Analytical Methods
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2025
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IF 2.6
Chen Lu,Jiayue Chen,Shengsheng Zhang,Yaru Tian,Xinyu Ying,Dan Zhang,Zhidan Luo,Xinxin Si,Ming Li
Two single nucleotide polymorphisms (SNPs) in the human SLCO1B1 gene, c.388A>G (rs2306283) and c.521T>C (rs4149056), are independent determinants of the efficacy and side effects of statin drugs. Multinational clinical guidelines recommend testing for SLCO1B1 genotypes before the initial use of statins. Current SLCO1B1 SNP identification methods, primarily based on quantitative fluorescence PCR, rely on expensive equipment, are time-consuming, and require cold-chain storage for reagents, making them unsuitable for use in resource-limited healthcare settings. In this study, we developed a CRISPR/Cas12b-based amplification-free genotyping technique for SLCO1B1 SNPs. Within 30 minutes of the isothermal reaction, genotyping of the c.388A>G and c.521T>C SNPs in the SLCO1B1 gene can be observed by the naked eyes under blue light. Additionally, maltodextrin was identified as an effective vitrification stabilizer for the CRISPR/Cas12b premix. A low-cost vitrification process was optimized to prepare a glass like solid reagent via room-temperature vacuum drying. The vitrified CRISPR/Cas12b reagent retained approximately 88% of its activity after 30 days of storage at 37 °C, eliminating the need for cold-chain storage and allowing for long-term preservation at room temperature. This vitrified CRISPR/Cas12b based rapid SNP detection technique is especially suitable for genotyping drug metabolism genes in primary healthcare settings, providing effective guidance for precision medicine in clinical practice.
研究领域:
分子生物学
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医学遗传学
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精准医疗
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产品关联
DOI
PubMed
高影响因子
BIOTECHNOLOGY PROGRESS
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2025
·
IF 2.5
Shumin Zhang,Huimin Huang,Jingwei Zhang,Yuanyuan Zhao,Wen-song Tan,Haibo Cai
Cell culture medium is an important factor affecting the expansion of NK cells in vitro. As an important component of cell culture medium, lipids participate in various complex physiological activities of cells and significantly affect the expansion of cells. Using NK-92 cells as a model, the lipid metabolism of NK cells in vitro was analyzed, and combined with the kinetic relationship between lipid metabolism and NK cell expansion. Four fatty acids, oleic acid, linoleic acid, palmitoleic acid, and α-linolenic acid, were preliminatively identified as the key lipid combinations. The combination was preliminarily verified on the self-developed serum-free medium. It was found that when the key lipid combination was added according to the concentration in the serum, NK-92 cells expansion reached 188.03 ± 33.34-folds, which was significantly higher than 105.28 ± 13.23-folds in the basic medium. Additionally, NK-92 cells expanded by adding key lipid combinations could maintain cell killing function. Overall, this research provides technical support for the development of NK cell serum-free medium.
研究领域:
免疫学
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细胞生物学
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产品关联
DOI
PubMed
高影响因子
Bioresources and Bioprocessing
·
2022
·Vol. 9
·期 1
·1-11
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IF 4.983
Liu Yangyang,Sun Qihao,Hao Mengyang,Tan Wen‑Song,Cai Haibo
The application of natural killer (NK) cells as potential antitumor effector cells appears to be valuable for immunotherapies. However, the clinical use of NK cells is limited because the technical difficulties associated with mass production NK cells at sufficiently high numbers represents a great challenge. Ex vivo expansion of NK cells is a key technology for cell therapy. Bioreactor systems can generate homogeneous culture condition and modulate the environmental and biochemical cues. In this study, a novel magnetically controlled bioreactor was developed for supporting NK cells ex vivo expansion. Using synthetic magnetic beads, the stirring device of the magnetically controlled bioreactor generated reduced shearing force. The intermittent magnetic field was applied for magnetic beads movement to homogenize the culture system. NK-92 cells were cultured in the magnetically controlled bioreactor and the expansion and function of expanded cells were investigated on day 8. The results showed that the expansion of NK-92 cells in the bioreactor was 67.71 ± 10.60-fold, which was significantly higher than that of the T25 culture flask (P < 0.05). Moreover, the proportions of CD3−CD56+ cells and cell killing activity of expanded cells in the bioreactor did not reveal any differences compared to T25 flasks. Taken together, this study demonstrated the possibility of magnetically controlled bioreactor as a potent strategy in NK cells production for facilitating cancer immunotherapy.
