Heparinized self-healing polymer coating with inflammation modulation for blood-contacting biomedical devices.

The current techniques for antithrombotic coating on blood-contacting biomedical materials and devices are usually complex and lack practical feasibility with weak coating stability and low heparin immobilization. Here, a heparinized self-healing polymer coating with inflammation modulation is introduced through thermal-initiated radical copolymerization of methacrylate esterified heparin (MA-heparin) with methyl methacrylate (MMA) and n-butyl acrylate (nBA), followed by the anchoring of reactive oxygen species (ROS)-responsive polyoxalate containing vanillyl alcohol (PVAX) onto the coating through esterification. The aspirin, which is readily dissolved in the solution of MMA and nBA, is encapsulated within the coating after copolymerization. The copolymerization of MA-heparin with MMA and nBA significantly increases the heparin content of the coating, effectively inhibiting thrombosis and rendering the coating self-healing to help maintain long-term stability. ROS-responsive PVAX and aspirin released in a temperature-dependent manner resist acute and chronic inflammation, respectively. The heparinized self-healing and inflammation-modulated polymer coating exhibits the ability to confer long-term stability and hemocompatibility to blood-contacting biomedical materials and devices. STATEMENT OF SIGNIFICANCE: Surface engineering for blood-contacting biomedical devices paves a successful way to reduce thrombotic and inflammatory complications. However, lack of effectiveness, long-term stability and practical feasibility hinders the development and clinical application of existing strategies. Here we design a heparinized self-healing and inflammation-modulated polymer coating, which possesses high heparin level and self-healing capability to maintain long-term stability. The polymer coating is practically feasible to varied substrates and demonstrated to manipulate inflammation and prevent thrombosis both in vitro and in vivo. Our work provides a new method to develop coatings for blood-contacting biomedical materials and devices with long-term stability and hemocompatibility.

An endothelium membrane mimetic antithrombotic coating enables safer and longer extracorporeal membrane oxygenation application.

Thrombosis and plasma leakage are two of the most frequent dysfunctions of polypropylene (PP) hollow fiber membrane (PPM) used in extracorporeal membrane oxygenation (ECMO) therapy. In this study, a superhydrophilic endothelial membrane mimetic coating (SEMMC) was constructed on polydopamine-polyethyleneimine pre-coated surfaces of the PPM oxygenator and its ECMO circuit to explore safer and more sustainable ECMO strategy. The SEMMC is fabricated by multi-point anchoring of a phosphorylcholine and carboxyl side chained copolymer (PMPCC) and grafting of heparin (Hep) to form PMPCC-Hep interface, which endows the membrane superior hemocompatibility and anticoagulation performances. Furthermore, the modified PPM reduces protein adsorption amount to less than 30 ng/cm2. More significantly, the PMPCC-Hep coated ECMO system extends the anti-leakage and non-clotting oxygenation period to more than 15 h in anticoagulant-free animal extracorporeal circulation, much better than the bare and conventional Hep coated ECMO systems with severe clots and plasma leakage in 4 h and 8 h, respectively. This SEMMC strategy of grafting bioactive heparin onto bioinert zwitterionic copolymer interface has great potential in developing safer and longer anticoagulant-free ECMO systems. STATEMENT OF SIGNIFICANCE: A superhydrophilic endothelial membrane mimetic coating was constructed on surfaces of polypropylene hollow fiber membrane (PPM) oxygenator and its ECMO circuit by multi-point anchoring of a phosphorylcholine and carboxyl side chain copolymer (PMPCC) and grafting of heparin (Hep). The strong antifouling nature of the PMPCC-Hep coating resists the adsorption of plasma bio-molecules, resulting in enhanced hemocompatibility and anti-leakage ability. The grafted heparin on the zwitterionic PMPCC interface exhibits superior anticoagulation property. More significantly, the PMPCC-Hep coating achieves an extracorporeal circulation in a pig model for at least 15 h without any systemic anticoagulant. This endothelial membrane mimetic anticoagulation strategy shows great potential for the development of safer and longer anticoagulant-free ECMO systems.

Drug delivery under cover of erythrocytes extends drug half-life: A thrombolytic targeting therapy utilizing microenvironment-responsive artificial polysaccharide microvesicles.

