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Biomedical subjects

C M Thomas

Publications and source records attributed to C M Thomas.

At least 217 records · Page 12Linked to original sources
PubMed1988

Characterization of a human ovarian carcinoma cell line, OTN 14, derived from a mucinous cystadenocarcinoma.

A human ovarian carcinoma cell line, OTN 14, has been established from malignant ascitic fluid of a patient with a well-differentiated mucinous cystadenocarcinoma of the left ovary. The cell line has been maintained in vitro for 6 months through 23 passages, growing in monolayers as well as in 3-dimensional clusters, with a population doubling time of 28 1/2 hr. The number of chromosomes per cell varied from 67 to 88, with a modal number of 86. Two characteristic marker chromosomes were recognized, consisting of partially deleted chromosome I. With a DNA index of 1.934 the tumour cell line was near tetraploid. The epithelial character of the OTN 14 cells was confirmed by a positive immunofluorescence reaction with monoclonal antibodies (MAbs) against different keratins, and when (immuno)electron microscopy was used, keratin filaments and small junctional complexes were observed. Vimentin was also expressed in these cells, while desmin was not detected. Cultured tumour cells reacted (weakly) positive with MAb OV-TL 3 as a marker for ovarian carcinomas, while reactivity with the anti-ovarian carcinoma MAb OC 125 was limited to a few cells, not permitting the detection of shed CA 125 antigen in the culture supernatant. Cells stained heterogeneously positive for CEA marker BW 431/31, the presence of which was confirmed by detection of CEA shed into the culture medium. The cell line released estradiol at a concentration of 130,000 pmol/L in the culture medium, while no progesterone or dehydroepiandrosterone sulphate were found. Electron microscopical evidence for steroid production was suggested in some cells showing "dense-core" vesicles near the Golgi areas. The OTN 14 tumour cells formed poorly differentiated tumour nodules in nude mice, and metastatic cells were also found in blood capillaries. Cell types with mucinous as well as endocrine characteristics were found.

Animals↗
PubMed1988

Gene regulation on broad host range plasmid RK2: identification of three novel operons whose transcription is repressed by both KorA and KorC.

The product of the korA gene of broad host range plasmid RK2 is a key transcriptional repressor which regulates not only the expression of the essential replication gene trfA but also its own expression and that of the kilA operon. It has previously been proposed that korA also encodes a positive activator of transcription of the korC gene, which may act as a transcriptional antiterminator. Here we show that the action of korA in relation to korC can be explained entirely through the korA protein's property as a transcriptional repressor. The limited ability of the previously cloned korC gene to suppress kilC on its own is shown to be due to the fact that korC in RK2 is transcribed from the bla promoter of Tn1 which was deleted in the original korC clones. We demonstrate that korA is a second repressor along with korC of three operons, one of which encodes kilC, the other two not having been described previously and serving an as yet unknown function. We have designated these operons kcrA, B and C for KorC-regulated. Putative kilC is designated kcrC. The homology between the expression signals of these operons suggests that they have arisen by duplication. This is confirmed in the case of kcrA and B by the existence of considerable homology between the products of the first ORFs in each of these operons.

Amino Acid Sequence↗
PubMed1988

5S rRNA genes in Pisum: sequence, long range and chromosomal organization.

We have employed a combination of techniques to examine the organization of pea 5S rRNA genes. These include the analysis of length variant interspersion patterns in cosmid clones, sequence analysis, Southern analysis of both conventional gels and field inversion gels and in situ hybridization. From these analyses we conclude that the 5S rRNA genes of pea are arranged in three major tandem arrays which are represented by three large EcoRI fragments and that these correspond to the three sites of in situ hybridization in the haploid pea complement.

Base Sequence↗
PubMed1988

Inverse relationships between steroid concentration and volume in preovulatory follicles of the golden hamster.

