention and feedback has attracted much attention. In this review, we summarize the related research on the surface modification of materials by endothelialization in recent years, and analyze the advantages and challenges of current endothelialization design ideas, explain the relationship between materials, cells, and vascular remodeling in order to find a more ideal endothelialization surface modification strategy for future researchers to meet the requirements of clinical biocompatibility of cardiovascular materials.

NO released via both a Cu-MOF-based donor and surface-catalyzed generation enhances anticoagulation and antibacterial surface effects.

The anticoagulation and antibacterial functions of implant and interventional catheters during indwelling will determine their success or failure. Here, an amino-containing copper-based metal-organic framework (Cu-MOF) coating was prepared on the thermoplastic polyurethane substrate (TPU) surface by spin coating for anti-thrombotic and anti-infection effects. The adhesion properties of the polyurethane prepolymer coating (PC) enhanced the binding force of Cu-MOF particles and TPU surface and improved stability. Due to the coordination affinity of Cu2+ with nitric oxide (NO) and the NO loading capacity of the amino group, it showed that a large amount of NO was loaded in the coating. Meanwhile, the coordinated Cu2+ in the coating also catalyzed endogenous NO donors to generate NO, which prolonged the NO release for up to 30 days. The results of antibacterial experiments showed that the NO released from the coating had good antibacterial effects on both E. coli and S. epidermidis. An obvious antibacterial ring can be seen and the antibacterial rate was higher than 96%. It also showed inhibiting platelet adhesion and activation, prolonged in vitro clotting time and inhibited thrombus formation. In summary, for the first time, NO release from the coating was realized by the combined ways of NO donor and catalytic endogenous NO donor. It can not only meet the high NO release rate required for early anticoagulation and antibacterial of the catheter but also maintain normal anticoagulation requirements in the later period.

[Study on immobilization of heparin on surface of Ti-O films and its antithrombogenicity].

Photoreactive heparin was synthesized by reaction of 4-azidoaniline and heparin. An organic layer was introduced on the surface of Ti-O by 3-aminopropylphosphonic acid assembling, and then the modified heparin was immobilized on the surface by UV irradiation. Water contact angle was used to characterize the hydrophilicity, quantitive assay was done by azure staining methods, and blood compatibility was evaluated by platelet adhesion experiment. Water contact angle of heparinized surface was smaller than that of Ti-O film, which indicated more hydrophilic property of heparinized surface. The surface density of heparin increased with the prolonging of irradiation time and the density was 2.1 microg/cm2 when irradiated for 300s. It showed the heparinized surface was effective in resisting platelets from adhesion and aggregation.

Supramolecular medical antibacterial tissue adhesive prepared based on natural small molecules.

In the field of biomedicine, tissue bio-adhesives require the use of polymer materials with integrated functions to meet changing practical applications. However, the currently available tissue glues cannot balance mechanical properties and biocompatibility. Inspired by the conversion of lipoic acid from small molecular biological sources into high-performance supramolecular polymeric materials, thioctic acid (TA) was modified and polyethylene glycol diacrylate (PEGDA) was introduced. Successfully constructed a dry gel with antibacterial effect and promote infection for wound regeneration. The prepared modified lipoic acid is mixed with PEGDA, melted under mild heating and self-assembled, and then directly extruded on both sides of the wound. It quickly cures at 37 °C and firmly adheres to both sides of the wound. The material exhibits good processability and rapid self-healing ability due to the cross-linked structure of the internal disulfide bonds and thioether bonds. In addition, the characteristics of TA make the prepared xerogels have good tissue adhesion and good antibacterial properties. This work proposes an innovative material with mechanical strength and biocompatible tissue glue, which provides broad prospects for application in wound treatment.

Heparin/poly-l-lysine nanoplatform with growth factor delivery for surface modification of cardiovascular stents: The influence of vascular endothelial growth factor loading.

