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John J O'Leary

Publications and source records attributed to John J O'Leary.

13 recordsLinked to original sources
PubMed2006

Effect of ret/PTC 1 rearrangement on transcription and post-transcriptional regulation in a papillary thyroid carcinoma model.

BACKGROUND: microRNAs (miRNAs) are a group of non-coding single stranded RNAs measuring approximately 22 nt in length that have been found to control cell growth, differentiation and apoptosis. miRNAs negatively regulate their target genes and recently have been implicated in tumourigenesis. Furthermore, miRNA expression profiling correlates with various cancers, with these genes thought to act as both tumour suppressors and oncogenes. ret/PTC 1 is an oncogene with constitutive kinase activity implicated in the development of papillary thyroid carcinoma (PTC). This rearrangement leads to aberrant MAPK activation that is implicated in PTC tumourigenesis. AIM: The aim of this study was to identify the effect that ret/PTC 1 has on transcription and post-transcriptional regulation in PTC by using DNA microarray and microRNA analysis. RESULTS: DNA microarray analysis revealed a group of genes differentially expressed between normal thyroid cell lines and those harbouring a ret/PTC 1 rearrangement.Furthermore, a unique miRNA expression signature differentiated between PTC cell lines with ret/PTC 1 and a normal thyroid cell line. 21 miRNAs showed significant overexpression and 14 miRNAs showed underexpression in these cell lines when compared to normal thyroid. Several of these up/down regulated miRNAs may be involved in PTC pathogenesis.

Biomarkers, Tumor↗
PubMed2006

Distinct array comparative genomic hybridization profiles in oral squamous cell carcinoma occurring in young patients.

BACKGROUND: Oral cancer typically affects smokers older than 50 years of age. Recently, however, a marked increase in the number of patients 40 years old and younger, many with no history of tobacco smoking, has been noted. Studies in this age group have so far been restricted to genomic areas well recognized as abnormal in typical patients with oral cancer. The aim of this study was to assess genomic aberrations in oral cancer, using comparative genomic hybridization (CGH) microarray technology, and to compare the genomic aberration profile of patients older than 40 years old with those 40 years old and younger. METHODS: Tumor samples from 20 patients with oral cancer (age range, 21-78; 10 smokers and 10 nonsmokers) were laser microdissected, and array CGH was used to identify genomic imbalances in these two cohorts. RESULTS: The older cohort showed high numbers of gains and losses in contrast to very few copy number changes in the younger nonsmoker cohort. In concurrence with the literature, tumors from the older cohort manifested deletions involving 3p and 9p21 and gains involving 3q, 5q, 7p, 8q, 11q, and 20q. The younger group, particularly the nonsmokers, showed very few changes overall, and the aberrations were not in the sites classically associated with oral cancer. Deletion of CDKN2A (p16) was completely absent in the younger group but was present in 50% of the older cohort. CONCLUSIONS: We have demonstrated that there is far less genomic instability in young nonsmokers with oral cancer than found in typical patients with oral cancer. These observations indicate that oral cancer presenting at a younger age, particularly in nonsmokers, has a genomic profile different from the classically described oral cancer.

Adult↗
PubMed2006

Real-time reverse transcription-PCR for detection of rotavirus and adenovirus as causative agents of acute viral gastroenteritis in children.

Viral pathogens are the most common cause of gastroenteritis in developed countries. Human rotavirus and adenovirus infections are major causes of acute outbreaks and sporadic cases of gastroenteritis, occurring primarily among children less than 2 years of age. Patient hospitalization is often required, with enormous infection control implications. This work describes the development of real-time PCR assays for the detection of group F adenovirus, rotavirus A, and rotavirus C from stool specimens. Two hundred twenty stool samples from pediatric patients exhibiting symptoms of diarrhea and/or vomiting were examined. PCR results were compared with those of virus detection by electron microscopy and latex agglutination antigen detection. The incorporation of an internal-control RNA that was spiked into individual stool extracts functioned as an internal validation for the reporting of PCR-negative results. Rotavirus C was not detected by real-time PCR in the patient stool samples examined. Real-time reverse transcription-PCR resulted in 175% and 111% increases in the rates of detection of adenovirus F and rotavirus A, respectively, compared with latex agglutination testing. Molecular detection increased the number of stool specimens in which causative agents of gastroenteritis were identified by 155% compared to electron microscopy. Genotyping of a proportion of the rotavirus and adenovirus strains identified only genotype G1 rotavirus and both adenovirus genotypes 40 and 41 in circulation within the patient cohort examined. The results highlight the significance of rapid molecular methods for the routine screening of stool samples in hospital laboratories to provide rapid definitive diagnoses.

