Comparative studies of nuclear DNA content in benign and malignant thyroid lesions.

Currently available data suggest that DNA aneuploidy is associated with aggressive behavior of and unfavorable prognosis in several malignant human tumors as compared with diploid malignancies. However, the diagnostic and prognostic importance of flow cytometric DNA measurements in the case of thyroid neoplasms remains controversial. Therefore, the aim of our study was to evaluate utility of DNA index (DI) and proliferative index (PI) in distinguishing benign from malignant thyroid lesions taking into account the possible influence of intra-tumor heterogeneity and tissue preparation mode on DNA flow-cytometry measurements. A retrospective study was performed on 71 paraffin-embedded specimens from 57 patients with benign and malignant thyroid pathologies: 13 colloid goitres, 12 parenchymatous goitres, 19 adenomas and 13 carcinomas. In 14 of 57 cases two separate specimens taken from different areas of the same lesion were analysed and DNA parameters were compared. Additionally, flow cytometry DNA analysis was parallelly performed on 3 adjacent but differently processed tissue sections (fresh, formalin-fixed and paraffin-embedded) taken from each of 26 surgically excised thyroid lesions. DNA content was also analysed in both fresh and formalin-fixed twin specimens of normal pig thyroid glands (N = 6). We demonstrated that all tumors diagnosed as thyroid carcinomas were associated with abnormal nuclear DNA content although aneuploidy was not found specific to malignant thyroid tumors. Aneuploid samples of benign thyroid lesions exhibited higher proliferative activity, expressed as mean PI values, than diploid ones. In carcinomas the mean PI values were significantly higher than in benign lesions, independently whether they concerned aneuploid or diploid tissues. Considering intra-tumor heterogeneity, the flow cytometric DNA parameters can be assumed as reproducible despite differences in the mode of tissue fixation and preparation for analysis.

Fructosamine in human and bovine semen.

We detected the presence of fructosamine in human and bovine semen. In seminal plasma of healthy normozoospermic men (N = 17) fructosamine was found in 53% of the cases (fru+). In fru+ semen samples the concentration of fructosamine was (mean +/- S.E.M., N = 9) 0.45 +/- 0.09 mmol/L and varied from 0.15 to 0.75 mmol/L. It was 3-12 times lower than in blood serum of healthy men. In semen of infertile men (N = 57) fructosamine was present only in 21% of the cases and its concentration was lower than in fertile men i.e. (mean +/- S.E.M., N = 12) 0.27 +/- 0.007 mmol/L. In bulls (N = 98) fructosamine was found in semen of 82% of animals. In fru+ semen samples the concentration of fructosamine was (mean +/- S.E.M., N = 80) 0.77 +/- 0.12 mmol/L and varied from 0.30 to 1.15 mmol/L. We did not find any correlation between the concentration of fructosamine on one hand, and that of fructose and glucose on the other hand, in either human or bull semen. The difference in the frequency of fructosamine appearance in semen of fertile and infertile men suggests that fructosamine may be in some way involved in the process of fertilisation.

[Multi-indicator biochemical and clinical evaluation of occupational exposure of workers in the petrochemical industry. II. Results of the search for early clinico-biochemical signs of exposure to toxic substances present in the petrochemical industry. Analysis of the results of 5-year epidemiological studies]. / Wielowskaznikowa ocena biochemiczno-kliniczna narazenia pracownikówprzemyslu petrochemiczego. Cz. II. Wyniki poszukiwan wczesnych objawów kliniczno-biochemicznych narazenia na toksyczny wplyw substancji wystepujacych w przemysle petrochemicznym. Analiza wyników 5-letnich badan epidemiologicznych.