In order to investigate variations in the microenvironment of oocytes within a cohort of maturing follicles the follicular volumes as well as the intrafollicular concentrations of oestradiol (E2) and progesterone (P) were measured in the golden hamster. At 10 h before ovulation the follicular volumes varied from 0.009 to 0.037 mm3 (mean +/- SD: 0.0187 +/- 0.0071 mm3; n = 36). Large follicles (greater than 0.025 mm3; n = 8) contained statistically significantly lower E2 and P levels (30.1 +/- 10.4 and 517 +/- 113 mumol/l, respectively) than the medium sized group (less than 0.025 and greater than 0.015 mm3; n =20): 46.9 +/- 16.0 (P less than 0.02) and 919 +/- 264 (P less than 0.0001) mumol/l, respectively. Small follicles (less than 0.015 mm3) showed the highest steroid levels: 97.0 +/- 33.3 and 1590 +/- 517 mumol/l for E2 and P (P less than 0.001 versus the medium sized group values). Correlation coefficients for the steroid concentrations and the follicular volumes appeared to be -0.674 for E2 and -0.612 for P (P less than 0.001). At the time studied a positive correlation between E2 and P concentrations in the follicles was found: r = 0.655 (P less than 0.001). The mean ratios of intrafollicular over serum steroid concentrations appeared to be approx 36 x 10(3) in the case of E2 and about 17 x 10(3) in the case of P. These results clearly show that there is an inverse relationship between follicular volume and intrafollicular steroid concentrations. The presence of a fine regulatory mechanism for a collective maturation of follicles is hypothesized.

Animals↗
PubMed1988

Clinical applications of monoclonal antibodies against ovarian cancer-associated antigens.

Monoclonal antibodies (McAbs), reactive with tumour-associated antigens (TAAs) present on tumour cells, appear to offer new possibilities in the diagnosis and treatment of cancer (Table I). In addition to these prospects for clinical application, monoclonal antibodies also serve as useful instruments in basic cancer research. The hybridoma technology initiated by Köhler and Milstein in 1975, underwent a very rapid development and has now shown its potential in the field of oncology. This technique made it possible to produce very large quantities of homogeneous antibodies of a stable quality. These McAbs often recognize only one antigenic determinant, or epitope, of cell surface and other molecules. This high specificity is essential for in vivo applications, especially in therapeutic immunotargeting. A central question is whether the antibodies can reach and identify those antigens on ovarian tumour cells that are not shared with normal tissues. Various antibodies have been described in the field of gynaecological oncology, which are assumed to be capable of recognizing such ovarian tumour-related antigens. These McAbs, single or in combination, are capable of showing, unambiguously, the presence of various tumour-associated antigens on ovarian carcinoma cells either in tissue or, when antigen shedding occurs, in blood. However, these McAbs may also react with tumour-associated antigens present on endometrial, cervical, colorectal, breast or other carcinoma cells. The original immunogens used to generate these McAbs differ as to their origin: ovarian cancer cells, breast cancer cells, human milk-fat preparations, trophoblastic cells, endometrial cancer cells have been used as well as osteogenic sarcoma cells, epidermoid carcinoma cells and small-cell lung cancer, colorectal, pancreatic and laryngeal carcinoma cells. The histological distribution patterns of the antigens recognized by these McAbs vary widely: cross-reactions with normal tissue and with carcinomas different from those used as immunogen are frequently seen.

Antibodies, Monoclonal↗
PubMed1988

Human chorionic gonadotrophin measurements in the luteal phase of the menstrual cycle of infertile and fertile women.

Serum human chorionic gonadotrophin (HCG) determinations were performed every other day during the luteal phase of the menstrual cycle of 20 women suffering from unexplained infertility and of 20 women assumed to be fertile. HCG was determined by a specific radioimmunoassay which applied an antiserum directed against the HCG-beta subunit. The group of infertile patients demonstrated detectable serum HCG in only three cases. No clinically confirmed pregnancies were achieved in this group. Eighteen of the 20 women presumed to be fertile showed detectable HCG levels. Five of these achieved ongoing pregnancies, while 13 women revealed increased HCG levels without delayed menstrual periods. The first day of detection of HCG during the luteal phase did not differ between the two groups. In the five women with ongoing pregnancies the time of detection of HCG ranged from day 5 until day 12 after ovulation. It is concluded that the high rate of increased HCG values in the fertile group may represent early conceptual loss, but the apparently disorderly HCG results indicate the need for cautious interpretation of HCG determinations during the luteal phase. Furthermore, early conceptual loss forms no major explanation for unexplained infertility.

