[Heredity and cancer]. / Hérédité et cancer.

A personal or family history of cancer has now become the primary cause of genetic consultations. In recent years, various genes have been identified that are associated with a more or less marked genetic predisposition to the development of cancers. The syndrome associated with the hereditary risk of breast and ovarian cancer and the Lynch syndrome are the most frequent ones, but there are many other, much less common, situations associated with familial cancer risk. In most cases, there are clear recommendations regarding the indications for genetic testing and the follow-up of patients identified as having a predisposition to cancer. At the CHU of Liège, we currently perform more than 1.400 oncogenetic consultations per year and we maintain a positivity rate of genetic tests performed in this indication higher than 10%. In this way, we allow a multidisciplinary care of patients with a high oncological risk and participate in a prevention and surveillance activity. We also pay increasing attention to the hereditary risk associated with pediatric cancers and to patients with multiple cancers, especially when these develop at an early age. Finally, the oncogenetic consultation must consider the psychological, ethical and legal aspects of a diagnosis that involves the patient and his or her future, but also the whole family.

Une histoire personnelle ou familiale de cancer est aujourd'hui devenue la première cause de consultation en génétique. Au cours de ces dernières années, en effet, différents gènes associés à une prédisposition génétique plus ou moins marquée au développement de pathologies cancéreuses ont été identifiés. Le syndrome de prédisposition héréditaire au cancer du sein et de l'ovaire (HBOC : Hereditary Breast and/or Ovarian Cancer) et le syndrome de Lynch sont les plus fréquents. Mais il existe une multitude d'autres situations beaucoup moins fréquentes associées à un risque familial de cancer. Dans la plupart des cas, des recommandations claires existent quant à l'indication des tests génétiques et quant au suivi proposé au patient chez qui une prédisposition au cancer est identifiée. Au CHU de Liège, nous réalisons actuellement plus de 1.400 consultations d'oncogénétique par an et nous maintenons un taux de positivité des tests génétiques réalisés dans cette indication supérieur à 10 %. De cette façon, nous permettons une prise en charge multidisciplinaire des patients avec un haut risque oncologique et participons à une activité de prévention et de surveillance. Nous portons une attention croissante aux enfants et adolescents présentant un cancer, d'une part, et aux adultes ayant présenté de multiples cancers, d'autre part, particulièrement lorsque ceux-ci se développent à un âge précoce. Enfin, la consultation d'oncogénétique doit tenir compte des aspects psychologiques, éthiques et légaux, liés à un diagnostic qui implique le patient et son avenir, mais également l'ensemble de la famille.

[Hereditary diffuse gastric cancer : case serie of 8 patients from a single family and literature review]. / Cancer gastrique diffus héréditaire. Série de 8 patients appartenant à une même famille et revue de la littérature.

Hereditary diffuse gastric cancer is a form of gastric cancer associated, in about 40 % of cases, with a germline mutation of the CDH1 gene. The management of patients with a pathogenic mutation of this gene is based on total prophylactic gastrectomy because, until proven otherwise, endoscopic monitoring is insufficient. We report a series of eight patients with pathogenic CDH1 mutation who underwent total prophylactic gastrectomy in our centre.

Le cancer gastrique diffus héréditaire est une forme de cancer gastrique associé, dans 40 % des cas environ, à une mutation germinale du gène CDH1. La prise en charge des patients porteurs d'une mutation pathogène de ce gène repose sur la gastrectomie totale prophylactique car, jusqu'à preuve du contraire, la surveillance endoscopique est insuffisante. Nous rapportons une série de huit patients porteurs d'une mutation pathogène de CDH1 ayant bénéficié d'une gastrectomie totale prophylactique dans notre centre. Mots-clés : Gastrectomie prophylactique - Cancer gastrique diffus héréditaire - Mutation CDH1.

[How to avoid overmedicalization of the elderly patient ?] / Comment éviter la surmédicalisation du patient âgé ?