Acute Disease↗
PubMed2006

Molecular profiling of cervical neoplasia.

Cervical cancer, a potentially preventable disease, remains the second most common malignancy in women worldwide. Human papillomavirus is the single most important etiological agent in cervical cancer, contributing to neoplastic progression through the action of viral oncoproteins, mainly E6 and E7, which interfere with critical cell cycle pathways, p53 and retinoblastoma. However, evidence suggests that human papillomavirus infection alone is insufficient to induce malignant changes and that other host genetic variations are important in the development of cervical cancer. This article will discuss the latest molecular profiling techniques available and review the published literature relating to their role in the diagnosis and management of cervical dysplasia and cancer. It is hoped that these techniques will allow the detection of novel biomarkers at DNA, RNA, microRNA and protein levels, which may ultimately play a role in facilitating early disease diagnosis and in predicting response to therapies, thus allowing the development of personalized treatment strategies.

Biomarkers, Tumor↗
PubMed2005

Oestrogen regulated gene expression in normal and malignant endometrial tissue.

OBJECTIVES: The aim of this study was to examine the expression of oestrogen regulated genes in premenopausal and postmenopausal normal and malignant endometrial specimens. The molecular mechanisms and the role of these genes in endometrial carcinogenesis are poorly understood. METHODS: Normal and malignant endometrial specimens were collected from patients undergoing hysterectomy. Real time TaqMan PCR was used to examine the mRNA expression levels of oestrogen receptor a (ERa) and b (ERb), progesterone receptor (PR), insulin like growth factor 1 (IGF-1) and vascular endothelial growth factor (VEGF). RESULTS: Expression analysis was carried out on 60 patients. ERa was more predominantly expressed in the endometrial samples than ERb, 28% of the specimens did not express ER. Normal pre and postmenopausal tissue expressed higher levels of ERa, PR and IGF-1 than malignant tissue. ERa and PR expression was significantly higher in the proliferative phase endometrium compared to the secretory phase (P < 0.05). PR mRNA expression was significantly correlated with ERa in all tissue types. CONCLUSIONS: ERa expression may play an important role in the regulation of PR in normal and malignant endometrium. Further work is needed to establish if IGF-1 plays a role in a subset of endometrial cancers and if isoforms of VEGF play a role in endometrial cancer.

Adenocarcinoma↗
PubMed2005

Quantitation of CDC6 and MCM5 mRNA in cervical intraepithelial neoplasia and invasive squamous cell carcinoma of the cervix.

CDC6 and MCM5 play essential roles in eukaryotic DNA replication. Several studies have highlighted the potential of these proteins as molecular markers of dysplastic and malignant cells in histopathological diagnosis. The mode of expression of CDC6 and MCM5 mRNA and their significance in normal, dysplastic and malignant cervical cells remains to be elucidated. Using a quantitative real-time RT PCR assay, we compared CDC6 and MCM5 mRNA expression in normal cervical epithelium, cervical intraepithelial neoplasia and invasive squamous cell carcinoma of the cervix. Our study cohort comprised 20 normal cervical biopsies, 20 CIN3 and eight invasive squamous cell carcinomas. All samples were formalin fixed and paraffin embedded. Total RNA was extracted and analysed for expression of GAPDH, CDC6 and MCM5 using real-time quantitative TaqMan RT-PCR. A linear increase in MCM5 and CDC6 mRNA expression is observed in normal cervix, CIN3 and invasive cervical carcinoma. The overall difference in MCM5 mRNA expression in the normal cervix, CIN3 and invasive cohort groups is highly statistically significant (P=0.001). An increase in CDC6 mRNA expression in CIN3 and invasive cervical squamous cell carcinoma was observed; however, the overall difference between cohort groups was not found to be statistically significant (P=0.104). Increased transcription of MCM5 and CDC6 occurs as a consequence of cervical neoplastic progression. This pattern of increased mRNA expression in CIN3 and invasive cervical carcinoma directly correlates with findings at the phenotypic protein expression level. This study further confirms the importance of MCM5 and CDC6 in malignant transformation and in the pathogenesis of cervical dysplasia.

