Showing papers by "Paul J. van Diest published in 2006"

Expression pattern of a homeotic gene, HOXA5, in normal breast and in breast tumors.

Abstract: Introduction: Homeotic (HOX) gene products are now known to be functionally associated with breast cancer biogenesis. Recent evidence has indicated that HOXA5 regulates both p53 and progesterone receptor expression levels in breast cancer cells. In addition, HOXA5 has been shown to interact and regulate the activity of another protein referred to as Twist. As homeotic genes play a pivotal role in development, we sought to decipher the expression pattern in both normal breast tissues and in breast carcinomas. Methods: RT-PCR and immunohistochemistry were performed, to assay the levels of HOXA5 expression, on a panel of normal breast tissue and its corresponding primary breast tumors. Results and Conclusions: We show that HOXA5 expression was maintained at stable levels at different reproductive stages of a woman's life, except during lactation. This evidence indicates that HOXA5 may play a role in maintaining the differentiated state within the breast epithelium. However, nearly 70% of all breast carcinomas had decreased HOXA5 protein levels as compared to normal breast tissues. In addition, we demonstrate that HOXA5 protein expression levels in breast carcinomas inversely co-relates with Epidermal Growth Factor Receptor (EGFR) expression. Furthermore, we found that the survival rate amongst the different low levels of HOXA5 expressing breast tumors was not significant, indicative of an early tumorigenesis process in the absence of innate levels of HOXA5 in normal breast cells.

Low prevalence of (pre) malignant lesions in the breast and high prevalence in the ovary and fallopian tube in women at hereditary high risk of breast and ovarian cancer

Abstract: To analyse the prevalence of (pre) malignant lesions occurring in breast and adnexal tissue at prophylactic surgery in women at hereditary high risk of breast and/or ovarian cancers. Tissue was obtained from 85 women who underwent prophylactic bilateral salpingo-oophorectomy (pBSO) and from 59 women who underwent prophylactic mastectomy (pM). Control tissue samples were obtained from women undergoing breast reduction surgery (N = 99) or adnexal surgery for benign reasons (N = 72). In women with a BRCA1/2 mutation, the prevalence of a (pre) malignant adnexal lesion was 50% (95% CI 26-74) if older than 40 years and 14% (95% CI 0-58) if younger. The prevalences of (pre) malignant breast lesions in women older than 40 years, with and without a BRCA1/2 mutation, were 0% (95% CI 0-16) and 47% (95% CI 21-73), respectively. No association was found between (pre) malignant lesions in breast and adnexal tissue occurring in 28 women who underwent surgery on both organs (R = 0.155, p = 0.432), but the prevalence of lesions was significantly higher in adnexal tissue than in the breast (p = 0.023). Compared to controls, women at hereditary high risk had a higher chance of (pre) malignant lesions in the breast and an even higher chance of such lesions in the adnexal tissue. There was no indication for concomitant presence of such lesions in both organs at the time of prophylactic surgery. The high frequency of (pre) malignant lesions in the adnexal tissue stresses further the importance of pBSO from the age of 40 onwards in women at hereditary high risk.

Serous borderline tumor of the ovary presenting with cervical lymph node involvement: a report of 3 cases.

Abstract: Supradiaphragmatic lymhadenopathy is extremely rare in patients with a serous borderline ovarian tumor (BOT), and clinically difficult to recognize. We describe 3 cases of serous BOT that primarily presented with arm thrombosis due to supradiaphragmatic lymphadenopathy. In all the 3 cases, fine needle aspiration cytology initially indicated metastatic adenocarcinoma. The primary tumor was not immediately apparent, and multiple diagnostic examinations had to be done before the definitive diagnosis of serous BOT, International Federation of Gynecology and Obstetrics stage IV could be made. In the meanwhile, erroneous therapies had been given in 1 case. After surgical removal of the adnexal masses and full surgical staging, all the 3 patients remained free of disease after a follow-up period of 48 to 84 months. In conclusion, supradiaphragmatic lymph node involvement can be present in patients with serous BOTs, and can even be the presenting symptom. When fine needle aspiration cytology of such a lymph node is compatible with adenocarcinoma of unknown primary, serous BOT should be included in the differential diagnosis and pelvic examination should be performed.

One-time general consent for research on biological samples: opt out system for patients is optimal and endorsed in many countries.

Abstract: EDITOR—Wendler's valuable overview of empirical studies of patients' preferences about the use of residual (leftover) tissue for research is not complete.1 The discussion has already been settled by legislation in many countries. In 2004 the Danish act on patient rights was amended with an opt out system for using residual tissue for research. In 2004 the US Office for Human Research Protections (OHRP) issued guidance …

EGFR Expression Predicts BRCA1 Status in Patients with Breast Cancer

Abstract: To the Editor: In their article, Lakhani et al. ([1][1]) report on the value of basal phenotype markers for the prediction of BRCA1 status. One of the useful features pointing to “BRCA1-ness” appeared to be high expression of the epidermal growth factor receptor (EGFR). No rationale is given by

Synergistic effect of interstitial laser coagulation and doxorubicin in a murine tumor recurrence model of solitary colorectal liver metastasis.

