Cloning and sequence of the cDNA coding for rat type II Fc gamma receptor of mast cells.

A clone encoding the rat type II Fc gamma receptor was isolated from a cDNA library of the rat mucosal mast cell line RBL-2H3 and its sequence determined. The predicted amino acid sequence is highly homologous with the mouse type II as well as rat type III Fc gamma receptors. The site of alternative splicing which generates the mouse Fc gamma Rb2 isoform is completely conserved. Hence we consider the new sequence to encode a rat Fc gamma RII b2 isoform. In further analogy to the mouse receptor, no consensus motif known to be involved in accessory chain association was observed in the transmembranal domain. The importance of the identification of this receptor for investigation of the immunological stimulation of mast cells is discussed.

Sweeping of neutral analytes in electrokinetic chromatography with high-salt-containing matrixes

The concept of sweeping neutral analytes using a high-conductivity matrix or under a reduced electric field in micellar electrokinetic chromatography (MEKC) using anionic micelles and in the presence of electroosmotic flow is presented. Three important processes are identified. First, stacking of the micelles at the cathodic interface between the sample solution (S) and background solution (BGS) zones is identified. This is then followed by the sweeping of analyte molecules by the stacked micelles that enter the S zone. Finally, the destacking of the stacked micelles at the anodic interface between the S and BGS zones occurs. The stacking of the micelles improves the focusing effect of sweeping by a factor approximately equal to the ratio of conductivities between the S and BGS zones (ratio = enhancement factor = gamma'). However, the destacking of the stacked micelles broadens the swept zones by a factor approximately equal to 1/gamma'. In effect, the focusing effect of sweeping using a matrix with equal or higher conductivity compared to the BGS will be roughly the same. The micelle stacking and destacking mechanisms are verified experimentally. This paper also provides comments on the mechanism of neutral analyte focusing under similar conditions proposed by another group.

Ranitidine or dobutamine alone or combined has no effect on gastric intramucosal-arterial PCO(2) difference after cardiac surgery.

OBJECTIVE: To test the hypothesis that ranitidine, either alone or in combination with dobutamine, modifies the gastric intramucosal-arterial PCO(2) difference. DESIGN: Full factorial design (double-blinded for ranitidine). SETTING: Intensive Care Unit of a university hospital. PATIENTS: Sixty-four haemodynamically stable coronary artery bypass surgery patients. INTERVENTIONS: Ranitidine (150 mg preoperatively per os and 50 mg intravenously postoperatively) and dobutamine (4 micro g. kg(-1). min(-1) for 3 h postoperatively) were administered in four randomised groups of patients: preoperative and postoperative ranitidine, either alone (n = 15) or in combination with dobutamine (n = 17), dobutamine alone (n = 15) or neither ranitidine nor dobutamine (n = 17). MEASUREMENTS AND RESULTS: Gastric intramucosal-arterial PCO(2) difference was measured during the first 5 postoperative hours. No differences in the postoperative pattern of gastric intramucosal-arterial PCO(2) difference were found among the groups. CONCLUSIONS: Ranitidine and dobutamine have no effect on the gastric tonometry results on intramucosal-arterial PCO(2) difference after uncomplicated cardiac surgery. Hence, the routine use of H(2)-antagonists for gastrointestinal tonometry is not warranted. Our results must be limited to results obtained by tonometry; they do not allow any conclusions on the effects of these drugs on splanchnic blood flow or its distribution.

Calculation of stability constants for the chiral selector-enantiomer interactions from electrophoretic mobilities.

Interactions of chiral selectors with enantiomers or with other chiral analytes, underlying their electrophoretic separations, are particular cases of interactions of dissolved species. The interaction model that describes these reversible, fast interactions is simple. Equations describing the model are also simple and applying equations for the calculation of respective equilibrium constants to experimental data is easy using computers. Obtaining the experimental mobility data, necessary for the calculation, is the critical step on the way to stability constants quantitating the strengths of interactions of chiral selectors with enantiomers and other analytes. These data are decisive for both the accuracy and precision of the calculated constants. The meaning and applicability of the particular constant depend on the type of the constant. The common method for the determination of stability constants from electrophoretic migration data is reasonable for low and medium stability constants. For stronger complexing, characterised by stability constants of the order of 10(4) l/mol, typical of affinity chiral selectors, the method becomes unreliable. For strong complexing giving constants of the order of 10(5) or higher of or higher the method is not applicable in its commonly used form.

Injection bias of DNA fragments in capillary electrophoresis with sieving.

The relative amounts of DNA fragments in a mixture injected into the capillary by electromigration or hydrodynamically by pressure were compared. Even if the electrophoretic mobilities of DNA fragments with different sizes are the same in a free solution in the sample vial, the size bias is brought about by the different mobilities in a sieving medium and by the electroosmosis. The experiments were performed in capillaries filled with a solution of liquified agarose, a replaceable sieving medium. The experimental results were compared with a theoretical model.

Capillary zone electrophoresis in phosphate buffer--known or unknown?

