Inter-laboratory validation of PCR-based detection of Mycobacterium tuberculosis in formalin-fixed, paraffin-embedded tissues

The present study is based on the initiative for quality assurance in pathology of the German Society of Pathology and the Professional Association of German Pathologists. Four panel laboratories with experience and expertise in polymerase chain reaction (PCR) detection of Mycobacterium tuberculosis were selected to establish the prerequisites for continuous external laboratory trials, in particular, by providing pre-tested specimens and evaluation criteria for participating institutes. In the first step, the four panel laboratories performed an internal trial to test their own reliability and reproducibility. Paraffin sections and DNA preparations from 34 tissues (25 clinical specimens and 9 controls) totalling to 66 samples were evaluated by each panel institute according to their own protocols. The methodologies differed and are described in detail. Despite these differences, a high degree of inter-laboratory reliability was achieved. In this report, we summarise our results including the correlation with the histology and provide recommendations for applying PCR-based methodology for the detection of mycobacterial DNA in surgical specimens. Supplementary data are available online at (rubric Forschung). Pre-tested specimens are now available for the external trial and can be ordered from the steering institute via Oligene (). All molecular pathology laboratories are invited to participate in this quality assurance initiative.     

Absence of peripheral clonal deletion and anergy in immune responses of T cell-reconstituted athymic mice

Superantigens induce clonal deletion of reactive T cells in the thymus and clonal deletion and anergy in the periphery of euthymic mice. In this report we have assessed the ability of Staphylococcal enterotoxin B (SEB) to induce peripheral tolerance in nude mice reconstituted with normal, syngeneic T cells. Immunization of reconstituted nude mice with SEB resulted in lethal toxic shock in a large fraction of the animals. Such lethality was never observed in the normal donor mouse strain. Analysis of lymphokine production in response to SEB showed that reconstituted nude mice produced higher levels of interleukin-2 and tumor necrosis factor-α, but lower levels of interleukin-4, than euthymic control mice. Furthermore, SEB was unable to promote either clonal elimination or induction of anergy in the SEB-responsive peripheral T cells, despite the fact that reconstituted nude mice did produce high levels of corticosterone upon treatment with SEB. These results imply a lack of control over immune responses to superantigen in T cell-reconstituted athymic mice.     

Regulation of hepatocyte activator inhibitor-1 expression by androgen and oncogenic transformation in the prostate

Hepatocyte activator inhibitor-1 (HAI-1) is a transmembrane serine protease inhibitor that regulates the conversion of latent to active hepatocyte growth factor (HGF). Studies supporting a role for the HGF pathway in prostate carcinogenesis prompted an analysis of HAI-1 expression in the prostate. Here we analyze the regulation of HAI-1 expression by androgen, oncogenic transformation, and cancer progression. Immunohistochemical analysis revealed that HAI-1 expression was restricted to prostate epithelium, where staining occurred primarily in basal and atrophic luminal epithelial cells. Compared to normal glands, HAI-1 expression was significantly increased in localized prostate cancer and was present in most prostate cancer metastases. HAI-1 protein expression levels were sensitive to androgen in normal epithelium but not in cancer. Although androgen did not increase HAI-1 protein expression levels in LNCaP cells, it decreased HAI-1 surface expression, consistent with previous data from our group (Martin DB, Gifford DR, Wright ME, Keller A, Yi E, Goodlett DR, Aebersold R, Nelson PS: Quantitative proteomic analysis of proteins released by neoplastic prostate epithelium. Cancer Res 2004, 64:347-355). HAI-1 overexpression in cancer was predictive of prostate-specific antigen recurrence (relative risk, 1.24). These results suggest that HAI-1 regulates the HGF Met axis on prostate epithelial cells and influences HGF mediated tumor invasion and metastasis.     

Limiting factors for cytopathological diagnosis of high-grade squamous intraepithelial lesions: a cytohistological correlation between findings in cervical smears and loop electrical excision procedure.

The present study sought possible factors leading to the cytological diagnosis of atypical squamous cells of uncertain significance (ASCUS) in cases of high-grade squamous intraepithelial lesions (HSIL). Based on retrospective histopathological analysis of loop electrical excision procedure (LEEP) products that diagnosed HSIL, two study groups were randomly selected. The first was consisted of cases with two consecutive Papanicolaou (Pap) smears with the diagnosis of ASCUS. The second (control) group was represented by cases diagnosed as HSIL by cytology. From the Pap smears diagnosed as ASCUS, the sampling limitations was different from control group (P < 0.05). The median size of the largest lesion in each case with ASCUS was 2.66 mm (+/- 1.71 mm). In the control group, the median size of the largest lesion was 5.15 mm (+/-2.58 mm) (P < 0.05). The size of the lesion and sample limitations led patients with cervical intraepithelial neoplasms to be diagnosed as ASCUS for two consecutive times, after a 6-mo period.     

