Fetal sex and preterm birth.

Rates of preterm birth vary between different populations and ethnic groups. Epidemiologic studies have suggested that the incidence of preterm birth is also higher in pregnancies carrying a male fetus; the male:female difference is greater in earlier preterm pregnancy. Placental or chorion trophoblast cells from pregnancies with a male fetus produced more pro-inflammatory TNFα in response to LPS stimulation and less anti-inflammatory IL-10 and granulocyte colony stimulating factor (G-CSF) than cells from pregnancies with a female fetus, more prostaglandin synthase (PTGS-2) and less prostaglandin dehydrogenase (PGDH). These results suggest that in the presence of a male fetus the trophoblast has the potential to generate a more pro-inflammatory environment. Maturation of the fetal hypothalamic-pituitary-adrenal axis and expression of placental genes, particularly 11ß hydroxysteroid dehydrogenase-2 are also expressed in a sex dependent manner, consistent with the sex-biasing influences on gene networks. Sex differences in these activities may affect clinical outcomes of pre- and post-dates pregnancies and fetal/newborn wellbeing. These factors need consideration in studies of placental function and in the development of personalized strategies for the diagnosis of preterm labor and postnatal health.

Prospective comparison of endoscopic ultrasound-guided fine-needle aspiration and surgical histology in upper gastrointestinal submucosal tumors.

STUDY AIM: To assess the accuracy of ultrasound-guided fine-needle aspiration biopsy in the differential diagnosis of gastrointestinal stroma cell tumors (GIST) from other submucosal tumors, using both cytology and histology. PATIENTS AND METHODS: We conducted a prospective study from May 2005 to September 2008 in all patients presenting with upper gastrointestinal submucosal tumors. Only patients in whom surgical resection was carried out were included in the final analysis. In cases of mesenchymal tumor, immunocytochemistry was attempted for further differentiation between GIST and non-GIST. Surgical histopathology served as the gold standard. RESULTS: A total of 47 patients were analyzable, with a final histologic diagnosis of 35 mesenchymal tumors. Sufficient tissue for conventional cytologic diagnosis was obtained only in the 35 patients with mesenchymal tumors; in this subgroup, immunocytochemistry was possible in 46 %. If and only if enough material was available for immunocytochemistry, the sensitivity for (correct recognition of) GIST tumors was 93 %. In all 12 patients with nonmesenchymal tumors and lesions, cytology was nondiagnostic and the diagnosis had to be based on clinical suspicion and the appearance on endoscopy and endoscopic ultrasound (EUS). On an intention-to-diagnose basis, endoscopic ultrasound-guided fine-needle aspiration (EUS-FNA) had a positive predictive value for mesenchymal tumors of 100 %, but no value for the diagnosis of other lesions; using immunocytochemistry, a GIST tumor was recognized among the mesenchymal tumors with a sensitivity of 58 % and a specificity of 8 %. CONCLUSIONS: EUS-FNA-based cytology is safe and has only limited value for the differential diagnosis of submucosal tumors, mainly because insufficient material is harvested. Better tissue acquisition techniques are necessary for better differential diagnosis.

[Immunocytochemical identification of carcinomas of unknown primaries on fine-needle-aspiration-biopsies]. / Immunzytochemische Abklärung des initialen CUP-Syndroms an Feinnadelpunktaten.

Clinical investigations with imaging- and endoscopic techniques in order to identify the primary tumor sites in patients with CUP syndrome generally entail a significant diagnostic effort. If costs exceed 800.00, a financial loss ensues for German hospitals, as public health insurance companies do not reimburse above this amount. Combined cytological/immunocytochemical investigation of metastatic cancer cells represents a cost-effective, minimally invasive procedure to identify the probable primary cancer site that can be applied on an outpatient basis. We report on 85 fine needle aspiration biopsies of metastases to the liver, 30 to the lymph nodes and over 180 serous effusions and/or ascites with metastatic cancer cells in CUP. After conventional microscopic inspection, a routine panel of six monoclonal antibodies was applied (CK 5/6, CK 7, CK 20, Cdx 2, TTF 1 and CA 125). We were thus able to correctly identify the primary tumor sites in 90.3%, 92.0% and 85.1%, respectively, within three days. In total, 23 primary hepatocellular carcinomas could all be classified correctly, applying the antibodies HepPar 1, BerEp 4, AFP, CD 31, CD 68 and Ki 67. In addition, 141 malignant epithelial mesotheliomas were typed correctly in 97.1%, using the antibodies BerEp 4, Calretinin, Mesothelin, EMA and WT. Therefore, immunocytochemical investigation of metastatic cancer cells from fine needle aspiration biopsies or in serous effusions offers an efficient, cost-effective diagnostic alternative to imaging and endoscopic techniques in the workup of patients with CUP syndrome.

