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The contribution of the slow waves (SWs) on high-density plasma production by m=0 helicon waves is investigated in H2, D2, He, N2, Ne, Ar, and Xe plasmas. The density-jump is observed in all the gases used at optimum external magnetic fields B0. The measured plasma density np as a function of B0 peaks at a condition close to the lower-hybrid resonance in H2, D2, He, N2, and Ne, whereas, the measured dispersion relation indicates the excitation of helicon wave. Dispersion relation of electromagnetic waves is solved using dielectric tensor elements of a cold uniform plasma including ion terms, which shows two branches: the SW and fast (helicon) wave. The transition from the ionization by SWs, which are excited in the low-density plasma produced by an antenna induction field, to that by helicon waves has been predicted to explain the mechanism to produce the high-density plasma in the helicon wave discharges.

Antenna coupling has been controlled in high-density plasma production by m=0 helicon waves using multiple-loop antennas and varying the phase and the distance between antennas. When two single-loop antennas are connected in series with the phase between the two being zero, threshold power for the density jump Pth is reduced by a factor of 2–3 compared with that of the one-turn loop antenna. This reduction in Pth is caused by the increase in antenna coupling at the lower plasma density. The observed differences in antenna coupling among different antennas have been examined by using a two-dimensional wave code.

Using expression cloning to screen a human fetal kidney cDNA library for regulator(s) of pro-matrix metalloproteinase (MMP)-2 processing mediated by membrane-type (MT) 1 MMP, we isolated a cDNA whose product interfered with pro-MMP-2 activation. It encodes the NH(2)-terminal 313-amino acid region of a calcium-binding proteoglycan, testican 3, with a 3-amino acid substitution at the COOH terminus and thus was named N-Tes. N-Tes comprises a signal peptide, a unique domain, a follistatin-like domain, and a Ca(2+)-binding domain but lacks a COOH-terminal thyroglobulin domain and two putative glycosaminoglycan attachment sites of testican 3. Pro-MMP-2 activation by MT3-MMP was also inhibited by the coexpression of N-Tes. Immunoprecipitation analysis demonstrated direct interaction of N-Tes with either MT1-MMP or MT3-MMP. Expression of testican 1 or testican 3 but not testican 2 also inhibited pro-MMP-2 activation by either MT1-MMP or MT3-MMP. Deletion and substitution of amino acids residues in N-Tes revealed that the unique NH(2)-terminal domain of N-Tes is responsible for the inhibition of pro-MMP-2 activation by MT-MMPs. Expression of N-Tes and testican 3 was detected in normal brain but down-regulated in glioma tissues. Transfection of either the N-Tes or testican 3 gene into U251 glioma cells or Madin-Darby canine kidney cells transformed by erbB2 suppressed their invasive growth in collagen gel. These results suggest that both N-Tes and testican 3 would interfere with tumor invasion by inhibiting MT-MMPs.

Ets transcription factors are associated with tumor malignancy. We reported previously that the stable transfection of the dominant-negative form of Ets-1 (Ets-DN) in the glioma cell line U251 induced down-regulation of urokinase-type plasminogen activator mRNA expression and invasiveness (M. Nakada et al., J. Neuropathol. Exp. Neurol., 58: 329-334, 1999). Here we analyzed effects of Ets-DN expression on cell adhesion, migration, and phosphorylation of focal adhesion kinase. U251 cells expressing Ets-DN (U251-DN) showed reduced cell adhesion, spreading, and extension of actin stress fibers on dishes coated with fibronectin but not on dishes coated with collagen. Migration of U251-DN cells was found to be significantly inhibited compared with that of parental cells when examined by wound-induced migration assay on fibronectin-coated dishes. Phosphorylation levels of focal adhesion kinase in U251-DN cells were also attenuated on dishes coated with fibronectin. Reduced expression level of integrin alpha5 subunit in U251-DN cells was demonstrated by semiquantitative reverse transcription-PCR analysis. Semiquantitative reverse transcription-PCR of surgical samples of brain tumors revealed that the expression level of Ets-1 mRNA correlated with that of integrin alpha5 mRNA in glioma. The experimental metastatic ability of U251-DN cells examined in chick embryo was considerably lower than that of parental cells. These results suggest that Ets-1 contributes to glioma malignancy by up- regulating expression of the integrin alpha5 subunit, which composes integrin alpha5beta1 and mediates intracellular signaling and the subsequent acceleration of the invasive process, including cell adhesion and migration.

