Renal Pathology Lab, Hankook Kidney and Diabetes Institute, Seoul, Korea.
J Cell Physiol. 2017 Dec;232(12):3209-3217. doi: 10.1002/jcp.25867. Epub 2017 May 3.
Studies have shown that high levels of serum free fatty acids (FFAs) are associated with lipotoxicity and type 2 diabetes. Palmitic acid (PA) is the predominant circulating saturated FFA, yet its role in the pathogenesis of diabetic nephropathy (DN) is not clear. Recently, one study suggested that mitochondrial superoxide production is related to AMP-activated protein kinase (AMPK) activity in diabetic mice kidneys. To elucidate the link between PA and oxidative stress and AMPK activity in DN, we compared the cultured murine podocytes exposed to PA and oleic acid (OA). Incubation of cells with 250 μM PA or OA induced a translocation of CD36, a fatty acid transport protein, with intracellular lipid accumulation. PA, but not OA, induced mitochondrial superoxide and hydrogen peroxide (H O ) generation in podocytes, as shown by enhanced fluorescence of MitoSOX Red and dichlorofluorescein (DCF), respectively. Costimulation of PA-treated cells with the H O scavenger catalase abolished the PA-induced DCF fluorescence. Only PA induced mitochondrial damage as shown by electron microscopy. The AMPK activity was determined by immunoblotting, measuring the ratio of phosphorylated AMPK (p-AMPK) to total AMPK. Only PA significantly increased the p-AMPK levels compared with controls. Addition of catalase to PA-treated cells did not affect the PA-stimulated p-AMPK levels. Collectively, our results indicate that PA induces mitochondrial superoxide and H O generation in cultured podocytes, which may not be directly linked to AMPK activation. Given that, PA seems to play an important role in the pathogenesis of DN through lipotoxicity initiated by mitochondrial superoxide overproduction.
研究表明，血清游离脂肪酸（FFAs）水平升高与脂毒性和 2 型糖尿病有关。软脂酸（PA）是主要的循环饱和 FFA，但它在糖尿病肾病（DN）发病机制中的作用尚不清楚。最近的一项研究表明，糖尿病小鼠肾脏中线粒体超氧化物的产生与 AMP 激活蛋白激酶（AMPK）活性有关。为了阐明 PA 与氧化应激和 AMPK 活性在 DN 中的关系，我们比较了暴露于 PA 和油酸（OA）的培养鼠足细胞。用 250μM PA 或 OA 孵育细胞会导致脂肪酸转运蛋白 CD36 易位和细胞内脂质积累。PA 但不是 OA 会诱导足细胞中线粒体超氧化物和过氧化氢（H O ）的产生，这可以通过 MitoSOX Red 和二氯荧光素（DCF）的荧光增强分别显示。用 H O 清除剂过氧化氢酶对 PA 处理的细胞进行共刺激，消除了 PA 诱导的 DCF 荧光。只有 PA 通过电子显微镜显示出诱导的线粒体损伤。通过免疫印迹法测定 AMPK 活性，测量磷酸化 AMPK（p-AMPK）与总 AMPK 的比值。只有 PA 与对照组相比显著增加了 p-AMPK 水平。向 PA 处理的细胞中添加过氧化氢酶不会影响 PA 刺激的 p-AMPK 水平。总之，我们的结果表明，PA 可诱导培养的足细胞中线粒体超氧化物和 H O 的产生，这可能与 AMPK 激活无关。鉴于此，PA 似乎通过线粒体超氧化物过度产生引发的脂毒性在 DN 的发病机制中发挥重要作用。
