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Updated: Aug 29, 2026
Rapid In Vitro Cytotoxicity Evaluation of Jurkat Expressing Chimeric Antigen Receptor using Fluorescent Imaging
Published on: October 27, 2023
[Construction of subtracted cDNA library in human Jurkat T cell line induced by arsenic trioxide in vitro]
Yue-xin Zhang1, Yan Liu, Kai-tai Liu
1Department of Infectious Diseases, First Affiliated Hospital, Xinjiang Medical University, Urumuqi, Xinjiang 830054, China.
Objective:
To understand the differentially expressed genes in human T lymphocytes induced by arsenic trioxide (As(2)O(3)) and to explore mechanism of its immunotoxicity and immune suppression.
Methods:
Human Jurkat T cell line was treated by arsenic trioxide (5 micromol/L, 24 h) in vitro, as a sample model. Then, the differentially expressed genes were cloned and the subtractive cDNA library from Jurkat T cell line was constructed by suppression subtractive hybridization (SSH). Polymerase chain reaction (PCR) and sequencing techniques were applied to identify positive clones.
Results:
The forward subtracted cDNA library contained differentially expressed genes from Jurkat T cell line induced by arsenic trioxide was constructed, including 29 different gene fragments and only replicated one in the subtracted cDNA library identified by PCR and sequencing analysis. These gene sequences were 95%-100% analogous to the genes in public database (GenBank/EMBL). The cDNA library contained oxidative metabolic genes in mitochondria (triose phosphate dehydrogenase, NADH4, pyrophosphate synthase, 16S rRNA ribosome, succinate-CoA ligase and ATP synthase 6); transcriptional and translation genes poly (A) binding protein, t-RNA-guanine transglycoslase, ribosomal protein L23, ribosomal protein S15A, eukaryotic translation initiation factor 3, Rab interaction protein 5, splicing factor-arginine serine rich 5, and ADP-ribosylation factor-like 6 interacting protein), oxide stress related genes (ferritin high chain and high-mobility group protein 2); protein activating and signaling pathway related genes (casein kinase, serine kinase 2 and phosphatidylinositol-four-phosphate adaptor protein-1-associated protein); cell differentiation and apoptosis associated genes (NB4 cell apoptosis related protein and myeloid differentiation primary response protein) and five genes with unknown function (KIAA0092, CGI-147protein, GCI-35, nucleolar phosphoprotein Nopp34 and Mus muscular partial mRNA for hypothetical protein), as well as a novel gene unmatched to the sequence in GenBank.
Conclusions:
The forward subtracted cDNA library contained differentially expressed genes from Jurkat T cell line induced by arsenic trioxide was successfully constructed. And, genes not involved in previous research on arsenic were found. Results of analysis for these genetic function suggested that there should be many genes involved in process of T lymphocytes apoptosis or injury induced by arsenic trioxide and that there should still be many genes associated with arsenic that were not reported in the past.
Insights
Arsenic trioxide exposure alters gene expression in human T lymphocytes, revealing new genes involved in apoptosis and immune suppression. This study identifies novel targets for understanding arsenic
Area of Science:
- Immunology
- Molecular Biology
- Toxicology
Background:
- Arsenic trioxide (As(2)O(3)) is known to affect immune cells.
- The specific genes and mechanisms underlying its immunotoxicity and immune suppression in T lymphocytes require further elucidation.
Purpose of the Study:
- To identify differentially expressed genes in human T lymphocytes induced by arsenic trioxide.
- To explore the mechanisms of arsenic trioxide-induced immunotoxicity and immune suppression.
Main Methods:
- Human Jurkat T cell line treated with arsenic trioxide (5 micromol/L, 24 h).
- Construction of a subtractive cDNA library using suppression subtractive hybridization (SSH).
- Identification of differentially expressed genes via Polymerase Chain Reaction (PCR) and sequencing.
Main Results:
- A subtractive cDNA library was successfully constructed, containing 29 unique gene fragments.
- Identified genes include those involved in oxidative metabolism, transcription/translation, oxidative stress, signaling pathways, and apoptosis.
- Novel genes with unknown functions and a gene not found in GenBank were discovered, suggesting new roles in arsenic toxicity.
Conclusions:
- The study successfully constructed a subtractive cDNA library of genes differentially expressed in T lymphocytes treated with arsenic trioxide.
- Several novel genes involved in arsenic-related processes, particularly T lymphocyte apoptosis and injury, were identified.
- These findings expand the understanding of arsenic's impact on the immune system and suggest previously unreported genes associated with arsenic toxicity.