The mechanisms of the lymph node metastasis remain unclear. We demonstrate the role of MT1-MMP on the lymph node metastasis using in vivo experimental model of lymph node metastasis by orthotopic implantation of MT1-MMP transfected gastric cancer cell line in the stomach of nude rats. TMK-1 cell line without expression of MT1-MMP was transfected with the pcDNA3 plasmid containing a 3.4-kb MT1-MMP cDNA fragment by calcium phosphate method, and the transfected cell line was designated as TMK-MT. Western blot and RT-PCR analyses showed the specific bands corresponding to MT1-MMP in the TMK-MT cells. By gelatin zymography, the activated form (62-kDa) of MMP-2 was identified in the medium of TMK-MT cell line, but was not detected in TMK-1 cells. Six weeks after orthotopic implantation of TMK-1 and TMK-MT xenografts of nude mouse-subcutaneus tumor into the stomach of nude rats, gastric tumors were found in all the animals. Histologically, the lymphatic invasion was found in the submucosa of the TMK-MT gastric tumors. Lymph node metastasis was not detected in nude rats bearing TMK-1 gastric tumor (0/8). In contrast, lymph node metastasis was detected in five out of 8 rats, bearing TMK-MT gastric tumor. MT1-MMP immunoreactivity was found on the cell membrane and cytoplasm of TMK-MT cells not only in the lymph node metastasis but also in the stomach tumor. These results suggest that MT1-MMP overexpression induced by transfection of its gene may promote lymph node metastasis of transformed cells.

Thirty-two patients with chronic active hepatitis C were initially treated with 9 million units of interferon (IFN)-alpha 2a daily for 4 weeks and then thrice weekly for 20 weeks. The incidence of development and influence on clinical effectiveness of anti-IFN alpha 2a neutralizing antibodies measured by bioassay were investigated. Thirteen (41%) of the 32 patients developed antibodies, 4 (12.5%) of whom exhibited a high titer of 128 NU or greater. There was a significant difference (p0.05) between responders and non-responders in the low viral load group (less than 10(5) copies/50 microliters) in the incidence of antibody development, with antibodies present in 2 of the 9 responders (22%) and 5 of the 7 non-responders (71%). There was also a significant difference between the antibody-negative and antibody-positive patients in circulating HCV-RNA, 2-5AS activity and average ALT levels at the latter half of treatment. In addition, 2-5AS activity was significantly lower (p0.05) at completion of treatment in the high antibody titer group (128 NU or greater) than in the low antibody titer group. These results indicate that the development of anti-IFN neutralizing antibodies had a definite influence on the clinical course of IFN treatment for chronic hepatitis C.

Membrane-type matrix metalloproteinase 1 (MT1-MMP) is a member of the recently identified unique membrane-type subgroup in the matrix metalloproteinase (MMP) family. MT1-MMP has proteolytic activity against components in the extracellular matrix and activates progelatinase A (72-kDa type IV procollagenase/proMMP-2) on the cell surface. Because MT1-MMP is frequently expressed in a variety of tumors, we examined its contribution to their metastatic potential. The mouse lung carcinoma cell line Madison 109 was transiently transfected with a MT1-MMP expression plasmid and inoculated into the tail vein of BALB/c mouse. Fate of the transfected cells was monitored by the neo(r) gene in the plasmid using the quantitative PCR method. The survival rate of the parental cells in lung was 0.7% of the inoculated cells. It was increased by 3-fold with the MT1-MMP transfected cells and the number of the lung nodules increased accordingly. Immunostaining of the consecutive tissue sections revealed that lung nodules expressing MT1-MMP were positive for gelatinase A as well, whereas MT1-MMP-negative cells were not stained for gelatinase A at all. Thus, MT1-MMP-expressing cells acquire specific ability to bind exogenous progelatinase A.