Innate immune defences in the human uterus during pregnancy.

The prevention of uterine infection is critical to appropriate fetal development and term delivery. The innate immune system is one component of the uterine environment and has a role in prevention of uterine infection. Natural antimicrobials are innate immune molecules with anti-bacterial, anti-viral and anti-fungal activity. We discuss two groups of natural antimicrobials in relation to pregnancy: (i) the defensins; and (ii) the whey acidic protein motif containing proteins, secretory leukocyte protease inhibitor (SLPI) and elafin. Human beta-defensins (HBD) 1-3 are expressed by placental and chorion trophoblast, amnion epithelium and decidua in term and preterm pregnancy. Elafin shows a similar pattern of localisation while SLPI is produced only by amnion epithelium and decidua. Evidence suggests that there is aberrant production of some natural antimicrobials in pathologic conditions of pregnancy. In preterm premature rupture of membranes (PPROM) levels of SLPI and elafin are reduced in amniotic fluid and fetal membranes, respectively. Elafin and HBD3 increase in chorioamnionitis and levels of the alpha-defensins, HNP1-3, increase in maternal plasma and amniotic fluid in women affected by microbial invasion of the uterus. In vitro culture studies have suggested a mechanism for increased production of natural antimicrobials in chorioamnionitis. Elafin, SLPI, HBD2 and 3 are all upregulated by inflammatory molecules in cells derived from gestational tissues. In summary, production of natural antimicrobials at key sites within the pregnant uterus suggests an important role in prevention of uterine infection during pregnancy and labour. Aberrant production of these molecules in PPROM and chorioamnionitis suggests that they also have a role in pathologic conditions. In particular, upregulation of these molecules by inflammatory molecules present in chorioamnionitis will ensure a robust response to infection.

[Quantitative methylation-specific PCR for the diagnosis of lung cancer]. / Quantitative methylierungsspezifische PCR zur Lungenkarzinomdiagnostik.

Efficient, preferably early diagnosis of lung cancer represents a major challenge. Under this aspect the sensitivity of conventional histomorphology and cytomorphology procedures is unsatisfactory. This review highlights technical aspects, possibilities and drawbacks of the application of aberrant promoter methylation as a biomarker for lung cancer diagnostics using specimens of pulmonary exfoliative cytology.

Towards fully automatic acquisition of multimodal cytopathological microscopy images with autofocus and scene managing.

OBJECTIVES: To increase the chance for a cure, cancer must be detected as early as possible. This can be achieved with cytopathological diagnostic methods. For a further increase of the diagnostic accuracy of these methods we introduced the multimodal cell analysis, viz, cells on the slide have to be relocalized to enable successive analysis of identical cells in different stains. For practical reasons the relocalization step must be automated. METHODS: For a fully automatic acquisition of successive cell images we use a passive autofocus that is adaptive to the material, i.e., to the cells, followed by a comparison of the scenes, i.e., the cell constellation, of two such obtained images from different stains. In case that no sub-scene match can be found the search is extended to the surrounding area. A set of 1556 scenes from seven specimens have been subject to our algorithm. The automatically relocalized and acquired images from a second stain have been manually compared to the images from a first stain. RESULTS: An overall relocalization rate of 85.4% is achieved. 14.3% of the images could not be relocalized and are lost for the following diagnostic process, while the critical case of erroneously matched images was observed in only 0.3% of cases. CONCLUSIONS: We could show that it is possible to automatically acquire images of successive stains of identical cells on cytopathological specimens. The method presented achieves acceptable relocalization rates. Wrong image acquisitions are very rare and can mostly be ascribed to images with single cells, i.e., without scene information.

Expression of natural antimicrobials by human placenta and fetal membranes.

Preterm birth associated with infection is a major clinical problem. We hypothesized that this condition is associated with altered expression of natural antimicrobial molecules (beta-defensins (HBD), elafin). Therefore, we examined expression of these molecules and their regulation by proinflammatory cytokines in placentae and fetal membranes from term pregnancy. HBD1-3 and elafin were localized by immunohistochemistry in fetal membranes and placenta. Real-time quantitative PCR was used to examine mRNA expression in primary trophoblast cells treated with inflammatory molecules. HBD1-3 and elafin were immunolocalized to placental and chorion trophoblast layers of fetal membranes and placenta. Immunoreactivity was also observed in amnion epithelium and decidua. No differences were noted between samples from women who were not in labour compared to those in active labour. In in vitro cultures of primary trophoblast cells, HBD2 and elafin mRNA expression was upregulated by the proinflammatory cytokine, IL-1beta. These results suggest that the chorion and placental trophoblast layers may be key barriers to the progression of infection in the pregnant uterus. Natural antimicrobial expression may be altered in response to inflammatory mediator expression associated with the onset of labour and/or uterine infection, providing increased protection when the uterus may be particularly susceptible to infection.

