In vitro efficacy and fungicidal activity of voriconazole againstAspergillus andFusarium species

Twenty-nineAspergillus isolates and 25Fusarium isolates underwent in vitro antifungal susceptibility testing by a broth macrodilution procedure adapted from the National Committee for Clinical Laboratory Standards guidelines. The MIC50s of both voriconazole and amphotericin B were 0.5 g/ml and 1 g/ml against species ofAspergillus andFusarium, respectively, while the MIC90s of both agents were 1 and 2 g/ml. Voriconazole was more active in vitro than amphotericin B: the geometric mean MICs of voriconazole and amphotericin B againstAspergillus spp. were 0.36 g/ml and 0.64 g/ml, respectively. Voriconazole also demonstrated fungicidal activity againstAspergillus spp., with 86% (24/29) of isolates exhibiting minimum lethal concentrations of 4 g/ml.     

In vitro activity of voriconazole and other antifungal agents against clinical isolates of <Emphasis Type="Italic">Candida glabrata</Emphasis> and <Emphasis Type="Italic">Candida krusei</Emphasis>

The antifungal susceptibility of 309 Candida glabrata and 63 Candida krusei clinical isolates was tested via the Sensititre YeastOne-3 system (Trek Diagnostic Systems, East Grinstead, UK) to compare the in vitro activity of voriconazole with that of five other antifungal agents (amphotericin B, fluconazole, itraconazole, ketoconazole, and flucytosine). Voriconazole was highly active (MIC90, 0.5 g/ml) against isolates of both species, including those for which the MICs of itraconazole and fluconazole were high (MIC90s of itraconazole, 2 g/ml for C. glabrata and 0.5 g/ml for C. krusei; MIC90s of fluconazole, 32 g/ml for C. glabrata and 64 g/ml for C. krusei). Ketoconazole MIC90 values for both species were identical to those of voriconazole. The MIC90 of amphotericin B was similar for both species (0.125 g/ml for C. glabrata and 0.25 g/ml for C. krusei). As expected, flucytosine was only moderately active against C. krusei isolates (MIC90, 16 g/ml) but was highly active against C. glabrata isolates (MIC90, 0.03 g/ml). Potential cross-resistance within the azole class was noted for some strains of C. glabrata (5.5%) that presented high MIC values for all the azoles tested. In order to consider voriconazole a viable alternative to other triazoles for the treatment of infections caused by Candida species, susceptibility testing of all clinically significant isolates of C. glabrata and C. krusei is recommended because of the potential for azole cross-resistance. The Sensititre YeastOne-3 seems to be a suitable commercial tool for this purpose.     

Activity of trimethoprim/ sulfamethoxazole plus polymyxin B against multiresistantStenotrophomonas maltophilia

The inhibitory activity of eight antibiotics and the inhibitory and bactericidal activities of combinations of trimethoprim/sulfamethoxazole (TMP/SMX) plus three fixed concentrations of polymyxin B (0.01 g/ml, 0.1 g/ml and 0.5 g/ml) against 30 multiresistant strains ofStenotrophomonas maltophilia were tested. Polymyxin B at 0.01 g/ml modified the inhibitory activity of TMP/SMX against only 40% of strains. At 0.1 g/ml and 0.5 g/ml, polymyxin B enhanced the inhibitory activity of TMP/SMX activity against all strains. Polymyxin B enhanced the bactericidal activity of TMP/SMX only at concentrations near the minimum inhibitory concentration of polymyxin B alone.     

