Ji Xin-Yi, Chen Yang, Ye Guang-Hua, Dong Miao-Wu, Lin Ke-Zhi, Han Jun-Ge, Feng Xiang-Ping, Li Xing-Biao, Yu Lin-Sheng, Fan Yan-Yan
Department of Forensic Medicine, School of Basic Medical Science, Wenzhou Medical University, Higher Education District, Wenzhou, 325035, Zhejiang Province, People's Republic of China.
Forensic Center, Wenzhou Medical University, Wenzhou, 325027, Zhejiang Province, People's Republic of China.
Int J Legal Med. 2017 May;131(3):691-698. doi: 10.1007/s00414-016-1529-7. Epub 2017 Jan 11.
With the prevalence of diabetes, it is becoming important to analyze the diabetic wound age in forensic practice. The present study investigated the time-dependent expression of receptor for advanced glycation end products (RAGE) during diabetic wound healing in mice and its applicability to wound age determination by immunohistochemistry, double immunofluorescence, and Western blotting. After an incision was created in genetically diabetic db/db mice and control mice, mice were killed at posttraumatic intervals ranging from 6 h to 14 days, followed by the sampling of wound margin. Compared with control mice, diabetic mice showed the delayed wound healing. In control and diabetic wound specimens, RAGE immunoreactivity was observed in a small number of polymorphonuclear cells (PMNs), a number of macrophages, and fibroblasts. Morphometrically, the positive ratios of RAGE in macrophages or fibroblasts considerably increased in diabetic wounds during late repair, which exceeded 60% at 7 and 10 days post-injury. There were no control wound specimens to show a ratio of >60% in macrophages or fibroblasts. By Western blotting analysis, the ratios of RAGE to GAPDH were >1.4 in all diabetic wound samples from 7 to 10 days post-injury, which were >1.8 at 10 days after injury. By comparison, no control wound specimens indicated a ratio of >1.4. In conclusion, the expression of RAGE is upregulated and temporally distributed in macrophages and fibroblasts during diabetic wound healing, which might be closely involved in prolonged inflammation and deficient healing. Moreover, RAGE is promising as a useful marker for diabetic wound age determination.
随着糖尿病患病率的上升，在法医学实践中分析糖尿病伤口的形成时间变得愈发重要。本研究调查了晚期糖基化终末产物受体（RAGE）在小鼠糖尿病伤口愈合过程中的时间依赖性表达，以及其在通过免疫组织化学、双重免疫荧光和蛋白质印迹法确定伤口形成时间方面的适用性。在基因糖尿病db/db小鼠和对照小鼠身上制造切口后，在创伤后6小时至14天的不同时间点处死小鼠，随后采集伤口边缘样本。与对照小鼠相比，糖尿病小鼠的伤口愈合延迟。在对照和糖尿病伤口标本中，在少量多形核细胞（PMN）、大量巨噬细胞和成纤维细胞中观察到RAGE免疫反应性。形态计量学分析显示，在糖尿病伤口晚期修复过程中，巨噬细胞或成纤维细胞中RAGE的阳性率显著增加，在受伤后7天和10天时超过60%。对照伤口标本中，巨噬细胞或成纤维细胞的阳性率均未超过60%。通过蛋白质印迹分析，在受伤后7至10天的所有糖尿病伤口样本中，RAGE与甘油醛-3-磷酸脱氢酶（GAPDH）的比值均>1.4，在受伤后10天时>1.8。相比之下，对照伤口标本中该比值均未超过1.4。总之，在糖尿病伤口愈合过程中，RAGE在巨噬细胞和成纤维细胞中的表达上调且呈时间性分布，这可能与炎症持续时间延长和愈合不足密切相关。此外，RAGE有望成为确定糖尿病伤口形成时间的有用标志物。
