Background

Spinal cord injury (SCI) is one of the serious neurological diseases with high morbidity which may be treated with hematopoietic stem cell (HSC) transplants. Circular RNAs (circRNAs) play vital roles in SCI. The study aimed to reveal the function and mechanism of circRNA homeodomain interacting protein kinase 3 (HIPK3) in SCI.

Methods

SCI model in vitro was established by treating neuronal cells AGE1.HN with oxygen-glucose deprivation (OGD) and CoCl2. The levels of circHIPK3, miR-382-5p and dual specificity phosphatase 1 (DUSP1) were examined using quantitative real-time PCR (qRT-PCR) or western blot assay. Enzyme linked immunosorbent assay (ELISA) was used to detect the levels of inflammatory factors (IL-6 and TNF-α). Cell proliferation and apoptosis were evaluated by 5′-ethynyl-2′-deoxyuridine (EdU) assay and flow cytometry. Caspase-3 Colorimetric Assay Kit was used to detect aaspase-3 activity. The interactions among circHIPK3, miR-382-5p and DUSP1 were confirmed by dual-luciferase reporter and RNA immunoprecipitation assays.

Results

CircHIPK3 and DUSP1 were down-regulated, while miR-382-5p was up-regulated in OGD-induced AGE1.HN cells. Overexpression of circHIPK3 suppressed inflammatory response and cell apoptosis and promoted proliferation in OGD-induced AGE1.HN cells by sponging miR-382-5p. CircHIPK3 regulated DUSP1 expression by targeting miR-382-5p. MiR-382-5p inhibition hindered inflammatory response of IL-6 and TNF-α and neuronal apoptosis and promoted apoptosis via targeting DUSP1.

Conclusion

CircHIPK3 overexpression alleviated OGD-induced AGE1.HN cell inflammatory response and neuronal apoptosis via regulating miR-382-5p/DUSP1 axis, indicating that circHIPK3 might be a promising therapeutic target for SCI.

中文翻译：

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CircHIPK3 通过调节 miR-382-5p/DUSP1 轴减轻脊髓损伤中的炎症反应和神经元凋亡

背景

脊髓损伤（SCI）是一种高发病率的严重神经系统疾病，可以通过造血干细胞（HSC）移植治疗。环状 RNA (circRNA) 在 SCI 中起着至关重要的作用。该研究旨在揭示circRNA同源域相互作用蛋白激酶3（HIPK3）在SCI中的功能和机制。

方法

通过用氧-葡萄糖剥夺（OGD）和CoCl2处理神经元细胞AGE1.HN建立体外SCI模型。使用定量实时 PCR (qRT-PCR) 或蛋白质印迹法检测 circHIPK3、miR-382-5p 和双特异性磷酸酶 1 (DUSP1) 的水平。酶联免疫吸附试验（ELISA）用于检测炎症因子（IL-6和TNF-α）的水平。通过5'-乙炔基-2'-脱氧尿苷（EdU）测定和流式细胞术评估细胞增殖和凋亡。Caspase-3 Colorimetric Assay Kit 用于检测 aaspase-3 活性。circHIPK3、miR-382-5p 和 DUSP1 之间的相互作用通过双荧光素酶报告基因和 RNA 免疫沉淀测定得到证实。

结果

CircHIPK3 和 DUSP1 下调，而 miR-382-5p 在 OGD 诱导的 AGE1.HN 细胞中上调。circHIPK3 的过表达抑制炎症反应和细胞凋亡，并通过海绵化 miR-382-5p 促进 OGD 诱导的 AGE1.HN 细胞的增殖。CircHIPK3 通过靶向 miR-382-5p 调节 DUSP1 的表达。MiR-382-5p 抑制抑制 IL-6 和 TNF-α 的炎症反应和神经元凋亡，并通过靶向 DUSP1 促进细胞凋亡。

结论

CircHIPK3 过表达通过调节 miR-382-5p/DUSP1 轴减轻 OGD 诱导的 AGE1.HN 细胞炎症反应和神经元凋亡，表明 circHIPK3 可能是 SCI 的一个有希望的治疗靶点。