The development of thrombolytic drug carriers capable of thrombus-targeting, prolonged circulation time, intelligent responsive release, and the ability to inhibit thrombotic recurrences remains a promising but significant challenge. To tackle this, an artificial polysaccharide microvesicle drug delivery system (uPA-CS/HS@RGD-ODE) was constructed. It is composed of cationic chitosan and anionic heparin assembled in a layer by layer structure, followed by surface modification using RGD peptide and 2-(N-oxide-N,N-diethylamino) ethylmethacrylate (ODE) before encapsulation of urokinase-type plasminogen activator (uPA). The effect of chitosan on the basic performances of uPA-CS/HS@RGD-ODE was estimated. The in vitro results suggest the uPA carrier, CS/HS@RGD-ODE, displayed outstanding targeting specific to activated platelets (61 %) and microenvironment-responsiveness at pH 6.5, facilitating thrombus-targeting and a controlled drug release, respectively. Most importantly, in vivo experiment suggests ODE from uPA-CS/HS@RGD-ODE substantially extends the half-life of uPA (120 min), as uPA-CS/HS@RGD-ODE can adhere onto erythrocytes and deliver uPA under cover of erythrocytes enabling a prolonged circulation time in the bloodstream. Further tail vein and abdominal aorta thrombosis models confirmed uPA-CS/HS@RGD-ODE exhibited superior targeting and thrombolysis capabilities compared to systemic administration of free uPA. To the knowledge of authors, this may be the first study to develop new drug carriers for delivery of thrombolytic drugs under the cover of erythrocytes for extended drug half-lives.

Under Heparin-Free Conditions Unsaturated Phospholipids Inhibit the Aggregation of 1N4R and 2N4R Tau.

A progressive aggregation of Tau proteins in the brain is linked to both Alzheimer's disease (AD) and various Tauopathies. This pathological process can be enhanced by several substances, including heparin. However, very little if anything is known about molecules that can inhibit the aggregation of Tau isoforms. In this study, we examined the effect of phosphatidylserines (PSs) with various lengths and saturations of fatty acids (FAs) on the aggregation properties of Tau isoforms with one (1N4R) and two (2N4R) N-terminal inserts that enhance binding of Tau to tubulin. We found that PS with unsaturated and short-length FAs inhibited Tau aggregation and drastically lowered the toxicity of Tau oligomers that were formed in the presence of such phospholipids. Such an effect was not observed for PS with fully saturated long-chain FAs. These results suggest that a short-chain irreversible disbalance between saturated and unsaturated lipids in the brain could be the trigger of Tau aggregation.

Butyrate Increases Heparin Synthesis and Storage in Human Mast Cells.

Sulphated glycosaminoglycans (GAGs) such as heparin are a major component of mast cell granules and form the matrix within which biogenic mediators are stored. Since GAGs released from mast cells also play an important role in helminth expulsion, understanding GAG storage can offer new insights into mast cell function. Sodium butyrate (NaBu), a short-chain fatty acid, causes ultrastructural changes within the granules of human mast cells (HMC-1) and increases their histamine content. Therefore, we hypothesized that NaBu treatment would also modify the storage of polysaccharides such as GAGs. NaBu (1 mM) significantly increased GAG content and granularity in a time- and concentration-dependent manner without affecting cell viability and metabolic activity. NaBu increased the expression of enzymes associated with heparin biosynthesis (GLCE, NDST1, NDST2, HS6ST1, and GALT1) in a time-dependent manner. A cholesteryl butyrate emulsion (CholButE) increased heparin content after 24 and 48 h and modestly altered the expression of genes involved in heparin biosynthesis. Similar to NaBu, CholButE reduced cell proliferation without significantly altering viability or metabolic activity. These data show that butyrate increases the synthesis and storage of heparin in human mast cells, perhaps by altering their metabolic pathways.

Interleukin-4-Loaded Heparin Hydrogel Regulates Macrophage Polarization to Promote Osteogenic Differentiation.

In bone tissue engineering, biological scaffolds are designed with structural and functional properties that closely resemble the extracellular environment, aiming to establish a microenvironment conducive to osteogenesis. Macrophages hold significant potential for promoting osteogenesis and modulating the biological behavior of tumor cells. Multiple coculture experiments of macrophages and osteoblasts have demonstrated that macrophage polarization significantly impacts osteogenesis. Therefore, exploring bone biomaterials that can modulate macrophage polarization holds great clinical significance. In this study, heparin was modified with maleimide and was used as a raw material to form a hydrogel with 4-am-PEG-SH. The compound was used to polarize macrophages and promote osteogenesis after combining with interleukin 4 (IL-4) by taking advantage of the electronegativity of heparin. The results revealed overexpressed M2 macrophage-related phenotypic genes and cocultivation with MC3T3-E1 cells demonstrated the osteogenesis-promoting effect of the loaded IL-4 heparin hydrogel. Previous research reported that hydrogel loaded with IL-4 can be used as a biomaterial for osteogenesis promotion. Heparin materials used in this paper are derived from clinically anticoagulant drugs and feature a simple operation. The synthesized hydrogel effectively binds cytokines, regulates macrophages to induce osteogenesis and has many potential clinical applications.

