Li Yufeng, Zhao Mingming, He Dan, Zhao Xuyang, Zhang Wenjing, Wei Lixin, Huang Edgar, Ji Liang, Zhang Meng, Willard Belinda, Fu Zuodi, Wang Lijuan, Pan Bing, Zheng Lemin, Ji Linong
Department of Endocrinology and Metabolism, Peking University People's Hospital, No.11 Xizhimen Nan Dajie, Xicheng District, Beijing, 100044, China.
Department of Endocrinology and Metabolism, Capital Medical University Pinggu Teaching Hospital, Beijing, 101200, China.
Lipids Health Dis. 2016 Apr 14;15:76. doi: 10.1186/s12944-016-0246-z.
Diabetic nephropathy has a high cardiovascular risk with a low-level HDL(high density lipoprotein) in epidemiologic studies. Glycated HDL in diabetes can diminish the capacity to stimulate endothelial cell migration, but the mechanism has not been adequately explored in diabetic nephropathy. We performed this study to find out whether HDL in diabetic nephropathy is more dysfunctional than HDL in diabetes without complications.
Endothelial cells were treated with N-HDL (normal), D-HDL (T2DM[type 2 diabetes mellitus] without complications), DN-HDL (T2DM nephropathy), N-apoA-I (normal apoA-I), and G-apoA-I (glycated apoA-I in vitro). Cell migration capacity was measured with wound-healing and transwell migration assay in vitro and electric carotid injury model in vivo. Protein glycation levels were measured with nanoLC-MS/MS. PI3K expression and Akt phosphorylation were analyzed by western blot.
In wound-healing assay, DN-HDL showed a 17.12% decrease compared with D-HDL (p < 0.05). DN-HDL showed a 29.85% decrease in comparison with D-HDL (p < 0.001) in transwell assay. In the electric carotid injury model, D-HDL and DN-HDL impaired the re-endothelialization capacity; DN-HDL was less effective than D-HDL. Meanwhile, DN-HDL was found to have a significantly higher protein glycation level than D-HDL (p < 0.001). PI3K expression and Akt phosphorylation were reduced significantly in DN-HDL in comparison with D-HDL and N-HDL.
We found that HDL from diabetic nephropathy has a higher level of glycation and induced less cell migration in vitro and in vivo compared with that from diabetes without nephropathy. This finding suggests that diabetic nephropathy has higher levels of glycated HDL and partially explains why patients with DN have a higher risk of cardiovascular disease.
在流行病学研究中，糖尿病肾病具有较高的心血管风险且高密度脂蛋白（HDL）水平较低。糖尿病中的糖化HDL可降低刺激内皮细胞迁移的能力，但在糖尿病肾病中该机制尚未得到充分研究。我们开展这项研究以确定糖尿病肾病中的HDL是否比无并发症糖尿病中的HDL功能更失调。
用N-HDL（正常）、D-HDL（无并发症的2型糖尿病）、DN-HDL（2型糖尿病肾病）、N-载脂蛋白A-I（正常载脂蛋白A-I）和G-载脂蛋白A-I（体外糖化载脂蛋白A-I）处理内皮细胞。通过体外伤口愈合和Transwell迁移试验以及体内颈动脉电损伤模型测量细胞迁移能力。用纳升液相色谱-串联质谱法测量蛋白质糖化水平。通过蛋白质印迹法分析PI3K表达和Akt磷酸化。
在伤口愈合试验中，与D-HDL相比，DN-HDL降低了17.12%（p<0.05）。在Transwell试验中，与D-HDL相比，DN-HDL降低了29.85%（p<0.001）。在颈动脉电损伤模型中，D-HDL和DN-HDL损害了再内皮化能力；DN-HDL的效果不如D-HDL。同时，发现DN-HDL的蛋白质糖化水平显著高于D-HDL（p<0.001）。与D-HDL和N-HDL相比，DN-HDL中的PI3K表达和Akt磷酸化显著降低。
我们发现，与无肾病的糖尿病患者相比，糖尿病肾病患者的HDL糖化水平更高，且在体外和体内诱导的细胞迁移更少。这一发现表明糖尿病肾病患者的糖化HDL水平更高，部分解释了糖尿病肾病患者心血管疾病风险更高的原因。