Persistent efficacy of topical eprinomectin against nematode parasites in cattle

Six studies were conducted to evaluate the persistent efficacy of eprinomectin pour-on against experimental challenges with infective nematode larvae in calves. In each study, calves were randomly assigned to one untreated group and up to four test groups, which were treated with eprinomectin at 500 μg/kg body weight at weekly intervals before single bolus challenge. The calves were necropsied approximately 4 weeks after challenge infection for nematode recovery. Eprinomectin pour-on provided ≥90% efficacy against challenge with Haemonchus placei, Trichostrongylus axei and T. colubriformis at 21 days after treatment and against Cooperia oncophora, C. punctata, C. surnabada, Dictyocaulus viviparus, Nematodirus helvetianus, Oesophagostomum radiatum and Ostertagia ostertagi at 28 days after treatment. Received: 14 April 2000 / Accepted: 18 May 2000     

Immunohistochemical characterization of fibroblast subpopulations in normal peritoneal tissue and in peritoneal dialysis-induced fibrosis

Peritoneal fibrosis is one of the most common morphological changes observed in continuous ambulatory peritoneal dialysis (CAPD) patients. Both resident fibroblasts and new fibroblast-like cells derived from the mesothelium by epithelial-to-mesenchymal transition are the main cells involved fibrogenesis. In order to establish markers of peritoneal impairment and pathogenic clues to explain the fibrogenic process, we conducted an immunohistochemical study focused on peritoneal fibroblasts. Parietal peritoneal biopsies were collected from four patient groups: normal controls (n=15), non-CAPD uremic patients (n=17), uremic patients on CAPD (n=27) and non-renal patients with inguinal hernia (n=12). To study myofibroblastic conversion of mesothelial cells, -smooth muscle actin (SMA), desmin, cytokeratins and E-cadherin were analyzed. The expression of CD34 by fibroblasts was also analyzed. Fibroblasts from controls and non-CAPD uremic patients showed expression of CD34, but no myofibroblastic or mesothelial markers. The opposite pattern was present during CAPD-related fibrosis. Expression of cytokeratins and E-cadherin by fibroblast-like cells and -SMA by mesothelial and stromal cells supports that mesothelial-to-myofibroblast transition occurs during CAPD. Loss of CD34 expression correlated with the degree of peritoneal fibrosis. The immunophenotype of fibroblasts varies during the progression of fibrosis. Myofibroblasts seem to derive from both activation of resident fibroblasts and local conversion of mesothelial cells.Manuel López-Cabrera and Rafael Selgas contributed equally to the article.     

Surgical treatment of portal hypertension in schistosomiasis

Patients with mansonic schistosomiasis and portal hypertension are usually young, with good liver functional reserve, huge splenomegaly, and hypersplenism detectable only by alterations seen in laboratory tests. Patients who are not operated on do not develop portosystemic encephalopathy (PSE). Angiographic alterations are characteristic and totally different from those observed in patients with liver atrophy or cirrhosis. Digestive tract hemorrhage is the most severe complication. The patient with schistosomiasis is, therefore, a good model for the study of the consequences caused by the surgical treatment of portal hypertension. The evaluation, however, of results published in the literature regarding schistosomiasis is practically impossible. In a review of 130 publications referring to 4,516 surgically treated patients, the absence of prospective or retrospective studies with adequate controls was observed, and only 3.2% of the patients were followed for 5 years. According to available data, a total of 29 different surgical techniques has been employed, 81.6% of which are represented by simple splenectomy, esophagogastric devascularization (EGDS), and splenorenal shunt (SRS). Splenectomy was the surgical procedure showing the highest rate of recurrent hemorrhage (54.3%), and the portocaval shunt had the highest PSE incidence (60%). Portosystemic encephalopathy incidence after SRS was 29%. Thus, in view of a lack of objective information, in 1977 our group started a prospective, randomized study comparing the results of the surgical procedures most widely used in our country: selective portal decompression (SPD), EGDS, and SRS. The randomization was interrupted after 94 patients had been operated on (32 SRS, 30 SPD, and 32 EGDS) because of a highly significant PSE incidence in the SRS group. Preliminary results as of January, 1984, refer to an average follow-up period of 53 months (minimum 29, maximum 77 months). In the SRS group, the late mortality rate was 18.7%, and the incidence of PSE was 31.2%. Among the patients included in this group, 18.7% had arterial hypertension. In the SPD group, late mortality occurred in 10% of the cases, and PSE in 13.3%, while a mild hyperbilirubinemia was seen in 43.3% of patients. Recurrent hemorrhage and ascites had an approximately similar incidence in the 3 groups.