Hydrogel coating containing heparin and cyclodextrin/paclitaxel inclusion complex for retrievable vena cava filter towards high biocompatibility and easy removal.

Aiming to improve the retrieval rate of retrievable vena cava filters (RVCF) and extend its dwelling time in vivo, a novel hydrogel coating loaded with 10 mg/mL heparin and 30 mg/mL cyclodextrin/paclitaxel (PTX) inclusion complex (IC) was prepared. The drug-release behavior in the phosphate buffer solution demonstrated both heparin and PTX could be sustainably released over approximately two weeks. Furthermore, it was shown that the hydrogel-coated RVCF (HRVCF) with 10 mg/mL heparin and 30 mg/mL PTX IC effectively extended the blood clotting time to above the detection limit and inhibited EA.hy926 and CCC-SMC-1 cells' proliferation in vitro compared to the commercially available bare RVCF. Both the HRVCF and the bare RVCF were implanted into the vena cava of sheep and retrieved at at 2nd and 4th week after implantation, revealing that the HRVCF had a significantly higher retrieval rate of 67 % than the bare RVCF (0 %) at 4th week. Comprehensive analyses, including histological, immunohistological, and immunofluorescent assessments of the explanted veins demonstrated the HRVCF exhibited anti-hyperplasia and anticoagulation properties in vivo, attributable to the hydrogel coating, thereby improving the retrieval rate in sheep. Consequently, the as-prepared HRVCF shows promising potential for clinical application to enhance the retrieval rates of RVCFs.

Oleanolic acid derivative self-assembled aggregates based on heparin and chitosan for breast cancer therapy.

Oleanolic acid is an active ingredient from natural products with anti-breast cancer activity. However, the poor solubility in water and low bioavailability have limited its effectiveness in clinic. To improve the anticancer activity of oleanolic acid, we synthesized a novel oleanolic quaternary ammonium (QDT), which, driven by electrostatic interactions, was introduced into heparin and coated with chitosan to obtain a QDT/heparin/chitosan nanoaggregate (QDT/HEP/CS NAs). QDT/HEP/CS NAs showed the negative zeta potential (-35.01 ± 4.38 mV), suitable mean particle size (150.45 ± 0.68 nm) with strip shape, and high drug loading (36 %). The coated chitosan had strong anti-leakage characteristics toward QDT under physiological conditions. More importantly, upon sustained release in tumor cells, QDT could significantly decrease the mitochondrial membrane potential and induce apoptosis of breast cancer cells. Further in vivo antitumor study on 4 T1 tumor-bearing mice confirmed the enhanced anticancer efficacy of QDT/HEP/CS NAs via upregulation of caspase-3, caspase-9 and cytochrome C, which was attributed to the high accumulation in tumor via the enhanced permeability and retention effect. Moreover, QDT/HEP/CS NAs significantly enhanced the biosafety and biocompatibility of QDT in vitro and in vivo. Collectively, the development of QDT/HEP/CS NAs with high antitumor activity, favorable biodistribution and good biocompatibility provided a safe, facile and promising strategy to improve the anti-cancer effect of traditional Chinese medicine ingredients.

Heparan sulfate-dependent phase separation of CCL5 and its chemotactic activity.

Secreted chemokines form concentration gradients in target tissues to control migratory directions and patterns of immune cells in response to inflammatory stimulation; however, how the gradients are formed is much debated. Heparan sulfate (HS) binds to chemokines and modulates their activities. In this study, we investigated the roles of HS in the gradient formation and chemoattractant activity of CCL5 that is known to bind to HS. CCL5 and heparin underwent liquid-liquid phase separation and formed gradient, which was confirmed using CCL5 immobilized on heparin-beads. The biological implication of HS in CCL5 gradient formation was established in CHO-K1 (wild-type) and CHO-677 (lacking HS) cells by Transwell assay. The effect of HS on CCL5 chemoattractant activity was further proved by Transwell assay of human peripheral blood cells. Finally, peritoneal injection of the chemokines into mice showed reduced recruitment of inflammatory cells either by mutant CCL5 (lacking heparin-binding sequence) or by addition of heparin to wild-type CCL5. Our experimental data propose that co-phase separation of CCL5 with HS establishes a specific chemokine concentration gradient to trigger directional cell migration. The results warrant further investigation on other heparin-binding chemokines and allows for a more elaborate insight into disease process and new treatment strategies.

