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PubMed1989

Extracellular adenosine triphosphate increases cation permeability of chronic lymphocytic leukemic lymphocytes.

Extracellular adenosine triphosphate (ATP) is known to reversibly increase the cation permeability of a variety of freshly isolated and cultured cell types. In this study the effects of extracellular ATP were studied using peripheral blood lymphocytes (PBL) isolated from both normal subjects and from patients with chronic lymphocytic leukemia (CLL). Changes in the permeability to Na+, Rb+, and Li+ ions were measured using conventional isotope and flame photometry techniques. In addition, changes in cytosolic (Ca2+) were fluorimetrically monitored to assess possible changes in net Ca2+ influx. ATP produced a 12-fold increase in 22Na+ influx into CLL cells but only a 3.5-fold increase in this flux in PBL cells. A maximal response was produced by 0.1 mmol/L ATP in the absence of Mg2+, while a twofold molar excess of Mg2+ over ATP abolished the response. ATP had no effect on the passive (ouabain-insensitive) 86Rb+ influx into PBL cells but stimulated this flux by fivefold in the CLL cells. Li+ influx into CLL cells was also stimulated threefold by ATP. Under these same conditions ATP also produced a net increase in total cell Na and a decrease in total cell K in the CLL cells. Exclusion of two normally impermeable dyes, trypan blue and ethidium bromide, was not altered in the ATP-treated CLL cells. Finally, extracellular ATP (3 mmol/L) produced no significant change in the cytosolic (Ca2+) of normal, monocyte-depleted populations of PBL. Conversely, this same concentration of ATP produced a very rapid (complete within 30 seconds) and a significant (an average threefold peak change) increase in the cytosolic (Ca2+) of cell preparations derived from five out of nine CLL patients. In these latter CLL cells, the ATP-induced elevation in cytosolic (Ca2+) appeared to be due to a net increase in Ca2+ influx, since no elevations were observed when the extracellular (Ca2+) was reduced to less than 0.1 mmol/L. These actions of ATP were specific in that equimolar concentrations of other nucleotides were without effect. These data indicate that treatment of CLL lymphocytes with extracellular ATP4 produces large increases in cation permeability. In contrast, there is less or no ATP-induced permeabilization of normal PBL.

Adenosine Triphosphate↗

PubMed1989

[Action of trichlormethiazide and amiloride on cellular Na+, K+ and Mg+ concentrations].

While diuretic-induced changes of plasma electrolyte concentrations have often been described, comparatively few data exist on intracellular electrolyte concentrations under diuretic treatment. Therefore, we studied the effect on intracellular Mg2+, Na+ and K+ concentrations of a thiazide diuretic (trichlormethiazide 4 mg/d) in red blood cells of 14 patients with mild essential hypertension, and of a combination of a thiazide diuretic and a potassium-sparing diuretic (trichlormethiazide and amiloride 2 mg/d each) in red blood cells of 11 patients with mild essential hypertension. Measurements were performed by atomic absorption spectroscopy and flame photometry before starting treatment and after 4 and 8-12 weeks' diuretic treatment. There was no significant change in intracellular Na+ and K+ concentrations under either form of diuretic treatment. Intracellular Mg2+ concentrations decreased significantly under thiazide therapy, whereas there was a significant increase in intracellular Mg+ concentrations under thiazide and potassium-sparing combined diuretic therapy. The results show that the combination of a thiazide diuretic and a potassium-sparing diuretic is a useful means to avoid intracellular Mg2+ loss.

Amiloride↗

PubMedDate not supplied

[A comparative evaluation of methods for the isolation of khlozepid from human cadaveric liver].

Comparative evaluation of two isolation methods in fatal chlozepid poisoning was performed using expert material. Isolation by neutral acetone from visceral tissues showed that chlozepid and its metabolites are extracted in amounts 6-13 times greater (in terms of chlozepid) than using A. A. Vasil'eva's method. Quantitative determination was performed by photometry according to the reaction of azo dye formation after hydrochloric acid hydrolysis of the extracts.

