Involvement of Rab13 and JRAB/MICAL-L2 in epithelial cell scattering.

Epithelial cell scattering recapitulates the first steps of carcinoma invasion/metastasis. While the balance between cell-cell adhesive activity and cell motility ultimately determines this process, its molecular mechanisms remain unclear. Adherence junctions and tight junctions (TJs) are primarily responsible for cell-cell adhesive activity and subjected to dynamic remodeling. We previously showed that Rab13 and its effector protein JRAB/MICAL-L2 mediate the endocytic recycling of the integral TJ protein occludin and the assembly of functional TJs. In this study, we examined the role of Rab13 and JRAB/MICAL-L2 in the scattering of Madin-Darby canine kidney (MDCK) cells in response to 12-O-tetradecanoylphorbol-13-acetate (TPA). Knockdown of Rab13 in canine MDCK cells suppressed the TPA-induced scattering, and this phenotype was restored by re-expression of human Rab13. During TPA-induced MDCK cell scattering, Rab13 was transiently activated and returned to its basal level, and both Rab13 and JRAB/MICAL-L2 were colocalized with F-actin at cell-cell contact sites and then accumulated at emerging lamellipodial structures. TPA-induced MDCK cell scattering was also inhibited by knockdown of canine JRAB/MICAL-L2 and rescued by re-expression of mouse JRAB/MICAL-L2. These results indicate that Rab13 and JRAB/MICAL-L2 are involved in epithelial cell scattering.

Successful treatment with reduced-intensity stem cell transplantation in a case of relapsed refractory central nervous system lymphoma.

A 33-year-old male with refractory relapsed central nervous system lymphoma underwent allogeneic hematopoietic stem cell transplantation (allo-HSCT) from an HLA-identical sibling after reduced-intensity conditioning chemotherapy. The preparative regimen for allo-HSCT consisted of fludarabine and busulfan. Cyclosporine (CsA) and short-term methotrexate were used as prophylaxis for acute graft-versus-host disease (GVHD). Although CsA was quickly reduced to induce a graft-versus-lymphoma (GVL) effect, no symptoms of GVHD and GVL effect were evident. Donor lymphocyte infusion (DLI) was performed on day +40 following transplantation. The patient developed acute GVHD (grade III) after DLI, and lymphoma regression was observed after the occurrence of GVHD. Four months after transplantation, complete remission was achieved with extensive chronic GVHD, and the patient continues to be disease free at 15 months after transplantation.

E-cadherin expression in lymph nodes of three patients with non-Hodgkin's lymphoma.

The E-cadherins are a family of cell-cell adhesion molecules. These molecules exhibit Ca2+ dependent cell adhesion and are expressed on epithelial cells. E-cadherin levels in serum are known to be significantly elevated in patients with epithelial carcinomas. We determined serum E-cadherin levels in 30 patients with non-Hodgkin's lymphoma (NHL) using an enzyme immunoassay and then investigated whether E-cadherin is expressed on lymphoma cells in lymph nodes of three cases selected to analyze from 15 cases of serum E-cadherin levels over mean + 2SD with monoclonal antibody immunohistochemistry. Results indicated that E-cadherin antigen is expressed on the lymphoma cells in these three patients with NHL, and that soluble E-cadherin might be released into blood from lymphoma cells. Expression of E-cadherin may contribute to the morphological appearance of some malignant lymphoma, although no conclusion can be drawn based on such small number of patients analyzed.

Clinical significance of serum E-cadherin levels in patients with haematological malignancies.

E-cadherin is a transmembrane glycoprotein that mediates Ca2+-dependent intracellular adhesion in normal epithelial cells. E-cadherin levels in serum are known to be significantly elevated in patients with epithelial carcinomas. However, the role of E-cadherin in haematopoietic cells is less clear. In this study, serum E-cadherin levels were therefore determined in patients with acute or chronic leukaemia, malignant lymphoma or myelodysplastic syndromes. Significant elevation of serum E-cadherin levels was detected in patients with haematological malignancies, and between types of acute leukaemias or subtypes of myelodysplastic syndromes, stages of malignant lymphoma, and phases of chronic leukaemia, respectively, compared with those in healthy adult volunteers. These findings suggest that E-cadherin might be expressed in malignant haematopoietic cells and might be useful as a diagnostic indicator in haematological malignancies.

Acute rhabdomyolysis following administration of high-dose cyclophosphamide: case report.

