Phenotype-dependent expression of cadherin 6B in vascular and visceral smooth muscle cells.

We used mRNA subtraction of differentiated and dedifferentiated smooth muscle cells (SMCs) to reveal the molecular mechanisms underlying the phenotypic modulation of SMCs. With this approach, we found that a 10 kb mRNA encoding a homotypic cell adhesion molecule, cadherin 6B, was strongly expressed in differentiated vascular and visceral SMCs, but not in the dedifferentiated SMCs derived from them. In vivo, cadherin 6B was expressed in vascular and visceral SMCs, in addition to brain, spinal cord, retina and kidney, at a late stage of chicken embryonic development. These results suggest that cadherin 6B is a novel molecular marker for vascular and visceral SMC phenotypes and is involved in the late differentiation of SMCs.

Calcification in untreated non-Hodgkin's lymphoma of the jejunum.

A 69-year-old Japanese female was admitted to our hospital due to a 2-month history of vomiting after eating. Examination of the small intestinal tract revealed a tumor with calcification in the inner portion, from the horizontal portion to the ascending portion of the duodenum, and jejunojejunostomy was performed. The pathological findings of the tumor gave a diagnosis of non-Hodgkin's lymphoma, diffuse small cleaved cell (Working Formulation classification), B cell type, of the jejunum. Calcification is rarely found in untreated malignant lymphoma and 15 cases of untreated malignant lymphoma with calcification have been reviewed.

Differentiated phenotype of smooth muscle cells depends on signaling pathways through insulin-like growth factors and phosphatidylinositol 3-kinase.

Under conventional culture conditions, smooth muscle cells display their phenotypic modulation from a differentiated to a dedifferentiated state. Here, we established a primary culture system of smooth muscle cells maintaining a differentiated phenotype, as characterized by expression of smooth muscle-specific marker genes such as h-caldesmon and calponin, cell morphology, and ligand-induced contractility. Laminin retarded the progression of dedifferentiation of smooth muscle cells. Insulin-like growth factors (IGF-I and IGF-II) and insulin markedly prolonged the differentiated phenotype, with IGF-I being the more potent. In contrast, serum, epidermal growth factor, transforming growth factors, and platelet-derived growth factors potently induced dedifferentiation compared with angiotensin II, arginine-vasopressin, and basic fibroblast growth factor. Using the present culture system, we investigated signaling pathways regulating a phenotype of smooth muscle cells. In cultured cells, IGF-I specifically activated phosphatidylinositol 3-kinase (PI3-kinase) and its downstream target, protein kinase B, but not mitogen-activated protein kinases. Specific inhibitors of PI3-kinase (wortmannin and LY294002) induced dedifferentiation of smooth muscle cells even when they were cultured on laminin under IGF-I-stimulated conditions. The sole effect of laminin to retard the dedifferentiation was completely blocked by anti-IGF-I antibody, and laminin promoted the endogenous expression of IGF-I in cultured cells. The reduced promoter activity of the caldesmon gene induced by platelet-derived growth factor BB was overcome by the forced expression of the constitutive active form of PI3-kinase p110alpha catalytic subunit. These findings suggest that an IGF-I signaling pathway through PI3-kinase plays a critical role in maintaining a differentiated phenotype of smooth muscle cells.

[Complete remission of liver metastasis from rectal cancer following continuous oral administration of tegafur-uracil after intrahepatic arterial single injection of CDDP and 5-FU].

An intrahepatic arterial injection of CDDP, 5-FU, followed by ten months of oral tegafur-uracil administration (2g/day), induced remission for 3 months or more in a 72-year-old male with rectal cancer and synchronous liver metastasis subsequent to anterior resection of the rectum. Tegafur-uracil showed an excellent anticancer effect against colorectal metastatic liver cancers without loss of QOL because a single-low dose of intraarterial anticancer injection was followed by continuous oral administration of tegafur-uracil, and the chemotherapy could be managed to obtain complete remission of the hepatic lesion.

c-Myc gene single-strand binding protein-1, MSSP-1, suppresses transcription of alpha-smooth muscle actin gene in chicken visceral smooth muscle cells.

The expression of alpha-smooth muscle actin is coordinately regulated by positive and negative cis- elements in the promoter region. Although cis -elements and trans -acting factors involved in the positive regulation of the alpha-smooth muscle (alpha-SM) actin gene have been well characterized, details of negative regulation remain unclear. In functional analyses using cultured gizzard smooth muscle cells, we identified a sequence ranging from -238 to -219 in the promoter region as a novel negative element. Mutation and deletion analyses further revealed that a sequence, TATCTTA (-228 to -222), is essential for negative regulation. Gel shift assay and Southwestern blotting indicated that a nuclear protein factor specifically interacts with single- or double-strand DNA including this sequence, and the protein factor displays a highly potent binding to the sense strand DNA. cDNA cloning and gel shift analysis using anti-MSSP-1 antibodies revealed that this protein factor is a chicken homolog of human MSSP-1 (c- myc gene single-strand binding protein-1). In fact, overexpression of MSSP-1 in cultured smooth muscle cells suppresses the promoter activity. These results suggest a novel function of MSSP-1 regarding the transcriptional regulation of alpha-sm actin gene.

