Biomedical subjects

C L Paul

Publications and source records attributed to C L Paul.

16 recordsLinked to original sources

PubMed1999

Detailed methylation analysis of the glutathione S-transferase pi (GSTP1) gene in prostate cancer.

Glutathione-S-Transferases (GSTs) comprise a family of isoenzymes that provide protection to mammalian cells against electrophilic metabolites of carcinogens and reactive oxygen species. Previous studies have shown that the CpG-rich promoter region of the pi-class gene GSTP1 is methylated at single restriction sites in the majority of prostate cancers. In order to understand the nature of abnormal methylation of the GSTP1 gene in prostate cancer we undertook a detailed analysis of methylation at 131 CpG sites spanning the promoter and body of the gene. Our results show that DNA methylation is not confined to specific CpG sites in the promoter region of the GSTP1 gene but is extensive throughout the CpG island in prostate cancer cells. Furthermore we found that both alleles are abnormally methylated in this region. In normal prostate tissue, the entire CpG island was unmethylated, but extensive methylation was found outside the island in the body of the gene. Loss of GSTP1 expression correlated with DNA methylation of the CpG island in both prostate cancer cell lines and cancer tissues whereas methylation outside the CpG island in normal prostate tissue appeared to have no effect on gene expression.

Alleles↗

PubMed1998

Print material as a public health education tool.

Despite the widespread use of print materials in public health education, little is known about the costs and processes involved in developing these materials and their effectiveness in practice. We examined a sample of printed health education materials, using interviews and checklists. The most cost-effective processes for developing materials were not being used and the effectiveness of materials was rarely evaluated.

Cost-Benefit Analysis↗

PubMed1997

Detection and measurement of PCR bias in quantitative methylation analysis of bisulphite-treated DNA.

Methylation analysis of individual cytosines in genomic DNA can be determined quantitatively by bisulphite treatment and PCR amplification of the target DNA sequence, followed by restriction enzyme digestion or sequencing. Methylated and unmethylated molecules, however, have different sequences after bisulphite conversion. For some sequences this can result in bias during the PCR amplification leading to an inaccurate estimate of methylation. PCR bias is sequence dependent and often strand-specific. This study presents a simple method for detection and measurement of PCR bias for any set of primers, and investigates parameters for overcoming PCR bias.

5-Methylcytosine↗

PubMed1997

Extensive DNA methylation spanning the Rb promoter in retinoblastoma tumors.

The retinoblastoma gene (Rb) is one of the best characterized tumor suppressor genes, and its inactivation is associated with a number of cancers. Previous studies have shown, by restriction enzyme analysis, that the promoter region of the Rb gene is methylated in a significant proportion of primary retinoblastoma tumors. We now report the first detailed methylation sequence analysis of the CpG island spanning the retinoblastoma promoter from hypermethylated retinoblastoma tumors. Our results show methylation is not confined to a specific CpG site, as detected by restriction enzyme studies, but extends to essentially all 27 CpG dinucleotides spanning the retinoblastoma CpG island, including the core promoter. The methylation pattern from each tumor DNA sample is different, ranging from densely to sparsely methylated profiles. Single CpG sites, in particular the E2F transcription factor binding site, as well as blocks of CpGs, were undermethylated in some tumor samples. Possible interference of methylation could be due to the binding of sequence-specific protein factors at these sites in the tumor cells. This study highlights that the dynamics of DNA methylation in cancer cells are clearly different from normal cells and gives an insight into the mechanism of abnormal methylation of CpG islands in cancer cells.

Base Sequence↗

PubMed1997

Prospective, randomized, double-blind study of high-dose aprotinin in pediatric cardiac operations.

BACKGROUND: Perioperative aprotinin decreases postoperative blood loss in adults undergoing cardiac operations, but its role is less clear in children. Therefore, a trial of aprotinin in pediatric cardiac operations was conducted to study the efficacy of its use in children. METHODS: Forty-two patients were randomly assigned to receive either high-dose aprotinin or placebo. Aprotinin efficacy was assessed using time from protamine administration to skin closure, postoperative blood loss and hemoglobin loss, and postoperative transfusion requirements. Measures of fibrinolysis (fibrin degradation product titers) and platelet preservation (beta-thromboglobulin levels) were also assessed. RESULTS: There were no statistically significant differences between groups in any of the blood loss or transfusion parameters. Fibrin degradation product levels, measured 4 hours postoperatively, had increased significantly for control patients, but remained unchanged for the aprotinin group (p < 0.02). beta-Thromboglobulin levels increased more rapidly during cardiopulmonary bypass in the control group (p = 0.03). CONCLUSIONS: Aprotinin appears to provide no clinical benefit in routine pediatric cardiac operations. A reduction in fibrinolysis, with perhaps an early preservation of platelet structure, is seen in the aprotinin group.

Aprotinin↗

PubMed1996

Cytosine methylation: quantitation by automated genomic sequencing and GENESCAN analysis.

