探讨恩格列净导致的远端肾小管高糖环境是否会诱导远端肾小管细胞衰老及其可能通路。

选取9周龄雄性C57BL/6小鼠，随机分为恩格列净组[10 mg/(kg· d)恩格列净灌胃]与对照组(等体积生理盐水灌胃)，每组6只。连续干预4周后取材，检测血糖与尿糖，免疫荧光与Western印迹检测远端肾小管衰老相关指标p16、p21和p53表达，利用转录组测序筛选潜在通路并进行基因集富集分析。体外培养的小鼠远端肾小管上皮细胞分为对照组(无干预)、甘露醇组(30 mmol/L甘露醇)和高糖组(30 mmol/L葡萄糖)，孵育24 h后采用衰老相关β-半乳糖苷酶染色评估细胞衰老，实时定量PCR检测衰老标志物p21和铁死亡相关基因Alox15、Ncoa4、Slc11a2、Gpx4等表达。采用GraphPad Prism软件进行数据分析。

与对照组相比，恩格列净组小鼠尿糖升高(P<0.05)；免疫荧光与Western印迹结果显示，恩格列净组小鼠远端肾小管中p21与p53表达水平均显著上调(P均<0.05)；转录组测序及富集分析显示，铁死亡相关通路在干预组中显著激活(校正后q值均<0.05)。在小鼠肾组织及体外培养小鼠远端小管上皮细胞均检测到铁死亡相关基因Alox15、Ncoa4、Slc11a2表达升高，Gpx4表达下降(P均<0.05)。

恩格列净导致的小鼠远端肾小管尿糖升高可以诱导细胞衰老，其机制可能与铁死亡相关通路激活有关。

To investigate whether the high-glucose environment in the distal renal tubule induced by empagliflozin triggers senescence of distal tubular cells as well its potential signaling pathways.

Nine-week-old male C57BL/6 mice were selected and randomly divided into an empagliflozin group [intragastric administration of empagliflozin at 10 mg/(kg·d) and a control group (intragastric administration of an equal volume of normal saline), with six mice in each group. After 4 weeks of consecutive intervention, tissues were harvested, and blood glucose and urine glucose were measured. Immunofluorescence staining and Western blot were performed to detect the expression of senescence-related markers p16, p21, and p53 in distal renal tubules. Transcriptome sequencing was used to screen potential signaling pathways, and gene set enrichment analysis was carried out subsequently. Mouse distal renal tubular epithelial cells were divided into a control group (untreated), a mannitol group (30 mmol/L mannitol), and a high-glucose group (30 mmol/L glucose). After 24 h of incubation, cellular senescence was assessed by senescence-associated β-galactosidase staining. The mRNA expression of the senescence marker p21 and ferroptosis-related genes (Alox15, Ncoa4, Slc11a2, Gpx4) was determined by RT-qPCR. All data were analyzed using GraphPad Prism software.

Compared with the control group, urinary glucose was significantly increased in the empagliflozin group (P<0.05). Immunofluorescence and Western blot analysis showed that the expression levels of p21 and p53 were markedly upregulated in the renal distal tubules of empagliflozin-treated mice (all P<0.05). Transcriptome sequencing and gene set enrichment analysis revealed that ferroptosis-related pathways were significantly activated in the empagliflozin group (adjusted q<0.05). The expression of ferroptosis-related genes Alox15, Ncoa4, and Slc11a2 was upregulated, while Gpx4 expression was downregulated in both mouse renal tissues and cultured mouse distal renal tubular epithelial cells (all P<0.05).

Empagliflozin induced elevation of urinary glucose in mouse renal distal tubules promoted cellular senescence, which may be mediated by the activation of ferroptosis-related pathways.