Membrane-type 1 matrix metalloproteinase that is associated with the proteolytic activation of progelatinase A was expressed as a recombinant fusion protein in Escherichia coli. The recombinant enzyme cleaved the propeptide sequence of gelatinase A in a sequence-specific manner. A mutant progelatinase A that has a substitution of Asn(66)-Leu to Ile-Val was not processed at all. The processing was blocked by tissue inhibitor of metalloproteinases-2 or BB-94 but not by tissue inhibitor of metalloproteinases-1. Thus, membrane-type 1 matrix metalloproteinase is a direct activator of progelatinase A without requiring additional proteases.

Membrane-type matrix metalloproteinase (MT-MMP), which we have identified recently, is unique in its transmembrane (TM) domain at the C terminus and mediates activation of pro-gelatinase A on the cell surface (Sato, H., Takino, T., Okada, Y., Cao, J., Shinagawa, A., Yamamoto, E., and Seiki, M. (1994) Nature 370, 61-65; Takino, T., Sato, H., Yamamoto, E., and Seiki, M. (1995) Gene (Amst.) 115, 293-298). In addition to MT-MMP, a novel MMP-related cDNA of 2.1 kilobases was isolated from a human placenta cDNA library. The cDNA contains an open reading frame for a new MMP. The deduced protein composed of 604 amino acids was closely related to MT-MMP in the amino acid sequence (66% homology at the catalytic domains) and has a potential TM domain at the C terminus. Monoclonal antibodies raised against the synthetic peptide recognized a 64-kDa protein as the major product in the transfected cells. TIMP-1 fused with the potential TM domain was localized on the cell surface while native TIMP-1 is in the culture medium. Thus, we called the second membrane-type MMP, MT-MMP-2 and renamed MT-MMP, MT-MMP-1. MT-MMP-1 and -2 are thought to form a distinct membrane-type subclass in the MMP family since all the others are secreted as soluble forms. Like MT-MMP-1, expression of MT-MMP-2 induced processing of pro-gelatinase A (68-kDa in gelatin zymography) into the activated form of 62-kDa fragments through a 64-kDa intermediate form. Expression of MT-MMP-2 mRNA was at the highest levels in the brain where MT-MMP-1 was at the lowest level compared to other tissues. MT-MMP-1 and -2 are thought to be utilized for extracellular matrix turnover on the surface of cells under different genetic controls.

Magnetic resonance imaging (MRI) findings in cases with symptomatic and asymptomatic Schmorl's nodes have been analysed. In all symptomatic cases, the vertebral body marrow surrounding the Schmorl's node was seen as low signal intensity on T1-weighted images and as high signal intensity on T2-weighted images. It was confirmed by histological examination that the MRI findings indicated the presence of inflammation and oedema in the vertebral bone marrow. These MRI findings were not seen in asymptomatic individuals. Inflammatory changes in the vertebral body marrow induced by intraosseous fracture and biological reactions to intraspongious disc materials might cause pain. We postulate that after fracture healing and subsidence of inflammation, the Schmorl's nodes become asymptomatic, in analogy with old vertebral compression fractures. MRI is not only useful in detecting the recently developed Schmorl's nodes but also in differentiating between symptomatic and asymptomatic Schmorl's nodes.

The hepatic disposition of lactosaminated bovine serum albumin (Lac-BSA) in rats was studied at the whole body, isolated liver, and isolated parenchymal cell levels. After intravenous injection, 111In-Lac-BSA (1 mg/kg) was rapidly eliminated from the plasma due to extensive uptake by liver parenchymal cells; however, a significant decrease in hepatic clearance was observed at high dose (50 mg/kg). In a single-pass, constant infusion experiment in the isolated liver, 111In-Lac-BSA was continuously extracted. The extraction ratio at steady state (Ess) for 111In-Lac-BSA was significantly decreased by coadministrating galactose, NH4Cl, or chloroquine, and at low temperature, suggesting that hepatic uptake of Lac-BSA proceeds via receptor-mediated endocytosis for asialoglycoprotein. Kinetic analysis of 111In-Lac-BSA binding with isolated parenchymal cells at 4 degrees C yielded a dissociation constant (Kd) of 2.5 x 10(-8) M and a value of 3.5 x 10(5) maximal binding sites/cell (Bmax). The internalization rate constant (kint) for 111In-Lac-BSA was calculated to be 0.46 min-1 in liver perfusion experiments using the EDTA-wash method.

