Abstract：
BACKGROUND: Focal segmental glomerulosclerosis (FSGS) is a common glomerulopathy with an unclear mechanism. The demand for FSGS clinical diagnostic biomarkers has not yet been met. Circular RNA (circRNA) is a novel non-coding RNA with multiple functions, but its diagnostic value for FSGS remains unexplored. This study aimed to identify circRNAs that could aid in early clinical diagnosis and to investigate their mechanisms in podocyte injury.
METHODS: The signature of plasma circRNAs for FSGS was identified by circRNA microarray. The existence of circRNAs was confirmed by quantitative real-time polymerase chain reaction (qRT-PCR), RNase R assay, and DNA sequencing. Plasma levels of circRNAs were evaluated by qRT-PCR. The diagnostic value was appraised by the receiver operating characteristic curve. The circRNA-miRNA-mRNA network was built with Cytoscape 7.3.2. Statistically significant differences were calculated by the Mann-Whitney U test.
RESULTS: A total of 493 circRNAs (165 upregulated, 328 downregulated) were differentially expressed in the plasma of FSGS patients (n = 3) and normal controls (n = 3). Eight candidate circRNAs were demonstrated to be circular and stable transcripts. Among them, hsa_circ_0001230 and hsa_circ_0023879 were significantly upregulated in FSGS patients (n = 29) compared to normal controls (n = 51). The areas under the curve value of hsa_circ_0001230 and hsa_circ_0023879 were 0.668 and 0.753, respectively, while that of the two-circRNA panel was 0.763. The RNA pull-down analysis revealed that hsa_circ_0001230 and hsa_circ_0023879 could sponge hsa-miR-106a. Additionally, hsa_circ_0001230 and hsa_circ_0023879 positively regulated hsa-miR-106a target genes phosphatase and tensin homolog (PTEN) and Bcl-2-like protein 11 (BCL2L11) in podocytes.
CONCLUSIONS: hsa_circ_0001230 and hsa_circ_0023879 are novel blood biomarkers for FSGS. They may regulate podocyte apoptosis by competitively binding to hsa-miR-106a.
METHODS: The signature of plasma circRNAs for FSGS was identified by circRNA microarray. The existence of circRNAs was confirmed by quantitative real-time polymerase chain reaction (qRT-PCR), RNase R assay, and DNA sequencing. Plasma levels of circRNAs were evaluated by qRT-PCR. The diagnostic value was appraised by the receiver operating characteristic curve. The circRNA-miRNA-mRNA network was built with Cytoscape 7.3.2. Statistically significant differences were calculated by the Mann-Whitney U test.
RESULTS: A total of 493 circRNAs (165 upregulated, 328 downregulated) were differentially expressed in the plasma of FSGS patients (n = 3) and normal controls (n = 3). Eight candidate circRNAs were demonstrated to be circular and stable transcripts. Among them, hsa_circ_0001230 and hsa_circ_0023879 were significantly upregulated in FSGS patients (n = 29) compared to normal controls (n = 51). The areas under the curve value of hsa_circ_0001230 and hsa_circ_0023879 were 0.668 and 0.753, respectively, while that of the two-circRNA panel was 0.763. The RNA pull-down analysis revealed that hsa_circ_0001230 and hsa_circ_0023879 could sponge hsa-miR-106a. Additionally, hsa_circ_0001230 and hsa_circ_0023879 positively regulated hsa-miR-106a target genes phosphatase and tensin homolog (PTEN) and Bcl-2-like protein 11 (BCL2L11) in podocytes.
CONCLUSIONS: hsa_circ_0001230 and hsa_circ_0023879 are novel blood biomarkers for FSGS. They may regulate podocyte apoptosis by competitively binding to hsa-miR-106a.
摘要：
简介局灶节段肾小球硬化（FSGS）是一种常见的肾小球病，其机制尚不清楚。对FSGS临床诊断生物标志物的需求尚未得到满足。环状RNA(circularRNA,circRNA)是一种具有多种功能的新型非编码RNA。但其对FSGS的诊断价值仍有待探索。本研究旨在鉴定有助于早期临床诊断的circRNAs，并研究其在足细胞损伤中的作用机制。方法通过circRNA微阵列鉴定FSGS的血浆circRNAs的特征。通过qRT-PCR证实了circRNAs的存在，RNaseR测定,和DNA测序。通过qRT-PCR评估circRNA的血浆水平。通过受试者工作特征曲线评估诊断价值。使用Cytoscape7.3.2构建circRNA-miRNA-mRNA网络。通过Mann-WhitneyU检验计算统计学上的显著差异。结果共493个circRNAs(165个上调，328下调）在FSGS患者（n=3）和正常对照（n=3）的血浆中差异表达。八个候选circRNAs被证明是环状和稳定的转录物。其中,与正常对照组(n=51)相比，FSGS患者(n=29)的hsa_circ_0001230和hsa_circ_0023879显著上调。hsa_circ_0001230和hsa_circ_0023879的曲线下面积值分别为0.668和0.753,而两个circRNAs小组的结果是0.763。RNA下拉分析表明，hsa_circ_0001230和hsa_circ_0023879可以海绵hsa-miR-106a。此外,hsa_circ_0001230和hsa_circ_0023879正调控足细胞中的hsa-miR-106a靶基因磷酸酶和张力蛋白同源物(PTEN)和Bcl-2样蛋白11(BCL2L11)。结论Hsa_circ_0001230和hsa_circ_0023879是FSGS的新型血液生物标志物。它们可以通过竞争性结合hsa-miR-106a来调节足细胞凋亡。
