Publications by authors named "Christine Kottaridi"

46 Publications

Animal Coronaviruses Induced Apoptosis.

Life (Basel) 2021 Feb 26;11(3). Epub 2021 Feb 26.

Department of Biomedical Sciences, University of West Attica, 12243 Athens, Greece.

Apoptosis is a form of programmed death that has also been observed in cells infected by several viruses. It is considered one of the most critical innate immune mechanisms that limits pathogen proliferation and propagation before the initiation of the adaptive immune response. Recent studies investigating the cellular responses to SARS-CoV and SARS-CoV-2 infection have revealed that coronaviruses can alter cellular homeostasis and promote cell death, providing evidence that the modulation of apoptotic pathways is important for viral replication and propagation. Despite the genetic diversity among different coronavirus clades and the infection of different cell types and several hosts, research studies in animal coronaviruses indicate that apoptosis in host cells is induced by common molecular mechanisms and apoptotic pathways. We summarize and critically review current knowledge on the molecular aspects of cell-death regulation during animal coronaviruses infection and the viral-host interactions to this process. Future research is expected to lead to a better understanding of the regulation of cell death during coronavirus infection. Moreover, investigating the role of viral proteins in this process will help us to identify novel antiviral targets related to apoptotic signaling pathways.
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http://dx.doi.org/10.3390/life11030185DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7996831PMC
February 2021

Erdheim-Chester Disease and Acute Myeloid Leukemia with Mutated in a Patient with Clonal Hematopoiesis: A Case Report.

Onco Targets Ther 2020 16;13:11689-11695. Epub 2020 Nov 16.

2nd Department of Pathology, National and Kapodistrian University of Athens, Medical School, University General Hospital "Attikon", Haidari, Athens, Greece.

Background: Erdheim-Chester Disease (ECD) is a clonal non-Langerhans histiocytosis, classified as a macrophage-dendritic cell neoplasm in the 2016 WHO classification. The exact cell of origin of ECD is unknown, although some limited evidence suggests that it arises from myeloid progenitors.

Case Presentation: A 43-year-old patient, diagnosed with mutated ECD, developed acute myeloid leukemia (AML) with wild-type , 15 months after the initial ECD diagnosis. The patient received intensive chemotherapy plus midostaurin, followed by midostaurin maintenance. Six months into maintenance, the patient remains in complete remission with low-level measurable residual disease, whereas ECD shows a sustained partial metabolic response. Molecular karyotype at several distinct timepoints, namely ECD diagnosis, AML diagnosis, and following treatment of AML, highlighted a molecular signature, indicative of a persistent, underlying clonal hematopoiesis.

Conclusion: This case report suggests that ECD and AML might represent an expansion of two distinct clones in a background of clonal hematopoiesis, indicating their shared origin. Moreover, molecular karyotype might serve as a strong, inexpensive tool for revealing clonal hematopoiesis in cases of negative targeted next-generation sequencing. Finally, the moderate response of ECD to midostaurin suggests that kinase inhibition might have a potential role in ECD treatment.
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http://dx.doi.org/10.2147/OTT.S276497DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7678692PMC
November 2020

Alterations of HPV-Related Biomarkers after Prophylactic HPV Vaccination. A Prospective Pilot Observational Study in Greek Women.

Cancers (Basel) 2020 May 5;12(5). Epub 2020 May 5.

Department of Obstetrics & Gynaecology, University Hospital of Larisa, Mezourlo, 41334 Larisa, Greece.

The objective of this study was to investigate the hypothesis that HPV vaccination administered in patients with low-grade (LG) cytology shortly after an initial colposcopic assessment could prospectively alter HPV-related biomarkers. This was a prospective pilot observational study involving women attending a colposcopy clinic for evaluation of abnormal LG cytology that were advised to undergo HPV vaccination and proceeded accordingly. These women were compared with a matched unvaccinated group. Women requiring cervical biopsies or CIN treatment were excluded.

Intervention: A full three-dose HPV vaccination was undertaken with either the 2-valent or the 4-valent anti-HPV VLP vaccine. LBC samples were obtained prior and after the completion of the vaccination regimen and tested for HPV DNA genotyping (CLART-2 HPV test) and E6 and E7 mRNA (NASBA technique).

Results: Alterations of HPV-related biomarkers at a colposcopy reassessment appointment 12 months later.

Analysis: The -values, relative risk (RR), absolute relative risk (ARR), number needed to treat (NNT) and 95% confidence intervals for each biomarker in each group were assessed.

Results: A total of 309 women were included in the analysis. One hundred fifty-two women received the vaccine. HPV vaccination reduced in a statistically significant manner ( < 0.05) HPV DNA positivity rates for genotypes 16, 18, and 31, RR = 1.6 (95% CI: 1.1 to 2.3), RR = 1.7 (95% CI: 1.1 to 2.8), and RR = 1.8 (95% CI: 1.0 to 2.9), in women who only tested DNA-positive for HPV16, 18, and 31 genotypes, respectively, prior to vaccination. A less pronounced, statistically insignificant reduction was shown for women who tested positive for both HPV DNA and mRNA E6 and E7 expression for HPV16, 18, and 33 subtypes. Statistically significant reduction in HPV mRNA positivity was solely documented for genotype 31 ( = 0.0411).

Conclusions: HPV vaccination appears to significantly affect the rates of HPV16, 18, and 31 DNA-positive infections in the population testing HPV DNA-positive for the aforementioned genotypes. The above findings deserve verification in larger cohorts.
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http://dx.doi.org/10.3390/cancers12051164DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7281708PMC
May 2020

Searching HPV genome for methylation sites involved in molecular progression to cervical precancer.

J Cancer 2019 7;10(19):4588-4595. Epub 2019 Aug 7.

2 nd Department of Pathology, University General Hospital "ATTIKON", School of Medicine, National and Kapodistrian University of Athens, Athens 12464, Greece.

