hsa_circ_0005389参与新生儿急性呼吸窘迫综合征炎症过程的实验研究
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1.南京医科大学第一附属医院儿科;2.空军军医大学附属唐都医院

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国家自然科学(编号:81871195)


Experimental study on the involvement of hsa_circ_0005389 in the inflammatory process of neonatal acute respiratory distress syndrome
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Affiliation:

1.the First Affiliated Hospital of Nanjing Medical University, Department of Pediatrics;2.the First Affiliated Hospital of Nanjing Medical University,Department of Pediatrics,

Fund Project:

the National Natural Science Foundation of China ([grant number.81871195])

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    摘要:

    目的:在人类circRNA高通量芯片测序及相关生物信息学分析的基础上研究hsa_circ_0005389与新生儿ARDS之间的联系。 方法:建立脂多糖诱导急性肺损伤细胞模型。通过 qRT-PCR 及 Western Blot 检测炎性标记物及相关通路指标(IL-6、IL-8、sTNFR1、TNFα、PI3K、Akt)在空白组、阴性对照组和敲低 hsa_circ_0005389 表达的干预组中的表达变化情况,同时用CCK8 与 V-APC/7-AAD 检测敲低 hsa_circ_0005389表达后的空白A549 细胞与建立急性肺损伤模型的 A549 细胞的增殖与凋亡。 结果:qPCR 检测及 Western Blot 结果显示 A549 细胞暴露 10 μg/cm2 LPS 48 h 时,IL-6、IL-8、sTNFR1、TNFα、PI3K、Akt 等炎性标记物及相关通路指标高表达(P<0.05)。与空白组及阴性对照组相比,敲低hsa_circ_0005389表达,sTNFR1 mRNA 相对表达量和 TNFα 蛋白表达量显著降低;在急性肺损伤模型中,各炎性标记物及相关通路指标 mRNA 相对表达量均下降(P<0.0001)。CCK8 结果显示,与阴性对照组相比,培养 48 h、72 h 时敲低 hsa_circ_0005389 表达的A549 细胞显著增殖( P<0.05 )。同时与阴性对 照组的凋亡率相比,敲低hsa_circ_0005389 表达的 A549 细胞凋亡率显著降低(P<0.05)。 结论:急性肺损伤细胞模型中,hsa_circ_0005389 促进了肺损伤炎性标记物的表达,如 sTNFR1、TNFα等;敲低 hsa_circ_0005389 表达促进了肺上皮细胞的增殖并抑制凋亡;hsa_circ_0005389 参与了新生儿 ARDS 的炎症过程,可能作为潜在的干预靶标。

    Abstract:

    Objective: To gain a new direction about distinguishing and treating neonatal ARDS, the relationship between hsa_circ_0005389 and neonatal ARDS, based on human circRNA high-throughput sequencing and the results of bioinformatics analysis, was studied. Methods: The acute lung injury model was induced by lipopolysaccharide. The expression of inflammatory markers were detected by qRT-PCR and Western blot in the A549 cell knocked down the expression of hsa_circ_0005389 and (or no) exposure LPS. And then the proliferation and apoptosis of A549 cells knocked down the expression of hsa_circ_0005389 were detected by CCK8 and flow cytometer. Result: The mRNA relative expression and the protein expression of all the research indexes increased significantly in the group of 10 μg/cm2 LPS(P<0.0001). Compare with the black and negative control (NC) group, it is decreased significantly that the mRNA relative expression of sTNFR1 and the protein expression of TNFα in A549 cells knocked down the expression of hsa_circ_0005389 (P<0.05). But all the research indexes decreased significantly (P<0.0001) in the group which knocked down the expression of hsa_circ_0005389 and then exposed LPS. In addition, the results of CCK8 showed that the proliferation of si-hsa_circ_0005389 group, compared with other groups, was different after 48 hours and 72 hours of culture (P<0.05). Compared with the apoptosis rate of the NC group, the apoptosis rate of the si-hsa_circ_0005389 group decreased significantly. Conclusion: In the acute lung injury model, hsa_circ_0005389 promotes the expression of inflammatory factors, such as TNFR1,TNFα and so on. At same time, it inhibits the proliferation of A549 and boosts the apoptosis. So hsa_circ_0005389 accelerates the inflammatory process of neonatal ARDS.

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  • 收稿日期:2022-12-31
  • 最后修改日期:2023-05-09
  • 录用日期:2023-07-13
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