Adult↗

PubMedDate not supplied

[A new in vitro test of endothelial viability. Vital staining using Janus green with spectrophotometric analysis of the color extract].

A new in vitro technique has been developed to measure by simple photometry the amount of Janus Green extracted with absolute alcohol from isolated corneas after vital staining. The amount of stain corresponds to the percentage of damaged endothelial cells and is a reliable parameter of endothelial viability. More rapid and easier to perform than the usual cell counting of devitalized cells this technique may be used to study the endothelial damage induced e.g. by experimental surgery, intraocular solutions, IOL material, cornea storage solutions.

Animals↗

PubMed1989

Clinical occurrence and virulence testing of coagulase-negative staphylococci.

A total of 650 coagulase-negative staphylococcal strains of miscellaneous clinical origin were isolated and identified during a 7-year-period. In all kinds of samples Staphylococcus epidermidis was the most frequently found species. In wounds the incidence of haemolytic species (S. cohnii, S. epidermidis, S. haemolyticus, S. simulans, S. warneri) was 79.3%. Among strains derived from male genitals (urethra, semen, etc.) S. epidermidis and S. simulans predominated. Only 8.2% of the strains were found not to harbour resistance determinants, while 63% proved to be multiply resistant. Only the incidence of resistance to oxacillin and first and second generation cephalosporins was low (12.8, 14.9 and 4.5%, respectively). Vancomycin was the only drug effective against all strains. Extracellular slime was produced by 42.2% of the strains. The frequency of slime-positivity among isolates causing clinical infection was twice as high as among contaminations. Slime production seems to be a stable characteristic of the isolates. The comparison of two methods used for detection of slime factor showed that incubation in test tube in Tryptone Soya Broth, or its application to microtiter plates evaluated by photometry are of equal diagnostic value.

Drug Resistance, Microbial↗

PubMed1989

[Cardiospermum--a new phytogenic drug in dermatology?].

In a double blind study on persons with healthy skin, we tested the efficacy of cardiospermum ointment against the ointment base only. There were no significant differences between placebo and active substance regarding clinical evaluation, light reflection photometry, and transepidermal water loss. By means of doppler laser flowmetry, we found a decrease in corial blood supply after 16 hours' application of cardiospermum, which suggests a possible antiphlogistic effect.

Adult↗

PubMed1989

A microfluorometric viability assay for isolated human and rat islets of Langerhans.

A microfluorometric viability assay for isolated human and rat islets of Langerhans has been developed using the fluorochromes fluorescein diacetate and propidium iodide. Fluoroscein diacetate causes live cells to fluoresce green under blue light excitation (490 nm); propidium iodide causes dead cells to fluoresce red under green light excitation (545 nm). The fluorescence intensity from the live and dead cells within a single islet was selectively measured by photometry using 520 nm and 610 nm barrier filters with blue and green light excitation respectively. All measurements were corrected for background fluorescence. It was necessary to incubate single islets with the fluorochrome mixture for 105 min in order to achieve maximum fluorescence intensity. It was found that when 50 microliters of a fluorochrome mixture containing 0.67 mumol/l fluorescein diacetate and 4.0 mumol/l propidium iodide was incubated with a single islet and the fluorescence from live (blue light excitation) and dead (green light excitation) cells measured, then the proportion of dead cells within the islet was equal to the (propidium iodide fluorescence)--(0.04 x fluorescein fluorescence) divided by the sum of the (fluorescein fluorescence) and (propidium iodide fluorescence--(0.04 x fluorescein fluorescence]. The proportion of dead cells within single human or rat islets measured by microfluorometry was found to correlate highly significantly (r = 0.99, p less than 0.001) with the proportion of dead cells measured by dissociating the same islet into a single cell suspension and counting the actual proportion of dead cells. This assay therefore provides a rapid, accurate and objective measurement of the proportion of dead cells within isolated human and rat islets.

Animals↗

PubMed1989

[In vitro changes in the lens epithelium and corneal endothelium caused by the cytostatic drug daunomycin].