Over-medicalization is a broad concept, which also concerns the elderly patient. It encompasses both over-diagnosis and over-treatment. An increasing awareness of this issue has emerged since 2013, with the first " Preventing Overdiagnosis " conference. Currently, Evidence-Based Medicine does not prevent over-diagnosis. Indeed, the presence of geriatric characteristics such as multiple comorbidities, polypharmacy and frailty can lead to misdiagnosis and to potentially deleterious treatment. Subclinical hypothyroidism and Alzheimer's disease are two examples of pitfalls in the interpretation of biological and para-clinical data that may lead to the administration of useless treatment. Different issues are discussed to identify the causes of over-medicalization and to better prevent it.

La surmédicalisation est un concept large, qui concerne également le patient âgé. Elle englobe à la fois le surdiagnostic et sa conséquence à savoir le surtraitement. Une sensibilisation à ce sujet a émergé depuis 2013, date du premier congrès " Preventing Overdiagnosis ". Actuellement, l'Evidence-Based Medicine ne permet pas d'éviter le surdiagnostic chez le patient âgé. En effet, la présence de caracté- ristiques gériatriques telles que les multiples comorbidités, la polymédication et la fragilité peut mener à l'élaboration d'un diagnostic erroné et à l'instauration d'un traitement potentiellement délétère. L'hypothyroïdie subclinique et la maladie d'Alzheimer sont deux exemples de pièges potentiels à l'interprétation de données biologiques et paracliniques pouvant mener à l'administration d'un traitement futile. Différentes pistes sont abordées pour identifier les causes de la surmédicalisation et mieux la prévenir.

Genetic assessment and folate receptor autoantibodies in infantile-onset cerebral folate deficiency (CFD) syndrome.

INTRODUCTION: Cerebral folate deficiency (CFD) syndromes are defined as neuro-psychiatric conditions with low CSF folate and attributed to different causes such as autoantibodies against the folate receptor-alpha (FR) protein that can block folate transport across the choroid plexus, FOLR1 gene mutations or mitochondrial disorders. High-dose folinic acid treatment restores many neurologic deficits. STUDY AIMS AND METHODS: Among 36 patients from 33 families the infantile-onset CFD syndrome was diagnosed based on typical clinical features and low CSF folate. All parents were healthy. Three families had 2 affected siblings, while parents from 4 families were first cousins. We analysed serum FR autoantibodies and the FOLR1 and FOLR2 genes. Among three consanguineous families homozygosity mapping attempted to identify a monogenetic cause. Whole exome sequencing (WES) was performed in the fourth consanguineous family, where two siblings also suffered from polyneuropathy as an atypical finding. RESULTS: Boys (72%) outnumbered girls (28%). Most patients (89%) had serum FR autoantibodies fluctuating over 5-6 weeks. Two children had a genetic FOLR1 variant without pathological significance. Homozygosity mapping failed to detect a single autosomal recessive gene. WES revealed an autosomal recessive polynucleotide kinase 3´phosphatase (PNKP) gene abnormality in the siblings with polyneuropathy. DISCUSSION: Infantile-onset CFD was characterized by serum FR autoantibodies as its predominant pathology whereas pathogenic FOLR1 gene mutations were absent. Homozygosity mapping excluded autosomal recessive inheritance of any single responsible gene. WES in one consanguineous family identified a PNKP gene abnormality that explained the polyneuropathy and also its contribution to the infantile CFD syndrome because the PNKP gene plays a dual role in both neurodevelopment and immune-regulatory function. Further research for candidate genes predisposing to FRα-autoimmunity is suggested to include X-chromosomal and non-coding DNA regions.

A next-generation newborn screening pilot study: NGS on dried blood spots detects causal mutations in patients with inherited metabolic diseases.