It has been shown recently that the analysis records in capillary electrophoresis may involve regions with extremely strong electromigration dispersion of peaks. Such a fundamental effect is due to the existence of more centers of symmetry in a given electrolyte system. This paper shows that even such a simple and frequently used electrolyte system as phosphate buffer may exhibit more than one center of symmetry. By using the peak shape diagram approach we have revealed that neutral and alkaline phosphate buffers have two centers of symmetry and one center of extreme dispersion. Model experiments confirmed this new important discovery.

Highly alkaline electrolyte for single-stranded DNA separations by electrophoresis in bare silica capillaries.

A new, highly denaturing electrolyte system based on a solution containing 0.01 M NaOH, 0.0015 M Na2B4O5(OH)4 and a replaceable polymer sieving medium was designed for the separation of single-stranded DNA fragments in bare fused-silica capillaries. Extreme denaturing power, together with the optimized composition of the electrolyte, allows for a separation efficiency as high as 2,300,000 height equivalents to a theoretical plate per meter. Sample denaturation in alkaline solutions provides single-stranded DNA fragments without any intra- or intermolecular interactions at room temperature. Their electrophoretic mobilities were found to be twice those of fragments denatured by dimethylformamide or HCl. This can be interpreted in terms of an increased effective charge on the DNA molecules. The surprisingly weak electroosmosis (6 x 10(-10) m2 V-1 s-1) of polymer solutions at pH 12 or higher is considered to be the result of the dissolution of the silica capillary wall. A highly viscous thin layer of dissolved silica probably causes a shift of the slipping plane further away from the wall to the lower value of the zeta potential. Applications of the electrolyte in clinical diagnostics demonstrate its remarkable properties.

Analysis of orotic acid in human urine by on-line combination of capillary isotachophoresis and zone electrophoresis.

The techniques of the on-line combination of capillary isotachophoresis with zone electrophoresis in two coupled capillaries (ITP-CZE) and a single capillary zone electrophoresis (CZE) were used for the sensitive determination of orotic acid (OA) in human urine. The simple CZE system was successfully applied for fast and reliable analyses of urine of healthy adult volunteers (the detection limit 1.7.10(-6) M OA, the total time of analysis 6 min). However, this method failed in analyses of OA in urine of ill children due to more complex matrix of the samples. Here, the ITP preconcentration and preseparation step coupled on-line with CZE proved to serve well with an electrolyte system developed and optimized for this purpose. The maximum selectivity and resolution of OA from other sample constituents in ITP-CZE was achieved by use of an electrolyte system of very low pH 2.15 both for ITP and CZE stage. The sensitivity of detection and simplicity of OA identification were enhanced by use of an external UV scanning detector. High sensitivity of ITP-CZE combination (limit of detection 3.10(-7) M OA), low sample consumption (1 microliter), good reproducibility of migration times (inter-day RSD < 1.86%) and acceptable reproducibility of the determination of OA in urine samples (average RSD = 7.27%) make this technique suitable for routine determination of trace concentration of OA especially in urine of ill children under various pathological conditions and medication.

Why robust background electrolytes containing multivalent ionic species can fail in capillary zone electrophoresis.

In this paper it is demonstrated that system peaks are induced by multivalent weak ionic species in background electrolytes. Their existence is derived from SystCharts and from Peak Shape Diagrams and the theory is confirmed experimentally. If analytes are present in a sample, with mobilities approximately equal to the mobility of a system peak, they interact, resulting in a strong increase of electromigration dispersion. This leads to strong peak broadening, peak deformation and a loss of resolution. Typical background electrolytes containing multivalent ionic species, e.g. phosphate and phthalate buffers, often reported to be robust electrolytes, are therefore not always universally applicable and can fail for the application of specific analytes. This paper reports a systematic study of the above phenomena and shows both theoretical and experimental results for background electrolytes containing phosphoric acid and phthalic acid.

Determination of dissociation constants of cytokinins by capillary zone electrophoresis.

A method for the pKa determination, based on mobility data measured by capillary zone electrophoresis, was applied to cytokinins and their analogs. The combination of charged mobility standards with an uncharged electroosmosis marker, injected in the uncoated capillary simultaneously with the measured substances, allows one to minimize the number of runs, reduce their duration and, in addition, to inform on the run-to-run stability of electroosmosis and on contingent side-effects. pKa values of investigated cytokinins and their analogs ranged from 2.8 to 4.0 at 25 degrees C in the phosphate and acetate buffers of ionic strength 0.015 M. Standard deviations of the constants, obtained by the non-linear fitting of equations for the pKa calculation, were 3-5-times lower than standard deviations from the linear fitting or from the point-to-point calculation utilizing the Hendersson-Haselbalch equation. The equation of Boltzman sigmoid offers two checks on reliability of effective mobilities that serve as the raw data in the pKa calculation.

Oncogene amplification and expression in pediatric solid tumors.

