Steinbrech D S, Longaker M T, Mehrara B J, Saadeh P B, Chin G S, Gerrets R P, Chau D C, Rowe N M, Gittes G K
Department of Surgery, New York University School of Medicine, New York, New York, 10016, USA.
J Surg Res. 1999 Jun 15;84(2):127-33. doi: 10.1006/jsre.1999.5627.
A number of studies have demonstrated the critical role of angiogenesis for successful wound repair in the surgical patient. Vascular disruption from tissue injury due to trauma or surgery leads to a hypoxic zone in the healing wound. In this dynamic process, angiogenesis is vital for the delivery of oxygen, nutrients, and growth factors necessary to initiate the synthetic processes of wound healing. Fibroblasts, invading the wound early in the healing process, are involved in extracellular matrix (ECM) deposition as well as wound contraction. However, the exact mechanisms by which important genes are regulated remain unknown. In order to examine these processes, we studied the effects of hypoxia on fibroblasts for the expression of VEGF, type IalphaI collagen, and matrix-metalloproteinase-3, three genes essential for the regulation of angiogenesis, ECM deposition, and ECM degradation in wound healing. Primary cell cultures of normal human dermal fibroblasts (NHDFs) were placed in hypoxia for varying periods of time. Northern blot hybridization was performed with [alpha32P]dCTP-labeled cDNA probes for VEGF, type IalphaI collagen, and MMP-3. The results demonstrated a time-dependent VEGF mRNA upregulation (470% of baseline) under hypoxia. Type IalphaI collagen increased (170% of baseline) at 24 h, but was then abruptly downregulated to 3.8% of baseline at 48 h. MMP-3 was incrementally downregulated to 2.2% of baseline at 48 h. These experiments focused on the effect of hypoxia on genes thought to play a role in wound repair. VEGF upregulation in the hypoxic microenvironment of the early wound may serve to stimulate angiogenesis. Type IalphaI collagen, though upregulated early on, was abruptly downregulated at 48 h. This downregulation may reflect the in vivo requirement for angiogenesis to deliver oxygen for successful hydroxylation and collagen synthesis in the wound. MMP-3, also downregulated at 48 h, may also implicate the need for angiogenesis. These data support the theory that hypoxia-driven angiogenesis is critical for ECM formation and remodeling in successful soft tissue repair. Furthermore, they may represent the role of hypoxia as an important regulator to efficiently balance these complex processes in the healing wound.
多项研究已证明血管生成在手术患者伤口成功修复中起关键作用。创伤或手术导致的组织损伤引起的血管破坏会在愈合伤口中形成缺氧区。在这个动态过程中，血管生成对于输送启动伤口愈合合成过程所需的氧气、营养物质和生长因子至关重要。成纤维细胞在愈合过程早期侵入伤口，参与细胞外基质（ECM）沉积以及伤口收缩。然而，重要基因被调控的确切机制仍不清楚。为了研究这些过程，我们研究了缺氧对成纤维细胞中血管内皮生长因子（VEGF）、Ⅰα1型胶原蛋白和基质金属蛋白酶-3表达的影响，这三个基因对于伤口愈合中血管生成、ECM沉积和ECM降解的调控至关重要。将正常人皮肤成纤维细胞（NHDFs）的原代细胞培养物置于缺氧环境中不同时间段。用[α32P]dCTP标记的VEGF、Ⅰα1型胶原蛋白和基质金属蛋白酶-3的cDNA探针进行Northern印迹杂交。结果表明，在缺氧条件下，VEGF mRNA呈时间依赖性上调（为基线的470%）。Ⅰα1型胶原蛋白在24小时时增加（为基线的170%），但随后在48小时时突然下调至基线的3.8%。基质金属蛋白酶-3在48小时时逐渐下调至基线的2.2%。这些实验聚焦于缺氧对被认为在伤口修复中起作用的基因的影响。早期伤口缺氧微环境中VEGF的上调可能有助于刺激血管生成。Ⅰα1型胶原蛋白虽然早期上调，但在48小时时突然下调。这种下调可能反映了体内血管生成对于为伤口中成功的羟基化和胶原蛋白合成输送氧气的需求。基质金属蛋白酶-3在48小时时也下调，这也可能意味着需要血管生成。这些数据支持以下理论：缺氧驱动的血管生成对于成功的软组织修复中ECM的形成和重塑至关重要。此外，它们可能代表了缺氧作为重要调节因子在有效平衡愈合伤口中这些复杂过程的作用。