The effect of electric charge on the hepatic disposition of macromolecules was studied in the rat. Charged derivatives of dextran (T-70) and bovine serum albumin (BSA), mitomycin C-dextran conjugates (MMC-D), and lactosaminated BSA (Lac-BSA) were employed as model macromolecules. After intravenous injection, cationic macromolecules were rapidly eliminated from plasma because of their extensive hepatic uptake, while anionic and neutral macromolecules were slowly eliminated. Cationic macromolecules were recovered from parenchymal and nonparenchymal hepatic cells at a cellular uptake (per unit cell number) ratio of 1.4-3.2, while that of Lac-BSA was 14. During liver perfusion using a single-pass constant infusion mode, cationic macromolecules were continuously extracted by the liver, with extraction ratios at steady-state (Ess) ranging between 0.03 and 0.54, whereas anionic and neutral macromolecules were almost completely recovered in the outflow at steady state. The Ess for cationized BSA (Cat-BSA) and cationic MMC-Dcat were concentration dependent and decreased at low temperatures and in the presence of colchicine and cytochalasin B. The possible participation of the internalization process in the uptake of cationic macromolecules by hepatocytes was suggested.

The blood flow in esophageal varices was measured by endoscopic laser Doppler velocimetry (LDV) in 27 patients histologically or clinically diagnosed as having liver cirrhosis. The effect of endoscopic injection sclerotherapy (EIS), and of intravenous oral administration of propranolol, on the output signal of LDV was evaluated. EIS was performed intravariceally after the measurement, with a 50% glucose solution, human thrombin (1000 U), and Aethoxysclerol as sclerosants.

Es wird uber einen 22jahrigen Japaner mit einer hereditaren, nichtspharozytaren hamolytischen Anamie berichtet. Es bestand eine konstante Anamie mit Retikulozytose, Ikterus und Mangel an Glucose-6-Phosphat-Dehydrogenase in den Erythrozyten.

Recently, there has been much concern that cancer chemotherapy may have undesirable consequences in the form of secondary malignancies. At present, secondary leukemia or therapy-linked leukemia constitutes a well delineated clinical syndrome characterized by several distinct symptoms, that was illustrated by the presentation of cases recently observed. In an attempt to understand the basis for this therapy-linked leukemia, mutagenic activity of various anticancer agents were studied by the induction of sister chromatid exchanges and risk of a certain drug as a possible candidate causing secondary leukemia was discussed from the molecular cytogenetic point of view. From both clinical and cytogenetic aspects it seemed that bifunctional alkylating agents were potentially relevant to the development of secondary leukemia.

Sister chromatid exchanges (SCEs) induced by chemotherapeutic agents currently used for treating various malignancies were studied in vitro and in vivo. Whether the agents are potentially mutagenic/carcinogenic is discussed on the basis of recent clinicopathological observations on secondary malignancy. In the in vitro studies, the induction of SCEs by various anticancer agents, including three anticancer antibiotics, six alkylating agents, and three antimetabolites, was examined and compared with induction by two known carcinogens, 4-nitroquinoline 1-oxide and N-ethyl-N'-nitronitrosoguanidine. In the in vivo studies, SCE frequencies were examined in the lymphocytes from 74 blood samples drawn from 40 patients with cancer or leukemia at various times after the agents had been administered alone or in combination. The SCE frequencies induced by the anticancer antibiotics were variable because the drugs had different modes of action: generally, the drugs have strong cytotoxicity that must affect the viability of cells. On the other hand, alkylating agents induced significantly high levels of SCE frequencies in vitro; the modes of SCE inducibility were similar to those of the two carcinogens. The fact that cells were viable up to relatively high molar concentrations of the agents and that high SCE levels persisted for a long period suggests that the lesions induced by alkylating agents are long-lived and may therefore be more frequently involved in mutagenesis. This finding may be compatible with the clinical observation that the vast majority of patients with second malignancy are found in the group treated with alkylating agents. Antimetabolites generally showed only a weak SCE induction or none in vivo and in vitro.

A 3-year-old boy with partial No. 9 tetrasomy is described. The patient showed markedly retarded physical and mental development as well as multiple congenital anomalies. Routine chromosome analysis revealed an extra C-group chromosome. It had a pronounced secondary constriction at the proximal part of its long arm. Based on studies by a variety of banding techniques, the extra chromosome was identified to be an iso-dicentric No. 9 chromosome with inactivation of one of the two centromeres, the karyotype being 47,XY, + DIC (9)(Q2101). The value of BrdUrd treatment was emphasized in the detection of a very small piece of euchromatin within a long stretch of constitutive heterochromatin.