The activities of GGT (EC 2.3.2.1.) and glycine transaminidase (EC 2.6.2.1.) increased in all studied groups after at minimum four years of exposure to: acetobenzene, furfurol, ethylene--derivatives, polypropylene and butadiene. Besides these universal indicators the specific ones were found for a particular group of exposure: for acetobenzene--sialic acid, haptoglobine and IgA, for furfurol--sialic acid and haptoglobine, for ethylene--derivatives--haptoglobine, ceruloplasmine and the activity of arginase of the red blood cells, for polypropylene--IgA, for butadiene--proteolytic activity. These changes are treated as a manifestation of adaptive processes--the answer of the body to the environmental stimuli but not as the signs of early damage. The need of verification of the results by prospective studies is stressed.

[Multi-indicators in the biochemical and clinical evaluation of exposure of workers in the petrochemical industry. I. Preliminary epidemiologic screening studies]. / Wielowskaznikowa ocena biochemiczno-kliniczna narazenia pracowników przemyslu petrochemicznego. CZ. I. Wstepne badania epidemiologiczno-skriningowe.

Eleven non-routine biochemical and clinical indicators (selected of 26 preliminarily estimated) in Petrochemical Plant workers and in controls, localized in an increasing distance from the Plant (3-18-40 km), have been measured. Even in the 3 km distance, no effects of the Petrochemical Plant have been found. All the exposed groups exhibited changes in enzymes GGT and TA. We estimate these indicators as universal. The concentration of haptoglobin (Hp) changed in 3 groups, whereas sialic acid and arginase--in 2 exposed groups.

Studies on L-arginase in developing rat small intestine, brain, and kidney. I. Ontogenic evolution of arginase isoenzymes.

The adult patterns of arginase isoenzymes in rat intestine, kidney, and brain are nearly identical and consist of two forms, cationic A1 and anionic A4. In this paper, the organ-specific maturation of the enzyme equipment in these tissues is reported. The activity of arginase in all tissues studied could be detected on the 13th to 16th days of gestation. In fetal intestine and kidney the arginase activity is low, and persists up to the weaning time when the rapid, 10-fold rise of the enzyme activity occurs. However, the adult pattern of arginase isoenzymes in these tissues is accomplished in different ways. In the intestine, arginase A1 appears in fetal life and is the only form of the enzyme till the 19th to 21st days of postnatal life when the second form of arginase, A4, appears and rapidly accumulates, being exclusively responsible for the rise of the total enzyme activity at the time of weaning. In kidney, arginase A1 alone is present in the early fetal period. Arginase A4 appears 3-4 days before birth and its activity persists unchanged within the first 2 weeks of postnatal life. The intensive rise in total specific activity of kidney arginase at weaning is due to the accumulation of preexisting arginase A4. In brain, the adult pattern of arginase isoenzymes is achieved earlier than in other tissues. Both forms, A1 and A4, occur on Days 13-14 of gestation.

Studies on L-arginase in developing rat small intestine, brain, and kidney. II. Effect of hydrocortisone and thyroxine.

The influences of hydrocortisone and thyroxine on the developmental changes of arginase activity in intestine, kidney, and brain of suckling rats were studied. A single injection of hydrocortisone (50 mg/kg) into rats aged 9 days evoked premature increase of jejunal arginase activity due to precocious formation of arginase A4. Arginase A4 can be detected about 48 hr after hydrocortisone injection, whereas in intact rats the enzyme appears in the intestinal mucosa on the 19th-21st days of postnatal life. After hydrocortisone administration to rats aged 6 days, a similar pattern of arginase activity in jejunum was observed. Under the same conditions, the influence of hydrocortisone on kidney arginase was weaker. The hormone did not have any influence on the activity of brain arginase. Daily injection of thyroxine (2 mg/kg) to 6-day-old rats (for 6 consecutive days) caused a precocious increase of the arginase activity in intestine. Under the same conditions, only a slight increase of the arginase activity was observed in kidney, whereas in brain the activity was unaffected.

A simple quantitative micromethod of arginase assay in blood spots dried on filter paper.