研究领域:
生物工程
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免疫学
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肿瘤学
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DOI
高影响因子
Journal of Materials Chemistry B
·
2024
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IF 6.1
Jing Hong,Zhanchi Zhu,Leisha Cui,Zhaojun Wang,Ying Hao,Xiaopeng Tian,Guosheng Cheng
Hematopoietic stem cell (HSC) expansion offers a key strategy to address the source limitation and donor shortages of HSCs for the treatment of various blood disorders. Specific remodeling of the complex bone marrow microenvironment that contributes to efficient in vitro expansion of HSCs remains challenging. Here, inspired by the regions with different stiffness levels in the bone marrow niche, a three dimensional (3D) bone marrow-mimicking composite scaffold created based on gelatin–hyaluronic acid (Gel–HA) hydrogels and graphene foams (GFs) was engineered to support the in vitro expansion of HSCs. The composite scaffold was prepared by forming a photo-cross-linked Gel–HA hydrogel surrounding the GF. The “soft” Gel–HA hydrogel and “stiff” GF replicate the structure and stiffness of the vascular niche and endosteal niche in the bone marrow, respectively. Furthermore, HSCs cultured in the Gel–HA/GF scaffold proliferated well and retained the CD34+CD38− immunophenotype and pluripotency, suggesting that the Gel–HA/GF composite scaffold supported the in vitro expansion of HSCs, maintaining the primitive phenotype and the ability to differentiate into functional blood cells. Thus, the hydrogel/graphene composite scaffold offers a means of facilitating HSC expansion through structurally and mechanically mimicking bone marrow niches, demonstrating great promise for HSC transplantation.
研究领域:
生物材料
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组织工程
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干细胞生物学
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生物医学工程
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产品关联
DOI
PubMed
高影响因子
Stem Cell Reviews and Reports
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2022
·Vol. 18
·期 3
·1168-1180
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IF 6.692
Wang Jin,Xiong Minghao,Sun Qihao,Tan Wen-Song,Cai Haibo
The existing approaches of hematopoietic stem cells (HSCs) expansion in vitro were difficult to meet the needs of clinical application. While in vivo, HSCs efficiently self-renew in niche where they interact with three dimension extracellular matrix and stromal cells. Osteoblasts (OBs) are one of most significant stromal cells of HSCs niche. Here, we proposed a three-dimensional environment based on gallic acid grafted-chitosan ( 2c ) scaffold and OBs differentiated from human umbilical cord mesenchymal stem cells (HUMSCs) to recapitulate the main components of HSCs niche. The results of alkaline phosphatase staining and alizarin red staining demonstrated that HUMSCs were successfully induced into OBs. The results showed that the expansions of CD34 + cells, CD34 + CD38 − cells and CD34 + CD38 − CD45RA − CD49f + CD90 + cells (primitive hematopoietic stem cells, pHSCs) harvested from the biomimetic HSCs niche based on 2c scaffold and OBs ( IV) group were larger than those harvested from other three culture groups. Importantly, it was found that the CD34 + cells harvested from IV group had better secondary expansion capability and colony forming potential, indicating better self-renewal ability. In addition, the biomimetic HSCs niche based on 2c scaffold and OBs protected HSCs apoptosis and promoted HSCs division. Taken together, the biomimetic HSCs niche based on 2c scaffold and OBs was an effective strategy for ex vivo expansion of HSCs in clinical scale. Graphical abstract
研究领域:
生物医学工程
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干细胞生物学
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组织工程
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再生医学
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DOI
PubMed
高影响因子
Stem Cell Research & Therapy
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2019
·Vol. 10
·期 1
·1-11
·
IF 4.627
Liao Naishun,Shi Yingjun,Zhang Cuilin,Zheng Youshi,Wang Yingchao,Zhao Bixing,Zeng Yongyi,Liu Xiaolong,Liu Jingfeng
Background Adipose tissue-derived mesenchymal stem cells (ADSCs) are promising candidates for regenerative medicine. However, long-term in vitro passaging leads to stemness loss and cell senescence of ADSCs, resulting in failure of ADSC-based therapy. Methods In this study, ADSCs were treated with low dose of antioxidants (reduced glutathione and melatonin) with anti-aging and stem cell protection properties in the in vitro passaging, and the cell functions including stem cell senescence, cell migration, cell multidirectional differentiation potential, and ROS content were carefully analyzed. Results We found that GSH and melatonin could maintain ADSC cell functions through reducing cell senescence and promoting cell migration, as well as by preserving stemness and multidirectional differentiation potential, through inhibiting ROS generation during long-term expansion of ADSCs. Conclusions Our results suggested that antioxidant treatment could efficiently prevent the dysfunction and preserve cell functions of ADSCs after long-term passaging, providing a practical strategy to facilitate ADSC-based therapy.
