Turner Neil A, Ho Selina, Warburton Philip, O'Regan David J, Porter Karen E
Institute for Cardiovascular Research, School of Medicine, University of Leeds, Leeds, United Kingdom.
J Vasc Surg. 2007 May;45(5):1022-8. doi: 10.1016/j.jvs.2007.01.061.
Smooth muscle cell (SMC) proliferation, invasion, and matrix metalloproteinase (MMP) secretion are key events in the development of intimal hyperplasia, the lesion that causes coronary artery bypass graft (CABG) failure. Saphenous vein (SV) grafts are the most commonly used bypass conduits but are markedly more susceptible to intimal hyperplasia than internal mammary artery (IMA) grafts. We hypothesized that this may be due to inherent functional differences between SV-SMCs and IMA-SMCs. In this study we used paired cultures of SV-SMCs and IMA-SMCs from the same patients and compared their rates of proliferation, invasion, migration, and MMP-2 and MMP-9 secretion.
SMCs were cultured from explants of paired SV and IMA from 22 patients undergoing CABG. SMC populations of equivalent passage were used to determine proliferation in response to 10% fetal calf serum (FCS), 10 ng/mL platelet-derived growth factor (PDGF), and 10 ng/mL basic fibroblast growth factor (bFGF) by counting cells during a 7-day period. Immunoblotting was used to quantify phosphorylation of p44/42-mitogen-activated protein kinase (MAPK). Invasion and migration rates of paired SMCs were quantified using a modified Boyden chamber technique in the presence or absence of a Matrigel basement membrane barrier (BD Biosciences, Oxford, UK). Conditioned media from invasion assays were analyzed for secretion of MMP-2 and MMP-9 by gelatin zymography.
Analysis of areas under curves for 7-day proliferation assays revealed that the number of SV-SMCs in response to FCS, PDGF, and bFGF was 2.1, 2.0, and 2.3 times higher, respectively, than that of paired IMA-SMCs. Basal MAPK activation in SV-SMCs was approximately double that of paired IMA-SMCs. SV-SMCs exhibited a 2.1-fold increase in invasion rate (Matrigel barrier) compared with IMA-SMCs, but migration rates (no Matrigel barrier) and MMP-2 and MMP-9 secretion were similar for the two cell types.
Human SV-SMCs are inherently more proliferative and invasive than paired IMA-SMCs, likely due to a relative increase in p44/42-MAPK activation. These inherent functional differences between SMC of different origins may contribute to the increased prevalence of intimal hyperplasia in SV grafts compared with IMA grafts.
平滑肌细胞（SMC）增殖、侵袭及基质金属蛋白酶（MMP）分泌是内膜增生发展过程中的关键事件，内膜增生是导致冠状动脉旁路移植术（CABG）失败的病变。大隐静脉（SV）移植物是最常用的旁路管道，但与乳内动脉（IMA）移植物相比，其对内膜增生的易感性明显更高。我们推测这可能是由于SV-SMC与IMA-SMC之间存在内在功能差异。在本研究中，我们使用来自同一患者的SV-SMC和IMA-SMC配对培养物，比较它们的增殖、侵袭、迁移速率以及MMP-2和MMP-9的分泌情况。
从22例行CABG患者的配对SV和IMA外植体中培养SMC。使用同等传代的SMC群体，通过在7天内计数细胞来确定其对10%胎牛血清（FCS）、10 ng/mL血小板衍生生长因子（PDGF）和10 ng/mL碱性成纤维细胞生长因子（bFGF）的增殖反应。采用免疫印迹法对p44/42-丝裂原活化蛋白激酶（MAPK）的磷酸化进行定量。在有或无基质胶基底膜屏障（英国牛津BD生物科学公司）存在的情况下，使用改良的博伊登小室技术对配对SMC的侵袭和迁移速率进行定量。通过明胶酶谱法分析侵袭试验的条件培养基中MMP-2和MMP-9的分泌情况。
7天增殖试验曲线下面积分析显示，SV-SMC对FCS、PDGF和bFGF反应的细胞数量分别比配对的IMA-SMC高2.1倍、2.0倍和2.3倍。SV-SMC的基础MAPK激活水平约为配对IMA-SMC的两倍。与IMA-SMC相比，SV-SMC的侵袭率（有基质胶屏障）增加了２.1倍，但两种细胞类型的迁移速率（无基质胶屏障）以及MMP-2和MMP-9分泌情况相似。
人SV-SMC本质上比配对的IMA-SMC更具增殖性和侵袭性，这可能是由于p４４/４２-MAPK激活相对增加所致。不同来源SMC之间的这些内在功能差异可能导致与IMA移植物相比，SV移植物中内膜增生的发生率更高。
