Analysis of the lymphatic development using mutant mice

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The lymphatics are thought to be responsible for edematous condition in patients, especially in patients suffering from primary lymphedema. Resent studies show that lymphangiogenesis, as well as angiogenesis, also plays some roles on tumor metastasis. However, the lymphatic development in mammals has been unknown from lack of reliable anatomical / histological procedure for the detection of lymphatic vassals and useful mutant animal that has obvious lymphatic abnormality.In order to understand the mechanism of lymphatic development and functions, we are generating "reporter" strains by transgenesis or targeted mutagenesis, using marker genes (lacZ, GFP, PLAP) under transcriptional regulation of the lymphatic endothelial-specific genes, such as LYVE-1, prox-1 and VFGFR-3. We have cloned mouse LYVE-1 gene and determined the genomic structure and chromosomal location of the gene in mice. For the construction of the transgenes using Prox-1 gene or VEGFR-3 gene, we purchased and re-analyzed their P1 or BAC clones. We are now introducing some constructs into mouse fertilized eggs. We have also started on the conditional knockout analyses of potential growth factor / receptor and transcriptional factors in lymphangiogenesis.We are also under investigation of an original spontaneous mutant mouse line developing chylous ascites, intestinal lymphedema and edematous hindfoot that are thought to be due to lymphatic abnormality. The blood flow is found not only in blood vessels but also in lymphatic vessels in the homozygous mutants. According to the confocal microscopic analysis of TRITC-conjigated gelatin injected into blood vessels, the peripheral capillary-lacteal shunt at the intestinal villi is one of the cause for blood flow observed lymphatics of the homozygous mutant mice. We are trying forward genetic approaches to find the candidate gene (s) for this mutation.