Abstract：
BACKGROUND: Dysfunction in the processes of autophagy and apoptosis within renal tubular epithelial cells (RTEc) contributes to renal ischemia-reperfusion injury (IRI). However, the factors influencing this dysfunction remain unclear. Leucine-rich alpha-2-glycoprotein 1 (Lrg1) plays a role in the progression of diabetic nephropathy and kidney fibrosis by modulating the activin receptor-like kinase 1 (ALK1)-Smad1/5/8 and TGF-β1/Smad3 pathways, respectively. Therefore, we aimed to investigate whether Lrg1 is involved in the pathological mechanisms of renal IRI and whether its effects are related to the dysregulation of autophagy and apoptosis in RTEc.
METHODS: We conducted in vitro and in vivo experiments using CoCl2-induced hypoxic human kidney-2 (HK-2) cells and mice with renal IRI, respectively. Lrg1 was silenced using siRNA and lentiviral vectors in HK-2 cells and mouse kidneys. Rapamycin (Rapa) and methyladenine were applied to regulate autophagy in renal IRI models.
RESULTS: Increased Lrg1 expression was observed in hypoxic HK-2 cells and in the kidneys of mice with renal IRI. Silencing of Lrg1 through siRNA and lentiviral approaches restored autophagy and suppressed apoptosis in CoCl2-induced hypoxic HK-2 cells and renal IRI models. Additionally, reduced Lrg1 expression alleviated kidney damage caused by renal IRI. The downregulation of Lrg1 expression restrained the TGFβ-Smad1/5 signaling pathway in hypoxic-induced HK-2 cells and renal IRI by reducing ALK1 expression. Lastly, the enhancement of autophagy, achieved through Rapa treatment, provided protection against renal IRI in mice.
CONCLUSIONS: Our findings suggest that Lrg1 silencing can be applied as a potential therapeutic target to inhibit the TGFβ1-Smad1/5 pathway, thereby enhancing autophagy and decreasing apoptosis in patients with acute kidney injury.
METHODS: We conducted in vitro and in vivo experiments using CoCl2-induced hypoxic human kidney-2 (HK-2) cells and mice with renal IRI, respectively. Lrg1 was silenced using siRNA and lentiviral vectors in HK-2 cells and mouse kidneys. Rapamycin (Rapa) and methyladenine were applied to regulate autophagy in renal IRI models.
RESULTS: Increased Lrg1 expression was observed in hypoxic HK-2 cells and in the kidneys of mice with renal IRI. Silencing of Lrg1 through siRNA and lentiviral approaches restored autophagy and suppressed apoptosis in CoCl2-induced hypoxic HK-2 cells and renal IRI models. Additionally, reduced Lrg1 expression alleviated kidney damage caused by renal IRI. The downregulation of Lrg1 expression restrained the TGFβ-Smad1/5 signaling pathway in hypoxic-induced HK-2 cells and renal IRI by reducing ALK1 expression. Lastly, the enhancement of autophagy, achieved through Rapa treatment, provided protection against renal IRI in mice.
CONCLUSIONS: Our findings suggest that Lrg1 silencing can be applied as a potential therapeutic target to inhibit the TGFβ1-Smad1/5 pathway, thereby enhancing autophagy and decreasing apoptosis in patients with acute kidney injury.
摘要：
背景：肾小管上皮细胞（RTEc）自噬和凋亡过程中的功能障碍导致了肾缺血再灌注损伤（IRI）。然而,影响这种功能障碍的因素尚不清楚.富含亮氨酸的α-2-糖蛋白1（Lrg1）通过调节活化素受体样激酶1（ALK1）-Smad1/5/8和TGF-β1/Smad3途径在糖尿病肾病和肾纤维化的进展中发挥作用。分别。因此,我们旨在研究Lrg1是否参与肾IRI的病理机制，以及其作用是否与RTEc中自噬和凋亡的失调有关。
方法：我们使用CoCl2诱导的低氧人肾-2（HK-2）细胞和具有肾IRI的小鼠进行了体外和体内实验，分别。在HK-2细胞和小鼠肾脏中使用siRNA和慢病毒载体沉默Lrg1。雷帕霉素（Rapa）和甲基腺嘌呤用于调节肾IRI模型中的自噬。
结果：在低氧HK-2细胞和肾IRI小鼠的肾脏中观察到Lrg1表达增加。通过siRNA和慢病毒方法沉默Lrg1可恢复CoCl2诱导的低氧HK-2细胞和肾IRI模型中的自噬并抑制凋亡。此外,Lrg1表达降低减轻肾IRI引起的肾损害。Lrg1表达下调通过降低ALK1表达抑制低氧诱导的HK-2细胞TGFβ-Smad1/5信号通路和肾IRI。最后,自噬的增强，通过Rapa治疗，对小鼠肾脏IRI提供保护。
结论：我们的研究结果表明，Lrg1沉默可以作为抑制TGFβ1-Smad1/5通路的潜在治疗靶点，从而增强急性肾损伤患者的自噬和减少细胞凋亡。
方法：我们使用CoCl2诱导的低氧人肾-2（HK-2）细胞和具有肾IRI的小鼠进行了体外和体内实验，分别。在HK-2细胞和小鼠肾脏中使用siRNA和慢病毒载体沉默Lrg1。雷帕霉素（Rapa）和甲基腺嘌呤用于调节肾IRI模型中的自噬。
结果：在低氧HK-2细胞和肾IRI小鼠的肾脏中观察到Lrg1表达增加。通过siRNA和慢病毒方法沉默Lrg1可恢复CoCl2诱导的低氧HK-2细胞和肾IRI模型中的自噬并抑制凋亡。此外,Lrg1表达降低减轻肾IRI引起的肾损害。Lrg1表达下调通过降低ALK1表达抑制低氧诱导的HK-2细胞TGFβ-Smad1/5信号通路和肾IRI。最后,自噬的增强，通过Rapa治疗，对小鼠肾脏IRI提供保护。
结论：我们的研究结果表明，Lrg1沉默可以作为抑制TGFβ1-Smad1/5通路的潜在治疗靶点，从而增强急性肾损伤患者的自噬和减少细胞凋亡。