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Resumen Los pacientes con esquistosomiasis mansónica e hipertensión portal generalmente son jóvenes con buena reserva funcional hepática, enorme esplenomegalia e hipersplenismo detectable solamente por las pruebas de laboratorio. Los pacientes nó operados no desarrollan encefalopatia portosistémica (EPS). Los cambios angiográficos son característicos y totalmente diferentes de los que se presentan en pacientes con cirrosis y atrofia hepática. La complicación más grave es la hemorragia digestiva. Por consiguiente, el paciente con esquistosomiasis es un buen modelo para el estudio de las consecuencias que se derivan del tratamiento quirúrgico de la hipertensión portal. Sinembargo, la evaluación de los resultados en la esquistosomiasis que han sido publicados en la literatura es prácticamente imposible. En una revisión de 130 publicaciones relativas a 4516 pacientes operados, se pudo comprobar ausencia de estudios prospectivos o retrospectivos con controles adecuados, y apenas el 3.2% de los casos fué seguido por 5 años. De acuerdo con los datos disponibles, se empleó un total de 29 técnicas quirúrgicas diferentes, 81.6% de los cuales estuvo representado por simple esplenectomía, devascularización esofagogástrica (DEG) y shunt espleno-renal (SER). La esplenectomía fué el procedimiento quirúrgico que tuvo la más alta incidencia de hemorragia recurrente (54.3%) y el shunt portacava la más alta incidencia de EPS (60%). La incidencia de EPS después de SER fué de 29%.Por consiguiente, en vista de la ausencia de información objetiva, nuestro grupo inición en 1977 un estudio prospectivo y aleatorizado para comparar los resultados de los procedimientos quirúrgicos mayormente utilizados en nuestro país: descompresión portal selectiva (DPS), DEG y SER. La aleatorización fué interrumpida después de haber operado los primeros 94 pacientes (32 SER, 30 DPS y 32 DEG) debido a una muy significativa incidencia de la EPS en el grupo con SER. Los resultados preliminares (enero de 1984) se refieren a un período promedio de seguimiento de 53 meses (mínimo 29, máximo 77 meses). En el grupo de SER la mortalidad tardía fué de 18.7% y la incidencia de EPS fué de 31.2%.El 18.7% de los pacientes incluidos en este grupo exhibió hipertensión arterial. En el grupo de DPS la mortalidad tardía fué de 10% y la incidencia de EPS de 13.3%; se observó hiperbilirrubinemia leve en 43.3% de los pacientes. La incidencia de hemorragia recurrente y de ascitis fué aproximadamente igual en los tres grupos.

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Résumé Les malades qui accusent une hypertension portale d'origine bilharzienne sont généralement jeunes. Leurs fonctions hépatiques sont bonnes mais ils présentent une splénomégalie très importante et une hypersplénisme qui se manifeste seulement par des altérations des tests biologiques. Lorsqu'ils ne sont pas opérés ils ne présentent pas de manifestations d'encéphalopathie. Les caractères angiographiques sont très particuliers et totalement différents de ceux qui sont observés chez les sujets dont le foie est atrophié ou cirrhotique. L'hémorragie digestive représente la complication la plus sévère. Le malade atteint de bilharziose hépatique est par conséquent un bon modèle d'étude des conséquences provoquées par le traitement chirurgical de l'hypertension portale. Cependant les résultats du traitement publiés dans la littérature sont ininterprétables. Dans une revue de 130 publications concernant 4516 opérations, l'absence d'études prospectives ou rétrospectives bénéficiant d'un contrôle de qualité a été constatée, 3,2% seulement des malades ayant été suivis pendant 5 ans. En tenant compte des données disponibles 29 techniques chirurgicales ont été employées, 81,6% desquelles sont repésentées par les opérations suivantes: splénectomie, devascularisation et anastomose spléno-rénale. La splénectomie a été suivie du taux le plus élevé de récidive hémorragique (54,3%) et l'anastomose portocave du taux le plus important d'encéphalopathie (60%), l'anastomose spléno-rénale n'étant suivie d'encéphalopathie que dans 29% des cas. En raison du défaut d'information objective notre équipe a entrepris en 1977 une étude prospective faite selon la méthode du choix au hasard des 3 techniques chirurgicales les plus employées au Brésil: l'anastomose spléno-rénale, la décompression portale sélective et la dévascularisation. L'étude fut interrompue alors que 94 malades avaient été opérés (32 anastomose spléno-rénales, 30 décompressions portales sélectives et 32 dévascularisations oesophagogastriques) en raison du taux particulièrement élevé de l'encéphalopathie postopératoire. Les résultats de cette étude menée jusqu'en janvier 1984 répondent à une période postopératoire moyenne de 53 mois (minimum 29, maximum 77 mois). Dans le groupe traité par anastomose spléno-rénale le taux de la mortalité tardive fut de 18,7% et celui de l'encéphalopathie de 31,2%. Parmi les malades de ce groupe, 18,7% présentaient une hypertension artérielle. Dans le groupe traité par décompression portale selective le taux de la mortalité tardive fut de 10%, celui de l'encéphalopathie de 13,3%, cependant qu'une hyperbilirubinémie fut constatée chez 43,3% de ces opères. La récidive hémorragique et l'ascite atteignent un taux très voisin pour les trois opérations.

     

Correction to: Longitudinal associations of physical fitness and affect with depression,anxiety and life satisfaction in adult women with fibromyalgia

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