The range of applications performed on dried blood spots (DBS) widely broadened during the past decades to now include next-generation sequencing (NGS). Previous publications provided a general overview of NGS capacities on DBS-extracted DNA but did not focus on the identification of specific disorders. We thus aimed to demonstrate that NGS was reliable for detecting pathogenic mutations on genomic material extracted from DBS. Assuming the future implementation of NGS technologies into newborn screening (NBS), we conducted a pilot study on fifteen patients with inherited metabolic disorders. Blood was collected from DBS. Whole-exome sequencing was performed, and sequences were analyzed with a specific focus on genes related to NBS. Results were compared to the known pathogenic mutations previously identified by Sanger sequencing. Causal mutations were readily characterized, and multiple polymorphisms have been identified. According to variant database prediction, an unexplained homozygote pathogenic mutation, unrelated to patient's disorder, was also found in one sample. While amount and quality of DBS-extracted DNA are adequate to identify causal mutations by NGS, bioinformatics analysis revealed critical drawbacks: coverage fluctuations between regions, difficulties in identifying insertions/deletions, and inconsistent reliability of database-referenced variants. Nevertheless, results of this study lead us to consider future perspectives regarding "next-generation" NBS.

[The clinical use of the IWG diagnostic criteria for Alzheimer's disease: A retrospective study in a memory clinic]. / L'application en clinique des critères diagnostiques IWG pour la maladie d'Alzheimer: Etude rétrospective dans une clinique de la mémoire.

INTRODUCTION: In recent years knowledge about Alzheimer's disease (AD) presented a major change since the availability of new diagnostic criteria incorporating biomarkers within the known cl inical criteria. Such criteria were elaborated by NIA-AA (National Institute on Aging - Alzheimer's Association) for clinical use and IWG (International Working Group) for research purpose. The aim of this study is to investigate the use of complementary examinations included in the NIA-AA and IWG criteria in a memory clinic characterized by a heterogenous population of patients. METHODS: We retrospectively reviewed patient's characteristics of cases consulting between 01/01/2010 and 31/12/2013 based on the implementation of three additional tests: neuropsychological testing, measurement of cerebrospinal fluid (CSF) biomarkers and brain MRI. Patients were compared according to their origins. RESULTS: 919 patients' medical records were included in the study, 20 % of which were from immigrant background. Non-European Union migrants underwent less neuropsychological testing and more lumbar punctures in comparison with the local population. CONCLUSION: CSF biomarkers allow defining an objective diagnostic component of AD by suppressing the variability due to language, culture, education and the clinical presentation of the disease. Despite the known use in the research domain but not yet validated for clinical use, the IWG criteria are applied in heterogeneous populations with a high percentage of migrants, especially when the results of the neuropsychological testing are poorly contributive. We expect recommendations for clinical use of the CSF biomarkers or better cognitive assessment methods for these populations in the future.

INTRODUCTION: A ce jour, notre vision de la maladie d'Alzheimer (MA) connaît un changement majeur depuis l 'apparition de nouveaux critères diagnostiques incorporant des biomarqueurs aux critères cliniques connus. La NIA-AA (National Institute on Aging - Alzheimer's Association) en a élaboré pour la pratique courante et l'IWG (International Working Group) pour le domaine de la recherche. L'objecti f de cette étude est d'analyser la réalisation des examens complémentaires cités dans les critères NIA-AA et IWG au sein d'une clinique de la mémoire rassemblant une population hétérogène. METHODES: Nous avons revu de manière rétrospective les caractéristiques de patients ayant consulté entre le 1/01/2010 et le 31/12/2013 selon 3 paramètres : la réalisation ou non de l'examen neuropsychologique, du dosage de biomarqueurs dans le liquide céphalo-rachidien (LCR) et de l'imagerie par résonance magnétique cérébrale. Les patients ont ensuite été comparés selon leur origine. RESULTATS: Sur les 919 dossiers analysés dans l'étude, 20 % des patients sont issus de l'immigration. Les migrants non européens ont reçu moins d'examens neuropsychologiques et plus de ponctions lombaires que les patients belges. CONCLUSIONS: Les biomarqueurs du LCR permettent de définir une composante diagnostique objective de la MA en supprimant les variables liées à la langue, la culture, la scolarité et l'expression de la maladie. Malgré leur application connue dans le domaine de la recherche mais actuellement non validée en clinique, les critères IWG prédominent en pratique dans les populations hétérogènes comprenant un haut taux de migrants lorsque les examens neuropsychologiques sont peu contributifs. Des recommandations d'utilisation des biomarqueurs du LCR en pratique courante ou des outils d'évaluation cognitive plus performants dans ces types de population sont attendues.

