Basic information ofHuman glioblastoma U251MG
Human glioblastoma U251MG is a cell line derived from patients with glioblastoma (GBM), primarily used for cancer research. The U251MG cell line has the following characteristics:
1. Source and characteristics: The U251MG cell line is derived from the brain tissue of a 75 year old male patient, specifically a grade III-IV astrocytoma. These cells are fibroblast like, polymorphic, and grow adherent to the wall. The cell morphology is diverse and usually appears polygonal or fibroblast like under culture conditions.
2. Cultivation conditions: The culture medium for U251MG cells is usually DMEM medium, which contains a mixture of 10% fetal bovine serum (FBS) and 1% antibiotics [6]. The growth conditions are 95% air and 5% carbon dioxide, with a temperature of 37℃. The passage ratio is generally 1:2 to 1:3, and the passage period is 2-3 days.
3. Gene expression and markers: U251MG cells express various molecular markers, such as PDGFR alpha, EGFR, and GFAP. These markers help confirm the characteristics of astrocytes in cells. In addition, U251MG cells also showed positive reactions to astrocyte markers such as GFAP, S100 β, and vimentin [5].
4. Application areas: U251MG cells are widely used in cancer research, especially in studying the mechanisms of glioblastoma, drug screening, and gene editing. Due to its easy transfection property, U251MG cells have also been used to study the growth and expression of tumor cells.
5. Freezing and transportation: The freezing solution formula for U251MG cells is usually 90% basic medium, 10% FBS, and 10% DMSO [2]. The freezing condition is liquid nitrogen storage, which is effective for a long time.
GMP grade non programmed cell cryopreservation solution (exclusively for immune cells, free of protein/serum/animal derived components)
6. Precautions: U251MG cells are for scientific research purposes only and should not be used for clinical diagnosis or treatment. When publishing research papers or results, users should indicate the source of the cell line and comply with relevant laws and regulations.
Human glioblastoma U251MG is an important experimental model widely used in the study of glioblastoma, with clear sources, specific culture conditions, and diverse application fields.
What is the detailed gene expression profile of human glioblastoma U251MG cell line?
The detailed gene expression profile of human glioblastoma U251MG cell line can be analyzed using various methods. The U251MG cell line used CRISPR technology in gene knockout studies, and achieved the knockout of RSK1 and RSK2 genes by designing gRNA sequences targeting these two genes [7]. In addition, cell expression profiles were analyzed through microarray chip experiments. The integrity of RNA samples was checked using Agilent 2100 Bioanalyzer, and the hybridized chips were scanned using Affymetrix Gene Chip Array Scanner 3000-7G to extract expression intensity data and store it as a CEL file. Normalize CEL files using the SST-RMA-GENE-FULL algorithm and perform differential expression analysis [7].
In addition, the U251MG cell line has been reported to express genes such as PDGFR alpha, EGFR, and GFAP. The expression of these genes may vary under different experimental conditions, but they are one of the important features of this cell line.
What are the specific application cases of U251MG cell line in glioblastoma research?
The U251MG cell line has a wide range of applications in the study of glioblastoma. Here are several specific application cases:
The U251MG cell line was used to study resistance to temozolomide (TMZ). By gradually adapting to the TMZ concentration, a drug-resistant variant U251MG-R cell line was obtained. These cell lines are used to study resistance mechanisms and explore new therapeutic strategies, such as the study of miR-204 reversing temozolomide resistance [11].
Use U251MG cell line for apoptosis and necrosis testing to evaluate the effects of different drugs on glioblastoma cells. For example, in the study of the interaction between Nb206 and TUFM, the Annexin V-FITC/Propidium Iodide kit was used to perform apoptosis and necrosis tests on U251MG cells to observe the effect of Nb206 on cell survival [9].
The U251MG cell line was used to screen for potential anti-tumor drugs. For example, studies on the effects of glucosinolates (SFN) on U251MG cells have shown that SFN can significantly inhibit the survival rate of U251MG cells, induce apoptosis, and suppress their invasive ability [15]. In addition, OM (oxidized brucine) also showed significant anti-tumor effects on U251MG cells, effectively inhibiting proliferation and invasion, and inducing apoptosis [10].
