Purpose: Ion channel activity is involved in several basic cellular behaviors that are integral to metastasis (e.g., proliferation, motility, secretion, and invasion), although their contribution to cancer progression has largely been ignored. The purpose of this study was to investigate voltagegated Na + channel (VGSC) expression and its possible role in human breast cancer. Experimental Design: Functional VGSC expression was investigated in human breast cancer cell lines by patch clamp recording. The contribution of VGSC activity to directional motility, endocytosis, and invasion was evaluated by in vitro assays. Subsequent identification of the VGSC a-subunit(s) expressed in vitro was achieved using reverse transcription-PCR, immunocytochemistry, and Western blot techniques and used to investigate VGSCa expression and its association with metastasis in vivo. Results:VGSC expression was significantly up-regulated in metastatic human breast cancer cells and tissues, and VGSC activity potentiated cellular directional motility, endocytosis, and invasion. Reverse transcription-PCR revealed that Na v 1.5, in its newly identified ''neonatal'' splice form, was specifically associated with strong metastatic potential in vitro and breast cancer progression in vivo. An antibody specific for this form confirmed up-regulation of neonatal Na v 1.5 protein in breast cancer cells and tissues. Furthermore, a strong correlation was found between neonatal Na v 1.5 expression and clinically assessed lymph node metastasis. Conclusions: Up-regulation of neonatal Na v 1.5 occurs as an integral part of the metastatic process in human breast cancer and could serve both as a novel marker of the metastatic phenotype and a therapeutic target.Breast cancer is the most common cancer of women and the second leading cause of female cancer mortality, accounting for about 10% of all cancer deaths in the western world (1, 2). To date, several breast cancer metastasis -associated genes have been identified both individually and in combination in microarray analyses (3, 4). These include oncogenes (e.g., ras and c-myc), cell cycle -associated markers (e.g., Ki67), adhesion molecules (e.g., E-cadherins), motility factors (e.g., hepatic growth factor), growth factors and their receptors (e.g., epidermal growth factor/Her-2 and fibroblast growth factor), and the well-established steroid hormones (e.g., estrogen and progesterone; refs. 3, 4). However, indirect measures of metastatic progression (including size of primary carcinoma, assessment of intratumoral vascular invasion, and lymph node involvement) remain the most widely used methods in clinical management. At present, although it is possible to detect micrometastases, approximately one third of women who seem disease-free at primary diagnosis eventually develop overt metastases (5, 6). Clinicians, therefore, require a more accurate diagnosis to predict the development of metastatic disease.Ion channels are major signaling molecules expressed in a wide range of tissues where they hav...

Background: In the present study, a technique for computer-assisted, normothermic, oxygenated, ex vivo, recirculating small bowel perfusion was established as a tool to investigate organ pretreatment protocols and ischemia/reperfusion phenomena. A prerequisite for the desired setup was an organ chamber for ex vivo perfusion and the use of syngeneic whole blood as perfusate. Methods: The entire small bowel was harvested from Lewis rats and perfused in an organ chamber ex vivo for at least 2 h. The temperature was kept at 37°C in a water bath. Three experimental groups were explored, characterized by different perfusion solutions. The basic perfusate consisted of syngeneic whole blood diluted with either NaCl, Krebs’ solution or Krebs’ solution and norepinephrine to a hematocrit of 30%. In addition, in each group l-glutamine was administered intraluminally. The desired perfusion pressure was 100 mm Hg which was kept constant with a computer-assisted data acquisition software, which measured on-line pressure, oxygenation, flow, temperature and pH and adjusted the pressure by changing the flow via a peristaltic pump. The viability of the preparation was tested by measuring oxygen consumption and maltose absorption, which requires intact enzymes of the mucosal brush border to break down maltose into glucose. Results: Organ perfusion in group 1 (dilution with NaCl) revealed problems such as hypersecretion into the bowel lumen, low vascular resistance and no maltose uptake. In contrast a viable organ could be demonstrated using Krebs’ solution as dilution solution. The addition of norepinephrine led to an improved perfusion over the entire perfusion period. Maltose absorption was comparable to tests conducted with native small bowel. Oxygen consumption was stable during the 2-hour perfusion period. Conclusions: The ex vivo perfusion system established enables small bowel perfusion for at least 2 h. The viability of the graft could be demonstrated. The perfusion time achieved is sufficient to study leukocyte/lymphocyte interaction with the endothelium of the graft vessels. In addition, a viable small bowel, after 2 h of ex vivo perfusion, facilitates testing of pretreatment protocols for the reduction of the immunogenicity of small bowel allografts.

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