Susceptibility to beta-lactam antibiotics in septicemia isolates from twenty-nine European laboratories

In 1984 the European Study Group on Antibiotic Resistance (ESGAR) consecutively collected gram-negative bacilli and staphylococci blood isolates and performed susceptibility testing with 11 antibiotics using the microdilution method. In all 2,578 isolates were collected: 68% gram-negative bacilli and 32% staphylococci. The MICs of ampicillin and cefazoline for the susceptible gram-negative bacilli were 1–8g/ml; of piperacillin0.5–4; of Sch 34343, cefotaxime, moxalactam, ceftazidime and aztreonam0.5–2g/ml; of cefoxitin, cefuroxime and cefamandole0.5–8g/ml. For susceptible staphylococci the MICs of cefazoline and cefuroxime were0.5–1g/ml, and of cefoxitin, moxalactam, ceftazidime and cefotaxime,0.5–32 g/ml. The resistance levels varied between laboratories and countries, being lower in Northern Europe. In clinical protocols on patients with gram-negative septicemia from whom cefazoline-resistant strains were isolated, cefotaxime was the beta-lactam most commonly used (12%). In protocols on patients with staphylococcal septicemia from whom gentamicin-resistant or cefazoline-resistant strains were isolated, the most commonly used beta-lactam was cloxacillin (6%).     

On the regulation of the expressed L-type calcium channel by cAMP-dependent phosphorylation

The Ca²⁺ channel subunits _1C-a and _1C-b were stably expressed in Chinese hamster ovary (CHO) and human embryonic kidney (HEK) 293 cells. The peak Ba²⁺ current (I _Ba) of these cells was not affected significantly by internal dialysis with 0.1 mM cAMP-dependent protein kinase inhibitor peptide (mPKI), 25 M cAMP-dependent protein kinase catalytic subunit (PKA), or a combination of 25 M PKA and 1 M okadaic acid. The activity of the _1C-b channel subunit expressed stably in HEK 293 cells was depressed by 1 M H 89 and was not increased by superfusion with 5 M forskolin plus 20 M isobutylmethylxanthine (IBMX). The _1C-a·₂·₂/ complex was transiently expressed in HEK 293 cells; it was inhibited by internal dialysis of the cells with 1 M H 89, but was not affected by internal dialysis with mPKI, PKA or microcystin. Internal dialysis of cells expressing the _1C-a·₂·₂/ channel with 10 M PKA did not induce facilitation after a 150-ms prepulse to +50 mV. The Ca²⁺ current (I _Ca) of cardiac myocytes increased threefold during internal dialysis with 5 M PKA or 25 M microcystin and during external superfusion with 0.1 M isoproterenol or 5 M forskolin plus 50 M IBMX. These results indicate that the L-type Ca²⁺ channel expressed is not modulated by cAMP-dependent phosphorylation to the same extent as in native cardiac myocytes.     

Effect of natural inhibitors of free radical reactions on adrenalin autoxidation

Water-soluble antioxidants (1 M reduced glutathione, 3 M cysteine, 1 M ascorbate), in the presence of EDTA, inhibit the adrenalin autoxidation reaction at pH 10.2, whereas their oxidized forms, and also -tocopherol (40 M), have no such effect. The inhibiting power of superoxide dismutase is many times greater than that of the substances tested. Adrenochrome formation during free-radical oxidation of adrenalin takes place without the participation of the hydroxyl radical.Department of Biochemistry, Second Moscow Medical Institute. (Presented by Academician of the Academy of Medical Sciences of the USSR V. N. Orekhovich.) Translated from Byulleten' Éksperimental'noi Biologii i Meditsiny, Vol. 81, No. 1, pp. 33–35, January, 1976.     

Provisional quality control parameters and interpretive criteria for testing susceptibility ofStreptococcus pneumoniae andHaemophilus influenzae to quinupristin/dalfopristin (RP59500)

Studies were undertaken to select tentative criteria for susceptibility testing of quinupristin/dalfopristin againstStreptococcus pneumoniae andHaemophilus influenzae. Against 612 isolates ofStreptococcus pneumoniae, MICs of quinupristin/dalfopristin were 1.0 g/ml for all but one strain. With a tentative MIC breakpoint of either 1.0 g/ml or 2.0 g/ml for susceptible, a disk diffusion zone diameter breakpoint of 19 mm embraced all but two of the susceptible pneumococci; 16 mm included all strains. ForHaemophilus influenzae, MICs of quinupristin/dalfopristin clustered near the tentative breakpoints; 91.5% of the MICs were 2.0 to 8.0 g/ml. This precluded satisfactory performance of the disk diffusion test in discriminating between resistant and susceptible isolates unless MIC breakpoints are modified for this species: clinical experience will be needed before that can be justified. Based on data from a multilaboratory study, the following quality control limits are proposed forStreptococcus pneumoniae ATCC 49619 when testing quinupristin/dalfopristin: 0.25 to 1.0 g/ml for broth microdilution tests and 19 to 24 mm for disk diffusion tests. For tests ofHaemophilus influenzae ATCC 29247, MIC limits are 2.0 to 16 g/ml; disk tests were very reproducible but are not yet recommended.     