Human Papilloma Virus has been considered as the main cause for cervical cancer. In this study we investigated epigenetic changes and especially methylation of specific sites of HPV genome. The main goal was to correlate methylation status with histological grade as well as to determine its accuracy in predicting the disease severity by establishing optimum methylation cutoffs. : In total, sections from 145 cases genotyped as HPV16 were obtained from formalin- fixed, paraffin-embedded tissue of cervical biopsies, conization or hysterectomy specimens. Highly accurate pyrosequencing of bisulfite converted DNA, was used to quantify the methylation percentages of promoter, enhancer and 5' UTR, CpGs 494, 502, 506 and CpGs 765, 780, 790. The samples were separated in different groupings based on the histological outcome. Statistical analysis was performed by SAS 9.4 for Windows and methylation cutoffs were identified by MATLAB programming language. The most important methylation sites were at the enhancer and especially 7535 and 7553 sites. Specifically for CIN3+ (i.e. HSIL or SCC) discrimination, a balanced sensitivity vs. specificity (68.1%, 66.2% respectively) with positive predictive value (PPV) and negative predictive value (NPV) (66.2%, 68.2% respectively) was achieved for 7535 methylation of 6.1% cutoff with overall accuracy 67.1%, while for 7553 a sensitivity 60.9%, specificity 69.0%, PPV=65.6%, NPV=64.5% and overall accuracy=65.0% at threshold 10.1% was observed. Viral HPV16 genome was found methylated in NF-1 binding sites of in cases with high grade disease. Methylation percentages of and CpG sites were elevated at the cancer group.
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http://dx.doi.org/10.7150/jca.30081DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6746133PMC
August 2019

A prospective study on the epidemiology and clinical significance of viral respiratory infections among pediatric oncology patients.

Pediatr Hematol Oncol 2019 Apr 19;36(3):173-186. Epub 2019 Jun 19.

a Third Department of Pediatrics , National and Kapodistrian University of Athens, "ATTIKON" University Hospital , Athens , Greece.

Respiratory infections in oncology are both common and potentially severe. However, there is still a gap in the literature, regarding the epidemiology of viral respiratory infections in children with cancer. We prospectively enrolled 224 patients, from September 2012 to August 2015. The cohort included children with hematologic or solid malignancies receiving chemotherapy, or undergoing hemopoietic stem cell transplantation, outpatients/inpatients exhibiting signs/symptoms of febrile/afebrile upper/lower respiratory infection. Viral infection was diagnosed by detection of ≥1 viruses from a sample at time of enrollment, using the CLART kit (GENOMICA, Spain). Α detailed questionnaire including demographics and medical history was also completed. Samples were processed in batches, results were communicated as soon as they became available. Children recruited in whom no virus was detected composed the no virus detected group. Viral prevalence was 38.4% in children presenting with respiratory illness. A single virus was found in 30.4%, with RSV being the most frequent. Viral coinfections were detected in 8%. Children with viral infection were more likely to be febrile upon enrollment and to present with lower respiratory signs/symptoms. They had longer duration of illness and they were more likely to receive antibiotics/antifungals. Only 22% of children with influenza received oseltamivir. Mortality was low (2.7%), however, pediatric intensive care unit (PICU) admission and death were correlated with virus detection. In our study mortality was low and PICU admission was related to virus identification. Further research is needed to clarify whether antibiotics in virus-proven infection are of value and underline the importance of oseltamivir's timely administration in influenza.
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http://dx.doi.org/10.1080/08880018.2019.1613462DOI Listing
April 2019

The prognostic value of multiple electrode aggregometry and light transmittance aggregometry in stable cardiovascular patients with type 2 diabetes mellitus.

Thromb Res 2019 08 4;180:47-54. Epub 2019 Jun 4.

Second Cardiology Department, Attikon University Hospital, School of Medicine, National and Kapodistrian University of Athens, Athens, Greece.

Aim: Limited data are available regarding the clinical relevance of platelet function measurements in stable patients with coronary artery disease (CAD). Our aim is to evaluate the agreement between multiple electrode aggregometry (MEA) and light transmittance aggregometry (LTA) in detecting clopidogrel low responders and their prognostic value in CAD patients with type 2 diabetes mellitus (T2DM) on dual platelet inhibition.

Methods: LTA and MEA were performed in 122 stable cardiovascular patients with T2DM. The upper quartile of patients according to maximum LTA (LTAmax) and MEA measurements were defined as clopidogrel low responders. Agreement between the two methods was evaluated by kappa statistics. We assessed the potential correlation between antiplatelet response and clinical outcome and the optimal cutoff value according to ROC analysis to predict the occurrence of major adverse cardiovascular events (MACE), during 1-year follow-up period.

Results: Cohen's kappa coefficients (0.214) indicated fair agreement (70.2%) between LTA and MEA. A total of 25 MACE occurred in 108 patients (23.1%). Patients with MACE had higher LTAmax than those without (57.1 ± 16.5 vs 49.3 ± 18.3, respectively, p = 0.023). MEA measurements were similar between patients with and without MACE (30.1 ± 15.4 vs 30.6 ± 20.8, respectively; p = 0.84). Multiple logistic regression showed LTAmax response as an independent predictor of death from cardiovascular causes (Odds Ratio, adjusted:0.2;0.05-0.81). ROC analysis indicated that LTAmax cutoff of 62.5% best predicted death (AUC = 0.67, sensitivity = 78%, specificity = 61.5%).

Conclusions: The assessment of platelet responsiveness remains highly test-specific. Our results support the prognostic role of LTA, but not MEA testing, for death risk evaluation in stable cardiovascular T2DM patients.
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http://dx.doi.org/10.1016/j.thromres.2019.06.001DOI Listing
August 2019

Mutation Status in Primary, Recurrent, and Metastatic Malignant Melanoma and Its Relation to Histopathological Parameters.

Dermatol Pract Concept 2019 Jan 31;9(1):54-62. Epub 2019 Jan 31.

Second Department of Pathology, National and Kapodistrian University of Athens, School of Medicine, Attikon University Hospital, Athens, Greece.

Background: mutations are a common finding in malignant melanoma (MM). Nevertheless, apart from their significance as a therapeutic target in advanced melanoma, their prognostic value is still debated.

Objective: To assess mutation status in primary, recurrent, or metastatic MM and its correlations with histopathological findings.

