Bile salt stimulates intestinal epithelial cell migration through TGFbeta after wounding.

BACKGROUND: In addition to aiding in the digestion of fats, luminal bile salts have been shown to modulate gastrointestinal epithelial growth, differentiation, and other functions. We hypothesized that bile acids could modulate the intestinal mucosal repair process of restitution. We investigated the effect of the bile salt taurodeoxycholic acid on epithelial migration and identified a role for TGFbeta, a widely expressed cytokine in the intestinal villus, in this repair process. METHODS: Using a well-established model of epithelial restitution, IEC-6 cells were plated on 60-mm Matrigel-coated plastic dishes and grown to confluence. The epithelium was wounded by scraping with a 6-mm-wide blade to create a smooth denuded edge and cell migration was measured 8 h later. Cells were grown in control DMEM with 5% FBS with or without 0.01-2 mM taurodeoxycholic acid (TDCA). In parallel experiments, cells were harvested for Northern analysis of TGFbeta and GAPDH expression; [3H]thymidine uptake was used to measure proliferation. Anti-TGFbeta antibody was added to cells grown in the presence of 0.05 mM TDCA and migration was measured at 8 h. RESULTS: TDCA at physiologic luminal concentrations augments IEC-6 cell migration, with a maximal effect at 0.05 mM. TDCA inhibited proliferation at these concentrations. TGFbeta expression increased in response to bile acid, while wounding had less of an effect on TGFbeta expression. Blockade of TGFbeta function with TGFbeta antibody eliminated the effect of bile on cell migration. CONCLUSIONS: Bile acid at physiologic concentrations augments small intestinal epithelial cell migration. The process is dependent on TGFbeta and is independent of cell division. The data further support a role for bile acids and TGFbeta in differentiated intestinal cell function and in preservation of an intact mucosa.

Ca2+-RhoA signaling pathway required for polyamine-dependent intestinal epithelial cell migration.

Expression of voltage-gated K(+) (Kv) channel genes is regulated by polyamines in intestinal epithelial cells (IEC-6 line), and Kv channel activity is involved in the regulation of cell migration during early restitution by controlling membrane potential (E(m)) and cytosolic free Ca2+ concentration ([Ca2+](cyt)). This study tests the hypothesis that RhoA of small GTPases is a downstream target of elevated ([Ca2+](cyt)) following activation of K(+) channels by increased polyamines in IEC-6 cells. Depletion of cellular polyamines by alpha-difluoromethylornithine (DFMO) reduced whole cell K+ currents [I(K(v))] through Kv channels and caused membrane depolarization, which was associated with decreases in ([Ca2+](cyt)), RhoA protein, and cell migration. Exogenous polyamine spermidine reversed the effects of DFMO on I(K(v)), E(m), ([Ca2+](cyt)), and RhoA protein and restored cell migration to normal. Elevation of ([Ca2+](cyt)) induced by the Ca2+ ionophore ionomycin increased RhoA protein synthesis and stimulated cell migration, while removal of extracellular Ca2+ decreased RhoA protein synthesis, reduced protein stability, and inhibited cell motility. Decreased RhoA activity due to Clostridium botulinum exoenzyme C(3) transferase inhibited formation of myosin II stress fibers and prevented restoration of cell migration by exogenous spermidine in polyamine-deficient cells. These findings suggest that polyamine-dependent cell migration is partially initiated by the formation of myosin II stress fibers as a result of Ca2+-activated RhoA activity.

Glypican-3 expression in Wilms tumor and hepatoblastoma.

BACKGROUND: Glypican-3 (GPC3) is a heparan sulfate proteoglycan. When it is disrupted, it causes the X-linked gigantism-overgrowth Simpson-Golabi-Behmel syndrome. Its involvement in growth control is consistent with recent reports that it can bind to growth factors, possibly including insulin-like growth factor 2. Further, it has been hypothesized that it may function as a tumor suppressor gene in breast and ovarian carcinomas and mesotheliomas. PATIENTS AND METHODS: RNA and protein were extracted from Wilms tumor and hepatoblastoma tissue samples and GPC3 levels were measured in these extracts by Northern blotting, reverse transcription polymerase chain reaction, and immunoblotting. RESULTS: In contrast to published results with carcinomas, high levels of GPC3 expression were found in Wilms tumor and hepatoblastoma. Low or undetectable expressions of this gene were found in normal tissue surrounding the tumor. CONCLUSIONS: Increased expression of GPC3 in Wilms tumor and hepatoblastoma suggests a growth-promoting or neutral activity for this gene product rather than a growth-suppressive effect.

Retropsoas hernia as a cause of chronic abdominal pain: CT diagnosis.

Congenital retropsoas small bowel herniation is reported as the cause of long-standing recurrent abdominal pain in a teenage girl. Knowledge of this entity is important for differential diagnosis of abdominal pain, mass, or retroperitoneal gas and fluid, and for avoiding complications of percutaneous renal interventions.

Combined hyperthermia and immunotherapy treatment of multiple pulmonary metastases in mice.

The combination of immunotherapy and hyperthermia results in a greater reduction in tumour growth compared to either therapy used alone in a murine subcutaneous tumour model. To evaluate this combination further we tested it in a murine pulmonary metastasis model. Mice were given 5 x 10(5) MCA-105 sarcoma cells on day 0 intravenously resulting in the formation of pulmonary metastases. Mice were treated with local hyperthermia to the left hemithorax with a transcutaneous microwave applicator or with whole-body hyperthermia to 41 degrees C for 30 min on days 3 and 6. Immunotherapy included 5 x 10(7) syngeneic LAK cells administered on days 3 and 6 and interleukin-2 given intraperitoneally three times daily on days 3-7. Animals were killed on day 12 and pulmonary nodules enumerated. While the addition of whole-body hyperthermia to immunotherapy had no significant effect on tumour growth, the combination of local hyperthermia and immunotherapy significantly decreased the number of pulmonary nodules by 94% compared to controls in combined experiments. The mechanism of this beneficial effect may be related to increased trafficking of immune active cells to the tumour-bearing site, an increase in the sensitivity of tumour cells to lysis, or perhaps a local release of cytokines induced by hyperthermia. This study established the efficacy of combined immunotherapy and hyperthermia for the treatment of visceral metastases and provides impetus for the initiation of clinical trials.