Hibernoma of the antero-lateral thigh.

Hibernoma is a rare benign tumour of brown fat. In most cases it presents as a voluminous slow-growing mass in the regions where remnants of brown fat can remain in adults. We report a case of a hibernoma on the thigh and present the diagnostic and therapeutic elements of this type of tumour, whose differential diagnosis of liposarcoma.

[Kaposi disease, limited to the skin, in ulcerated hemorrhagic rectocolitis]. / Maladie de Kaposi strictement cutanée au cours d'une rectocolite ulcérohémorragique.

We report the first case of limited cutaneous Kaposi's sarcoma during ulcerative colitis, in a HIV-negative 68 year-old man without any risk factors for this tumor. In our case, the steroid-therapy necessitated by the colitis is the most important aetiological factor of the sarcoma and its withdrawal leads to a complete remission of the tumor. The link between steroid-therapy and Kaposi's sarcoma is well known. A link between ulcerative colitis and Kaposi's sarcoma can be discussed but not proven. The possible occurrence of this cutaneous tumor during ulcerative colitis is of interest.

[Impalpable cancers of the breast. Histological and immunohistochemical study]. / Les cancers du sein impalpables. Etude histologique et immunohistochimique.

Breast cancer screening enables the diagnosis of impalpable lesions detectable by mammography or ultrasonography. The diagnosis of impalpable cancers lacking macroscopic disorders is difficult. However the diagnosis is facilitated by X-raying of surgical specimens, which shows that screened breast lesions have effectively been excised and which exactly localize these lesions. In addition, the X-raying of sliced (5 mm thick) fresh surgical specimens during surgery, enables the intraoperative diagnosis on frozen section of 72% of breast carcinomas and a one step surgical treatment (clear specimen edges, axillary lymph node excision). Moreover, the intraoperative examination of surgical specimens enables appropriate sampling of tissue for delayed immunodetections on frozen sections. These immunodetections provide prognostic information complementary to current microscopic investigation, particularly relevant in small node negative tumors.

[Granular cell tumors of the breast]. / Tumeurs à cellules granuleuses du sein.

The granular cell tumor was first described by Abrikossoff who referred to that lesion as myoblastoma. This lesion is mainly observed in soft tissues and is exceptionally found in the mammary gland. Among the 159 cases of breast granular cell tumor reported in the literature, only 121 have been well documented. Tumors occurred in 15 to 74 year old patients and in only 9.8% of the tumors discovered in males. In breast this tumor may mimic a carcinoma like in the three cases reported. The intra-operative diagnosis on frozen sections may be particularly difficult and the risks to overdiagnose a carcinoma are not negligible and must be known. The diagnosis on paraffin sections is much more easier than on frozen sections. The immunostaining procedures (anti p S100) and electron easier than on frozen section. The immunostaining procedures (anti p S100) and electron microscopy are mainly interesting to document the histogenetic hypotheses. The schwannian origin of the tumor is favoured in most of the recent studies. The granular cell tumors are usually benign and malignant tumors account for only 2.5% of those diagnosed in the breast.

Non palpable breast carcinomas. Histological and immunohistochemical studies of 160 cases.

A series of 160 impalpable breast carcinomas was collected from 1979 to 1991. Mammographs showed microcalcifications (64%), or opaque images (36%). Surgical specimens were X-rayed during the intervention in order (i) to ascertain that the lesions detected on mammographs were removed, and (ii) to guide the pathologist in sampling tissue fragment for an appropriate microscopic evaluation of the lesions. During the intervention, the peroperation histological diagnosis was correct in 63% of the cases, whereas malignancies were underscored in 37%. No false positive diagnosis was recorded. A large majority (92%) of false diagnoses stated during surgery were in situ carcinomas diagnosed as epitheliosis and invasive carcinomas diagnosed as in situ carcinomas. In 63% of the cases the axillary lymph node could be removed during the first intervention. In 91% of the cases "in sano" margins of resection were evaluated as such during the intervention. The size of tumors ranged from 1 to 60 mm (m = 10 mm - SD = 8.45), 70% measuring less than 10 mm. Carcinomas were in situ (23.75%), microinvasive (13.75%) and invasive (62.5%). Carcinomas were ductal (78.1%), lobular (18.7%) and of other types (14.2%). A majority of intraductal carcinoma (68%) were comedocarcinomas. Invasive carcinomas accounted for grade I in 37% of the cases, grade II in 56%, grade III in 7%, ductal carcinomas and for tubular carcinomas in 15%. Immunodetection could be performed on frozen sections in 78 cases. Tumors were receptor positive in 58% of the cases. The greater growth fraction (Ki-67) and higher detection of HER-2/neu oncogene product were observed in comedocarcinomas. Diploid tumors accounted for 52% of those evaluated (n = 48).