Researchers investigated the role of miR-218 in glioblastoma cells by constructing a stable U251MG cell line transfected with miR-218. The results indicate that miR-218 inhibits the proliferation of glioblastoma cells through the YY1/p53 signaling pathway [12]. Similarly, miR-340 was also constructed as a stable expression U251MG cell line for studying its predictive role in glioblastoma [13].
The U251MG cell line was used to establish a xenograft model and study the sensitivity of undifferentiated glioblastoma cell populations to anti-tumor agents. For example, U251MG-P1 cells were used to evaluate the sensitivity of cisplatin, and M-CTX Fc bound liposomes were designed to encapsulate cisplatin for enhanced anti-tumor efficacy [14].
How to optimize the culture conditions of U251MG cells to improve their stability and experimental efficiency?
In order to optimize the culture conditions of U251MG cells and improve their stability and experimental efficiency, adjustments can be made from the following aspects:
1. Selection and composition of culture medium:
According to reference [16], U251MG cells are typically cultured in DMEM medium supplemented with 10% fetal bovine serum (FBS), penicillin, and streptomycin. In addition, adding MEM non essential amino acids and antifungal agents such as amphotericin B can also be considered to ensure comprehensive nutritional support and prevent contamination.
In some experiments, high glucose DMEM/F12 medium was used and supplemented with 10% heat inactivated fetal bovine serum and 1% penicillin streptomycin solution [21]. This medium formula may be more suitable for certain specific experimental needs, so the appropriate medium can be selected according to the specific experimental objectives.
2. Control of cultivation environment:
Cells should be cultured under conditions of 37℃, 5% CO2, and controlled humidity [16]. These conditions help maintain the physiological state and metabolic activity of cells.
For studies that require three-dimensional culture models, the hanging drop method can be used to form cell spheres [16]. This method helps to simulate the tumor microenvironment in vivo and improve the biological relevance of experiments.
3. Cell density and stratification time:
Cells should be stratified near confluence, typically around 80% [19]. This can avoid cell death or growth inhibition caused by overgrowth.
During stratification, 0.25% trypsin EDTA solution was used and incubated at 37℃C to promote cell separation [19]. This helps maintain the integrity and activity of cells.
4. Frequency of medium replacement:
Change the culture medium every 2-3 days to ensure that cells receive sufficient nutrition and timely removal of metabolic waste [18,19]. Frequent replacement of culture medium helps maintain the health status of cells and the accuracy of experimental results.
5. Cell viability and stability testing:
Regularly detect cell viability using Alamar blue cell viability analysis [19]. This method can monitor the growth status of cells in real-time and their response to experimental treatments.
Cell survival rate can be measured by MTT reduction assay to evaluate the effects of different compounds or treatments on cells [18].
6. Transfection and gene editing:
For experiments that require gene editing or transfection, Lipofectamine 3000 or other transfection reagents can be used for operation [21]. Choosing appropriate transfection methods and reagents can improve transfection efficiency and cell stability.
When using CRISPR technology for gene editing, stable cell lines can be established through FACS sorting [17]. This method ensures the acquisition of homozygous cell lines with the desired gene mutations.
How effective is the U251MG cell line in drug screening, and what are its advantages and disadvantages compared to other glioblastoma cell lines?
The U251MG cell line has shown significant efficacy in drug screening, but there are certain advantages and disadvantages compared to other glioblastoma (GBM) cell lines.
Advantages:
1. High expression of TUFM: Studies have shown that the expression level of TUFM (a microRNA) in U251MG cell lines is significantly higher than that in control tissues, which may make it more sensitive in certain drug screening [9].
2. Reactivity to multiple compounds: In drug screening experiments, U251MG cell lines exhibit different reactivity to various targeted compounds and chemotherapy drugs. For example, Nb206 significantly inhibited the proliferation of U251MG cells in a dose-dependent manner [9]. In addition, the U251MG cell line has low resistance to temozolomide (TMZ), making them an ideal model for studying the mechanism of TMZ resistance [8].
3. Cell cycle arrest: The U251MG cell line has a cell retention rate of up to 90% in the G2/M phase, indicating that they are particularly sensitive to certain drug treatments, such as SynB1-ELP-DOXO and free cisplatin [23].
inferiority:
1. Drug resistance issue: Although the U251MG cell line has a high sensitivity to TMZ, its resistant sub line exhibits significant chemical resistance, especially under neurosphere conditions, with significantly increased IC50 values [27]. This means that in long-term experiments, drug resistance may limit its application.
