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Biomedical subjects

T Nakashima

Publications and source records attributed to T Nakashima.

At least 19 recordsLinked to original sources

Effects of gelsolin on human platelet cytosolic phosphoinositide-phospholipase C isozymes.

The effective resolution of human platelet cytosolic phosphoinositide-phospholipase C (PLC) revealed five distinct activity peaks by Q-Sepharose and heparin-Sepharose column chromatographies when assayed using phosphatidylinositol (PI) and phosphatidylinositol 4,5-bisphosphate (PIP2). The results of Western blotting analysis with various antibodies against PLC isozymes showed that peak-Ia (PLC-delta type), peak-Ib (PLC-gamma 1 type), and peak-IIc (PLC-beta type) and two unidentified activity peaks (PLC-IIa and PLC-IIb) were present in human platelet cytosol. A protein with guanosine 5'-3-O-(thio)triphosphate-binding activity was coeluted with the PLC-IIa and was purified to homogeneity. It exhibited 86- and 42-kDa polypeptide bands upon sodium dodecyl sulfate-polyacrylamide gel electrophoresis which were identified as gelsolin and actin by immunostaining, respectively. Large amounts of gelsolin/actin (1:1) complex "gelsolin complex" were detected in the PLC-delta and PLC-gamma 1 fractions. The PLC-gamma 1 and the gelsolin complex were co-immunoprecipitated by the antibody raised against PLC-gamma 1. Furthermore, the partially purified bovine brain PLC-gamma 1 fraction also was found to be associated with the gelsolin complex and the association was released by the addition of 1% sodium cholate. This finding has prompted us to examine effects of the gelsolin complex and the free gelsolin on activities of the above PLC isoforms from platelet cytosol. The gelsolin complex did not affect the PIP2 hydrolyzing activities of all PLC isoforms. In contrast, the purified gelsolin inhibited distinctly PIP2 hydrolyses by PLC-Ia (delta), PLC-Ib (gamma 1), and PLC-IIa (unidentified), whereas the inhibitory effects for PLC-IIb (unidentified) and PLC-IIc (beta) were moderate. The inhibitory effect of gelsolin on PIP2-hydrolysis by PLC-gamma 1 was diminished by a large amount of PIP2 substrate. These results suggested that the inhibition of PLC by gelsolin is due to sequestration of substrate PIP2 by its competitive binding.

Animals

Chemosensitivity and DNA ploidy in head and neck squamous cell carcinomas.

The succinate dehydrogenase activity and cellular DNA content of human head and neck squamous cell carcinomas were examined, and the chemosensitivity and ploidy status were compared with histologic differentiation. The average decrease of enzyme activity in the poorly differentiated squamous cell carcinomas was significantly greater than that of well- and moderately differentiated squamous cell carcinomas. Statistically significant differences were also noted with relation to the original site of the primary tumor. The chemosensitivity of a tumor with DNA aneuploidy tended to be lower among the well- and moderately differentiated squamous cell carcinomas and higher among the poorly differentiated type. We conclude from this study that simultaneous analysis of the chemosensitivity and DNA ploidy will aid not only in selecting effective antitumor drugs but also in predicting changes in cellular characteristics during the course of disease.

Antineoplastic Agents

Endonuclease analyses of DNA of human herpesvirus-6 isolated from blood before and after bone marrow transplantation.

Three strains of human herpesvirus-6 (HHV-6) were isolated from peripheral blood mononuclear cells of a leukemic child with the antibody of HHV-6 before and after bone marrow transplantation (BMT); two strains were obtained before BMT and one after BMT. The DNA extracted from the three isolates was analyzed by six different restriction endonucleases. The cleavage profiles of two strains obtained before BMT were different, but the third strain isolated after BMT was identical with one of the two, which suggest reactivation of HHV-6 from the recipient's own body after BMT and possible mutation or super-infection of the virus in an immunocompromised patient.

Bone Marrow Transplantation

Activation of human herpesvirus-6 in children with acute measles.

Virological and serological studies were carried out prospectively to evaluate the possible activation of human herpesvirus-6 (HHV-6) in 50 infants and children with acute measles by isolation of HHV-6 from peripheral blood and by determining neutralizing antibodies to the virus. All but 5 patients (90%) were seropositive to HHV-6 in the acute stage of measles and 18 (40%) had a significant increase in HHV-6 antibody titers thereafter, whereas only 2 of 27 patients who were initially seropositive to Epstein-Barr virus (EBV) viral capsid antigen (VCA) had a significant rise in antibody titers to EBV VCA. Among 18 patients with a significant increase in HHV-6 titers, the virus was isolated from the peripheral blood mononuclear cells of three patients in the early convalescent stage of measles. These results indicate that activation of HHV-6 may occur frequently a few weeks after primary infection with the measles virus.

