Effect of Extracellular Matrix Coating on Cancer Cell Membrane-Encapsulated Polyethyleneimine/DNA Complexes for Efficient and Targeted DNA Delivery In Vitro.

Polyethyleneimine (PEI) has a good spongy proton effect and is an excellent nonviral gene vector, but its high charge density leads to the instability and toxicity of PEI/DNA complexes. Cell membrane (CM) capsules provide a universal and natural solution for this problem. Here, CM-coated PEI/DNA capsules (CPDcs) were prepared through extrusion, and the extracellular matrix was coated on CPDcs (ECM-CPDcs) for improved targeting. The results showed that compared with PEI/DNA complexes, CPDcs had core-shell structures (PEI/DNA complexes were coated by a 6-10 nm layer), lower cytotoxicity, and obvious homologous targeting. The internalization and transfection efficiency of 293T-CM-coated PEI70k/DNA capsules (293T-CP70Dcs) were 91.8 and 74.5%, respectively, which were higher than those of PEI70k/DNA complexes. Then, the internalization and transfection efficiency of 293T-CP70Dcs were further improved by ECM coating, which were 94.7 and 78.9%, respectively. Then, the internalization and transfection efficiency of 293T-CP70Dcs were further improved by ECM coating, which were 94.7 and 78.9%, respectively. Moreover, the homologous targeting of various CPDcs was improved by ECM coating, and other CPDcs also showed similar effects as 293T-CP70Dcs after ECM coating. These findings suggest that tumor-targeted CPDcs may have considerable advantages in gene delivery.

Improving antibiotic removal and anaerobic digestion performance of discarded cefradine pellets through thermo-alkaline pretreatment.

Discarded cefradine pellets (DCP) as the hazardous wastes contain lots of bioavailable sucrose. Anaerobic digestion (AD) may be a promising technology for treating DCP, achieving dual goals of waste treatment and resource recovery. However, high concentration of cefradine will inhibit the AD process. This study applied thermo-alkaline pretreatment (TAP) to remove cefradine and improve the AD performance of DCP. Around 95% cefradine could be degraded to different intermediate degradation products (TPs) in TAP at optimal condition, and hydrolysis and hydrogenation were the main degradation pathways. Quantitative structure-activity relationship analysis indicated that the main TPs exhibited lower toxicity than cefradine, and DCP residues after TAP were almost not toxic to E. coli K12 and B. subtilis growth by antibacterial activity analysis. Therefore, TAP promoted the biomethane yield in AD of DCP residues (274.74 mL/g COD), which was 1.91 times that of control group. Besides, compared to control group, final cefradine concentrations in liquids and sludge were significantly decreased in AD system with TAP, lowering environmental risk and indicating stronger prospect for process application. Microbiological analysis revealed that acidogens (Macellibacteroides, Bacteroides), syntrophs (Syntrophobacter, Syntrophorhabdus), and acetoclastic Methanosaeta were enriched in AD system with TAP, which contributed to improving AD performance of DCP.

Enhanced cadmium removal by growing Bacillus cereus RC-1 immobilized on different magnetic biochars through simultaneous adsorption and bioaccumulation.

Biosorption of cadmium by growing bacteria immobilized on the three magnetic biochars derived from rice straw (MRSB-pellet), sewage sludge (MSSB-pellet), and chicken manure (MCMB-pellet) was investigated, respectively. Total biosorption capacity of the pellets was tested under varying range of pH, culture time, and initial Cd2+ concentration. The maximum biosorption capacity of 93.02 mg/g was obtained with MRSB-pellet, followed by MSSB-pellet (68.02 mg/g) and MCMB-pellet (63.95 mg/g). The biosorption by these immobilized bacterial pellets was more effective than free bacteria; this enhancement could be the result of simultaneous adsorption and bioaccumulation, mainly resulting from magnetic biochar carrier and active bacteria, respectively. The biosorption process by immobilized pellets was primarily driven by ion exchange and complexation, which jointly contributed 73.56% (MRSB-pellet) to 78.62% (MSSB-pellet) of the total adsorption, while the mechanisms of chemical precipitation and physical adsorption could averagely contribute 6.91% (MSSB-pellet) and 11.24% (MRSB-pellet), respectively. Intracellular accumulation was comparably tiny among these mechanisms accounting for 4.30-5.92% of total biosorption; in turn, it would keep intracellular Cd2+ concentration below a toxic threshold to maintain cell activity. These suggested that magnetic biochar immobilized bacteria, particularly MRSB-pellet, could be used as an effective biosorbent to remove the Cd2+ from the growth medium. This study further deepened our understanding of biosorption process by microorganism immobilized onto magnetic biochar for the metal removal.