Chorionic Gonadotropin↗
PubMed1988

Molecular cloning, identification and transcriptional analysis of genes involved in acetate utilization in Neurospora crassa.

Four Neurospora crassa genomic clones have been selected as hybridizing much more strongly to labelled mRNA isolated from acetate-grown mycelium than to mRNA from sucrose-grown mycelium. Hybridization of restriction fragments with acetate-specific mRNA or cDNA has been used to delimit the transcribed region(s) of each clone. The transcription of all four clones is strongly induced by transfer of growing mycelium from sucrose to acetate as sole carbon source. In wild-type mycelium, mRNAs corresponding to the four clones reach maximum levels after four hours of induction. They accumulate more rapidly and reach higher levels in an acetate non-utilizing mutant, acu-7, which has been found to overproduce enzymes of the glyoxylate cycle and to have a partial block in the TCA cycle. Molecular transformation of a Neurospora acu-5 mutant and of an Aspergillus nidulans acuE mutant by DNA of clone 2 and clone 1, respectively, strongly suggests that clone 2 codes for acetyl-coenzyme A synthetase and that clone 1 codes for malate synthase. The transcribed segments of clones 1 and 2 each hybridize to corresponding clones from Aspergillus nidulans (R. A. Sandeman and M. J. Hynes, personal communication).

Acetates↗
PubMed1988

Endocrine effects of 17 alpha-hydroxyprogesterone caproate during early pregnancy: a double-blind clinical trial.

The clinical and endocrine effects of progestogen therapy in early pregnancy were investigated using a double-blind randomized trial in 64 patients who had a viable fetus at 6 weeks gestation and had an increased risk of miscarriage. The patients were randomly allocated to receive either 17 alpha-hydroxyprogesterone caproate or a placebo between 7 and 12 weeks gestation. Four fetal ultrasonographic variables and 17 maternal endocrine variables were studied in each woman. Only four maternal serum variables (17 alpha-hydroxyprogesterone, prolactin, thyroxin and thyroxin binding globulin) rose significantly. The serum progesterone levels in the hormone supplemented group were on average 20% higher than in the placebo group but the difference was not statistically significant. However, the relation between the progesterone levels and the fetal outcome was not clear. Therefore it is not advisable to prescribe 17-OHP-C during early pregnancy to prevent a miscarriage.

17 alpha-Hydroxyprogesterone Caproate↗
PubMed1988

Endocrine effects of CV 205-502, a new dopamine agonist, in hyperprolactinemic women.

CV 205-502 (CV) is a potent dopaminergic compound which exerts a strong and sustained suppression of prolactin secretion in healthy volunteers. In a prospective double-blind, randomized, placebo-controlled trail, 12 hyperprolactinemic women (greater than or equal to 2000 mU/l), divided into 2 groups of 6 women each, were treated for 4 weeks. Combined pituitary challenge tests (GnRH, TRH, CRH and GHRH) were performed before and after treatment. The 6 CV-treated women (0.05 micrograms daily) showed approximately a 64% decrease of their initial prolactin serum concentrations after 4 weeks of capsule intake. Placebo-treated women showed no change in their prolactin serum level. After TRH administration, a blunted prolactin response was present in all women before treatment. After CV treatment a trend towards normalization of the prolactin response to TRH was seen, whereas the response pattern in the placebo group remained unaltered. The responses of GH, TSH, LH and ACTH to their releasing hormones and cortisol showed no significant changes after administration. FSH, however, showed a significant decrease in response to LH-RH, which could be explained by an increase in estradiol (E2) as ovarian function normalized in CV-treated women. In conclusion, CV shows strong dopamine agonistic properties in hyperprolactinemic women treated with 0.05 micrograms daily. The profile of this new quinoline compound, as judged from the pituitary challenge tests, does not differ from that of dopamine agonists of the ergoline type.

Adult↗
PubMed1988

The urinary excretion of prostaglandins and thromboxane B2 in healthy volunteers: a study in males and females, and on the influence of seminal fluid contamination.

