Molecular mechanism study of acute kidney injury induced by aristolochic acid I
Aristolochic acid (AA) is naturally found in Aristolochiaceae plants, and has anti-tumor, anti infection, anti-inflammatory and other effects. Traditional Chinese medicine and patent medicine preparations containing AA are widely used in clinical practice, including Aristolochia, Fangji, Vermillion, Asarum and Shanci Mushroom of Asarum, as well as traditional Chinese patent medicines and simple preparations such as Compound Weitong Capsules, Chuanxiling Capsules, Shisanwei Shugan Capsules. Studies have also shown that exposure to AA is associated with a high risk of developing kidney disease and upper urinary tract cancer; As one of the main components of AA, aristolochic acid I (AAI) also has nephrotoxicity and carcinogenicity; Long term use of traditional Chinese medicine containing such substances can cause aristolochic acid nephropathy (AAN). There have been many studies on the nephrotoxicity of AAI, but there are not many reports that comprehensively characterize its toxic mechanism, mostly focusing on specific cells or signaling pathways. Therefore, further exploration of the toxicity mechanism of AAI is still needed. Transcriptome is the sum of all gene transcription products within a cell, and transcriptomics is the study of transcriptome, covering RNA related topics such as transcription, expression, and function. In order to gain a more comprehensive understanding of the mechanism of AAI nephrotoxicity, this study used the IIIumina high-throughput sequencing platform to perform transcriptomic sequencing of renal tissue, differential expression, and GO/KEGG enrichment analysis to clarify the molecular mechanism of AAI induced nephrotoxicity.

 

The toxic effects of AA were reported as early as the 1990s and subsequently classified as a first-class carcinogen by the International Agency for Research on Cancer of the World Health Organization. At present, China has banned the use of medicinal herbs with high AA content such as Guangfangji and Guanmutong, while relatively low content Aristolochiaceae plants and their preparations are still used in clinical treatment. AAI is the main component of AA, and the content of AAI varies among different Chinese medicinal materials. For example, the AAI content in Asarum is around 10 μ g/g, while the AAI content in cinnabar lotus ranges from 0.9386 to 3.5675mg/g. When cinnabar lotus is clinically administered with 5-10g, the AAI amount reaches between 9.386-35.675mg. This article refers to the research of others. After being administered with AAI20mg/kg for 5 days, mice showed significant kidney damage: blood creatinine and urea nitrogen both increased significantly; Glomerular edema, glomerular consolidation, and detachment of proximal tubular epithelial cells.
RNA seq is a technology based on high-throughput sequencing to perform high-throughput sequencing analysis on all mRNA in tissue cells. This study used transcriptomics to analyze the RNA of AAI treated mouse kidney tissue and found a total of 4975 differentially expressed genes compared to the normal group. Among them, 2511 genes were upregulated and 2464 genes were downregulated. The top 5 differentially expressed genes were kap Spp1、Aldh1a2、Serpine1、Tnc。
Kap is a renal androgen regulatory protein, which is highly specific and strictly regulated in renal proximal tubular cells. Studies have shown that the interaction between Kap and cyclophilin B can protect proximal tubular cells from the toxicity of cyclosporine A. Spp1, also known as osteopontin, is an extracellular matrix chemokine like phosphoglycoprotein that is associated with immune regulation, tumorigenesis, and cell signaling. Studies have found that upregulation of Spp1 can activate the NF – κ B signaling pathway to promote cancer progression. Aldh1a2 belongs to the aldehyde dehydrogenase 1 family and is involved in the synthesis of endogenous all trans retinoic acid (ATRA) during kidney development. As an active metabolite of vitamin A, ATRA plays an important role in cell proliferation, differentiation, and apoptosis. Serpine 1 is a type 1 plasminogen activator inhibitor. Many studies have pointed out that Serpine 1 has abnormal expression in cancer, such as oral squamous cell carcinoma, esophageal carcinoma, bladder cancer carcinoma, etc. Tnc is a large molecule extracellular matrix (ECM) glycoprotein that is highly expressed during embryonic development. It can bind to ECM structural proteins and cell surface receptors (such as EGFR, integrin), activate downstream pathways, and regulate cell adhesion, diffusion, migration, and proliferation. Studies have shown that Tnc is the main component that promotes fibroblast proliferation and plays a key role in renal fibrosis. However, during acute kidney injury, Tnc can also be induced, thereby enhancing Wnt/β – catenin signaling to protect the kidneys. There are few reports on the correlation between the above genes and AAI nephrotoxicity.
The genes differentially expressed in AAI nephrotoxicity are mainly significantly enriched in GO classifications such as small molecule metabolism, extracellular matrix tissue, and immune responses involving neutrophils. KEGG analysis showed that differentially expressed genes are involved in the JAK-STAT and NF – κ B signaling pathways; Peroxidation reaction; Glycolysis/gluconeogenesis; Insulin resistance; Significant enrichment is observed in the degradation pathways of valine, leucine, and isoleucine. This suggests that the nephrotoxicity caused by AAI may be related to pathways such as inflammatory response, oxidative stress, and glucose metabolism.
Given the significant toxic side effects of AAI on the kidneys, it is important to monitor renal function during the clinical use of traditional Chinese medicines containing aristolochic acid, such as Qingmu Xiang, Zhusha Lian, and Xungufeng; At the same time, the genes and related signaling pathways revealed by transcriptomics for AAI nephrotoxicity may provide some therapeutic ideas for AAI nephrotoxicity.