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  • Title: Increased expression of metastasis-related genes in hypoxic cells sorted from cervical and lymph nodal xenograft tumors.
    Author: Chaudary N, Hill RP.
    Journal: Lab Invest; 2009 May; 89(5):587-96. PubMed ID: 19308047.
    Abstract:
    Solid tumors contain regions of poor oxygenation that relate to the abnormal vascular network. Clinical investigations in cervical carcinoma have shown that positive lymph node status in patients with cervical carcinoma correlates with hypoxia. Earlier, in an orthotopic cervical cancer model, we had shown that exposure to acute hypoxia enhances lymph node metastasis. This study describes a technique for sorting hypoxic cells directly from the cervical xenograft model and reports the expression of 'metastasis-related' genes in hypoxic cells from xenografted cervix and lymph node tumors. Tumor cells were sorted on the basis of DsRed fluorescence and the sub-population of hypoxic cells was sorted on the basis of carbonic anhydrase-9 (CA-9) expression. Quantitative RT-PCR was conducted to measure changes in gene expression in the hypoxic cells sorted from primary cervix tumors and lymph node metastases. Immunohistochemistry was used to track changes in protein expression in sections of the same tumors. Metastasis-related genes, CXCR4, uPAR, VEGFC, Hdm2, and OPN, were observed to be upregulated at gene and protein levels in the primary tumors and nodal metastasis from the orthotopic transplants. In particular, the hypoxic cells sorted from orthotopically transplanted cervix tumors and their lymph node metastases from mice exposed to cyclic (intermittent) hypoxia showed higher levels of expression of these genes. These results are consistent with the hypothesis that these genes may be involved in regulating lymph node metastasis in cervical cancers under hypoxic conditions and provide support to the concept cyclic hypoxia that plays an important role in this process. Our methodological study emphasizes the technique of cell sorting to identify hypoxic cells using CA-9, which may aid in improving prognostic capabilities and in designing rational therapeutic strategies by focusing on hypoxia-specific gene expression profiles of patients. The technique can be applied to identify other potential 'hypoxia-related' genes of interest for tumor growth and metastasis.
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