Methods: We analyzed 203 samples from 178 consecutive patients: 129 primary cutaneous MM, 49 metastatic and recurrent MM of unknown primary site, and 25 cases of recurrences or metastases of primary MM. mutations in exon 15 were identified with real-time polymerase chain reaction and/or direct sequencing or pyrosequencing. Histopathological examination was performed according to standard procedures.

Results: We observed a 42.1% prevalence of mutations at codon 600 among our patients, 84% of whom harbored the V600E mutation. Mutations showed a statistically significant increase in younger patients (P = 0.011), in ulcerated tumors (P = 0.020), and in tumors lacking solar elastosis in adjacent dermis (P = 0.008). Mutations were also more common in male patients, as well as in primary MMs of the torso, and in nonvisceral metastases, however without reaching statistical significance. Logistic regression analysis identified type and ulceration as the only significant predictors of mutation. The highest frequencies of mutated were identified in superficial spreading and nodular types, and the lowest in acral lentiginous and lentigo maligna types. In situ MM and primary dermal melanoma displayed intermediate frequencies.

Conclusion: Frequency of mutated is type-related and correlated with ulceration, a known adverse prognostic factor.
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http://dx.doi.org/10.5826/dpc.0901a13DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6368075PMC
January 2019

Th17 serum cytokines in relation to laboratory-confirmed respiratory viral infection: A pilot study.

J Med Virol 2019 06 4;91(6):963-971. Epub 2019 Feb 4.

4th Department of Internal Medicine, University of Athens Medical School, Athens, Greece.

Background: Th17 cytokines are associated with modulation of inflammation and may be beneficial in clearing influenza infection in experimental models. The Th17 cytokine profile was evaluated in a pilot study of respiratory virus infections.

Methods: Consecutive patients with symptoms of respiratory tract infection visiting the emergency department of a tertiary care hospital during the winter influenza season of 2014 to 2015 were evaluated. CLART PneumoVir kit, (GENOMICA, Madrid, Spain) was used for viral detection of all known respiratory viruses. Th17 cytokine profile was evaluated with the MILLIPLEX MAP Human TH17 Magnetic Bead Panel (Millipore Corp., Billerica, MA). Correlation of the TH17 profile with viral detection was performed with univariate and multivariate analysis.

Results: Seventy-six patients were evaluated (median age 56 years, 51.3% female); a respiratory virus was identified in 60 (78.9%) patients; 45% had confirmed influenza. Influenza A (H3N2) correlated with higher levels of granulocyte-macrophage colony-stimulating factor (GM-CSF), interleukin 1β (IL-1β), IL-17A, IL-17E, IL-17F, IL-21, IL-22, and IL-23 (P < 0.05 by analysis of variance [ANOVA]) compared with respiratory syncytial virus (RSV). Parainfluenza virus (PIV) similarly had higher levels of GM-CSF, IL-1b, IL-17A, IL-22 compared with those detected in RSV, influenza B and any other virus infection ( P < 0.05; ANOVA). Increasing age (β-coefficient = 1.11, 95% CI, 1.04-1.2, P < 0.01) as well as IL-17A levels (β-coefficient = 1.03, 95% CI, 1.001-1.05, P = 0.04) predicted hospital admission.

Conclusion: Main Th17 cell effector cytokines were upregulated in laboratory-confirmed A(H3N2) influenza and PIV. Excessive amounts of Th17 cytokines may be implicated in the pathogenesis and immune control of acute influenza and PIV infection in humans and may predict the severity of disease.
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http://dx.doi.org/10.1002/jmv.25406DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7166444PMC
June 2019

Comparison between Nageotte and flow cytometric counting of residual leucocytes in freshly prepared leucocyte-reduced red blood cell components.

Transfus Apher Sci 2018 Aug 7;57(4):544-548. Epub 2018 Jun 7.

Laboratory of Haematology & Blood Bank Unit, "Attikon" Hospital, School of Medicine, National and Kapodistrian University of Athens, 1 Rimini Str., 12462, Athens, Greece. Electronic address:

Background: Flow cytometry (FC) and Nageotte hemocytometry represent the most widely accepted methods for counting residual white blood cells (rWBCs) in leucocyte-reduced (LR) blood components. Our aim was to study the agreement between the two methods, under real working blood bank conditions.

Materials And Methods: 94 freshly produced LR red blood cell (RBC) units were tested for rWBC concentrations by FC and Nageotte. To assess the precision of each method, we calculated the intra-assay coefficients of variation (CV), and followed the Bland-Altman methodology to study the agreement between the two methods.

Results: CV was 18.5% and 26.2% for the Nageotte and the FC, respectively. However, the agreement between the duplicate observations, using the binary cut-off threshold of 1 × 10 WBCs per unit to define the results as "pass/fail", was 71.9% for the Nageotte and 93.3% for the FC. Linear regression analysis did not show any correlation (R-squared = 0.01, p = 0.35) between the two methods, while the Bland-Altman analysis for the measuring agreement showed a bias toward a higher Nageotte count of 0.77 × 10 leucocytes per unit (p < 0.001) with the 95% limits of agreement (d ± 2 sd) ranging from -0.40 × 10 to 1.94 × 10 leucocytes per unit.

Conclusion: The absence of agreement between Nageotte and FC method, with the differences within d ± 2 sd being of high clinical importance, suggests that the two methods cannot be used for clinical purposes interchangeably. The Nageotte seems unsuitable for quality control even with a pass-fail criterion, under real working blood bank conditions.
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http://dx.doi.org/10.1016/j.transci.2018.06.002DOI Listing
August 2018

The association between sexually transmitted infections, human papillomavirus, and cervical cytology abnormalities among women in Greece.

Int J Infect Dis 2018 Aug 11;73:72-77. Epub 2018 Jun 11.

Fourth Department of Internal Medicine, 'Attikon' University Hospital, National and Kapodistrian University of Athens, Medical School, Athens, Greece. Electronic address:

Objective: To investigate the diagnosis of sexually transmitted infections (STIs) with human papillomavirus (HPV) infection and the presence of cytological changes in the cervix in a cohort of sexually active women in Greece.

Methods: Cervical cytology testing and the molecular typing of HPV and other STIs were performed for 345 sexually active women aged between 18 and 45 years (mean 33.2±7.2years) visiting a gynaecology clinic for routine cervical screening. The association of HPV and STI detection with cytological findings was investigated.

