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Biomedical subjects

D Whittaker

Publications and source records attributed to D Whittaker.

27 records · Page 2Linked to original sources
PubMed1980

Erythrocyte pyrimidine 5'-nucleotidase.

In this study 31 family members of a patient with erythrocyte pyrimidine 5'-nucleotidase deficiency were studied. The activity of this enzyme in their erythrocytes is compared with levels in normal subjects and the problems surrounding heterozygote detection are discussed. The mean erythrocyte pyrimidine 5'-nucleotidase activity in 158 normal fresh blood samples was 130 +/- SD 29.8 mU/gHb. There was no significant difference between males and females. The enzyme level in a patient with non-spherocytic haemolytic anaemia was 6 mU/gHb. Of 31 relatives of the enzyme deficient patient examined five were clearly heterozygous for the enzyme defect. Their enzyme levels were below 50 mU/gHb. The father, who is an obligatory heterozygote, had an enzyme level of 87 mU/gHb which falls within the low values of the normal range. The distribution of enzyme activity in 26 family members having enzyme activities greater than 50 mU/gHb suggests that six of these may be carriers of the defective gene. We were unable to identify carriers by enzyme kinetic studies, electrophoresis, chromatographic examination of acid extractable nucleotides or measurement of enzyme levels in young and old erythrocyte populations. The last-mentioned technique showed that erythrocyte 5'-nucleotidase activity in reticulocytes may be as high as 1785 mU/gHb and declines rapidly as the cell ages reaching about 50 mU/gHb in the oldest cells. Blood samples which had been stored frozen were examined to see whether such samples were satisfactory for population studies. The mean enzyme activity 151 blood samples stored frozen for more than 12 months was 153+/-SD 44.7 mU/gHb. The increase in enzyme levels in the frozen samples appears to be greatest in samples showing haemolysis. In spite of the increased enzyme level frozen samples could be used to detect subjects with enzyme deficiency and some heterozygotes.

5'-Nucleotidase↗
PubMed1979

Distribution of erythrocyte nucleotides in pyrimidine 5'-nucleotidase deficiency.

In pyrimidine 5'-nucleotidase deficiency, erythrocytes contain elevated levels of pyrimidine nucleotides. The composition of this nucleotide pool was examined by ion exchange chromatography on Dowex formate columns using a linear ammonium formate elution gradient. In contradistinction to normal erythrocytes, adenine nucleotides accounted for only 32% of the nucleotide pool. The remainder consisted of 50% cytidine and 16% uridine nucleotides. The remaining 2% was not identified. The most abundant compound appeared to be UDP glucose whilst high levels of CTP, CMP and an unidentified cytidine compound less polar than CMP accounted for most of the cytidine nucleotide pool. The possibility that the abnormal nucleotides were due to an elevated reticulocyte count was excluded and it was also shown that erythrocytes from subjects heterozygous for pyrimidine 5'-nucleotidase deficiency did not have detectable levels of the abnormal nucleotides.

Adenosine Triphosphate↗
PubMed1977

Purification and properties of human erythrocyte pyrimidine 5'-nucleotidase.

A 250,000-fold purification of pyrimidine 5'-nucleotidase from human erythrocytes has been achieved using a combination of DEAE-cellulose chromatography, ammonium sulfate fractionation, gel filtration, and isoelectric focusing. Polyacrylamide disc and starch gel electrophoresis of the purified material show two strong protein bands. On starch gel these bands exhibited pyrimidine 5'-nucleotidase activity. Two faint protein bands devoid of enzyme activity were also found in the case of polyacrylamide electrophoresis. The enzyme has a pH optimum at 7.5 and is most stable between pH 6 and 7.5. The enzyme has pI of 5.0 and a molecular weight of 28,000 by gel filtration. The Km of the purified enzyme was 10 muM, compared to 40 muM when measured in hemolysate. The higher Km in the hemolysate is due to the presence of an inhibitor. Inorganic phosphate was shown to be a competitive inhibitor of pyrimidine 5'-nucleotidase and inorganic phosphate in the hemolysate may be responsible for increasing the Km of the enzyme for the substrate cytidine monophosphate.

Erythrocytes↗
PubMed1977

Fate of prostaglandins E(1) and A(1) in the human pulmonary circulation.

It is recognized that the lung extracts norepinephrine and 5-hydroxytryptamine from the pulmonary circulation and that this process is affected by cardiopulmonary bypass. Since alterations in the lung's processing of vasoactive substances may be a mechanism of pulmonary injury sustained during operation, we investigated the lung's ability to extract or metabolize prostaglandin A1 (ga1) and prostaglandin E1 (PGE 1). Sixteen patients undergoing cardiac surgery were studied. In five patients, just before going on bypass, a 10 ml of blood was withdrawn at a constant rate, simultaneously from the pulmonary artery and left atrium. In 11 patients, 3H-PGE1 was injected just prior to bypass and, in five of these, again after coming off bypass. Extraction was calculated from tritium activity in the samples. Metabolites were quantitated by thin-layer chromatography after being identified by marker compounds run simultaneously in each chromatogram. The pulmonary extraction of PGA1 was 11.3 +/- 2.3% and there were no detectable metabolites in left atrial blood. Before bypass the extraction of PGE1 was 42.3 +/- 14.3% and after bypass 24.8 +/- 10.0% (P less than 0.005; Student's paired t test). PGE1 was extensively metabolized with 79.7 +/- 7.1% of total radioactivity appearing in the left atrium as metabolites before bypass and 89.1 +/- 2.0% appearing after bypass. This study indicates that PGA(1) is not metabolized by the lung and is only slightly extracted. On the other hand, PGE(1) is extensively extracted and metabolized. While the rate of metabolism is not significantly affected by cardiopulmonary bypass, the extractiom before bypass was significantly greater than after bypass.

Adult↗
PubMedDate not supplied

Sex determination by PCR analysis of DNA extracted from incinerated, deciduous teeth.

Establishing the biological sex of human remains is a very important part of identifying victims of fire when severe soft tissue destruction has occurred. Deciduous (children's) teeth were exposed to a range of incineration temperatures 100-500 degrees C for 15 minutes. Polymerase Chain Reaction (PCR) amplification was used to identify specific human amelogenin regions. There was successful identification of human biological sex, from deciduous teeth exposed to incineration temperatures of 200 degrees C and below, using standard ethidium bromide gel staining. There was greater sensitivity using fragment analysis by laser induced fluorescence which achieved sex identification from some teeth heated to 400 degrees C.

DNA↗