THE L-ARGININE: NO PATHWAY IN MAMMARY TUMOR PROGRESSION

尽管已有共识认为一氧化氮(NO)调节肿瘤的发展，但它是抑制还是刺激血管生成和转移扩散还没有定论。NO的细胞抑制作用支持了它具有抗肿瘤作用的信念。然而，来自不可比较的实验系统的相互矛盾的数据使得NO在肿瘤进展中的作用悬而未决。因此，需要对NO在肿瘤进展中的作用进行全面研究。它需要一个适当的模型系统来考虑两个协同产生NO的事件：(I)NOS2基因表达的激活，和(Ii)足够数量的NO底物L-精氨酸的运输。一个非常有用的肿瘤模型系统采用了多瘤中T癌基因(PYV-Mt)。它能在100%的女性中诱发快速、可复制、多灶性转移的乳腺腺癌。持续和丰富的NO产生是通过转录控制的诱导型一氧化氮合酶(NOS2)酶来实现的。转运的唯一底物为NO的产生，L-精氨酸是由特定的转运蛋白(CAT)介导的。当PYV-mt在NOS2-/-和Cat2-/-小鼠中表达时，这些模型用于确定它们在乳腺肿瘤进展中的单独角色。我们推测CAT2精氨酸转运体在通过NOS2调节NO的产生中起核心作用。由NOS2产生的NO和通过CAT2转运的精氨酸的后果将通过测量乳腺癌的发生、生长、血管生成和转移来评估，方法是使用“设计”的双基因和三基因小鼠。PYV-mt转基因是受体酪氨酸激酶(特别是cErbB2)的替代品，是一种高度致癌的转基因。然而，cErbB2转基因是一种更惰性的癌基因，表现出较慢的肿瘤动力学。移植研究将确定NO对乳腺肿瘤形成的刺激作用是否是上皮细胞自主的。或者，移植实验将揭示循环和/或间质组织是否对观察到的NOS2-/-和NOS2+/+小鼠之间的肿瘤进展差异负责。微血管密度的分析将用于测量血管生成，并将评估细胞凋亡的程度。CAT2介导的L精氨酸转运对NO产生的作用将在肿瘤细胞系和适当的荷瘤小鼠中进行评估。建议的实验直接测试NOS2和Cat2基因在该模型中对血管生成、肿瘤生长和转移的作用。这些模型系统显示的乳腺肿瘤进展与人类乳腺癌的发展具有重要的形态和病因学相似之处。因此，所获得的知识将是有价值的，因为精氨酸的运输和NO的产生都可以通过改变NOS2活性或底物通量的饮食和/或精氨酸类似物来调节。

Although there is consensus that nitric oxide (NO) modulates tumor development, it is unsettled whether it inhibits or stimulates angiogenesis and metastatic spread. The cytostatic effect of NO lends support to the belief that it has antitumor action. However, conflicting data from non-comparable experimental systems leave unsettled the effect of NO on tumor progression. Hence, a comprehensive investigation of the effects of NO on tumor progression is needed. It requires an adequate model system that takes into account two coordinate events that cooperate to produce NO: (i) The activation of NOS2 gene expression, and (ii) The transport of adequate amounts of the NO substrate L-arginine. A highly useful tumor model system employs the polyoma middle T oncogenic transgene (PyV-mT). It elicits rapid, reproducible, multifocal metastatic mammary adenocarcinoma in 100 percent of females. Sustained and copious NO production occurs via the transcriptionally controlled inducible nitric oxide synthase (NOS2) enzyme. Transport of the sole substrate for NO production, L-arginine is mediated by specific transporters (CATs). When PyV-mT is expressed in either Nos2-/- and Cat2-/- mice, the models serve to define their individual roles in breast tumor progression. We postulate that the CAT2 arginine transporter plays a central role in the regulated production of NO via NOS2. The consequences of NO produced from NOS2 and arginine transport via CAT2, will be assessed by measuring the occurrence, growth, angiogenesis and metastasis of mammary adenocarcinoma using "designer" bigenic and trigenic mice. The PyV-mT transgene is a surrogate for receptor tyrosine kinase (specifically cErbB2) and is an extremely oncogenic transgene. However, the cErbB2 transgene is a more indolent oncogene exhibiting slower tumor kinetics. Transplantation studies will determine whether the stimulatory effect of NO on mammary tumor formation is epithelial cell autonomous. Alternatively, transplant experiments will reveal if circulating and/or stromal tissues are responsible for the differences in tumor progression observed between Nos2-/- and Nos2+/+ mice. An analysis of the microvascular density will be used to measure angiogenesis and the extent of apoptosis will be assessed. The role of CAT2 mediated L-arginine transport on NO production will be assessed in tumor cell lines and in appropriate tumor bearing mice. The proposed experiments directly test the role of Nos2 and Cat2 genes on angiogenesis, tumor growth, and metastasis in this model. The mammary tumor progression exhibited by these model systems have important morphological and etiological similarities with human breast cancer development. Hence, the knowledge gained will be worthwhile since both the transport of arginine and the production of NO can be modulated by diet and/or arginine analogues that modify NOS2 activity or substrate flux.