Results: HPV was detected in 61 women (17.7%) and STIs in 82 (23.8%). Ureaplasma spp was the most frequently detected pathogen, which was found in 63 (18.2%) women, followed by Mycoplasma spp (21 women, 25.6%) and Chlamydia trachomatis (five women, 6.1%). HPV positivity only (with no co-presence of STI) was associated with an abnormal cytology (odds ratio 6.9, p<0.001), while women who were negative for both HPV and STIs had a higher probability of a normal cytology (odds ratio 0.36, p<0.01). Sixteen out of the 63 (25.4%) women who tested positive for Ureaplasma spp, harboured a high-risk HPV type (odds ratio 2.3, p=0.02).

Conclusions: In a population with a high prevalence of Ureaplasma spp, there was an association of this pathogen with high-risk HPV infection, a finding that needs further elucidation.
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http://dx.doi.org/10.1016/j.ijid.2018.06.001DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7128516PMC
August 2018

Neonatal screening for congenital CMV infection stresses the importance of maternal nonprimary infection even in an area where prenatal serology testing is common.

J Matern Fetal Neonatal Med 2019 Jun 27;32(11):1901-1904. Epub 2017 Dec 27.

h Second Department of Pediatrics , National and Kapodistrian Univeristy of Athens Medical School, P&A Kyriakou Childrens' Hospital , Athens , Greece.

Aim And Methods: Dried blood spots from 2149 newborns were examined to diagnose congenital cytomegalovirus (cCMV).

Results: Prenatal CMV-IgG antibodies had been measured during prenatal care in 1287 (60.3%) of mothers and 980 (76.1%) of them were found seropositive. cCMV incidence was 0.47%. All newborns were asymptomatic; 9/10 were born post nonprimary maternal infection; two developed sensorineural hearing loss.

Conclusions: In a country where prenatal CMV testing is common and therefore a false sense of control might prevail, nonprimary maternal infection should not be overlooked. Indeed, women of childbearing age should be educated on CMV prevention measures irrespectively to their serostatus.
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http://dx.doi.org/10.1080/14767058.2017.1416605DOI Listing
June 2019

Mixed viral infections of the respiratory tract; an epidemiological study during consecutive winter seasons.

J Med Virol 2018 04 17;90(4):663-670. Epub 2018 Jan 17.

4th Department of Internal Medicine, University Hospital Attikon, National and Kapodistrian University of Athens Medical School, Athens, Greece.

The current study aimed to describe the molecular epidemiology of mixed respiratory viral infections during consecutive winter seasons in a tertiary care hospital. Patients with symptoms of respiratory tract infection were evaluated during the 2009-2011 and 2013-15 winter seasons. A clinical microarray technique was used for viral detection. Clinical and epidemiological data were correlated with mixed viral detection and the need for hospitalization. In 332 out of 604 (54.4%) evaluated patients (17.6% children) a respiratory virus was identified. Mixed viral infections were diagnosed in 68/332 (20.5%) patients with virus detection (66.2% mixed Influenza-RSV infections). Mixed viral infections were more commonly detected in children (OR 3.7; 95%CI 1.9-5.6, P < 0.01) and patients with comorbidities. In logistic regression analyses, mixed viral infections were associated with younger age (mean age 30.4 years vs. 41.8 years, P ≤ 0.001) and increased rates of fever (OR: 2.7; 95%CI 1.04-7.2, P < 0.05) but no adverse outcomes or increased rates of hospitalization. High rates of mixed viral infections were noted during all winter seasons (especially Influenza and RSV) and were more common in younger patients. The clinical significance of mixed respiratory viral infection needs further elucidation.
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http://dx.doi.org/10.1002/jmv.25006DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7167177PMC
April 2018

Association of p16 (CDKN2A) polymorphisms with the development of HPV16-related precancerous lesions and cervical cancer in the Greek population.

J Med Virol 2018 05 11;90(5):965-971. Epub 2017 Dec 11.

University of Thessaly, School of Health Sciences, Department of Biochemistry & Biotechnology, Microbiology-Virology Laboratory, BIOPOLIS, Larissa, Greece.

The tumor suppressor protein p16 plays a fundamental role in cell cycle regulation and exerts a protective effect against tumor growth. Two different polymorphisms at positions 540 and 580 at the 3'UTR of exon 3 of p16 gene are implicated in several types of cancer, while their role in cervical cancer development remains rather vague. In the present study, we investigated for the impact of p16 genotypes/haplotypes on patients' vulnerability to cervical disease and examined whether these factors can be used as progression markers in the Greek population. A total of 96 HPV16 positive samples and histologically confirmed as LSIL (42 samples), HSIL (44 samples), and cervical cancer cases (10 samples) along with 50 control cases were tested. The identification of p16 polymorphisms was performed by PCR-RFLP methodology. The present analysis revealed that women with p16 540 CG/GG genotype are at a 2.7-fold higher risk of developing HPV16-associated HSIL (OR = 2.7, 95%CI: 1.01-6.6, P = 0.028). The G allele can be regarded as a risk factor of developing HSIL in the Greek population (OR = 2.7, 95%CI: 1.2-5.9, P = 0.012). Moreover, p16 polymorphism C580T is not associated with the growth of cervical lesion in Greek patients, while 540G/580C haplotype can be regarded as a risk haplotype of developing HSIL (OR = 3.67, 95%CI: 1.56-8.6, P = 0.0019). Our results demonstrated that p16 C540G polymorphism influence patients' susceptibility to more severe dysplasia and consequently this polymorphism could potentially emerge as a valuable biomarker for HSIL development in the Greek population.
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http://dx.doi.org/10.1002/jmv.24996DOI Listing
May 2018

Corrigendum: Association of codon 72 polymorphism of p53 with the severity of cervical dysplasia, E6-T350G and HPV16 variant lineages in HPV16-infected women.

J Med Microbiol 2017 10;66(10):1521

Department of Biochemistry and Biotechnology, Microbiology-Virology Laboratory, School of Health Sciences, University of Thessaly, Biopolis, 41500 Larissa, Greece.

