A high molecular weight silk fibroin scaffold that resists degradation and promotes cell proliferation.

The regulation of the biodegradation rate of 3D-regenerated silk fibroin scaffolds and the avoidance of premature collapse are important concerns for their effective applications in tissue engineering. In this study, bromelain, which is specific to sericin, was used to remove sericin from silk, and high molecular weight silk fibroin was obtained after the fibroin fibers were dissolved. Afterwards, a 3D scaffold was prepared via freeze-drying. The Sodium dodecyl sulfate-polyacrylamide gel electrophoresis results showed that the average molecular weight of the regenerated silk fibroin prepared by using the bromelain-degumming method was approximately 142.2 kDa, which was significantly higher than that of the control groups prepared by using the urea- and Na2 CO3 -degumming methods. The results of enzyme degradation in vitro showed that the biodegradation rate and internal three-dimensional structure collapse of the bromelain-degumming fibroin scaffolds were significantly slower than those of the two control scaffolds. The proliferation activity of human umbilical vein vascular endothelial cells inoculated in bromelain-degumming fibroin scaffolds was significantly higher than that of the control scaffolds. This study provides a novel preparation method for 3D-regenerated silk fibroin scaffolds that can effectively resist biodegradation, continuously guide cell growth, have good biocompatibility, and have the potential to be used for the regeneration of various connective tissues.

Plasmid containing VEGF-165 and ANG-1 dual genes packaged with fibroin-modified PEI to promote the regeneration of vascular network and dermal tissue.

Reducing the cytotoxicity of cationic polymers is the major issue to their use as a gene delivery carrier. In this study, plasmids containing encoding vascular endothelial cell growth factor 165 and angiopoietin-1 were packaged with the conjugates of cationic fibroin (CSF) and polyethylenimine (PEI), instead of packaging pDNA with PEI alone, to prepare nanocomplexes (CSF+PEI)/pDNA. The complexes were loaded into a silk fibroin scaffold to enhance its function to induce microvascular network generation and dermal tissue regeneration. The results of transfecting EA.hy926 cells with the complexes in vitro showed that (CSF+PEI)/pDNA had a stronger transfection ability than PEI/pDNA. Importantly, compared with PEI as the gene carrier alone, the cell viability was significantly increased and the cytotoxicity was effectively reduced after the conjugate of CSF and PEI was used as the gene carrier. The results of angiogenesis in chick embryo chorioallantoic membranes showed that compared with scaffolds loaded with PEI/pDNA, the neovascularization ratio in scaffolds loaded with (CSF+PEI)/pDNA was significantly increased. In vivo experimental results of scaffolds implantation for full-thickness skin defects in SD rats showed that, compared with loading PEI/pDNA complex, loading (CSF+PEI)/pDNA complex in the scaffold more effectively promoted the formation of vascular network in the scaffold and accelerated the regeneration of dermal tissue. The gene delivery system established in this study has application potential not only in the regeneration of vascular-containing tissues, but also in tumor gene therapy.

Response process and adaptation mechanism of estuarine benthic microbiota to polyvinyl chloride microplastics with and without phthalates.

This study aimed to explore the response mechanisms of the microbiota in estuarine sediments amended with polyvinyl chloride (PVC) microplastics (MPs) with and without phthalates (PAEs) through a 60-day microcosm experiment. The results indicated that addition of MPs increased the porosity of the sediment. However, the sediment porosity decreased with the length of the amendment period. Following amendment with MPs containing PAEs, the sediment PAE content increased over time. The addition of MPs without PAEs increased the relative abundance of the dominant phyla of bacteria (Actinobacteria, Bacteroidetes, Chloroflexi, Firmicutes, Gemmatimonadetes, and Planctomycetes) and eukaryotes (Ascomycota, Bacillariophyta, Chordata, and Streptophyta), whereas the relative abundance decreased over time following the addition of MPs containing PAEs. The PAEs released from MPs had greater effects on these phyla than the MPs themselves. The dominant bacteria were more sensitive to MPs than the dominant eukaryotes. After a 60-day amendment with MPs containing PAEs, the bacterial and eukaryotic species numbers were lower by 5.4% and 3.4%, respectively, the relative abundance of certain genes involved in metabolism was lower, and the relative abundance of stress-related genes was higher. These findings provide insight into the microbial response and adaptation mechanisms in estuarine environments polluted with MPs.

