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21.
SR Marengo RA Sikes P Anezinis SM Chang LW Chung 《Canadian Metallurgical Quarterly》1997,19(3):165-175
The effects of the composition of bacteriological growth media on the light output in a chemiluminometric assay of beta-galactosidase in Escherichia coli using 1,2-dioxetane substrates has been studied. In this assay a basic conflict exists between conditions that promote optimal bacterial growth and those conducive to maximal chemiluminescence. Common medium ingredients such as yeast or beef extract, protein hydrolysates and lactose suppress light emission and/or lead to high backgrounds. Quenching of light emission is probably partly due to light absorption by medium ingredients such as oxgall, and partly to interference with the reaction triggering the chemiluminescent process. Elevated backgrounds are caused by the presence of high concentrations of protein hydrolysates, which interact with the alkali in the accelerator solution. Only two purposely developed media, i.e. ILM and Colicult are shown to reconcile the requirements of growth support with that of optimal luminescent properties. 相似文献
22.
MJ Gorman DW Severson AJ Cornel FH Collins SM Paskewitz 《Canadian Metallurgical Quarterly》1997,146(3):965-971
A Plasmodium-refractory strain of Anopheles gambiae melanotically encapsulates many species of Plasmodium, whereas wild-type mosquitoes are usually susceptible. This encapsulation trait can also be observed by studying the response of refractory and susceptible strains to intrathoracically injected CM-Sephadex beads. We report the results of broad-scale quantitative trait locus (QTL) mapping of the encapsulation trait using the bead model system. Interval mapping using the method of maximum likelihood identified one major QTL, Pen1. The 13.7-cM interval containing Pen1 was defined by marker AGH157 at 8E and AGH46 at 7A on 2R. Pen1 was associated with a maximum LOD score of 9.0 and accounted for 44% of the phenotypic variance in the distribution of phenotypes in the backcross. To test if this QTL is important for encapsulation of Plasmodium berghei, F2 progeny were infected with P. berghei and evaluated for degree of parasite encapsulation. For each of the two markers that define the interval containing Pen1, a significant difference of encapsulation was seen in progeny with at least one refractory allele in contrast with homozygous susceptible progeny. These results suggest that Pen1 is important for melanotic encapsulation of Plasmodium as well as beads. 相似文献
23.
MR Garcia S Graham RA Harris SM Beverley PM Kaye 《Canadian Metallurgical Quarterly》1997,27(4):1005-1013
The activation of CD8+ T cell responses is commonplace during infection with a number of nonviral pathogens. Consequently, there has been much interest in the pathways of presentation of such exogenous antigens for major histocompatibility complex class I-restricted recognition. We had previously shown that Leishmania promastigotes transfected with the ovalbumin (OVA) gene could efficiently target OVA to the parasitophorous vacuole (PV), with subsequent recognition by class II-restricted T cells. We now report the results of studies aimed at evaluating the PV as a route of entry into the exogenous class I pathway. Bone marrow-derived macrophages can present soluble OVA (albeit at high concentrations) to the OVA(257-264)-specific T cell hybridoma 13.13. In contrast, infection with OVA-transfected Leishmania promastigotes failed to result in the stimulation of this hybridoma. This appeared unrelated to variables such as antigen concentration, parasite survival, and macrophage activation status. These results prompted an analysis of the effects of promastigotes on class I peptide binding using RMA-S cells and OVA(257-264). Our data indicate that the major surface protease of Leishmania, gp63, inhibits this interaction by virtue of its endopeptidase activity against the OVA(257-264) peptide. The data suggest that this activity, if maintained within the PV, would result in loss of the OVA(257-264) epitope. Although we can therefore draw no conclusions from these studies regarding the efficiency of the PV as a site of entry of antigen into the exogenous class I pathway, we have identified a further means by which parasites may manipulate the immune repertoire of their host. 相似文献
24.