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http://dx.doi.org/10.1099/jmm.0.000605DOI Listing
October 2017

Evaluation Analysis of miRNAs Overexpression in Liquid-Based Cytology Endometrial Samples.

J Cancer 2017 22;8(14):2699-2703. Epub 2017 Aug 22.

Department of Cytopathology, University General Hospital "ATTIKON", School of Medicine, National and Kapodistrian University of Athens, Athens 12464, Greece.

miRNAs have an important role as their deregulation is linked to endometrial cancer. : A custom miScript® miRNA PCR Array was used to investigate for the first time the expression of eight miRNAs in forty-nine histologically confirmed Liquid Based cytology endometrial samples. The expression profile of the same miRNAs was also examined in sixty formalin-fixed tissue samples. Expression of seven miRNAs was significantly higher in malignant samples with three of them (mir-182, mir-141 and mir-205) performing optimally. These results suggest the potential use of this non-invasive method of sampling for miRNA expression studies. Furthermore miRNA overexpression could serve as an ancillary or reflex test for optimal identification of malignant samples especially in morphologically inadequate samples.
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http://dx.doi.org/10.7150/jca.19947DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5604201PMC
August 2017

Association of codon 72 polymorphism of p53 with the severity of cervical dysplasia, E6-T350G and HPV16 variant lineages in HPV16-infected women.

J Med Microbiol 2017 Sep;66(9):1358-1365

Department of Biochemistry and Biotechnology, Microbiology-Virology Laboratory, School of Health Sciences, University of Thessaly, Biopolis, 41500 Larissa, Greece.

Polymorphic variability in the tumour-suppressor protein p53 at codon 72 has a considerable impact on cervical cancer development. The present study clarified the association between p53 codon 72 genotypes and the risk of cervical disease in Greek patients. We also examined whether the presence of specific p53 genotypes in combination with HPV16 variants or E6 T350G sequence variation can modify an individual's susceptibility to cervical disease. The analysis of p53 genotypes was performed through PCR-RFLP. Sequence and phylogenetic tree analyses of the HPV16 E6 gene were also performed in order to identify HPV16 variants and T350G sequence variation. The outcomes of the present analysis revealed that women who are homozygous for the arg genotype are at a 4.17-fold higher risk of developing HPV16-associated HSIL+ (OR=4.17, 95 % CI:1.48-4.9, =0.0049). Moreover, p53 arg/arg patients infected by an HPV16 prototype strain were associated with an increased risk of more severe lesions, while a significant relationship between the p53 arg/arg genotype in patients with T350G sequence variation and the risk of high-grade squamous intraepithelial lesions (HSILs) was revealed. The oncogenic potential of the virus is increased by the presence of the p53 arg/arg genotype in the Greek population in such a way that the specific protein interaction E6 (L83V)-p53 (Arg-72) can modify an individual's susceptibility to cervical disease.
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http://dx.doi.org/10.1099/jmm.0.000563DOI Listing
September 2017

Quantitative Measurement of L1 Human Papillomavirus Type 16 Methylation for the Prediction of Preinvasive and Invasive Cervical Disease.

J Infect Dis 2017 03;215(5):764-771

Department of Cytopathology, National and Kapodistrian University of Athens Medical School, "ATTIKON" University Hospital, Athens, Greece.

Background: Methylation of the human papillomavirus (HPV) DNA has been proposed as a novel biomarker. Here, we correlated the mean methylation level of 12 CpG sites within the L1 gene, to the histological grade of cervical precancer and cancer. We assessed whether HPV L1 gene methylation can predict the presence of high-grade disease at histology in women testing positive for HPV16 genotype.

Methods: Pyrosequencing was used for DNA methylation quantification and 145 women were recruited.

Results: We found that the L1 HPV16 mean methylation (±SD) significantly increased with disease severity (cervical intraepithelial neoplasia [CIN] 3, 17.9% [±7.2] vs CIN2, 11.6% [±6.5], P < .001 or vs CIN1, 9.0% [±3.5], P < .001). Mean methylation was a good predictor of CIN3+ cases; the area under the curve was higher for sites 5611 in the prediction of CIN2+ and higher for position 7145 for CIN3+. The evaluation of different methylation thresholds for the prediction of CIN3+ showed that the optimal balance of sensitivity and specificity (75.7% and 77.5%, respectively) and positive and negative predictive values (74.7% and 78.5%, respectively) was achieved for a methylation of 14.0% with overall accuracy of 76.7%.

Conclusions: Elevated methylation level is associated with increased disease severity and has good ability to discriminate HPV16-positive women that have high-grade disease or worse.
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http://dx.doi.org/10.1093/infdis/jiw645DOI Listing
March 2017

γH2Ax Expression as a Potential Biomarker Differentiating between Low and High Grade Cervical Squamous Intraepithelial Lesions (SIL) and High Risk HPV Related SIL.

PLoS One 2017 24;12(1):e0170626. Epub 2017 Jan 24.

2nd Department of Pathology, University Hospital "Attikon", School of Medicine, National and Kapodistrian University of Athens, Athens, Greece.

Background: γH2AX is a protein biomarker for double-stranded DNA breakage; its expression was studied in cervical squamous intraepithelial lesions and carcinomas.

Methods: Immunostaining for phospho-γH2AX was performed in sections from histologically confirmed cervical SIL and carcinomas, as well as from normal cervices used as controls. In total, 275 cases were included in the study: 112 low grade SIL (LGSIL), 99 high grade SIL (HGSIL), 24 squamous cell carcinoma (SCC), 12 adenocarcinoma and 28 cervical specimens with no essential lesions. Correlation of histological grading, high risk vs. low risk HPV virus presence, activated vs. non-activated status (by high risk HPV mRNA expression) and γH2AX expression in both basal and surface segments of the squamous epithelium was performed.

Results: Gradual increase of both basal and surface γH2AX expression was noted up from normal cervices to LGSIL harboring a low risk HPV type, to LGSIL harboring a high risk virus at a non-activated state (p<0.05). Thereafter, both basal and surface γH2AX expression dropped in LGSIL harboring a high risk virus at an activated state and in HGSIL.

