Heparin‐Controlled Growth of Polypyrrole Nanowires

Summary: Heparin, a potent anticoagulant, has been used for the first time for the synthesis of PPy nanowires serving not only as an anion dopant but also as an effective morphology‐directing agent. The obtained PPy nanowires exhibit long and fine structures with smooth surface and the average diameter of the nanowires is about 90–100 nm and lengths are several hundred nanometers to micrometers. The possible formation mechanism of PPy nanowires may be related to the chain structure of heparin with functional groups ( SO and  COO⁻) on the surface. The effect of concentrations of pyrrole monomers and heparin on the morphology and size of PPy nanowires has been investigated.

SEM image of PPy nanowires synthesized in the presence of heparin.     

Preactivation‐Based,One‐Pot Combinatorial Synthesis of Heparin‐like Hexasaccharides for the Analysis of Heparin–Protein Interactions

Heparin (HP) and heparan sulfate (HS) play important roles in many biological events. Increasing evidence has shown that the biological functions of HP and HS can be critically dependent upon their precise structures, including the position of the iduronic acids and sulfation patterns. However, unraveling the HP code has been extremely challenging due to the enormous structural variations. To overcome this hurdle, we investigated the possibility of assembling a library of HP/HS oligosaccharides using a preactivation‐based, one‐pot glycosylation method. A major challenge in HP/HS oligosaccharide synthesis is stereoselectivity in the formation of the cis‐1,4‐linkages between glucosamine and the uronic acid. Through screening, suitable protective groups were identified on the matching glycosyl donor and acceptor, leading to stereospecific formation of both the cis‐1,4‐ and trans‐1,4‐linkages present in HP. The protective group chemistry designed was also very flexible. From two advanced thioglycosyl disaccharide intermediates, all of the required disaccharide modules for library preparation could be generated in a divergent manner, which greatly simplified building‐block preparation. Furthermore, the reactivity‐independent nature of the preactivation‐based, one‐pot approach enabled us to mix the building blocks. This allowed rapid assembly of twelve HP/HS hexasaccharides with systematically varied and precisely controlled backbone structures in a combinatorial fashion. The speed and the high yields achieved in glycoassembly without the need to use a large excess of building blocks highlighted the advantages of our approach, which can be of general use to facilitate the study of HP/HS biology. As a proof of principle, this panel of hexasaccharides was used to probe the effect of backbone sequence on binding with the fibroblast growth factor‐2 (FGF‐2). A trisaccharide sequence of 2‐O‐sulfated iduronic acid flanked by N‐sulfated glucosamines was identified to be the minimum binding motif and N‐sulfation was found to be critical. This provides useful information for further development of more potent compounds towards FGF‐2 binding, which can have potential applications in wound healing and anticancer therapy.     

Synthesis of the Heparin‐Based Anticoagulant Drug Fondaparinux

Fondaparinux, a synthetic pentasaccharide based on the heparin antithrombin‐binding domain, is an approved clinical anticoagulant. Although it is a better and safer alternative to pharmaceutical heparins in many cases, its high cost, which results from the difficult and tedious synthesis, is a deterrent for its widespread use. The chemical synthesis of fondaparinux was achieved in an efficient and concise manner from commercially available D ‐glucosamine, diacetone α‐D ‐glucose, and penta‐O‐acetyl‐D ‐glucose. The method involves suitably functionalized building blocks that are readily accessible and employs shared intermediates and a series of one‐pot reactions that considerably reduce the synthetic effort and improve the yield.     

Inhibition of Inflammation‐Associated Thrombosis with ROS‐Responsive Heparin‐DOCA/PVAX Nanoparticles

Inflammation‐associated thrombosis is a non‐negligible source of mortalities and morbidities worldwide. To manipulate inflammation‐associated coagulation, nanoparticles that contain anti‐inflammatory polymer (copolyoxalate containing vanillyl alcohol, PVAX) and anti‐thrombotic heparin derivative deoxycholic acid (Hep‐DOCA) are prepared. The strategy takes advantage of the reducted side effects of heparin through heparin conjugation, achievement of long‐term anti‐inflammation by inflammation‐trigged release of anti‐inflammatory agents, and formation of PVAX/heparin‐DOCA nanoparticles by co‐self‐assembly. It is demonstrated that the Hep‐DOCA conjugate and PVAX are synthesized successfully; PVAX and Hep‐DOCA nanodrugs (HDP) are obtained by co‐assembly; the HDP nanoparticles effectively reduce the inflammation and coagulation without inducing lethal bleeding both in vivo and in vitro. The method provided here is versatile and effective, which paves new way to develop nanodrugs to treat inflammation‐associated thrombosis safely.     

