Modification of Athferoma by cytokin gene transfer

我们建立了大鼠双球囊损伤模型，并用免疫组织化学和RT-PCR方法研究增厚的新生内膜。大鼠左侧颈总动脉球囊损伤后14天，在同一部位造成第二次球囊损伤。单球囊损伤和双球囊损伤后14d取材，行免疫组织化学染色，计算内膜/中膜面积比，作为新生内膜形成的指标。提取部分动脉总RNA，用RT-PCR方法检测细胞外基质(ECM)及与ECM代谢相关的细胞因子受体的表达。大鼠双球囊损伤模型(II)的I/M比值(1.84±0.62，均值±SE，n=5)显著高于单球囊损伤模型(I)的I/M比值(1.30±0.19，n=10)。在平滑肌细胞表型方面，α-Actin染色显示，II组新生内膜以收缩为主多于合成为主，而I组以合成为主。II组新生内膜中层粘连蛋白和纤维连接蛋白呈强阳性染色，而I组新生内膜染色较弱。与此相一致的是，层粘连蛋白和纤维连接蛋白mRNAs在II组新生内膜中均有明显表达。I组和II组新生内膜中PDGF1(α和β)和转化生长因子-β(I型和II型)受体也呈相同程度的阳性表达。这些结果表明：细胞外基质尤其是层粘连蛋白和纤维粘连蛋白的积聚是双气囊损伤模型的特征；该模型中细胞外基质的显著积聚是由于除PDGf或转化生长因子-β信号通路外的其他机制；该模型类似于PTCA术后再狭窄的病变，因此为其研究提供了关键。

We developed a rat double-balloon injury model and studied the thickened neointima using immunohistochemical and RT-PCR methods. Fourteen days after the first balloon injury of the rat left common carotid artery, a second balloon injury was inflicted in the same place. Histochemical sections taken 14 days after the single- and double-balloon injuries were used for calculating intimal/medial (I/M) area ratios, as an indicator of neointimal formation, and were subjected to immunohistochemical staining. Total RNA was also purified from some arteries and mRNA expression of some extracellular matrices (ECM) and cytokine receptors related to ECM metabolism was estimated using the RT-PCR method. The I/M ratio in the rat double-balloon injury model (II) (1.84±0.62(mean±SE, n=5)) was significantly (p<0.05) higher than that in the single-balloon injury model (I) (1.30±0.19, n=10). The cell number per neointimal area was less in II than in I. As for the phenotype of smooth muscle cells, α-actin staining showed that the neointima of II consisted of more contractile form than synthetic, whereas that of I consisted of more synthetic form than contractile. The neointima of II was strongly stained with laminin and fibronectin, but that of I was stained only weakly. Consistent with these data, laminin and fibronectin mRNAs were markedly expressed in the neointima of II. Neointimas of both I and II were also stained positively with PDGF (α and β) and TGF-β (types I and II) receptors to the same extent. These results show: that ECM accumulation, particularly of laminin and fibronectin, characterizes the double-balloon injury model; that the marked accumulation of ECM in this model is due to a mechanism other than the PDGF or TGF-β signalling pathway; and that this model resembles the lesion of post-PTCA re-stenosis, and therefore provides the key for its investigation.