Insights into the microbial response mechanisms to ciprofloxacin during sulfur-mediated biological wastewater treatment using a metagenomics approach.

The fate and removal of ciprofloxacin, a class of fluoroquinolone antibiotic, during sulfur-mediated biological wastewater treatment has been recently well documented. However, little is known regarding the genetic response of microorganisms to ciprofloxacin. Here, a lab-scale anaerobic sulfate-reducing bioreactor was continuously operated over a long term for ciprofloxacin-contaminated wastewater treatment to investigate the response of the microorganisms to ciprofloxacin by adopting a metagenomics approach. It was found that total organic carbon (TOC) removal and sulfate reduction were promoted by approximately 10% under ciprofloxacin stress, along with the enrichment of functional genera (e.g., Desulfobacter, Geobacter) involved in carbon and sulfur metabolism. The metagenomic analytical results demonstrated that ciprofloxacin triggered the microbial SOS response, as demonstrated by the up-regulation of the multidrug efflux pump genes (8-125-fold higher than that of the control) and ciprofloxacin-degrading genes (4-33-fold higher than that of the control). Moreover, the contents of ATP, NADH, and cytochrome C, as well as related functional genes (including genes involved in energy generation, electron transport, carbon metabolism, and sulfur metabolism) were markedly increased under ciprofloxacin stress. This demonstrated that the carbon and sulfur metabolisms were enhanced for energy (ATP) generation and electron transport in response to ciprofloxacin-induced stress. Interestingly, the microbes tended to cooperate while being subjected exposure to exogenous ciprofloxacin according to the reconstructed metabolic network using the NetSeed model. Particularly, the species with higher complementarity indices played more pivotal roles in strengthening microbial metabolism and the SOS response under long-term ciprofloxacin stress. This study characterized the response mechanisms of microorganisms to ciprofloxacin at the genetic level in sulfur-mediated biological wastewater treatment. These new understandings will contribute the scientific basis for improving and optimizing the sulfur-mediated bioprocess for antibiotics-laden wastewater treatment.

Development of a novel DNA delivery system based on rice bran polysaccharide-Fe(III) complexes.

Metal complexes, as a type of potential non-virus gene carriers, have gained much attention due to their properties of high charge density and unique three-dimensional structure. This study investigated the potential of polyethyleneimine (PEI)-modified rice bran polysaccharide-Fe(III) complex (PEI-PI) as a safe gene delivery system and explored the effect of Fe(III) on the efficiency of gene transfection mediated by PEI modified rice bran polysaccharide (PEI-P) and PEI-PI. Gel retardation assay was used to study the DNA binding and protection capability, MTT assay was performed to evaluate the biocompatibility, and PEI-PI complex-mediated EGFP gene transfection was studied in vitro. Results showed the PEI-PI could induce DNA condensation and protect DNA from degradation by DNase I at a low weight ratio (vector/DNA) of 2. At the same weight ratio, PEI exhibited the strongest DNA binding capability but PEI-PI exhibited the highest gene transfection efficiency among all carrier systems. Compared with the PEI-P + Fe(III) system, PEI-PI not only had a more significant capability to condense DNA but also presented higher gene transfection efficiency. Moreover, PEI-PI exhibited no obvious cytotoxicity to cells. This work provides a strategy for the design and development of gene vectors based on PI complexes.

TAT-functionalized PEI-grafting rice bran polysaccharides for safe and efficient gene delivery.

Polysaccharides are considered to be promising candidates for non-viral gene delivery because of their molecular diversity, which can be modified to fine-tune their physicochemical properties. In this work, transcriptional activator protein (TAT) functionalized PEI grafted polysaccharide polymer (PRBP) was prepared by using rice bran polysaccharide as the starting material, and characterized by various methods. The potential of TAT functionalized PRBP (PRBP-TAT) as gene vector was studied in vitro, including DNA loading capacity, DNA protection ability and biocompatibility. The cell uptake and transfection efficiency of the PRBP-TAT/pDNA polyplexes were studied. The results showed that PRBP-TAT could completely condense DNA at N/P 2. The PRBP-TAT/pDNA polyplexes could protect DNA from degrading by DNase and were efficiently internalized by cells. Biocompatibility result showed that PRBP-TAT had no significant cytotoxicity and effect on cell proliferation. At low N/P ratios of 1-3.5, PRBP-TAT showed higher transfection efficiency than PEI30k and PEI30k-grafted rice bran polysaccharide. PRBP-TAT and PEI showed the highest transfection efficiency of 42.8% and 28.1% when pDNA is 2 µg and N/P ratio is 1.5, respectively, while PRBP showed the highest transfection efficiency of 37.3% at N/P 2.5. These results indicate that PTA is a promising candidate vector for safe and efficient gene delivery.

