安徽医科大学学报2025，Vol.60Issue(11)：2026-2034,9.DOI:10.19405/j.cnki.issn1000-1492.2025.11.006
TYROBP通过抑制ERK通路减缓糖尿病肾病的进展
TYROBP attenuates the progression of diabetic kidney disease by inhibiting the ERK signaling pathway
摘要
Abstract
Objective To investigate whether TYRO protein tyrosine kinase-binding protein(TYROBP)affects the progression of diabetic kidney disease(DKD)through the extracellular signal-regulated kinase(ERK)pathway.Methods Key genes in DKD were identified through bioinformatics analysis.Immunohistochemical staining and quantitative real-time PCR(qPCR)were used to validate the expression levels of TYROBP in a DKD mouse model and high glucose-stimulated NRK-52E cells.NRK-52E cell models with stable TYROBP overexpression/knockdown and their corresponding empty vector(ev)/scrambled sequence(ss)controls were established via lentiviral trans-fection.Cells were treated with 5.5 mmol/L or 30.0 mmol/L glucose for 72 hours to mimic normal glucose(NG)and high glucose(HG)conditions,respectively.High glucose medium containing 3.5 μmol/L FR180204 was used for ERK inhibitor intervention.The experiment included seven groups:ev+NG,ev+HG,oe-TYROBP+HG,ss+NG,ss+HG,sh-TYROBP+HG,and sh-TYROBP+HG+ERK inhibitor.Western blot was used to de-tect the expression levels of phosphorylated ERK/total ERK(p-ERK/ERK),apoptosis-related proteins B-cell lym-phoma-2(Bcl-2)and Bcl-2-associated X protein(Bax),and epithelial-mesenchymal transition(EMT)-related proteins E-cadherin and α-smooth muscle actin(α-SMA).Tetramethylrhodamine ethyl ester(TMRE)staining and Annexin V-fluorescein isothiocyanate/propidium iodide(Annexin V-FITC/PI)flow cytometry were performed to as-sess mitochondrial membrane potential and apoptosis levels.Results Bioinformatics analysis identified TYROBP as a key gene in DKD.In vivo and in vitro validation showed increased TYROBP mRNA levels in DKD models.The results from the HG model indicated that,compared to the ev+NG/ss+NG group,the ev+HG/ss+HG group demonstrated increased p-ERK/ERK expression,reduced mitochondrial membrane potential,elevated apoptosis,and enhanced EMT.In TYROBP-perturbed NRK-52E cells,compared to the ev+HG group,the oe-TYROBP+HG group showed decreased p-ERK/ERK expression(P＜0.01),increased mitochondrial membrane potential(P＜0.05),reduced apoptosis(P＜0.001),and attenuated EMT;whereas compared to the ss+HG group,the sh-TYROBP+HG group exhibited increased p-ERK/ERK expression(P＜0.001),decreased mitochondrial mem-brane potential(P＜0.01),elevated apoptosis(P＜0.001),and enhanced EMT.Furthermore,compared to the sh-TYROBP+HG group,the sh-TYROBP+HG+ERK inhibitor group displayed reduced p-ERK/ERK expression(P＜0.01),increased mitochondrial membrane potential(P＜0.001),decreased apoptosis(P＜0.001),and suppressed EMT.Compared with the scrambled sequence control+high glucose group,the TYROBP knockdown+high glucose group showed elevated p-ERK/ERK expression(P＜0.001),reduced mitochondrial membrane potential(P＜0.01),increased apoptosis level(P＜0.001),and enhanced EMT.Compared with the TYROBP knockdown+high glucose group,the TYROBP knockdown+high glucose+ERK inhibitor group demonstrated decreased p-ERK/ERK expression(P＜0.01),restored mitochondrial membrane potential(P＜0.001),reduced apoptosis level(P＜0.001),and suppressed EMT.Conclusion TYROBP may regulate the ERK signaling path-way to modulate apoptosis-and EMT-related proteins,thereby influencing mitochondrial membrane potential,apop-tosis,and EMT in renal tubular epithelial cells and contributing to DKD progression.关键词
糖尿病肾病/肾小管细胞/凋亡/上皮间质转化/TYROBP/ERKKey words
diabetic nephropathy/renal tubular cell/apoptosis/epithelial-mesenchymal transition/TYROBP/ERK分类
医药卫生引用本文复制引用
李亮,黄杰,王心灵,闫丽平,於慧清,李治国..TYROBP通过抑制ERK通路减缓糖尿病肾病的进展[J].安徽医科大学学报,2025,60(11):2026-2034,9.基金项目
河北省自然科学基金(编号:H2020209243) （编号:H2020209243）
中央引导地方科技发展资金项目(编号:236Z7712G) Natural Science Foundation of Hebei Province(No.H2020209243) （编号:236Z7712G）
Central Government Guid-ing Local Scientific and Technological Development Fund(No.236Z7712G) （No.236Z7712G）
