Biomedical subjects

M J Scanlon

Publications and source records attributed to M J Scanlon.

36 records · Page 2Linked to original sources

PubMed1995

Primary sequence determination and molecular modelling of the variable region of an antiMUC1 mucin monoclonal antibody.

Polymerase chain reaction (PCR) products representative of the DNA sequence coding for the variable heavy (VH) and the variable light (VL) chains of an antiMUC1 mucin monoclonal antibody, C595, have been produced. These products were cloned, sequenced, and the primary amino acid sequences of the VH and VL regions deduced. The hypervariable complementarity determining regions (CDRs) and framework regions in the heavy and light chains were located, and homologies with canonical forms for the CDR loops L1, L2, L3, H1 and H2 were identified by database searching. The structure for the H3 loop was calculated directly. Computational molecular modelling was accomplished using the fully automated AbM package (Oxford Molecular, Oxford, U.K.). Energy minimisation was performed using the program InsightII (Biosym, San Diego, California, U.S.A.). The investigation provides a basis for the molecular analysis of the antigen binding site of the C595 antibody with the aim to identify key residues and interactions involved in the immune recognition of the C595 antibody defined epitope, which is expressed in the majority of breast and ovarian carcinomas.

Amino Acid Sequence↗

PubMed1994

Multiple conformations of the sea anemone polypeptide anthopleurin-A in solution.

Anthopleurin-A (AP-A) is a member of a family of sea anemone-derived polypeptides that interact with sodium channels in a voltage-dependent manner, producing a positive inotropic effect on the mammalian heart. There has been considerable interest in this molecule as a lead compound for the development of novel therapeutic agents. Earlier attempts to define the 3-dimensional structure of AP-A were complicated by the fact that it was found to exist in 2 conformations in solution. Using 1H- and 13C-NMR spectroscopy, we have now shown that this conformational heterogeneity arises from cis-trans isomerization about the Gly 40-Pro 41 peptide bond and that in the major form of the protein this peptide bond adopts a cis conformation. Furthermore, the increased sensitivity afforded by higher-field NMR has allowed identification of additional minor conformations of AP-A, the origin of which is presently unknown. We believe there will be many more examples of the detection by high-field NMR of previously unobserved minor conformations of proteins in solution.

Amino Acid Sequence↗

PubMed1994

Genetic analysis of 63 mutations affecting maize kernel development isolated from Mutator stocks.

Sixty-three mutations affecting development of the maize kernel were isolated from active Robertson's Mutator (Mu) stocks. At least 14 previously undescribed maize gene loci were defined by mutations in this collection. Genetic mapping located 53 of these defective kernel (dek) mutations to particular chromosome arms, and more precise map determinations were made for 21 of the mutations. Genetic analyses identified 20 instances of allelism between one of the novel mutations and a previously described dek mutation, or between new dek mutations identified in this study; phenotypic variability was observed in three of the allelic series. Viability testing of homozygous mutant kernels identified numerous dek mutations with various pleiotropic effects on seedling and plant development. The mutations described here presumably arose by insertion of a Mu transposon within a dek gene; thus, many of the affected loci are expected to be accessible to molecular cloning via transposon-tagging.

Alleles↗

PubMed1993

DNA sequence and transcript analysis of transposon MuA2, a regulator of Mutator transposable element activity in maize.

The 4942 bp DNA sequence of Zea mays transposon MuA2 was determined. Previous evidence indicated MuA2 controls activity of the Mu1 transposon located in the mutable allele a1-mum2. MuA2 contains two large, ATG-initiated open reading frames (ORFs) of 612 and 232 codons, respectively, located on opposite strands. MuA2 produces two transcripts, each containing one of these ORFs. Four different tandem direct repeat sequences are located downstream of the 612 codon ORF. The restriction map of MuA2 is identical to that of transposon MuR1, which also is known to regulate mutability of a1-mum2. Furthermore, except for a single nucleotide, MuA2 is identical to the Mutator element Mu9.

Amino Acid Sequence↗

PubMed1993

Hypersecretion of luteinizing hormone and ovarian steroids in women with recurrent early miscarriage.