The rapid re-endothelialization of the vascular stent surface is desirable for preventing thrombosis or reducing restenosis. Many biological factors that promote the biological behavior of endothelial cells have been used for the surface modification of stents. Vascular endothelial growth factor (VEGF), which plays an important role in angiogenesis, induces strong vascular growth. In this study, we investigated different VEGF concentrations (50 to 500 ng/ml) to determine the optimum concentration for biocompatibility. First, VEGF-loaded heparin/poly-l-lysine (Hep-PLL) nanoparticles were created by electrostatic interactions. Then, the VEGF-loaded nanoparticles were immobilized on dopamine-coated 316 L stainless steel (SS) surfaces. The physical and chemical properties of the modified surface were characterized and the biocompatibility was evaluated in vitro. The results indicated that the VEGF-loaded nanoparticles were immobilized successfully on the 316LSS surface, as evidenced by the results of Alcian Blue staining and water contact angle (WCA) measurements. The low platelet adhesion and activation indicated that the modified surfaces had good blood compatibility. The modified surfaces showed a good inhibitory effect on smooth muscle cells, indicating that they inhibited tissue hyperplasia. In addition, the modified surfaces significantly promoted endothelial cell adhesion, proliferation, migration, and biological activity, especially VEGF concentration was 350 ng/ml (NPV350). The optical VEGF concentration of the surface modified Hep-PLL nanoparticles was 350 ng/ml. The proposed method shows promise for potential applications for cardiovascular devices.

[The effect of immobilized laminin on titanium-oxide films to the behavior of human umbilical vein endothelial cells].

In this study, Ti-O films were synthesized using magnetron sputtering, and were pretreated using NaOH solution for improving surface activity from hydroxyl. The laminin(LN) biomacromolecule was further immobilized to the surface through an anminosilane linker. The surface characteristics of these samples were analyzed by Fourier Transform Infrared Spectroscopy, Scanning Electron Microscopy, Atomic Force Microscopy and the contact angle method. Finally, human umbilical vein endothelial cells (HUVEC) were in vitro seeded to the modified and unmodified Ti-O films surface for evaluating the cell compatibility. Survey results suggested that the functional group of hydroxyl was presented onto Ti-O film surface after being pretreated, and laminin could be covalently immobilized to Ti-O film surface by anminosilane linker. The in vitro cell culture results reveal that the biological behaviors of ECs on biochemical modified Ti-O film surface are excellent. The adherence, growth and proliferation of ECs on laminin-immobilized surface were obviously improved when compared to control one. It implies that the laminin immobilizing is helpful to increasing the endothelialization of Ti-O films.

[The influence on the growth of endothelial cells onto titanium-oxide films modified by poly-L-lysine and immobilized with fibronectin].

In this work, a surface biological and chemical modification method was used for improving the biological behavior of endothelial cells onto titanium-oxide films. The titanium-oxide films were first activated by HCl and H2O2 to produce hydroxyl group, then coated with poly-L-lysine and further immobilized with fibronectin. The surface characteristics of samples were analyzed by Fourier Transfer Infrared Spectrum, X-ray Photoelectron Spectroscopy and contact angle method. The biological behavior of cultured human umbilical vein endothelial cell (HUVEC) seeding onto different samples surface was evaluated by the in vitro HUVEC original cultured experiment. The results showed that the method of coating with poly-L-lysine and immobilizing with fibronectin can promote the adhesion and growth of endothelial cells onto titanium-oxide film.

[Study on a novel vascular stent material (titanium oxide, Ti-O) coated with albumin and heparin: is it hemocompatible with fibrinogen].