Carcinoma, Squamous Cell↗
PubMed2005

Analysis of DNA in endometrial cancer cells treated with phyto-estrogenic compounds using comparative genomic hybridisation microarrays.

The aim of this study was to identify genomic aberrations in endometrial cancer cells treated with the phyto-estrogenic compounds tectorigenin, irigenin and apigenin and to compare with those treated with beta-estradiol using array-based comparative genomic hybridisation (array CGH). The microarray contains 287 targets and includes telomeres, microdeletions, oncogenes and tumour suppressor genes and has increased mapping resolution compared to conventional CGH. An endometrial cancer cell line (Ishikawa) was cultured and treated with the phyto-estrogens. Treated cells were examined using the CGH microarray. Over 20 % of the array genes were aberrated in the cells treated with beta-estradiol, tectorigenin and irigenin compared to 3 % in those treated with the same concentration of apigenin. Protein kinase c zeta form, insulin, insulin receptor and protein-tyrosine phosphatase non-receptor-type 1 which are involved in insulin metabolism were aberrated by tectorigenin and irigenin. Apigenin may play a role in the treatment of endometrial cancer and in the treatment of postmenopausal women. Further studies in normal endometrium and primary endometrial cancer cells are needed to elucidate the role of the phyto-estrogens.

Apigenin↗
PubMed2005

Womb with a view.

Zygote transfer is a significant limiting factor in pregnancy rates after Assisted Reproductive Technology (ART) treatment. Adequate technique appraisal should be a high priority task for any medical facility that offers this treatment. Although many clinicians have extensive personal experience, it is difficult for trainees to understand the anatomical and mechanical process of safe and adequate placing of zygotes in the uterine cavity. Ultimately, even with ultrasound help, it is a "blind procedure". The experimental module described here allows for direct visualisation of the mechanical passage of a catheter through the cervix and final placement of the tip in relation to the intravaginal visible part of the catheter. It has worldwide practical applicability for trainees and experienced clinicians accompanying the theory and allowing adequate training and retraining of all staff performing zygote transfers.

Education, Medical↗
PubMed2005

Increased fetal RhD gene in the maternal circulation in early pregnancy is associated with an increased risk of pre-eclampsia.

OBJECTIVE: To determine whether the fetal RhD gene is present in the maternal circulation in early pregnancy prior to the clinical manifestation of pre-eclampsia. DESIGN: This is a nested case-control study. SETTING: Blood samples were obtained from patients attending for a first antenatal visit. SAMPLE: Cases were asymptomatic RhD negative women (n= 23) who subsequently developed pre-eclampsia matched to RhD negative controls (n= 23) for parity and gestational age. METHODS: Real time PCR using TaqMan primers and probes directed against the RhD gene quantified fetal DNA in the maternal circulation. MAIN OUTCOME MEASURES: Quantity of RhD gene detected. RESULTS: As the copy number of RhD gene per millilitre of whole blood at 15 weeks of gestation increased, there was a significantly increased risk of developing pre-eclampsia. There was a graded association between copy number of RhD gene in early pregnancy and severity of disease with controls having 6942, mild pre-eclamptics 83,273 and severe pre-eclamptics 285,793 copies/mL (logscale 3.6, 4.0 and 4.5, respectively). CONCLUSION: Increased fetal RhD gene is present in the maternal circulation in early pregnancy in women who subsequently develop pre-eclampsia and there appears to be a graded response between the quantity of fetal DNA and severity of pre-eclampsia.

Adult↗
PubMed2004

Detection and quantitation of HER-2 gene amplification and protein expression in breast carcinoma.

We compared fluorescence in situ hybridization (FISH), immunohistochemical analysis, immunocytochemical analysis, and relative quantification assays using polymerase chain reaction (PCR) as methods for estimating HER-2 gene amplification and protein overexpression infine-needle aspirate (FNA) specimens and paraffin-embedded tissue samples from 49 cases of breast cancer: FISH can be performed successfully on FNA smears. Immunohistochemical and immunocytochemical staining intensity of 3+ corresponds to a FISH ratio of more than 2.5. Immunohistochemical and immunocytochemical staining of 2+ and 1+ are not necessarily associated with gene amplification. Increased DNA PCR ratios might be seen without amplification, reflecting polysomy. HER-2 messenger RNA relative quantitation scores correlate well with HER-2 gene amplification. Owing to the ease with which it can be performed and interpreted, we conclude that FISH is the test of choice for HER-2 estimation and, when possible, should be performed on whole nuclei, which are readily available in FNA smears or imprint cytology. FISH may be used primarily or to confirm immunohistochemical, immunocytochemical, and PCR results.