Comparison of disk diffusion,the E test,and detection ofmecA for determination of methicillin resistance in coagulase-negative staphylococci

The aim of this study was to find a reliable, fast, and simple alternative to the methicillin disk method for determination of methicillin resistance in coagulase-negative staphylococci, since results of this method are often difficult to read due to growth within the zone of inhibition. The sensitivity of 319 strains of coagulase-negative Staphylococci to a 5 g methicillin disk on Mueller-Hinton agar using an incubation period of 48 h was compared with that of 1 (1 g and 5 g oxacillin disks on Mueller-Hinton agar with or without 2% NaCl, using an incubation period of 24 h. The detection ofmecA (MecAgen) by the polymerase chain reaction was used as a standard. Minimum inhibitory concentrations were determined by means of the E test. Of the 225mecA-positive strains, 190, 215, and 193 were resistant to 5 g methicillin, 1 g oxacillin and 5 g oxacillin disks on Mueller-Hinton agar, respectively, and 216, 218, and 223 were resistant on Mueller-Hinton agar with 2% NaCl. Of the 94mecA-negative strains, 89, 93, and 94 were susceptible to 5 g methicillin, 1 g oxacillin, and 5 g oxacillin disks on Mueller-Hinton agar, respectively, and 92, 93, and 94 were susceptible on Mueller-Hinton agar with 2% NaCl. Using breakpoints of 2 g/ml for oxacillin resistance and 8 g/ml for methicillin resistance, the E test yielded sensitivities of 99.6 and 99.1% and specificities of 97.9 and 98.9% after 48 h of incubation. The 5 g oxacillin disk was faster and easier to read than the methicillin disk and correlated better with detection ofmecA than the methicillin disk or the 1 g oxacillin disk.     

Anti-inflammatory and immuno-modulatory effects of novel retinoidlike 2,4,6,8-nonatetraenoic acids (NTA) in adjuvant-induced arthritis

Prophylactic treatment (p.o.) of rats with adjuvant-induced arthritis (AA) with two retinoid-like 2,4,6,8-nonatetraenoic acids (NTA), Ro 23-6457 and Ro 23-2895, significantly reduced hind paw swelling between days 10–23 and the level of plasma fibrinogen (MED 25 moles/kg). When given therapeutically (75 moles/kg between day 21 and 28) either NTA arrested the progression of the disease (MED, 25–75 moles/kg).Unseparated and adherent cell (AC) depleted spleen cells from rats with AA (day 12–15) responded poorly to the T cell mitogen, Con A (2.5 g/ml) and the B cell mitogen, LPS (10 g/ml). The responses were partially restored (30% of normal responses) in AC-depleted (but not unseparated) spleen cells from Ro 23-6457 treated rats (75 and 250 moles/kg/day). These data demonstrate an immunomodulatory effect of Ro 23-6457 in the adjuvant rat which may contribute to its anti-inflammatory activity in AA.     

