精子相关抗原6基因以非P53依赖方式促进TRAIL诱导的骨髓增生异常综合征细胞凋亡

朱春霞; 王利; 罗小华; 刘林

doi:10.16016/j.1000-5404.201709167

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精子相关抗原6基因以非P53依赖方式促进TRAIL诱导的骨髓增生异常综合征细胞凋亡

基础医学 | 更新时间：2026-01-09

- 精子相关抗原6基因以非P53依赖方式促进TRAIL诱导的骨髓增生异常综合征细胞凋亡
- Sperm-associated antigen 6, independent on P53, regulates cell apoptosis of myelodysplastic syndromes induced by TRAIL
- 第三军医大学学报 2018年40卷第3期页码：198-204
- 作者机构：
  
  400016 重庆，重庆医科大学附属第一医院血液内科
- 作者简介：
  
  [ 刘林，电话：(023)89011532，E-mail:liul7776@aliyun.com LIU Lin，Tel: 86-23-89011532，E-mail:liul7776@aliyun.com
- 基金信息：
  
  国家自然科学基金面上项目(81570109)
- DOI：10.16016/j.1000-5404.201709167
  中图分类号： R394-33;R394.3;R551.3
- 收稿：2017-09-21，
  
  修回：2017-11-06，
  
  纸质出版：2018-02-15
- 稿件说明：
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朱春霞, 王利, 罗小华, 等. 精子相关抗原6基因以非P53依赖方式促进TRAIL诱导的骨髓增生异常综合征细胞凋亡[J]. 第三军医大学学报, 2018,40(3):198-204.

Chunxia ZHU, Li WANG, Xiaohua LUO, et al. Sperm-associated antigen 6, independent on P53, regulates cell apoptosis of myelodysplastic syndromes induced by TRAIL[J]. Journal of Third Military Medical University, 2018, 40(3): 198-204.
朱春霞, 王利, 罗小华, 等. 精子相关抗原6基因以非P53依赖方式促进TRAIL诱导的骨髓增生异常综合征细胞凋亡[J]. 第三军医大学学报, 2018,40(3):198-204. DOI： 10.16016/j.1000-5404.201709167.

Chunxia ZHU, Li WANG, Xiaohua LUO, et al. Sperm-associated antigen 6, independent on P53, regulates cell apoptosis of myelodysplastic syndromes induced by TRAIL[J]. Journal of Third Military Medical University, 2018, 40(3): 198-204. DOI： 10.16016/j.1000-5404.201709167.

摘要

目的

研究精子相关抗原6(sperm-associated antigen 6，SPAG6)基因是否以P53依赖方式促进TRAIL诱导的骨髓增生异常综合征(myelodysplastic syndrome，MDS)细胞凋亡。

方法

通过P53

RNA慢病毒转染人结肠癌细胞株HCT116筛选P53基因沉默的最佳靶点，利用筛选出来的最佳靶点序列转染SKM-1细胞；加入嘌呤霉素提高转染效率，RT-PCR及Western blot检验P53基因的干扰效果，构建SKM-1 P53

细胞模型。用SPAG6

RNA慢病毒载体转染SKM-1和SKM-1 P53

细胞，RT-PCR及Western blot验证。用CCK-8法检测重组人可溶性TRAIL蛋白(rTRAIL)对SKM-1细胞的适宜作用浓度。Western blot检测rTRAIL处理后SKM-1和SKM-1 P53

细胞株中Caspase-8、cleaved-PARP、PARP的表达水平。

结果

构建了SKM-1 P53

细胞模型，转染率在90%以上，P53基因在mRNA和蛋白表达水平较对照组均显著降低(

0.01)。SPAG6

RNA慢病毒载体成功感染SKM-1细胞和SKM-1 P53

细胞，转染率均在80%以上，SPAG6基因表达较对照组明显降低(

0.01)。随着rTRAIL浓度的增加，细胞生长逐渐被抑制，30 ng/mL TRAIL诱导的细胞凋亡与对照组差异没有统计学意义，100 ng/mL和300 ng/mL TRAIL显著诱导SPAG6

细胞凋亡(

0.05)，而P53基因沉默与否并没有影响该结果，rTRAIL处理后细胞凋亡标志分子明显激活或水平升高，细胞中Caspase-8、cleaved-PARP、PARP的表达水平在SKM-1和SKM-1 P53

细胞中差异无统计学意义(

0.05)。

结论

SPAG6基因以非P53依赖方式促进TRAIL诱导的MDS细胞凋亡。

Abstract

Objective

To investigate the sperm-associated antigen 6 (SPAG6) regulating cell apoptosis of myelodysplastic syndromes (MDS) induced by the TRAIL whether is dependent on P53.

Methods

The P53-knoc

k down RNA (P53

RNA) lentiviral vector was transfected into HCT116 cells to obtain the most effective shRNA-containing plasmid. To construct SKM-1 P53

cells model

the most effective interference sequence was transferred into SKM-1 cells

with the transfection efficiency being increased by purinomycin. Then the interference efficiency was determined by quantitative reverse transcription PCR (RT-PCR) and Western blotting. Next

the SPAG6

RNA-expressing lentiviral vector was transferred into SKM-1 cells and SKM-1 P53

cells

and the interference efficiency was verified by RT-PCR and Western blotting. Cell apoptosis was measured after they were treated with recombinant human sTRAIL (rTRAIL) of different concentrations by CCK-8 assay. The expression levels of PARP

cleaved PARP and Caspase-8 were measured by Western blotting after the cells were treated with rTRAIL.

Results

The SKM-1 P53

cell model was successfully constructed

with the transfection rate over 90%

and both RT-PCR and Western blotting indicated the significant decrease of P53 expression (

0.01). The SPAG6

RNA lentiviral vector was also successfully infected the SKM-1 cells and SKM-1 P53

cells

and the transfection rate was more than 80%. The expression of the SPAG6 was decreased significantly after the transfection (

0.01). With the concentration of TRAIL was increased

cell growth was inhibited gradually. When the concentration was 30 ng/mL

there was no difference in cell apoptosis between the treated cells and the control cells

but 100 and 300 ng/mL obviously induced cell apoptosis (

0.05)

and P53 knockdown showed no effect on the results. rTRAIL treatment increased the expression of apoptotic molecules and enhanced cell apoptosis

but there were no significant differences in the expression levels of PARP

cleaved PARP and Caspase-8 between the SKM-1 cells and

SKM-1 P53

cells (

0.05).

Conclusion

SPAG6 is not depend on P53 in regulating cell apoptosis of MDS induced by TRAIL.

关键词

Keywords

references

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