2. Cell Heterogeneity: The U251MG cell line exhibits high cell heterogeneity, which may affect the reproducibility and reliability of drug screening results. For example, cell populations under different conditions may exhibit different drug reactivity [26].
3. Gene expression differences: Compared with other GBM cell lines, U251MG cell line has differences in certain gene expression. For example, the high expression of FREM2 and SPRY1 proteins in U251MG cells may affect the results of drug screening [25].
The U251MG cell line has significant advantages in drug screening, especially in high expression of TUFM and responsiveness to multiple compounds. However, its drug resistance and cellular heterogeneity also pose challenges.
What are the latest research advances on the U251MG cell line?
The latest research progress on U251MG cell line can be explained in detail from the following aspects:
Researchers successfully knocked out the PLEKHG5 gene in U251-MG cells and validated this process through genomic PCR and immunohistochemistry analysis. The results showed that after knockout, the expression of PLEKHG5 gene in cells significantly decreased, and the cell morphology changed, losing its typical spindle shape and cilia, and the cell division time was significantly prolonged. In addition, the study also found that the accumulation of autophagosomes in cells significantly increased after knockout, which was correlated with a significant decrease in LAMP-1 protein levels. These changes may be related to the proliferation and survival of tumor cells [30].
Multiple proteomic and transcriptomic analyses were conducted on the U251-MG cell line. Proteins in cell lysate were isolated and identified using SDS-PAGE and immunoblotting techniques. In addition, NF - κ B activated Luciferase activity was evaluated by transient transfection of U251-MG cells, and GST-RBD pulldown experiments were conducted to assess the specific activity of RhoA. These experiments revealed the methylation status of the MGMT promoter and performed sequence analysis using the Therascreen MGMT Pyro Kit [30].
The study analyzed the ultrastructural changes of U251 cells under different drug treatments using transmission electron microscopy (TEM), and identified three characteristics representing different cell death pathways: apoptosis, autophagy, and necrotic cell apoptosis. Especially the increased expression of caspase-3 and caspase-8 indicates the activation of the apoptotic pathway. In addition, the study also investigated the activation of caspase-3 substrate PARP-1 [29].
The study investigated the effect of GDNF (brain-derived neurotrophic factor) on proN cadherin expression and cell survival rate in U251MG cells. The results showed that after GDNF treatment, the expression levels of proN cadherin in the membrane and cytoplasm of U251MG cells were significantly increased, while the survival rate of cells was also improved [32].
Shanghai Gene Chemical Company synthesized four shRNA sequences for transfection into U251 MG cells and conducted cell proliferation experiments. The results showed that different concentrations of TNF - α stimulated the proliferation of U251MG cells within 24 hours, with 10 ng/mL TNF - α showing the best effect. In addition, the study also evaluated the relationship between ANXA1 and Ki-67, as well as the significance of ANXA1 expression and clinical pathological variables [28].
Research has found that RC-7 has a significant inhibitory effect on the cell survival rate of U251MG cells at concentrations of 12.5 nM and 50 nM, and RC-7 mainly acts on the mitochondria of the cells. This indicates that RC-7 has significant inhibitory effects on cell growth and induces apoptosis in U251MG cells [20].
The study investigated the effects of Hirsutinolite series compounds on Stat3 activity and their impact on various proteins. Through cell cycle distribution analysis and protein immunoblotting analysis, it was found that U251MG cells treated with Cmpd1 exhibited significant changes in cell cycle and protein expression levels [31].
Single cell analysis using C1 Fluidigm technology revealed the persistent characteristics of U251 cells after treatment with temozolomide (TMZ), and identified four key genes associated with it: CHI3L1, FAT2, HB-EGF, and KLK5. The functions of these genes may be related to inflammation, extracellular tissue remodeling, and tumor drug resistance mechanisms [8].
The growth, proliferation, migration, and invasion abilities of U-251 MG and U-87 MG cell lines with stable knockdown of USP32 gene were observed through various cell biology methods, such as colony formation assay, EdU incorporation assay, flow cytometry, and Transwell assay.
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