[PHARMACOGENOMICS AND PERSONALIZED MEDICINE: TOWARDS A SYSTEMATIC GENOMIC SCREENING?]. / PHARMACOGÉNOMIQUE ET MÉDECINE PERSONNALISÉE : VERS UN SCREENING SYSTÉMATIQUE DE LA POPULATION ?

Recent advances in medical genomics open new perspectives for personalized medicine through the identification of genetic variants that influence drug response and/or the risk of side effects. Today, the clinical applications of pharmacogenetics remain scarce as a consequence of the cost and turn-around-time of genetic tests. However, a few tests are recommended, for instance before the prescription of some anti-cancer agents or the anti-retroviral agent abacavir. In the future, we will probably move either towards rapid targeted tests or towards a large screening, before any diagnosis, of all the genetic factors influencing the therapeutic response. In that case, physicians will have to consult the patient genomic data before drug prescription in order to personalize the choice of the therapeutic agent or its dosage. However, such a genomic approach brings economical and ethical questions and will require further progress in our capacity to interpret and store the personal genomic data without compromising their confidentiality.

Dynamic reconstruction of complex abdominal wall defects with the pedicled innervated vastus lateralis and anterolateral thigh PIVA flap.

BACKGROUND AND AIM: Reconstruction of large and chronically infected recurrent abdominal wall defects with exposed bowel in a scarred wound environment, when component release has been previously performed but failed, is a veritable challenge. We use a pedicled innervated vastus lateralis muscle with a fasciocutaneous anterolateral thigh flap (PIVA flap) to restore the continuity of the abdominal wall with vascularised tissues and create a dynamic component that improves the functional outcome. MATERIALS AND METHODS: A one-stage PIVA flap was used in 15 patients with grade 4 transmural chronically infected defects. They had a mean of 4.53 previous laparotomies and important co-morbidities. We determined post-operative reconstructive abdominal wall strength using a validated quality-of-life (QoL) hernia-related questionnaire and modified it to quantify donor-site morbidity at the thigh. We measured the maximal force generated at 60°/s and the force velocity at 120°/s by isokinetic dynamometric analysis at 3 and 12 months. Electromyography (EMG) was performed 12 months after the reconstruction to analyse the contractile integrity of the vastus lateralis segment. A two-sided sign test was used to analyse data. RESULTS: All transmural chronic wounds healed without recurrence. Dynamometric strength increased significantly in the abdominal wall musculature (p < 0.016) and in the donor thigh (p < 0.023) between 3 months and 12 months after the intervention, which reflected in the EMG outcome and the high scores in the QoL measurements after 12 months. CONCLUSIONS: The PIVA flap revascularises the scarred milieu, adds a dynamic component to improve function and may reach up to the xiphoid process. Donor-site morbidity is limited.

[Screening of the risk of functional decline performed by an inpatient geriatric consultation team in a general hospital]. / Dépistage du risque de déclin fonctionnel par une equipe gériatrique mobile au sein d'un hôpital général.

The Mobile Geriatric Team (MGT) is part of the Geriatric Care Program and aims to provide interdisciplinary geriatric expertise to other professionals for old patients hospitalized outside geriatric department. Our hospital has a MGT since 2008. Our objective is to retrospectively describe the population of patients of 75 years and older hospitalized outside the geriatric ward and screened for the risk of functional decline by the MGT between 1 October 2009 and 30 September 2011. We recorded the risk of functional decline, as indicated by the Identification of Senior At Risk score (ISAR) performed within 48 h after admission, place of living, discharge destination, Mini Mental State Examination (MMSE) and Geriatric Depression Scale (GDS) scores. In two years, 1.568 patients > or = 75 Y were screened with the ISAR score (mean age 82.5 Y, 60.7% of women). We identified 833 patients with a high-risk of functional decline (ISAR > or = 3). The majority of high-risk subjects (78%) were living at home before hospitalization and 58.7% returned home after discharge. Depression and cognitive impairment were identified among respectively 41% and 59% of high-risk subjects. Only 128 patients were admitted for fall. Most of the faller patients were living at home prior hospitalization and had an ISAR score > or = 3. The MGT allowed identifying many patients > or = 75 Y living at home and presenting with high-risk of functional decline and geriatric syndromes, confirming that good screening procedures are necessary to optimize management of hospitalized olders. Most of faller patients have an ISAR score > or = 3 and should benefit a comprehensive geriatric assessment.