Conclusions: γH2AX could serve as a potential biomarker discriminating between LGSIL and HGSIL, as well as between LGSIL harboring high risk HPV at an activated state.
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http://journals.plos.org/plosone/article?id=10.1371/journal.pone.0170626PLOS
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5261776PMC
August 2017

Many children aged two to five years have a persistent presence of respiratory viruses in their nasopharynx.

Acta Paediatr 2016 Feb 8;105(2):e89-92. Epub 2015 Dec 8.

Pediatric Allergy and Respiratory Unit, 3rd Department of Pediatrics, University General Hospital 'Attikon', School of Medicine, National and Kapodistrian University of Athens, Athens, Greece.

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http://dx.doi.org/10.1111/apa.13259DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7159633PMC
February 2016

A Pyrosequencing Assay for the Quantitative Methylation Analysis of GALR1 in Endometrial Samples: Preliminary Results.

Biomed Res Int 2015 4;2015:756359. Epub 2015 Oct 4.

Department of Cytopathology, "ATTIKON" University Hospital, University of Athens Medical School, 1 Rimini, Haidari, 12462 Athens, Greece.

Endometrial cancer is the most common malignancy of the female genital tract while aberrant DNA methylation seems to play a critical role in endometrial carcinogenesis. Galanin's expression has been involved in many cancers. We developed a new pyrosequencing assay that quantifies DNA methylation of galanin's receptor-1 (GALR1). In this study, the preliminary results indicate that pyrosequencing methylation analysis of GALR1 promoter can be a useful ancillary marker to cytology as the histological status can successfully predict. This marker has the potential to lead towards better management of women with endometrial lesions and eventually reduce unnecessary interventions. In addition it can provide early warning for women with negative cytological result.
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http://dx.doi.org/10.1155/2015/756359DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4609388PMC
August 2016

Genital HPV in Children and Adolescents: Does Sexual Activity Make a Difference?

J Pediatr Adolesc Gynecol 2016 Jun 3;29(3):228-33. Epub 2015 Sep 3.

Division of Infectious Diseases, First Department of Pediatrics, University of Athens, Aghia Sophia Children's Hospital, Athens, Greece.

Study Objective: To compare the prevalence of human papillomavirus (HPV) genital infection among prepubertal children, sexually active and not sexually active adolescents, and assess potential risk factors for transmission.

Design: Prospective study.

Setting: Outpatient adolescent health clinic.

Participants: Ninety-five girls aged 2-21 years; 38 sexually active adolescents (group A), 28 not sexually active adolescents (group B), and 29 prepubertal children (group C).

Interventions: Participants' vaginal or cervical specimens were tested for HPV with the CLART HPV 2 assay (Clinical Array Technology, Genomica, Madrid, Spain) and for cytological abnormalities with liquid-based cytology.

Main Outcome Measures: Differences in prevalence of low- and high-risk HPV infections among the 3 groups.

Results: Genital HPV was detected in 37.9% (36/95) of all participants; 47.4% (18/38) of group A, 28.6% (8/28) of group B, and 34.5% (10/29)of group C (P = .27). Multiple HPV infection was detected in 26.3% (10/38), 10.7% (3/28), and 13.8% (4/29) of groups A, B, and C, respectively (P = .21). High-risk genotypes were detected in 47.4% (18/38), 28.6% (8/28), and 24.1% (7/29) of groups A, B, and C, respectively (P = .10). Main high-risk genotypes were HPV 16 (27%, 10/37), HPV 31 (21.6%, 8/37 ), HPV 35 (13.5%, 5/37), HPV 53 (13.5%, 5/37), and low-risk HPV 6 (18.9%, 7/37). Sexual activity was associated with increased risk for genital high-risk HPV infection (odds ratio = 3.41; 95% confidence interval, 1.19-9.78); specifically with HPV 33 and HPV 51. Forty percent of sexually active adolescents with normal cervical cytology were infected with high-risk HPV types. Family history of skin HPV was positively associated with genital HPV in the sexually active group (odds ratio = 2.01; 95% confidence interval, 1.17-3.46).

Conclusion: Timeline and target population for HPV vaccination might need to be reappraised, in view of significant nonsexual transmission of genital HPV so early in childhood.
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http://dx.doi.org/10.1016/j.jpag.2015.08.010DOI Listing
June 2016

Impact of the proton pump inhibitors and CYP2C19*2 polymorphism on platelet response to clopidogrel as assessed by four platelet function assays.

Thromb Res 2013 Aug 2;132(2):e105-11. Epub 2013 Jul 2.

Laboratory of Haematology and Blood Bank Unit, "Attiko" University Hospital, School of Medicine, National and Kapodistrian University of Athens, Athens - Greece. Electronic address:

Background: Previous studies suggested a possible negative interference of proton pump inhibitors (PPIs) on clopidogrel's antiplatelet effect because of the competitive inhibition of the CYP 2C19 isoenzyme. Moreover, carriers of the loss-of-function allele of CYP2C19 polymorphism (CYP2C19*2) display significantly lower responses to clopidogrel. In this study, we investigated the association between CYP2C19*2 genotype, PPI intake and clopidogrel resistance in patients with coronary artery disease (CAD) and their effect on clinical outcome.

Methods: We recruited 95 patients with CAD receiving chronic clopidogrel therapy in combination with aspirin. Platelet reactivity was simultaneously assessed by INNOVANCE PFA-100 P2Y, ADP-induced light transmission aggregometry (LTA), flow-cytometric vasodilator-stimulated phosphoprotein (VASP)-phosphorylation assay and multiple electrode aggregometry (Multiplate). Cardiovascular outcomes were recorded during 1-year follow-up period.

Results: Only platelet reactivity assessed by measuring platelet phosphorylated-VASP demonstrated a significant higher platelet reactivity in carriers of CYP2C19*2 (p=0.023). The other methods displayed higher - but not statistically significant - platelet reactivity in patients carrying the CYP2C19*2 variant as compared with non-carriers. Patients on PPIs demonstrated almost similar suppression of platelet reactivity in comparison with those not treated with PPIs by all platelet function assays. In logistic regression analysis none of the platelet function assays measurements were related with clinical outcomes. Similarly neither CYP2C19*2 genetic variant nor PPI treatment were associated with adverse clinical events.