Polycystic ovary syndrome is associated with hypersecretion of luteinizing hormone (LH) which has been implicated in the aetiology of early pregnancy loss. Although 82% of women with recurrent early loss have polycystic ovaries on ultrasound imaging, random serum LH concentrations are normal. In the present study, we have obtained further information from serial samples concerning the cyclical patterns of gonadotrophin and sex steroid secretion in these women. Twenty-one women with recurrent early pregnancy loss and 10 multiparous controls were investigated; 81% of them and one of ten control subjects had polycystic ovaries. Mean mid-follicular and mid-luteal serum LH and follicle stimulating hormone (FSH) levels were similar in both groups. Seventeen women with pregnancy loss had either raised urinary LH excretion or a premature LH surge; one control subject had a premature LH surge. Total LH excretion during the cycle and mean follicular phase serum testosterone was significantly greater with early pregnancy loss than in the control group, the difference in LH being greatest in the early luteal phase. Urinary oestrone-3-glucuronide excretion was raised in the early luteal phase of the cycle in the group with early pregnancy loss; there was no difference between the groups in pregnanediol-3 alpha-glucuronide excretion. These data demonstrate abnormalities in LH secretion in 81% of women with recurrent fetal loss. Inappropriately raised LH levels may have adverse effects on the developing oocyte or endometrium either directly, or indirectly by causing an elevation in testosterone and oestrogen levels.

Abortion, Habitual↗

PubMed1992

Structural and computational investigations of the conformation of antigenic peptide fragments of human polymorphic epithelial mucin.

Human polymorphic epithelial mucins (PEM) are complex glycoproteins that are associated with breast and ovarian carcinomas. The PEM core protein consists of variable numbers of a tandem repeat sequence which contains a short antigenic hydrophilic region (Pro1-Asp-Thr-Arg-Pro-Ala-Pro7). High-field n.m.r. studies undertaken on antigenic 20- and 11-amino acid fragments of the PEM core protein in dimethyl sulphoxide have identified a type-I beta-turn to be present in the region Pro1-Asp-Thr-Arg4. This region includes and overlaps the identified type-I (Asp2-Thr-Arg4) and type-II (Arg4-Pro-Ala6) epitopes of anti-PEM monoclonal antibodies. The studies indicate that the beta-turn is stabilized by the presence of a salt-bridge interaction between Asp-2 and Arg-4. In order to probe the conformations accessible to the PEM peptides a computational study was undertaken independently on the peptide Pro-Asp-Thr-Arg-Pro using a modified Metropolis Monte Carlo algorithm. This study identified the n.m.r.-observed salt-bridge type-I beta-turn as the major low-energy conformer. These results suggest that this structural motif may be involved in the immune recognition of PEM.

Amino Acid Sequence↗

PubMed1992

Peptide epitope binding and fluorescence quenching with the anti-mucin antibody C595.

The Fab fragment of the anti-polymorphic epithelial mucin antibody C595 (IgG3, kappa) has been produced by digestion with papain. The purified antibody fragment retained its binding activity for the mucin in an indirect radioisotopic antiglobulin assay. The binding constant of the Fab fragment for the synthetic peptide (PDTRPAPGSTAPPAHGVTSA) corresponding to the 20-amino acid tandem repeat sequence found in the mucin protein core was determined to be 0.3 x 10(6) M-1 by measuring the capacity of the peptide to quench the fluorescence of the Fab fragment.

Amino Acid Sequence↗

PubMed1990

Death by smothering and its investigation.

This case report provides details from an extensive death investigation, the results of which led to the certification of suicidal manner of death by means of smothering in a pillow, without additional assistance from other persons or mechanical devices. This was the death of a chronically mentally ill person who was under currently approved professional treatment. Major investigative aspects of this case were the death-scene observations of the body, elimination of the possibility of homicide, and the study and analysis of the subject's psychiatric illness. Reasons for the uncommonness of reported deaths in this category are discussed.

Adult↗

PubMed1988

Serum androstanediol glucuronide concentrations in normal and hirsute women and patients with thyroid dysfunction.