Rhabdomyolysis is an unusual complication of hematopoietic stem cell transplantation (HSCT). Cyclophosphamide has been one of the key drugs in the most common preparative regimen for HSCT. We present here a rare case of acute rhabdomyolysis following administration of high-dose cyclophosphamide. A 47-year-old woman with adult T-cell leukemia in remission was treated with high-dose cyclophosphamide as a preparative regimen for allogeneic bone marrow transplantation. Nineteen hours later, general convulsions and acidosis suddenly occurred. Levels of serum creatine kinase (skeletal muscle type), myoglobin, and aldolase were markedly elevated to 32870 IU/l, 640 ng/ml, and 240.3 IU/l, respectively. Rhabdomyolysis caused by high-dose cyclophosphamide was diagnosed, and the preparative chemotherapy was discontinued. Subsequently, her muscular signs and symptoms improved, and the results of laboratory examinations returned to normal after 2 weeks. She had previously been treated with conventional doses of cyclophosphamide, doxorubicin, vincristine, and prednisolone without evidence of rhabdomyolysis. Acute rhabdomyolysis may be an adverse effect specific to high-dose cyclophosphamide therapy.

[CD13- and CD33-negative acute myelocytic leukemia (FAB classification; M2) with morphological changes and CD13 expression on recurrence].

A 42-year-old man was diagnosed as having acute myelocytic leukemia in July 1998. The leukemic cells tended to be differentiated, and on the basis of positive peroxidase staining, this case was considered to be AML (M2) according to the FAB classification. t(8;21)(q22;q22) chromosomal abnormality was observed, but surface antigen analysis revealed no expression of either CD13 or CD33, a finding characteristic of myelocytic leukemia. Combination chemotherapy resulted in complete remission, and allogeneic bone marrow transplantation was performed with donor cells from the patient's sister. Unfortunately, however, the patient died about 18 months after the onset of leukemia. Comparison of the findings at recurrence with those at initial diagnosis revealed morphological changes in non-differentiated immature cells (AML-M1) and CD13 surface antigen expression. This was considered to be a rare case of AML with neither CD13 nor CD33 expression at onset, but with CD13 expression at recurrence.

Nodal gamma/delta T cell lymphoma in complete remission following allogeneic bone marrow transplantation from an HLA-matched unrelated donor.

Gamma/delta T cell lymphoma is very rare, and usually occurs as an extranodal tumor. We describe the case of a 16-year-old Japanese man with an unusual nodal gamma/delta T cell lymphoma with generalized lymphadenopathy and bone marrow involvement. No tumor involvement was observed in the liver, spleen, or nasal cavity. Examination for surface antigens on lymphoma cells revealed a unique phenotype, positive for CD3 and T cell receptor (TCR) gamma/delta, but negative for CD2. Genotypic analysis revealed the tumor to be of monoclonal origin and characterized by TCR gamma-chain gene rearrangement, but there was no rearrangement of the TCR beta-chain gene. Our patient's tumor responded to combination chemotherapy and subsequent allogeneic bone marrow transplantation from an HLA-matched unrelated donor. He has remained well and free of disease for 35 months.

Blastic NK-cell lymphoma/leukemia with T-cell receptor gamma rearrangement.

A 79-year-old Japanese man was admitted to our hospital with dyspnea in June 1999. Physical examination revealed general exanthema, hepatosplenomegaly, and lymphadenopathy. Increased numbers of abnormal cells were observed in peripheral blood; these cells were of lymphoblastic morphology with high nuclear/cytoplasm ratios and few azurophilic granules. Immunophenotypic analysis revealed positivity for CD2, CD4, CD56, and HLA-DR, and negativity for CD3, CD13, CD16, CD33, CD34, and T cell receptor (TCR). On genotypic analysis, TCRgamma chain was rearranged, but neither the TCRbeta chain nor TCRdelta chain. Despite an initial good response to chemotherapy the disease relapsed in the early stage, and the patient died 6 months after diagnosis.

Analysis of IL-18 bioactivity and IL-18 mRNA in three patients with adult T-cell leukaemia, acute mixed lineage leukaemia, and acute lymphocytic leukaemia accompanied with high serum IL-18 levels.

