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1.
Chinese Journal of Biotechnology ; (12): 1086-1095, 2022.
Artigo em Chinês | WPRIM | ID: wpr-927765

RESUMO

ERα-36 is a novel subtype of estrogen receptor α which promotes tumor cell proliferation, invasion and drug resistance, and it serves as a therapeutic target. However, only small-molecule compounds targeting ERα-36 are under development as anticancer drugs at present. Gene therapy approach targeting ERα-36 can be explored using recombinant adenovirus armed with decoy receptor. The recombinant shuttle plasmid pDC316-Ig κ-ERα-36-Fc-GFP was constructed via genetic engineering to express an Ig κ-signaling peptide-leading secretory recombinant fusion protein ERα-36-Fc. The recombinant adenovirus Ad-ERα-36-Fc-GFP was subsequently packaged, characterized and amplified using AdMaxTM adenovirus packaging system. The expression of fusion protein and functional outcome of Ad-ERα-36-Fc-GFP transduction were further analyzed with triple-negative breast cancer MDA-MB-231 cells. Results showed that the recombinant adenovirus Ad-ERα-36-Fc-GFP was successfully generated. The virus effectively infected MDA-MB-231 cells which resulted in expression and secretion of the recombinant fusion protein ERα-36-Fc, leading to significant inhibition of EGFR/ERK signaling pathway. Preparation of the recombinant adenovirus Ad-ERα-36-Fc-GFP provides a basis for further investigation on cancer gene therapy targeting ERα-36.


Assuntos
Adenoviridae/genética , Proliferação de Células , Receptor alfa de Estrogênio/metabolismo , Proteínas Recombinantes , Transfecção
2.
Chinese Journal of Biotechnology ; (12): 969-978, 2020.
Artigo em Chinês | WPRIM | ID: wpr-826879

RESUMO

Drugs targeting immune checkpoint are used for cancer treatment, but resistance to single drug may occur. Combination therapy blocking multiple checkpoints simultaneously can improve clinical outcome. Therefore, we designed a recombinant protein rPC to block multiple targets, which consists of extracellular domains of programmed cell death protein 1 (PD-1) and cytotoxic T lymphocyte-associated antigen 4 (CTLA-4). The coding sequence was inserted into expression vector and stably transfected into HEK293 cells. The culture supernatant was collected and rPC was affinity-purified. Real-time quantitative PCR was used to evaluate the expression levels of ligands for PD-1 and CTLA-4 in several human cancer cell lines. The binding of rPC with cancer cells was examined by immunofluorescence cell staining, the influence of rPC on cancer cell growth was assayed by CCK-8. The results showed that rPC could be expressed and secreted by stably transfected HEK293 cells, the purified rPC could bind to lung cancer NCI-H226 cells which have high levels of ligands for PD-1 and CTLA-4, no direct impact on cancer cell growth could be observed by rPC treatment. The recombinant protein rPC can be functionally assayed further for developing novel immunotherapeutic drugs for cancer.


Assuntos
Animais , Humanos , Antígeno CTLA-4 , Genética , Proliferação de Células , Células HEK293 , Neoplasias Pulmonares , Metabolismo , Receptor de Morte Celular Programada 1 , Genética , Ligação Proteica , Domínios Proteicos , Genética , Proteínas Recombinantes de Fusão , Genética , Metabolismo
3.
Protein & Cell ; (12): 213-224, 2012.
Artigo em Inglês | WPRIM | ID: wpr-757278

RESUMO

The self-renewal and multipotent potentials in neural stem cells (NSCs) maintain the normal physiological functions of central nervous system (CNS). The abnormal differentiation of NSCs would lead to CNS disorders. However, the mechanisms of how NSCs differentiate into astrocytes, oligodendrocytes (OLs) and neurons are still unclear, which is mainly due to the complexity of differentiation processes and the limitation of the cell separation method. In this study, we modeled the dynamics of neural cell interactions in a systemic approach by mining the high-throughput genomic and proteomic data, and identified 8615 genes that are involved in various biological processes and functions with significant changes during the differentiation processes. A total of 1559 genes are specifically expressed in neural cells, in which 242 genes are NSC specific, 215 are astrocyte specific, 551 are OL specific, and 563 are neuron specific. In addition, we proposed 57 transcriptional regulators specifically expressed in NSCs may play essential roles in the development courses. These findings provide more comprehensive analysis for better understanding the endogenous mechanisms of NSC fate determination.


Assuntos
Animais , Camundongos , Astrócitos , Biologia Celular , Metabolismo , Diferenciação Celular , Genética , Perfilação da Expressão Gênica , Redes Reguladoras de Genes , Células-Tronco Neurais , Biologia Celular , Metabolismo , Oligodendroglia , Biologia Celular , Metabolismo , Mapeamento de Interação de Proteínas
4.
China Pharmacy ; (12)1991.
Artigo em Chinês | WPRIM | ID: wpr-518522

RESUMO

OBJECTIVE:To study the extraction process of ginsenoside.METHODS:The optimal extraction process was selected with the orthogonal design.The content of ginsenoside in ginseng was determined by UV-Spectrophotometry.RESUL_TS:The amount and concentration of alcohol and extraction time showed significant influence on extract obtained.CONCLUSI_ON:The optimal extraction process is as follows:adding 75% alcohol into crude ginseng(6∶1 in weight) and extracting 6 times for 45 min each extraction.

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