X-raying of sliced surgical specimens during surgery: an improvement of the histological diagnosis of impalpable breast lesions with microcalfications.

Surgical specimens, from 78 patients operated for impalpable breast lesions containing microcalcifications detected on mammographs (breast cancer screening program) were systematically examined by a pathologist during the surgical intervention. The per-operative procedures encompassed (i) X-raying of the unfixed specimens to ascertain that the latter did enclose the screened lesion, (ii) slicing and subsequent X-raying of the specimens slices in order to locate the microcalcifications, (iii) histological evaluation of areas containing the microcalcifications on frozen sections. Thirty-two of the lesions were histologically benign, 55% were malignant and 13% borderline. Forty of the carcinomas were in situ and 60% were invasive. The per-operative histological diagnosis was correct in 65% of the case, erroneous in 10% and uncertain in 25%. Malignancy was never overscored. In 65% of the carcinomas diagnosed during the surgical intervention, the in sano margins excision and axillary lymph node removal could be performed (one stage surgical treatment). These results suggest that X-raying of sliced specimens and histological evaluation during the surgical intervention ascertains that lesions screened by mammographs are effectively and completely removed, and that they are precisely and extensively histologically examined after being identified and located.

Inflammatory breast carcinoma: an immunohistochemical study using monoclonal anti-pHER-2/neu, pS2, cathepsin, ER and PR.

Immunocytochemical assays were performed on frozen sections of inflammatory breast carcinomas (n = 22) using the following monoclonal antibodies (MoAb): anti-pHER2/neu, cathepsin, pS2, ER, PR and MoAb Ki67. The distribution of these proteins, known as prognostic indicators, was evaluated with an image analysis system (SAMBA, Alcatel TITN, France). On standard HE stained paraffin sections, only about 50% of inflammatory breast tumors exhibited intradermal tumor cell emboli. All tumors were strongly pHER/2neu positive. All tumors also, but to a lesser degree, were cathepsin and ki67 positive. Conversely, less than 40% were faintly ER, PR and pS2 immunoreactive. The results correlated with the high degree of malignancy of inflammatory breast carcinomas. Therefore the immuno-detection of these markers in addition to standard histological techniques appears to be a useful tool to evaluate the degree of malignancy of breast carcinomas.

Inflammatory carcinoma of the breast.

Inflammatory breast carcinoma has to be defined by accurate clinical and pathological criteria. The prognosis is very poor and improvements made by chemotherapy are limited to the increase in the disease-free period and overall survival for a few months.

Correlation between laminin and type IV collagen distribution in breast carcinomas, and estrogen receptors expression, lymph node and vascular involvement.

The laminin (Lam) and type IV collagen (Coll IV) and estrogen receptor (ER) immunodetections were assessed in a large series of 400 human breast carcinomas. In all the cases the patient's age, the tumor size, the histological type and grade, the presence or the absence of axillary lymph node metastasis and of vascular invasion in tumor borders, and ER tumor content were recorded. Monoclonal anti-Lam, anti-Coll IV were applied with the avidin-biotin-peroxidase complex and monoclonal anti ER with peroxidase anti-peroxidase complex, on frozen sections. A computerized system of image analysis referred to as SAMBA (TITN) with specific software for tissue sections analysis permitted a multiparametric quantitative analysis of immunostained surfaces. With this system, in each tumor, the cellularity, the percentage of Lam, Coll IV and receptor positive surfaces versus the total cell surface and versus the epithelial (keratin positive) surface, the integrated optical density IOD histograms were obtained and correlated to morphometrical and standard histological data. From this study, it was shown that: (1) Lam and Coll IV immunostained epithelial basement membranes in carcinomas were correlated to the presence of estrogen and progesterone receptor antigenic sites within the tumors, with a significant decrease of the positive staining in ER-ICA negative tumors in comparison to ER-ICA/PR-ICA positive tumors. (2) The combined densitometric and morphometric evaluation demonstrated a decrease of Lam and Coll IV immunostaining in malignant tumors, correlated to (i) the presence of peritumorous vascular invasion and (ii) keratin positive cells in bone marrow (iii) axillary lymph node involvement. It is concluded that the variations in Lam and Coll IV antigens distribution may be relevant indicators of tumor metastatic potential in breast carcinomas and that computerized image analysis enables the standardization of the evaluation antigens distribution.