The functional hemocompatibility between fibrinogen (FIG) and a novel vascular stent material (Ti-O film fixed with albumin and heparin) was investigated as follows: (1) Preparing the new biologic material (Ti-O) film; (2) Coating albumin and heparin on the Ti-O film; (3) Testing platelets (PL) adsorption; (4) Determining FIG adhesion number by use of enzyme linked immunoassay (ELISA); (5) Implanting the films from the test group of Ti-O film and from the comparison group of stainless steel (SS) film into the left and right femoral arteries respectively in 4 dogs. It was proved that albumin and heparin were fixed on Ti-O film. After 6 months, the femoral arteries of the dogs were resected. In the test group of Ti-O film coated with albumin and heparin, few PL adhered to the coat, their form did not change, and no thrombus was found by scanning electron microscopy; the result was better than that of plain Ti-O film, and was much better than that of SS film. Ti-O maintained normal transformation condition of FIG, and no C terminal of gamma chain in FIG was revealed. As it is known whether the hemocompatibility of a biomaterial is good depends upon its adsorption of FIG, and Ti-O has excellent reaction on albumin and heparin by chemical processes. In this study, the Ti-O film coated with albumin and heparin further reduced the absorption of FIG and PL when compared against the plain Ti-O film. So the Ti-O film coated with albumin and heparin has the insistent and permanent anticoagulant character.

[The effect of surface free energy parameters of diamond-like carbon films deposited on medical polyethylene terephthalate on bacterial adhesion].

Diamond-like carbon (DLC) films were deposited by acetylene plasma immersion ion implantation-deposition (PIII-D) on biomedical polyethylene terephthalate (PET). The capacities of Staphylococcus aureus (SA), Staphylococcus epidermidis (SE), Escherichia coli (EC), Pseudomonas aeruginosa (PA) and Candida albicans (CA) for adhesion to PETs are quantitatively determined by the plate counting and Gamma-ray counting of 125I radio labeled bacteria in vitro. The results indicate that the capacities of five types of bacteria for adhesion to PETs are all suppressed by C2H2 PIII-D (P<0.05). The surface energy components of the various substrates and bacteria are calculated based on measurements in water, formamide and diiodomethane and Lifshitz-van del Waals/acid-base approach (LW-AB). The surface free energies obtained are used to calculate the interfacial free energies of adhesion (deltaF(adh)) of five kinds of bacteria on various substrates, and the results show that it is energetically unfavorable for bacterial adhesion to the DLC films already deposited on PET by C2H2 PIII-D.

Design of Multiple Logic Gates Based on Chemically Triggered Fluorescence Switching of Functionalized Polyethylenimine.

In this study, two new functionalized polyethylenimine (PEI), PEIR and PEIQ, have been synthesized by covalently conjugating rhodamine 6G (R6G) or 8-chloroacetyl-aminoquinoline (CAAQ) and have been investigated for their sensing capabilities toward metal ions and anions basing on fluorescence on-off and off-on mechanisms. When triggered by protons, metal ions, or anions, functionalized PEIs can behave as a fluorescence switch, leading to a multiaddressable system. Inspired by these results, functionalized PEI-based logic systems capable of performing elementary logic operations (YES, NOT, NOR, and INHIBIT) and integrative logic operations (OR + INHIBIT) have been constructed by observing the change in the fluorescence with varying the chemical inputs such as protons, metal ions, and anions. Due to its characteristics, such as high sensitivity and fast response, developing functionalized PEI as a new material to perform logic operations may pave a new avenue to construct the next generation of molecular devices with better applicability for biomedical research.

Grafting polyethylenimine with quinoline derivatives for targeted imaging of intracellular Zn(2+) and logic gate operations.

