Longitudinal growth of French singleton children born after in vitro fertilization and intracytoplasmic sperm injection. Body mass index up to 5 years of age.

BACKGROUND: The literature presents conflicting results on the epigenetic effect of in vitro fertilization (IVF) on the short-term and mid-term growth of children. These contradictory results may be related to the use of heterogeneous methodologies and non-longitudinal data. The goal of this study was to compare the body mass index (BMI) of children conceived via IVF and spontaneous conception (SC) children, using longitudinal data from birth to 5 years. METHODS: This study compared 118 singleton children born after in vitro fertilization, with or without intracytoplasmic sperm injection (ICSI), selected from a pre-existing single-center cohort to 320 SC children from the same geographic area. BMI and its evolution were analyzed using the mixed-effect model during three periods: before standing acquisition (from birth to 1 year of age), during standing acquisition, and the following period from 2 to 5 years of age. RESULTS: BMI means were not significantly different between groups regardless of the period, when adjusting for confounding factors related to parents, pregnancy, and children's characteristics and lifestyle. Nevertheless, during the standing acquisition period, children born after IVF-ICSI presented a less significant decrease in BMI (P<0.05). In addition, for each period we identified influencing factors (maternal BMI, level of wealth indicator) associated with BMI. CONCLUSION: In the study population, the suspected epigenetic influence of IVF reported in the literature was not observed for BMI from birth to 5 years of age. Further investigations need to be conducted to determine if the suspected influence of IVF on adiposity could be expressed through other parameters.

[Reflection on the psychiatric financial allocation in France]. / Réflexion sur la péréquation financière de psychiatrie en France.

For 25years work has been underway in France for the implementation of an alternative to public financing of health care. In the absence of progress, some regional health agencies are engaged in work related to the reallocation of public finances between psychiatric institutions. We propose a reflection with suggestion on the method proposed by the Provence Alpes Côte d'Azur Regional Health Agency. Without questioning the need for a reallocation of resources between psychiatric institutions, the method proposed here needs to evolve further to be applied in a legitimate and appropriate manner. There is a kind of urgency for a reallocation of resources between psychiatric institutions in France, but it implies a collective thinking and especially the definition of evaluation procedures for the selected models. These conditions are necessary to guarantee the quality of French psychiatry and equity in access to psychiatric care.

Quantitative immunocytochemical assays of P-glycoprotein in breast carcinomas: correlation to messenger RNA expression and to immunohistochemical prognostic indicators.

BACKGROUND: Chemotherapy failure that is due to cellular drug resistance remains a major problem in most cancer patients. One type of drug resistance that has been characterized is the multidrug resistance phenomenon, which demonstrates a reduced ability of cancer cells to accumulate drugs as a result of the effects of an energy-dependent unidirectional drug efflux pump with a broad substrate specificity. This drug pump is composed of a 170-kd transmembrane glycoprotein referred to as the P-glycoprotein (P-gp) that uses energy in the form of adenosine triphosphate to transport drugs through a channel formed by transmembrane segments. PURPOSE: Our purpose was to detect the levels of P-gp expression in frozen untreated breast carcinomas by immunocytochemical assays and to correlate these levels to current prognostic indicators and, in a few cases, to MDR1 (also known as PGY1) mRNA expression by polymerase chain reaction (PCR). METHODS: The immunocytochemical expression of the multidrug resistance gene, P-gp, was investigated using a specific monoclonal antibody (JSB1) against P-gp in 5-microns frozen sequential sections of breast carcinomas obtained from 213 patients. Microscopic images of immunostained preparations were evaluated by image analysis and were compared with MDR1 transcription (mRNA) assessed by PCR in 16 patients. Quantitative P-gp immunocytochemical assays were correlated to histoprognostic factors and immunocytochemical indicators. RESULTS: Among the 213 breast carcinomas tested, 113 (53%) were P-gp positive, but in 28% of the tumors, the immunostained surface accounted for less than 5% of the total area stained. Quantitative immunocytochemistry reflecting the amount of intracellular P-gp antigen strongly correlated (r = 0.865; two-sided, P < .0001; Pearson's test) with the quantitative evaluation of the scanner analysis of mRNA transcripts. The P-gp expression was significantly (two-sided, P < .001) correlated with p53 expression in tumors, to cathepsin D and Ki67 (two-sided, P < .01) immunoreactivity, and to a lesser extent, the detection of estrogen receptor antigenic sites (two-sided, P = .019). P-gp expression was found to be independent of expression of progesterone receptor and pS2, pHER-2/neu, and CD31 in tumors and from patient age, tumor size, histologic types, grades and Nottingham prognostic index, and nodal status. CONCLUSIONS: The quantitative immunocytochemical assays of P-gp are correlated to PCR analysis of MDR1 expression, and such correlations can be useful in evaluating potential multidrug resistance in breast cancer. However, the clinical significance of P-gp immunodetections remains to be further determined.

