Evaluation of acoustic properties of the live human smooth-muscle cell using scanning acoustic microscopy

This study was performed to measure the acoustic propagation speed in live human aortic smooth-muscle cells (HASMC), using scanning acoustic microscopy (SAM) and a novel measurement theory that permits the measurement of the acoustic propagation speed in biological samples of unknown thickness. C-mode and X–Z-mode images of HASMC under three different conditions: growing (G); differential (D); and on hypotonic loading (H), were acquired using 100-MHz, 450-MHz and 600-MHz ultrasound. The images exhibit features related to the cell surface curvature and intracellular structure. The theory supporting the methodology is derived in this article and makes use of the interference fringes within the focusing lens of the high-frequency transducer. The propagation speed in the cells was calculated from the location of the interference fringe on the C-mode images and the fringe shift on the X–Y-mode images with 450-MHz ultrasound. The propagation speed in D (1624 ± 16 m/s) was significantly higher than those in G (1571 ± 14 m/s, p < 0.05) and H (1585 ± 8 m/s, p < 0.05). Scanning acoustic microscope measurements, along with the described theory, are useful for studying the acoustic properties of live cells ex vivo and have applications in both pathophysiology and biomechanics.     

Platelet glucose-6-phosphatase activity in patients with von Gierke's disease and their parents

The present report describes the results of an assay of platelet glucose-6-phosphatase (G-6-Pase) in patients with von Gierke's disease and their parents. The G-6-Pase activity of the patients was found to be about 10–25% of that of controls, while that of the parents showed an intermediate value between that of the patients and that of the controls, which led to the conclusion that the parents were the heterozygotes of the disease. These results suggest a possible method for the diagnosis of G-6-Pase deficiency as well as for detection of the heterozygotes of the disease by a platelet G-6-Pase assay without making a G-6-Pase assay of liver biopsy materials.As for the separation of platelets, a differential centrifugation was undertaken using Na₂ -EDTA as an anticoagulant. In the G-6-Pase assay, universally ¹⁴C-labelled glucose 6-phosphate ([U-¹⁴C]-G-6-P) was used as a substrate and the enzyme activity was measured by the amount of glucose liberated by the platelet suspension after the reaction. The effect of non-specific phosphatase was estimated by using universally ¹⁴C-labelled glucose 1-phosphate ([U-¹⁴C] -G -6-P) as substrate.In the diagnosis of von Gierke's disease, as well as the detection of the heterozygotes of the disease by the G-6-Pase assay using platelets as material, the effect of non-specific phosphatase was considered to be negligible.     

Nanomolar concentrations of tri-n-butyltin facilitate gamma-aminobutyric acidergic synaptic transmission in rat hypothalamic neurons

Tri-n-butyltin (TBT), an environmental pollutant, is accumulated in edible mollusks and fishes. It has also become a health concern in today's society. In the present study, to elucidate the possible neurotoxic action of TBT, the effect on spontaneous gamma-aminobutyric acid (GABA) release from GABAergic nerve terminals projecting to rat ventromedial hypothalamic neurons was examined using "synaptic bouton" preparation with a nystatin perforated patch recording mode under voltage-clamp conditions. The threshold concentration of TBT to affect the synaptic transmission was 10 to 30 nM. TBT at 30 nM or higher concentrations increased the frequency of GABAergic miniature inhibitory postsynaptic currents in a dose-dependent manner, whereas the current amplitude and current kinetics were not affected. The removal of either external Ca2+ or application of Cd2+ attenuated the TBT-induced facilitation of neurotransmission. TBT at 1 microM induced an inward current in more than one-half of the cells. This current persisted even after TBT was washed out. The present results indicate that TBT at environmentally relevant concentrations (30-100 nM) facilitates the GABAergic neurotransmission in the mammalian brain and the external Ca2+ is needed in this facilitation. Because the concentration of TBT accumulated in some mollusks and fishes has been reported to reach levels of 100 nM or more, such accumulation of TBT in some mollusks and fishes is thus suggested to be hazardous to the health of humans.     

Neutrophil-derived hydroxyl radicals mediate interleukin-8-induced increases in tetragastrin-stimulated acid secretion in rats

Until recently, the secretion of gastric acid, which plays an important role in the pathogenesis of peptic ulcer, was thought to be controlled by diet, the autonomic nerves and gut hormones. However, peptic ulcer is now known to be caused by the infection with Helicobacter pylori (H. pylori), so it is possible that inflammation modifies the secretion of gastric acid. We used gastric-lumen-perfused rats to first examine the effect of interleukin-8 (IL-8) on acid secretion and then the involvement of free radicals and neutrophils in the action of IL-8. IL-8 enhanced tetragastrin-stimulated acid secretion and free radical scavengers or inhibitors and the pretreatment with anti-rat neutrophil serum inhibited this effect, which indicates that IL-8 enhances gastrin-stimulated acid secretion and that neutrophil-derived hydroxyl radicals mediate the IL-8-induced increase in acid secretion.     

