Time Dependent Gene Expression Analysis of Vitamin D3 (1,25 (OH)2 D3) Treated HL–60 Cells by Q-RT-PCR


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Kanlı A. , Gülkaç M. D. , Savlı H.

Kocaeli Tıp Dergisi / Kocaeli Medical Journal, vol.9, no.1, pp.60-67, 2020 (National Refreed University Journal)

  • Publication Type: Article / Article
  • Volume: 9 Issue: 1
  • Publication Date: 2020
  • Title of Journal : Kocaeli Tıp Dergisi / Kocaeli Medical Journal
  • Page Numbers: pp.60-67

Abstract

ABSTRACT
INTRODUCTION: The treatment of Acute myelogenous leukemia (AML) with differential effect of 1alpha,25-dihydroxyvitamin D(3) (1,25(OH)2D3) has been one of the popular research topics. The aim of this study is to investigate the role of 1.25 (OH) 2D3 in leukemia cell cycle regulation and apoptosis in HL-60 cells.
METHODS: We analyzed 13 genes (TNFR1, Bcl-w, Bax, Bak, Caspase–6, Caspase–8, AIF, Survivin, Cdk1 (Cdc2), Cdk2, Cdk4, Cyclin-D1 and Cyclin-E) for changes in expression associated with the cell cycle and the apoptosis of human promyelocytic leukemia HL-60 cells induced by 1,25(OH)2D3 at 18th, 36th, 48th and 72nd h, using quantitative real-time PCR.
RESULTS: We did not find significantly down or up-regulated expression profiles at mentioned time points. In our experiments, we observed decreased expression of TNFR1, Cdk-4, Cyclin-D1, Cyclin-E and Survivin genes at 72nd hours, and increased expression of Caspase-8 and Bak genes. The tendency of cell cycle related genes decrease from 18th to 72nd hours suggested that HL60 cells differentiated. Decreased expression of anti-apoptotic Bcl-w and Survivin genes and increased expression of pro-apoptotic Bak and Caspase-8 genes suggested that apoptotic process stage starts around 72nd hours.
DISCUSSION AND CONCLUSION: In conclusion, exposure of leukemic HL60 cell line to vitamin D3 derived the cells into differentiation. The clonal development completed and apoptotic stage begun.
Keywords: HL–60 cells, 1, 25 (OH)2 D3, Cell Cycle, Differentiation, Apoptosis, Quantitative Real Time Polymerase Chain Reaction (Q-RT-PCR)