Phagocytosis and Chemiluminescence Response of Granulocytes to Monodisperse Latex Particles of Varying Sizes and Surface Coats

The response of human granulocytes to polystyrene latex beads of diameter 0.1–7 m was measured by luminol-dependent chemiluminescence. In all instances, the response to beads of 3–7 m was definitely higher than with smaller beads. In protein-free medium, the chemiluminescence response was slow compared to that of opsonized zymosan, and the highest response was only 9% of the response to opsonized zymosan. Scanning electron microscopy showed that granulocytes in suspension bound the particles, occasionally by extending rope-like protrusions. When the beads were coated with albumin, the chemiluminescence diminished to about 1/3 of that seen with uncoated beads; however, preincubating the beads in serum led to a large increase with beads of 1.1 m (to 25% of the maximal response to opsonized zymosan) and 3.19 m (to 42%), but with the smallest beads, no increase was noted. "Priming" of the cells with tumor necrosis factor- caused a further increase with serum-coated beads. When uncoated beads of 1.1 m were tested with "primed" cells, there was an increase of 6 times in the chemiluminescence compared to un-"primed" cells.     

In vitro activity of the aryl-fluoroquinolones A-56619 and A-56620 and evaluation of disk susceptibility tests

The activity of two new quinolones, A-56619 and A-56620, was compared in vitro to that of norfloxacin and ciprofloxacin against 6,699 bacterial isolates in four separate clinical laboratories. The overall percentage of strains susceptible to designated concentrations were as follows: 99.1% for norfloxacin (MIC4.0 g/ml), 96.1% for ciprofloxacin (MIC1.0 g/ml), 96.8% for A-56620 (MIC 2.0 g/ml) and 96.1% for A-56619 (MIC 4.0 g/ml). For disk diffusion susceptibility tests 10 g A-56619 disks are tentatively recommended with interpretive standards of 18mm for susceptibility and 13mm for resistance; 5 g A-56620 disks may be used with tentative standards of 19mm for susceptibility and 14mm for resistance.     

Roles of inositol trisphosphate and protein kinase C in the spontaneous outward current modulated by calcium release in rabbit portal vein

We examined the effects of heparin, guanosine nucleotides, protein kinase C (PKC) modulators, such as phorbol 12,13-dibutylate (PDBu) and H-7 on Ca²⁺-dependent K⁺ currents in smooth muscle cells of the rabbit portal vein using the whole-cell patch-clamp technique, to explore the effects of PKC on the oscillatory outward current (I _oo). Neomycin (30 M), an inhibitor of phospholipase C, and intracellular applications of heparin (10 g/ml) and guanosine 5-O-(2-thiodiphosphate) (GDP[S]; 1 mM) partly but consistently inhibited the generation of I _oo, whereas a higher concentration of heparin (100 g/ml) transiently enhanced then suppressed the generation of I _oo. Inhibition of I _oo generation by heparin was more powerful at the holding potential of + 20 mV than at –20 mV. Inositol 1,4,5-trisphosphate (InsP ₃; 30 M) continuously generated I _oo at holding potentials more positive than –60 mV. Noradrenaline (10 M) and caffeine (3–20 mM) transiently augmented, then reduced the generation of I _oo. Heparin (10 g/ml) completely inhibited responses induced by InsP ₃ and noradrenaline, but not those induced by caffeine. Intracellular application of guanosine 5-triphosphate (GTP; 200 M) or low concentrations of guanosine 5-O-(3-thiotriphosphate) (GTP[S]; 3 M) continuously augmented the generation of I _oo. High concentrations of GTP[S] (10 M) transiently augmented, then inhibited I _oo. Neither GTP[S] nor noradrenaline induced the transient augmentation or the subsequent inhibition of I _oo when applied in the presence of GDP[S] (1 mM), neomycin (30 M) or heparin (10 g/ml). PDBu (0.1 M) reduced the generation of I _oo but failed to produce an outward current following application of caffeine (3–5 mM). This action of PDBu was inhibited by pretreatment with H-7 (20 M). In the presence of H-7, GTP[S] continuously enhanced the generation of I _oo. The suppression of the generation of I _oo during application of noradrenaline (10 M) was reduced by pretreatment with H-7. Thus both InsP₃ and protein kinase C contribute to the generation of I _oo in smooth muscle cells of the rabbit portal vein and heparin is not a specific InsP ₃ antagonist on the InsP ₃-induced Ca²⁺-release channel (PIRC). InsP ₃ opens PIRC and protein kinase C may deplete the stored Ca²⁺ by either inhibiting the reuptake of Ca²⁺ or by enhancement of the releasing actions of InsP ₃.     