研究领域:
再生医学
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干细胞生物学
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细胞衰老
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氧化应激
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产品关联
DOI
PubMed
高影响因子
FOOD RESEARCH INTERNATIONAL
·
2024
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IF 7
Shengliang Zhang,Hongyun Lu,Hanghang Lou,Ying Shi,Donghong Liu,Qihe Chen
The large-scale production of cultured meat has been restricted by high cost of serum, batch difference and unclear composition, thus it is important to develop serum-free media for cell growth. Here we obtained a serum-free medium (SFM), which was suitable for ex vivo culturing of myoblasts from larimichthys crocea . It offered simple composition and high effectiveness that supporting myoblast attachment, expansion and differentiation. Our findings revealed that the optimal SFM consisted of DMEM/F12 base medium, 5 mg/mL bovine serum albumin (BSA), 20 mmol HEPES, 1 % penicillin/streptomycin mixture (PS) and the exogenous additive hydrocortisone. The living cells of SFM could occupy about 65 % of the serum-containing medium (SM). Additionally, differentiation assays showed that myoblasts cultured without serum still differentiated into more muscle tubes, with a higher fusion index than those cultured with serum. Finally, transcriptome sequencing was conducted to demonstrate that hydrocortisone might act as a ligand binding to G protein-coupled receptors, triggering the downstream PI3K/AKT signaling pathway and leading to cell proliferation. Our findings will provide a basis for large-scale cell expansion and cell culture meat production in the future.
研究领域:
细胞生物学
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组织工程
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生物技术
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食品科学
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产品关联
DOI
PubMed
高影响因子
iScience
·
2021
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IF 5.458
Yuying Zhang,Anqi Guo,Cheng Lyu,Ran Bi,Zhaozhao Wu,Wenjing Li,Peng Zhao,Yudi Niu,Jie Na,Jianzhong Jeff Xi,Yanan Du
Summary It is still a challenge for synthesizing ‘cellular niche-mimics’ in vitro with satisfactory reproducibility and fidelity to recreate the natural niche components (e.g., extracellular matrices and soluble factors) for stem cell cultivation. Inspired by the massive amplification of hepatic progenitor cells during liver fibrosis in vivo , here we optimized the in vitro liver fibrotic niches and subsequently harvested their bioactive ingredients as niche extracts (NEs). The fibrosis-relevant NE marginally outperformed Matrigel for phenotype maintenance of human embryonic stem cell (hESC)-derived hepatoblasts (HBs) and recapitulation of the pathological angiogenesis of hESC-derived endothelial cells both in 2D culture and 3D liver organoids. Finally, defined NE components (i.e., collagen III, IV, IL-17, IL-18 and M-CSF) were resolved by the quantitative proteomics which exhibited advantage over Matrigel for multi-passaged HB expansion. The pathology-relevant and tissue-specific NEs provide innovative and generalizable strategies for the discovery of optimal cellular niche and bioactive niche compositions.
研究领域:
干细胞生物学
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组织工程
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细胞微环境
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蛋白质组学
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类器官培养
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DOI
PubMed
高影响因子
危险品化学品经营许可证（带存储）