Biomarkers, Tumor↗
PubMed2003

Detection of fetal Rhesus D gene in whole blood of women booking for routine antenatal care.

OBJECTIVE: To determine if the RhD status of the fetus can be detected in an unselected group of RhD-negative women with ongoing pregnancies booking for routine antenatal care. MATERIAL AND METHODS: We obtained 2.5 ml of whole blood from 202 unselected women with a normal ongoing pregnancy who booked for antenatal care before 20 weeks' gestation. DNA was extracted and real time quantitative PCR performed. Primers and a specific probe were targeted at the Rhesus D gene. RESULTS: Of 194 women, 31 were RhD-negative with no evidence of prior isoimmunisation. They delivered 17 RhD-positive and 14 RhD-negative babies. Of the 17 RhD-negative women carrying a RhD-positive fetus, the RhD gene was detected in the maternal blood in 14 (82%). There were no false positives. The quantity of RhD gene detected in the maternal blood of RhD-negative mothers ranged from 2 x 10(2) to 4 x 10(6) copies/ml of whole blood. CONCLUSION: Using real time quantitative PCR assays to amplify free fetal DNA in the maternal circulation, identification of RhD-positive fetuses in RhD-negative mothers is feasible when they book for antenatal care. Before such RhD genotyping can be introduced into clinical practice, however, further studies are required to show that false negative results can be eliminated and to show that this diagnostic test is reliable outside a research setting.

DNA↗
PubMed2003

Fetal DNA quantitation in peripheral blood is not useful as a marker of disease severity in women with preeclampsia.

OBJECTIVE: Trophoblast migration into the maternal circulation is increased in preeclampsia and can trigger the endothelial dysfunction that characterizes the clinical disease. We hypothesised that 'fetal trafficking' is increased in women with severe preeclampsia and that the quantity of trafficking is greater in women with more severe disease. STUDY DESIGN: To test the hypothesis, we used a technique that quantifies a genetic marker specific for the fetomaternal unit, that is, the SRY gene in a pregnant woman carrying a male fetus. Thirty two women with pre-eclampsia and 32 control women (women without preeclampsia) were recruited. Preeclampsia was defined according to the International Society for the Study of Hypertension in Pregnancy (ISSHP). All subjects with preeclampsia had evidence of the multisystemic nature of the disease. DNA was extracted from maternal peripheral blood and RTQ PCR analysis was performed to quantify the fetal DNA (SRY) and the total DNA (beta-actin) in each sample. The ratio of fetal to total DNA was calculated and compared between women with preeclampsia and controls. RESULTS: The women with preeclampsia and the control women did not differ in parity, blood pressure at booking, and gestational age at sampling. The groups differed significantly in age (29 +/- 5.7 vs 25 +/- 5.1 years; P =.007), diastolic blood pressure (DBP) at sampling (101 +/- 9.5 vs 70 +/- 5.5 mm Hg; P <.0001), gestational age at delivery (33 +/- 4.3 vs 39 +/- 1.8 weeks; P <.001), and fetal weight (1.98 +/- 1 vs 3.35 +/- 0.5 kgs; P <.0001). SRY was detected in 31 out of 32 women with preeclampsia and in 24 out of 32 control women (P <.001). The median SRY copy number per micro L was greater in women with preeclampsia (10.6, interquartile range 12.89) than in the control women (8.6, interquartile range 20.1) but these differences were not statistically significant at P =.75. The median ratio of fetal to total DNA was almost identical in both groups (0.06, interquartile range 0.13) in PET compared to (0.06, interquartile range 0.17) the control women. No correlation was found between the quantity of fetal DNA and disease severity. CONCLUSION: Fetal trafficking is more likely to be detected in women with preeclampsia compared to control women but the quantity does not appear to correlate with disease severity.

Adult↗