J Korean Surg Soc.  2009 Oct;77(4):225-237. 10.4174/jkss.2009.77.4.225.

Apoptosis Gene Expression Pattern Analysis of Jurkat Cells Treated with FK506

Affiliations
  • 1Department of Surgery, Hanil Hospital, Seoul, Korea.
  • 2Korea Food & Drug Administration, Seoul, Korea.
  • 3Department of Surgery, Chonnam National University Medical School, Gwangju, Korea. sycpvts@jnu.ac.kr

Abstract

PURPOSE
FK506 (tacrolimus) is a widely used immunosuppressive agent in the treatment of various medical conditions, including autoimmune disease, bone marrow and organ transplantations. Previously FK506 was known to cause apoptotic death of human Jurkat T cells. METHODS: The current study was designed to analyze the gene expression pattern of Jurkat T cells after FK506 application by using cDNA microarray. Treatment of Jurkat T cells with FK506 resulted in a decrease of cell viability in a time- and dose-dependent manner. Next, total RNA of Jurkat T cells was extracted by using TRIzol reagent and used to carry out a confirmation test for the purity and integrity of total RNA. RESULTS: Gene expression levels related to apoptosis and cell cycle process were mainly focused to analyze in FK506-treated Jurkat T cells. According to the inhibition of calcineurin activity, MARCKS in PKC substrates and Sp3 transcription factor was markedly increased in FK506-treated cells. Also, cell cycle control gene Id1 and Id3 were induced in expression from FK506-treated Jurkat T cells. However, FK506 decreased the expression of Src homology 2, G protein, and MEK 2 genes in bioactive peptide induced signaling pathway. It also reduced the expression level of the insulin receptor, DRPLA and Bai1-associated protein 2 genes, which are involved in the regulation of cell motility and morphology control. CONCLUSION: The author will continue to pursue the exact functional roles of genes that are markedly changed in expression by FK506 in human Jurkat T cells in vitro and in vivo experimental models.

Keyword

Tacrolimus; Gene expression; Apoptosis; Jurkat cell

MeSH Terms

Apoptosis
Autoimmune Diseases
Bone Marrow
Calcineurin
Cell Cycle
Cell Cycle Checkpoints
Cell Movement
Cell Survival
Gene Expression
Gene Expression Profiling
GTP-Binding Proteins
Guanidines
Humans
Jurkat Cells
Models, Theoretical
Oligonucleotide Array Sequence Analysis
Organ Transplantation
Phenols
Receptor, Insulin
RNA
Sp3 Transcription Factor
T-Lymphocytes
Tacrolimus
Transplants
Calcineurin
GTP-Binding Proteins
Guanidines
Phenols
RNA
Receptor, Insulin
Sp3 Transcription Factor
Tacrolimus

Figure

  • Fig. 1 FK506 decreased the viability of Jurkat cells in dose- and time-dependent manners. Cells were treated with various concentration of FK506 for 96 hours, then cell viability was measured by MTT assay after FK506 treatment. Data represent the mean±standard deviation (SD) of triplicates.

  • Fig. 2 Modulation of gene expression following FK506-treated Jurkat cells. Total RNA was prepared from Jurkat using TRIzol as described by manufacturer. (A) Scatter plot analysis. The x and y axes represent Cy3 (Control) and Cy5 (FK506 10µM or 20µM) signal intensity values, respectively. These represent up-regulated (upper lines; red spots) or down-regulated (under lines; green spots) genes. (B) MA plot analysis. The x and y axes represent A (R+G/2) and M (R/G) signal intensity values. The R and G repesent Cy5-background and Cy3-background signal intensity, respectively. These represent up-regulated (upper lines; red spots) or down-regulated (under lines; green spots) genes.

  • Fig. 3 Cluster image showing the different classes of total gene expression profiles.


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