An accurate, precise and sensitive method has been developed for measurement of arginase activity in erythrocytes in dried blood spots. The assay is based on colorimetric measurement of ornithine produced by enzymatic hydrolysis of L-arginine. Only 10 microliter of capillary blood collected on filter paper are required. One disc of 3 mm in diameter punched from the dried blood spot is used for the arginase assay. The whole procedure is performed in one test-tube and does not need deproteinization. The second disc of the same diameter is used for hemoglobin (Hb) measurement. There was a good correlation between activities determined in dried blood spots and fresh erythrocytes of the same blood specimens taken from 101 healthy adults and 49 children (corre. coeff. 0.955 and 0.968, respectively). Arginase activity in dried blood specimens was 68.3 +/- 22.7 in adults and 62.7 +/- 15.7 U/g Hb in children. In 118 newborns, the activity was 101.9 +/- 29.2. In 1270 residents of nursing homes screened for hyperargininemia the activity was 21.5-171.2 U/g Hb. In screening for arginase deficiency, the method may be used as a simplified qualitative test without Hb assay.

Human salivary arginase and its deficiency in argininaemia.

Arginase from normal human mixed saliva was characterized. The enzyme was completely activated after a preincubation of 20 min at 55 degrees C and a Mn2+ final concentration of 5 mmol/l. The pH optimum was 9.6-9.8, and the Km for L-arginine was 4.2 +/- 0.7 mmol/l. In normal saliva only one form was found, which was chromatographically identical with the cationic form of arginase in liver and blood cells. Salivary arginase was completely precipitated by rabbit antiserum against human liver arginase. Arginase activity was not detectable in the saliva of patients suffering from argininaemia. Enzyme activities in the saliva of the heterozygous parents and the unaffected daughter were 0.08, 0.07 and 0.12 U/mg protein, respectively, whereas the activities in the saliva of 60 healthy adults and 8 children were 0.17 +/- 0.11 and 0.16 +/- 0.06 U/mg protein, respectively.

Isoenzyme pattern and immunological properties of arginase in normal and hyperargininemia fibroblasts.

Arginase deficiency is an inborn error of the last step in the urea cycle and leads to profound hyperargininemia. The enzyme deficiency has been demonstrated in the liver and red blood cells. In cultured patient fibroblasts, the activity is normal. Arginase exists in multiple molecular forms only one of which is missing in hyperargininemic patients. In fibroblasts, three arginase isoenzymes can be demonstrated by DEAE-cellulose column chromatography, two by electrophoresis and by immunoprecipitation methods. From the present data, it is improbable that part of the A1 isoenzyme in fibroblasts originates from fetal calf serum arginase which supplements the culture media. None of the techniques for the separation and analyses of arginase isoenzyme allows to differentiate between the normal and the arginase-deficient phenotype. A possible explanation would be that the defect in A1 arginase observed in the liver is the result of a regulatory defect.

Arginase activity in human fibroblast cultures.

Controversial results have been published as to the presence of arginase activity in human skin fibroblasts in normal cells and in argininemia. Experiments were undertaken to see if arginase is intrinsic to the fibroblasts or may be there as the result of exogenous contamination inherent to the mode of cell culturing. The mode of harvesting by scraping or trypsin treatment demonstrated similar arginase activity. No significant differences of arginase activity were found between the fibroblasts grown on fetal calf or human serum. In the conditioned serum-free medium arginase separated on DEAE-cellulose into an A1 and A4 form. These fractions are identical with those found in the cells. Arginase activity varies, however, with age of the fibroblasts. These observations are also valid for fibroblasts from a case of argininemia. Arginase activity is therefore intrinsic to diploid human fibroblasts.

A simple, rapid and accurate screening test for hyperchlororhoea.

A simple, rapid screening paper test for the quantitative estimation of Cl- concentration in feces is described. The measured time of decolorisation of the Chlorotrex indicator by aqueous extract of feces is inversely proportional to the chloride concentration. The technical details of the test are given. The test is very suitable for laboratory and mass screening.