Bisulfite treatment and PCR amplification of genomic DNA permits the methylation analysis of any cytosine residue in a target sequence. By cloning and sequencing the PCR product, the methylation of individual molecules can be determined, whereas direct sequencing of the PCR product can provide an average of the methylation status in the population of molecules. Reliable quantitation of cytosine methylation by direct sequencing, however, has not been possible with current methods. In this paper we describe an accurate and innovative protocol to directly quantitate the methylation of any cytosine residue in the target sequence by fluorescence-based automated genomic sequencing. Only the cytosine and thymine residues of bisulfite-treated and amplified genomic DNA are sequenced. The degree of methylation is obtained by direct comparison of the cytosine and thymine signals, which have been labeled with the same fluorescent dyes. GENESCAN analysis is employed to achieve a fast and accurate estimate of methylation at every cytosine in the target sequence. Combining direct bisulfite genomic sequencing and GENESCAN analysis permits the rapid survey of detailed DNA methylation profiles. Using this approach we have found the unexpected result that multicopy plasmid DNA grown in a Dcm host is not always fully methylated as suggested by restriction enzyme data.

Autoanalysis↗

PubMed1994

High sensitivity mapping of methylated cytosines.

An understanding of DNA methylation and its potential role in gene control during development, aging and cancer has been hampered by a lack of sensitive methods which can resolve exact methylation patterns from only small quantities of DNA. We have now developed a genomic sequencing technique which is capable of detecting every methylated cytosine on both strands of any target sequence, using DNA isolated from fewer than 100 cells. In this method, sodium bisulphite is used to convert cytosine residues to uracil residues in single-stranded DNA, under conditions whereby 5-methylcytosine remains non-reactive. The converted DNA is amplified with specific primers and sequenced. All the cytosine residues remaining in the sequence represent previously methylated cytosines in the genome. The work described has defined procedures that maximise the efficiency of denaturation, bisulphite conversion and amplification, to permit methylation mapping of single genes from small amounts of genomic DNA, readily available from germ cells and early developmental stages.

5-Methylcytosine↗

PubMed1994

Renal transplantation at Allegheny General Hospital with special reference to the use of extreme-age cadaveric donors.

We report our experience with 25 living-donor and 427 cadaveric-donor kidneys followed for over 6 years. Patient survival exceeds 90% at 5 years for all recipients. Overall one-year and 5-year graft survival rates were 82.7% and 65.5%, respectively. Approximately one-third of CAD were of extreme age (eg, < or = 10 and > or = 60 years old). One-year and 5-year GS rates of this group were 72.2% and 54.8%, respectively. Kidneys from elderly donors probably should be given to age-matched recipients. Infant en bloc kidneys had good GS. They also adapted well their growth and function to adult recipients and should be used more frequently to alleviate the organ shortage problem. HLA mismatches, transplant PRA level, original kidney disease, and immunosuppressive regimens significantly affected the GS. Race, sex, cytomegaloviral antibody states, unacceptable antigens, length of pretransplant dialysis, and CAD cold ischemic time had no significant effect on GS.

Adolescent↗

PubMed1992

A genomic sequencing protocol that yields a positive display of 5-methylcytosine residues in individual DNA strands.

The modulation of DNA-protein interactions by methylation of protein-binding sites in DNA and the occurrence in genomic imprinting, X chromosome inactivation, and fragile X syndrome of different methylation patterns in DNA of different chromosomal origin have underlined the need to establish methylation patterns in individual strands of particular genomic sequences. We report a genomic sequencing method that provides positive identification of 5-methylcytosine residues and yields strand-specific sequences of individual molecules in genomic DNA. The method utilizes bisulfite-induced modification of genomic DNA, under conditions whereby cytosine is converted to uracil, but 5-methylcytosine remains nonreactive. The sequence under investigation is then amplified by PCR with two sets of strand-specific primers to yield a pair of fragments, one from each strand, in which all uracil and thymine residues have been amplified as thymine and only 5-methylcytosine residues have been amplified as cytosine. The PCR products can be sequenced directly to provide a strand-specific average sequence for the population of molecules or can be cloned and sequenced to provide methylation maps of single DNA molecules. We tested the method by defining the methylation status within single DNA strands of two closely spaced CpG dinucleotides in the promoter of the human kininogen gene. During the analysis, we encountered in sperm DNA an unusual methylation pattern, which suggests that the high methylation level of single-copy sequences in sperm may be locally modulated by binding of protein factors in germ-line cells.

5-Methylcytosine↗

PubMed1992

The cost and availability of devices for preventing childhood injuries.

Accidental injury is the major cause of death among Australian children. Many childhood injuries are preventable through the use of safety devices such as those recommended by the Child Accident Prevention Foundation of Australia (CAPFA). It has been suggested that poor availability and the high cost of obtaining and installing safety devices may contribute to their low rates of use among families with young children. This paper assesses the availability and cost of recommended safety devices in a medium sized city in Australia. Of the 17 devices recommended by the CAPFA only 10 were readily available in Newcastle. The cost of purchasing and installing all the devices recommended by CAPFA amounted to $1516, or 21.4% of the annual disposable income, after the purchase of necessities, of families with children and 75.5% of the annual disposable income, after purchase of necessities, of single parents. It is argued that the cost of safety devices may be prohibitively expensive, particularly for those most at risk of suffering from injury.

Accident Prevention↗

PubMed1970