The Effects of Different Blood Sample and Anticoagulant Types on Quantitative Tests of B-Type Natriuretic Peptide.

BACKGROUND: BNP is a sensitive and widely used biomarker for an early diagnosis of heart failure. Currently, most commercial BNP detection products use EDTA plasma samples. The aim of this study was to evaluate the clinical performance of the BNP test by using whole blood samples compared to plasma samples, and to evaluate the effect of the anticoagulant type on the BNP test result. METHODS: In total, 106 patients with different BNP levels from the Dahua Hospital volunteered for this study. Clinically homogenous samples, including EDTA anticoagulant plasma, EDTA whole blood, and heparin anticoagulant plasma, were collected and analyzed by using i-Reader S automatic immuno-analyzer and its supporting reagent kits. Pearson's correlation and weighted least squares linear regression analysis, Bland-Altman plotting, and Kappa test were used for statistical analysis. RESULTS: Correlation analysis showed that BNP concentrations, measured from EDTA anticoagulated plasma samples, had a good linear regression relationship with BNP from whole blood samples, with a slope of 0.9477, r = 0.9978, p < 0.05. A similar correlation was observed between EDTA anticoagulated plasma samples and heparin anticoagulant plasma, with a slope of 0.8413, r = 0.9793, p < 0.05. The BNP concentration measured from the heparin plasma samples were lower than of the EDTA plasma samples. Bland-Altman analysis for assessing BNP concentration agreement showed there was no outlier ratio between EDTA whole blood and EDTA plasma within the range of the detection system, as well as no outlier between EDTA anticoagulated and heparin anticoagulant plasma. Kappa coefficient of BNP concentration between homologous EDTA anticoagulated and heparin anticoagulant plasma was 0.8553 (p < 0.001), and for EDTA anticoagulated plasma and homologous whole blood it was 0.8941 (p < 0.001). CONCLUSIONS: The diagnostic performance of EDTA anticoagulated whole blood samples did not differ significantly from EDTA anticoagulated plasma samples for the BNP test. This study showed no big significant difference between EDTA anticoagulated and heparin anticoagulated plasma measurements within 2 hours. The type of anticoagulant should be carefully chosen when performing the BNP test if BNP samples were in vitro for a long time.

PON1 enzyme activity assays for serum and heparinized plasma in horses and stability evaluation of the enzyme activity over different freeze-thaw cycles and mimic transportation.

Consistent information and standardization procedures regarding the time of storage for frozen samples and the effects of storage time on enzyme activity are still missing in the literature. Thus, we evaluated the effects of different storage temperatures (-20 °C and - 80 °C), three repetitive freeze/thaw cycles, and 24-h mimic transportation on the activities of PON1 (paraoxonase and arylesterase), enzymes involved in the protection and detoxification processes of reactive molecules. PON1 enzymes' activity was validated on serum and heparinized plasma in horses. The results revealed that conditions and time of storage of blood samples for PON1 analyses altered the activities of both enzymes in both sample types, evidencing that these conditions can lead to protein degradation or general alteration. Specifically, paraoxonase and arylesterase activities significantly decreased among storage temperatures, with major effects detected at -20 °C. The repeated freeze/thaw cycles at -20 °C and 24-h mimic transport conditions also generated an expected degradation of the arylesterase in both serum and heparinized plasma while freeze/thaw cycles at -80 °C caused an increase of both arylesterase and paraoxonase activities on both sample types. In general, similar enzyme responses were detected between serum and heparinized plasma.

Production of bovine embryos by piezo-ICSI using capacitated spermatozoa selected by fluorescence-activated cell sorting (FACS-piezo-ICSI).

