THE L-ARGININE: NO PATHWAY IN MAMMARY TUMOR PROGRESSION

虽然一氧化氮（NO）调节肿瘤的发展已达成共识，但它是否抑制或刺激血管生成和转移扩散尚不清楚。一氧化氮的细胞抑制作用支持了它具有抗肿瘤作用的观点。然而，来自非可比性实验系统的相互矛盾的数据未确定NO对肿瘤进展的影响。因此，需要对NO对肿瘤进展的影响进行全面的研究。它需要一个适当的模型系统，考虑到协同产生NO的两个协调事件：(i) NOS2基因表达的激活，以及（ii）足够量的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多瘤中间T致癌转基因（PyV-mT）。它在100%的女性中引起快速、可重复、多灶性转移性乳腺腺癌。通过转录控制的诱导型一氧化氮合酶（NOS2）产生持续和丰富的NO。生产NO的唯一底物，l -精氨酸的运输是由特异性转运体（CATs）介导的。当PyV-mT在Nos2-/-和Cat2-/-小鼠中表达时，该模型用于确定它们在乳腺肿瘤进展中的个体作用。我们假设CAT2精氨酸转运体在通过NOS2调节NO的产生中起核心作用。利用“设计”双基因和三基因小鼠，通过测量乳腺腺癌的发生、生长、血管生成和转移来评估NOS2产生的NO和精氨酸通过CAT2运输的后果。PyV-mT转基因是酪氨酸激酶受体（特别是cErbB2）的替代品，是一种极具致癌性的转基因。然而，cErbB2转基因是一种较惰性的癌基因，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。另外，移植实验将揭示循环和/或间质组织是否对Nos2-/-和Nos2+/+小鼠之间观察到的肿瘤进展差异负责。微血管密度分析将用于测量血管生成，并评估细胞凋亡的程度。CAT2介导的l -精氨酸运输对NO生成的作用将在肿瘤细胞系和适当的荷瘤小鼠中进行评估。本实验直接检测Nos2和Cat2基因在该模型血管生成、肿瘤生长和转移中的作用。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学和病因学相似性。因此，所获得的知识将是有价值的，因为精氨酸的运输和一氧化氮的产生都可以通过饮食和/或精氨酸类似物来调节，从而改变NOS2活性或底物通量。

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.