Immunocytochemical assays in human endometrial carcinomas: a multiparametric computerized analysis and comparison with nonmalignant changes.

Immunocytochemical assay (ICAs) were performed on frozen sections from human endometrial samples (n = 89) including normal endometrium, decidua, hyperplasia with and without atypia, and carcinomas. Monoclonal antiestrogen receptor (ER), anti-laminin (Lam), anti-type IV collagen (Coll IV), and anti-Ki67 were applied with avidin-biotin-peroxidase complex or peroxidase-antiperoxidase complex. The results of the ICAs were evaluated through a computerized system of image analysis referred to as SAMBA. It was shown that this system provided for an accurate reliable and reproducible analysis of ICAs in tissue sections. It is concluded that this multiparametric and standardized method of analysis of ICAs can further be applied in correlations with clinical and biochemical data.

Immunodetection in fine-needle aspirates and multiparametric (SAMBA) image analysis. Receptors (monoclonal antiestrogen and antiprogesterone) and growth fraction (monoclonal Ki67) evaluation in breast carcinomas.

Immunocytochemical assays (ICA) using monoclonal antiestrogen receptors (ER ICA), antiprogesterone receptors (PR ICA), and monoclonal antibody Ki67 (Ki67 ICA) were performed in 127 breast carcinomas. The immunostaining procedures were applied on frozen tissue sections, tumour imprints, and fine-needle aspirates in order to compare the variations in the distribution of the antigens detected in the three different types of preparations. Positive reactions detected with peroxidase-antiperoxidase and avidinbiotin-peroxidase, and alkaline phosphatase-antialkaline phosphatase complexes were evaluated through a computerized system of image analysis referred to as SAMBA 200 (SAMBA TITN, Grenoble, France). Application programs specifically developed for the analysis of tissue sections and of cytologic preparations were applied. This system allowed a multiparametric, accurate, reliable, reproducible and automatized evaluation of the heterogeneity of the antigenic sites in tumors. For each markers positive cell surface (PS), and integrated and mean optical densities (IOD, MOD) and IOD histograms were compared. It was shown that (1) there was no significant variation in optical densities in cell imprints and aspirates whereas PS significantly (P less than 0.01) differed in both preparations; (2) there were significant differences of the optical densities between tissue sections and cytological preparations, either imprints or aspirates, likely due to randomly cut nuclei in tissue sections; and (3) there was a significant difference between the PS of tissue sections and aspirates but no significant difference between tissue sections and imprints. It is concluded that fine-needle aspiration constitutes a convenient method for cell sampling, reliable for the diagnosis of malignancies. However, it may not reflect the heterogeneity of cell subpopulations in tissue.

Type IV collagen immunostaining and computerized image analysis (SAMBA) in breast and endometrial disorders.

Type IV collagen immunostaining was performed on tissue sections from a large series of non-malignant and malignant disorders of the breast and endometrium. The results were analysed by means of a computerized system of image analysis referred to as SAMBA. It was shown that this system provided an accurate, reliable, reproducible, automated and multiparameteric analysis of collagen IV immunoprecipitates. It was concluded that this standardized method of analyses can be routinely used for the measurement of collagen IV, thus enabling correlations to be sought with histopathological and clinical data.

[Growth fraction (Ki67), ploidy balance and proliferation index in tumors of the urogenital tract and breast]. / Fraction de croissance (Ki67), balance de ploidie et index de prolifération dans les tumeurs du tractus urogénital et du sein.

The practical applications of computer-assisted image analysis systems are multiple in oncology. The computerized system of image analysis referred to as SAMBA (TITN) is particularly relevant to analyse coloured images resulting from immunostaining or histochemical procedures assessed either on tissue sections of any type or cytological preparations (imprints, smears). The SAMBA analysis of positive Ki67 surfaces in tissue sections from breast, endometrial, ovarian, cervical or urinary bladder samples enables a multiparametric evaluation of the growth fraction (GF) in intraepithelial, borderline or invasive proliferations. Moreover, the SAMBA analysis after Feulgen staining procedures provides a parametric evaluation of the nuclei densitometry and morphological features and of the chromatin texture, which serve to compute the ploidy balance (BP) and the proliferation index (PI). In benign tumors, GF and PI are low and tumor cells are diploid with an overall high positive PB values. In malignant tumors, GF, PI and the percentage of aneuploid cells increase with tumor grade and stage whereas PB decreases. In borderline proliferations, FC, PI and PB intermediate values are recorded. These new criteria of prognosis should be assessed routinely in pathology departments and the results from these new investigations are likely to be soon implicated in the selection of patient therapy.