Interleukin-18 (IL-18) bioactivity in sera and IL-18 mRNA expression in leukaemia cells of three patients with adult T-cell leukaemia (ATL), acute mixed lineage leukaemia (AMLL) and acute lymphocytic leukaemia (ALL) accompanied with high serum IL-18 levels have been analysed. There was little serum IL-18 bioactivity in the three patients with ATL, AMLL and ALL, while IL-18 mRNA expression was detected in leukaemia cells of all three patients.

Automated bone marrow analysis using the CD4000 automated haematology analyser.

At present, bone marrow analysis is performed microscopically, but is time consuming and labour intensive. No automated methods have been successfully applied to classification of bone marrows cells because automated blood cell analysers have been incapable of identifying erythroblasts. The present study was designed to evaluate automated analysis of bone marrow aspirates with the CELL-DYN 4000 (CD4000) haematology analyser, which enables automated determination of erythroblast counts in both the normal mode (haemolytic time; 11.5 s) and the resistant RBC mode (34.0 s). The percentages of subpopulations including lymphocytes, neutrophils and erythroblasts were obtained with the CD4000, and as a reference, differential counts by microscopic observation of May-Grünwald-Giesa-stained films of bone marrow aspirates were performed (n=98). Significant correlations (P < 0.01) between the results obtained with the two methods were observed for total nucleated cell count and lymphocytes, neutrophils, erythroblasts and myeloid/erythroid (M/E) ratio. However, there were biases in the average percentages of erythroblasts, lymphocytes and M/E ratio obtained using the normal mode with the CD4000 toward values lower than those obtained with the microscopic method. Using the RBC resistant mode with the CD4000, the average percentages of erythroblasts, lymphocytes and M/E ratio approximated those obtained with the microscopic method. In conclusion, the CD4000 in resistant RBC mode is more useful for analysis of bone marrow aspirates than is the normal mode, because the former better approximates the M/E ratio than the latter.

[Human parvovirus B19-induced aplastic crisis in a patient treated with fibrin sealant].

A 42-year-old woman underwent a myomectomy on March 31, 1998. On the 10th postoperative day, leukopenia and reticulocytopenia were observed. Bone marrow aspiration revealed severe erythroblastopenia with giant proerythroblasts, suggesting a recent parvovirus infection. Both anti-parvovirus B19 IgM antibody and IgG antibody seroconversion was observed, and human parvovirus B19 DNA was detected by polymerase chain reaction (PCR) methods. The hematologic data on the patient rapidly improved thereafter. It was determined that acute-phase serum had inhibited CFU-E and BFU-E derived colony formation. Based on these findings, parvovirus B19-induced aplastic crisis was diagnosed. Fibrin sealant, which is a typical hemostatic agent produced from blood, had been during the operation. Human parvovirus B19 DNA was detected in the fibrin sealant by PCR. Our case report documents the transmission of human parvovirus B19 by fibrin sealant.

1513-DMIa and 1513-DMIb, DNA methyltransferase inhibitors produced by Streptomyces sp. strain No. 1513.

Two new methyltransferase inhibitors were isolated from the culture filtrate of Streptomyces sp. strain No. 1513 and named 1513-DMIa and 1513-DMIb. 1513-DMIa and 1513-DMIb were distinguished in certain properties from DMI-1, DMI-2, DMI-3 and DMI-4 previously reported. The molecular weight of 1513-DMIa and 1513-DMIb were estimated to be 576 and 8400 from the results of FAB-MS and gel filtration, respectively. The inhibitory activities of 1513-DMIa and 1513-DMIb were shown to be pH- and temperature-dependent and both inhibited M. EcoRI in an uncompetitive manner with respect to DNA or S-adenosylmethionine (SAM).

Peripheral blood stem cell transplantation for non-Hodgkin's lymphoma in complete remission.

High-dose chemotherapy with PBSCT as consolidation therapy was administered to 15 patients with non-Hodgkin's lymphoma admitted to our department between November 1991 and March 1998. The average number of CD34-positive cells harvested was 3.1 x 10(6)/kg and the median time to a granulocyte count above 500/microliter was 12.8 days. Of the patients, one died from cytomegalovirus pneumomonia after WBC count recovered, 2 relapsed, and 12 patients are still alive and disease-free. The average disease-free survival period was 50.0 months. Stem cell harvesting is relatively simple and hematopoietic recovery is mory rapid with PBSCT than with ABMT. Our findings suggest that PBSCT can be used for consolidation therapy in NHL patients who have achieved complete remission with standard chemotherapy.

Interrelationship of fever, immune response and aging in mice.

