Biomedical subjects
Publications and source records attributed to D M Sheppard.
A der(1)t(1;7)(p11;p11) was observed in bone marrow chromosome analyses of four patients with myeloproliferative disorders. Three patients had developed secondary leukemias or preleukemias following chemotherapeutic exposure, and one patient was diagnosed with M4 de novo.
Karyotypes of three sublines of the human cell line U937 showed considerable variation, but all contained four consistent marker chromosomes (i.e., 3q-, 11q-, 16p+, and 17p- chromosomes). The 11q- chromosome appeared to be derived from either an interstitial deletion in bands 11q21-23 or from a translocation with an unidentified chromosome. The presence of this chromosome was of particular interest because rearrangements of chromosome #11 at band 11q23 are often associated with malignancies of the monocytic lineage. The possible significance of these findings is discussed.
Four cell lines derived from adenocarcinomas of the ovary, including three recently established cell lines, have been karyotyped. Chromosomes #1, #3, and #6 were found to be frequently involved in translocations with various other chromosomes, in agreement with results of other investigators, strongly implicating genes on these chromosomes in ovarian tumorigenesis.
Four choriocarcinoma cell lines were karyotyped and examined for genetic homozygosity or heterozygosity using chromosomal polymorphisms. The BeWo, Jar, and ElFa lines had modal chromosomes in the hypertriploid range, while the DoSmi line was hypotetraploid. A number of chromosomal rearrangements were seen in all lines but there was no common rearrangement. The BeWo and Jar cell lines, derived from tumors following term births, were shown to be heterozygous by the presence of both X and Y chromosomes. The ElFa and DoSmi lines, established following molar pregnancies, were shown to be heterozygous; the ElFa line by the presence of both X and Y chromosomes and the DoSmi line by the examination of Q-band polymorphisms. Thus, all four lines, whatever their origin, were shown to be genetically heterozygous.
Using in situ hybridization, c-erbA1 has been mapped immediately distal to the translocation breakpoint on chromosome 17 in fibroblasts with a karyotype 46,XX, t(15;17)(q22;q11). Previous work has shown that c-erbA1 is proximal to the translocation breakpoint on chromosome 17 in the t(15;17)(q22;q12-21) in acute promyelocytic leukaemia. The oncogene can therefore be localized to the region of chromosome 17 between the breakpoints in 17q11 and 17q12-21.
In a study to correlate the genetic origin of hydatidiform moles with the subsequent fate of the patients, 62 complete moles were examined for cytogenetic and/or biochemical polymorphisms. Comparisons of polymorphisms in molar tissue with those seen in white blood cells from the parents have shown that the majority of complete hydatidiform moles are androgenetic in origin and homozygous for all polymorphisms examined. Where karyotyped, these moles were found to be 46,XX. Three exceptions were found in which the molar tissue was heterozygous for enzyme polymorphisms. Cytogenetic analysis of two of these cases confirmed heterozygosity and also revealed a 46,XY karyotype which was androgenetic in origin, most likely arising by dispermy. The two patients with 46,XY complete moles have been followed up for a minimum of 9 months and neither has required treatment for persistent trophoblastic disease.
Uroporphyrinogen decarboxylase (EC 4.1.1.37) has been purified 4419-fold to a specific activity of 58.3 nmol of coproporphyrinogen III formed/min per mg of protein (with pentacarboxyporphyrinogen III as substrate) from human erythrocytes by adsorption to DEAE-cellulose, (NH4)2SO4 fractionation, gel filtration, phenyl-Sepharose chromatography and polyacrylamide-gel electrophoresis. Progressive loss of activity towards uroporphyrinogens I and III occurred during purification. Experiments employing immunoprecipitation, immunoelectrophoresis and titration with solid-phase antibody indicated that all the uroporphyrinogen decarboxylase activity of human erythrocytes resides in one protein, and that the substrate specificity of this protein had changed during purification. The purified enzyme had a minimum mol.wt. of 39 500 on sodium dodecyl sulphate/polyacrylamide-gel electrophoresis. Gel filtration gave a mol.wt. of 58 000 for the native enzyme. Isoelectric focusing showed a single band with a pI of 4.60. Reaction with N-ethylmaleimide abolished both catalytic activity and immunoreactivity. Incubation with substrates or porphyrins prevented inactivation by N-ethylmaleimide. An antiserum raised against purified erythrocyte enzyme precipitated more than 90% of the uroporphyrinogen decarboxylase activity from human liver. Quantitative immunoprecipitation and crossed immunoelectrophoresis showed that the erythrocyte and liver enzymes are very similar but not identical. The differences observed may reflect secondary modification of enzyme structure by proteolysis or oxidation of thiol groups, rather than a difference in primary structure.