Oncogene amplification and expression and their mutual relationship was analyzed in 92 pediatric tumors by Southern and Northern blot hybridization with N-MYC, ERB A, ERB B, N-RAS and Shb probes. Amplification and overexpression was associated with more advanced clinical stages of tumor, especially in neuroblastomas, rhabdomyosarcomas and ganglioneuroblastomas. The most frequent alteration observed was N-MYC amplification together with overexpression. N-RAS amplification was not detected, while the overexpression of this oncogene was found in 3 cases. Neither amplification nor overexpression was revealed in any specimen of hepatoblastoma or hepatocellular carcinoma. We suggest that oncogenes overexpression provides more accurate prognostic information than amplification.

[Amplification of oncogenes in solid tumors in children]. / Amplifikace onkogenu v detských solidních nádorech.

BACKGROUND: The objective of the work was detection of amplification of oncogenes N-MYC, N-RAS, C-ERB A, C-ERB B and adaptor tyrosine kinase Shb in a group of 92 child age tumours in an attempt to reveal clinical and histopathological associations. METHODS AND RESULTS: Amplifications of oncogenes were detected by means of Southern's transfer, hybridization with labelled probes and densitometric evaluation. Amplification of the N-MYC oncogene in child tumours can be considered a manifestation of progression of the disease with an adverse prognosis, in particular in neuroblastomas, where it corresponds also with the adverse histological finding. In a group of sarcomas N-MYC amplification was detected in advanced clinical stages, while in malignant lymphogranulomas of the Hodgkin type it was not found. In Wilms tumour it was detected sporadically. Amplifications of oncogenes ERB A and ERB B are rare, amplifications of the oncogene RAS were not observed. Coamplifications characterized progression of the disease, in case of neuroblastoma even very short survival. In hepatic malignancies oncogene amplification was not found even in advanced stages. CONCLUSIONS: Oncogene amplification characterizes progression in a number of child tumours and its application in clinical oncology is prognostically useful.

[Late sequelae of comprehensive antineoplastic therapy in children and adolescents with solid extracranial tumors. Effect on growth, pubertal development and gonadal function]. / Pozdní následky komplexní protinádorové lécby u detí a mladistvých s extrakraniálními solidními nádory. Vliv na rust, pubertální vývoj a funkci gonád.

BACKGROUND: Modern treatment of oncological diseases increases markedly the chance of long-term survival and permanent recovery. Due to frequently highly aggressive treatment it is however associated with the risk of late sequelae in the surviving patients. Comprehensive care of patients includes therefore not only control of the neoplastic disease but also efforts of maximal improvement of the quality of life of the patients. In young subjects, in view of their long-term perspective, this problem is particularly important. METHODS AND RESULTS: In 32 patients (25 boys and 7 girls) with extracranial solid tumours without primary endocrinological symptomatology (m. Hodgkin, neuroblastoma, ganglioneuroblastoma, nephroblastoma, Ewings sarcoma and others) a single examination was made assessing height, body weight, grade of sexual maturation according to Tanner, in boys testicular volume by means of a orchidometer and 20 other anthropometric dimensions. The mean age at the time of examination was 16.5 +/- 4.1 years, the mean age at the onset of treatment 6.1 +/- 4.8 years. The patients height, -0.4 +/- 0.9 SD, differs from the Czech national standard (p = 0.025). Impaired growth was recorded in 12.5% patients and had heterogenous causes. The authors proved a negative effect of radiotherapy on the growth of the spine, most markedly in children subjected to irradiation of the abdomen and chest and a highly significant reduction of the testicular volume in boys after cytostatic treatment of m. Hodgkin. CONCLUSIONS: The results are consistent with studies made abroad and indicate the necessity of comprehensive long-term follow-up of somatic growth and development of the gonads in oncological child patients.

[Aortic connectors--the beginning or the end?]. / Aortální konektory--zacátek nebo konec?

In a group of 90 consecutive patients a total of 140 anastomoses were made, using the system of Aortic Connector Symmetry, and the results were compared with a control group of another 90 consecutive patients. Statistically significant differences were found in the period of extracorporeal circulation, the number of tight anastomoses and surgical revisions in favour of the investigated group. The other assessed parameters--period of cardiac ischaemia, number of peripheral anastomoses, mortality and blood losses were not significant.

New aspects of buffering with multivalent weak acids in capillary zone electrophoresis: pros and cons of the phosphate buffer.

Phosphate buffer is frequently used as background electrolyte in capillary electrophoresis. It can cover a broad range of pH due to the three dissociation constants (pK1 = 2.0, pK2 = 7.2, and pK3 = 11.0) of phosphoric acid and because it is UV-light transparent. This contribution brings a theoretical study of the analytical separation performance (sample window, regions of peak symmetry, regions of fronting and tailing peaks) of phosphate buffer, serving as a model of buffering with multivalent weak acids. The study is based on the use of peak shape diagrams and covers the pH range 2.55-11.43. New important general knowledge has been revealed that single multivalent weak acids mimic the performance of background electrolytes with multiple coanionic species for anionic analyses. It is shown that simple phosphate buffers prepared by mixing phosphoric acid and potassium hydroxide may exhibit up to two regions of symmetry, of fronting as well as of tailing zones, on the mobility scale inside the sample window. Moreover they may exhibit a "schizophrenic" region of enormous electromigration dispersion.