The mechanism of matrix-synthesis and-degradation in vascular smooth muscle cells.

为了解血管平滑肌细胞(SMC)表型转化与基质重塑过程的关系，建立了SMC表型转化的体外培养模型。通过血清浓度和最终细胞密度的变化来验证培养条件。用Western blotting和酶免疫方法检测基质金属蛋白酶(MMPs)及其抑制物(TIMPs)的表达。测定了凝胶原活性和凝胶溶解活性。在表现为增殖性表型的细胞中，MMP1、TIMP-1-3和MMP1rIMP-1的表达和活性最高。MMP2的表达与SMC表型无关。制备了表达TIMP-1的腺病毒载体，并利用该载体对SMC的表型或功能进行了分析，细胞外基质的量也通过细胞周期调控基因的功能来调节SMC的表型。表达模式…无论是在培养的SMC还是在人类动脉粥样硬化病变中，都观察到p21Waf-1，周期蛋白依赖的激酶抑制物的Rn增加。P2lWaf-1在增殖细胞中的表达增强，符合G0-G1转变的规律。在体内，该基因的表达主要在新生内膜动脉粥样硬化病变中检测到。P21Waf-1可能不仅在细胞周期停滞中起作用，而且在适当的细胞周期进程中也起作用。携带p21Waf-1的腺病毒载体可诱导SMC细胞周期停滞和肥大，但不能促进细胞的再分化和凋亡。结论：基质降解是SMC的重要功能，与SMC表型协同调节。这些SMC功能可能由内源性和外源性生长因子信号介导。增殖被认为受到细胞周期调节蛋白的严格调控，如p21Waf-1，其功能可能调节SMC的表型。这些发现可能有助于理解血管平滑肌细胞的生物学。本项目所采用的腺病毒介导的基因转移方法，可能为临床改变人类动脉粥样硬化的治疗提供参考。较少

To understanding the relationship between phenotypic modulation of vascular smooth muscle cells (SMC), and matrix remodeling process, in vitro culture model for the phenotypic modulation of SMC was established. The culture conditions were verified by the alteration of serum-concentration and final cell densities. The expressions of matrix metalloproteinases (MMPs) and their inhibitors (TIMPs) were determined with Western blotting and enzyme immunoassays. Collogenolytic activity and gelationolytic activity were also measured. The Expressions and activities of MMP-1, TIMP-1-3, and MMP-1rrIMP-1 expressions were maximal in the cells displaying the proliferative phenotype. MMP-2 expression was not changed with SMC phenotypes. The adenovirus vector expressing TIMP-1 was prepared and the analysis for SMC phenotypes or functions using this vector is addressing.The amount of extra-cellular matrix also regulates SMC phenotypes via the function of cell-cycle regulatory genes. The expression patte … More rn of p2lWaf-1, cyclin-dependent kinase inhibitor was observed in either cultured SMC or human atherosclrotic lesions. p2lWaf-1 expression was enhanced in the proliferating cells in accordance with G0-G1 transition. In vivo, the expression was dominantly detected in neo-intimal atherosclerotic lesions. p21Waf-1 may contribute not only in cell-cycle arrest, but also in an appropriate cell cycle progression. Adenovirus vector expressing p21Waf-1 induced cell cycle arrest and hypertrophy in SMC, but not promoted re-differentiation and apoptosis.In conclusion, Matrix-degradation is an important function of SMC, which is coordinately regulated with SMC phenotypes. These SMC functions may be mediated by both endogenous and exogenous growth factor signalings. Proliferation is considered to be tightly regulated by cell-cycle regulatory proteins, such as p21Waf-1, of which function may modulate SMC phenotypes. These finding may contribute to understanding the biology of vascular smooth muscle cells. The adenovirus-mediated gene transfer method used in this project, may imply clinical alteration of human atherosclerosis in the therapeutic implications. Less