Multiparametric evaluation (SAMBA) of growth fraction (monoclonal Ki67) in breast carcinoma tissue sections.

Breast tissue samples, including normal breast, nonmalignant disorders, and breast carcinomas (n = 257), were tested with monoclonal antibody Ki67 to define the growth fraction in each tissue subgroup. Immunocytochemical assays using anti-Ki67 and avidin-biotin-peroxidase complex and/or alkaline phosphatase anti-alkaline phosphatase were applied in frozen sections. The immunoreactions were analyzed with a computerized system of image analysis referred to as SAMBA (Systeme d'Analyse Microphotometrique à Balayage Automatique). This system permitted a multiparametric and automatized analysis of colored images. The results obtained were: (a) the SAMBA analysis of Ki67-positive staining was accurate, reliable, and reproducible; (b) the anti-Ki67 immunostaining was significantly (P less than 0.01) increased in malignancies and was related to the tumors' degree of differentiation, the vascular invasion, and the presence of axillary lymph node metastases; (c) anti-Ki67 immunostaining is increased (P less than 0.01) in tumors in which estrogen receptor and progesterone receptor antigenic sites are not detected. It is concluded that the SAMBA analysis of the anti-Ki67 immunocytochemical assay provides relevant information in selecting subgroups of patients with higher risk for relapse.

[Medical emergency care units in France]. / Les maisons médicales de garde en France.

UNLABELLED: The objective of this article is to take inventory of the different medical emergency care units in France as of June 30, 2003, and to describe their main characteristics. METHODOLOGY: A telephone survey was conducted which posed questions based on a standard questionnaire to the Regional Unions of Liberal Doctors, the County Advisory Boards of the Medical Orders, and the directors and managers of the existing emergency care units. RESULTS: 97 medical safe houses were inventoried, 46 of which were operational and 51 which were in development. The Ile-de-France region is the region with the highest concentration of these units either operating or under construction. A large majority of the emergency care units were created since the year 2000, and half of them are located within health care centres and the other half into separate cells either close or distant from existing health care centres. More than half of them have no other staff aside from doctors, whilst the others comprise a spectrum of personnel including secretaries, nurses, and social assistants amongst others. More than half of the doctors are paid at the cost of each consultation and per treatment and do not have a technical platform at their disposal. Usually, these emergency care units are open all night during the week and on the weekends. One-third of them collaborate with the emergency centre and ambulance service (those who respond to calls to 15) and three-quarters of them are funded by the city's Assistance Fund for the Quality of Care. CONCLUSION: It would seem advantageous that a Charter for operation and management of these establishments be drafted and implemented in order standardise these types of structures and that their supervisors implement a national and regional follow-up mechanism for the establishments in order to better evaluate the evolution in terms of health care organisation, in particular with respect to raising the level and capacity of response.

Bromocriptine effects on cultured human prolactin-producing pituitary adenomas: in vitro ultrastructural, morphometric, and immunoelectron microscopic studies.

In an effort to better characterize the ultrastructural, morphometric, and immunocytochemical changes induced by 10(-8) M bromocriptine (BR), tumor cells from three surgically removed PRL-producing pituitary adenomas were cultured on an extracellular matrix in serum-free medium. In each instance, the treated cultures were compared to control cells at the end of 24 h and 16 days. PRL RIAs were performed on culture medium. A decrease in cell and nucleus surface area was found on day 16 in two cultures. This supports the well known shrinkage of BR-treated PRL-producing adenomas. BR induced no change in these parameters in the tumor from a third patient who was partly resistant to the drug. Changes in the secretory process were discernible as of day 1 in all three tumors, with a dramatic reduction of exocytosis and intracellular accumulation of PRL-immunoreactive granules. This induced delayed inhibition of protein synthesis, demonstrated by preembedding immunocytochemistry on day 16. These results, obtained for the first time in human PRL-producing adenomas, are informative as to the subcellular events subsequent to short term BR treatment and illustrate that secretory inhibition and tumor shrinkage are not necessarily linked.