There is currently much interest in measurements of 5 alpha-androstane-3 alpha, 17 beta-diol glucuronide (AdiolG) as a marker of peripheral androgen metabolism. We have therefore developed an assay to measure serum AdiolG levels and report that mean concentrations in hirsute (2.9 +/- 1.9 nmol/l, mean +/- SD, n = 15) and non-hirsute (1.9 +/- 0.6 nmol/l, n = 7) women with polycystic ovaries do not differ significantly from concentrations in normal women (2.2 +/- 0.8 nmol/l, n = 20). However, a correlation was found between serum AdiolG levels and Body Mass Index (r = 0.48, P less than 0.05) for women with polycystic ovaries, suggesting that weight may be an important factor in determining concentrations of this steroid conjugate. Serum AdiolG levels were significantly reduced in hypothyroid women (0.6 +/- 0.4 nmol/l, n = 5) and women receiving oral contraceptive therapy (0.6 +/- 0.4 nmol/l, n = 28) but increased in hyperthyroid women (4.0 +/- 0.6 nmol/l, n = 5). The results from this study do not support the hypothesis that serum AdiolG levels provide a marker of peripheral androgen metabolism in hirsute women and show that it is essential to exclude from such investigations any women with thyroid abnormalities or receiving oral contraceptive therapy.

Adult↗

PubMed1988

Serum 11 beta-hydroxyandrostenedione as an indicator of the source of excess androgen production in women with polycystic ovaries.

Serum 11 beta-hydroxyandrostenedione levels (11-OHA) were measured in normal women and women with polycystic ovaries (PCO) to assess their value in localizing the source of excessive androgen production in women with PCO. Serum 11-OHA was undetectable (less than 1.5 nmol/L) in an adrenalectomized woman, a woman with 11-hydroxylase deficiency, and a woman receiving chronic dexamethasone therapy, confirming the specificity of the antiserum used in this study. Serum 11-OHA concentrations were similar in normal women [mean, 5.0 +/- 2.3 (+/- SD) nmol/L] and women with PCO (5.0 +/- 2.1 nmol/L); serum androstenedione concentrations were increased in women with PCO. Thus, the ratio of androstenedione to 11-OHA was significantly higher (P less than 0.001) in women with PCO (2.0 +/- 0.7) than in normal women (1.1 +/- 0.5). Serum 11-OHA levels after adrenal suppression or stimulation were similar in women with PCO who had an ovulatory response and those who failed to ovulate after clomiphene administration. Administration of dexamethasone (1 mg) and injection of ACTH (250 micrograms) were associated with marked suppression and subsequent stimulation of serum 11-OHA levels in both normal women and women with PCO, and the responses were similar in the two groups. Also, the hour to hour and diurnal variations in serum 11-OHA were similar to those of androstenedione and cortisol during a 24-h period, indicating the adrenal origin of 11-OHA. Our finding of similar serum 11-OHA levels in the presence of increased serum androstenedione levels in women with PCO supports the concept that the ovary is the major source of excess androgen production in women with PCO.

Adrenal Glands↗

PubMed1988

Androstenedione concentrations following dexamethasone suppression: correlation with clomiphene responsiveness in women with polycystic ovary syndrome.

It is difficult to predict clomiphene responsiveness in women with polycystic ovary syndrome (PCOS) but it has been suggested that women with evidence of excess adrenal androgen are less likely to respond to clomiphene. To investigate this further we performed a short Synacthen test following overnight dexamethasone suppression, using 11 beta-hydroxyandrostenedione (11-OHA) as a specific marker of adrenal androgen secretion in women with anovulatory infertility due to PCOS (n = 19) compared with a normal group (n = 7). Women with PCOS were subsequently divided into 2 groups according to whether or not they ovulated after clomiphene. On day 1 blood was taken at 9.00 hours for measurement of androstenedione (A), 11-OHA and cortisol, and 1 mg dexamethasone was given at 22.00 hours. On day 2 blood was taken at 9.00 hours and at 30 and 60 minutes after intravenous administration of 250 micrograms Synacthen. Before dexamethasone was given, concentrations of A but not of 11-OHA or cortisol were significantly higher in women with PCOS than in controls but there was no difference in A levels between clomiphene responders and non-responders. After 1 mg dexamethasone had been given, concentrations of A, 11-OHA and cortisol were suppressed in all 3 groups and there were no differences between the groups in the post-dexamethasone concentrations of 11-OHA or cortisol.(ABSTRACT TRUNCATED AT 250 WORDS)

Amenorrhea↗

PubMed1986

The relationship between oestradiol metabolism and adrenal steroids in the endometrium of postmenopausal women with and without endometrial cancer.