Intracytoplasmic sperm injection (ICSI) remains inefficient in cattle. One reason could lie in the injection of oocytes with sperm that have not undergone molecular changes associated with in vivo capacitation and fertilizing ability. This study aimed to enhance the efficiency of bovine intracytoplasmic sperm injection (piezo-ICSI) by employing fluorescent-activated cell sorting (FACS) to select the sperm population before injection based on capacitation markers. First, we evaluated the effects of incubating thawed sperm for 2 hours with different capacitating inductors: heparin, methyl-beta-cyclodextrin (MßCD), and dibutyryl cyclic AMP (dbcAMP), alone or in combinations in a basal capacitating (C) medium (Sp-TALP). Sperm capacitation and quality markers were evaluated by flow cytometry, revealing heparin as the most effective inducer of sperm capacitation changes. It, therefore, this treatment was chosen as the sperm pretreatment for FACS-piezo-ICSI. Two cell populations showing high capacitating levels (Heparin-HCL) and low capacitating levels (Heparin-LCL) of the markers associated with sperm capacitation i(Ca2+) levels and acrosome integrity were selected by FACS and used for sperm injection. Pronuclear formation was significantly higher when ICSI was performed with Heparin-HCL sperm than with Heparin-LCL and the control group (Heparin unsorted) groups (50 %, 10 %, and 20 %, respectively). Furthermore, injecting Heparin-HCL sperm resulted in a higher blastocyst rate (22.5 %) than Heparin-LCL (10 %) and the control group (15.2 %). In conclusion, heparin treatment effectively induced changes associated with sperm capacitation. The combination of Heparin-HCL treatment and FACS enabled precise selection of capacitated sperm before ICSI, enhancing the efficiency of this technology in the bovine species.

Photoresponsive heparin ionic complexes toward controllable therapeutic efficacy of anticoagulation.

Controllable heparin-release is of great importance and necessity for the precise anticoagulant regulation. Efforts have been made on designing heparin-releasing systems, while, it remains a great challenge for gaining the external-stimuli responsive heparin-release in either intravenous or catheter delivery. In this study, an azobenzene-containing ammonium surfactant is designed and synthesized for the fabrication of photoresponsive heparin ionic complexes through the electrostatic complexation with heparin. Under the assistance of photoinduced trans-cis isomerization of azobenzene, the obtained heparin materials perform reversible athermal phase transition between ordered crystalline and isotropic liquid state at room temperature. Compared to the ordered state, the formation of isotropic state can effectively improve the dissolving of heparin from ionic materials in aqueous condition, which realizes the photo-modulation on the concentration of free heparin molecules. With good biocompatibility, such a heparin-releasing system addresses photoresponsive anticoagulation in both in vitro and in vivo biological studies, confirming its great potential clinical values. This work provides a new designing strategy for gaining anticoagulant regulation by light, also opening new opportunities for the development of photoresponsive drugs and biomedical materials based on biomolecules.

Heparin as a Potential Therapeutic Substance for Post-Traumatic Stress Disorder.

In sexually mature male Wistar rats with modeled post-traumatic stress disorder, personalized characteristics of neurobiological reactions in the population of predator-induced stress-resilient and stress-susceptible heparinized animals were determined. Characteristics of the systemic response of immune mechanisms, hypothalamic-pituitary-adrenal axis, behavioral manifestations, as well as basic properties of the CNS (excitation/inhibition) are presented. The study demonstrated encouraging positive results of the course administration of unfractionated heparin at a dose below the therapeutic and prophylactic doses. The inclusion of heparin drugs into the clinical practice for the treatment of post-traumatic stress disorder will not require large-scale clinical trials, because many effects of heparin as a nonspecific adaptogen are well studied. Moreover, these properties were confirmed at a higher technological level during the COVID-19 pandemic.

The impact of heparin and direct thrombin inhibitors on cell-penetrating polymer siRNA transfection.

Gene therapy using siRNA has become a promising strategy to achieve targeted gene knockdown for treatment of cardiovascular pathologies. However, efficient siRNA transfection often relies on cationic delivery vectors such as synthetic cell-penetrating polymers which are susceptible to interference by negatively charged molecules. Anticoagulants such as heparin, which is negatively charged and widely used in cardiovascular applications, may pose a significant barrier to effective siRNA delivery. We therefore conducted in vitro studies utilizing human smooth muscle and endothelial cells transfected with glyceraldehyde-3-phosphate dehydrogenase (GAPDH) and ß2-microglobulin (B2M) siRNA in the presence of heparin, argatroban, and bivalirudin in order to determine which anticoagulant therapy is most compatible for siRNA delivery. We observed that while heparin, at clinical doses, decreases the efficiency of siRNA targeted mRNA knockdown, mRNA knockdown is not inhibited in the presence of either argatroban or bivalirudin. Our data suggests that heparin should be avoided during siRNA therapy with cationic transfection agents, and argatroban and bivalirudin should be used in its stead.

