RESUMO
In chicken, primordial germ cells (PGC) are crucial for the preservation and manipulation of genetic resources in poultry production. The HiS and FAcs culture systems are two important methods for the in vitro cultivation of chicken PGCs. The purpose of this study was to compare and analyze the two cultivation systems for PGCs (His and FAcs culture systems) to assess their efficacy and applicability in supporting PGC growth, maintaining PGC characteristics, and lineage transmission ability. The study found that both HiS and FAcs culture systems could maintain the basic biological characteristics of chicken PGCs, including the simultaneous expression of pluripotency and reproductive marker genes, as well as the presence of abundant glycogen granules. Subsequently, we identified 2,145 differentially expressed genes (DEG) through RNA sequencing. GO and KEGG analysis revealed a large number of DEGs enriched in the cell adhesion and calcium ion binding pathways, and the analysis found that these genes maintained a higher level in HiS-PGCs. Further personalized analysis found that the regulatory genes for maintaining PGC pluripotency were highly expressed in HiS-PGCs, while germ cell-related genes showed similar expression in both systems. Additionally, through RNA sequencing data and cell proliferation ability, it was found that PGCs in the FAcs system had a higher proliferation rate and a faster cell cycle. Finally, it was discovered that the expression of cell migration-related genes was maintained at a higher level in HiS-PGCs, but the migration efficiency of HiS-PGCs did not show a significant difference compared to FAcs-PGCs. These results suggest that both HiS and FAcs culture systems can maintain the proliferation and basic characteristics of chicken PGCs, but differences exist in cell proliferation, pluripotency regulation, and cell adhesion. These findings provide new information for optimizing PGC cultivation systems and are important for the preservation and genetic improvement of chicken PGCs.
RESUMO
The identification of accurate gene insertion sites on chicken sex chromosomes is crucial for advancing sex control breeding materials. In this study, the intergenic region NC_006127.4 on the chicken Z chromosome and the non-repetitive sequence EE0.6 on the W chromosome were selected as potential gene insertion sites. Gene knockout vectors targeting these sites were constructed and transfected into DF-1 cells. T7E1 enzyme cleavage and luciferase reporter enzyme analyses revealed knockout efficiencies of 80.00% (16/20), 75.00% (15/20), and 75.00% (15/20) for the three sgRNAs targeting the EE0.6 site. For the three sgRNAs targeting the NC_006127.4 site, knockout efficiencies were 70.00% (14/20), 60.00% (12/20), and 45.00% (9/20). Gel electrophoresis and high-throughput sequencing were performed to detect potential off-target effects, showing no significant off-target effects for the knockout vectors at the two sites. EdU and CCK-8 proliferation assays revealed no significant difference in cell proliferation activity between the knockout and control groups. These results demonstrate that the EE0.6 and NC_006127.4 sites can serve as gene insertion sites on chicken sex chromosomes for gene editing without affecting normal cell proliferation.
Assuntos
Galinhas , Edição de Genes , Cromossomos Sexuais , Animais , Galinhas/genética , Edição de Genes/métodos , Cromossomos Sexuais/genética , Mutagênese Insercional , Sistemas CRISPR-Cas , Linhagem Celular , Técnicas de Inativação de Genes/métodos , Feminino , MasculinoRESUMO
The utilization of chicken embryonic-derived pluripotent stem cell (PSC) lines is crucial in various fields, including growth and development, vaccine and protein production, and germplasm resource protection. However, the research foundation for chicken PSCs is relatively weak, and there are still challenges in establishing a stable and efficient PSC culture system. Therefore, this study aims to investigate the effects of the FGF2/ERK and WNT/ß-catenin signaling pathways, as well as different feeder layers, on the derivation and maintenance of chicken embryonic-derived PSCs. The results of this study demonstrate that the use of STO cells as feeder layers, along with the addition of FGF2, IWR-1, and XAV-939 (FIX), allows for the efficient derivation of chicken PSC-like cells. Under the FIX culture conditions, chicken PSCs express key pluripotency genes, such as POUV, SOX2, and NANOG, as well as specific proteins SSEA-1, C-KIT, and SOX2, indicating their pluripotent nature. Additionally, the embryoid body experiment confirms that these PSC-like cells can differentiate into cells of three germ layers in vitro, highlighting their potential for multilineage differentiation. Furthermore, this study reveals that chicken Eyal-Giladi and Kochav stage X blastodermal cells express genes related to the primed state of PSCs, and the FIX culture system established in this research maintains the expression of these genes in vitro. These findings contribute significantly to the understanding and optimization of chicken PSC culture conditions and provide a foundation for further exploration of the biomedical research and biotechnological applications of chicken PSCs.