Conclusions: PPI co-administration did not influence clopidogrel's antiplatelet effect on laboratory testing by all platelet function assays used. On the contrary, patients carrying CYP2C19*2 genotype had significantly higher residual platelet reactivity as estimated by VASP-phosphorylation assay.
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http://dx.doi.org/10.1016/j.thromres.2013.06.015DOI Listing
August 2013

mRNA and DNA detection of human papillomaviruses in women of all ages attending two colposcopy clinics.

PLoS One 2012 15;7(11):e49205. Epub 2012 Nov 15.

Department of Cytopathology, University General Hospital "ATTIKON", School of Medicine, National and Kapodistrian University of Athens, Chaidari, Greece.

Objective: HPV infection is a common finding, especially in young women while the majority of infections are cleared within a short time interval. The aim of this study was to examine the efficacy of HPV DNA and mRNA testing in a population attending colposcopy units of two University hospitals.

Methods: 1173 liquid based cervical samples from two colposcopy clinics were tested for HPV DNA positivity using a commercial typing kit and HPV E6/E7 mRNA positivity with a flow cytometry based commercial kit. Statistic measures were calculated for both molecular tests and morphological cytology and colposcopy diagnosis according to histology results.

Results: HPV DNA, high-risk HPV DNA, HPV16 or 18 DNA and HPV mRNA was detected in 55.5%, 50.6%, 20.1% and 29.7% of the cervical smears respectively. Concordance between the DNA and the mRNA test was 71.6% with their differences being statistically significant. Both tests' positivity increased significantly as lesion grade progressed and both displayed higher positivity rates in samples from women under 30 years old. mRNA testing displayed similar NPV, slightly lower sensitivity but significantly higher specificity and PPV than DNA testing, except only when DNA positivity for either HPV16 or 18 was used.

Conclusions: Overall mRNA testing displayed higher clinical efficacy than DNA testing, either when used as a reflex test or as an ancillary test combined with morphology. Due to enhanced specificity of mRNA testing and its comparable sensitivity in ages under 25 or 30 years old, induction of mRNA testing in young women could be feasible if a randomized trial verifies these results.
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http://journals.plos.org/plosone/article?id=10.1371/journal.pone.0049205PLOS
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3499555PMC
May 2013

Dasatinib inhibits proliferation and induces apoptosis in the KASUMI-1 cell line bearing the t(8;21)(q22;q22) and the N822K c-kit mutation.

Leuk Res 2013 Feb 10;37(2):175-82. Epub 2012 Nov 10.

Second Department of Internal Medicine and Research Institute, Attikon University Hospital, Rimini 1, Haidari, Athens 12462, Greece.

Activating mutations of the c-kit gene are frequently found in CBF (core binding factor) leukemias. We evaluated the effect of tyrosine kinase inhibitor dasatinib in leukemic cell lines bearing or not c-kit mutations. Our data demonstrate that in the AML Kasumi-1 cell line, bearing the N822K c-kit mutation, dasatinib is a potent suppressor of c-kit and Src kinase activity and inhibits the phosphorylation of their downstream target AKT, possibly through the Src-mediated VEGF/VEGFR receptor type 2 pathway. Dasatinib also effectively blocks proliferation and induces apoptosis through caspase-3 activation in Kasumi-1 cells. These data further encourage the integration of dasatinib in the treatment of CBF AML with c-kit mutations in the context of clinical trials, which are eagerly anticipated.
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http://dx.doi.org/10.1016/j.leukres.2012.10.011DOI Listing
February 2013

Identification of women for referral to colposcopy by neural networks: a preliminary study based on LBC and molecular biomarkers.

J Biomed Biotechnol 2012 3;2012:303192. Epub 2012 Oct 3.

Department of Cytopathology, Medical School Attikon University Hospital, University of Athens, 12462 Athens, Greece.

Objective of this study is to investigate the potential of the learning vector quantizer neural network (LVQ-NN) classifier on various diagnostic variables used in the modern cytopathology laboratory and to build an algorithm that may facilitate the classification of individual cases. From all women included in the study, a liquid-based cytology sample was obtained; this was tested via HPV DNA test, E6/E7 HPV mRNA test, and p16 immunostaining. The data were classified by the LVQ-NN into two groups: CIN-2 or worse and CIN-1 or less. Half of the cases were used to train the LVQ-NN; the remaining cases (test set) were used for validation. Out of the 1258 cases, cytology identified correctly 72.90% of the CIN-2 or worst cases and 97.37% of the CIN-1 or less cases, with overall accuracy 94.36%. The application of the LVQ-NN on the test set allowed correct classification for 84.62% of the cases with CIN-2 or worse and 97.64% of the cases with CIN-1 or less, with overall accuracy of 96.03%. The use of the LVQ-NN with cytology and the proposed biomarkers improves significantly the correct classification of cervical precancerous lesions and/or cancer and may facilitate diagnosis and patient management.
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http://dx.doi.org/10.1155/2012/303192DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3470889PMC
January 2013

Evaluation of the role of the new INNOVANCE PFA P2Y test cartridge in detection of clopidogrel resistance.

Platelets 2012 30;23(6):481-9. Epub 2012 May 30.

Laboratory of Haematology and Blood Bank Unit, "Attikon" University Hospital, School of Medicine, National and Kapodistrian University of Athens, Athens, Greece.