Rapid,Label‐free Optical Spectroscopy Platform for Diagnosis of Heparin‐Induced Thrombocytopenia

The use of surface‐enhanced Raman spectroscopy (SERS) to determine spectral markers for the diagnosis of heparin‐induced thrombocytopenia (HIT), a difficult‐to‐diagnose immune‐related complication that often leads to limb ischemia and thromboembolism, is proposed. The ability to produce distinct molecular signatures without the addition of labels enables unbiased inquiry and makes SERS an attractive complementary diagnostic tool. A capillary‐tube‐derived SERS platform offers ultrasensitive, label‐free measurement as well as efficient handling of blood serum samples. This shows excellent reproducibility, long‐term stability and provides an alternative diagnostic rubric for the determination of HIT by leveraging machine‐learning‐based classification of the spectroscopic data. We envision that a portable Raman instrument could be combined with the capillary‐tube‐based SERS analytical tool for diagnosis of HIT in the clinical laboratory, without perturbing the existing diagnostic workflow.     

Super‐Anticoagulant Heparin‐Mimicking Hydrogel Thin Film Attached Substrate Surfaces to Improve Hemocompatibility

In this study, heparin‐mimicking hydrogel thin films are covalently attached onto poly(ether sulfone) membrane surfaces to improve anticoagulant property. The hydrogel films display honeycomb‐like porous structure with well controlled thickness and show long‐term stability. After immobilizing the hydrogel films, the membranes show excellent anticoagulant property confirmed by the activated partial thromboplastin time values exceeding 600 s. Meanwhile, the thrombin time values increase from 20 to 61 s as the sodium allysulfonate proportions increase from 0 to 80 mol%. In vitro investigations of protein adsorption and blood‐related complement activation also confirm that the membranes exhibit super‐anticoagulant property. Furthermore, gentamycin sulfate is loaded into the hydrogel films, and the released drug shows significant inhibition toward E. coli bacteria. It is believed that the surface attached heparin‐mimicking hydrogel thin films may show high potential for the applications in various biological fields, such as blood contacting materials and drug loading materials.

     

Heparin‐gold Nanoparticles and Liquid Crystal Applied in Label‐free Electrochemical Immunosensor for Prostate‐specific Antigen

A label‐free electrochemical immunosensor based on the liquid crystal (E)‐1‐decyl‐4‐[(4‐decyloxyphenyl)diazenyl]pyridinium bromide (Br−Py), together with heparin‐stabilized gold nanoparticles (AuNP‐Hep) and Nafion is proposed for the determination of prostate‐specific antigen (PSA). The Br−Py liquid crystal presented redox properties and good film‐forming abilities on the electrode surface, and thus it is a suitable alternative as a redox probe for a label‐free electrochemical immunosensor, which could simplify the analysis methodology. The stepwise construction of the immunosensor and the incubation process (immunocomplex formation) were characterized by voltammetry and electrochemical impedance spectroscopy. The proposed immunosensor could directly detect PSA concentrations in the incubation samples, based on the suppression of the Br−Py redox peak (‘base peak’) current. After optimization, the immunosensor exhibited a linear response to PSA concentrations in the range of 0.1 to 50 ng mL⁻¹, with a calculated detection limit of 0.08 ng mL⁻¹. The reproducibility (coefficient of variance less than 3.0 %), selectivity and accuracy of the methodology were adequate. The immunosensor was satisfactorily applied in the quantification of PSA in human blood plasma samples.     

