TET1基因对小鼠uNK细胞增殖及IFN-γ、VEGF-C和TGF-β1转录水平的影响

doi:10.11843/j.issn.0366-6964.2023.03.033

Abstract

Abstract: The purpose of this study was to evaluate the effect of ten eleven translocation 1 (TET1) on the proliferation and cytokine secretion and expression of mouse uterine natural killer (uNK) cells. The endometrium of pregnant mice was collected on gestational day 9-12, and lymphocytes were isolated. uNK cells were further sorted by streptavidin magnetic bead method for culture. The RNA interference sequence of TET1 gene was synthesized and connected to RNA interference vector GM-19167 to construct interference expression plasmid, then lentivirus was packaged and virus titer was determined. uNK cells were transfected according to the ratio of lentivirus to cell number of (10-200)∶1. After 168 h, the transfection effect was detected by fluorescence microscope. uNK cells were collected and fluorescent quantitative PCR (qPCR) method was used to verify the interference effect of TET1 gene while β- actin was used as an internal reference. Cell counting kit-8 (CCK8) was used to detect the proliferation of uNK cells. Primers of interferon-γ (IFN-γ), vascular endothelial growth factor (VEGF-C) and transforming growth factor β1 (TGF-β1) were synthesized respectively and were used to detect the expression of these three cytokines by qPCR method. The results showed that the expression of TET1 in uNK cells decreased significantly after 168 h of lentivirus infection (P<0.05), indicating that the expression of TET1 gene was significantly down regulated under RNA interference. When compared with the blank control groups, the uNK cells proliferation was not affected after TET1 interference (P>0.05) which shown by CCK8 detection results. Fluorescent qPCR results showed that after TET1 interference, the expression level of TGF-β1 was increased significantly (P<0.01), while IFN-γand VEGF-C were both decreased significantly (P<0.05). These findings revealed that interference with down-regulation of TET1 gene expression did not affect the proliferation of uNK cells, but had an effect on the transcriptional level of cytokines, thereby playing an important role in the innate immunity of animals in the uterus.

Key words: TET1, uNK cell, cellular proliferation, cytokine, mouse

CLC Number:

S857.2

YANG Xiaowei, ZHAO Ziliang, FU Yu, YU Zixiao, ZHAO Yongju. Effects of TET1 Gene on the Proliferation of Mouse uNK Cells and the Transcriptional Level of IFN-γ, VEGF-C and TGF-β1[J]. Acta Veterinaria et Zootechnica Sinica, 2023, 54(3): 1221-1228.

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Effects of TET1 Gene on the Proliferation of Mouse uNK Cells and the Transcriptional Level of IFN-γ, VEGF-C and TGF-β1

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