Aims/hypothesis

Glomerular lipid accumulation is a defining feature of diabetic kidney disease (DKD); however, the precise underlying mechanism requires further elucidation. Recent evidence suggests a role for proprotein convertase subtilisin/kexin type 9 (PCSK9) in intracellular lipid homeostasis. Although PCSK9 is present in kidneys, its role within kidney cells and relevance to renal diseases remain largely unexplored. Therefore, we investigated the role of intracellular PCSK9 in regulating lipid accumulation and homeostasis in the glomeruli and podocytes under diabetic conditions. Furthermore, we aimed to identify the pathophysiological mechanisms responsible for the podocyte injury that is associated with intracellular PCSK9-induced lipid accumulation in DKD.

Methods

In this study, glomeruli were isolated from human kidney biopsy tissues, and glomerular gene-expression analysis was performed. Also, db/db and db/m mice were used to perform glomerular gene-expression profiling. We generated DKD models using a high-fat diet and low-dose intraperitoneal streptozocin injection in C57BL/6 and Pcsk9 knockout (KO) mice. We analysed cholesterol and triacylglycerol levels within the kidney cortex. Lipid droplets were evaluated using BODIPY staining. We induced upregulation and downregulation of PCSK9 expression in conditionally immortalised mouse podocytes using lentivirus and siRNA transfection techniques, respectively, under diabetic conditions.

Results

A significant reduction in transcription level of PCSK9 was observed in glomeruli of individuals with DKD. PCSK9 expression was also reduced in podocytes of animals under diabetic conditions. We observed significantly higher lipid accumulation in kidney tissues of Pcsk9 KO DKD mice compared with wild-type (WT) DKD mice. Additionally, Pcsk9 KO mouse models of DKD exhibited a significant reduction in mitochondria number vs WT models, coupled with a significant increase in mitochondrial size. Moreover, albuminuria and podocyte foot process effacement were observed in WT and Pcsk9 KO DKD mice, with KO DKD mice displaying more pronounced manifestations. Immortalised mouse podocytes exposed to diabetic stimuli exhibited heightened intracellular lipid accumulation, mitochondrial injury and apoptosis, which were ameliorated by Pcsk9 overexpression and aggravated by Pcsk9 knockdown in mouse podocytes.

Conclusions/interpretation

The downregulation of PCSK9 in podocytes is associated with lipid accumulation, which leads to mitochondrial dysfunction, cell apoptosis and renal injury. This study sheds new light on the potential involvement of PCSK9 in the pathophysiology of glomerular lipid accumulation and podocyte injury in DKD.

Graphical Abstract

中文翻译：

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PCSK9在糖尿病肾病肾小球脂质沉积和肾损伤中的作用

 目标/假设

肾小球脂质蓄积是糖尿病肾病 (DKD) 的一个决定性特征；然而，确切的潜在机制需要进一步阐明。最近的证据表明前蛋白转化酶枯草杆菌蛋白酶/kexin 9 型 (PCSK9) 在细胞内脂质稳态中发挥作用。尽管 PCSK9 存在于肾脏中，但其在肾细胞内的作用以及与肾脏疾病的相关性在很大程度上仍未得到探索。因此，我们研究了细胞内 PCSK9 在糖尿病条件下调节肾小球和足细胞脂质积累和稳态的作用。此外，我们的目的是确定 DKD 中与细胞内 PCSK9 诱导的脂质积累相关的足细胞损伤的病理生理机制。

 方法

在这项研究中，从人肾活检组织中分离出肾小球，并进行了肾小球基因表达分析。此外，还使用​​db/db和db/ m 小鼠进行肾小球基因表达谱分析。我们使用高脂肪饮食和低剂量腹腔注射链脲佐菌素在 C57BL/6 和Pcsk9敲除 (KO) 小鼠中生成 DKD 模型。我们分析了肾皮质内的胆固醇和三酰甘油水平。使用 BODIPY 染色评估脂滴。在糖尿病条件下，我们分别使用慢病毒和 siRNA 转染技术诱导条件永生化小鼠足细胞中 PCSK9 表达的上调和下调。

 结果

在 DKD 患者的肾小球中观察到PCSK9转录水平显着降低。糖尿病条件下动物的足细胞中 PCSK9 的表达也有所减少。我们观察到与野生型（WT）DKD 小鼠相比， Pcsk9 KO DKD 小鼠肾组织中的脂质积累明显更高。此外，与 WT 模型相比， Pcsk9 KO DKD 小鼠模型的线粒体数量显着减少，同时线粒体大小显着增加。此外，在WT和Pcsk9 KO DKD小鼠中观察到蛋白尿和足细胞足突消失，其中KO DKD小鼠表现出更明显的表现。暴露于糖尿病刺激的永生化小鼠足细胞表现出细胞内脂质积累增加、线粒体损伤和细胞凋亡，这些现象因小鼠足细胞中Pcsk9过表达而改善，而因Pcsk9敲低而加剧。

结论/解释

足细胞中 PCSK9 的下调与脂质积累有关，从而导致线粒体功能障碍、细胞凋亡和肾损伤。这项研究为 PCSK9 在 DKD 肾小球脂质积累和足细胞损伤的病理生理学中的潜在参与提供了新的线索。

 图解摘要