Контроль гіпотетичного прояву хронічного синдрому втоми: попередні спостереження

Предмет:

Медицина

Тип роботи:

Стаття

К-сть сторінок:

20

Мова:

Українська

Оцінка:

subjects involved several steps. All subjects underwent an extensive medical evaluation, consisting of a standard physical examination and medical history, an exercise capacity test, a symptom checklist and routine laboratory tests. The laboratory tests included a complete blood cell count, determination of the erythrocyte sedimentation rate, a serum electrolyte panel, measures of renal, hepatic, and thyroid function, and rheumatological and virological screenings. In a number of cases further neurological, gynaecological, endocrinological, cardiac, psychiatric and/or gastro-intestinal evaluation was performed. When positive results were found in any of the evaluations that met the Fukuda (2) exclusionary criteria, the patients were not included in this study. The medical records were reviewed to determine if patients suffered from organic or psychiatric illnesses that could explain their symptoms. All patients completed a questionnaire which included demographic information, dates of onset and curren health status. Afterwards the subjects were examined by one physiciai (KDM), who interviewed the patients with respect to their signs anc symptoms. Subjects were excluded if they were < 18 or > 66 years o: age, using one of the medications listed in Table 1, or reported an episode of diarrhoea or vomiting. All patients and controls were Caucasian. Demographic characteristics of the sample are presented in Table 2.

Measurement of Whole Body Potassium by Gamma-Ray Spectrometry

Whole body potassium was measured by gamma-ray spectrometry, which assesses isotope 40K. A naturally occurring gamma-radiation-emitting isotope, 40K exists in the human body at a constant 0. 012% of total body potassium (8, 9, 10). Gamma-ray spectrometry occurred at the isotope-centre of Brugmann hospital (AZ-V. U. B. & U. L. B.), using a whole body scintillation counter (model 8102A, Nuclear Enterprises, Brussels), consisting out of four detectors (voltage sensitive preamplifier, model NE 5288B). The same equipment was used in the study of Burnet et al. (7). The counter consists out of four detectors (voltage sensitive preamplifier, model).

TABLE 1

List of Medications Used for Exclusion

Diurética

Angiotensine-Converting enzyme inhibitor

Potassium supplements

Therapeutic steroids*

Antibiotics*

* glucocorticoids and some forms of antibiotics stimulate secretion of renal potassium, which in turn causes hypokalemia

The detectors were enclosed within a lead lined pre-1945 shield of wall thickness approximately 15 cm, to reduce background radiation to a minimum and to increase sensitivity. Subjects were dressed in clean hospital gowns, and laid supine for 30 minutes on a transparent perspex sheet. The latter enables the operator to accurately align the lower detectors with a selected axis or area of the patient. Data acquisition is performed by an AccuSpec Nal Plus board computer interface (Canberra Industries).

TABLE 2

Demographic Features of Our Sample

NPatientsControlsP-value

2720

# fulfilling Holmes et al. 160

# fulfilling Fukuda et al. 270

# males (%) 2 (7, 4) 2 (10) , 759

# females (%) 25 (92, 6) 18 (90)

Mean age41, 138, 6, 371

SDage8, 610, 4

Calibration was performed using known concentrations of isotope in water phantoms (plastic bottles). Phantom counts showed a coefficient of variation (CV) of 1. 1% and background CV counts of 2. 3%. Total body potassium results are expressed in mEq/kg, and also as a percentage of expected normal value. Expected normal values are predicted on the basis of height, weight, age and sex from a database of 61 healthy subjects, previously measured using the same whole body counter (12).

RNase L-Ratio Determination

Within four hours of phlebotomy, peripheral blood mononuclear cell extracts (PBMC) were separated from heparinized blood (30 ml) by Ficoll-Hypaque density gradient centrifugation. In addition, PBMCs were stored at -70°C until cytoplasmic extraction preparation (3). The latter was performed in the presence of protease inhibitors aprotinin, leupeptin, pefabloc-SC and EDTA (Roche Biochemicals, Mannheim, Germany). Standard laboratory procedures were used to separate serum from coagulated blood, and to store it at -70°C until analysis. A modified Bradford assay method (Bio-Rad Laboratories, Hercules, CA) was used for quantification of total proteins in the patients' cell extracts and serum. Briefly, 200 ug of PBMC extract was incubated with a meta-periodate (10 mM final concentration, pH 4. 75) oxidized 2-5A trimer radiolabeled at the 3' end with 32P-pCp as the receptor ligand, at 2-4°C for 15 minutes. In addition, it was covalently attached to the binding proteins by the addition of cyanoborohydride (20 mM in 100 mM phosphate buffer, pH 8. 0). This reduction reaction was allowed to progress for 20 minutes at 2-4°C. Sodium dodecylsulfate polyacrylamide gel electrophoresis (SDS-PAGE) buffer and a tracking dye were added to the samples, and incubated at 95°C for 5 minutes followed by separation using standard SDS-PAGE with a 4% stacking and a 10% separating gel. The gel was dried and autoradiography was performed