Acute Disease

Nicotine-induced sensitization to ambulatory stimulant effect produced by daily administration into the ventral tegmental area and the nucleus accumbens in rats.

Bilateral injections of nicotine (30 micrograms/side) into the ventral tegmental area (VTA) and the nucleus accumbens (NACC) increased the ambulatory activity in rats. Moreover, daily injections of nicotine (10, 20 and 30 micrograms/side) into the VTA and the NACC for 6 successive days produced sensitization to the ambulatory stimulant effect of nicotine. Sensitization produced by daily injections of nicotine (20 micrograms/side) into both the sites was maintained for withdrawal periods of 10 days. Mecamylamine (2 mg/kg, i.p.), SCH23390 (0.05 mg/kg, i.p.) and spiperone (0.1 mg/kg, i.p.) antagonized nicotine-induced sensitization to the ambulatory stimulant nicotine-induced sensitization to the ambulatory stimulant effect produced by daily injections into the VTA. These results suggest that nicotine-induced sensitization to the ambulatory stimulant effect involves the stimulation of the mesolimbic dopaminergic pathway through the nicotinic acetylcholine receptor (nAChR) in the VTA and the NACC.

Animals

The responses to phorbol esters which stimulated protein kinase C in canine Purkinje fibers.

1. The effects of phorbol esters on canine Purkinje fibers were examined using conventional microelectrode techniques. 2. 12-O-Tetradecanoylphorbol-13-acetate (TPA) and 4-beta-phorbol-12,13-dibutyrate (PDB), which are specific activators of protein kinase C (PKC), decreased the action potential amplitude and the maximum rate of depolarization (Vmax) at 3 x 10(-7) M or higher. These phorbol esters had little effect on the resting potential. 3. PDB (1-3 x 10(-7) M) also reduced the contractile force, accompanied with initial increase (in 5 out of 8 experiments), whereas TPA did not decrease it to any significant extent. 4. An inactive analog of phorbol esters, 4-alpha-phorbol-12,13-didecanoate (PDD), decreased the action potential amplitude and Vmax, and slightly increased the action potential duration. However, PDD failed to produce any inotropic effect. 5. Post-rest potentiation of the contractile force after a rest from stimulation for 30 sec was inhibited in the presence of 3-10 x 10(-7) M TPA or 3 x 10(-7) M PDB. 6. Isoproterenol 10(-7) M augmented the action of PDB 3 x 10(-7) M. 7. These results suggest that activation of PKC may modulate myocardial Ca2+ homeostasis and influence the excitation-contraction process.

Action Potentials

The effects of m-octopamine on salivary flow rates and protein secretion by rat submandibular glands.

1. m-Octopamine given i.v. or i.p. was a potent sialogogue for rat salivary glands. 2. Salivation in response to i.v. m-octopamine was completely abolished by prazosin and phenoxybenzamine. 3. The alpha-type of proteins were secreted in response to all doses of i.v. and i.p. m-octopamine and these were converted into the beta-type with prazosin, but not with yohimbine. 4. m-Octopamine stimulated both alpha- and beta-adrenoceptors and was a much more selective alpha 1-agonist than was the p-isomer.

Adrenergic beta-Antagonists

Composition of urinary calculi related to urinary tract infection.

The composition of 3,084 urinary calculi was determined using an infrared spectrophotometer. Mixed calcium oxalate-calcium phosphate stones were most frequently implicated. Of the urinary calculi analyzed 199 were associated with urinary tract infection. Escherichia coli was most frequently isolated (43 strains) and urease-producing organisms, such as Proteus mirabilis, were cultured from 40 patients. The core culture of 20 staghorn calculi yielded 15 isolates from 14 stones. There were 13 identical species isolated from the urine and stone specimens of 13 patients (65%), including 7 strains of P. mirabilis. These results suggest that cultures of urine specimens of urolithiasis patients, especially those with staghorn calculi, may help to elucidate the bacteriology of the stones.

Adolescent

Activation of phospholipase D by platelet-derived growth factor (PDGF) in rat C6 glioma cells: possible role in mitogenic signal transduction.

The effects of platelet-derived growth factor (PDGF) on phospholipase D (PLD) activity and deoxyribonucleic acid (DNA) synthesis in rat C6 glioma cells have been investigated. Pretreatment of serum-starved C6 cells with PDGF results in enhanced choline production and the phosphatidylethanol (PEt) formation in the presence of ethanol, indicating the activation of PLD acting on phosphatidylcholine (PC). The dose-response curve for choline generation and DNA synthesis were comparable. In addition, the effects of PDGF on both PEt formation and [3H]thymidine incorporation into acid-precipitable material was blocked by the potent protein kinase C (PKC) inhibitor 1-(5-isoquinolinesulphonyl)-2-methylpiperazine (H-7) but not by N-(2-guanidinoethyl)-5-isoquinolinesulphonamide (HA1004), a relatively weak inhibitor of PKC, suggesting that PDGF plays an important role as a positive regulator of glioma cell growth via a PLD-mediated mitogenic signal transduction cascades, which depends largely on the activation of PKC.