Immunoreactive uroporphyrinogen decarboxylase was measured by rocket immuno-electrophoresis in haemolysates from 7 unrelated patients with familial porphyria cutanea tarda (PCT), 6 patients with sporadic PCT, and 7 normal subjects. In all patients with familial PCT immunoreactive enzyme protein was decreased (51% of normal), to the same extent as catalytic activity (56% of normal), whereas in sporadic PCT both measurements were normal. These results show that familial PCT is commonly caused by a mutation which does not lead to the production of non-catalytic cross-reactive immunological material. Familial PCT can be distinguished from other types of PCT by a simple immunoelectrophoretic method that does not involve measurement of uroporphyrinogen decarboxylase activity and which is therefore likely to be suitable for routine diagnostic use.
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In a case of complete hydatidiform mole with fetus the genetic origins were defined by the use of chromosomal polymorphisms. The fetus had a normal 46,XY karyotype with evidence of the presence of both maternal and paternal chromosomes. The mole was 46,XX and of androgenetic origin. There was no evidence of a maternal contribution, and duplication of paternal chromosomes was shown. In such atypical molar pregnancies examining genetic polymorphisms yields much more information than do sex chromosome studies and karyotyping, particularly in confirming the diagnosis and defining the origin and aetiology of the condition.
Cytogenetic and biochemical polymorphisms have been used to determine the origin of a tetraploid conceptus. Genetic polymorphisms were found in chromosomes 1, 6, 9, 15, 16, 22 and the sex chromosomes. The conceptus was found to have one maternal and three paternal contributions, indicating an origin other than a failure of cytokinesis at the first cleavage division of the zygote. The results presented are in favour of a trispermic origin of the conceptus.
Enzyme assays for phenylalanine mono-oxygenase have been performed on a variety of non-hepatic human cell types under varying conditions. The cell types studied were cultured fibroblasts, short-term lymphocyte cultures, long-term lymphoblastoid cultures, cultured amniotic fluid cells, hair roots and placental extracts. The cultured cells were assayed after growth under standard conditions and in the presence of a high concentration of substrate, a low concentration of end-product, added hydrocortisone or dexamethasone and under combinations of these conditions. In no instance was a significant enzyme activity obtained.
1. Erythrocyte uroporphyrinogen decarboxylase activity has been measured in 27 patients with porphyria cutanea tarda, of whom 11 had a family history of overt porphyria cutanea tarda. 2. Eight patients from six families had erythrocyte uroporphyrinogen decarboxylase activities that were decreased to about half of control values. This decrease was shown by family studies to be inherited as an autosomal dominant characteristic. Two of these patients had no family history of overt porphyria cutanea tarda. 3. Nineteen patients had uroporphyrinogen decarboxylase activities close to or within the range found in 18 control subjects. Of these, five patients had a family history of porphyria cutanea tarda. 4. Inheritance of an autosomal dominant gene which decreases uroporphyrinogen decarboxylase activity in erythrocytes and liver is an uncommon cause of porphyria cutanea tarda and may not explain all cases of familial porphyria cutanea tarda. The hepatic enzyme defect in the common type of porphyria cutanea tarda, in which erythrocyte uroporphyrinogen decarboxylase activity is normal, may be caused either by inheritance of a gene whose effect is restricted to the liver or by gene whose effect is restricted to the liver or by chemicals that selectively inhibit the hepatic enzyme.
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