Radioimmunoassay measurements of prostaglandins (PGs) E2, F2 alpha, 6-keto-PGF1 alpha and thromboxane (Tx) B2 in 24 h urine specimens from a male and a female healthy volunteer on several consecutive days revealed a dramatic increase of PGE2, PGF2 alpha, 6-keto-PGF1 alpha on days, upon which they had sexual intercourse; only TxB2 remained stable. Furthermore, the PGE2/PGF2 alpha ratio rose to values greater than 0.5 on days with sexual intercourse. This was found to be due to contamination of the urine samples by seminal fluid. Two 24 h urine samples from each of 26 healthy male and female volunteers (HV) revealed higher (p less than 0.01) mean PGE2 and PGF2 alpha values in males than in females. The results show that the interpretation of the urinary PG excretion as a measure of renal PG synthesis should be considered carefully, and that a PGE2/PGF2 alpha ratio greater than 0.5 indicates probable seminal contamination of urine.

Coitus↗
PubMed1987

Nucleotide sequence of the transcriptional repressor gene korB which plays a key role in regulation of the copy number of broad host range plasmid RK2.

The product of the korB gene of broad host range plasmid RK2 is one of at least two proteins which repress transcription of the essential replication gene trfA. We report here the nucleotide sequence of korB and the properties of its predicted polypeptide product KorB which has a molecular weight of 39,011 Da. KorB is likely to be a soluble protein with an overall net negative charge. However, consistent with a role in transcriptional regulation, there is a region with extensive homology to the alpha helix-turn-alpha helix motif of many DNA binding proteins. This region shows no significant homology to equivalent regions of the TrfB protein which is the primary transcriptional repressor of RK2 and which binds to an operator whose half sites show considerable homology to the half sites of the korB operator.

Amino Acid Sequence↗
PubMed1987

Comparison of the organisation of the genomes of phenotypically diverse plasmids of incompatibility group P: members of the IncP beta sub-group are closely related.

Comparison of physical maps of the broad host range plasmids R751, R906 and R772, belonging to the IncP beta sub-group of the Escherichia coli incompatibility group P, reveals two large regions of similarity, separated by dissimilar regions which contain the majority of the cleavage sites for restriction endonucleases with hexanucleotide recognition sites. Mapping of the regions of these plasmids which show homology to probes specific for genetically characterised segments of the distantly related IncP alpha plasmid RK2, involved in plasmid maintenance or conjugal transfer, reveals that all four plasmids share a similar genetic organisation. In each case the homologous plasmid backbone is interrupted by heterologous segments both between the essential replication loci oriV and trfA, and between the conjugal transfer regions tra1 and tra2, although in the case of R772 the segment of the backbone carrying the trfA and tra2 regions is inverted relative to that of the other plasmids. However, in the case of pJP4, shown to be a fourth member of the IncP beta sub-group, the backbone is interrupted only by a single large segment adjacent to the trfA region. Mapping of the regions of the four IncP beta plasmids which show homology to Tn501 and nucleotide sequence determination at the ends of the homologous regions reveals that R906, R772 and pJP4 share a common mercury resistance region. This region, which appears to have been inactivated in R772, was probably inserted into a common ancestor of these plasmids by the transposition of an element related to an ancestor of Tn501. R751 shows no trace of the mercury resistance region, but contains a short relict of Tn501, derived from an independent insertion event.

DNA Restriction Enzymes↗
PubMed1987

Analysis of transcription from the trfA promoter of broad host range plasmid RK2 in Escherichia coli, Pseudomonas putida, and Pseudomonas aeruginosa.

Reverse transcriptase mapping has been used to analyze transcription from the trfA promoter of broad host range plasmid RK2. The results show that trfA operon mRNA has the same 5' end in Pseudomonas aeruginosa, Pseudomonas putida, and Escherichia coli. The strengths of wild-type and mutant trfA promoters, which differ by defined base substitutions, have been compared and the positions of their transcriptional start sites determined. While these base substitutions do not alter the transcriptional start site, they do have marked effects on promoter strength which are broadly similar in each of the host species. A single base pair substitution, which lies in the region corresponding to the E. coli promoter consensus, brings about a large reduction in gene expression while the introduction of a second mutation, at a locus outside this region, has no further effect on promoter strength. The results indicate that these Pseudomonas species possess an RNA polymerase which recognizes the same region of the trfA promoter as that utilized by E. coli RNA polymerase. Within the limits of these observations it is clear that the trfA operon is transcribed from a single promoter which can function efficiently in diverse species, a property which may be important for its broad host range.

