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昼夜节律分子BMAL1通过调节免疫细胞毒性促进骨肉瘤的预后
Circadian rhythms molecule BMAL1 promotes the prognosis of osteosarcoma
via regulating immune cell cytotoxicity- 第三军医大学学报 2020年42卷第18期 页码：1787-1794
- 作者机构：
1.410000 长沙，中南大学湘雅三医院检验科
2.510062 广州，南部战区空军医院血管内科
3.410000 长沙，长沙血液中心
4.410007 长沙，湖南省儿童医院儿科医学研究所
- 作者简介：
[ 张树菊，E-mail:zhangshuju22@126.com ZHANG Shuju, E-mail:zhangshuju22@126.com
- 基金信息：国家自然科学基金青年科学基金(81900055);湖南省自然科学基金(2020JJ5283);湖南省卫健委基金(B2019021);湖南省儿童医院基金(2019ZSJ)
DOI：10.16016/j.1000-5404.202006005
中图分类号： R394.3; R730.23; R738.1收稿：2020-06-01，
修回：2020-08-12，
纸质出版：2020-09-30
- 稿件说明：
移动端阅览
袁金玲, 陈新建, 廖凯, 等. 昼夜节律分子BMAL1通过调节免疫细胞毒性促进骨肉瘤的预后[J]. 第三军医大学学报, 2020,42(18):1787-1794.
Jinling YUAN, Xinjian CHEN, Kai LIAO, et al. Circadian rhythms molecule BMAL1 promotes the prognosis of osteosarcoma
via regulating immune cell cytotoxicity[J]. Journal of Third Military Medical University, 2020, 42(18): 1787-1794.袁金玲, 陈新建, 廖凯, 等. 昼夜节律分子BMAL1通过调节免疫细胞毒性促进骨肉瘤的预后[J]. 第三军医大学学报, 2020,42(18):1787-1794. DOI： 10.16016/j.1000-5404.202006005.
Jinling YUAN, Xinjian CHEN, Kai LIAO, et al. Circadian rhythms molecule BMAL1 promotes the prognosis of osteosarcoma
via regulating immune cell cytotoxicity[J]. Journal of Third Military Medical University, 2020, 42(18): 1787-1794. DOI： 10.16016/j.1000-5404.202006005.
摘要
目的
2
探讨核心节律分子在骨肉瘤预后中的作用及可能机制。
方法
2
在骨肉瘤细胞中过表达节律基因BMAL1，qRT-PCR检测BMAL1基因的表达；克隆形成实验和Transwell实验检测细胞的克隆形成和侵袭能力；骨肉瘤细胞与外周血单个核淋巴细胞(peripheral blood mononuclear cell，PBMCs)体外共培养后，钙黄素染色法检测PBMCs对骨肉瘤细胞的直接杀伤作用，流式细胞术检测PBMCs中CD8
+
T和NK细胞脱颗粒(CD107a蛋白表达)的影响。
结果
2
BMAL1 mRNA高表达的骨肉瘤患者的生存时间明显高于低表达组(
P
<
0.000 1)。BMAL1过表达对U2OS细胞的侵袭和克隆形成无显著影响，但相较于对照组而言，过表达BMAL1的U2OS在体外与外周血单核血细胞(PBMCs)共培养4 h，其细胞死亡明显增加[PBMCs:U2OS细胞为5:1时，对照组和BMAL1组分别为(73.33±0.67)%和(81.00±0.58)%，
P
<
0.01；PBMCs:U2OS细胞为10:1时，对照组和BMAL1组分别为(76.67±0.33)%和(82.67±1.86)%，
P
<
0.01
]
。过表达BMAL1的骨肉瘤细胞组与PBMCs共培养后，T细胞和NK细胞CD107a表达差异无统计学意义(E:T=5:1，CD3
+
CD56
-
T细胞
P
=0.46; CD3
-
CD56
+
NK细胞
P
=0.96；E:T=10:1
CD3
+
CD56
-
T细胞
P
=0.93; CD3
-
CD56
+
NK细胞
P
=0.21)。
结论
2
骨肉瘤细胞BMAL1表达水平与骨肉瘤患者预后呈正相关，其机制主要是通过增加免疫细胞对其免疫监视作用，而非通过影响骨肉瘤细胞的克隆形成能力和侵袭。
Abstract
Objective
2
To investigate the role and possible mechanism of the core rhythm molecule BMAL1 in the prognosis of osteosarcoma (OS).
Methods
2
After BMAL1 was overexpressed in U2OS cells by Lipofectamine3000
the expression of BMAL1 was detected with qRT-PCR
and the clone formation and invasion ability of the overexpressed cells were observed with clonogenic assay and Transwell assay. Then
the BMAL1 overexpressed U2OS cells or the cells with control vector transfection were cocultured with peripheral blood mononuclear cells (PBMCs)
the direct killing effects of PBMCs on U2OS cells was observed with Calcein AM staining
and the CD107a expression by CD8
+
T and NK cells was measured with flow
cytometry.
Results
2
The OS patients with higher BMAL1 mRNA level had significantly longer survival time when compared with those with lower expression (
P
<
0.001). BMAL1 overexpression had no obvious effects on the invasion and clone formation of U2OS cells. However
cell death was significantly increased in BMAL1 overexpressed U2OS cells than the control vector transfected U2OS cells when co-cultured with PBMCs
in vitro
[PBMCs:U2OS cells=5:1
(81.00±0.58)%
vs
(73.33±0.67)%
P
<
0.01; PBMCs:U2OS cells=10:1
(82.67±1.86)%
vs
(76.67±0.33)%
P
<
0.01
]
. There was no significant difference in the expression level of CD107a between T cells and NK cells in the BMAL1 overexpressed U2OS cells than the control vector transfected U2OS cells when co-cultured with PBMCs (E:T=5:1
CD3
+
CD56
-
T cells
P
=0.46
CD3
-
CD56
+
NK cells
P
=0.96; E:T=10:1
CD3
+
CD56
-
T cells
P
=0.93; CD3
-
CD56
+
NK cells
P
=0.21).
Conclusion
2
The expression level of BMAL1 is positively correlated with the prognosis of OS patients
which might be mainly due to the enhanced immunological surveillance
but not by affecting the ability of invasion and clone formation of OS cells.
关键词
Keywords
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