Table8_APOC1 exacerbates renal fibrosis through the activation of the NF-κB signaling pathway in IgAN.DOCX

Introduction: IgA nephropathy (IgAN) is the most common disease leading to end-stage renal disease, and tubular fibrosis represents an important risk factor for disease progression. However, research on early molecular diagnostic indicators of tubular fibrosis and the mechanisms underlying disease progression is still lacking.
Methods: The GSE93798 dataset was downloaded from the GEO database. DEGs were screened and analyzed for GO and KEGG enrichment in IgAN. The least absolute shrinkage and selection operator (LASSO) and support vector machine recursive feature elimination (SVM-RFE) algorithms were applied to screen for hub secretory genes. The expression and diagnostic efficacy of hub genes were confirmed by the GSE35487 dataset. ELISA was applied to detect the expression of APOC1 in serum. The expression and localization of hub genes in IgAN were verified by the expression of IHC and IF in human kidney tissues, and the correlation of expression with clinical data was verified in the Nephroseq database. Finally, cellular experiments clarified the role of hub genes in the signaling pathway.
Results: A total of 339 DEGs were identified in IgAN, of which 237 were upregulated and 102 downregulated. The KEGG signaling pathway is enriched in the ECM–receptor interaction and AGE-RAGE signaling pathway. APOC1, ALB, CCL8, CXCL2, SRPX2, and TGFBI identified six hub secretory genes using the LASSO and SVM-RFE algorithms. In vivo and in vitro experiments demonstrated that APOC1 expression was elevated in IgAN. The serum concentration of APOC1 was 1.232 ± 0.1812 μg/ml in IgAN patients, whereas it was 0.3956 ± 0.1233 μg/ml in healthy individuals. APOC1 exhibited high diagnostic efficacy for IgAN (AUC of 99.091%, specificity of 95.455%, and sensitivity of 99.141%) in the GSE93798 dataset. APOC1 expression negatively correlated with eGFR (R2 = 0.2285, p = 0.0385) and positively correlated with serum creatinine (R2 = 0.41, p = 0.000567) in IgAN. APOC1 exacerbated renal fibrosis, possibly in part by activating the NF-κB pathway in IgAN.
Conclusion: APOC1 was identified as the core secretory gene of IgAN, which was closely associated with blood creatinine and eGFR and had significant efficacy in the diagnosis of IgAN. Mechanistic studies revealed that the knockdown of APOC1 could improve IgAN renal fibrosis by inhibiting the NF pathway, which may be a potential therapeutic target for improving renal fibrosis in IgAN.

引言：IgA肾病（IgA nephropathy, IgAN）是引发终末期肾病的最常见疾病，而肾小管纤维化是疾病进展的重要危险因素。然而，目前针对肾小管纤维化的早期分子诊断指标以及疾病进展机制的研究仍较为匮乏。
方法：从GEO数据库下载GSE93798数据集。对IgAN中的差异表达基因（differentially expressed genes, DEGs）进行筛选，并开展基因本体（Gene Ontology, GO）富集分析与京都基因与基因组百科全书（Kyoto Encyclopedia of Genes and Genomes, KEGG）富集分析。采用最小绝对收缩和选择算子（least absolute shrinkage and selection operator, LASSO）以及支持向量机递归特征消除（support vector machine recursive feature elimination, SVM-RFE）算法筛选核心分泌基因。通过GSE35487数据集验证核心基因的表达水平与诊断效能。采用酶联免疫吸附测定（enzyme-linked immunosorbent assay, ELISA）检测血清中APOC1的表达水平。通过人体肾脏组织的免疫组化（immunohistochemistry, IHC）与免疫荧光（immunofluorescence, IF）实验验证核心基因在IgAN中的表达与定位，并在Nephroseq数据库中验证基因表达与临床数据的相关性。最后，通过细胞实验阐明核心基因在信号通路中的作用。
结果：本研究共在IgAN中筛选得到339个差异表达基因，其中237个为上调基因，102个为下调基因。KEGG信号通路富集分析显示，差异基因主要富集于细胞外基质-受体相互作用（ECM-receptor interaction）与晚期糖基化终末产物-晚期糖基化终末产物受体（AGE-RAGE）信号通路。通过LASSO与SVM-RFE算法，共筛选得到APOC1、ALB、CCL8、CXCL2、SRPX2及TGFBI共6个核心分泌基因。体内与体外实验结果表明，APOC1在IgAN中的表达水平显著升高。IgAN患者血清中APOC1的浓度为1.232 ± 0.1812 μg/ml，而健康个体血清中APOC1浓度为0.3956 ± 0.1233 μg/ml。在GSE93798数据集中，APOC1对IgAN具有较高的诊断效能（曲线下面积area under the curve, AUC为99.091%，特异性为95.455%，灵敏度为99.141%）。在IgAN患者中，APOC1的表达水平与估算肾小球滤过率（estimated glomerular filtration rate, eGFR）呈负相关（R²=0.2285，p=0.0385），与血清肌酐水平呈正相关（R²=0.41，p=0.000567）。APOC1可能通过激活核因子κB（nuclear factor kappa-B, NF-κB）通路加剧IgAN患者的肾纤维化。
结论：本研究鉴定APOC1为IgAN的核心分泌基因，其与血清肌酐水平及eGFR密切相关，且对IgAN具有显著的诊断效能。机制研究显示，敲低APOC1可通过抑制核因子κB（NF-κB）通路改善IgAN患者的肾纤维化，这表明APOC1或可作为改善IgAN肾纤维化的潜在治疗靶点。