In this study, a highly sensitive and selective fluorescent Zn(2+) probe which exhibited excellent biocompatibility, water solubility, and cell-membrane permeability, was facilely synthesized in a single step by grafting polyethyleneimine (PEI) with quinoline derivatives. The primary amino groups in the branched PEI can increase water solubility and cell permeability of the probe PEIQ, while quinoline derivatives can specifically recognize Zn(2+) and reduce the potential cytotoxicity of PEI. Basing on fluorescence off-on mechanism, PEIQ demonstrated excellent sensing capability towards Zn(2+) in absolute aqueous solution, where a high sensitivity with a detection limit as low as 38.1nM, and a high selectivity over competing metal ions and potential interfering amino acids, were achieved. Inspired by these results, elementary logic operations (YES, NOT and INHIBIT) have been constructed by employing PEIQ as the gate while Zn(2+) and EDTA as chemical inputs. Together with the low cytotoxicity and good cell-permeability, the practical application of PEIQ in living cell imaging was satisfactorily demonstrated, emphasizing its wide application in fundamental biology research.

Tailoring of the dopamine coated surface with VEGF loaded heparin/poly-L-lysine particles for anticoagulation and accelerate in situ endothelialization.

Coronary artery disease is a great threat to human health and is the leading killer worldwide. Percutaneous coronary intervention is the most effective therapy; however, thrombus, and restenosis caused by endothelium injury continue to be problematic after treatment. It is widely accepted that surface biofunctional modification to improve blood compatibility and accelerate endothelialization may be an effective approach to prevent the occurrence of adverse cardiac events. In this study, novel VEGF-loaded heparin/poly-L-lysine (Hep/PLL) particles were developed and immobilized on a dopamine coated titanium surface. The size, distribution, zeta potential, and morphology of the prepared particles were subsequently characterized. The influence of changes in the surface physicochemical properties after particle immobilization was assessed for blood compatibility and cytocompatibility. Surface-modified VEGF-loaded particles significantly inhibited platelet adhesion and activation and were effective in promoting the proliferation and survival of endothelial progenitor cells and endothelial cells. Moreover, Hep/PLL particles were also beneficial for controlling the long-term release of VEGF, which may facilitate endothelium regeneration. In conclusion, VEGF-loaded Hep/PLL particles were successfully immobilized on the Ti surface, and the biocompatibility was significantly improved. This study demonstrates a potential application for the multifunctional modification of stent surfaces for clinical use.

Dynamic Constitution of the Pathogens Inducing Encephalitis in Hand, Foot and Mouth Disease in Kunming, 2009-2011.

Hand, foot and mouth disease (HFMD), caused by various viral pathogens, is an emerging infectious disease in children in Asia. Understanding the composition of these pathogens is necessary to prevent and control this disease. In the present study, the pathogens in 436 HFMD patients (from 2009 to 2011) with concurrent clinical indications of encephalitis, meningoencephalitis, or both, were defined using the semi-nested PCR. A systematic analysis of the composition of these pathogens was performed. Various enteroviruses that are capable of inducing central nervous system (CNS) damage in HFMD patients were identified, including enterovirus 71, coxsackievirus A16, and Echovirus 9. Most of these pathogens were found co-infecting the patients. The composition of the pathogens that induced CNS damage in the HFMD patients was dynamically modulated in the cases.

[Research of plasma adsorption and action of platelet adhesion of Dacron modified by plasma surface modification].

In this paper, polyethylene glycol (PEG) of different molecular weight was grafted on the polyethylene terephthalate (PET, Dacron) films by plasma surface grafting modification. The competitive adsorption relation of plasma (fibrinogen and albumin) adsorbing on materials surface was analyzed in light of surface energy and interface free energy. The results indicated that the PET films grafted PEG long chain molecular possesses the characteristic of preferentially adsorbing albumin and this adsorption tendency of grafted PEG6000 sample is most distinct. The platelet adhesion tests of the PET films whose surfaces were pre-set in contact with fibrinogen and albumin indicated that the surface adsorbing albumin can distinctly inhibit platelet adhesion and aggregation and possess favorable blood compatibility, but the surface adsorbing fibrinogen can enhance platelet adhesion and aggregation.

Endothelialization of implanted cardiovascular biomaterial surfaces: the development from in vitro to in vivo.