Introduction of nonselectible 2μ plasmid into [cir°] cells of the yeast S. cerevisiae by DNA transformation and in vivo site-specific resolution

Summary The 2 DNA plasmid of the yeast Saccharomyces cerevisiae does not confer any known selectable phenotype to the host cell carrying it. Selection of cells transformed with purified 2 DNA therefore cannot be achieved, and the intracellular presence of 2 can only be assessed by molecular analysis of the DNA complement. In addition, 2 alone does not replicate in bacterial hosts, thus rendering its amplification by conventional methods impossible. We have isolated a shuttle plasmid, pBH-2L, generated by in vivo sites-pecific recombination between the endogenous 2 DNA plasmid and pRL, a pBR322 derivative containing the yeast LEU2 gene and one 2 repeat sequence associated with the origin of replication. This new shuttle plasmid has the property, when transformed into yeast, of undergoing site-specific recombinational resolution between its two direct repeat sequences. This releases 2 plasmid and pRL as individual molecules. The latter can undergo progressive mitotic loss during growth in nonselective medium, ultimately leaving leucine auxotrophic transformants that contain only 2 DNA plasmid. This system can be utilized to introduce 2 DNA alone into cells lacking it, thereby providing a novel means to study the biology and the molecular genetics of the plasmid and its potential practical applications as a vector.     

In vitro activity of nonsteroidal antiinflammatory agents,phenotiazines, and antidepressants againstBrucella species

The in vitro antimicrobial activity of streptomycin, rifampicin, tetracycline, seven nonsteroidal antiinflammatory agents (acetyl-salicylic acid, piroxicam, indomethacin, ibuprofen, ketoprofen, sulindac, and diclofenac), and eight phenotiazine derivatives and antidepressant agents (clorpromazine, fluphenazine, amitryptiline, clomipramine, imipramine, maprotiline, sertraline, and diazepam) against 62 strains ofBrucella spp. was tested. Diclofenac was the most active of the anti-inflammatory agents (MIC90 = 16 g/ml). The activity of the phenotiazines and antidepressants was heterogeneous, with MIC90S ranging from 16 g/ml for sertraline and 32 g/ml for fluphenazine and clomipramine to > 512 g/ml for diazepam. When the six most active anti-inflammatory agents and the six most active psychiatric drugs were tested at pH 5 and pH 4, the MICs remained unchanged except for those of fluphenazine; the MIC50 and MIC90 of this agent increased by one dilution.     

Modulation of bradykinin responses in airway smooth muscle by epithelial enzymes

We have studied the effect of epithelium removal on responses of guinea pig trachea to bradykinin (BK). BK (1 nM–10 M) gave a concentration-dependent relaxation when epithelium was present (E+: EC₅₀=10±3 nM). Epithelium removal resulted in a biphasic response to BK with relaxation at low concentrations (E–: EC₅₀=3.0±1.0 nM) and a recontraction to baseline at higher concentrations (EC₅₀=2.0±1 M). Phosphoramidon (10 M), an inhibitor of neutral endopeptidase (NEP), which cleaves BK into inactive peptides, potentiated relaxation (EC₅₀=1.0±0.9 nM and 0.1±0.1 nM in E+ and E respectively) and contraction in trachea with intact epithelium (EC₅₀=0.08±0.03 M). Inhibition of cyclooxygenase by indomethacin (5 M), inhibited relaxation to BK in E+ tracheal segments, resulting in a slight contraction (EC₅₀=1.0 M), whereas a potent contractile response was observed in E–segments (EC₅₀ 1.6 M, maximal contraction >1 g). In the presence of both indomethacin and phosphoramidon BK caused contraction, even in the presence of epithelium (EC₅₀=0.2±0.11 M), and the response in the absence of epithelium was similar to the response observed in trachea with intact epithelium (EC₅₀=0.25±0.1 M). The contractile effect of BK on airway smooth muscle may be inhibited by a protective role of epithelium, due to release of relaxant prostanoids and by degradation by epithelial NEP. In asthma, bronchoconstrictor responses to BK may be partly explained by loss of airway epithelium.     

