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| Status |
Public on Dec 31, 2023 |
| Title |
Glutaminase (GLS) was a target gene downstream of lithocholic acid inhibiting gallbladder cancer |
| Organism |
Homo sapiens |
| Experiment type |
Expression profiling by high throughput sequencing
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| Summary |
Purpose:To further explore the molecular mechanisms of LCA treatment underlying its tumor suppressive functions.
Methods:For the NOZ cells cultures, there were two different conditions performed in biological triplicates: (I) mock (DMSO), (II) 150 μM LCA for 24 h. A total amount of 1 µg RNA per sample was used as input material for the RNA sample preparations. Sequencing libraries were generated using NEBNext® UltraTM RNA Library Prep Kit for Illumina® (NEB, USA) following manufacturer’s recommendations and index codes were added to attribute sequences to each sample. The clustering of the index-coded samples was performed on a cBot Cluster Generation System using TruSeq PE Cluster Kit v3-cBot-HS (Illumina) according to the manufacturer’s instructions. After cluster generation, the library preparations were sequenced on an Illumina Novaseq platform and 150 bp paired-end reads were generated. Paired t-test was used to normalize and statistically analyze the data. The threshold we set to screen for upregulated and downregulated genes with a fold change >0 and a padj-value < 0.05. Hierarchical clustering was performed and a heat map was constructed with edgeR R package (3.18.1) to visualize differential mRNA expression. Gene set enrichment analysis was conducted with the local version of the GSEA analysis tool http://www.broadinstitute.org/gsea/index.jsp.
Results: By means of RNA-seq and subsequent data analysis with volcano plot and hierarchical clustering, we found that LCA treatment led to a different gene expression profile with the upregulation of 3607 genes and the downregulation of 3501genes compared with those in cells treated with mock. Gene Set Enrichment Analysis (GSEA) suggested that alanine, aspartate, and glutamate metabolism pathways were top-listed in the inhibitory regulation of LCA, and the heatmap plots showed that the pathways involved a set of 35genes. In addition, qRT-PCR was used to confirm the expression of several metabolism genes including GLS, ASS1, CPS1, ASNS, ABAT, and ADSSL. We were particularly interested in the commonly downregulated GLS in both NOZ and EH-GB1 cells, which mediates glutamate and glutamine metabolism. As the result, protein levels of GLS were dramatically inhibited by LCA in NOZ and EH-GB1 cells in a time-dependent manner as compared with the untreated cancer cells,
Conclusion: GLS was a downstream effector of LCA treatment in gallbladder cancer.
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| Overall design |
mRNA profiles of LCA treated gallbladder cancer cell line
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| Contributor(s) |
Li W |
| Citation missing |
Has this study been published? Please login to update or notify GEO. |
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| Submission date |
Dec 24, 2021 |
| Last update date |
Dec 31, 2023 |
| Contact name |
Weijian Li |
| E-mail(s) |
liweijian@sjtu.edu.cn
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| Phone |
+8615800386985
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| Organization name |
Renji Hospital Affiliated to Shanghai Jiao Tong University School of Medicine
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| Street address |
NO. 160, Pujian Road, Pudong New District, Shanghai, China
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| City |
Shanghai |
| State/province |
Shanghai |
| ZIP/Postal code |
200127 |
| Country |
China |
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| Platforms (1) |
| GPL11154 |
Illumina HiSeq 2000 (Homo sapiens) |
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| Samples (6)
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| Relations |
| BioProject |
PRJNA792090 |
| Supplementary file |
Size |
Download |
File type/resource |
| GSE192577_gene_count.txt.gz |
2.2 Mb |
(ftp)(http) |
TXT |
| GSE192577_gene_fpkm.txt.gz |
3.2 Mb |
(ftp)(http) |
TXT |
SRA Run Selector |
| Raw data are available in SRA |
| Processed data are available on Series record |
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