Animals

A prospective study of human herpesvirus-6 infection in renal transplantation.

Sixty-five kidney transplant recipients and their (22 living related and 43 cadaveric) donors were studied prospectively to determine the relationship between kidney transplantation and human herpesvirus-6 (HHV-6) infection. The virus isolation from peripheral blood and other tissues and sequential determination of neutralizing antibodies to HHV-6 were performed during 3 months following the transplantation. All of the donors and their recipients examined had neutralizing antibodies to HHV-6 at the time of renal transplantation and the virus was not isolated from them. HHV-6 was isolated from 3 renal tissues (2 living related and 1 cadaveric) obtained during transplant surgery, but not from their blood at that time. HHV-6 viremia occurred in 9 (14%) of the 65 recipients around 2 to 4 weeks after the transplantation. An additional 27 recipients showed a significant rise in the antibody titer. Thus, the infection with HHV-6 was confirmed in 36 (55%) of the 65. These results indicate that the virus is activated in many cases in the early posttransplant period and that HHV-6 establishes in vivo latency in the kidney tissue. There was no correlation between HHV-6 infection and acute rejection or the antirejection prophylaxis.

Adolescent

IgM neutralizing antibody responses to human herpesvirus-6 in patients with exanthem subitum or organ transplantation.

The assay for detecting IgM neutralizing (NT) antibody activity to human herpesvirus-6 (HHV-6) was developed by using pretreatment of blood sample with staphylococcal protein A. The activity was mostly present in IgM fractions of serum but not in IgA fractions separated by ultracentrifugation. The assay was used for seroepidemiological studies for HHV-6 infection. In primary HHV-6 infection, IgM NT antibodies appeared 5 to 7 days after onset of exanthem subitum, reached maximum titers at 2 to 3 weeks, and tended to decline to undetectable levels after 2 months. In contrast, reactivation of HHV-6 observed in organ transplants showed somewhat greater degree of IgM NT antibody responses that persisted for 2 to 3 months and became undetectable 5 to 6 months after transplantation. The level and persistence of NT antibody titers measured by the conventional method was generally greater than those of the IgM titers. The prevalence of the IgM NT antibodies was examined in healthy individuals. The antibody was first detected at 4 to 7 months of age (5%), reached maximum level at 8 to 11 months (40%), and was detectable by 4 to 6 years (17%). A few (4 to 5%) of adolescents and adults were positive for the antibody.

Adolescent

Comparison of severity of viremia and antibody responses between infants and children with measles.

Severity of viremia and neutralizing antibody responses were compared between nine young infants (less than or equal to 10 months) with measles and 18 infants and children (greater than or equal to 11 months) with ordinary measles. Peripheral blood mononuclear cell (PBMC)-associated viremia was detected between the first day of elevation of fever (day 0) and day 5 of the disease in the former group, whereas PBMC-associated and cell-free viremia were detected between day 0 and day 14 in the latter group. The number of infected PBMC during the first 7 days of the disease was 3.22 +/- 1.07 (log10, mean +/- s.d.) per 10 million PBMC in the former group, which was significantly smaller (P = 0.02) than that of the latter group (4.21 +/- 1.18). The former group reached the maximum level of antibody earlier than the latter group.

Age Factors

Thiazide treatment for calcium urolithiasis in patients with idiopathic hypercalciuria.

In a randomised trial based on a parallel design to determine the prophylactic effect of thiazide on stone formation, 210 calcium urolithiasis patients with idiopathic hypercalciuria were allocated either to treatment with trichlormethiazide (4 mg/day) or no treatment with only close follow-up; 35 patients were excluded for various reasons, including voluntary withdrawal. The background of the remaining 175 patients (82 in the thiazide group and 93 in the control group), including age and sex, was similar for both groups. In patients treated with thiazide there was a statistically significant fall in urinary calcium output. Statistical analyses also demonstrated that the stone formation rate in the thiazide group was significantly less than that in the control group. Adverse clinical reactions probably due to the drug were observed in 9 patients. These findings indicate that trichlormethiazide has a prophylactic effect on calcium urolithiasis in patients with idiopathic hypercalciuria.

Adult

Thermal and PGE2 sensitivity of the organum vasculosum lamina terminalis region and preoptic area in rat brain slices.

