支链氨基酸氨基转移酶1通过P53信号调节过氧化氢导致的肾小管上皮细胞损伤

刘慧; 张玲; 王正阳; 孙芳芳; 张富洋; 陶凌

doi:10.12125/j.chj.202411119

支链氨基酸氨基转移酶1通过P53信号调节过氧化氢导致的肾小管上皮细胞损伤

Branched chain amino acid aminotransferase-1 regulates tubular epithelial cell injury via P53 signaling.

摘要

摘要:
目的探讨支链氨基酸氨基转移酶1（branched chain amino acid aminotransferase-1，BCAT1）调控肾小管上皮细胞（tubular epithelial cell，TEC）损伤的作用和机制。
方法从C57BL/6J小鼠肾组织中分离并培养TEC。通过腺病毒转染TEC分别实现BCAT1过表达和敲低；通过过氧化氢（hydrogen dioxide，H₂O₂）处理诱导TEC凋亡；通过CCK-8评估TEC细胞活力；通过检测Cleaved-caspase3表达水平和原位末端转移酶标记技术（TdT-mediated dUTP nick-end labeling，TUNEL）染色评估TEC凋亡；使用P53抑制剂PFT-1α阻断P53信号评估其在BCAT1调节TEC损伤中的作用。
结果在H₂O₂处理的TEC中，BCAT1 mRNA和蛋白水平均显著降低（均P<0.01）。沉默BCAT1后，H₂O₂诱导的TEC凋亡显著加重（P<0.01）；而过表达BCAT1显著减轻了H₂O₂诱导的TEC凋亡（均P<0.01）。沉默BCAT1导致H₂O₂处理的TEC中P53下游基因表达水平进一步升高，提示沉默BCAT1进一步激活了P53信号（均P<0.05）。P53抑制剂PFT-1α可以明显消除由沉默BCAT1导致的P53信号活化和TEC凋亡（均P<0.01）。
结论 BCAT1通过P53信号调控H₂O₂诱导的TEC损伤，可能是一个防治慢性肾脏病有潜力的治疗靶点。

Abstract:
AIM Explore the effects and molecular mechanism of branched chain amino acid aminotransferase-1 (BCAT1) in regulating renal tubular epithelial cell (TEC) injury.
METHODS Cortices from mice with C57BL/6J were used to isolate TEC.BCAT1 was overexpressed or knocked down in TEC, which was performed by adenovirus vectors. Hydrogen dioxide (H₂O₂) stimulation induced TEC apoptosis. TEC viability was assessed by cell counting kit-8 (CCK-8). TEC apoptosis was evaluated by Cleaved-caspase3 expression and TdT-mediated dUTP Nick-End Labeling (TUNEL) staining. The molecular mechanism of BCAT1 regulating TEC injury by P53 signaling was investigated by using PFT-1α, a P53 inhibitor.
RESULTS In H₂O₂-induced apoptotic TEC, a declined expression of BCAT1 mRNA and protein was observed (P<0.01). BCAT1 knockdown exacerbated H₂O₂-induced TEC apoptosis (P<0.01), whereas overexpressing BCAT1 ameliorated it (P<0.01). Expression of P53 downstream target genes further elevated when silencing BCAT1 in H₂O₂-induced TEC, suggesting the activation of P53 signaling (P<0.05). PFT-1α reversed detrimental effects of silencing BCAT1 in H₂O₂-induced TEC apoptosis (P<0.01).
CONCLUSION BCAT1 regulates TEC injury through P53 signaling and might serve as a therapeutic target for chronic kidney disease.

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