Multifunctional ß-Cyclodextrin-Poly(ethylene glycol)-Cholesterol Nanomicelle for Anticancer Drug Delivery.

Nanoparticle drug delivery systems have drawn considerable attention worldwide due to their unique characteristics and advantages in anticancer drug delivery. Herein, the curcumin (Cur) loaded nanomicelles with two-stage drug release behavior were developed. ß-Cyclodextrin (ß-CD) and cholesterol were conjugated onto both ends of the poly(ethylene glycol) (PEG) chain to obtain an amphiphilic ß-CD-PEG-Chol. The Cur was loaded into the cavities of ß-CD and nanomicelle when the ß-CD-PEG-Chol self-assembled to the Cur@ß-CD-PEG-Chol nanomicelles (Cur@CPC NMs). These Cur@CPC NMs are spherical particles with a particle size of 120.9 nm. The Cur drug loading capacity of Cur@CPC NMs are 61.6 ± 6.9 mg/g. The release behavior of Cur from Cur@CPC NMs conformed to a two-stage mode of "burst-release followed by sustained-release". The prepared Cur@CPC NMs possess high storage stability and excellent hemocompatibility. Moreover, these Cur@CPC NMs exhibit satisfactory antioxidant activity and anticancer activity, resulting in significant reduction in intracellular H2O2-induced ROS and a nearly 50% lethality rate of HepG-2 cells. Meanwhile, the Cur@CPC NMs show good anti-inflammatory activity, by which the secretion of inflammatory factors of IL-6 and TNF-α are inhibited. Overall, the developed Cur@CPC NMs show application prospects in anticancer drug delivery systems.

Cleavable poly(ethylene glycol) branched chain-modified Antheraea pernyi silk fibroin as a gene delivery carrier.

Aim: To obtain a gene carrier that can effectively deliver loaded therapeutic genes to tumor cells, avoid toxic effects on normal cells and reduce nonspecific adsorption of plasma proteins. Methods: The conjugate of poly(ethylene glycol) (PEG) and MMP2SSP (PEG-MMP2SSP) was covalently coupled to cationized Antheraea pernyi silk fibroin (CASF) through disulfide bond exchange reaction to obtain a PEG-MMP2SSP-modified CASF (CASFMP). Results: The PEG chains were effectively cleaved from the CASFMP by MMP2. CASFMP/pDNA complexes inhibited human fibrosarcoma cell proliferation, and its cytotoxicity to human normal embryonic kidney cells was significantly lower than that of poly(ethylenimine)/pDNA after coculturing with cells for 24 h. Conclusion: CASFMP is a promising compound for use in gene therapy.

Fate and abundance of antibiotic resistance genes on microplastics in facility vegetable soil.

Microplastics (MPs) and antibiotic resistance genes (ARGs) coexist widely in farmland soils, but the fate and abundance of ARGs on MPs is rarely explored. In this study, high-throughput fluorescent quantitative polymerase chain reaction was used to determine ARGs on MPs in facility vegetable soil. The results indicated that when the particle size of the MPs was larger, the weathering was more serious, or the MPs came from soils with a long vegetable cultivation period, the levels of antibiotics and heavy metals on the MPs were higher. The distribution of the detected ARGs types on distinct MPs showed changes. Compared with weakly weathered MPs, the detected beta lactamase and aminoglycoside resistance genes on strongly weathered MPs were decreased by 2.6% and 1.7%, while the detected sul-ARGs and Macrolide-Lincosamide-Streptogramin B (MLSB) resistance genes were increased by 1.5% and 2.8%. Compared with smaller MPs, the detected MLSB and vancomycin resistance genes on larger MPs were decreased by 2.0% and 1.4%, while the detected fluoroquinolone, quinolone, florfenicol, chloramphenicol, and amphenicol (FCA) resistance genes and sul-ARGs were increased by 1.2% and 1.0%. Compared with MPs in soil after three years of vegetable cultivation, the detected FCA resistance genes and sul-ARGs on MPs in soil after ten years of vegetable cultivation were decreased by 1.3% and 1.6%, while the detected beta lactamase and aminoglycoside resistance genes were increased by 1.0% and 1.7%. This study suggests that MPs with larger size, stronger weathering or from soil after long-term vegetable cultivation adsorb more antibiotics and heavy metals and cause more mobile genetic elements, which can contribute to antibiotic resistance on the MPs.

