J. Venizelos

Human embryonal epithelial cells of the developing small intestinal crypts can express the Hodgkin-cell associated antigen Ki-1 (CD30) in spontaneous abortions during the first trimester of gestation

BackgroundKi-1 (CD30) antigen expression is not found on peripheral blood cells but its expression can be induced in vitro on T and B lymphocytes by viruses and lectins. Expression of CD30 in normal tissues is very limited, being restricted mainly to a subpopulation of large lymphoid cells; in particular, cells of the recently described anaplastic large cell lymphoma (ALCL), the Reed-Sternberg (RS) cells of Hodgkins lymphoma and scattered large parafollicular cells in normal lymphoid tissues. More recent reports have described CD30 expression in non-hematopoietic and malignant cells such as cultured human macrophages, human decidual cells, histiocytic neoplastic cells, mesothelioma cells, embryonal carcinoma and seminoma cells.ResultsWe investigated the immunohistochemical expression of CD30 antigen in 15 paraffin-embedded tissue samples representing small intestines from fetuses after spontaneous abortion in the 8th, 10th and 12th weeks using the monoclonal antibody Ki-1. Hormones had been administered to all our pregnant women to support gestation. In addition, a panel of monoclonal antibodies was used to identify leukocytes (CD45/LCA), B-lymphocytes (CD20/L-26) and T-lymphocytes (CD3). Our findings were correlated with those obtained simultaneously from intestinal tissue samples obtained from 15 fetuses after therapeutic or voluntary abortions.ConclusionsThe results showed that: (1) epithelial cells in the developing intestinal crypts express the CD30 (Ki-1) antigen; (2) CD30 expression in these epithelial cells is higher in cases of hormonal administration than in normal gestation. In the former cases (hormonal support of gestation) a mild mononuclear intraepithelial infiltrate composed of CD3 (T-marker)-positive cells accompanies the CD30-positive cells.

Imbalance of mononuclear cell infiltrates in the placental tissue from foetuses after spontaneous abortion versus therapeutic termination from 8th to 12th weeks of gestational age

Placental macrophages (Hofbauer cells) are located close to trophoblastic cells and foetal capillaries, which make them perfect candidates for involvement in regulatory processes within the villous core. Their capacity of producing several cytokines and prostaglandin-synthesising enzymes, and expressing vascular endothelial growth factor, indicate a possible role in placental development and angiogenesis in order to support pregnancy. Common cells to Hofbauer macrophages sharing similar cell surface markers (HLA-A, -B, -C and leukocyte common antigen) have been reported in the stroma, decidua and amnion, indicating additional foetal protection. Yet this is not always the case. Most spontaneous abortions occur before 12 weeks’ gestation, and most are due to chromosomal errors in the conceptus. Relatively few truly spontaneous abortions take place between 12 and 20 weeks’ gestation. Thereafter, between 20 and 30 weeks, another type of premature spontaneous termination becomes prevalent, which is due to ascending infection. The numbers of cells expressing the various markers of the monocytemacrophage lineage change throughout pregnancy. In the present study, we investigated the immunohistochemical expression of mononuclear infiltrations in paraffin-embedded placentas, from foetuses after spontaneous abortion (8th, 10th and 12th weeks of gestational age), and those after therapeutic abortion at the same time, using a panel of monoclonal antibodies for the identification of leukocytes (CD45/LCA), B-lymphocytes (CD20/L-26), T lymphocytes (CD45RO/UCHL1), CD68 and CD14 cells. Immunologic factors in human reproductive failure are plausible mechanisms of infertility and spontaneous abortion. Approximately 25% of cases of premature ovarian failure appear to result from an autoimmune aetiology. Unfortunately, current therapeutic options for these women are limited to exogenous hormone or gamete substitution. Local inflammations at the sites of endometriosis implants are postulated to mediate the pain and reduced fecundability associated with this clinical syndrome. The recruitment of immune cells, particularly monocytes and T-cells, neovascularisation around foci of invading peritoneal lesions, and the possible development of antiendometrial autoantibodies support an immunologic basis of this disorder. To date, treatment of pain and infertility associated with endometriosis is primarily surgical, although immune-based adjuvants are theoretical possibilities for the future. Finally, although hypotheses supporting immunologic mechanisms of recurrent pregnancy loss have been popular over the past decade, most clinical investigations in this area do not provide compelling evidence for this position. Reputable specialists in reproductive medicine use experimental immunotherapies judiciously in selected cases of repetitive abortion. For example, the use of anticoagulation therapy can be beneficial in cases with documented antiphospholipid antibodies. At present, however, efficacious immunotherapy protocols for general application have not been established. Despite these caveats, continued strides in our understanding of human reproductive immunology should yield considerable future progress in this field. During the physiological changes that occur in the first and in the beginning of the second trimester of pregnancy, spiral arteries of the placental bed are converted into the uteroplacental arteries. The essence of this conversion consists of losing the muscular elements in the vessel walls, making them unable to respond to vasomotor influences. Cells that infiltrate the walls of spiral arteries and replace their normal elements are called migratory, non-villous or intermediate trophoblastic cells. Besides infiltrating and replacing the anatomic structures of spiral arteries, intermediate trophoblastic cells also penetrate into the lumina of these vessels forming endovascular plugs. These plugs are one of the reasons why early uteroplacental blood flow cannot be visualised, even with transvaginal ultrasound, during the first 12 weeks of gestation. In uncomplicated pregnancies, the endovascular trophoblast is bound to disappear by the end of the second trimester of pregnancy, but the literature on this topic is scarce. Here we describe the detection, isolation and characterisation of CD45RO-, L26- and CD68/CD14-positive cells from human early pregnancy deciduas. These cells were found in close vicinity to endometrial glands, with preference to the basal layer of the decidua. We conclude that (1) maternal cells, apparently CD45RO/UCHL1-positive cells, cross the maternofoetal barrier and participate in spontaneous (involuntary) abortions, and (2) a small proportion of maternal cells (approximately 30%), apparently CD68/CD14-positive cells, also cross the maternal-foetal barrier and cause growth delay and recurrent reproductive failure. Further investigation of involvement of the intercellular adhesion molecules 1 and 2, platelet endothelial cell adhesion molecule, vascular cell adhesion molecule and E-selectin in leukocyte accumulation will be needed to support the passage of maternal cells to the foetus. The results were statistically significant (P<0.0001, Student’s t-test).

