Nitric oxide (NO) plays a critical role in vascular endothelial growth factor (VEGF)-induced angiogenesis and vascular hyperpermeability. However, the relative contribution of different NO synthase (NOS) isoforms to these processes is not known. Here, we evaluated the relative contributions of endothelial and inducible NOS (eNOS and iNOS, respectively) to angiogenesis and permeability of VEGF-induced angiogenic vessels. The contribution of eNOS was assessed by using an eNOS-deficient mouse, and iNOS contribution was assessed by using a selective inhibitor [L-N 6 -(1-iminoethyl) lysine, L-NIL] and an iNOS-deficient mouse. Angiogenesis was induced by VEGF in type I collagen gels placed in the mouse cranial window. Angiogenesis, vessel diameter, blood flow rate, and vascular permeability were proportional to NO levels measured with microelectrodes: Wild-type (WT) > WT with L-NIL or iNOS ؊/؊ > eNOS ؊/؊ > eNOS ؊/؊ with L-NIL. The role of NOS in VEGF-induced acute vascular permeability increase in quiescent vessels also was determined by using eNOS-and iNOS-deficient mice. VEGF superfusion significantly increased permeability in both WT and iNOS ؊/؊ mice but not in eNOS ؊/؊ mice. These findings suggest that eNOS plays a predominant role in VEGF-induced angiogenesis and vascular permeability. Thus, selective modulation of eNOS activity is a promising strategy for altering angiogenesis and vascular permeability in vivo.

The large size of many novel therapeutics impairs their transport through the tumor extracellular matrix and thus limits their therapeutic effectiveness. We propose that extracellular matrix composition, structure, and distribution determine the transport properties in tumors. Furthermore, because the characteristics of the extracellular matrix largely depend on the tumor-host interactions, we postulate that diffusion of macromolecules will vary with tumor type as well as anatomical location. Diffusion coefficients of macromolecules and liposomes in tumors growing in cranial windows (CWs) and dorsal chambers (DCs) were measured by fluorescence recovery after photobleaching. For the same tumor types, diffusion of large molecules was significantly faster in CW than in DC tumors. The greater diffusional hindrance in DC tumors was correlated with higher levels of collagen type I and its organization into fibrils. For molecules with diameters comparable to the interfibrillar space the diffusion was 5-to 10-fold slower in DC than in CW tumors. The slower diffusion in DC tumors was associated with a higher density of host stromal cells that synthesize and organize collagen type I. Our results point to the necessity of developing site-specific drug carriers to improve the delivery of molecular medicine to solid tumors.

NO has been shown to mediate angiogenesis; however, its role in vessel morphogenesis and maturation is not known. Using intravital microscopy, histological analysis, α-smooth muscle actin and chondroitin sulfate proteoglycan 4 staining, microsensor NO measurements, and an NO synthase (NOS) inhibitor, we found that NO mediates mural cell coverage as well as vessel branching and longitudinal extension but not the circumferential growth of blood vessels in B16 murine melanomas. NO-sensitive fluorescent probe 4,5-diaminofluorescein imaging, NOS immunostaining, and the use of NOS-deficient mice revealed that eNOS in vascular endothelial cells is the predominant source of NO and induces these effects. To further dissect the role of NO in mural cell recruitment and vascular morphogenesis, we performed a series of independent analyses. Transwell and under-agarose migration assays demonstrated that endothelial cell-derived NO induces directional migration of mural cell precursors toward endothelial cells. An in vivo tissue-engineered blood vessel model revealed that NO mediates endothelial-mural cell interaction prior to vessel perfusion and also induces recruitment of mural cells to angiogenic vessels, vessel branching, and longitudinal extension and subsequent stabilization of the vessels. These data indicate that endothelial cell-derived NO induces mural cell recruitment as well as subsequent morphogenesis and stabilization of angiogenic vessels.

Angiogenesis inhibitors produced by a primary tumor can create a systemic anti-angiogenic environment and maintain metastatic tumor cells in a state of dormancy. We show here that the gallbladder microenvironment modulates the production of transforming growth factor (TGF)-beta1, a multifunctional cytokine that functions as an endogenous anti-angiogenic and anti-tumor factor in a cranial window preparation. We found that a wide variety of human gallbladder tumors express TGF-beta1 irrespective of histologic type. We implanted a gel impregnated with basic fibroblast growth factor or Mz-ChA-2 tumor in the cranial windows of mice without tumors or mice with subcutaneous or gallbladder tumors to study angiogenesis and tumor growth at a secondary site. Angiogenesis, leukocyte-endothelial interaction in vessels and tumor growth in the cranial window were substantially inhibited in mice with gallbladder tumors. The concentration of TGF-beta1 in the plasma of mice with gallbladder tumors was 300% higher than that in the plasma of mice without tumors or with subcutaneous tumors. In contrast, there was no difference in the plasma levels of other anti- and pro-angiogenic factors. Treatment with neutralizing antibody against TGF-beta1 reversed both angiogenesis suppression and inhibition of leukocyte rolling induced by gallbladder tumors. TGF-beta1 also inhibited Mz-ChA-2 tumor cell proliferation. Our results indicate that the production of anti-angiogenesis/proliferation factors is regulated by tumor-host interactions.