[Bilateral large conjunctival tumours as primary manifestation of sarcoidosis--successful treatment with steroid-depot-injections]. / Beidseitige ausgeprägte konjunktivale Tumoren als Primärmanifestation einer Sarkoidose -- erfolgreiche Therapie mit Steroid-Depot-Injektionen.

INTRODUCTION: Ocular manifestations of sarcoidosis vary enormously. They include the conjunctiva, lacrimal gland, orbita, intraocular structures and eye-lid, either isolated or combined. We describe a female patient who presented with unusually large, bilateral conjunctival tumours as a primary manifestation of sarcoidosis. PATIENT: A 79-year-old white woman was referred to us for further management of a persisting "conjunctivitis", which had been refractory to treatment with multiple medications. Initial examination disclosed swollen eye-lids and bilateral large hard tumours of the inferior fornix. The obtained brush smear, which was cytopathologically evaluated, revealed epitheloid cells and multinucleate giant cells. After 4 weeks she developed three reddish-brown maculopapular lesions on her face. The subsequent biopsy from the left inferior fornix and the skin showed histopathologically a granulomatous epitheloid cell inflammation without central necrosis and without acid-proof bacilli. Therefore a sarcoidosis was included into the differential diagnosis. The systemic evaluation revealed no other manifestation. At first we tried to reduce the chronically inflammatory tumours with different immunomodulating local treatment forms. Only the repeated intralesional injection of a steroid depot showed a complete disappearance of all conjunctival and skin tumours. CONCLUSION: An isolated bilateral primary manifestation of sarcoidosis with large massive conjunctival tumours is very rare and clinically not typical. The non-invasive, cytopathological examination by means of brush smears offers a new perspective in the fast diagnosis of conjunctival manifestation of sarcoidosis. The tumours respond excellently to the intralesional injection of steroid depots.

Prognostic impact of DNA-image-cytometry in neuroendocrine (carcinoid) tumours.

Establishing prognosis proves particularly difficult with neuroendocrine tumours (NETs) as a benign looking histology can be associated with a malignant behaviour. In order to identify prognostic factors we examined 44 gastrointestinal and pulmonary, paraffin-embedded NETs histologically and immunohistochemically. DNA-image-cytometry was used to examine 40 of these. We found that poor differentiation (corresponding to a Soga and Tazawa type D) and infiltrative growth correlated with a poorer prognosis. Moreover, parameters determined by diagnostic DNA cytometry like the 5c-exceeding rate, the 2c-deviation index, DNA-grade of malignancy, DNA-entropy and the type of DNA histogram were found to be of prognostic relevance. Morphometric parameters like the form factor and the mean nuclear area were relevant for survival, tumour recurrence and metastasis. However, in the multivariate analysis the only independent risk factor was the histological differentiation. The 5c-exceeding rate is a good objective risk factor, which can be used particularly in cases in which only a fine needle biopsy is available. Direct comparison of the histology and the 5c-exceeding rate in the multivariate analysis suggests that the 5c-exceeding rate taken as sole prognostic factor might be of higher prognostic relevance than the histology but larger studies are needed to confirm this.

[Noninvasive brush biopsy as an innovative tool for early detection of oral carcinomas]. / Nichtinvasive Bürstenbiopsie als innovative Methode in der Früherkennung des Mundhöhlenkarzinoms.

PURPOSE: The aim of this prospective study was to investigate the diagnostic accuracy of DNA image cytometry in combination with non-invasive brush biopsies taken from suspicious oral lesions. MATERIAL AND METHODS: Cytological diagnoses obtained from 1328 exfoliative smears of 332 different lesions were compared with histology and/or clinical follow-ups of the respective patients. Additionally, nuclear DNA contents were measured after Feulgen restaining using a TV image analysis system. DNA aneuploidy was assumed if abnormal DNA stemlines or cells with DNA content greater than 9c were observed. RESULTS; The sensitivity of our cytological diagnosis in addition to DNA image cytometry on oral smears for the detection of cancer cells was 97.8%, specificity 100%, positive predictive value 100%, and negative predictive value 98.1%. CONCLUSION: The application of DNA image cytometry with DNA aneuploidy as a marker for neoplastic transformation in oral smears secures cytologic diagnosis of carcinomas. Smears from brushings of all visible oral lesions are an easily practicable, cheap, noninvasive, painless, and safe screening method for detection of oral precancerous lesions and squamous cell carcinoma in all stages. We conclude that DNA image cytometry is a very sensitive and highly specific, objective, and reproducible adjuvant tool for identification of neoplastic cells in oral smears.