Rice bran polysaccharide-metal complexes showed safe antioxidant activity in vitro.

The effect on the intracellular reactive oxygen species (ROS) generation, and the antioxidant and cytotoxicity properties of rice bran polysaccharides (RBP) and RBP-metal complexes RBP-Fe(III), RBP-Cu, RBP-Zn and RBP-Ca, were evaluated using atomic absorption spectroscopy (AAS), scavenging activity assays, cell viability assay and fluorescence microscopy. The RBP-metal complexes were prepared using the hydrothermal method. The RBP-Fe(III) complexes were found to be potent scavengers for superoxide (O2-) free radicals. The RBP alone and RBP-Ca complex showed high scavenging activity for 2,2-diphenyl-1-picrylhydrazyl (DPPH) free radicals. In addition, the RBP-Fe(III) complex also showed good biocompatibility and lowered the intracellular ROS levels, while RBP alone, RBP-Zn and RBP-Ca complexes were observed to increase the intracellular ROS level. Our findings suggest that among the tested RBP-metal complexes, RBP-Fe(III) complex is a strong candidate as an antioxidant therapeutic.

Safe and efficient gene delivery based on rice bran polysaccharide.

Polysaccharides are capable of being modified by polycations to adjust their physical and chemical properties, which accordingly are considered as potential candidate materials for safe and efficient gene delivery. Here, we extracted and purified polysaccharides from rice bran, and their physicochemical properties were determined by various methods. Polyethyleneimine (PEI) modified rice bran polysaccharide (PRBP) was prepared by grafting RBP with low molecular weight PEI and the preparation was determined by FTIR. The potential of PRBP as a gene vector was systematically evaluated in vitro. The results show that PRBP can compact DNA and form PRBP/DNA polylexes with a particle size of 50-100 nm. The PRBP/DNA polylexes can protect DNA degradation from DNase I efficiently. Compared with PEI, higher transfection efficiency was achieved by the PRBP. At weight ratio of 3, the highest efficiency of gene transfection mediated by PRBP-2000 was obtained, which was 37.5% and significantly higher than PEI and commercial reagents (calcium phosphate cell transfection kit) and was closed to lipo6000. Furthermore, according to MTT results, the cytotoxicity of PRBP is much lower than that of PEI, especially for PEI2000. We hope these results will provide new strategy for rice bran polysaccharides development and application as biomaterials.

Development of a safe and efficient gene delivery system based on a biodegradable tannic acid backbone.

Finding a safe and efficient gene delivery vector is a major international challenge facing the development of gene therapy. Tannic acid (TA) is a natural cross-linker owing to its hydroxyl and carboxyl groups that can interact with biopolymers for different biomaterial design. In this work, three polyethyleneimine-modified TA polymers were prepared, and the polymers were characterized by FTIR, UV-vis, elemental analysis and 1H NMR. The potential of PTAs as gene vector was studied in vitro, including DNA loading capacity, DNA protection ability and biocompatibility. In addition, the particle size, zeta potential, DNA encapsulation efficiency, cell uptake and transfection efficiency of the PTA-pDNA polyplexes were also studied. The results showed that PTA2k and PTA30k could completely condense DNA at N/P of 2, and PTA600 could only completely condense DNA at N/P of 50. The PTA/pDNA polyplexes could protect DNA from degrading by DNA enzymes and could be efficiently uptaked by cells. Biocompatibility assay showed that PTA had no significant cytotoxicity and effect on cell proliferation compared to PEI. At low N/P ratios of 1-4, PTA showed higher transfection efficiency than PEI, and the transfection efficiency increased with the increase of PEI molecular weight in PTA. At N/P of 3, PTA30k showed the highest transfection efficiency of 23.8%, while PEI30k showed only 6.7%. These results indicate that PTA is a promising candidate vector for safe and efficient gene delivery.