Development and application of haploid embryonic stem cells.

Haploid cells are a kind of cells with only one set of chromosomes. Compared with traditional diploid cells, haploid cells have unique advantages in gene screening and drug-targeted therapy, due to their phenotype being equal to the genotype. Embryonic stem cells are a kind of cells with strong differentiation potential that can differentiate into various types of cells under specific conditions in vitro. Therefore, haploid embryonic stem cells have the characteristics of both haploid cells and embryonic stem cells, which makes them have significant advantages in many aspects, such as reproductive developmental mechanism research, genetic screening, and drug-targeted therapy. Consequently, establishing haploid embryonic stem cell lines is of great significance. This paper reviews the progress of haploid embryonic stem cell research and briefly discusses the applications of haploid embryonic stem cells.

Generation of a human haploid neural stem cell line for genome-wide genetic screening.

BACKGROUND: Haploid embryonic stem cells (haESCs) have been established in many species. Differentiated haploid cell line types in mammals are lacking due to spontaneous diploidization during differentiation that compromises lineage-specific screens. AIM: To derive human haploid neural stem cells (haNSCs) to carry out lineage-specific screens. METHODS: Human haNSCs were differentiated from human extended haESCs with the help of Y27632 (ROCK signaling pathway inhibitor) and a series of cytokines to reduce diploidization. Neuronal differentiation of haNSCs was performed to examine their neural differentiation potency. Global gene expression analysis was con-ducted to compare haNSCs with diploid NSCs and haESCs. Fluorescence activated cell sorting was performed to assess the diploidization rate of extended haESCs and haNSCs. Genetic manipulation and screening were utilized to evaluate the significance of human haNSCs as genetic screening tools. RESULTS: Human haESCs in extended pluripotent culture medium showed more compact and smaller colonies, a higher efficiency in neural differentiation, a higher cell survival ratio and higher stability in haploidy maintenance. These characteristics effectively facilitated the derivation of human haNSCs. These human haNSCs can be generated by differentiation and maintain haploidy and multipotency to neurons and glia in the long term in vitro. After PiggyBac transfection, there were multiple insertion sites in the human haNSCs' genome, and the insertion sites were evenly spread across all chromosomes. In addition, after the cells were treated with manganese, we were able to generate a list of manganese-induced toxicity genes, demonstrating their utility as genetic screening tools. CONCLUSION: This is the first report of a generated human haploid somatic cell line with a complete genome, proliferative ability and neural differentiation potential that provides cell resources for recessive inheritance and drug targeted screening.

Injectable self-healing carboxymethyl chitosan-zinc supramolecular hydrogels and their antibacterial activity.

Injectable and self-healing hydrogels have found numerous applications in drug delivery, tissue engineering and 3D cell culture. Herein, we report an injectable self-healing carboxymethyl chitosan (CMCh) supramolecular hydrogels cross-linked by zinc ions (Zn2+). Supramolecular hydrogels were obtained by simple addition of metal ions solution to CMCh solution at an appropriate pH value. The mechanical properties of these hydrogels were adjustable by the concentration of Zn2+. For example, the hydrogel with the highest concentration of Zn2+ (CMCh-Zn4) showed strongest mechanical properties (storage modulus~11,000Pa) while hydrogel with the lowest concentration of Zn2+ (CMCh-Zn1) showed weakest mechanical properties (storage modulus~220Pa). As observed visually and confirmed rheologically, the CMCh-Zn1 hydrogel with the lowest Zn2+ concentration showed thixotropic property. CMCh-Zn1 hydrogel also presented injectable property. Moreover, the antibacterial properties of the prepared supramolecular hydrogels were studied against Staphylococcus aureus (S. aureus) and Escherichia coli (E. coli) by agar well diffusion method. The results revealed Zn2+ dependent antibacterial properties against both kinds of strains. The inhibition zones were ranging from ~11-24mm and ~10-22mm against S. aureus and E. coli, respectively. We believe that the prepared supramolecular hydrogels could be used as a potential candidate in biomedical fields.

Facile fabrication of moldable antibacterial carboxymethyl chitosan supramolecular hydrogels cross-linked by metal ions complexation.

Herein, carboxymethyl chitosan (CMCh) supramolecular hydrogels cross-linked by metal ions (Ag+, Cu2+ and Zn2+) are reported. The hydrogels were formed within a few seconds by simple mixing of CMCh solution with metallic salt solutions at an appropriate pH. The prepared hydrogels were characterized by using FT-IR, XRD, SEM and rotational rheometery. FT-IR measurements suggested that the facile complexation of metal ions with carboxylic, amino and hydroxyl groups of CMCh chains promoted the rapid hydrogelation. SEM images revealed a cross-linked structure of hydrogels, while microstructural openings were observed by cross-sectional studies of the freeze-dried hydrogels. Moreover, the hydrogels showed a remarkable moldability to form free standing objects. The antibacterial activities of the hydrogels were also studied against Escherichia coli and Staphylococcus aureus by agar well diffusion method. The results demonstrated an excellent antibacterial activity of the supramolecular hydrogels. Therefore, the developed CMCh supramolecular hydrogels might be used effectively in biomedical field.

Preparation, characterization and antibacterial applications of carboxymethyl chitosan/CuO nanocomposite hydrogels.