Digital image-analysis of nuclear morphometry, DNA-ploidy and AgNORs in breast-carcinoma cell imprints.

A series (n = 322) of breast carcinomas was investigated from 1991 to 1993 using digital image analysis. Nuclear morphometry and DNA content, and AgNORs were evaluated on cell imprints from fresh tissue samples which were further stored frozen (-80-degrees-C). Data were correlated to morphological prognostic factors and immunocytochemical expression of cell markers assessed on frozen sections and evaluated by densitometry after image analysis processing. Nuclear morphometric parameters, DNA nuclear content and AgNORs were independent from the tumor size, histological grades, and the tumor content of immunodetectable pHER-2/neu, Cathepsin D, ER, PR, pS2, and p53. But, DNA index and hyperploidy degree correlated with the mitosis index (p < 0.01) and Ki67 immunostaining (p = 0.003, p < 0.0001) whereas the shape factor and nucleus large diameter correlated with the degree of cell anisocytosis (p < 0.01). Nuclear surface and large diameter were greater in ductal carcinomas (p = 0.028) and in comedocarcinomas (p < 0.001) than in lobular carcinomas. These results suggest that image analysis processing provides accurate data to refine histoprognostic grading and additional parameters to evaluate tumor proliferative activity.

Quantitative imaging of ps2 immunocytochemical assays in breast carcinomas.

pS2 expression was studied in 212 breast carcinomas. Immunocytochemical assays (ICAs) were performed on frozen sections and evaluated by digital image analysis. pS2 immunostaining was compared in frozen sections and paraffin sections. The pS2 positivity was observed in 45% of the cases on frozen sections. But in pS2 positive tumors, the tumor surface which was immunostained was small (m=14.3%). In 22% of immunoreactive tumors the positive surface was 5% or less. In contrast to frozen sections, the pS2 positivity on paraffin sections could not be evaluated by image analysis system because of the background. No significant correlation was observed between pS2 expression and patient age, tumor size, histological type and grade, nor with lymph node status. The pS2 positivity was significantly correlated to ER and PR positive immunodetection on frozen sections. But pS2 was independent from pHER-2/neu and cathepsin expression whereas there was a significant inverse relationship between pS2 and Ki67. This study shows that pS2-ICAs on frozen sections and image analysis are optimal and standardized conditions for the evaluation of pS2 expression in breast carcinomas, and suitable for selecting patients for hormone therapy.

Prognostic value of Ki 67/MIB1 automated and quantitative immunolabelling in primary operable breast carcinomas.

The prognostic significance of Ki 67/MIB1 immunohistochemical assays (ICAs) was investigated in optimal technical conditions in 139 breast carcinomas. Automated ICAs (immunoperoxidase/Ventana device) was performed on frozen sections. Immunoprecipitates were quantified by computerized analysis (SAMBA) of digitized microscopic images. Mean positive surfaces (%) and quantitative immunocytochemical (QIC) index were correlated to the patients survival (8-year survival). The results showed that Ki 67/MIB1 large surfaces (cutpoint, 20%) and high QIC index (cutpoint, 12) correlated with poor overall survival (Kaplan Meier, log rank test, p=0.011 and p=0.037, BMDP software). In node positive, but not in node negative patients, large Ki 67/MIB1 surface (cutpoint, 20%) and high QIC index (cut-off 12) correlated with the overall patient survival (p=0.0037 and p=0.049). Also large Ki 67/MIB1 positive surface (cut-off, 20%) correlated with disease-free survival in all patients and node positive patients (p=0.022 and p=0.0057) but not in node negative patients. It is concluded that in optimal technical conditions (automated and quantitative immunohistochemical assays on frozen sections) Ki 67 antigen immunohistochemical expression in breast carcinomas tissue has a prognostic significance in node positive patients but not in node negative patients.

Expression of her-2/neu oncogene in breast-cancer - correlation of quantitative immunocytochemistry and prognostic factors.