Intrahepatic Extramedullary Hematopoietic Tumor Mimicking Metastatic Carcinoma from a Colonic Primary

Background: Extramedullary hematopoiesis (EMH) is associated with a number of diseases in which the normal function of the bone marrow is disturbed. While organs with hemopoietic capacity like the liver and spleen are most commonly involved, EMH has also occasionally been found in other organs like the adrenal gland, lymph nodes, breast, thymus, small bowel and central nervous system. However, presentation of a myeloproliferative disorder, such as EMH in these organs is a rare event. Case Report: We report clinical and fine-needle aspiration (FNA) findings in a patient who presented with intrahepatic EMH which mimicked metastatic carcinoma from a colonic primary. Results: Ultrasound-guided FNA of the intrahepatic mass revealed megakaryocytes and myelocytes thus establishing the diagnosis of EMH. Conclusions: EMH is an unusual condition that can mimic other solid masses of the liver. Because radiologic findings are not specific, EMH should be considered in the differential diagnosis, especially in patients with a myeloproliferative disorder. FNA and subsequent cytopathological interpretation of the aspirates enables avoidance of unnecessary potentially hazardous surgery.

Prognostic significance of HLA-DR antigen in serous ovarian tumors

Abstract.The antigens encoded by the major histocompability complex (HLA-DR) are cell glycoproteins that play a fundamental role in the regulation of the immune response. The prognosis of ovarian cancer is dependent on the histological type and on the clinical stage at diagnosis. Our study reports the value of HLA-DR antigen as a prognostic marker of ovarian serous adenocarcinoma. We studied 31 cases of serous ovarian cystadenoma, 12 cases of serous ovarian borderline cystadenoma, and 39 cases of well-differentiated cystadenocarcinoma for HLA-DR monoclonal antigen. We also studied the T helper marker (CD4) in the tumor stroma of the relevant cases, given that it is now known that the dependence of immune responsiveness on the class II antigens reflects the central role of these molecules in presenting antigen to T helper cells. HLA-DR was expressed in 20 of 31 cystadenomas (64.5%), 4 of 12 borderline tumors (33.3%), and in 10 of 39 invasive carcinomas (25.6%). CD4 was expressed in 9 of 31 cystadenomas (29%), 5 of 12 borderline tumors (42%), and in 26 of 39 invasive carcinomas (67%). There was a statistically significant difference for the two examined antigens in cystadenomas (p<0.001) and invasive carcinomas (p<0.001), whereas there was no statistical difference in borderline tumors (p<0.5). The results showed decreased expression of HLA-DR and increased expression of CD4 as the lesion progressed to malignancy. The aberrant expression of HLA-DR by epithelial cells of cystadenomas, of borderline tumors, and of invasive adenocarcinomas agrees with the hypothesis of the adenoma/adenocarcinoma sequence. The immune attraction mechanism by low HLADR signaling seems to be of minor importance in the malignant and metastatic potential of serous ovarian tumors.