Hemocompatible and Bioactive Heparin‐Loaded PCL‐α‐TCP Fibrous Membranes for Bone Tissue Engineering

The combination of bioactive components such as calcium phosphates and fibrous structures are encouraging niche‐mimetic keys for restoring bone defects. However, the importance of hemocompatibility of the membranes is widely ignored. Heparin‐loaded nanocomposite poly(ε‐caprolactone) (PCL)‐α‐tricalcium phosphate (α‐TCP) fibrous membranes are developed to provide bioactive and hemocompatible constructs for bone tissue engineering. Nanocomposite membranes are optimized based on bioactivity, mechanical properties, and cell interaction. Consequently, various concentrations of heparin molecules are loaded within nanocomposite fibrous membranes. In vitro heparin release profiles reveal a sustained release of heparin over the period of 14 days without an initial burst. Moreover, heparin encapsulation enhances mesenchymal stem cell (MSC) attachment and proliferation, depending on the heparin content. It is concluded that the incorporation of heparin within TCP–PCL fibrous membranes provides the most effective cellular interactions through synergistic physical and chemical cues.     

Electrospun Heparin‐Loaded Core–Shell Nanofiber Sutures for Achilles Tendon Regeneration In Vivo

Achilles tendon reconstruction surgery is the primary clinical method for repairing acute Achilles tendon ruptures. However, the efficacy of the postoperative healing process and the recovery of physiological function are inadequate. This study examines the healing mechanism of ruptured rat Achilles tendons seamed with heparin‐loaded core–shell fiber sutures fabricated via near‐field electrospinning. High‐heparin‐concentration sutures (PPH3.0) perform better than the low‐heparin‐concentration sutures and commercial sutures (CSs). The PPH3.0 suture recruits fewer inflammatory cells and shows good histocompatibility in peritoneal implantation experiments. Staining of the Achilles tendon rupture repair zone demonstrates that a high heparin concentration in sutures reduces immune‐inflammatory responses. Immunohistochemical analysis reveals that the transforming growth factor‐β staining scores of the PPH3.0 sutures are not significantly different from those of the corresponding control group but are significantly different from those of the CSs and non‐heparin‐loaded‐suture groups. According to vascular endothelial growth factor (VEGF) analysis, the concentration of VEGF in the group treated with the PPH3.0 suture increases by 37.5% compared with that in its control group. No significant difference in tension strength is observed between the PPH3.0 group and healthy Achilles tendons. These findings illustrate that this novel method effectively treats Achilles tendon rupture and promotes healing and regeneration.     

Structure of the Complex between a Heparan Sulfate Octasaccharide and Mycobacterial Heparin‐Binding Hemagglutinin

Heparin‐binding hemagglutinin (HBHA) is a 199 amino acid virulence factor at the envelope of Mycobacterium tuberculosis that contributes to latent tuberculosis. The binding of HBHA to respiratory epithelial cells, which leads to extrapulmonary dissemination of the pathogen, is mediated by cell‐surface heparan sulfate (HS). We report the structural characterization of the HBHA/HS complex by NMR spectroscopy. To develop a model for the molecular recognition, the first chemically synthesized uniformly ¹³C‐ and ¹⁵N‐labeled HS octasaccharide and a uniformly ¹³C‐ and ¹⁵N‐labeled form of HBHA were prepared. Residues 180–195 at the C‐terminal region of HBHA show large chemical shift perturbation upon association with the octasaccharide. Molecular dynamics simulations conforming to the multidimensional NMR data revealed key electrostatic and even hydrophobic interactions between the binding partners that may aid in the development of agents targeting the binding event.     

Amides and Heparin‐Like Polymer Co‐Functionalized Graphene Oxide Based Core @ Polyethersulfone Based Shell Beads for Bilirubin Adsorption

Excessive bilirubin in the body of patient with liver dysfunction or metabolic obstruction may cause jaundice with irreversible brain damage, and new type of adsorbent for bilirubin is under frequent investigation. Herein, graphene oxide based core @ polyethersulfone‐based shell beads are fabricated by phase inversion method, amides and heparin‐like polymer are introduced to functionalize the core‐shell beads. The beads are successfully prepared with obvious core‐shell structure, adequate thermostability and porous shell. Clotting times and protein adsorption are investigated to inspect the hemocompatibility property of the beads. The adsorption of bilirubin is systematically investigated by evaluating the effects of contacting time, initial concentration and temperature on the adsorption, which exhibits improved bilirubin adsorption amount for the beads with amides contained cores or/and shells. It is worth believing that the amides and heparin‐like polymer co‐functionalized core‐shell beads may be utilized in the field of hemoperfusion for bilirubin adsorption.     

