Small-amplitude dynamic force microscopy using a quartz cantilever with an optical interferometer.

We propose dynamic force microscopy (DFM) that employs a quartz cantilever in combination with an interferometric deflection sensor. The high stiffness of the quartz cantilever allows DFM to be performed at oscillation amplitudes as small as 1 A. DFM topographic images can be obtained for a Si(111)-(7 x 7) surface even when a blunt tip is used at room temperature. The low-noise interferometer with a deflection noise floor of n = 15 fm Hz(-1/2) exhibits a high force sensitivity in force spectroscopy. By subtracting the huge van der Waals force contribution, we obtain a short-range force curve that is in good agreement with the theoretical curve of the covalent bonding force. Simultaneous measurement of the tunneling current during DFM imaging is also demonstrated.

Intra-parenchymal tumor biopsy using neuroendoscopy with navigation.

BACKGROUND AND OBJECTIVES: Neuroendoscopy has allowed us to biopsy tumors located in the ventricle and the para-ventricle. With this technique we can obtain specimens under direct visual monitoring and examine tumor dissemination. For intra-parenchymal tumors, however, we normally use stereotactic procedures to collect tissues or perform open biopsies. We now report that we have successfully combined neuroendoscopy with navigational guidance to biopsy intra-parenchymal tumors. We explain our methods and discuss the advantages and disadvantages of this technique. CASES AND METHODS: We carried out intra-parenchymal tumor biopsies or resection using neuroendoscopy and guided navigation in three patients. We advanced a transparent sheath, containing a removable inner tube on which a navigation tool was mounted, to target tissues. We directly monitored procedures with a neuroendoscope placed in the transparent sheath. We collected specimens just in front of and in the tumor by forceps. In one patient with bleeding, we used the aspirator tip as a monopolar coagulator. We were also able to check for bleeding along the sheath tract using endoscopy. Without causing any new neurological deficits, we collected tissues for histological diagnosis of astrocytoma in two patients and radiation necrosis in one. CONCLUSIONS: We believe that combining neuroendoscopy with navigation guidance is a safe and precise method for obtaining biopsies of intra-parenchymal tumors. Tumors with rich vasculature will not benefit from this procedure until better hemocoagulation instruments have been developed.

Checkpoint and physiological apoptosis in germ cells proceeds normally in spaceflown Caenorhabditis elegans.

It is important for human life in space to study the effects of environmental factors during spaceflight on a number of physiological phenomena. Apoptosis plays important roles in development and tissue homeostasis in metazoans. In this study, we have analyzed apoptotic activity in germ cells of the nematode C. elegans, following spaceflight. Comparison of the number of cell corpses in wild type or ced-1 mutants, grown under either ground or spaceflight conditions, showed that both pachytene-checkpoint apoptosis and physiological apoptosis in germ cells occurred normally under spaceflight conditions. In addition, the expression levels of the checkpoint and apoptosis related genes are comparable between spaceflight and ground conditions. This is the first report documenting the occurrence of checkpoint apoptosis in the space environment and suggests that metazoans, including humans, would be able to eliminate cells that have failed to repair DNA lesions introduced by cosmic radiation during spaceflight.

Long-term survival and late-onset complications of cancer patients treated with high-dose chemotherapy followed by autologous peripheral blood stem cell transplantation.

The antitumor effect of high-dose chemotherapy (HDC) followed by autologous peripheral blood stem cell transplantation (auto-PBSCT) is considered superior to that of conventional chemotherapy. However, the long-term benefits of this strategy in Japan remain unclear. Therefore, in this study, 109 cancer patients enrolled between 1989 and 1999 were treated with HDC and auto-PBSCT. Patients were evaluated for long-term survival and late-onset complications, including secondary malignancy. The mean number of CD34+ cells harvested per apheresis was larger in the group receiving high-dose cytosine arabinoside or high-dose etoposide plus granulocyte colony-stimulating factor (G-CSF) than in the group receiving conventional chemotherapy plus G-CSF. The 5-year overall survival rates for non-Hodgkin's lymphoma patients in first complete remission (CR) (83.2%), second or subsequent CR (74.1%), or first partial remission (PR) (66.7%) at the time of transplantation were significantly higher than those with no remission (35.7%) at the time of transplantation (first CR, P < .05; second or subsequent CR, P < .05; first PR, P < .05). The 5-year overall survival (OS) rates for breast cancer was 40.8%, and the disease-free survival rate was extremely low (8.8%). The 5-year OS rates for chemotherapy-sensitive and chemotherapy-resistant diseases at the time of transplantation were 32.7% and 35.7%, respectively, a difference that was not considered significant. The 5-year OS for germ cell tumor was 80.0%, and the disease-free survival rate was 77.9%. The rate of therapy-related death was 8.2%. The occurrence rate of secondary malignancy was 0.9%. Late-onset complications were observed in 4 cases (glomerulonephritis, interstitial pneumonitis, ulcerative colitis, and acute myelogenous leukemia). At 3.7%, the occurrence rate was not very high, but most complications of auto-PBSCT were life threatening and interfered with patients' quality of life. A careful follow-up is required for at least 2 years after transplantation, because the mean occurrence time of late-onset complications is 16.7 months posttransplantation.