[Effects of osmoconditioning on membrane lipid components and fatty acid content of cold-sensitive soybean seeds].

Cold-sensitive soybean (Glycisne max Zhonghuang No.22) seeds were used to investigate the effects of osmoconditioning for invigoration of seeds chilling tolerance and the changes in membrane lipid by thin layer chromatography and gas chromatography to disclose the role of membrane lipid played in imbibitional chilling resistance capacity. Results are as follows: being treated with 33% PEG6000, seeds germination index, vigor index and root dry weight presented the trend of upward as the osmocondition time prolonged. In 72 h, the percentage of unsaturated phospholipid (e.g. PC and PE) in soybean seed increased, composition of lipid fatty acid changed: the saturated fatty acid 16:0 content in PC, PE, PI significantly decreased, while unsaturated fatty acid 18:2 increased greatly, which led to increase index of unsaturation fatty acid of lipid. These changes were positively corrected with osmoconditioning time. After treated with PEG, membrane fluidity increased to provide precondition for preventing imbibitional chilling, keeping the integrity of membrane system and normal metabolism.

A pH-responsive cyclodextrin-based hybrid nanosystem as a nonviral vector for gene delivery.

The absence of safe, efficient, cost-effective, and easily scalable delivery platforms is one of the most significant hurdles and critical issues that limit the bench to bedside translation of oligonucleotides-based therapeutics. Acid-labile materials are of special interest in developing nonviral vectors due to their capability of intracellularly delivering therapeutic payload. In this study, a nanovector was designed by integrating a pH-responsive cyclodextrin material and low molecular weight polyethylenimine (PEI). Antisense oligonucleotide (ASON) Bcl-xl could be encapsulated into this hybrid nanosystem with extremely high loading efficiency by a nanoemulsion technique. The developed pH-responsive ASON nanotherapeutics could be efficiently transfected into human lung adenocarcinoma cells in a time- and dose-dependent manner, resulting in effective cell growth inhibition, significant suppression on the expression of Bcl-xl mRNA/protein, and efficient cell apoptosis. Importantly, the new nanovector showed drastically higher efficacy and lower cytotoxicity when compared with PLGA-based counterpart and commonly used cationic vectors like branched PEI (25,000 Da) and Lipofectamine 2000. This pH-responsive hybrid nanosystem may serve as a safe and efficient nonviral vector that may find wide applications in gene therapy.

Synthesis and characterization of novel blood-compatible soluble chemically cross-linked polyurethanes with excellent mechanical performance for biomedical applications.

A controlled cross-linking polymerization system was designed, and soluble chemically cross-linked polyurethane was synthesized using laurylamine, n-octylamine, n-pentylamine, and ethylenediamine chain extenders. The mechanical analysis showed that the polyurethane materials synthesized in this paper have very excellent mechanical properties with a breaking elongation of 1914% and a tensile strength of 4303 N/cm(2). Such good mechanical properties must enable it to have good longevity when used as biomaterials. The polyurethane materials with n-pentylamine and n-octylamine chain extenders show reduced platelet adhesion than that with an ethylenediamine chain extender after sustaining 200 000 times of load cycles, indicating that polyurethanes introduced with an alkyl side chain onto the hard segments keep good antithrombogenic properties after sustaining load cycles. This might be because the hard segments are shielded by the alkyl side chain when the micro-phase-separation structure is destroyed in the repeated deformation of the polyurethane materials. The present investigation reveals that the influence of introducing long alkyl side chains into the backbone of the polyurethane macromolecule has been shown to reduce platelet deposition and to enhance in vitro albumin adsorption. However, in this paper, it has been observed that the polyurethane material introduced with a proper-length alkyl side chain onto the hard segment has the best antithrombogenic properties after the fatigue test.