Livers from 12 mice fed griseofulvin for 4 to 6 months were perfused in situ with a detergent solution to extract lipid membranes leaving the cytoskeleton intact. Seven control mice were similarly studied. After 30 to 120 minutes perfusion, liver samples were examined by scanning electron microscopy and transmission electron microscopy. By light microscopy, Mallory bodies (MBs) were observed in pericentral hepatocytes. These were confirmed by transmission electron microscopy. Intermediate filaments (IFs) were observed in close apposition to MBs. Numerous IFs were seen throughout the cytoplasm. The 3-dimensional organization of the cytoskeleton and MBs were clearly visualized by scanning electron microscopy. The IFs were disorganized in the hepatocytes and formed small MBs in griseofulvin-treated mice. In the case of hepatocytes containing large MBs, there was an apparent decrease in the concentration of IFs. Transition forms of dense networks of IFs between the normal cytoskeleton and the MBs were noted in the cytoplasm between small MBs and the nucleus and also at the cell border. The IFs connected to the nucleus and invaded the MBs. The MBs appeared to form as a result of condensation or collapse of the IFs.

Twenty-four autopsy cases of liver cell carcinomas were investigated by Feulgen-DNA cytofluorometry (NIKON SPM-RF 1-D) on paraffin-embedded tissue materials. The methods employed for tissue preparation and cytofluorometry were previously reported by us. The ploidy pattern based on cytofluorometry of the liver cell carcinomas exhibited wide DNA content distributions, in which many S phase cells having intermediate DNA content values ranging from diploid level to various, higher ploidy levels were included. From the comparative studies on both the histopathological findings and the ploidy pattern of the liver cell carcinomas, it was indicated that the peak value of the DNA content distribution was shifted from the diploid to the tetraploid level with the increasing appearance of the polyploid cells, and also that the degree of polyploidization appeared to correspond to the extent of the histological malignancy.

Forty one patients with infections associated with leukemia and related disorders were treated with cefpiramide (CPM). In 26 patients among them, we were able to evaluate the effectiveness of CPM against infections. Fifteen patients were not evaluated, because 6 patients were subjected to additional therapy such as gamma-globulin and other antibiotics, 5 were prophylactically treated, 2 had fever episode which were retrospectively reviewed to be originated from tumor mass, 1 received too short a duration of administration of CPM (2 days) to evaluate its effectiveness, and 1 with whom no precise data were recorded. Excellent responses were observed in 10 patients (38.5%) and good responses in 6 (23.1%) among these 26, with a total efficacy rate of 61.5%. Whereas, we found only one patient who showed an unfavorable side effect out of 31 patients including the 26 and 5 other patients who were prophylactically treated. The side effect observed was a mild bleeding tendency occurred in 77 years old female at 11 days after CPM was administrated. The bleeding tendency was easily diminished with the cessation of CPM treatment and a parenteral use of vitamin K. These results suggest that CPM is an effective and safe antibiotic for the treatment of infections in patients with leukemia and related disorders.

To clarify the clinical efficacy of SPG, a beta-1, 3 glucan extracted from cultured Schizophyllum commune Fries, a randomized comparative study was performed in combination with mitomycin C+5-fluorouracil (MF protocol) or tegafur (F protocol). A total of 514 cases with inoperable or recurrent gastric cancer were randomly allocated to either the SPG group or the control group, 367 of which were finally assessed for clinical efficacy. SPG was intramuscularly given at a dose of 20 mg twice a week or 40 mg once a week. Significant prolongation of life span was confirmed in the SPG group (MF protocol: p less than 0.01, F protocol: p less than 0.05), although the combination of SPG did not demonstrate a remarkable antitumor effect in both MF and F protocols. Among immune response parameters tested, the positive reactions in the PHA skin test were maintained in the SPG group and decreases in lymphocyte counts were inhibited by SPG in the MF protocol. Side effects associated with SPG therapy were noted in 6 of 258 cases treated with SPG. From these results, it is indicated that the combination of SPG and the chemotherapeutic agents may be useful in treating patients with inoperable or recurrent gastric cancer.