HER-2/neu oncogene expression by breast carcinomas (n = 208) was investigated on frozen sections using monoclonal anti-p185 HER-2/neu protein. Results were evaluated by computer-assisted image analysis and correlated with morphological prognostic factors, hormone receptor antigenic sites, Ki 67 antigen and cathepsin content, nuclear morphometry, DNA content and Ag NORs, which were also evaluated by image analysis. All tumors were anti-p185 HER-2/neu immunoreactive, but in 40% of the cases, less than 20% of the tumor cell surface was immunostained. In terms of both extent and intensity, immunostaining which was greatest in comedocarcinomas correlated with tumor size (p = 0.019) and Ki 67 (p = 0.0012) and cathepsin (p<0.0001) content. No correllation was found with tumor grade, axillary lymph node involvement, hormone receptor sites, nuclear DNA content and Ag NORs distribution and morphometry.

p53 quantitative immunocytochemical analysis in breast carcinomas.

A series of 200 breast carcinomas was investigated on frozen sections using PAb 1801 p53 monoclonal antibody and streptavidin biotin peroxidase complex. Densitometric analysis of the immunoprecipitates was assessed by processing digitized microscopic images. p53 was observed in the nucleus of 48% of the tumors. Some tumors (14 of 91) tested in parallel on paraffin sections were negative, although positive on frozen sections. Image analysis showed that the surfaces positive with anti-p53 and the staining intensity were decreased (P < .01) on paraffin sections. The p53 tumor expression was independent of patient age, tumor size, axillary lymph node status, HER-2/neu and cathepsin D expression, and nuclear morphometric parameters. However, p53 correlated with high histological grade (P < .01), lack of estrogen receptor (ER) (P = .0015) and progesterone (PR) (P = .0065) antigenic sites, pS2 detection (P = .03), high Ki-67 immunoreactivity (P = .018), large silver-stained nucleolar organizer region (AgNOR) nuclear surface ratio (P < .02), and degree of hyperploidy (P < .03), and was more often observed in the comedocarcinomas. The results suggest that p53 expression in breast carcinomas is not a totally independent prognostic indicator and that the clinical relevance and prognostic significance of p53 expression in breast carcinomas can be reliably assessed provided that the procedures are standardized, particularly with regard to the use of frozen sections and image analysis processing of the immunodetection.

Automated and quantitative immunocytochemical assays of CD44v6 in breast carcinomas.

CD44 variants carrying sequences encoded by exon v6 are preferentially expressed in metastatic animal cancer cell lines. CD44v6 overexpression correlates tumor dedifferentiation and progression in some human carcinomas, but the relationship of CD44v6 overexpression with metastatic behavior of tumor observed in animal models is controversial, particularly in breast carcinomas. The discrepancies probably result from analytical bias. We investigated CD44v6 and CD44s expression in 218 frozen samples of primary breast carcinomas. Immunocytochemical procedure was performed under optimal technical conditions using commercially available 2F-10 monoclonal antibody (MAb), a microprocessor-controlled automated device (Ventana Medical Systems, Tucson, AZ), and quantitative evaluation of results by processing digitized-colored microscopic images (SAMBA, Grenoble, France). CD44v6 expression in tissue sections was shown to be independent of the patient age, tumor size, histological types and grades, and the lymph node status. CD44v6 expression was also independent of the expression of molecules endowed with poor prognostic significance detected by MAbs (anti-p53, anti-c-erb B-2 protein, MIB1) on consecutive sections. No significant relationship could be evidenced either between CD44v6 expression, and CD31 involved stromal angiogenesis and cathepsin D. Finally, CD44v6 was independent of markers of hormone dependence (estrogen and progesterone receptors, pS2) and of multidrug resistance (P-glycoprotein). Similar results were observed with anti-CD44s. We conclude that the true prognostic significance of CD44v6 overexpression still remains to be shown under rigorous technical conditions (frozen samples, well-documented MAbs, and optimal standardization of procedure using automation and quantitative analysis) providing data appropriate for further correlation with long-term patient follow-up.

CD31 quantitative immunocytochemical assays in breast carcinomas. Correlation with current prognostic factors.