Restenosis and thrombosis formation after cardiovascular devices implantation continue to be problematic. Although various platforms and parameters of cardiovascular devices have been designed and optimized over the years, postoperative complications are hard to avoid. The native vascular endothelium always provide a nonthrombogenic surface as well as prevent intimal overproliferation, thereby, the presence of a confluent endothelial cell layer on material surfaces have been widely accepted as an ideal approach to improve the biocompatibility of implanted cardiovascular materials. Endothelialization on biomaterial surfaces is initially developed by in vitro cell seeding. However, numerous no-perfect parts of this method are existed for clinical use. The emergency of endothelial progenitor cells may provide a promising way for setting these limitations. Over the last decades, countless researches about EPCs-based in vivo induced self-endothelialization have been reported and mainly focused on cellular therapy, pharmacological therapy, materials designing, or surface biofunctional modification. This review details the development of endothelialization on cardiovascular material surfaces from in vitro to in vivo. Endothelialization progress on the basis of molecular biological level and bioinformatics theory is expected to be the key point in the coming decades.

Surface modification of implanted cardiovascular metal stents: from antithrombosis and antirestenosis to endothelialization.

Driven by the complications occurring with bare metal stents and drug-eluting stents, concerns have been raised over strategies for long-term safety, with respect to preventing or inhibiting stent thrombosis, restenosis, and in-stent restenosis in particularly. Surface modification is very important in constructing a buffer layer at the interface of the organic and inorganic materials and in ultimately obtaining long-term biocompatibility. In this review, we summarize the developments in surface modification of implanted cardiovascular metal stents. This review focuses on the modification of metal stents via coating drugs or biomolecules to enhance antithrombosis, antirestenosis, and/or endothelialization. In addition, we indicate the probable future work involving the modification of the metallic blood-contacting surfaces of stents and other cardiovascular devices that are under development.

Immobilization of heparin/poly-(L)-lysine nanoparticles on dopamine-coated surface to create a heparin density gradient for selective direction of platelet and vascular cells behavior.

Restenosis, thrombosis formation and delayed endothelium regeneration continue to be problematic for coronary artery stent therapy. To improve the hemocompatibility of the cardiovascular implants and selectively direct vascular cell behavior, a novel kind of heparin/poly-l-lysine (Hep/PLL) nanoparticle was developed and immobilized on a dopamine-coated surface. The stability and structural characteristics of the nanoparticles changed with the Hep:PLL concentration ratio. A Hep density gradient was created on a surface by immobilizing nanoparticles with various Hep:PLL ratios on a dopamine-coated surface. Antithrombin III binding quantity was significantly enhanced, and in plasma the APTT and TT times as coagulation tests were prolonged, depending on the Hep density. A low Hep density is sufficient to prevent platelet adhesion and activation. The sensitivity of vascular cells to the Hep density is very different: high Hep density inhibits the growth of all vascular cells, while low Hep density could selectively inhibit smooth muscle cell hyperplasia but promote endothelial progenitor cells and endothelial cell proliferation. These observations provide important guidance for modification of surface heparinization. We suggest that this method will provide a potential means to construct a suitable platform on a stent surface for selective direction of vascular cell behavior with low side effects.

New strategies for developing cardiovascular stent surfaces with novel functions (Review).

In this review, the authors summarize the developments in surface modification of cardiovascular materials especially in author's laboratory. The authors focus on three different strategies to construct multifunctional surfaces including coimmobilization of various biomolecules on stent surfaces, stem cell based therapy systems, and a single-molecule multipurpose modification strategy in vascular interventional therapy. The roles of various molecules like heparin, gallic acid, various aptamers, and nitric oxide are highlighted in the new strategies for developing cardiovascular stent surfaces with novel functions including excellent hemocompatibility, inhibiting smooth muscle cells proliferation, and native endothelium regeneration. The success of these multifunctional surfaces provides the tremendous potential in designing the next generation of vascular stents.