Outpatient chemotherapy and clinical pathway for non-small cell lung cancer]

We investigated the usefulness of outpatient chemotherapy in 54 cases of non-small cell lung cancer in which outpatient chemotherapy was performed between August 1999 and October 2001. This chemotherapy accounted for 67% of all chemotherapy. Assessment of therapeutic effect revealed a PR in 14 of the 54 cases, and the efficacy rate was 26%. Therapeutic effect according to chemotherapy regimen revealed the highest efficacy rate, 50%, for paclitaxel + CBDCA. The median survival time was 14.7 months, and the 1-year survival rate was 61.1%. On the basis of the above results, a 16-day inpatient clinical pathway using weekly paclitaxel + CBDCA was devised for non-small cell lung cancer. The aim was to shorten the number of inpatient days, standardize treatment, and introduce outpatient chemotherapy. The clinical pathway was introduced in 8 patients with recurrent non-small cell lung cancer between August and October 2002. Variance was found only in one patient whose hospital discharge had to be postponed by two days because of a Grade 3 side effect. Introduction of a clinical pathway with weekly paclitaxel + CBDCA successfully reduced the inpatient days to an average of 16.3 days.     

Discovery of immunostimulatory CpG-DNA and its application to tuberculosis vaccine development

DNA containing an unmethylated CpG motif has a potent immunostimulatory effect on the vertebrate immune system. Because such CpG motifs are relatively common in bacterial DNA, but rare in mammalian animal and plant DNA, they may be an evolutionary adaptation augmenting innate immunity, most likely in response to pathogens that replicate within the host cells, such as viruses and intracellular bacteria. Microbial infection induces innate immunity by triggering pattern-recognition systems. The infected cells produce proinflammatory cytokines that directly combat microbial invaders and express costimulating surface molecules, which develop adaptive immunity by inducing distinct T cell differentiation. Bacterial DNA with unmethylated CpG-DNA stimulates vertebrate immature immune cells to induce maturation and to produce TNF-alpha as well as Th1-type cytokines, IL-12 and IFN-gamma. Therefore, CpG-DNA functions as an adjuvant for regulating the initiation of Th1 differentiation. The roles of immunostimulatory CpG motifs in DNA vaccine developments and in therapeutic applications have been discussed.     

Imbalance between thrombin and plasmin activity in disseminated intravascular coagulation. Assessment by the thrombin-antithrombin-III complex/plasmin-alpha-2-antiplasmin complex ratio.

We investigated the imbalance between thrombin and plasmin activity in vivo with various grades of severity of disseminated intravascular coagulation (DIC) in relation to the underlying diseases. Plasma thrombin-antithrombin-III complex (TAT) and plasmin-alpha 2-antiplasmin complex (PAP) levels were measured in 133 blood samples obtained from patients with DIC. The TAT/PAP ratio was higher in patients with sepsis or solid cancer than in those with hematologic malignancies. In acute promyelocytic leukemia (APL), the TAT levels were the highest, but the PAP levels were even higher and the TAT/PAP ratio was the lowest. As for the severity of DIC, in mild DIC, both thrombin and plasmin activities were increased. In moderate DIC, the TAT/PAP ratio increased, and thrombin activity was much more predominant. However, in severe DIC, the ratio decreased, and plasmin activity became excessive. In 3 patients with acute myeloblastic leukemia, APL and pancreatic cancer, respectively, the PAP level remained high during heparin therapy although the TAT level was decreased. When tranexamic acid was given, the PAP level was selectively reduced, and the TAT/PAP ratio was markedly decreased along with clinical improvement. These results indicate that monitoring of the TAT/PAP ratio may contribute to decisions regarding the institution and performance of combination therapy for DIC using anticoagulants and antifibrinolytic agents.     

Remission induction therapy containing rituximab markedly improved the outcome of untreated mature B cell lymphoma

Many controlled clinical trials have proven that rituximab improves the clinical outcome of patients with mature B cell lymphoma. This study was conducted to assess the contribution of rituximab in the actual clinical practice. Patients with newly diagnosed mature B cell lymphoma treated at 20 National Hospital Organization hospitals from January 2000 to December 2004 were consecutively registered. Rituximab was approved in September 2002 for indolent B cell lymphoma and in September 2003 for aggressive B cell lymphoma in Japan. The patients were divided into two groups depending on whether they received induction therapy containing rituximab. The endpoint was to evaluate the rituximab benefit based on 2-year progression-free survival (PFS) and 2-year overall survival (OS). A total 1126 patients received chemotherapies. Of these, 762 were diagnosed as diffuse large B cell lymphoma (DLBCL) and 215 as follicular lymphoma (FL). PFS and OS were markedly improved in the rituximab group compared with the non-rituximab group in patients with DLBCL (both P < 0.001) and in patients with FL (P < 0.001 and P = 0.003 respectively). Rituximab, when used for remission induction therapy, significantly improved the clinical outcome of the mature B cell lymphoma patient in actual clinical practice.     

Virus-associated haemophagocytic syndrome responsive to steroid pulse therapy and double filtration plasmapheresis

We present an adult patient with haemophagocytic syndrome (HPS) successfully treated with a combination of steroid pulse therapy and double filtration plasmapheresis (DFPP). A 58-year-old male was admitted with high fever, severe renal dysfunction, liver dysfunction and an increased level of lactate dehydrogenase. A serological test for Epstein-Barr (EB) virus showed an elevation of EBNA-IgM antibody titre. There were increased haemophagocytic histiocytes in the bone marrow in addition to thrombocytopenia and disseminated intravascular coagulation (DIC) accompanied by organ dysfunction. EB virus associated haemophagocytic syndrome was diagnosed. On admission, interferon (IFN)-gamma, interleukin (IL)-6, IL-8, granulocyte colony-stimulating factor (G-CSF) and macrophage (M)-CSF were elevated, and were promptly normalized after steroid pulse therapy was initiated. G-CSF and M-CSF gradually decreased after DFPPs was started. To control hypercytokinaemia until treatment for the underlying disease is initiated, steroid pulse therapy and double filtration plasmapheresis are useful.