The distribution of PECAM-1/CD31 molecule was investigated in 133 breast carcinomas using monoclonal antibody and frozen sections. Anti-CD31 labels endothelial cells and reflects stromal angiogenesis. The CD31 immunoreactivity was evaluated by computer-assisted analysis of digitized microscopic images. The automatic screening of the whole preparation and the measurements of the mean CD31 immunostained surface was performed in each case. A similar procedure was achieved for p53, cathepsin D, P-gp, pHER-2/neu, Ki67, pS2 estrogen and progesterone antigenic sites immunodetection. The image analysis of positive CD31 surface was variable, ranging from 4% to 33% (mean 14.7%, SD = 5.43). The CD31 positive surface correlated (P < .01) with the Nottingham prognostic index, but not with the tumor size, the node status, the tumor grade, nor with the patient age. Also the CD31 immunoreactivity was independent of the pHER-2/neu, Ki67 antigen, p53, ER, PR and pS2 immunodetectable expression in tumors, but correlates with that of cathepsin D (P = .024) and P-gp (P = .028), which reflects the multi-drug resistance capacity of tumor cells. In conclusion, CD31 positive vessels assessed on frozen sections by image analysis constitute an excellent method of evaluating tumor stromal angiogenesis, and can be further used for clinical purposes. The results also suggest that the CD31/PECAM molecule may be involved in the spread of tumor by interacting with extracellular matrix lysis that results from the tumor cell proteasic activity and with multidrug resistance.

Quantitative immunocytochemical assays on frozen sections of p53: correlation to the follow-up of patients with breast carcinomas.

A series of 222 tumor samples stored at -80 degrees C in the authors' tumor library were investigated with anti-p53 (PA 1801) and streptavidin-biotin-peroxidase complex. The p53 immunoprecipitates were quantified by densitometry assessed by image analysis of digitized microscopic images. Two parameters, percentage of positive surface and mean optical densities, were compared with the patient's outcome (follow-up = 96.8 months) (life table method, Mantel Cox test, BMDP statistical software). The p53 expression significantly correlated with a poor overall survival (P = .0063), metastasis-free survival (P = .024), and recurrence-free survival (P = .022) at a 20% cutoff point of positive immunoreactive tumor surface. A strong prognostic significance was observed in the node-positive subset of patients but not in the node-negative subset, except for recurrence-free survival (P = .047). The results indicate the clinical relevance p53 evaluated by quantitative immunocytochemistry.

Quantitative imaging of estrogen and progesterone receptors, estrogen-regulated protein, and growth fraction: immunocytochemical assays in 52 meningiomas. Correlation with clinical and morphological data.

Quantitative imaging of estrogen receptors (ER's), progesterone receptors (PR's), estrogen-regulated protein (pS2), and growth fraction (Ki67) immunocytochemical assays were performed in 52 meningiomas. The results were correlated with clinical (age, sex, hormonal status, and tumor volume and location) and morphological (histological types and grades) data. The authors observed a lack of ER's in all meningiomas but the presence of PR's in 53% of these meningiomas. The immunoreactivity was restricted to tumor cell nuclei. The PR immunocytochemical assay was correlated with tumor location, histological type, histological grade, and pS2 immunocytochemical assay, but not with Ki67 immunocytochemical assay; high PR content was observed in cisternae, transitional, meningothelial, and low-grade meningiomas. Only 11 meningiomas showed more than 1% Ki67 immunoreactive nuclei. These meningiomas were usually located in the convexity and were of high histological grade. Estrogen-regulated protein immunoreactivity was observed in 34 meningiomas but the number of immunoreactive nuclei was low. The pS2 immunocytochemical assay was not related to clinicopathological features but was preferentially observed in PR-negative meningiomas. The results of this study are compared with those previously reported, and the function and regulation of PR's in meningiomas is discussed. The results indicate that 1) regulation of PR's and pS2 proteins in meningiomas differs from regulation in estrogen-dependent tissues such as breast or endometrium; 2) interruption of hormonal therapy in women presenting with a meningioma is not absolutely necessary; 3) meningiomas have different biological properties according to their clinicopathological features; and 4) future studies of hormonal clinical trials should be performed on well-defined meningioma subgroups.

Immunodetection of HER-2/neu protein in frozen sections evaluated by image analysis: correlation with overall and disease-free survival in breast carcinomas.