We evaluated the effects of the antiproliferative drug daunomycin on the viability and proliferation of cultured porcine lens epithelial cells. After a single short-term application, daunomycin penetrates the lens epithelial cells within 5 min. At a concentration of 2.5-7.5 mg/l, daunomycin has a low level of acute cytotoxicity and significantly suppresses the mitotic activity of the epithelial cells in culture. The daunomycin-induced endothelial lesions in freshly excised pig corneas were quantified with the Janus green photometry technique (n = 30) and studied with scanning electron microscopy (n = 9) in comparison to Ringer solution, which served as the control (n = 39). Whether the endothelial lesions were exposed to daunomycin or Ringer solution for 10 or 30 min seemed to make no significant difference. In addition, daunomycin did not seem to induce any morphological changes in the endothelial surface. These in vitro results confirm our clinical experience that daunomycin is apparently not harmful to the adult human corneal endothelium. Our results indicate that short-term endocapsular application of daunomycin during extracapsular cataract extraction, with or without IOL implantation, may prevent and/or delay the formation of capsular opacification.

Animals↗

PubMed1987

Simultaneous determination of magnesium and potassium in lymphocytes, erythrocytes and thrombocytes.

A standardized method for the determination of magnesium and potassium in the lymphocytes, erythrocytes, and thrombocytes of normal subjects is described. The procedure consists of four steps: 1) the removal of platelets from the remaining cells and heparin plasma, 2) the isolation of platelet-poor lymphocytes by means of density-gradient centrifugation of the buffy-coat, 3) the purification of blood cells by repeated washings, and 4) the determination in cell lysates of magnesium by atomic absorption spectrometry and of potassium by flame photometry. In order to get reproducible lymphocyte magnesium values, it was necessary to obtain a mean lymphocyte recovery of 60% with a purity of 97% (nucleated cells), this yield containing less than 5 thrombocytes per lymphocyte and less than 4 erythrocytes per 100 lymphocytes. Under these conditions, the normal content of magnesium in lymphocytes was 3.5-5.7 fmoll/cell (n = 18), that of potassium was 38.8-63.5 fmol/cell. The precision (CV) of the whole procedure measured with 10 identical samples from each of 5 persons was 5.7% for lymphocyte magnesium and 7.8% for potassium, respectively. The concentration of magnesium in erythrocytes was 1.99-2.43 mmol/l and that of potassium was 85.8-102.5 mmol/l. The precision was 3.6% for magnesium and 2.2% for potassium. The content of magnesium in thrombocytes was found to be 0.07-0.12 fmol/cell.

Adult↗

PubMed1985

Intra-erythrocytic sodium content in normotensive offspring of normotensive and hypertensive subjects: an epidemiological study.

An increase in intra-erythrocytic sodium (IENa) content has been proposed as a genetic marker of essential hypertension. Intra-erythrocytic sodium was studied using hypotonic lysis and flame photometry after four washings with isotonic MgCl2 in 240 normotensive subjects (aged 10-45 years) on a free diet with (F+, 121 patients) or without (F-, 119 patients) hypertensive parents, recruited from a random sample of the general population. Systolic blood pressure was significantly higher in males F+ than in males F- (130 +/- 2 versus 125 +/- 2 mmHg, mean +/- s.e.m., P < 0.05), while IENa did not differ. In contrast, intra-erythrocytic potassium content (IEK) was significantly lower and red cell sodium potassium (Na:K) ratio significantly higher in F+ than F-. This might reflect decreased NaK pump activity, or increased membrane permeability to cations which causes increased K leakage. No differences in blood pressure, IENa or IEK showed in female F+ versus F-. It is concluded that IENa is not a genetic marker of hypertension, and that it is probably influenced by exogenous factors. Being associated with differences in blood pressure, the abnormalities of IEK and Na:K ratio might be pathogenetically linked to an early increase in blood pressure.

Adolescent↗

PubMed1987

[The effect of magnesium on the formation of toxic shock syndrome toxin 1 (TSST-1) by Staphylococcus aureus].