The aim of the present study was to investigate the hypothesis that adrenal androgens are contributory to the development of endometrial cancer either by the oestrogenic action of 5-androstene-3 beta, 17 beta-diol (androstenediol) or through the inhibition of oestradiol metabolism. Concentrations of androstenediol, dehydroepiandrosterone (DHA), DHA sulphate (DHAS), oestrone and oestradiol were measured in plasma and endometrium from postmenopausal women with and without endometrial cancer. There was no difference between normal postmenopausal women and endometrial cancer patients with respect to either tissue or plasma adrenal androgens although there was a tendency for plasma DHAS levels to be increased in cancer patients (normal women: 640 +/- 156 ng/ml; cancer patients: 808 +/- 159 ng/ml). There was a positive correlation between endometrial tissue concentrations of androstenediol and DHA in both normal women (P less than 0.05) and cancer patients (P less than 0.01) but for DHAS the relationship was only significant for non-malignant tissue (androstenediol: DHAS, P less than 0.05; DHA: DHAS, P less than 0.02). A significant positive correlation was found between all three plasma adrenal androgens for both groups. In cancer patients there was a trend towards an inverse correlation between endometrial tissue concentrations of DHAS and the enzyme 17 beta-hydroxysteroid dehydrogenase (17OHSD) although the relationship was not significant (r = 0.49). In endometrium, oestradiol was present in significantly higher concentrations than oestrone whereas in plasma the reverse was the case. There was also a tendency for plasma oestradiol levels to be elevated in the cancer subjects. These data do not support a substantial role for adrenal androgens in endometrial cancer but suggest that a relationship may exist between DHAS and 17OHSD and that an imbalance between sulphatase and sulphotransferase activities may be involved.

17-Hydroxysteroid Dehydrogenases↗

PubMed1986

The use of nonradiolabelled steroid infusions to investigate the origin of oestrone sulphate in postmenopausal women.

Infusion of nonradiolabelled dehydroepiandrosterone sulphate (DHA-S) has been used to investigate the possible formation of oestrone sulphate via a sulphated conjugate of androstenedione. The metabolic clearance rate (MCR) of DHA-S also was measured and the mean value (25 1/24h) was similar to values reported using isotopic techniques. Although conversion of DHA-S to 5-androstenediol, a steroid with oestrogenic properties, was detected during infusion of DHA-S, there were no significant increases in plasma levels of conjugated androstenedione or oestrone sulphate. The MCR's oestrone sulphate measured using infusion of nonradiolabelled steroid in two menopausal women were 99 1/24h and 121 1/24h. For one woman, the production rate of oestrone sulphate, calculated from the conversion of oestrone and oestradiol to oestrone sulphate (151 nmol/day) was similar to the measured production rate of oestrone sulphate (144 nmol/day). It is concluded that in menopausal women, oestrone sulphate is derived from conversion of oestrone and oestradiol with no formation occurring via conjugated androstenedione.

Androstenediols↗

PubMed1984

The interrelationship between plasma 5-ene adrenal androgens in normal women.

Plasma concentrations of 5-androstene-3 beta,17 beta-diol (ADIOL) dehydroepiandrosterone (DHA) dehydroepiandrosterone sulphate (DHAS) and cortisol were measured by radioimmunoassay in a group of women aged between 27 and 88 years of age. There was a significant negative correlation with increased age for all three adrenal androgens but not for cortisol. The decrease in adrenal androgens was not related to an excessive divergence from ideal body weight. There was a highly significantly positive correlation between plasma concentrations of all three adrenal androgens which supports a metabolic interrelationship.

Adult↗

PubMed1984

Adrenal androgen concentrations in breast tumours and in normal breast tissue. The relationship to oestradiol metabolism.