In vitro activity of the two new 4-quinolones A56619 and A56620 againstNeisseria gonorrhoeae,Chlamydia trachomatis,Mycoplasma hominis,Ureaplasma urealyticum andGardnerella vaginalis

The in vitro activity of tetracycline, ciprofloxacin and two recently developed 1-aryl-fluoro-quinolones, A56610 and A56620, was tested against 65 beta-lactamase-negative and 35 betalactamase-positive Neisseria gonorrhoeae strains, 12 Chlamydia trachomatis,50 Mycoplasma hominis,28 Ureaplasma urealyticum and 50 Gardnerella vaginalis strains. In the case of Chlamydia trachomatis and Mycoplasma hominis both the MIC and the MBC were determined. The MIC90 of ciprofloxacin for Neisseria gonorrhoeae was 0.008 g/mland of A56619 and A56620 0.03 g/ml.No difference was observed between the activity against beta-lactamase-negative and beta-lactamase-positive strains. The MIC90 values of ciprofloxacin and A56620 for Chlamydia trachomatis, Mycoplasma hominis and Ureaplasma urealyticum were identical, the values being 2 g/ml, 1g/mland 4 g/mlrespectively. The MIC90 of A56619 for Chlamydia trachomatis and Ureaplasma urealyticum was 0.5 g/mland 1 g/mlrespectively. The MBC90 values of the three quinolones for Chlamydia trachomatis and Mycoplasma hominis were 2 g/ml.The activity of the quinolones against Gardnerella vaginalis was rather low, the MIC90 being 4 g/ml.It is concluded that A56619 and A56620 might be useful for single-dose therapy of gonococcal infections.     

In vitro activity of LY146032 (Daptomycin), a new peptolide

The in vitro activity of LY146032, a new peptolide antibiotic, was compared with those of vancomycin, teicoplanin, imipenem, amoxicillin and erythromycin. LY146032 inhibited 90% ofStaphylococcus aureus andStaphylococcus epidermidis, including methicillin-resistant isolates at 1g/ml. Its activity was comparableto those of vancomycin and teicoplanin. MIC90s for the beta-hemolytic streptococci varied from 0.25 g/ml for group B streptococci to 4g/ml for some group C and F streptococci. MICs forStreptococcus faecalis were in the range of 0.5 to 8 g/ml,and the MIC90 4 g/ml,compared to 4 g/ml for vancomycin and 1g/ml for teicoplanin. For some viridans streptococci the MICs were 4g/ml, whereasStreptococcus pneumoniae were inhibited by 0. 5g/ml.Corynebacterium JK species were inhibited by 0. 5g/ml, similar to vancomycin, andListeria monocytogenes by 4g/ml.Neisseria species,Haemophilus species and enteric species were not inhibited. Most MBCs were within two-fold of the respective MICs. After 14 days passage in subinhibitory concentrations of LY146032,Staphylococcus aureus, Staphylococcus epidermidis andStreptococcus faecalis showed minimal increase in MICs. The activity of LY 146032 was increased by adding Ca²⁺ and was reduced in an anaerobic environment. Overall, LY146032 is an extremely interesting new agent that inhibits gram-positive species.     

Effect of auranofin,a new antiarthritic agent,on immune complex-induced release of lysosomal enzymes from human leukocytes