Caenorhabditis elegans Chk2-like gene is essential for meiosis but dispensable for DNA repair.

A Chk2-like gene was identified in the genome of Caenorhabditis elegans. The putative gene product, termed Ce-chk-2 consists of 450 amino acid residues, and shows good homology with the Chk2/Cds1 gene family. The results of RNA-mediated interference (RNAi) indicated that the F1 generation from dsRNA injected animals grew to adulthood, but approximately 95% of their eggs (F2) died during early embryogenesis. Among the few surviving progeny, males (XO animals) arose at an abnormally high frequency (30%). In addition, 12 univalents were observed in full grown oocytes of the F1, while six bivalents were normally observed in wild-type oocytes. Ce-chk-2 gene expression increased in the adult stage, and their expression level decreased in the glp-4 mutant, which is defective in germ line proliferation. The radiation sensitivity of F1 embryos carrying Ce-chk-2 RNAi was not significantly affected.

Molecular cloning and sequence analysis of the gene encoding the collagenase from Cytophaga sp. L43-1 strain.

Cytophaga sp. strain L43-1 secretes a collagenase [Y. Sasagawa et al., Biosci. Biotech. Biochem., 57, 1894-1898 (1993)]. A cog gene encoding the collagenase from this strain was cloned, and the nucleotides were sequenced. The structural gene of cog consisted of 3846 base pairs, which encoded a polypeptide consisting of 1282 amino acid residues with a predicted molecular mass of 130 kDa which was synthesized as a pre-matured enzyme. The deduced N-terminal 14 amino acids sequence, molecular mass of 120 kDa, and pI of 4.96 of the predicted matured enzyme were consistent with those previously found for the collagenase purified from the strain. The cog gene was expressed in Escherichia coli using the lac promoter and ribosomal binding sequence in plasmid vector pUC119 or pKK223-3, but not its own putative promoter and Shine-Dalgarno sequence. The consensus amino acid sequence (His-Glu-Xaa-Xaa-His) of an active site of the metal proteases including the collagenase from Vibrio arginolyticus and a series of human MMPs was found in the Cog protein of the strain.

Purification and properties of collagenase from Cytophaga sp. L43-1 strain.

A collagenolytic bacterial strain was isolated from soil and was identified as Cytophaga sp. It produced several kinds of collagenase and protease. From the supernatant of a culture, a collagenase was purified as a single protein band upon SDS-PAGE and its molecular mass was estimated to be 120 kDa. Collagen and gelatin were good substrates for this enzyme. beta-Casein was cleaved by this enzyme at several sites.

A clinico-pathological study on progress factors in lupus nephritis.

The prognosis of lupus nephritis was investigated in 147 patients who underwent renal biopsy, in an attempt to elucidate the progress factors of this disease clinico-pathologically. Twenty-two patients received a follow-up biopsy and were observed for time-related histopathological changes. At the first examination, only the serum creatinine and mean blood pressure were correlated with prognosis. The anti-DNA antibody and serum complement titer measured at the same time did not correlate with prognosis, and neither showed a significant improvement in the poor prognosis group. Profuse proteinuria tended to last longer in the poor prognosis group. The renal biopsy findings were found to correlate with prognosis. The prognosis was especially poor in patients exhibiting marked mesangial cell proliferation, glomerulosclerosis and interstitial lesions.

[Follicular lymphoma associated with t(14;18)(q32;q21) chromosome translocation and bcl-2 gene rearrangement: report of a case].