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.

Multiparametric analysis (SAMBA 200) of the progesterone receptor immunocytochemical assay in nonmalignant and malignant breast disorders.

An immunocytochemical assay using monoclonal anti-progesterone receptor (PR-ICA) was performed in nonmalignant (N = 57) and malignant (N = 200) breast disorders. The results were analyzed with a computerized system of image analysis referred to as SAMBA and correlated with binding assays (DCC), and with standard histopathologic findings. It was shown that there was a correlation between 1) the PR-ICA and the binding assays (91.5%), and 2) between the binding assays and the multiparametric computerized (SAMBA) analysis of the PR-ICA. It was also shown that SAMBA provides an accurate, reliable, and reproducible evaluation of PR-ICA that is complementary to binding assays and constitutes a standardized method of evaluating the heterogeneity of the progesterone receptor (PR) distribution in tumors. It is concluded that SAMBA analysis of both the PR-ICA and estrogen receptor (ER-ICA) should improve the prognostic evaluation and the prediction of responsiveness to endocrine therapy in breast carcinomas.

[Immunohistochemical detection of estrogen receptors and image analysis (SAMBA 200) in breast carcinomas]. / Détection immunohistochimique des récepteurs oestrogéniques et analyse d'image (SAMBA 200) dans les carcinomes mammaires.

Estrogen receptor (ER) immunocytochemical assay (ER-ICA) was assessed human breast carcinomas. The patient's age, the tumor size, the histological type and SBR grade, the presence or the absence of axillary lymph node metastases and of vessel invasion in tumor borders were recorded in all the cases included in the series (n = 469) estrogen and progesterone receptors were concomitantly evaluated (DCC method) and or immunoenzymatic assays. Monoclonal H222 sp gamma and PAP procedure (Abbott kit) were applied in frozen sections, tumor imprints and fine needle aspirates. A computerized system of image analysis referred to as SAMBA (TITN), permitted to achieve a multiparametric quantitative analysis of ER positive surfaces. With this system, in each tumor, the cellularity, the percentage ER surface versus the total cell surface and versus the epithelial (keratin positive) surface, integrated optical density (IOD), mean optical density, index of the concentration of labelled objects, and IOD histograms, were obtained and correlated to histological and biochemical data. It was shown that: 1) ER antigenic sites were heterogeneously distributed in ER positive tumors, with a specific nuclear localization in epithelial cells; 2) the SAMBA 200 multiparametric analysis of the ER sites distribution in tissue was appropriate, accurate, reproductible and therefore more reliable than the semi-quantitative analysis; 3) the standardization and the complete automation of this method of immunoprecipitates evaluation on tissue section permit to daily and to routinely analyse a large number of preparation; 4) there was a correlation between ER binding sites evaluation (DCC) and ER antigenic sites immunodetection (ER-ICA and ER-EIA); 5) there was a correlation between the SAMBA evaluation of ER-ICA and other histological prognostic factors such as small tumor size, low SBR grade, the absence of axillary lymph node metastasis and vessel invasion; 6) the preliminary SAMBA analysis of ER-ICA in tissue sections, imprints and fine needle aspirates suggest that fine needle aspirates may not reflect accurately the tumor cell heterogeneity.

[Microphotometric system of analysis with automatic scanning and immunodetection: application to tissue sections (breast, endometrium, uterine cervix, ovary]. / Système d'analyse microphotométrique à balayage automatique (SAMBA) et immunodétection: applications sur coupes de tissu (sein, endomètre, col utérin, ovaire).

In the present study the immunodetection of various markers in a large series of tissue samples have been analysed through the computerized system of image analysis referred to as SAMBA 200. The immunodetection of estrogen and progesterone receptors, Laminin and type IV collagen, Ki 67, keratin, papilloma virus have been performed in frozen sections or in imprints and fine needle aspirates from tumorous and non tumorous disorders of human breast, endometrium, cervix and ovaries. Staining procedures varied with the type of the marker studied (PAP, ABC-P, ABC-GO, APAAP). The results obtained by the multiparametric SAMBA analysis were accurate, reproducible and reliable. Therefore, the SAMBA analysis providing a standardized and automatic analysis of immunodetection performed in tissue sections can be routinely used in pathology laboratories particularly for the prognostic evaluation and for the prediction of hormonotherapy and chemotherapy response of patients with solid malignant disorders.