Extracellular Matrix Mimicking Wound Microenvironment Responsive Amyloid-Heparin@TAAgNP Co-Assembled Hydrogel: An Effective Conductive Antibacterial Wound Healing Material.

Bioengineered composite hydrogel platforms made of a supramolecular coassembly have recently garnered significant attention as promising biomaterial-based healthcare therapeutics. The mechanical durability of amyloids, in conjunction with the structured charged framework rendered by biologically abundant key ECM component glycosaminoglycan, enables us to design minimalistic customized biomaterial suited for stimuli responsive therapy. In this study, by harnessing the heparin sulfate-binding aptitude of amyloid fibrils, we have constructed a pH-responsive extracellular matrix (ECM) mimicking hydrogel matrix. This effective biocompatible platform comprising heparin sulfate-amyloid coassembled hydrogel embedded with polyphenol functionalized silver nanoparticles not only provide a native skin ECM-like conductive environment but also provide wound-microenvironment responsive on-demand superior antibacterial efficacy for effective diabetic wound healing. Interestingly, both the cytocompatibility and antibacterial properties of this bioinspired matrix can be fine-tuned by controlling the mutual ratio of heparin sulfate-amyloid and incubated silver nanoparticle components, respectively. The designed biomaterial platform exhibits notable effectiveness in the treatment of chronic hyperglycemic wounds infected with multidrug-resistant bacteria, because of the integration of pH-responsive release characteristics of the incubated functionalized AgNP and the antibacterial amyloid fibrils. In addition to this, the aforementioned assemblage shows exceptional hemocompatibility with significant antibiofilm and antioxidant characteristics. Histological evidence of the incised skin tissue sections indicates that the fabricated composite hydrogel is also effective in controlling pro-inflammatory cytokines such as IL6 and TNFα expressions at the wound vicinity with significant upregulation of angiogenesis markers like CD31 and α-SMA.

Heparin-Functionalized Bioactive Glass to Harvest Endogenous Growth Factors for Pulp Regeneration.

Pulp and periapical diseases can lead to the cessation of tooth development, resulting in compromised tooth structure and functions. Despite numerous efforts to induce pulp regeneration, effective strategies are still lacking. Growth factors (GFs) hold considerable promise in pulp regeneration due to their diverse cellular regulatory properties. However, the limited half-lives and susceptibility to degradation of exogenous GFs necessitate the administration of supra-physiological doses, leading to undesirable side effects. In this research, a heparin-functionalized bioactive glass (CaO-P2O5-SiO2-Heparin, abbreviated as PSC-Heparin) with strong bioactivity and a stable neutral pH is developed as a promising candidate to addressing challenges in pulp regeneration. Fourier transform infrared spectroscopy, X-ray photoelectron spectroscopy, and thermogravimetric analysis reveal the successful synthesis of PSC-Heparin. Scanning electron microscopy and X-ray diffraction show the hydroxyapatite formation can be observed on the surface of PSC-Heparin after soaking in simulated body fluid for 12 h. PSC-Heparin is capable of harvesting various endogenous GFs and sustainably releasing them over an extended duration by the enzyme-linked immunosorbent assay. Cytological experiments show that developed PSC-Heparin can facilitate the adhesion, migration, proliferation, and odontogenic differentiation of stem cells from apical papillae. Notably, the histological analysis of subcutaneous implantation in nude mice demonstrates PSC-Heparin is capable of promoting the odontoblast-like layers and pulp-dentin complex formation without the addition of exogenous GFs, which is vital for clinical applications. This work highlights an effective strategy of harvesting endogenous GFs and avoiding the involvement of exogenous GFs to achieve pulp-dentin complex regeneration, which may open a new horizon for regenerative endodontic therapy.

In vitro degradation, swelling, and bioactivity performances of in situ forming injectable chitosan-matrixed hydrogels for bone regeneration and drug delivery.