Abstract: Interstitial laser coagulation (ILC) is gaining acceptance for treatment of unresectable colorectal liver metastases. However, local recurrence rates are still high. To overcome this problem, we investigated the potential of additional systemic therapy after ILC in a murine model. Single C26 colon carcinoma nodules (∼1 mm3) expressing firefly luciferase were implanted in the left liver lobe of 32 BALB/c mice. Seven days after implantation, tumors were treated with either ILC alone (neodymium–yttrium aluminum garnet; 6 W/cm; 800 J/cm) or ILC followed by 1 mg/kg of doxorubicin intravenously. Controls received either doxorubicin alone or sham treatment. Tumor load was measured by in vivo bioluminescent imaging. Solitary colorectal liver metastases developed over 7 days after tumor implantation in the liver. Extrahepatic disease was not observed. The ILC dose was set to ablate the liver metastases with recurrent tumor growth in 9 of 16 mice after 7 days. After ILC plus doxorubicin, complete tumor destruction occurred without recurrence (0 of 14). Sham treatment or treatment with doxorubicin alone showed an exponential increase in tumor load. A murine tumor recurrence model after local ablative treatment of solitary liver metastasis was developed. The combination of ILC and doxorubicin had a strong synergistic effect that led to complete tumor remission in all animals treated.

Confocal 3D DNA Cytometry: Assessment of Required Coefficient of Variation by Computer Simulation

Abstract: Background: Confocal Laser Scanning Microscopy (CLSM) provides the opportunity to perform 3D DNA content measurements on intact cells in thick histological sections. So far, sample size has been limited by the time consuming nature of the technology. Since the power of DNA histograms to resolve different stemlines depends on both the sample size and the coefficient of variation (CV) of histogram peaks, interpretation of 3D CLSM DNA histograms might be hampered by both a small sample size and a large CV. The aim of this study was to analyze the required CV for 3D CLSM DNA histograms given a realistic sample size. Methods: By computer simulation, virtual histograms were composed for sample sizes of 20000, 10000, 5000, 1000, and 273 cells and CVs of 30, 25, 20, 15, 10 and 5%. By visual inspection, the histogram quality with respect to resolution of G0/1 and G2/M peaks of a diploid stemline was assessed. Results: As expected, the interpretability of DNA histograms deteriorated with decreasing sample sizes and higher CVs. For CVs of 15% and lower, a clearly bimodal peak pattern with well distinguishable G0/1 and G2/M peaks were still seen at a sample size of 273 cells, which is our current average sample size with 3D CLSM DNA cytometry. Conclusions: For unambiguous interpretation of DNA histograms obtained using 3D CLSM, a CV of at most 15% is tolerable at currently achievable sample sizes. To resolve smaller near diploid stemlines, a CV of 10% or better should be aimed at. With currently available 3D imaging technology, this CV is achievable.

Microvessel density and p53 in detecting cervical cancer by FDG PET in cases of suspected recurrence.

Abstract: Cervical cancer is the second most frequently diagnosed cancer in women worldwide. About one-third of patients experience recurrent disease. A better chance of survival might be achieved by the early detection of recurrent cervical cancer. [18F]fluoro-2-deoxy-D-glucose (FDG) PET could be a promising imaging modality for this purpose, given that FDG PET has high diagnostic efficacy. Ideally, pre-selection of patients should be performed before considering FDG PET. The purpose of this study was to investigate parameters of primary cervical cancer associated with recurrence as a basis for pre-selection of patients in whom FDG PET should be performed. Thirty-eight cervical cancer patients, clinically suspected of having recurrent disease, underwent FDG PET. Tissue from primary tumours and nine histologically confirmed metastases was analysed for biomarkers possibly related to glucose metabolism and prognosis (vascular endothelial growth factor, CD31 for microvessel density, glucose transporter-1, hexokinases I, II and III, Ki67, p53, hypoxia-inducible factor 1α, and degree of infiltration by lymphocytes and macrophages). Based on clinical outcome, sensitivity and specificity of FDG PET were 96% and 100%, respectively. Cox regression revealed microvessel density and p53 (tumour suppressor protein) to be the two most important biomarkers for prediction of recurrence (hazard ratios 2.54 and 2.28, respectively). By combining these two biomarkers in a parallel test, sensitivity and specificity in predicting recurrence were 87% and 71%, respectively. Leave-one-out cross-validation demonstrated predictive validity of a model based on microvessel density and p53. In this first study of its kind, we have demonstrated that microvessel density and p53 profiles could be important in pre-selecting cervical cancer patients for detection of recurrence by FDG PET.