Concentrations of ADIOL, DHA and DHAS were measured in human breast tumours and normal tissue from the same breast and related to 17 beta-hydroxysteroid dehydrogenase (17 beta HSD) activity in these tissues. ADIOL and DHA were significantly higher in tumour tissue compared to normal tissue from the same breast (paired t-test: P less than 0.05 and P less than 0.01 respectively) whereas the difference between concentrations of DHAS in normal tissue and tumour tissue was not significant. There was a positive correlation between ADIOL and DHA in both tissues (P less than 0.001) but for DHAS the relationship was only significant in normal tissue (ADIOL:DHAS, P less than 0.001; DHA:DHAS, P less than 0.002). An increase in 17 beta-HSD activity was associated with an increase in DHAS concentrations in both normal and tumour tissue (P less than 0.01 and P less than 0.001 respectively) and with an increase in DHA concentrations in normal tissue (P less than 0.05). These results might be explained by an impairment in the balance between sulphatase and sulphotransferase activity in breast tumours.

17-Hydroxysteroid Dehydrogenases↗

PubMed1984

Adrenal androgen concentrations in endometrium and plasma during the menstrual cycle.

Concentrations of 5-androstene-3 beta, 17 beta-diol (androstenediol), dehydroepiandrosterone (DHA) and DHA sulphate (DHAS) were measured in endometrium and plasma from normal premenopausal and perimenopausal women (average ages 37 and 48 years respectively) at different stages of the menstrual cycle. Plasma levels did not vary with the stage of the cycle for any of the three steroids. Mean plasma levels of androstenediol ranged between 2.03 and 2.92 nmol/l for premenopausal women and 1.38 and 1.58 nmol/l for perimenopausal women while mean concentrations of DHA were 20.80-36.41 nmol/l (premenopausal women) and 13.87-19.07 nmol/l (perimenopausal women). The values for DHAS were more variable and ranged between 3.20 and 4.56 and 2.94 and 4.25 mumol/l for pre- and perimenopausal women respectively. In premenopausal women endometrial tissue concentrations of androstenediol and DHA increased three to fourfold in the secretory phase while no increase was observed in DHAS. There was a similar increase in androstenediol but not DHA or DHAS during the secretory phase for perimenopausal women. A significant positive correlation was found for tissue androstenediol and DHA in both groups of women but the relationship between DHAS and the other androgens was significant only for perimenopausal women. We suggest that the increase in androstenediol and DHA concentrations could be due to an increase in a receptor or binding protein, possibly progesterone dependent, present in secretory phase endometrium.

Adult↗

PubMed1983

Endometrial tissue and plasma concentrations of 5-androstene-3 beta, 17 beta-diol during the menstrual cycle in normal premenopausal and perimenopausal women.

A radioimmunoassay for 5-androstene-3 beta, 17 beta-diol (ADIOL) in human endometrium and plasma is described. The recognised criteria of reliability have been fulfilled. Plasma and endometrial tissue concentrations of ADIOL were determined in samples obtained from normal premenopausal and perimenopausal women (average ages 37 and 48 years respectively) at different phases of the menstrual cycle. In perimenopausal women plasma concentrations of ADIOL did not vary throughout the cycle (proliferative phase: 411 +/- 95 (SEM) pg/ml; secretory phase: 462 +/- 28.5 (SEM) pg/ml). For the premenopausal group the pattern was similar (proliferative phase: 568.4 +/- 56.9 (SEM) pg/ml; secretory phase: 663.1 +/- 64.7 (SEM) pg/ml) although a significant difference (P less than 0.05) was noted between late proliferative and late secretory phase levels in these women. A different pattern was observed for endometrial tissue concentrations of ADIOL. In both groups of women a significant (3-4-fold) increase occurred during the secretory phase. There was no apparent relationship between plasma and tissue concentrations of ADIOL either during the proliferative or the secretory phase. There was, however, an age associated decrease for both tissue and plasma ADIOL. Theories are proposed to account for the increase in ADIOL concentration during the luteal phase.

Adult↗

PubMedDate not supplied

The mucin antigens: what are we measuring?

Serum of breast cancer patients contains high molecular weight, mucin-like glycoproteins which are held to be differentiation markers for certain types of normal epithelia, in particular mammary epithelium. These components have primarily been identified using monoclonal antibodies raised against human milk fat globule membranes, tumour extracts or purified mucins. Even so, many of the antibodies produced react with a discrete region of the mucin protein core involving the hydrophilic turn domain APDTRPAP. The present investigation using the anti-urinary mucin antibody, C595, illustrates both the clinical potential of the mucin antigens in breast cancer studies as well as the exquisite specificity of immune recognition of a complex polymorphic glycoprotein at the level of the individual amino acids.

Amino Acid Sequence↗