Fever is the cardinal manifestation of infection and may be blunted in certain infected elderly individuals. It is known that elevated body temperature enhances both the inflammation response and immune function, resulting in increased host resistance to infection. Recently, it has been suggested that the elevation of body temperature and the activation of lymphocytes by IL-1 are interrelated host effects. However, the question of whether fever response in vivo is closely correlated to cell-mediated immune parameters is unknown. In this study, a well-defined murine model was used to study the relationships between aging, fever and cell-mediated immune response. Thus, measurements of rectal temperature changes were made in individual young (4-6 months) and old (26-27 months) BALB/c mice to determine their ability to respond to endogenous pyrogen (recombinant IL-1). Splenic cells from these animals were used to assess T- and B-cell proliferation, production of interleukin 1 (IL-1) and IL-2. The results revealed that the proliferative capacity and the IL-1 and IL-2 producing capacity of splenic cells from old mice were markedly decreased. However, aging did not significantly affect the mean febrile responses in old mice following rIL-1 injections. Finally, there was no significant correlation between in vivo fever responses and the immune parameters measured in vitro in both young and old mice.

5838-DNI, a deoxyribonuclease inhibitor produced by Streptomyces sp. strain no. A-5838.

5838-DNI, an inhibitor of deoxyribonuclease (DNase) II from porcine spleen was produced by Streptomyces sp. strain No. A-5838. The structure of 5838-DNI was shown to be 1,4,4a,5,12,12a-hexahydro-4,4a,11,12a-tetrahydroxy-3,8-dimethoxy-9- methoxycarbonyl-10-methyl-1,5,12-trioxo naphthacene. Although similar in structure to tetracenomycin C, which is an antibiotic against Gram-positive bacteria, 5838-DNI has different antibacterial activity. 5838-DNI was distinguished from 5923-DNI, a previously reported DNase II inhibitor, in inhibitory activity against each enzyme. 5838-DNI showed dependency of inhibition on pH and temperature, and inhibited phosphodiesterase I in a competitive manner. These data suggest that 5838-DNI is the first reported example of an inhibitor of microbial origin which is able to inhibit DNase II and phosphodiesterase I.

Clinically inapparent (asymptomatic) bacteriuria in ambulatory elderly men: epidemiological, clinical, and microbiological findings.

In a prospective longitudinal study, ambulatory elderly men were followed from 1 to 4.5 years to gain insight into the prevalence rates, clinical characteristics, and patterns of clinically inapparent (asymptomatic) bacteriuria (CIB). The prevalence of CIB was 12% (29/238) and increases with age. Unlike the gram-negative organisms that cause overt urinary tract infection in this age group, gram-positive organisms dominated the CIB group. Both the CIB and abacteriuric patients have multiple chronic medical conditions and are indistinguishable on that basis. Twenty-nine elderly men with bacteriuria and 105 abacteriuric subjects were followed with serial urine cultures. During the study period the bacteriuric subjects exhibited spontaneous temporary or permanent resolution (76%, 22/29), intermittency (21%, 6/29), and probable bacterial persistence (38%, 11/29). No consistent pattern of bacteriuria was evident. Therefore, antimicrobial therapy is not warranted in the treatment of asymptomatic or clinically inapparent bacteriuria in ambulatory elderly men.

Fever response in old and young mice after injection of interleukin 1.

In an attempt to determine why certain elderly humans show a blunted febrile response to infection, a mouse model was developed. Interleukin 1 (IL1), previously called endogenous pyrogen (EP), the predominantly macrophage-derived mediator of fever, was obtained from supernatants that were generated by stimulating macrophages obtained from young and old mice. The dose versus febrile response curves were generated by injecting crude or partially purified supernatants that contained IL1 into the tail veins of young and old mice. Old mice responded with less fever than young mice to both crude and partially purified supernatants, suggesting that one mechanism for the blunted fever response observed with aging may be an altered response to EP.

Pyuria: its predictive value of asymptomatic bacteriuria in ambulatory elderly men.

A total of 664 urine samples collected from 254 asymptomatic, ambulatory, elderly men was examined for pyuria with a standardized glass slide microscopy method. The presence or absence of more than 10 polymorphonuclear leukocytes per high power field was highly predictive of the presence or absence of significant bacteriuria, respectively. The determination of pyuria in this select population may be a useful and inexpensive test to screen for bacteriuria.