Effect of the Substituents of the Neighboring Ring in the Conformational Equilibrium of Iduronate in Heparin‐like Trisaccharides

Based on the structure of the regular heparin, we have prepared a smart library of heparin‐like trisaccharides by incorporating some sulfate groups in the sequence α‐D ‐GlcNS‐ (1‐4)‐α‐L ‐Ido2S‐(1‐4)‐α‐D ‐GlcN. According to the 3D structure of heparin, which features one helix turn every four residues, this fragment corresponds to the minimum binding motif. We have performed a complete NMR study and found that the trisaccharides have a similar 3D structure to regular heparin itself, but their spectral properties are such that allow to extract very detailed information about distances and coupling constants as they are isotropic molecules. The characteristic conformational equilibrium of the central iduronate ring has been analyzed combining NMR and molecular dynamics and the populations of the conformers of the central iduronate ring have been calculated. We have found that in those compounds lacking the sulfate group at position 6 of the reducing end glucosamine, the population of ²S₀ of the central iduronate residue is sensitive to the temperature decreasing to 19 % at 278 K. On the contrary, the trisaccharides with 6‐O‐sulfate in the reducing end glucosamine keep the level of population constant with temperature circa 40 % of ²S₀ similar to that observed at room temperature. Another structural feature that has been revealed through this analysis is the larger flexibility of the L ‐IdoAS‐ D ‐GlcN glycosidic linkage, compared with the D ‐GlcNS‐L ‐IdoA. We propose that this is the point where the heparin chain is bended to form structures far from the regular helix known as kink that have been proposed to play an important role in the specificity of the heparin–protein interaction.     

Polyisobutylene‐based polyurethanes. VIII. Polyurethanes with ‐O‐S‐PIB‐S‐O‐ soft segments

We describe the synthesis, characterization, and select properties of a novel polyurethane (PU) prepared using a new polyisobutylene diol, HO‐CH₂CH₂‐S‐PIB‐S‐CH₂CH₂‐OH, soft segment and conventional hard segments. The diol is synthesized by terminal functionalization of ally‐telechelic PIB followed by low‐cost thiol‐ene click chemistry. Properties of ‐S‐ containing PU (PIB_S‐PU) containing 72.5% PIB were investigated and compared to similar PUs made with HO‐PIB‐OH (PIB_O‐PU). Hydrolytic resistance was studied by contact with phosphate‐buffered saline, oxidative resistance by immersing in concentrated HNO₃, and metal ion oxidation resistance by exposure to CoCl₂/H₂O₂. Hydrolytic and oxidative resistances of PIB_S‐PU and PIB_O‐PU are similar and superior to a commercial PDMS‐based PU, Elast‐Eon? E2A. According to ¹H NMR spectroscopy the ‐S‐ in PIB_S‐PUs remained unchanged upon treatment with HNO₃, however, oxidized mainly to ‐SO₂‐ by CoCl₂/H₂O_2. Static mechanical properties of PIB_S‐PU and PIB_O‐PU are similar, except creep resistance of PIB_S‐PU is surprisingly superior. The thermal stability of PIB_S‐PUs is ～15 °C higher than that of PIB_O‐PU. FTIR spectroscopy indicates H bonded S atoms (N‐H…S) between soft and hard segments, which noticeably affect properties. DSC and XRD studies suggest random low‐periodicity crystals dispersed within a soft matrix. Energy dispersive X‐ray spectroscopy–scanning electron microscopy indicates homogeneous distribution of S atoms on PIB_S‐PU surfaces. © 2015 Wiley Periodicals, Inc. J. Polym. Sci., Part A: Polym. Chem. 2016 , 54, 1119–1131