Spontaneous malignant transformation of conventional giant cell tumor.

Spontaneous malignant transformation of conventional giant cell tumor (GCT) of bone is exceedingly rare. We report on a case of GCT of the iliac crest in a 35-year-old woman with malignant change into a high-grade osteosarcoma 10 years after the first appearance of GCT on a radiograph. Since the patient refused therapy for personal reasons the tumor remained untreated until sarcomatous transformation occurred. Image cytometry showed DNA aneuploidy and a suspiciously high 2c deviation index (2cDI) in the primary bone lesion. A thorough review of the world literature revealed only seven fully documented cases of secondary malignant GCT which matched the definition of a "sarcomatous growth that occurs at the site of a previously documented benign giant cell tumor" and not treated by radiotherapy. These cases as well as the current one suggest that a spontaneous secondary malignant GCT presents as a frankly sarcomatous tumor in the form of an osteosarcoma or malignant fibrous histiocytoma. It usually appears at sites of typical GCTs-often without any recurrent intermediate state-and is diagnosed 3 or more years after the primary bone lesion. The prognosis is poor.

[Atopic keratoconjunctivitis: probably a risk factor for the development of conjuntival carcinoma]. / Atopische KeratokonjunktivitisWahrscheinlich ein Risikofaktor für die Entstehung von Bindehautkarzinomen.

BACKGROUND: Almost half of the patients with atopic dermatitis experience chronic inflammation of the eyelids, the conjunctiva and the cornea. Chronic inflammation is a possible cause for the development of malignancies, especially if associated with some kind of immunological defect as in atopic patients. So far, a correlation between atopic conjunctivitis and conjunctival malignancies has not yet been reported. Here, we present 7 atopic patients with conjunctival carcinoma or carcinoma in situ detected between February 2000 and August 2001. PATIENTS: All 7 patients had a long history of atopic dermatitis and chronic of inflammation of the conjunctiva and cornea. In all patients smears were examined by cytology and DNA cytometry. Furthermore, in 6 of the 7 patients a histopathological examination of conjunctival biopsies was performed. RESULTS: In 4 of the 7 patients invasive conjunctival carcinoma and in 2 carcinoma in situ were detected. Cytology and cytometry revealed conjunctival carcinoma or carcinoma in situ in the remaining patient. Histopathological examination could not be performed since the patient refused to have a conjunctival biopsy. CONCLUSIONS: These results suggest that atopic keratoconjunctivitis might be a risk factor for the development of conjunctival carcinoma.

Extracellular glutamate levels and neuropathology in cerebral white matter following repeated umbilical cord occlusion in the near term fetal sheep.

Umbilical cord occlusion causes fetal hypoxemia which can result in brain injury including damage to cerebral white matter. Excessive glutamate release may be involved in the damage process. This study examined the relation between extracellular glutamate levels in the cerebral white matter of the ovine fetus during and after intermittent umbilical cord occlusion and the degree of resultant fetal brain injury. Fetal sheep underwent surgery for chronic catheterisation and implantation of an intra-cerebral microdialysis probe at 130 days of gestation (term approximately 147 days). Four days after surgery (day 1), seven fetuses were subjected to 5x2 min umbilical cord occlusions, and on the following day (day 2) they were subjected to either 4 or 5x4 min umbilical cord occlusions; seven fetuses served as controls. Microdialysis samples were collected before, during and after the umbilical cord occlusions to determine extracellular glutamate levels in the cerebral white matter. Fetal blood gas status was measured and the fetal electrocorticogram was recorded continuously. During the periods of umbilical cord occlusions on both days 1 and 2, fetal arterial oxygen saturation, arterial partial pressure of oxygen and arterial pH decreased (P<0.05) while arterial partial pressure of carbon dioxide increased (P<0.05). All fetuses showed episodes of isoelectric electrocortical activity during umbilical cord occlusions on both days 1 and 2. In fetuses with patent microdialysis probes there were marked increases of glutamate efflux in the cerebral white matter following umbilical cord occlusion. Fetal brains were removed at autopsy on day 5 and subjected to histological assessment. Brain damage was observed in all fetuses exposed to cord occlusion, particularly in the periventricular white matter, with the most extensive damage occurring in the fetuses with the greatest increases in glutamate levels. We conclude that, in the unanesthetised fetus in utero, glutamatergic processes are associated with umbilical cord occlusion-induced brain damage in the cerebral white matter.