1. The effects of local applications of prostaglandin E2 (PGE2) on the unit activity of fifty-one neurones in the organum vasculosum lamina terminalis (OVLT) region and fifty-eight neurones in the preoptic area (POA) were investigated in small tissue slices from the rat hypothalamus containing the OVLT and POA isolated from each other. 2. Of these, thirty OVLT and twenty-eight POA neurones were warm sensitive and increased their discharge rate in response to a rise in tissue temperature. One OVLT neurone and one POA neurone were cold sensitive and showed the opposite type of responses to changes in temperature. The thermosensitivity of these neurones was still observed in a Ca2+ free-high Mg2+ solution. 3. Perfusion with PGE2 in doses between 1 and 250 nM changed the discharge rate in forty-two of fifty-one OVLT neurones and in thirty-two of fifty-eight POA neurones in a dose-dependent manner. The responses to PGE2 were not lost during synaptic blockade. The threshold dose of PGE2 to alter the discharge rate of the OVLT neurones (4.8 +/- 1.1 (S.E.M.) nM, n = 16) was significantly lower than that of the POA neurones (40.9 +/- 12.2 nM, n = 16). 4. Fifteen of forty-two OVLT neurones exhibited the responses with a slower onset (latency 5-13 min) and a longer duration (20 min to 3 h), but such responses were observed in only one of thirty-two POA neurones. 5. The responses of OVLT and POA neurones to PGE2 (50-250 nM) were reversibly blocked by a concurrent application of AH6809, a prostanoid EP1 and/or a DP receptor antagonist. 6. While there was no clear correlation between the type of thermosensitivity and the type of response to PGE2 among the POA neurones, a significantly higher incidence of inhibitory response to PGE2 was found among the warm-sensitive neurones in the OVLT region. 7. The lower threshold responses to PGE2 and the higher incidence of PGE2 responsiveness among OVLT neurones are consistent with previous findings which showed that the highest density of PGE2 receptor binding and the highest pyrogenic sensitivity to microinjected PGE2 were observed in the OVLT region. The results provide further evidence for the critical involvement of the OVLT region in mediating the febrile responses to blood-borne endogenous pyrogen through the local release of PGE2.

Animals

A clinical study on patients with urinary tract infection due to coagulase-negative staphylococci.

To assess the clinical significance of coagulase-negative staphylococci (CNS) in patients with urinary tract infection (UTI), the clinical characteristics of a total of 117 patients (106 complicated UTI patients, 11 uncomplicated UTI patients) from whom CNS were isolated at urinary colony counts of 10(5) or more per ml were studied. Of the complicated UTI patients, 95 patients (89.6%) suffered from no symptoms while 11 (10.4%) had fever of 38 degrees C or greater, which was strongly suspected to be due to genitourinary tract infections. Six of these patients were managed by indwelling urinary catheters. On the other hand, all of the patients with uncomplicated UTI were young women and had typical symptoms of acute cystitis. These results suggest that CNS, which hitherto have been considered mere contaminants or benign colonization rather than true pathogens, can also cause complicated UTI requiring chemotherapy under certain conditions such as indwelling urinary catheterization and acute cystitis in sexually active women.

Adult

Fatal encephalitis/encephalopathy in primary human herpesvirus-6 infection.

An encephalitic illness with a fatal outcome occurred in a 9 month old girl with virologically confirmed exanthem subitum. Human herpes-virus-6 (HHV-6) DNA was found in the cerebrospinal fluid at the acute stage of the disease by the polymerase chain reaction, but the virus antigen was not detected in her brain tissue. This suggests that HHV-6-induced encephalitis/encephalopathy may be due to a non-infectious process.

Antigens, Viral

Comparison of in vitro anticancer chemosensitivity between human squamous cell carcinoma and adenocarcinoma.

The chemosensitivities of squamous cell carcinoma (SCC) tissues from the head and neck area were compared to findings of adenocarcinoma, mainly from digestive organs. The sensitivity of each tissue was determined using the in vitro succinate dehydrogenase (SD) inhibition test, which shares a common principle with the 3-(4,5-dimethyl-thiazolyl)-2,5-diphenyl-2H-tetrazolium bromide (MTT) assay. Tumor tissues were obtained at surgery or biopsy. Anticancer drugs tested were carboquone, Adriamycin, mitomycin C, cisplatin (CDDP), aclacinomycin A, 5-fluorouracil and 1-hexylcarbamoyl-5-fluorouracil with 10 times the peak plasma concentration, respectively. The means +/- standard deviations of SD activities in SCC tissues were significantly lower than those in adenocarcinoma tissues (p less than 0.001), and the sensitivity rates of SD activity in SCC tissues had a higher value than those in adenocarcinoma tissues (p less than 0.05), against each drug. Our study showed that CDDP-based combination regimens might be effective for SCC tissues. The chemosensitivity of each excised tissue should be tested, in order to prescribe sensitive, effective drugs for each patient.

Adenocarcinoma