Division of Pediatric Surgery, Children's Hospital of Pittsburgh of the University of Pittsburgh Medical Center, Pittsburgh, Pennsylvania 15207, USA.
J Biol Chem. 2010 Feb 12;285(7):4995-5002. doi: 10.1074/jbc.M109.067454. Epub 2009 Dec 11.
Toll-like receptor-4 (TLR4) is the receptor for bacterial lipopolysaccharide, yet it may also respond to a variety of endogenous molecules. Necrotizing enterocolitis (NEC) is the leading cause of death from gastrointestinal disease in newborn infants and is characterized by intestinal mucosal destruction and impaired enterocyte migration due to increased TLR4 signaling on enterocytes. The endogenous ligands for TLR4 that lead to impaired enterocyte migration remain unknown. High mobility group box-1 (HMGB1) is a DNA-binding protein that is released from injured cells during inflammation. We thus hypothesize that extracellular HMGB1 inhibits enterocyte migration via activation of TLR4 and sought to define the pathways involved. We now demonstrate that murine and human NEC are associated with increased intestinal HMGB1 expression, that serum HMGB1 is increased in murine NEC, and that HMGB1 inhibits enterocyte migration in vitro and in vivo in a TLR4-dependent manner. This finding was unique to enterocytes as HMGB1 enhanced migration of inflammatory cells in vitro and in vivo. In seeking to understand the mechanisms involved, TLR4-dependent HMGB1 signaling increased RhoA activation in enterocytes, increased phosphorylation of focal adhesion kinase, and increased phosphorylation of cofilin, resulting in increased stress fibers and focal adhesions. Using single cell force traction microscopy, the net effect of HMGB1 signaling was a TLR4-dependent increase in cell force adhesion, accounting for the impaired enterocyte migration. These findings demonstrate a novel pathway by which TLR4 activation by HMGB1 delays mucosal repair and suggest a novel potential therapeutic target in the amelioration of intestinal inflammatory diseases like NEC.
Toll 样受体-4（TLR4）是细菌脂多糖的受体，但它也可能对各种内源性分子做出反应。坏死性小肠结肠炎（NEC）是新生儿胃肠道疾病死亡的主要原因，其特征是由于肠细胞 TLR4 信号的增加，导致肠黏膜破坏和肠细胞迁移受损。导致肠细胞迁移受损的 TLR4 的内源性配体仍不清楚。高迁移率族蛋白 B1（HMGB1）是一种在炎症期间从受损细胞中释放的 DNA 结合蛋白。因此，我们假设细胞外 HMGB1 通过激活 TLR4 抑制肠细胞迁移，并试图确定涉及的途径。我们现在证明，鼠和人 NEC 与肠道 HMGB1 表达增加有关，鼠 NEC 中血清 HMGB1 增加，HMGB1 以 TLR4 依赖的方式抑制体外和体内肠细胞迁移。这一发现是独特的，因为 HMGB1 增强了体外和体内炎症细胞的迁移。为了了解所涉及的机制，TLR4 依赖性 HMGB1 信号增加了肠细胞中 RhoA 的激活，增加了粘着斑激酶的磷酸化，增加了丝切蛋白的磷酸化，导致应力纤维和粘着斑增加。通过单细胞力牵引显微镜，HMGB1 信号的净效应是 TLR4 依赖性的细胞力附着增加，这解释了肠细胞迁移受损的原因。这些发现表明了一种通过 HMGB1 激活 TLR4 延迟黏膜修复的新途径，并为改善像 NEC 这样的肠道炎症性疾病提供了一个新的潜在治疗靶点。