Injectable, tissue mimetic, bioactive, and biodegradable hydrogels offer less invasive regeneration and repair of tissues. The monitoring swelling and in vitro degradation capacities of hydrogels are highly important for drug delivery and tissue regeneration processes. Bioactivity of bone tissue engineered constructs in terms of mineralized apatite formation capacity is also pivotal. We have previously reported in situ forming chitosan-based injectable hydrogels integrated with hydroxyapatite and heparin for bone regeneration, promoting angiogenesis. These hydrogels were functionalized by glycerol and pH to improve their mechano-structural properties. In the present study, functionalized hybrid hydrogels were investigated for their swelling, in vitro degradation, and bioactivity performances. Hydrogels have degraded gradually in phosphate-buffered saline (PBS) with and without lysozyme enzyme. The percentage weight loss of hydrogels and their morphological and chemical properties, and pH of media were analyzed. The swelling ratio of hydrogels (55%-68%(wt), 6 h of equilibrium) indicated a high degree of cross-linking, can be suitable for controlled drug release. Hydrogels have gradually degraded reaching to 60%-70% (wt%) in 42 days in the presence and absence of lysozyme, respectively. Simulated body fluid (SBF)-treated hydrogels containing hydroxyapatite-induced needle-like carbonated-apatite mineralization was further enhanced by heparin content significantly.

The mutation of Japanese encephalitis virus envelope protein residue 389 attenuates viral neuroinvasiveness.

The envelope (E) protein of the Japanese encephalitis virus (JEV) is a key protein for virus infection and adsorption of host cells, which determines the virulence of the virus and regulates the intensity of inflammatory response. The mutation of multiple aa residues in the E protein plays a critical role in the attenuated strain of JEV. This study demonstrated that the Asp to Gly, Ser, and His mutation of the E389 site, respectively, the replication ability of the viruses in cells was significantly reduced, and the viral neuroinvasiveness was attenuated to different degrees. Among them, the mutation at E389 site enhanced the E protein flexibility contributed to the attenuation of neuroinvasiveness. In contrast, less flexibility of E protein enhanced the neuroinvasiveness of the strain. Our results indicate that the mechanism of attenuation of E389 aa mutation attenuates neuroinvasiveness is related to increased flexibility of the E protein. In addition, the increased flexibility of E protein enhanced the viral sensitivity to heparin inhibition in vitro, which may lead to a decrease in the viral load entering brain. These results suggest that E389 residue is a potential site affecting JEV virulence, and the flexibility of the E protein of aa at this site plays an important role in the determination of neuroinvasiveness.

An evaluation of a Stenotrophomonas maltophilia outbreak due to commercial arterial blood gas collection kit.

BACKGROUND: Stenotrophomonas maltophilia is a gram-negative bacterium that can cause hospital infections and outbreaks within hospitals. This study aimed to evaluate an outbreak of Stenotrophomonas maltophilia, caused by ready-to-use commercial syringes containing liquid lithium and heparin for arterial blood gas collection in a university hospital. METHODS: Upon detecting an increase in Stenotrophomonas maltophilia growth in blood cultures between 15.09.2021 and 19.11.2021, an outbreak analysis and a case-control study (52 patients for the case group, 56 patients for the control group) were performed considering risk factors for bacteremia. Samples from possible foci for bacteremia were also cultured. Growing bacteria were identified by matrix-assisted laser desorption ionization time-of-flight mass spectrometry. The genetic linkage and clonal relationship isolates were investigated with pulsed-field gel electrophoresis (PFGE) in the reference laboratory. RESULTS: In the case-control study, the odds ratio for the central venous catheter [3.38 (95% confidence interval [CI]: 1.444, 8.705 ; p = 0.006)], for surgery [3.387 (95% confidence interval [CI]: 1.370, 8.373 ; p = 0.008)] and for arterial blood gas collection history [18.584 (95% confidence interval [CI]:4.086, 84.197; p < 0.001)] were identified as significant risk factors. Stenotrophomonas maltophilia growth was found in ready-to-use commercial syringes used for arterial blood gas collection. Molecular analysis showed that the growths in the samples taken from commercial syringes and the growths from blood cultures were the same. It was decided that the epidemic occurred because the method for sterilization of heparinized liquid preparations were not suitable. After discontinuing the use of the kits with this lot number, the outbreak was brought under control. CONCLUSIONS: According to our results, disposable or sterile medical equipment should be included as a risk factor in outbreak analyses. The method by which injectors containing liquids, such as heparin, are sterilized should be reviewed. Our study also revealed the importance of the cooperation of the infection control team with the microbiology laboratory.