A 31-year-old Japanese male who complained of low-grade fever, fatigue and generalized lymphadenopathy had shown an increase in peripheral white cell count, and 84% of peripheral blood cell and 76% of nuclear bone marrow cells consisted of small cleaved lymphoblastic abnormal cells with or without barely visible nucleoli. Cytogenic study of cervical lymph node biopsy specimens showed a t(14;18)(q32;q21) chromosome translocation. Histologically the lymph node cells were classified according to the International Working Formulation as follicular small cleaved-cell lymphoma. Molecular analysis of DNA fragments of peripheral lymphocytes revealed both J-H gene and bcl-2 oncogene rearrangements. Immunophenotypes of peripheral lymphocytes, bone marrow cells and lymph node cells expressed a clonally distinct B-cell population bearing surface immunoglobulin-G, kappa-chain, CD-10, CD-19, CD-20, CD-21 and OKIa-1 antigens. We diagnosed this case as follicular lymphoma in the leukemic phase,

Hypereosinophilic syndrome associated with obliterative left ventricular chamber and systemic obliterative arteriosclerosis.

An extremely rare case of hypereosinophilic syndrome is reported. The patient had congestive heart failure due to left ventricular obliteration by a giant thrombus in the apex. Arteriosclerosis obliterans resulted in serious lower extremity gangrene. Combination therapy with a corticosteroid and hydroxyurea caused a marked reduction in eosinophil count, and heart failure recovered in stages. Although the patient was discharged, he returned to our hospital 3 months later with acute myeloblastic leukemia and subsequently died of respiratory failure with pneumonia.

Diffuse infiltrating T-cell lymphoma of the colon associated with polyclonal hypergammaglobulinemia and hepatocellular carcinoma: report of a case.

A 55-year-old man with a chief complaint of melena had diffusely infiltrated lymphoma from the rectum to the sigmoid colon and polyclonal hypergammaglobulinemia. Biopsy specimen obtained from the rectum revealed diffuse medium-sized lymphoma cell with plasmocytosis. Histochemical analysis with monoclonal antibodies indicated that the origin of the tumor cell to be T-lymphocyte. Chemotherapy with cyclophosphamide, vincristine and prednisolone (VEP regimen) was effective for mucosal bleeding of the colonic lesion and reduction of polyclonal hypergammaglobulinemia. Five yrs later he showed recurrence of the disease and elevation of serum alphafetoprotein, and died of pulmonary infections. Autopsy finding confirmed the diagnosis of malignant lymphoma of the colon and disclosed the association of hepatocellular carcinoma.

[An echocardiographic study of alcoholic cardiomyopathy after total abstinence].

Left ventricular function of alcoholic cardiomyopathy with congestive heart failure was investigated by echocardiography during total abstinence for six months. In five of 13 cases, left ventricular ejection fraction (LVEF) was normalized to 55% or more and left ventricular end-diastolic dimension (LVEDd) was reduced to 5.7 cm or less during the study. In the remaining eight patients, no improvement was observed. In the responsive group, two patients showed an increase of LVEF and a reduction of LVEDd simultaneously, but three showed an earlier change in LVEDd. This normalization of LVEF and LVEDd was recognized from three to 12 weeks after abstinence. No significant differences were detected between the responsive and non-responsive groups concerning their age, daily intake of alcohol and period of drinking, NYHA cardiac functional class, the period of experiencing cardiac symptoms and the cardiothoracic ratio by chest radiography. The ratio of left ventricular radius to wall thickness (R/Th) remained 2.9 or less in all of the responsive group, but seven of eight in the non-responsive group showed 2.9 or more. Left atrial dimension was reduced to the normal range except in one, but its normalization did not correlate with the change of left ventricular function. Consequently, after total abstinence from alcohol intake, both left ventricular end-diastolic dimension and ejection fraction improved rapidly, and their normalization could be highly expected. Echocardiography is very helpful in predicting the effect of total abstinence on left ventricular function in alcoholic cardiomyopathy.

[IBL like T cell lymphoma associated with hemolytic anemia and polyploidy chromosomal abnormality--a case report].