Light transmittance aggregometry (LTA) has been extensively used in monitoring clopidogrel therapy. However, the availability of simple and rapid point-of-care platelet function assays is of great clinical importance. Thus, the manufacturer of the Platelet Function Analyzer (PFA)-100 System has recently produced the INNOVANCE PFA P2Y test cartridge. We assessed the ability of this new test to reliably detect clopidogrel resistance. We enrolled 90 consecutive patients with coronary artery disease receiving chronic clopidogrel maintenance therapy in combination with aspirin. Twenty healthy volunteers served as controls. Clopidogrel resistance was simultaneously analysed by the INNOVANCE PFA P2Y test cartridge, ADP-induced LTA, the flow-cytometric vasodilator-stimulated phosphoprotein (VASP)-phosphorylation assay and the multiple electrode aggregometry (Multiplate). Agreement among the four platelet function methods by two was assessed using Cohen's kappa coefficient. According to the cut-off points for clopidogrel resistance proposed by the literature, agreement was fair between INNOVANCE PFA-100 P2Y and LTA (74.4%) and Multiplate (75.6%), while poor agreement was noticed in VASP assay (63.3%). Based on cut-off points indicating a higher thrombotic risk, agreement between the PFA-100 System and the other three methods did not significantly differ compared to the previous cut-offs (72.2%, 71.1% and 55.1%, respectively). The INNOVANCE PFA-100 P2Y test seems to be comparable to other established platelet function assays in detecting clopidogrel resistance. However, the modest agreement among platelet function methods makes the performance of platelet function testing crucial with more than one technique in order to reliably identify poor responders to clopidogrel treatment.
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http://dx.doi.org/10.3109/09537104.2012.689037DOI Listing
December 2012

Clinical performance of human papillomavirus E6, E7 mRNA flow cytometric assay compared to human papillomavirus DNA typing.

Anal Quant Cytol Histol 2011 Dec;33(6):305-10

Department of Cytopathology, Attikon University Hospital, Athens University Medical School, 1 Rimini Street, 12462 Chaidari, Athens, Greece.

Objective: To use flow cytometry to screen cervical samples for the overexpression of human papillomavirus (HPV) E6 and E7 mRNA and compare the performance of this assay with an HPV DNA array for the detection of high-grade cervical lesions.

Study Design: Cervical samples were analyzed for HPV DNA by clinical arrays, and the overexpression of E6 and E7 viral oncogenes was monitored using an HPV mRNA detection kit that quantifies the intracellular HPV E6 and E7 mRNA on a cell-by-cell basis.

Results: HPV positivity increased with severity of histologic lesions. On the basis of histology-confirmed CIN 2+ cases the specificity of HPV assay was 73.9% (95% CI 66.07, 80.88), whereas it was 39.3% (95% CI 31.85, 47.1) for the DNA assay.

Conclusion: The HPV assay provides an early predictor of persistent HPV infection and may improve cervical cancer screening by increasing the specificity of detecting high-grade lesions.
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December 2011

Genetic variability and phylogeny of high risk HPV type 16, 18, 31, 33 and 45 L1 gene in Greek women.

Int J Mol Sci 2012 22;13(1):1-17. Epub 2011 Dec 22.

Department of Cytopathology, Attikon General University Hospital, Athens University Medical School, Chaidari, 12462, Greece; E-Mails: (C.K.N.); (C.K.); (A.C.); (A.S.).

The present study explores nucleotide variability, phylogeny and association with cervical neoplasia in high risk HPV types 16, 18, 31, 33 and 45 collected from Greek women. Of the 1894 women undergoing routine cervical cytology examination, 160 samples test positive for single infections of HPV type 16 (n = 104), HPV 31 (n = 40), HPV 33 (n = 7), HPV 18 (n = 5), and HPV 45 (n = 4) were typed by microarrays method, amplified by PCR then sequenced and phylogenetically analyzed. For HPV 16, 9 variants with nucleotide variations were included into the study. For HPV 31, 33, 18 and 45, nucleotide variations were identified in 6, 4, 2 and 3 variants, respectively. The Bayesian inference and Maximum Parsimony methods were used in order to construct the phylogenetic trees. When types were analyzed independently HPV 16 (European and non-European) and HPV 18 (African and non-African) formed distinct clades. The genomic characterization of HPV variants will be important for illuminating the geographical relatedness and biological differences and for the determination of their risk.
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http://dx.doi.org/10.3390/ijms13010001DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3269669PMC
February 2015

Evaluation of a multiplex real time reverse transcription PCR assay for the detection and quantitation of the most common human rotavirus genotypes.

J Virol Methods 2012 Mar 11;180(1-2):49-53. Epub 2012 Jan 11.

Department of Cytopathology, Attikon University Hospital, Athens University Medical School, Athens, Greece.

Group A rotaviruses (RVs) are important pathogens that cause acute, dehydrating gastroenteritis in infants and young children. In this study, a multiplex real-time polymerase chain reaction protocol using primers and TaqMan(®) probes specific for viral VP4 and VP7 genes was evaluated. This assay offers simultaneous genotyping and quantification of the most common RV genotypes G1P[8], G2P[4], G3P[8], G4P[8], and G9P[8]. It was compared to the molecular typing results provided by conventional PCR. A total of 92 archived stool specimens obtained from children younger than 5 years old with the diagnosis of acute gastroenteritis were examined. Real-time PCR assay detected rotavirus strains among the most common genotype combinations G4P[8] (70.7%), G1P[8] (10.9%), G2P[4] (5.4%), G9P[8] (2.2%). This new assay described has an acceptable sensitivity (low limit 6.3×10(2)copies/g of stool).
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http://dx.doi.org/10.1016/j.jviromet.2011.12.009DOI Listing
March 2012

Cell cycle analysis of colorectal brushings collected in liquid-based cytology.

Anal Quant Cytol Histol 2011 Feb;33(1):29-35

Department of Cytopathology, University General Hospital Attikon, School of Medicine, National and Kapodistrian University of Athens, Athens, Greece.

Objective: To evaluate the possibility of robust cell cycle analysis from liquid-based cytology samples.

Study Design: Brushings were obtained after surgical resection of tumor samples. For each patient, one brushing sample was obtained from the macroscopically identified tumor, and a matched sample from a distant normal-appearing site. Cell suspensions were prepared for a time course series of experiments and from formalin-fixed paraffin-embedded tissue samples.

Results: Optimal results could be obtained when the sample was analyzed on the day of collection, whereas cell cycle analysis could be easily performed when cells were stored in a methanol-water solution for < 3 days, with a small penalty on coefficiency of variation (CV) values. Further storage in liquid-based medium had a profound effect on cell cycle analysis. CV values of liquid-based medium out-performed those obtained from samples that were paraffin embedded. Aneuploidy was more common in men and from samples from the left colon. Near diploid cell populations were more common in samples from the right colon.

Conclusion: Liquid-based cytology fixation and preservation media that are methanol-based could serve as an ancillary collection medium, providing cell cycle data for colorectal carcinoma samples.
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February 2011
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