The influence of magnesium on the production of toxic shock syndrome toxin-1 (TSST-1) by Staphylococcus aureus was examined. As media we used: Standard-I-Nutrient Broth, Todd Hewitt Broth, Mueller-Hinton Broth, Isosensitest Broth and a chemically defined liquid medium. The magnesium content of these media was determined using flame photometry and was subsequently changed using magnesium sulfate to the magnesium concentrations as shown in table 1. In each of these media the TSST-1 positive S. aureus strains MN8 and T 40 were grown at 37 degrees C, 18 h, vigorously shaken. Then the colony forming units (cfu) were determined. Toxin assays were performed by immunodiffusion after concentrating the culture fluids 100-fold using ethanol precipitation. Concentrations of toxin per milliliter were determined by comparison with standard toxin preparations using hyperimmune-TSST-1 antisera as described. The cfu and the amount of TSST-1 produced are shown in table 1. No link could be demonstrated between TSST-1 production and magnesium concentration of the media used. Our results are in agreement with the ones published by Schlievert and disagree with the results from Mills.

Bacterial Toxins↗

PubMed1988

[Functional and structural changes in the surface of human erythrocytes after UV irradiation with rays of various wavelengths. IV. Changes in the physicochemical properties of autotransfused UV-irradiated blood].

Properties of erythrocyte surface were investigated for patients with ischemic heart disease in the course of treatment with the UV-irradiated blood autotransfusion (UVIBA). Application of methods of light-scattering, photometry and cytochemistry revealed rapid and significant changes in deformability and aggregation properties of the erythrocytes immediately following each UVIBA procedure, which was accompanied by considerable blood viscosity decrease.

Absorption↗

PubMed1988

Ion-selective potentiometry in clinical chemistry. A review.

Ion-selective potentiometry is used more and more in clinical medicine for the determination of electrolytes in various body fluids. With regard to K+, Na+, Ca2+ and Cl- this technique has almost completely displaced flame photometry, atomic absorption spectrophotometry and coulometry. Moreover, reliable automated devices have facilitated routine analyses. Until now there are 6 different types of ion-selective sensors: glass membrane, solid phase, fluid membrane, carrier, gas-sensitive, and enzyme electrodes with immobilized enzymes. The latter are particularly used for in vivo monitoring, especially for continuous blood glucose monitoring. The essential fields of application in the clinical laboratory are the determinations of the cations H+, K+, Na+, Ca2+, Mg2+ and NH3+, and the anions F-, I-, Br-, Cl- and HCO3-. Despite the wide-spread application of ion-selective potentiometry a number of disturbing factors have to be taken into account by the user as well as by the manufacturer in order to get satisfactory results. For instance, there are differences between direct and indirect potentiometry. Moreover, the activities measured cannot be extrapolated readily to the desired concentrations. A careful and accurate calibration, a suitable sample preparation and an adjustment of the measuring conditions to the characteristics of the specimen and the matrix of the sample is necessary before each measurement. Therefore, a consequent internal and external quality control is necessary to achieve an optimal quality of these methods determining vital parameters in medicine. Thus, the technique of ion-selective potentiometry represents an important milestone in clinical chemistry. Moreover, being a very rapid procedure it is indispensable to clinical diagnostics.

Body Fluids↗

PubMedDate not supplied

[Increasing the accuracy of the stereoroentgenophotometric method of studying mineral saturation of the bones].

Method of roentgenodensitometry of voluminous objects, developed in Moscow Research Institute of Tuberculosis, is described. The method provides for stereoroentgenography of the studied subject with a special screw wedge, served as a standard. The analysis of films obtained in stereocomparator permits to define an excess of the studied area of bone tissue and determine a corresponding area of the standard for photometry. Stereoroentgenophotometry with the use of a standardized screw-shaped wedge permits a 10-20% increase in the accuracy of determination of bone structure mineral saturation that contributes to improvement of diagnosis in various skeletal diseases.

Absorptiometry, Photon↗

PubMed1986

[Stool electrolyte concentration in acute infantile diarrhea in France].