BackgroundIntestinal duplication, a congenital malformation, is considered a rare condition, particularly in adults. Although it affects young children, a minority of patients remains asymptomatic until adulthood. Here, we describe a case of an intestinal duplication cyst that caused intussusception by a unique mechanism.Case presentationA 19-year-old man was admitted to our hospital for intermittent abdominal pain. Computed tomography revealed colonic intussusception induced by a nodular mass in the ileocecal region. Urgent ileocecal resection was performed because of the risk of colonic ischemia. The resected material comprised a stool-filled noncommunicating cyst that protruded into the enteric lumen at the ileocecal valve. Histological analyses revealed that the inner wall of the cyst was lined with colonic mucosa and that the muscle layer of the cyst was shared with that of the original enteric wall; furthermore, the cyst had a vestige of an opening site in the wall. We concluded that the cyst was an intestinal duplication that poured stool into its lumen through the tiny orifice, thereby triggering intussusception.ConclusionsThe present case suggests that stool-pouring can cause intussusception into the space of an intestinal duplication lesion.

We report a case of a woman with a metastatic liver tumor from gastric carcinoma, who has been successfully treated with concurrent proton beam therapy and systemic chemotherapy. A 76-year-old woman underwent distal gastrectomy with regional lymph node dissection for advanced gastric carcinoma on January 17, 2002. She received five courses of sequential chemotherapy with methotrexate-5-fluorouracil after the surgical resection. A metastatic liver tumor was detected in the caudate lobe of the liver by computed tomography at 6 months after the surgical resection. We employed concurrent proton beam therapy and systemic chemotherapy which consisted of 5-fluorouracil (250 mg/body per day, as a 24-h intravenous injection for 4 weeks) and low dose cisplatin (10 mg/body on days 1-5 every week for 4 weeks). Proton beam therapy targeting the metastatic liver tumor was performed in a daily fraction of 3 Gy, 5 days per week, with a total dose of 66 Gy over 30 days. The tumor disappeared 3 months after the treatment and no recurrence has been observed for 2 years after termination of the treatment. Throughout the entire course of treatment, the patient received injections of granulocyte stimulating factor subcutaneously for grade 3 leukopenia. She never complained of abdominal symptoms, such as epigastralgia, nausea or diarrhea. Liver failure related to proton irradiation has not been observed. This concurrent proton beam radiotherapy with systemic chemotherapy could be an effective treatment modality for metastatic liver tumor from gastric carcinoma.

Gastric carcinosarcomas are rare morphologically biphasic tumors, consisting of carcinoma and sarcoma components, with a poor clinical course. Here we report the case of a 70-year-old man with advanced Borrmann type III carcinosarcoma arising from the upper body of the stomach with extensive lymph node metastasis who underwent a total, but palliative, gastrectomy. Histology showed the tumor consisted of a biphasic structure of tubular adenocarcinoma and spindle cell sarcoma. Immunohistochemistry revealed sarcoma cells expressing c-kit (CD117) and CD34, which are criteria for gastrointestinal stromal tumors. Nine months after the surgical operation, tumor metastases had extended to the hepatohilar, retroperitoneal and mediastinal lymph nodes. Radiation therapy of 50 Gy markedly decreased the size of each of these nodes and reduced the risk of respiratory complications and jaundice. However, the patient died of respiratory failure due to bronchopneumonia with multiple lung metastases 22 mo after resection. Autopsy revealed severe necrosis in most of the lymph nodes with tumor metastases. Radiation therapy combined with gastrectomy should be considered to improve survival in patients with gastric carcinosarcomas that express c-kit.

A case of extensive extra- and intrahepatic portal tumor thrombosis, with no metastatic foci in liver parenchyma, secondary to advanced gastric carcinoma in a 69-year-old man is reported. The portal tumor thrombosis was characterized by enlargement of the thrombosed segment of the vein, decreased density mass without intraluminal enhancement of the involved vein, nonvisualization of the portal venous branch in the involved lobe, and the so-called cavernous transformation of the portal vein. The surgically resected gastric specimen showed Borrmann type 3 advanced papillary adenocarcinoma. The portal tumor thrombus is presumed to have arisen from vascular invasion in the primary foci of gastric carcinoma, and then to have permeated the portal vein without invasion of liver parenchyma.