Aquaporin 1 (AQP1) is a plasma membrane water-transporting protein expressed strongly in tumor microvascular endothelia. We previously reported impaired angiogenesis in implanted tumors in AQP1-deficient mice and reduced migration of AQP1-deficient endothelial cells in vitro. Here, we investigated the consequences of AQP1 deficiency in mice that spontaneously develop well-differentiated, luminal-type breast adenomas with lung metastases [mouse mammary tumor virus-driven polyoma virus middle T oncogene (MMTV-PyVT)]. AQP1(+/+) MMTV-PyVT mice developed large breast tumors with total tumor mass 3.5 ± 0.5 g and volume 265 ± 36 mm(3) (SE, 11 mice) at age 98 d. Tumor mass (1.6±0.2 g) and volume (131±15 mm(3), 12 mice) were greatly reduced in AQP1(-/-) MMTV-PyVT mice (P<0.005). CD31 immunofluorescence showed abnormal microvascular anatomy in tumors of AQP1(-/-) MMTV-PyVT mice, with reduced vessel density. HIF-1α expression was increased in tumors in AQP1(-/-) MMTV-PyVT mice. The number of lung metastases (5±1/mouse) was much lower than in AQP1(+/+) MMTV-PyVT mice (31±8/mouse, P<0.005). These results implicate AQP1 as an important determinant of tumor angiogenesis and, hence, as a potential drug target for adjuvant therapy of solid tumors.

SUMMARY Urea transporter (UT) proteins, including UT-A in kidney tubule epithelia and UT-B in vasa recta microvessels, facilitate urinary concentrating function. A screen for UT-A inhibitors was developed in MDCK cells expressing UT-A1, water channel aquaporin-1, and YFP-H148Q/V163S. An inwardly directed urea gradient produces cell shrinking followed by UT-A1-dependent swelling, which was monitored by YFP-H148Q/V163S fluorescence. Screening of ~90,000 synthetic small molecules yielded four classes of UT-A1 inhibitors with low micromolar IC50 that fully and reversibly inhibited urea transport by a non-competitive mechanism. Structure-activity analysis of >400 analogs revealed UT-A1-selective and UT-A1/UT-B non-selective inhibitors. Docking computations based on homology models of UT-A1 suggested inhibitor binding sites. UT-A inhibitors may be useful as diuretics (‘urearetics’) with a novel mechanism of action that may be effective in fluid-retaining conditions in which conventional salt transport-blocking diuretics have limited efficacy.

absorption and urinary exosomal excretion of sodium transporters, and (2) the profile of sodium transporter excretion related to blood pressure (BP) changes with salt intake. A 24-hour ambulatory BP monitoring and a 24-hour urine collection were performed after 1 week on a low-and 1 week on a high-salt diet. Results: Animal studies: urinary NKCC2 and NCC excretion rates correlated well with their abundance in the kidney. Human studies: 6 patients (15%) were classified as salt sensitive. The NKCC2 and NCC abundance did not decrease after the high-salt period, when the urinary sodium reabsorption decreased from 99.7 to 99.0%. In addition, the changes in BP with salt intake were not associated with a specific profile of exosomal excretion. Conclusions: Our results do not support the idea that excretion levels of NKCC2 and NCC via urinary exosomes are markers of tubular sodium reabsorption in hypertensive patients.Copyright © 2010 S. Karger AG, Basel Key WordsExosomes ؒ Na-Cl cotransporter ؒ Na-K-2Cl cotransporter ؒ Renal sodium transporters ؒ Salt sensibility ؒ Urine biomarkers Abstract Background: Altered renal sodium handling has a major pathogenic role in salt-sensitive hypertension. Renal sodium transporters are present in urinary exosomes. We hypothesized that sodium transporters would be excreted into the urine in different amounts in response to sodium intake in salt-sensitive versus salt-resistant patients. Methods: Urinary exosomes were isolated by ultracentrifugation, and their content of Na-K-2Cl cotransporter (NKCC2) and Na-Cl cotransporter (NCC) was analyzed by immunoblotting. Animal studies: NKCC2 and NCC excretion was measured in 2 rat models to test whether changes in sodium transporter excretion are indicative of regulated changes in the kidney tissue. Human studies: in hypertensive patients (n = 41), we investigated: (1) a possible correlation between sodium re-

Several experimental models of cirrhosis have shown dysregulation of renal aquaporins in different phases of liver disease. We investigated the urinary excretion of both aquaporin-1 and aquaporin-2 in patients with cirrhosis at different stages of the disease. Twenty-fourhour urine was collected from 11 healthy volunteers, 13 patients with compensated cirrhosis (without ascites), and 20 patients with decompensated cirrhosis (11 with ascites without renal failure and 9 with hepatorenal syndrome). Aquaporin-1 and aquaporin-2 excretion was analyzed by immunoblotting. Urinary aquaporin-2 excretion was reduced in patients with cirrhosis compared to healthy subjects. A progressive decrease in urinary aquaporin-2 excretion was observed as the severity of cirrhosis increased, from compensated cirrhosis to cirrhosis with ascites and hepatorenal syndrome. Patients with hyponatremia had lower urinary aquaporin-2 excretion than patients without hyponatremia. Vasopressin plasma level did not correlate with aquaporin-2 excretion. There were no differences between healthy subjects and patients with cirrhosis with or without ascites in urinary excretion of aquaporin-1, but urinary aquaporin-1 excretion of those with hepatorenal syndrome was extremely low. In conclusion, patients with cirrhosis appear to exhibit a decreased abundance of renal aquaporin-2 and therefore lower water permeability in the collecting tubules. This may represent an adaptive renal response to sodium retention, with expansion of extracellular fluid volume and dilutional hyponatremia observed in those who have cirrhosis with ascites. Finally, aquaporin-1 does not appear to play a role in the progressive dysregulation of extracellular fluid volume in cirrhosis. (HEPATOLOGY 2006;44:1555-1563