Learning curve in tracheal allotransplantation.

The first vascularized tracheal allotransplantation was performed in 2008. Immunosuppression was stopped after forearm implantation and grafting of the recipient mucosa to the internal site of the transplant. Nine months after forearm implantation, the allograft was transplanted to the tracheal defect on the radial blood vessels. Since then, four additional patients have undergone tracheal allotransplantation, three (patients 2-4) for long-segment stenosis and one (patient 5) for a low-grade chondrosarcoma. Our goal was to reduce the time between forearm implantation and orthotopic transplantation and to determine a protocol for safe withdrawal of immunosuppressive therapy. Following forearm implantation, all transplants became fully revascularized over 2 months. Withdrawal of immunosuppression began 4 months after graft implantation and was completed within 6 weeks in cases 2-4. Repopulation of the mucosal lining by recipient cells, to compensate for the necrosis of the donor mucosa, was not complete. This resulted in partial loss of the allotransplant in patients 2-4. In patient 5, additional measures promoting recipient cell repopulation were made. The trachea may be used as a composite tissue allotransplant after heterotopic revascularization in the forearm. Measures to maximize recipient cell repopulation may be important in maintaining the viability of the transplant after cessation of immunosuppression.

Deletion of Xpter encompassing the SHOX gene and PAR1 region in familial patients with Leri-Weill Dyschondrosteosis syndrome.

Heterozygote deletions or mutations of pseudoautosomal 1 region (PAR1) encompassing the short stature homeobox-containing (SHOX) gene cause Leri-Weill Dyschondrosteosis (LWD), which is a dominantly inherited osteochondroplasia characterized by short stature with mesomelic shortening of the upper and lower limbs and Madelung deformity of the wrists. SHOX is expressed by both sex chromosomes in males and females and plays an important role in bone growth and development. Clinically, the LWD expression is variable and more severe in females than males due to sex differences in oestrogen levels. Here, we report two familial cases of LWD with a large Xp terminal deletion (approximately 943 kb) of distal PAR1 encompassing the SHOX gene. In addition, the proband had mental retardation which appeared to be from recessive inheritance in the family.

Observational Aranesp Survey to Investigate the Q3W Schedule (OASIS): a prospective observational study of treatment of chemotherapy-induced anaemia with every 3 weeks darbepoetin alfa.

INTRODUCTION: This prospective observational study examined the adherence to published European guidelines on erythropoiesis-stimulating agents (ESAs) and the pattern of use and effect of darbepoetin alfa (DA) 500 microg once every 3 weeks (Q3W) for the treatment of chemotherapy-induced anaemia (CIA). MATERIALS AND METHODS: A total of 293 patients were included (263 solid tumour, 30 haematologic malignancy). Their mean age was 63 years, 51% were male, 57% had platinum-based chemotherapy. DA was started at a haemoglobin (Hb) level between 9 and 11 g/dL in 82% of patients. RESULTS AND DISCUSSION: In an analysis correcting for transfusions, 55% of patients achieved > or =2 g/dL increase in Hb, and a Hb level of >11 g/dL was reached in 81%. Transfusion rate was 27%. Most patients (70%) were treated in a Q3W chemotherapy, and planned synchronisation of chemotherapy and Q3W DA could be maintained in 76%. CONCLUSION: Adherence to European guidelines for DA treatment was good, and Q3W DA treatment was in synchronisation with Q3W chemotherapy in the majority of the patients, thereby reproducing the findings of a recent phase III study.

Goblet cell carcinoid of the appendix: laparoscopic appendectomy or right hemicolectomy?