Auranofin, an oral chrysotherapeutic agent effective in the treatment of rheumatoid arthritis (RA), was found to be a potent, noncytotoxic inhibitor of IgG-RF immune complex-induced lysosomal enzyme release (LER) from human leukocytes. At a concentration of 1 g Au/ml (5M), auranofin produced a marked reduction in-glucuronidase (100%), acid phosphatase (88%), and lysozyme (72%) release. In contrast, gold sodium thiosulfate (GST, an injectable gold compound) had no inhibitory activity on LER at equivalent gold concentrations (i.e., 1g Au/ml) and only modest activity (< 36% inhibition) at concentrations as high as 40g Au/ml. The 50% inhibitory dose (ID₅₀) of auranofin on LER was calculated to be 3–4M (0.6–0.8g Au/ml). Blood gold levels in auranofin-treated RA patients were found to be within the range required for in vitro inhibition of LER, and correlated with decreases in IgG, RF titers, and IgG-RF immune-complex formation in vitro. These results suggest that the therapeutic action of auranofin may be caused, at least in part, by inhibition of LER and/or decreases in immune-complex formation.SK&F D-39162 (2,3,4,6-tetra-O-acetyl-1-thio--D-glucopyranosato-S) (triethylphosphine) Gold.     

Effects of inhibitors and ion substitutions on oscillations of cell membrane potential in cells expressing the RAS oncogene

Previous studies revealed that in NIH fibroblasts expressing the ras oncogene but not in other NIH fibroblasts, bradykinin leads to sustained, calcium dependent oscillations of cell membrane potential by repetitive activation of calcium-sensitive K⁺ channels. The present study has been performed to test for ion and inhibitor sensitivity of these oscillations. Both, Lys-bradykinin (kallidin) and bradykinin, but not any shorter peptide tested, maintained the oscillations. The oscillations are abolished in the presence of the K⁺ channel blocker barium (10 nmol/l). The amplitude but not the frequency of the oscillations is dependent on the extracellular potassium concentration. The oscillations are not dependent on the presence of extracellular sodium, bicarbonate or chloride. The oscillations are abolished in the absence of extracellular calcium and their frequency is significantly decreased at reduced extracellular calcium (to 0.2 mmol/l). The oscillations are not inhibited by acute administration of ouabain (0.1 mmol/l), by dimethylamiloride (100 mol/l), furosemide (1 mmol/l) and hydrochlorothiazide (100 mol/l), by cobalt (100 mol/l), zinc (100 mol/l), gadolinium (100 mol/l), verapamil (10 mol/l) and diltiazem (10 mol/l), but are abolished in the presence of 100 mol/l lanthanum, 1 mmol/l cadmium, 10 mol/l nifedipine, 25 mol/l SK & F 96365 and 200 mol/l TMB-8. Stimulation of calcium entry by 10 mol/l ionomycin is frequently followed by oscillations of cell membrane potential even in the absence of bradykinin. In conclusion, in cells expressing the ras oncogene bradykinin leads to sustained activation of calcium channels at the cell membrane, which cause oscillations of the cell membrane potential by triggering intracellular calcium release.     

Association of Phenotypic Changes in B Cell Lymphocytes and Plasma Viral Load in Human Immunodeficiency Virus-Infected Patients

Many B cell abnormalities have been reported in human immunodeficiency virus (HIV)-infected patients, including changes in the expression of , , and CD22 molecules on the cell surface. Phenotypic changes in these markers on B cells isolated from HIV-seropositive patients with high or low levels of plasma viremia were measured. The phenotypic changes in B cells isolated from such patients were compared with the markers on B cells isolated from HIV-seronegative individuals using three-color flow cytometry. HIV patients showed a reduction in the proportion of mature B cells isolated from peripheral blood mononuclear cells compared with B cells isolated from HIV-seronegative individuals. An increase in the proportion of B cells expressing both and molecules on the cell surface was also seen in association with high-HIV plasma viremia. A low plasma viral load was accompanied by a reduction in the proportion of B cells expressing both and molecules to a level comparable to those seen in HIV-seronegative individuals. HIV-seropositive individuals demonstrated an increase in the proportion of committed B cells, as indicated by an increase in the proportion of B cells expressing molecules. This observation may explain the poor humoral response of HIV seropositive patients to neo-antigens. Our results demonstrate that phenotypic changes indicative of in vivo B cell activation and an increase in immature cells are associated with HIV infection, particularly with a high plasma viral load. Phenotypic changes in B cell markers may correlate with functional deficits of B cells.