Neutrophil-rich Anaplastic Large Cell Lymphoma (NR-ALCL) Mimicking Lymphadenitis: A Study by Fine-Needle Aspiration Biopsy

A 45-year-old man presented to his practitioner with a 25day history of a painful tender nodule, measuring up to 2.5 cm in greatest dimension, in the right inguinal region. He had occasional complaints of pain down the anterior and lateral aspects of his right leg. No other palpable masses, skin lesions, or hepatosplenomegaly was identified. He was given a broad-spectrum antibiotic therapy, but the tenderness of the nodule did not resolve. Fifteen days later, the patient reported increased fatigue, but no fever, or recent weight loss. The patient was admitted for further analysis. Amongst the others, the laboratory tests for isolation of Chlamydia trachomatis (Frei test, complement-fixing antibodies, and microimmunofluorescence test for Chlamydia), were negative. Imaging studies did not show pelvic or elsewhere lymphadenopathy. A FNA biopsy on the inguinal nodule, using standard techniques with a 25-gauge needle, was performed. Airdried smears were stained with hemacolor and ethanolfixed smears were stained with Papanicolaou stain. The remaining fluid was centrifuged and cytospin preparations were made. A paraffin-embedded cell block was also prepared. The cytospin preparations and the direct smears were highly cellular consisting of small mature to large pleomorphic lymphoid cells intimately admixed with large numbers of neutrophils, representing up to 75% of all cells in some microscopic fields. Findings were suggestive of lymphadenitis, due to the intense presence of neutrophils. The cell block preparations showed an effacement of the architecture of the lymph node, with areas of suppurative necrosis surrounded by a cellular rim consisting of large-sized atypical lymphoid cells (Fig. 1). Higher magnification showed many of the atypical cells to have nuclei with a vesicular chromatin pattern and one or more prominent nucleoli. The surrounding cytoplasm was sometimes vacuolated. Numerous mitotic figures were identified. Classic Reed – Sternberg cells were scanty or absent. Immunocytochemical analysis performed on the cell block preparations revealed a CD30 and AKLpositive ALCL, T-cell type. Nuclear and cytoplasmic ALK-1 expression was seen in all tumor cells (Fig. 2), while CD30 showed a strong membranous staining with a paranuclear dot (Fig. 3). The atypical cells stained with antibodies for CD4/Leu-3a, CD43/Leu-22 (Bectin-Dickinson, San Jose, CA, USA), CD45/LCA, CD45RO/ UCHL-1, CD30/Ber-H2, and ALK protein/ALK-1 (Dako). Additional stains for CD3, CD15, CD20/L26, EMA/E29, kappa and Lambda polyclonal light chains (Dako) were negative. Most of the tumor cells showed nuclear p53 (Dako) staining and a high proliferative rate, confirmed by MIB-1/Ki67 staining of greater than 85%. The negative staining results with antibodies for S-100, Melan A/Klon A103, HMB-45/melanosome, cytokeratin coctail, myeloperoxidase and CD34 (Dako) effectively ruled out epithelial metastatic carcinoma, melanoma and granulocytic sarcoma respectively. Neutrophils were stained with CD15 but, as already stated, there was no staining of tumor cells with this antibody. The in situ hybridization for EBV, which was performed using a

RETRACTION: Intraoperative Touch Imprint Cytological Analysis of Sentinel Lymph Nodes for the Presence of Metastases in Breast Cancer

Background: Imprint cytology may provide a fast and accurate method for intraoperative screening of sentinel lymph nodes, so a decision can be made regarding whether to perform axillary clearance during primary surgery. If the findings are negative, in many cases axillary dissection can be omitted. Patients and Methods: 128 sentinel nodes from a cohort of 87 patients that had been identified using technetium-99m nanocolloid as a radioactive tracer and Patent blue dye were disected for rapid Diff-Quick stained touch preparations. Intraoperative evaluation of sentinel node status by imprint cytology was correlated with histopathological results of permanent sections. Tumor-negative nodes in routine paraffin sections were further investigated with the employment of an anti-cytokeratin antibody. Results: 36 of all sentinel nodes harbored metastases in the paraffin sections, of which 32 were identified by imprint cytology (sensitivity 88.8%). 3 sentinel nodes were positive by imprint cytology and negative by histopathology of the paraffin sections. Comparison of the results of the touch preparations with the final histopathology (hematoxylin-eosin and anticytokeratin antibody stains) demonstrated a sensitivity of 83.3% and a negative predictive value of 92.5%. The specificity and positive predictive value were 100% each. Conclusions: Touch imprint cytology is potentially useful for intraoperative evaluation of sentinel lymph nodes in breast cancer patients.

Metachronous extranodal marginal zone B-cell lymphoma of mucosa-associated lymphoid tissue (MALT lymphoma) and peripheral T-cell lymphoma unspecified: histologic, immunophenotypic, and molecular documentation.