A 56-year-old man was admitted to Sapporo Kitano Hospital on May 30, 1987 because of fever, retention of ascites and pleural effusion, generalized lymphnode swelling and hepatosplenomegaly. Laboratory findings showed Coombs' positive hemolytic anemia, leukocytosis and polyclonal hypergammaglobulinemia. Serological test included positive RA factor, anti-DNA 16 U/ml, thyroglobulin Ab 1600 x and microsome Ab 3200 x. A cervical lymphnode specimen exhibited typical histologic picture of IBL like T cell lymphoma. The surface markers of lymphoma cells were CD2(+), CD8(+) and CD4(-). Chromosomal analysis of these cells revealed polyploidy abnormality for all chromosomes except for No. 14's which are disomy. All lymphoma cells have XXY sex chromosome and mar1, mar2, mar3. Gene rearrangement for beta-chain of T cell receptor was proved in these lymphoma cells. He was treated with prednisolone, vincristine, cyclophosphamide, adriamycin etc, but died of respiratory failure 171 days after admission. We reported a rare case of IBL like T cell lymphoma with polyploidy chromosomal abnormality accompanied with hemolytic anemia.

Juvenile cases of restrictive cardiomyopathy without eosinophilia.

Two cases of juvenile restrictive cardiomyopathy are presented, in which we highlight the useful role of noninvasive diagnostic techniques including the apexcardiogram, echocardiography, and magnetic resonance (MR) imaging.

A phase II study of etoposide in patients with hepatocellular carcinoma by the Tokyo Liver Cancer Chemotherapy Study Group.

Twenty-four patients with hepatocellular carcinoma were treated with 80 mg etoposide/m2 for five consecutive days at 4-week intervals. Of the 21 patients evaluated, only one showed a partial response (of 8 weeks, duration). The dose-limiting factor was bone marrow suppression. Grades 3 and 4 white blood cell count toxicity occurred in five (24%) and two (10%) patients, respectively. Alopecia (grades 1 and 2) was observed in 18 patients (57%). The The present study indicates that etoposide has very limited activity against hepatocellular carcinoma.

[Cardiac involvement of alcoholics and the effect of total abstinence].

Echocardiographic features of 221 alcoholics after total abstinence were investigated. At the time of admission, none demonstrated left ventricular (LV) enlargement. Noticeably, 26% patients out of the total showed LV wall thickening more than 1.4 cm in the interventricular septum. On the other hand, 19% of the alcoholics illustrated hypokinetic LV wall without chamber dilatation which was either less than 55% in ejection fraction or 30% in fractional shortening. Three months later, the LV wall thickness reduced significantly from 1.6 to 1.4 cm in average and the LV wall motion improved significantly from 27 to 31% in fractional shortening. In the cases with wall thickening, the change was well correlated with a decrease both in heart rate and in LV muscle volume, but not with a decrease in systemic blood pressure. No significant correlated factors could be detected in the hypokinetic cases.

[Thrombolytic therapy of pulmonary thromboembolism--result of 18 patients].

In Japan, pulmonary thromboembolism (PTE) is rare and the therapy of this disease, especially the thrombolytic therapy, has not been established yet. We studied the effect of thrombolytic therapy by urokinase (UK) in eighteen patients with PTE. We also compared the results with those in Europe and the United States. Eighteen patients (male 7, female 11) were 27 yo to 77 yo (mean 55 yo). Seven cases were mild and 11 cases were moderate to severe. The initial dosage of UK administration was 2.4 x 10(5) to 7.2 x 10(5) IU with the mean of 4.1 x 10(5) IU, which was approximately the double of those in Europe and the US. The duration of UK therapy was 1 to 9 days with the mean of 4.8 days. It was longer than in Europe and the US (12-24 hours). Total amount of UK in each case ranged 4.8 x 10(5) to 42 x 10(5) IU (mean 19.3 x 10(5) IU) with the daily dosage of 4.0 x 10(5) IU in average. They were almost half of those in Europe and the US. Clinical signs and laboratory findings subsided in 1-7 day by UK administration in 14 cases (87%) with the reduction of defects on perfusion lung scans and all of these cases survived. No severe complications were found in any cases. We conclude that the thrombolytic therapy with UK in Japan might be acceptable from the results of this present study. However, the prospective study of thrombolytic therapy on a larger scale remains to be done in the future.

Generation of lymphokine activated killer cells in a new high density dialyzing culture apparatus.

In order to establish an efficient culture system for the generation of lymphokine activated killer (LAK) cells, we have developed a new device which is essentially based on a continuous dialyzing culture vessel. LAK cells grown in such a system showed higher cytotoxicity than those grown under conventional culture conditions. By using this new apparatus with continuous regulation of infused interleukin 2, nutritional medium, and pO(2) and pCO(2), yields of 2×10(7) cells/ml were achieved and maintained for more than 21 days. These cells also showed a significant increase of LAK activity on a per cell basis.