The immunodetection of HER-2/neu oncogene product was performed in 108 breast carcinomas using immunoperoxidase technique. Monoclonal anti HER-2/neu protein was applied on frozen sections. Immunoprecipitates were evaluated by a computerized system of image analysis (SAMBA). The percentage of the immunostained tissue surfaces and mean optical densities were correlated with the 5 year overall and disease-free survival rates. It was shown that in optimal conditions of antigen preservation and standardized method of immunoprecipitates evaluation, 67% of breast carcinomas were more than 20% pHER-2/neu positive. The pHER-2/neu overexpression was not significantly correlated with the overall 5 year survival. However large positive anti pHER-2/neu surfaces were correlated with higher risk of recurrence (p = 0.0013) and of metastases (p = 0.035).

Comparison of the prognostic significance of current and modified histological grades in breast carcinomas.

Paraffin sections of 185 breast carcinomas were reexamined in order to compare the current SBR histoprognostic grade (SBR) with to modified methods of grading recently proposed by two groups, Le Doussal et al (MSBR) and Elston et al (SBR-Elston). In each tumor, the SBR, MSBR, SBR-Elston, and each of their components were correlated with recurrence, metastases and survival rates (follow up 7 to 96 months, m = 52, SD = 19) (Kaplan-Meier test). The Nottingham prognostic index (NPI) was computed for each patient, the histological type reevaluated and both were also correlated with the patient follow up. Our results show that the three methods of grading were significantly (p < 0.0001) correlated. The three grades, were significantly correlated with metastases and survival, but not with the recurrence rates. The differentiation, the nuclear pleomorphism and the mitoses count also correlated with the metastases rate and overall survival but the mitoses number appeared to be a stronger prognostic indicator. The MSBR grading made it possible to refine the prognostic of the tumors usually scored as SBR grade 2. The NPI significantly correlated with metastases and survival (p < 0.0001), whereas the histological types were found to have no prognostic significance.

Micronuclei kinetics of exfoliated urothelial cells.

Micronuclei observed in exfoliated cells result from DNA-damage of basal epithelium's cells by mutagens. Exfoliated urothelial cells can be collected by non-invasive procedure and may be used as target site to identify genotoxic effects of chemicals. Kinetic studies are important for any biomarker, especially those in which tissue differentiation and maturation processes will heavily influence the time between induction of damage and collection of damaged cells for analysis. This manuscript details the result of a longitudinal study of micronuclei induction in cells isolated from urine samples of 4 healthy women over 6 consecutive days. Three of them were former smokers. Results suggested that micronucleated cell rates were not influenced neither by the day nor by the time of sampling.

Correlation of nucleolar organizer regions and nuclear morphometry assessed by automatic image analysis in breast cancer with aneuploidy, K167 immunostaining, histopathologic grade and lymph node involvement.

Silver-stained nucleolar organizer regions (AgNORs) in human breast carcinoma were studied using a computer-assisted system of image analysis. Standardized, automatic measurements of 7 morphometric parameters (area, perimeter, shape factor, bend energy, angle, and small and large diameters) performed on paraffin sections and cell imprint were compared and correlated with nuclear morphometry, histopathological grading, tumor growth fraction, (monoclonal Ki67-immunostaining), DNA nuclear content (stoechiometric Feulgen staining) and axillary lymph node invasion. The major findings were as follows: (i) variations in AgNORs and nuclear parameters were correlated, (ii) the ratio of AgNOR area/nuclear area was significantly different in low and high grade tumors, (iii) mean AgNOR parameter values increased significantly with the tumor growth fraction and tumor hyperploidy and were significantly higher in patients with axillary lymph node metastases and (iv) AgNOR evaluation was more accurate for cell preparations than for tissue sections.

[Evaluation of cost of laparoscopic surgery in gynecology]. / Evaluation du coût de la chirurgie coelioscopique en gynécologie.

The objective of this work was to evaluate hospital and postoperative costs associated with gynecologic laparoscopic surgery for benign pathologies. Hospital costs were broken down into several categories: operating room, postoperative hospitalization, pharmacy, surgical instruments, sterilization of reused equipment, food and laundry, indirect costs. The mean total costs for the hospitalization time were 8547,2 francs. Medical charges in recovery time were calculated with the help of the Social Security. These charges represented less than 5% of the total cost associated with the procedure.