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Cr(III) as one of the most concerned potentially toxic elements, is discharged from relevant industries and Cr(VI) reduction. Hydrogel-based adsorption could be one of the promising approaches for Cr(III) removal. Featured with environmental friendliness and low cost, carboxymethyl cellulose (CMC) was employed for the hydrogel synthesis, and attapulgite (APT) could be used to strengthen its stability. However, the adsorption performance and mechanisms need to be examined. In the present study, carboxymethyl cellulose-g-poly(acrylic acid-co-acrylamide)/ attapulgite (CMC-g-p(AA-co-AM)/APT) was synthesised via in situ copolymerisation. Its efficacy for removing Cr(III) from an aqueous solution was investigated using batch adsorption experiments. Results showed that the introduction of APT enhanced the thermal stability but decreased the swelling performance of the hydrogel. The prepared hydrogel could strongly adsorb Cr(III) at a wide pH range of 3.0-7.0. Cr(III) can be efficiently removed by the composite hydrogel within 1-2â h. At low concentration, CMC-g-p(AA-co-AM)/APT could slightly adsorbed more Cr(III) than CMC-g-p(AA-co-AM). The maximum absorption of CMC-g-p(AA-co-AM) and CMC-g-p(AA-co-AM)/APT were 74.8 and 47.7â mg/g at 298â K, respectively. The negative value of ΔHo and ΔGo indicated the adsorption of Cr(III) onto the two studied hydrogels is an exothermic and spontaneous process. Ion exchange and complexation, as implied by EDS, FT-IR and XPS, combining with electrostatic attraction are the possible adsorption mechanisms for Cr(III) onto the prepared hydrogels. All the results above suggests that the composite hydrogel CMC-g-p(AA-co-AM)/APT can be a promising candidate for the removal of Cr(III) from waste water.
Assuntos
Hidrogéis , Poluentes Químicos da Água , Carboximetilcelulose Sódica , Adsorção , Acrilamida , Espectroscopia de Infravermelho com Transformada de Fourier , Poluentes Químicos da Água/análise , Concentração de Íons de Hidrogênio , Cinética , Cromo/análiseRESUMO
Panax notoginseng is the dried root and rhizome of Panax notoginseng, which has the effect of lowering blood lipid, lowering blood pressure and promoting blood circulation to remove blood stasis. At present, the research on Panax notoginseng is mainly focused on its pharmacological action and its compound preparation, but the research on the granule of Panax notoginseng is less. This paper mainly studied the clinical study of compound notoginseng nanoparticles in the treatment of local infection in patients with hydrocephalus after medium craniocerebral injury in neurosurgery. The purpose of this article is to investigate the effects of compound notoginseng nanoparticles on serum TNF-α, IL-2 and IL-6 in rats with craniocerebral injury and to verify the protective effect of compound notoginseng nanoparticles on the body after craniocerebral injury. In this paper, 90 patients admitted to a hospital in this city were divided into a control group, model group and compound notoginseng nanoparticle group. According to the Zealonga method, the neurological function deficit score of experimental rats in each group was evaluated. The levels of TNF-α, IL-2 and IL-6 in the serum of the three groups were observed 1, 3 and 5 days after treatment. RESULTS: Compared with serum TNF-α, IL-2 and IL-6 of the three groups, there were significant differences in the main effects of time and intervention (P < 0.05). CONCLUSIONS: Compound notoginseng nanoparticles can reduce the contents of TNF-α and IL-6 in serum and increase the expression of IL-2 in rats with craniocerebral injury.
Assuntos
Traumatismos Craniocerebrais , Hidrocefalia , Nanopartículas , Neurocirurgia , Animais , Traumatismos Craniocerebrais/tratamento farmacológico , Medicamentos de Ervas Chinesas/uso terapêutico , Hidrocefalia/tratamento farmacológico , Hidrocefalia/cirurgia , Interleucina-2 , Interleucina-6 , Nanopartículas/uso terapêutico , Panax notoginseng , Ratos , Fator de Necrose Tumoral alfaRESUMO
As a classical model system of embryo biology, the chicken embryo has been used to investigate embryonic development and differentiation. Delivering exogenous materials into chicken embryos has a great advantage for studying gene function, transgenic breeding, and chimera preparation during embryonic development. Here we show the method of in ovo intravascular injection whereby exogenous materials such as plasmid vectors or modified primordial germ cells (PGCs) can be transferred into donor chicken embryos at early developmental stages. The results show that the intravascular injection through the dorsal aorta and head allows injected materials to diffuse into the whole embryo through the blood circulatory system. In the presented protocol, the efficacy of exogenous plasmid and lentiviral vector introduction, and the colonization of injected exogenous PGCs in the recipient gonad, were determined by observing fluorescence in the embryos. This article describes detailed procedures of this method, thereby providing an excellent approach to studying gene function, embryo and developmental biology, and gonad-chimeric chicken production. In conclusion, this article will allow researchers to perform in ovo intravascular injection of exogenous materials into chicken embryos with great success and reproducibility.