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Biomedical subjects

Z Lojda

Publications and source records attributed to Z Lojda.

At least 19 recordsLinked to original sources
PubMed1979

The histochemical demonstration of brush border endopeptidase.

A histochemical method for the demonstration of a brush border endopeptidase is described based on results of biochemical and histochemical experiments. The substrate of choice is Glut-Ala-Ala-Ala-MNA which displays a very good localization ability and suitable kinetic properties. Km estimated in rat kidney homogenate amounts to 2.35 X 10(-4) M. pH optimum of this endopeptidase associated with the brush border membrane is in the alkaline range. The activity is dependent on the buffer used. In phosphate and cacodylate buffers of pH 7.2 about 30% lower activity in rat kidney and about 25% lower activity in rat small intestine than in Tris-HCl buffer of the same pH was found. The most suitable diazonium salt for the detection "in situ" is Fast Blue B. It inhibits the endopeptidase activity of rat kidney by about 85% at pH 7.2 AND BY ABOUT 55% AT PH 6.0. The best results are obtained in cryostat sections adherent to semipermeable membranes treated with chloroform-acetone before the incubation. A microdensitometric evaluation of the reaction product is possible and results are in good agreement with those of the biochemical determination. When Suc-Ala-Ala-Ala-INA is used as substrate hexazonium-p-rosaniline is the most suitable coupling agent although it inhibits more than Fast Blue B. The reaction using acylated trialanyl naphthylamides as substrates runs in two steps. Endopeptidase sets free Ala-NA which is attacked by aminopeptidase M. Aminopeptidase M is not reaction rate or localization limiting factor because its activity in the brush border is very high and the enzyme is anchored to the cell membrane very closely to endopeptidase. In homogenates of rat kidney and jejunal mucosa the endopeptidase activity was inhibted by EDTA (2X10(-3) M) by 75% in the kidney and by 68% in the jejunum, by DFP (10(-3) M) by 41% in the kidney and by 35% in the intestine, by Mn2+ (5X10(-3) M) by 25% in the kidney and by 30% in the intestine. No inhibition was exerted by E 600. In sections the results were similar. 1,10-phenanthroline (10(-2) M) caused a substantial inhibition. Endopeptidase activity was detected in the brush border of cells of proximal convuluted tubules of the kidney and in the brush border of differentiated enterocytes of the small intestine. In the same species enterocytes display a lower activity than kidney tubular cells. There are species differences in the distribution pattern of endopeptidase in the kidney. In the rabbit and man the positive reaction occurs in the whole cortex. It is distributed unevenly, however. In the rat the tubules of the inner cortex display a very high activity. In the outer cortex straight portions react strongly. In the rabbit kidney cells of the parietal layer of Bowman's capsule display a weak reaction as well. No sex differences were found in the distribution pattern of endopeptidase in the rat kidney. In the intestine of all species examined a proximo-distal gradient was found...

Adult↗
PubMed1979

Histochemistry of the human duodenal mucosa with special reference to the gradient of activities of the brush border enzymes.

Histochemical examination of aspiration biopsy specimens of the duodenum in 8 healthy volunteers and 23 patients with various gastrointestinal disorders revealed no characteristic abnormality except for a rather frequent absence of the activity of disacharidases especially of lactase. A distinct difference of disacharidase was established between the duodenum and the jejunum in some individuals. A systematic study of different levels of the upper gastrointestinal tract established the fact that the activity of brush border enzymes of enterocytes starts to appear in individually different parts of the duodenum and sometimes even as low as in the jejunum. The distribution of lysosomal enzymes and the activity of dehydrogenases display differences as well. Consequently specimens from different parts of the duodenal and jejunal mucosa are not equivalent and cannot be used alternatively for diagnostic purposes. Biopsies of duodenal mucosa are not adequate for the diagnosis of the malabsorption syndrome.

Adult↗
PubMed1977

Experimental study on the mechanism of hyperphosphatasaemia in bile flow obstruction.

Extrahepatic bile flow obstruction is followed by a several-fold increase in alkaline phosphatase activity in the liver. This activity passes into the blood. The increase in the activity of the enzyme in the liver can be prevented by inhibitors of RNA and protein synthesis. Inducible alkaline phosphatase was purified from rat liver and 1,000-fold purification was achieved. The incorporation of labbelled precursors (14C amino acids and 14C glucosamine) into the liver alkaline phosphatase was evaluated. No significant difference were found between specific alkaline phosphatase activities and specific radioactivities in the control rats and rats with 7.30 h after bile duct obstruction. The incorporation of 14C-amino acids into the total liver proteins was 20% lower in rats with cholestasis than in the control group. The results are evidence that the liver is the source of elevated alkaline phosphatase activity in serum in cholestasis, and that the production of alkaline phosphatase in the liver is prevented by inhibitors of RNA a protein synthesis. However, the enzyme might be synthesized before the 14C-labelled precursors were applied.

Alkaline Phosphatase↗
PubMed1977

Diagnostic significance of determination of serum alkaline phosphatase intestinal isoenzyme activity in coeliac sprue in childhood.

In a group of 26 child patients with coeliac sprue verified by clinical tests and biopsy, we observed a significant change in the proportion of the intestinal alkaline phosphatase isoenzyme in the serum in correlation to a gluten-free therapeutic diet. In untreated patients, in the active phase of the disease, intestinal AP isoenzyme activity rises in the serum and falls in the intestinal mucosa. In our experience, the change in activity is reversible within 3--5 days of instituting adequate treatment, i.e. a gluten-free diet. No such activity changes were observed in children with other forms of malabsorption syndrome (hepatogenic, pancreatogenic) or in the children in the control group. This test was found to be a suitable and sensitive method for indirect determination of the response of the intestinal mucosa to treatment in children with coeliac sprue.

Adolescent↗
PubMed1977

Histochemical diagnosis of lipidoses.

Contemporary possibilities for the histochemical diagnosis of lipidoses are demonstrated in examples of phospholipidoses, Gaucher's disease, Fabry's disease, sulphatidosis, gangliosidosis and neuronal ceroid-lipofuscinoses.

Biopsy↗
PubMed1977

Possibilities for the cytochemical diagnosis of enzymopathies.

The authors review the contemporary uses of histochemistry for the diagnosis of enzymopathies. Enterokinase, lactase, sucrase and trehalase deficiency can be diagnosed by histochemical methods. In glycogenoses, glycogen storage and glucose-6-phosphatase, acid alpha-glucosidase and phosphorylase deficiencies can be demonstrated. In mucopolysaccharidoses, the accumulation of acid muco-substances and changes in lysosomal enzyme activities can be demonstrated.

Biopsy↗