DI Rodenhiser JD Andrews DN Mancini JH Jung SM Singh 《Canadian Metallurgical Quarterly》1997,373(2):185-195
Glutamatergic synaptic potentials induced by micromolar concentrations of the potassium conductance blocker 4-aminopyridine (4-AP) were recorded intracellularly from rat neostriatal neurons in the presence of 10 microM bicuculline (BIC). These synaptic potentials originate from neostriatal cortical and thalamic afferents and were completely blocked by 10 microM 6-cyano-7-nitroquinoxaline-2,3-dione (CNQX) plus 100 microM D-2-amino-5-phosphonovaleric acid (2-APV). Their inter-event time intervals could be fitted to exponential distributions, suggesting that they are induced randomly. Their amplitude distributions had most counts around 1 mV and fewer counts with values up to 5 mV. Since input resistance of the recorded neurons is about 40 M omega, the amplitudes agree to quantal size measurements in mammalian central neurons. The action of a D2 agonist, quinpirole, was studied on the frequency of these events. Mean amplitude of synaptic potentials was preserved in the presence of 2-10 microM quinpirole, but the frequency of 4-AP-induced glutamatergic synaptic potentials was reduced in 35% of cases. The effect was blocked by the D2 antagonist sulpiride (10 microM). Input resistance, membrane potential, or firing threshold did not change during quinpirole effect, suggesting a presynaptic site of action for quinpirole in some but not all glutamatergic afferents that make contact on a single cell. The present experiments show that dopaminergic presynaptic modulation of glutamatergic transmission in the neostriatum does not affect all stimulated afferents, suggesting that it is selective towards some of them. This may control the quality and quantity of afferent flow upon neostriatal neurons. 相似文献
25.
The p53 tumor suppressor gene encodes a phosphoprotein which when overexpressed can induce growth arrest at the G1 and G2/M phases of the cell cycle, promote differentiation and apoptosis. This paper demonstrates that p53 can associate with trk tyrosine kinase. Expression of a murine temperature-sensitive (ts) p53 mutant in PC12 cells overexpressing trk (a model system to analyse cellular differentiation and signal transduction induced by NGF) induces morphological changes in the absence of NGF stimulation at 32 degrees C but not at 37 degrees C. In cells differentiated by p53, trk, but not EGFr, was hyperphosphorylated on tyrosine. Furthermore trk was not phosphorylated when expressed in Saos-2 cells (human osteosarcoma cells that lack expression of both endogenous trk and p53) at either temperature. However, transfection of ts p53 into these cells induces trk phosphorylation at 32 degrees C in the absence of NGF stimulation. Association of trk and p53 can be detected in NIH3T3 and PC12 cells co-expressing trk and the ts p53 mutant, in NIH3T3 and PC12 cells transfected with trk alone, and in untransfected PC12 cells, showing that overexpressed and/or endogenous trk associates with endogenous, low levels of p53. These data suggest a novel function for p53 which involves the stimulation of signal transduction pathways (mediating morphological properties of cells), possibly through association with and hyperphosphorylation of trk. 相似文献
26.
SM Kornblau E Estey T Madden HT Tran S Zhao U Consoli V Snell G Sanchez-Williams H Kantarjian M Keating RA Newman M Andreeff 《Canadian Metallurgical Quarterly》1997,15(5):1796-1802
PURPOSE: Expression of the multidrug resistance gene (MDR1) p170 protein is frequent in leukemic blasts from patients with relapsed acute myelogenous leukemia (AML). A phase I study using the nonimmunosuppressive MDR1 blocker SDZ PSC-833 (PSC) in combination with mitoxantrone (MITO) and etoposide (VP) was performed. PATIENTS AND METHODS: Starting doses (LVL0) of MITO (3.25 mg/m2/d on days 1 and 3 to 6) and VP (210 mg/m2/d on days 1 and 3 to 5) were 40% of the maximal-tolerated dose (MTD) from a prior study. A 1.5-mg/kg loading dose of PSC was followed by a 120-hour continuous infusion of 10 mg/kg/d on days 2 to 6. Blood samples for PSC, MITO, and VP pharmacokinetics (PK) were taken on days 1 and 3, and samples for MDR1 expression were taken on day 0. RESULTS: Severe mucositis developed in all patients at LVL0; therefore, MITO and VP doses were reduced to 2.5 and 170 mg/m2 (LVL-1) for the next seven patients, and this dose proved to be MTD. All LVL0 and three LVL-1 patients had transient elevations in the serum bilirubin level to > or = 4 mg/dL. Serum creatinine level increased to greater than 2 mg/dL in one case. There were no other grade 3 or 4 nonhematologic toxicities observed. The peripheral blood was cleared of leukemia in three LVL0 and four LVL-1 patients. The marrow was cleared of leukemic cells in one LVL0 and five LVL-1 patients, and a significant reduction in marrow leukemic infiltrate was observed in eight of 10. No patient achieved complete remission (CR), and all died of progressive disease (n = 8) or infection (n = 2). MDR1 expression was detected by fluorescent-activated cell sorter (FACS) analysis in five of seven cases. An elevated MDR1 mRNA level was detected by quantitative polymerase chain reaction (Q-PCR) in six of eight cases studied. Clearing of leukemia cells from the marrow occurred in four of six MDR1-positive and one of three MDR1-negative patients. Despite the fact that LVL0 doses had to be reduced due to toxicity, coadministration of PSC did not produce a consistent effect on MITO PK; however, it did repeatedly lead to increased levels of VP in the serum. CONCLUSION: We conclude that PSC-MITO-VP is a tolerable regimen with antileukemic activity. Addition of PSC necessitated a 66% reduction in MITO and VP doses from a prior study without PSC. 相似文献
27.
