Department of Surgery, Division of Vascular Surgery, University of Wisconsin School of Medicine, Madison, Wisconsin 53705, USA.
J Surg Res. 2011 Dec;171(2):871-6. doi: 10.1016/j.jss.2010.04.027. Epub 2010 May 11.
Advanced glycation end products (AGEs), formed from proteins and peptides by nonenzymatic glycoxidation after contact with aldose sugars, have been implicated in the pathogenesis of age-related cardiac and vascular dysfunction. Our previous study demonstrated significantly elevated levels of AGE and the receptor for AGE (RAGE) in human abdominal aortic aneurysm (AAA) tissues. Inhibition of AGE signaling by targeted gene deletion of RAGE markedly reduced the development of aneurysm in a mouse model of AAA. We also showed that AGE may stimulate aneurysm formation by promoting metalloproteinase (MMP)-9 expression. In this study, we investigated the molecular mechanism underlying this novel function of AGE.
The murine macrophage cell line RAW 264.7 was pretreated with AGE, TGF-β, and MAPK inhibitors. The protein was collected for Western blot analysis. Culture supernatants were collected to determine MMP-9 activity by gelatin zymography.
We found that AGE induced the production of MMP-9 in macrophages in a dose-dependent manner. This induction of MMP-9 was markedly diminished by pretreatment with TGF-β. To delineate the underlying molecular mechanism, we showed that AGE increased phosphorylation of p44/42 ERK, p38, JNK, and PI3K in macrophages. Moreover, AGE induced active p65 subunit of NF- κB. Inhibition of ERK (UO126) or p38 (SB203580), but not PI3K (LY294002 or wortmannin), blocked AGE-induced MMP-9 expression. In contrast, inhibition of JNK (SP-600125) significantly enhanced the stimulatory effect of AGE on MMP-9. Furthermore, TGF-β suppressed AGE-induced expression of the active p65 subunit of NF-κB.
Our data indicate that AGE induces MMP-9 through activation of ERK, p38 mitogen-activated protein and NF-κB, a pathway that is antagonized by TGF-β. This finding in conjunction with previously reported AGE functions in inflammation suggests that anti-AGE therapies could be effective in the prevention of human AAA development and progression.
高级糖基化终产物（AGEs）是蛋白质和肽与醛糖糖接触后通过非酶糖基化形成的，已被牵连到与年龄相关的心脏和血管功能障碍的发病机制中。我们之前的研究表明，在人类腹主动脉瘤（AAA）组织中，AGE 和 AGE 受体（RAGE）的水平显着升高。通过 RAGE 的靶向基因缺失抑制 AGE 信号可显着减少 AAA 小鼠模型中动脉瘤的发展。我们还表明，AGE 可能通过促进基质金属蛋白酶（MMP）-9 的表达来刺激动脉瘤的形成。在这项研究中，我们研究了 AGE 这一新型功能的潜在分子机制。
用 AGE、TGF-β 和 MAPK 抑制剂预处理鼠巨噬细胞系 RAW 264.7。收集蛋白质进行 Western blot 分析。收集培养上清液，通过明胶酶谱法测定 MMP-9 活性。
我们发现 AGE 以剂量依赖的方式诱导巨噬细胞中 MMP-9 的产生。TGF-β 的预处理显着减少了 MMP-9 的这种诱导。为了阐明潜在的分子机制，我们表明 AGE 增加了巨噬细胞中 p44/42 ERK、p38、JNK 和 PI3K 的磷酸化。此外，AGE 诱导 NF-κB 的活性 p65 亚单位。ERK（UO126）或 p38（SB203580）抑制剂而非 PI3K（LY294002 或 wortmannin）抑制 AGE 诱导的 MMP-9 表达。相反，JNK（SP-600125）抑制剂显着增强了 AGE 对 MMP-9 的刺激作用。此外，TGF-β 抑制了 AGE 诱导的 NF-κB 活性 p65 亚单位的表达。
我们的数据表明，AGE 通过激活 ERK、p38 丝裂原激活蛋白和 NF-κB 诱导 MMP-9，该途径被 TGF-β 拮抗。这一发现与之前报道的 AGE 在炎症中的功能相结合表明，抗 AGE 疗法可能有效预防人类 AAA 的发生和进展。