Functional involvement of serum response factor in the transcriptional regulation of caldesmon gene.

A 22-bp fragment including the CArG element (CArG1) is essential for the transcription of the caldesmon gene. In this study, we investigated the effects of serum response factor (SRF) on the functional regulation of caldesmon promoter in smooth muscle cells. Gel supershift assay revealed that SRF was one component of the CArG1-protein complex. Dominant-negative mutants of SRF suppressed the promoter activity of caldesmon, whereas wild-type SRF overcame this suppression. These results suggest that SRF functions as a core activating factor of the caldesmon promoter. Furthermore, fractionation of smooth muscle cells' nuclear extracts using DNA affinity paramagnetic particles suggests that SRF transactivates the caldesmon promoter in concert with additional factors in the flow-through fraction recruited to the CArG element.

Identification and characterization of a colon-cancer-associated antigen expressed on leukemia-lymphoma cell lines.

A total of 72 human leukemia-lymphoma cell lines were studied for reactivity with the monoclonal antibody (MAb) A7, an anti-human colon-cancer-cell-associated antigen reagent, by indirect membrane immunofluorescence. Nine of the 72 cell lines expressed the antigen recognized by A7 MAb. Five of the 34 T-cell lines, 2 of the 21 B-cell lines, and 2 of the 3 non-lymphoid-non-myeloid cell lines were reactive with A7 MAb. By means of SDS-PAGE and immunoblotting, the antigens isolated from both colon cancer cell lines (WiDr, SW1116 and LoVo) and leukemia cell lines (A3/KAWAKAMI, H9, RPMI 8226 and SPI-801) showed an identical MW of 42-43 kDa. The non-glycosylated antigen recognized by A7 MAb, which was expressed on both the colon cancer line (SW1116) and the leukemia line (H9) in the presence of tunicamycin, also showed an identical MW of 36 kDa. However, the quantity of the antigen in the leukemia cells was significantly lower than in the colon cancer cells. Although expression of this colon-cancer-associated antigen in the non-colon cancer cells is real, the significant expression of this antigen in colon-cancer cells makes it useful for clinical monitoring of colon cancer patients.

[Immunoresponses and efficacy after arterial infusion of immunoconjugate A7-NCS in patients with colon cancer].

We prepared an immunoconjugate, A7-NCS, of a mouse-derived anti-human colon cancer monoclonal antibody A7 and the macromolecular protein anticancer agent neocarzinostatin (NCS), and evaluated changes in human anti-mouse antibody (HAMA) by ELISA in the serum of patients intraarterially administered this agent. IgG and IgM class HAMA were detected in all patients, but no IgE class HAMA. In patients with stage V disease, the survival rate was higher in a group treated with A7-NCS at an NCS dose of 4,000 units or more than in that treated at an NCS dose of less than 4,000 units. In these patients, the survival rate was higher in a group treated with A7-NCS at an NCS dose of 4,000 units or more than in one not treated with it. These results suggest the usefulness of A7-NCS administration at high dose for prolonging survival of patients with advanced colon cancer.

[Host factors that changes the distribution of immunotoxin].

Mouse monoclonal antibody A 7, which was raised against human colon cancer, was used for preparing the conjugates with neocarzinostatin, mitomycin C and adriamycin. The tissue distribution of these three conjugates were examined in athymic nude mice transplanted with human colon cancer. The distribution in tumor was different in these three conjugates. The route of administration of the conjugate affected its distribution in tumor tissues. In the case of tumor transplanted in the back of mice, intravenous administration seemed to superior to intraperitoneal one. In clinical trials of immunoconjugate composed of A 7 and polypeptide anticancer drug neocarzinostatin (A 7-NCS), human anti-mouse antibody (HAMA) was observed in most cases without serious immune response such as anaphylactic shock. Human antibody against neocarzinostatin could not be detected in any case receiving A 7-NCS.

[A case report of five-year survival with repeated resection of metastases from jejunal leiomyosarcoma].

A case of five-year survival of jejunal leiomyosarcoma with metastases to the liver and mesentery treated with four times surgery was reported here. Four metastatic lesions were detected in the sigmoid mesocolon once and in the liver three times and resected with hope of cure. The patient, a 37 years old male, tolerated surgical procedures including an extended right hepatic lobectomy and recovered each time. He is leading a satisfactory daily life 5 years and 3 months after the initial operation although multiple liver metastases were detected 10 months after the fourth operation. A positive surgical therapy is advocated in selected patients who have spreading leiomyosarcoma of the bowel.