A restaining method to restore faded fluorescence in tissue specimens

Abstract: The aim of this study was to develop a procedure to remove the TO-PRO-3 fluorescent dye from tissue sections and restain with TO-PRO-3, still allowing calculation of 3-D cellular DNA content and nuclear chromatin texture features by confocal laser scanning microscopy. After staining adrenal tissue with TO-PRO-3 and image acquisition, three destaining approaches were tested based on incubation at different temperatures for different durations in the medium that is normally used to dissolve TO-PRO-3. The same areas were imaged again to measure residual fluorescence, the slides were then restained with TO-PRO-3 and imaged again. After image matching, the grey-values of the images acquired after initial and restaining were compared. A number of 3-D texture features computed after segmentation of nuclei were compared as well. The best destaining result was obtained by incubation of sections at 37°C in pre-heated medium twice for 20 minutes. On average, the 3-D feature values did not change much after de/restaining, except for some discrete texture features which are sensitive to contrast that was slightly lower after restaining. In conclusion, we present a protocol to remove TO-PRO-3 fluorescence from tissue sections that can subsequently successfully be restained with minimal influence on fluorescence intensity and nuclear chromatin distribution that allows repeated and reliable TO-PRO-3 fluorescence quantification (and probably other intercalating fluorescent dyes) on the same section. Andre Huisman (2006) 3-D Nuclear chromatin texture analysis using confocal laser scanning microscopy

One-time general consent for research on biological samples

Abstract: EDITOR—Wendler suggests that since most (79-95%, depending on the study) people were willing to provide a one-time general consent to further usage of their biological sample we should routinely include a request for the subject to consent to the further indefinite usage of their samples in consent forms and participant information sheets.1 This misconstrues the nature of autonomy by conflating it with majoritarian democracy. Simply because most people would hypothetically consent to something …

Mindere kwaliteit van cervixuitstrijkjes afgenomen door doktersassistentes in vergelijking met huisartsen

Abstract: Buis PAJ, Van den Heuvel L, Balfoort GAMA, Chorus RMH, Van Diest PJ. Mindere kwaliteit van cervixuitstrijkjes afgenomen door doktersassistentes in vergelijking met huisartsen. Huisarts Wet 2006;49(1):24-7.

Implementation of accurate and fast DNA cytometry

Abstract: Confocal Laser Scanning Microscopy provides the opportunity to perform 3-D DNA content measurements on intact cells in thick histological sections. The aim of this study was to describe the conditions for accurate and fast 3-D CLSM cytometry with a minimum of user interaction to arrive at sufficient throughput for pilot clinical applications. To achieve this aim, nuclear DNA was stained in 14 μm thick tissue sections of normal liver and adrenal stained with either YOYO-1 iodide or TO-PRO-3 iodide. Different pre-treatment strategies were evaluated: boiling in citrate buffer (pH 6.0) followed by RNase application for 1 or 18 hours, or hydrolysis. The image stacks obtained with CLSM at microscope magnifications of ×40 or ×100 were analyzed off-line using in-house developed software for semi-automated 3-D fluorescence quantification. As a measure of histogram quality, the coefficient of variation (CV) of the diploid peak was assessed. The lowest CV (10.3%) was achieved with a protocol without boiling, with 1 hour RNase treatment and TO-PRO-3 iodide staining, and a final image recording at ×60 or ×100 magnifications. A sample size of 300 nuclei was generally achievable. By filtering the set of automatically segmented nuclei based on volume, size and shape, followed by interactive removal of the few remaining faulty objects, a single measurement was completely analyzed in approximately 3 hours. The described methodology allows obtaining a largely unbiased sample of nuclei in thick tissue sections using 3-D DNA cytometry by confocal laser scanning microscopy within an acceptable time frame for pilot clinical applications, and with a CV small enough to resolve small near diploid stemlines. Andre Huisman (2006) 3-D Nuclear chromatin texture analysis using confocal laser scanning microscopy

Development of 3-D chromatin texture analysis

Abstract: The aim of the study described in this Chapter was to develop and implement 3-D texture features that quantitatively describe the nuclear chromatin distribution. Thirty-five features thoughtfully chosen from 4 categories of 3-D texture features (discrete texture features, Markovian features, fractal features, grey value distribution features) were selected and tested for invariance properties (rotation and scaling) using artificial images with a known grey value distribution. The discriminative power of the 3-D texture features was tested on artificially constructed benign and malignant 3-D nuclei with increasing nucleolar size and advancing chromatin margination towards the periphery of the nucleus. 10 Benign and 10 malignant human prostate nuclei were acquired by CLSM to evaluate whether there was more texture information present in 3-D whole nuclei compared to a single 2-D plane from the middle of the nucleus. A set of 35 3-D nuclear texture features was used successfully to assess nuclear chromatin patterns in 3-D images obtained by confocal laser scanning microscopy. Andre Huisman (2006) 3-D Nuclear chromatin texture analysis using confocal laser scanning microscopy