Chromosome Mapping↗
PubMed1987

Pharmacokinetics of intravenous luteinizing hormone-releasing hormone administration in men: effects of various dosages.

Exogenous luteinizing hormone-releasing hormone (LH-RH) was intravenously administered as a single-bolus injection to 26 healthy normal men. LH-RH doses were selected at 1, 2.5, 5, and 20 micrograms for exploration of the optimum LH-RH dose to obtain adequate pituitary stimulation. Blood samples (for LH-RH and LH determinations) were collected at frequent intervals from 10 minutes before to 60 or 90 minutes after injection. LH-RH peak levels varied, in a dose-dependent way, between 119 +/- 16 and 517 +/- 70 ng/l. Peak values were all reached between 1 and 3 minutes after injection, and elimination occurred rapidly, with half-lives between 2.6 +/- 0.4 and 5.2 +/- 1.0 minutes. The area under the curves increased significantly (P less than 0.01) if the doses of LH-RH had been augmented from 1 to 20 micrograms. Maximum LH values were reached more rapidly in the low-dose (1 and 2.5 micrograms) experiments (between 13.5 and 16.3 minutes), with an obvious decline afterwards. The area under the LH curves increased (P less than 0.01) if the doses of LH-RH had been elevated from 1 to 2.5 micrograms, but no further increase of LH release occurred (P greater than or equal to 0.21) if LH-RH doses were further elevated from 2.5 to 20 micrograms. The current study demonstrates that a 2.5-micrograms intravenous LH-RH dose is best suited for an adequate pituitary stimulation in normal men.

Adult↗
PubMed1987

Narrow-host-range IncP plasmid pHH502-1 lacks a complete IncP replication system.

Plasmid pHH502-1 shows incompatibility only towards members of the IncP group, but has a narrower host range than typical members of that group. In contrast to other IncP plasmids its replication was not affected by a high-copy-number plasmid carrying the replication origin (oriV) of IncP plasmid RK2. Southern blotting of pHH502-1 revealed homology to oriV, consistent with its incompatibility phenotype, but no homology to trfA, the essential replication gene of RK2. Thus it is probable that pHH502-1 does not possess a functional IncP replication system, accounting for its restricted host range. A restriction map of pHH502-1 was constructed and the mercury-resistance determinant was localized to Tn735, which also carries the trimethoprim-resistance determinant and is related to Tn21. The presence of a korB-like function on pHH502-1 was also demonstrated.

DNA Transposable Elements↗
PubMed1987

Analysis of mutational lesions of acetate metabolism in Neurospora crassa by 13C nuclear magnetic resonance.

The adaptation of Neurospora crassa mycelium to growth on acetate as the sole carbon source was examined by using 13C nuclear magnetic resonance. Extracts were examined by nuclear magnetic resonance at various times after transfer of the mycelium from medium containing sucrose to medium containing [2-13C]acetate as the sole carbon source. The label was initially seen to enter the alanine, glutamate, and glutamine pools, and after 6 h 13C-enriched trehalose was evident, indicating that gluconeogenesis was occurring. Analysis of the isotopomer ratios in the alanine and glutamate-glutamine pools indicated that substantial glyoxylate cycle activity became evident between 2 and 4 h after transfer. Immediately after transfer of the mycelium to acetate medium, the alanine pool increased to about four times its previous level, only a small fraction of which was enriched with 13C. The quantity of 13C-enriched alanine remained almost constant between 2 and 7.5 h after the transfer, whereas the overall alanine pool decreased to its original level. The selective catabolism of the unenriched alanine leads us to suggest that the alanine pool is partitioned into two compartments during adaptation. Two acetate-nonutilizing mutants were also studied by this technique. An acu-3 strain, deficient for isocitrate lyase (EC 4.1.3.1) activity, showed metabolic changes consistent with this lesion. An acp strain, previously thought to be deficient in an inducible acetate permease, took up [2-13C]acetate but showed no evidence of glyoxylate cycle activity despite synthesizing the necessary enzymes; the lesion was therefore reinterpreted.

Acetates↗