S Della Casa SM Corsello MA Satta CA Rota P Putignano V Vangeli C Colosimo C Anile A Barbarino 《Canadian Metallurgical Quarterly》1997,58(6):503-509
Smoking is associated with insulin resistance, dyslipidaemia and markers of the insulin resistance syndrome. This study investigated adipose tissue lipolysis in situ by subcutaneous microdialysis twice in 10 healthy, male smokers after smoking four cigarettes over 2 h and after the administration of an equal amount of nicotine given as nasal spray (NNS). Glucose and insulin levels, in situ lipolysis and adipose tissue blood flow were studied in the post-absorptive state and after a 75-g oral glucose tolerance test (OGTT). Post-absorptively, acute smoking and NNS increased neither subcutaneous adipose tissue glycerol production nor plasma free fatty acid (FFA) or glycerol levels. After the OGTT, plasma insulin and lactate levels were significantly higher after smoking, whereas FFA levels were higher after NNS. Normal smoking or the administration of a normal dose of NNS caused only minor metabolic changes. Thus, it does not seem likely that increased lipolysis is an important contributor to the dyslipidaemia seen in smokers. 相似文献
28.
FG Abath EM Xavier JD Silva MA Morais Júnior SM Montenegro 《Canadian Metallurgical Quarterly》1997,92(5):637-641
Sm15 and Sm13 are recognized by antibodies from mice protectively vaccinated with tegumental membranes, suggesting a potential role in protective immunity. In order to raise antibodies for immunochemical investigations, the genes for these antigens were expressed in pGEX and pMal vectors so that comparisons could be made among different expression systems and different genes. The fusion proteins corresponding to several parts of the gene for the precursor of Sm15 failed in producing antibodies recognizing the parasite counterpart. On the other hand, antibodies raised against Sm13 MBP-fusion proteins recognized the 13 kDa tegumental protein. Thus the peculiarities of the gene of interest are important and the choice of the expression system must sometimes be decided on an empirical basis. 相似文献
29.
MJ de Gortari BA Freking SM Kappes KA Leymaster AM Crawford RT Stone CW Beattie 《Canadian Metallurgical Quarterly》1997,28(4):274-290
We report the evaluation of 1036 bovine microsatellite primer pairs for their suitability as linkage markers in sheep. Approximately 58% (605/1036) of bovine primer pairs amplified a locus in sheep. Sixty-seven per cent (409/605) of amplified loci were detected as polymorphic. Marker heterozygosity, allele number and range of allele sizes were significantly lower in sheep than cattle sampled in this study. However, median fragment size was similar. These data suggest that high-resolution comparative linkage maps between closely related species can be constructed relatively efficiently. 相似文献
30.
We investigated the effect of repeated cold stress (RCS) on the capsaicin-evoked release of glutamate from the primary afferent fibers of the rat, and compared this with the effect of inoculation of complete Freund's adjuvant (adjuvant inoculation). The release of glutamate was measured using a fluorometric on-line continuous monitoring system in which the immobilized glutamate dehydrogenase column was connected to an in vitro superfusion system. In the presence of 0.3 microM tetrodotoxin, the application of 1 microM capsaicin to spinal dorsal horn slices evoked glutamate release (18.6 +/- 1.2 pmol mg(-1) protein, n = 11). In rats subjected to RCS (RCS rats), the release of glutamate evoked by 1 microM capsaicin was markedly increased to 272% (n = 6, P < 0.05) of the value for the control group, although the basal release was not significantly altered (n = 6, P > 0.05). Adjuvant inoculation produced a significant increase in the basal and capsaicin (1 microM) evoked release of glutamate to 141 and 344% (n = 6, P < 0.05) of the value for the control group, respectively. The present results suggest that the facilitated release of glutamate from capsaicin-sensitive primary afferent terminals in the spinal dorsal horn is, at least in part, involved in the hyperalgesia of RCS rats as well as the complete Freund's adjuvant-induced hyperalgesia. 相似文献