Goblet cell carcinoids are uncommon tumours with histological features of both adenocarcinoma and carcinoid tumour. They occur predominantly in the appendix and although the malignant potential remains unclear, adenocarcinoids appear to be more aggressive than conventional carcinoids. In this case report, we present a goblet cell carcinoid with laparoscopic operative treatment in two stages. A 43-year-old female patient with constant dullness in the right lower quadrant was diagnosed with acute appendicitis and underwent laparoscopic appendectomy. Macroscopically, a diffusely inflamed appendix was found with no sign of perforation. Histopathological examination revealed a goblet cell carcinoid with characteristics of aggressive behaviour, indicating the need for laparoscopic right hemicolectomy in which, however, neither residual tumour nor metastatic lymph nodes could be found. The postoperative course was uneventful. As they may present the same clinical presentation, pathological diagnosis is required to distinguish goblet cell carcinoid from acute appendicitis. Two-stage surgery for goblet cell carcinoid is advocated in the literature, but the debate still continues as to whether the goblet cell carcinoid should be treated by appendectomy alone, as for most carcinoids, or by right hemicolectomy, as for the appendiceal adenocarcinoma.

Defining the structure of membrane-bound human blood coagulation factor Va.

BACKGROUND: Blood coagulation factor (F) Va is the essential protein cofactor to the serine protease FXa. Factor Va stimulates the thrombin-to-prothrombin conversion by the prothrombinase complex, by at least five orders of magnitude. Factor Va binds with very high affinity to phosphatidylserine containing phospholipid membranes, which allows the visualization of its membrane-bound state by transmission electron microscopy (EM). METHODS: In this paper we present an averaged three-dimensional structure of FVa molecules attached to phosphatidylserine containing lipid tubes, as determined by EM and single particle analysis. The low-resolution FVa three-dimensional structure is compared with the available atomic models for FVa. RESULTS: The experimental data are combined with the most suitable atomic model and a membrane-bound FVaEM model is proposed that best fits the protein density defined by EM. In the FVaEM model, the C1 and C2 membrane-binding domains are juxtaposed onto the membrane surface and the model geometries indicate a deeper insertion of both C domains into the lipid bilayer than has been previously suggested. CONCLUSION: The present structure is a first step towards a higher-resolution experimental structure of a human FVa molecule in its membrane-bound conformation, allowing the visualization of individual domains within FVa and its association with the membrane.

Novel SACS mutation in a Belgian family with sacsin-related ataxia.

The authors describe the four patients in the first known Belgian family with autosomal recessive spastic ataxia of Charlevoix-Saguenay (ARSACS). A novel homozygous missense mutation, NM_014363.3: c.3491T>A in exon 9, of the SACS gene was identified in the present family, which results in an original amino acid of methionine to lysine substitution at amino acid residue 1164 (p.M1164K). Although the cardinal clinical features, i.e., spastic ataxia with peripheral neuropathy, in our patients were similar to those in Quebec patients, our patients exhibited some atypical clinical features, e.g., teenage-onset and absence of retinal hypermyelination. The present family is from Wallonia, and there could be shared ethnicity with the families of Charlevoix-Saguenay.

Cervical cytological screening and human papillomavirus DNA testing in Flanders.

The causal relationship between genital human papillomavirus (HPV) infection and cervical dysplasia/carcinoma has been recognised for some time. The aim of this study was to document the occurrence and distribution of HPV infection in the five provinces of the Flemish region in Belgium and to correlate the HPV DNA test results with the cytological results on simultaneously performed thin layer preparations of cervical cells. Out of a total screened group of 105107 samples, 1978 samples with cytological abnormalities were tested for HPV DNA using the MY09/MY11 consensus PCR. The mean age of the whole group was 36.9 years. The LSIL group, with a mean age of 33.6 years, was significantly younger than the other groups. There was no significant difference in HPV prevalence among the provinces. In four out of five provinces the HPV prevalence reached 100% in high-grade lesions. There is a significant increase in predominance of high-risk HPV types, with increasing abnormal cytology (17.9% WNL < 51.1% ASCUS < 83.8% LSIL < 97.2% HSIL). Three peaks of HPV DNA positivity were observed, a first at 22 yrs (82%), a second at 47 yrs (60%) and a third in women older than 65 yrs (52%). These results shed more light on HPV prevalence in Flanders and show that the MY09/MY11 consensus primer based detection system is very suitable for the detection of HPV infections in Flanders.