In this study, a series of antibacterial carboxymethyl chitosan/CuO nanocomposites were prepared by treating the carboxymethyl chitosan (CMCh) hydrogel with copper (II) chloride (CuCl2) solution followed by the oxidation of copper ions in the presence of sodium hydroxide. The resulting nanocomposite hydrogels were characterized by using FTIR, XRD and SEM techniques. The SEM micrographs revealed the uniform distribution of CuO nanoparticles in the hydrogel matrix with the size ranging from 20nm to 50nm, while XRD patterns confirmed the formation of monoclinic crystalline structures of CuO nanoparticles. The swelling behavior of the nanocomposite hydrogels were studied at a varied pH range. CMCh/CuO nanocomposite hydrogels showed rather higher swelling as compared to pure CMCh hydrogel. The antibacterial properties of the nanocomposite hydrogels were studied against Staphylococcus aureus and Escherichia coli. The results showed excellent antibacterial behavior of the nanocomposite hydrogels. Therefore, the developed CMCh/CuO nanocomposite hydrogels might be used effectively in numerous biomedical fields.

Recent Advances in Antimicrobial Hydrogels Containing Metal Ions and Metals/Metal Oxide Nanoparticles.

Recently, the rapid emergence of antibiotic-resistant pathogens has caused a serious health problem. Scientists respond to the threat by developing new antimicrobial materials to prevent or control infections caused by these pathogens. Polymer-based nanocomposite hydrogels are versatile materials as an alternative to conventional antimicrobial agents. Cross-linking of polymeric materials by metal ions or the combination of polymeric hydrogels with nanoparticles (metals and metal oxide) is a simple and effective approach for obtaining a multicomponent system with diverse functionalities. Several metals and metal oxides such as silver (Ag), gold (Au), zinc oxide (ZnO), copper oxide (CuO), titanium dioxide (TiO2) and magnesium oxide (MgO) have been loaded into hydrogels for antimicrobial applications. The incorporation of metals and metal oxide nanoparticles into hydrogels not only enhances the antimicrobial activity of hydrogels, but also improve their mechanical characteristics. Herein, we summarize recent advances in hydrogels containing metal ions, metals and metal oxide nanoparticles with potential antimicrobial properties.

[Construction of yeast strains expressing long-acting glucagon-like peptide-1 (GLP-1) and their therapeutic effects on type 2 diabetes mellitus mouse model].

Probiotics, i.e., bacteria expressing therapeutic peptides (protein), are used as a new type of orally administrated biologic drugs to treat diseases. To develop yeast strains which could effectively prevent and treat type 2 diabetes mellitus, we firstly constructed the yeast integrating plasmid pNK1-PGK which could successfully express green fluorescent protein (GFP) in Saccharomyces cerevisiae. The gene encoding ten tandem repeats of glucagon-like peptide-1(10 × GLP-1) was cloned into the vector pNK1-PGK and the resulting plasmids were then transformed into the S. cerevisiae INVSc1. The long-acting GLP-1 hypoglycemic yeast (LHY) which grows rapidly and expresses 10 × GLP-1 stably was selected by nutrition screening and Western blotting. The amount of 10 × GLP-1 produced by LHY reached 1.56 mg per gram of wet cells. Moreover, the oral administration of LHY significantly reduced blood glucose level in type 2 diabetic mice induced by streptozotocin plus high fat and high sugar diet.

The effects of stimulation at acupoint ST36 points against hemorrhagic shock in dogs.

The aim of this study is to investigate the effects of electroacupuncturing (EA) zusanli points on levels of basic hemodynamics, lactate, and cytokines in dogs with hemorrhagic shock. Thirty healthy dogs were randomly divided into 5 groups: sham hemorrhagic shocked group, hemorrhagic shocked group, EA group, nonacupuncturing group, and EA after vagotomy group. Zusanli points were electroacupunctured with constant voltage (10-15 V, 30 Hz) for 30 minutes immediately after the shock models were established. Before the stimulation, a blood pressure transducer was implanted into the right femoral artery for continuous recording of mean arterial pressure (MAP), and a 5F Swan-Ganz pediatric catheter was implanted into the pulmonary artery. The levels of serum tumor necrosis factor α (TNF-α) in the femoral artery were detected at 0, 120, and 180 minutes after hemorrhage. The levels of serum lactate in the femoral artery were detected before hemorrhage (-45 minutes), at 0 minute, and at 180 minutes. In the hemorrhagic shocked group, the levels of MAP, cardiac output, cardiac index, central venous pressure, and pulmonary arterial wedge pressure decreased significantly; at the same time, the levels of serum TNF-α and serum lactate increased significantly. There were no differences between these groups and the hemorrhagic group, but they were different from the sham hemorrhagic shocked group. In the EA group, the levels of MAP, cardiac output, cardiac index, central venous pressure, and pulmonary arterial wedge pressure gradually increased, but the content of serum TNF-α and lactate obviously decreased. The results suggested that EA zusanli points produce a protective effect on hemorrhagic shock in dogs.

In vitro free radical scavenging activities and effect of synthetic oligosaccharides on antioxidant enzymes and lipid peroxidation in aged mice.

In this study we synthesized oligosaccharides using glucose as reactant via a route assisted by microwave irradiation and evaluated their antioxidant activity in vivo and in vitro. The results show that the oligosaccharides exhibited antioxidant activity in vitro as compared to standard antioxidants such as butylated hydroxytoluene (BHT), and alpha-tocopherol. This antioxidant activity depended on concentration and increased with increasing dose of sample. In addition, increased endogenous lipid peroxidation and decreased total antioxidant capacity (TAOC) were observed in aged mice. Thirty-day intraperitoneal administration of the oligosaccharides significantly decreased the lipid peroxidation in a dose-dependent manner. Oligosaccharides treatment increased TAOC and the activity of superoxide dismutase (SOD) and glutathione peroxidase (GSH-Px) in all organs tested in aged mice. The present study suggests that the synthetic oligosaccharides possess promising future for their strong free radical scavenging activity. Therefore they can be employed in compensating the decline in TAOC and the activities of antioxidant enzymes and in reducing the risks of lipid peroxidation.