Department of Ophthalmology, 4th Affiliated Hospital of Harbin Medical University, Harbin, Heilongjiang Province, China.
Department of Cardiovascular Surgery, 2nd Affiliated Hospital of Harbin Medical University, Harbin, Heilongjiang Province, China.
Curr Eye Res. 2022 Apr;47(4):624-633. doi: 10.1080/02713683.2021.2007535. Epub 2021 Dec 29.
Circular RNAs (circRNAs) play essential roles in the progression of human tumors, including retinoblastoma (RB). In this study, we aimed to explore the functions and potential mechanisms of circ_0000527 in RB.
Quantitative real-time polymerase chain reaction (qRT-PCR), Western blot assay and immunohistochemistry (IHC) assay were conducted to determine the levels of circ_0000527, microRNA-1236-3p (miR-1236-3p) and SMAD family member 2 (SMAD2). RNase R assay and actinomycin D assay were conducted to analyze the characteristic of circ_0000527. Cell Counting Kit-8 (CCK-8) assay, 5-ethynyl-2'-deoxyuridine (EdU) assay, and colony formation assay were performed for cell proliferation ability. Wound healing assay and transwell assay were applied to assess cell migration and invasion. Tube formation assay was utilized for angiogenesis ability. Flow cytometry analysis was adopted to analyze cell apoptosis. Dual-luciferase reporter assay and RNA immunoprecipitation (RIP) assay were performed to analyze the relationships among circ_0000527, miR-1236-3p, and SMAD2. Murine xenograft model assay was conducted for the role of circ_0000527 .
Circ_0000527 was overexpressed in RB patients and related to advanced TNM stages, optic nerve invasion and choroidal invasion. Circ_0000527 knockdown suppressed cell proliferation, migration, invasion and angiogenesis and promoted apoptosis in RB cells . Circ_0000527 sponged miR-1236-3p, which directly targeted SMAD2. MiR-1236-3p level was decreased in RB tissues and cells. MiR-1236-3p inhibition reversed circ_0000527 knockdown-mediated effects on RB cell malignant behaviors. Moreover, miR-1236-3p overexpression suppressed RB cell progression, with SMAD2 elevation abrogated the effect. Additionally, circ_0000527 knockdown restrained tumor formation .
Circ_0000527/miR-1236-3p/SMAD2 axis played a positive role in the progression of RB.
环状 RNA（circRNAs）在人类肿瘤的进展中发挥着重要作用，包括视网膜母细胞瘤（RB）。在这项研究中，我们旨在探讨 circ_0000527 在 RB 中的功能和潜在机制。
采用实时定量聚合酶链反应（qRT-PCR）、Western blot 检测和免疫组织化学（IHC）检测来测定 circ_0000527、微小 RNA-1236-3p（miR-1236-3p）和 SMAD 家族成员 2（SMAD2）的水平。采用 RNase R 试验和放线菌素 D 试验分析 circ_0000527 的特征。采用细胞计数试剂盒-8（CCK-8）试验、5-乙炔基-2'-脱氧尿苷（EdU）试验和集落形成试验评估细胞增殖能力。进行划痕愈合试验和 Transwell 试验以评估细胞迁移和侵袭能力。进行管形成试验以评估血管生成能力。采用流式细胞术分析评估细胞凋亡。采用双荧光素酶报告基因试验和 RNA 免疫沉淀（RIP）试验分析 circ_0000527、miR-1236-3p 和 SMAD2 之间的关系。进行小鼠异种移植模型试验以研究 circ_0000527 的作用。
circ_0000527 在 RB 患者中表达上调，与晚期 TNM 分期、视神经侵犯和脉络膜侵犯有关。circ_0000527 敲低抑制 RB 细胞的增殖、迁移、侵袭和血管生成，并促进细胞凋亡。circ_0000527 吸附 miR-1236-3p，后者可直接靶向 SMAD2。miR-1236-3p 在 RB 组织和细胞中下调。miR-1236-3p 抑制逆转了 circ_0000527 敲低对 RB 细胞恶性行为的影响。此外，miR-1236-3p 过表达抑制 RB 细胞进展，而 SMAD2 升高则可消除这种作用。此外，circ_0000527 敲低抑制肿瘤形成。
circ_0000527/miR-1236-3p/SMAD2 轴在 RB 的进展中发挥着正性作用。