Background: Coexistence of B- and T-cell lymphoid malignancies has been reported sporadically. Case Report: A 68-year-old woman developed a lymphoid neoplasm in the large intestine and a second lymphoid neoplasm in the esophagus, 24 months after the diagnosis of the first lymphoma. Immunophenotypic analyses were consistent with extranodal marginal zone B-cell mucosa-associated lymphoid tissue type (MALT type) and peripheral T-cell unspecified lymphomas in the large intestine and the esophagus, respectively. The molecular analysis confirmed the B-clonal genotype of the first lymphoma, and disclosed a biclonal genotype of the second one (composite T- and B-cell lymphoma). No evidence of Epstein- Barr virus (EBV) association was shown in either tumor. Conclusion: B- and T-cell neoplasms represent two distinct malignancies rather than progression of the same neoplastic clone.

ALK-positive neutrophil-rich variant of anaplastic large cell lymphoma diagnosed after head trauma

Background: Anaplastic lymphoma kinase (ALK) expression has not been described in neutrophil-rich anaplastic large cell lymphoma (NR-ALCL). Case Report: A 12-year old female with a 4-weeks history of a non-resolving bump over the forehead resulting from injury, was diagnosed of stage IE cutaneous T-cell lymphoma, and radiation was employed. Shortly after completion of therapy, there was progress of the disease on the soft tissue of the right hand, and bone marrow involvement was also found. A fine-needle aspiration of the hand mass was performed, and a diagnosis of CD30+/ALK+ NR-ALCL, was rendered. Methods: We studied the morphological characteristics of CD30+/ALK+ NR-ALCL using histological methods. A panel of antibodies were used to establish diagnosis and subtyping. In addition EBV status and molecular cytogenetics were determined. Conclusions: ALK-ALCL arising in the skin represents a single disease with a broad spectrum of morphology; clinicians and pathologists should be aware of this neutrophil-rich (NR) variant with aggressive clinical presentation.

Distribution of somatostatin in pancreatic ductal adenocarcinoma remodels the normal pattern of the protein during foetal pancreatic development: an immunohistochemical analysis.

Aim:To determine the immunoreactivity of somatostatin during the development of the human fetal pancreas and pancreatic ductal adenocarcinoma, given that, somatostatin-positive cells were demonstrated either into its embryonic anlage or into pancreatic cancer.Methods:Tissue sections from 15 pancreatic fetal specimens, and an equal number of ductal adenocarcinoma specimens were assessed.Results:The density of positive cells in the primitive exocrine ductal epithelium and endocrine epithelium was significantly different from the relevant density in the neoplastic pancreatic tissue of mixed (ductal-endocrine) and pure ductal type (P1=0.021 P2=0.001, P3<0.0001, P4=0.003 respectively). The above values were estimated from the 8th to 10th week. There was no significant difference in the density of positive cells in the mantle zone of the islets from the 13th to the 24th week, and the neoplastic tissue of mixed (P5=0.16) and pure ductal type (P6=0.65).Conclusion:The immunostaining for somatostatin identifies a subgroup of pancreatic ductal adenocarcinomas with a neuroendocrine component, (initially considered as pure ductal tumors), and mixed ductal and neuroendocrine tumors. This pattern of expression in neoplasms recapitulates the normal pattern during the embryonal development of the organ, raising the question of therapeutic efficacy of somatostatin and analogues as monotherapy in pancreatic cancer management.

Erratum to: NCL-CD30 staining of epithelial cells in the basal germinative layer of the epidermis and epithelial buds during foetal skin development

The fact that the CD30 molecule can mediate signals for cell proliferation or apoptosis prompted us to perform a systematic investigation of CD30 antigen expression in embryonal tissues during proliferation and differentiation stages. We first targeted the foetal human intestinal cryptae cells with positive results. The epidermis is a dynamic epithelium that is constantly renewed throughout life. The basal layer, attached to the basement membrane, contains the dividing cells of the skin and as cells move up from this layer they undergo differentiation, ending in the formation of a terminally differentiated anucleate cell called squame. It is intriguing to find out if cells in the basal layer can express the CD30 antigen. We investigated the immunohistochemical expression of CD30 antigen in 15 paraffin-embedded tissue samples representing epidermis and epidermal buds from foetuses after spontaneous abortion in the 8th, 10th and 12th weeks of gestation, respectively, using the monoclonal antibody NCL-CD30. A Northern blotting analysis was additionally performed. The results showed that: (1) the epithelial cells of the epidermis in the developing skin express the CD30 antigen; (2) CD30 expression in these epithelial cells is higher in cases of hormonal administration than in normal gestation; (3) a similar positive reaction involved the epidermal buds associated with the development of the skin appendages. Northern blots of tissue sections using a CD30 cDNA probe detected mRNAs of the same molecular mass and variety similarly to those in the positive control cell line HUT 102.