Sodium, potassium and chloride stool content was studied in 107 children aged 1 to 32 months (11 +/- 8 months) presenting with acute diarrhea related to a rotavirus infection (34 cases), to an invasive pathogen (Salmonella or Shigella 14 cases, E. coli 4 cases), or of non-identified etiology (65 cases). The therapeutic protocol was the same in all cases: rehydration for the first 24 hours, progressive realimentation from the second or third day, no drugs being given. An average of 4 stools were analysed for each child (range 2-14), the fecal samples being collected over 2 to 4 consecutive days. Na+ and K+ (n = 366) were assayed by flame photometry and chloride (n = 88) by continuous colorimetry. In the stool samples taken as a whole, without taking into account etiology or day of sampling, the electrolyte concentration (mean +/- SD) was 42 +/- 20 mmol/l for Na+ (range 4-166), 51 +/- 24 mmol/l for K+ (range 5-195), and 24 +/- 11 mmol/l for Cl- (range 4-93). No significant variation of these values was observed according to etiology, duration, severity of the diarrheal syndrome, oral rehydration or nutrition.

Acute Disease↗

PubMedDate not supplied

Computer-assisted photometric microplate analysis.

The main algorithm of computer-assisted absorption and emission photometry of samples on a microplate is presented. The software can be used for the enzyme immunoassay (EIA) and other virological tests. The performances of an SPF-500 (Aminco) spectrofluorometer/Felix M18 microcomputer system are discussed on the ground of some results obtained by using the implemented programs.

Algorithms↗

PubMed1987

[Dry chemistry of reagent strips].

Quantitative analysis with dry reagent strips for the determination of clinical chemistry parameters and drug monitoring in serum, plasma and whole blood is expanding in small laboratories and in office laboratories. The systems predominantly used are Ektachem DT 60, the Seralyzer and the Reflotron. - The methods offered comprise substrates, enzymes, electrolytes, CO2 and drugs. The spectrum of feasible tests varies with the manufacturer. On the whole all the tests commonly used in small laboratories are covered. The instruments used for quantitative dry reagent strip chemistry are calibrated by the manufacturer. Calibration data are either inputted to the instrument with each method or already programmed in the apparatus. Thus calibration is outside the control of the user, a change of philosophy which does not correspond to present concepts of quality control in clinical chemistry. The analytical precision is satisfactory for all three instruments for the common parameters measured. A comparison between "dry-chemistry" determinations and the classical aqueous analyses show good accuracy as long as patient samples are used. This is, however, not true for all reference materials, presumably due to matrix effects. Our own evaluation of the Seralyzer showed a good correlation of potassium analysis compared with flame photometry; this was also true of gamma-glutamyltransferase using the Reflotron with patient samples. Internal quality control can only be performed with materials assessed by the instrument manufacturers, and the results will often differ for these materials when assayed with conventional methods or with other "dry-chemistry" methods. External quality control also has its limitations due to the fact that no sera are available which could be used universally.(ABSTRACT TRUNCATED AT 250 WORDS)

Chemistry, Clinical↗

PubMed1987

A method for simultaneously estimating plasma, erythrocyte, and leukocyte sodium, potassium, and magnesium: reference values and considerations from biological variation data.

We describe a method for measuring plasma, erythrocyte, and leukocyte sodium (Na+), potassium (K+), and magnesium (Mg2+) concentrations in 10-mL blood specimens. After separating cells with Ficoll-Hypaque and washing with isotonic choline chloride, erythrocytes and leukocytes are counted, so that results can be expressed as amount of substance per cell and also to monitor cell integrity and possible contamination. Plasma and cell lysates were analyzed (CV less than or equal to 7.0%) with flame photometry and atomic absorption spectrometry. Reference intervals for an elderly population with values for plasma electrolytes within reference intervals are similar to those for younger healthy subjects. From data on biological variation, reference values for erythrocyte cations are not of much use, and analytical goals for precision are not met, but the results might be useful for monitoring disease progression in individual patients. In contrast, reference values for leukocyte cations are theoretically of use and goals are achieved, but large changes are required before consecutive results can confidently be said to be significantly different.

Aged↗