Ki-67 immunocytochemistry in liquid based cervical cytology: useful as an adjunctive tool?

AIMS: To test the ability of Ki-67 to detect cytological lesions in a screening setting and its use as a surrogate marker of human papillomavirus (HPV) infection. METHODS: A study of liquid based cytology, HPV DNA testing by MY09/MY11 consensus polymerase chain reaction (PCR), type specific PCRs, and Ki-67 immunocytochemistry on a randomly selected series of 147 patients. RESULTS: Comparison of the number of Ki-67 immunoreactive cells/1000 cells in the different cytological groups showed that the HSIL group yielded a significantly higher mean count than did the other groups. The number of Ki-67 immunoreactive cells/1000 cells was significantly higher in HPV-16 positive samples than in samples containing infections with other high risk types. Receiver operating characteristic curves indicated a test accuracy (area under curve) of 0.68, 0.72, and 0.86 for atypical squamous cells of undetermined significance (ASCUS), low grade squamous intraepithelial lesions (LSIL), and high grade squamous intraepithelial lesions (HSIL), respectively. Thresholds for 95% sensitivity were 0.07, 0.08, and 0.15 Ki-67 immunopositive cells/1000 cells for ASCUS, LSIL and HSIL, respectively. The threshold for 95% specificity was 1.9 Ki-67 immunopositive cells/1000 cells. CONCLUSIONS: Ki-67 immunocytochemistry can be applied to liquid based cytology. The accuracy and diagnostic indices of the test are good when compared with those of other techniques. As part of a panel of screening procedures, it could be used as an adjunct to liquid based cytology to identify HSIL, and as a surrogate marker of HPV-16 infection.

Thin-layer liquid-based cervical cytology and PCR for detecting and typing human papillomavirus DNA in Flemish women.

The objective of this study was to document the occurrence and to correlate the prevalence of different human papillomavirus (HPV) types with the cytological results on simultaneously performed thin-layer preparations in a large population of Flemish women. During 1 year, 69 290 thin-layer preparations were interpreted using the Bethesda classification system. Using an algorithm for HPV testing based on consensus primers and type-specific PCRs in combination with liquid-based cytology, we determined the occurrence and distribution of 14 different oncogenic HPV types (16, 18, 31, 33, 35, 39, 45, 51, 52, 56, 58, 59, 66 and 68). Reflex HPV testing was performed on cytologically abnormal samples and on an age matched randomly selected control group with normal cervical cytology (n=1351). Correlation between cytology, age and prevalence for the 14 different high-risk HPV types is given. There is a significant increase in predominance of high-risk HPV types, with increasing abnormal cytology. Coinfection with multiple HPV types also increased with cytological abnormalities, and was highest in HSIL (16.7%). In Flanders, HSIL was most often associated with HPV types 16, 33, 35, 31, 18 and 51. Using thin-layer liquid-based cytology and PCR to detect HPV, it is feasible to screen large numbers of women.

Comparative sequence analysis of the imprinted Dlk1-Gtl2 locus in three mammalian species reveals highly conserved genomic elements and refines comparison with the Igf2-H19 region.

The Dlk1-Gtl2 domain on mouse chromosome 12 contains reciprocally imprinted genes with the potential to contribute to our understanding of common features involved in imprinting control. We have sequenced this conserved region in the mouse and sheep and included the human sequence in a three species comparison. This analysis resulted in a precise conservation map and identification of highly conserved sequence elements, some of which we have shown previously to be differentially methylated in the mouse. Additionally, this analysis facilitated identification of a CpG-rich tandem repeat array located approximately 13-15 kb upstream of Gtl2. Furthermore, we have identified a third imprinted transcript that overlaps with the last Dlk1 exon in the mouse. This transcript lacks a conserved open reading frame and is probably generated by cleavage of extended Dlk1 transcripts. Because Dlk1 and Gtl2 share many of the imprinting properties of the well-characterized Igf2-H19 domain, it has been proposed that the two regions may be regulated in the same way. Comparative genomic examination of the two domains indicates that although there are similarities, other features are very different, including the location of conserved CTCF-binding sites, and the level of conservation at regulatory regions.