Methodological aspects of DNA image cytometry in formalin-fixed paraffin-embedded material from pancreatic adenocarcinoma

Deparaffinized and disintegrated material from conventionally formalin-fixed and paraffin-embedded surgical specimens of 100 cases of ductal adenocarcinoma of the pancreas was Feulgen-stained, and the cytochemical DNA distribution patterns of at least 100 single tumour cells and 50 "control" cells (fibrocytes) were assessed by means of image cytometry (ICM). In 77 cases a sufficient number of neoplastic cells could be obtained for these DNA assessments. The fairly high number (23) of cases that had to be excluded due to too small amounts of disintegrated cells or cell nuclei may be explained by the high content of connective tissue stroma in these pancreatic adenocarcinomas. The tumour cell nuclei in 76 of these 77 cases showed cytochemically a clear-cut "non-diploid" DNA distribution pattern. This observation reflects the well-known highly malignant growth potential of this carcinoma. Despite the fact that about 1/4 of the tumours had to be excluded, the main result of our methodological study is, after all that conventionally formalin-fixed paraffin-embedded specimens of most pancreatic adenocarcinomas can be successfully used for the deparaffinization-disintegration procedure preceding the nuclear DNA assessments by means of ICM. Additional studies are, however, required to obtain the diagnostic and prognostic impact of the results of such cytochemical analyses of the DNA distribution pattern in adenocarcinomas of the pancreas.     

Distribution of a formalin-resistant myelomonocytic antigen (L1) in human tissues. II. Normal and aberrant occurrence in various epithelia

L1 is an approximately 36-kd protein present in virtually all resting peripheral blood neutrophils and monocytes. It is particularly well preserved in formalin-fixed and paraffin-embedded routine material. In a recent immunohistochemical study, the authors showed that L1 has a restricted distribution within the monocyte-derived cell lineage, being mainly confined to reactive histiocytes (infiltrating macrophages). A protein sharing physicochemical and antigenic properties with L1 was identified in extracts of epidermal scales obtained from patients with psoriasis. This epithelial L1 was generally not expressed by normal epidermis, but its production was abundant in several skin diseases. Moreover, mucosal squamous epithelium normally expressed L1. No other epithelia showed signs of L1 production, although occasional patchy uptake was indicated, particularly in kidney tubular epithelium.     

CORTICAL SLOW POTENTIAL CHANGES IN MAN RELATED TO INTERSTIMULUS INTERVAL AND TO PRE TRIAL PREDICTION OF INTERSTIMULUS INTERVAL

Two experiments were performed to explore further the relationship between the cortical slow potential change known as the “contingent negative variation” (CNV) and the concept of “expectancy.” In Experiment I, 24 male Ss were presented click pairs, with inter-click intervals of 800, 1600 and 4800 msec (2 blocks of 10 trials each, counterbalanced between Ss for order), and instructed to press a key after the second click. Interval by order by trials analysis of variance showed interval to be the only significant factor: CNVs were lower and RTs longer as interval increased. In Experiment II, 8 female Ss given 60 pairs of clicks, 30 each with separations of 1200 and 2400 msec, were instructed to respond as in Experiment I, and were asked to make a pretrial prediction of the interval they would next receive. Analysis of variance of RTs showed that Ss responded slower when the interval was other than that predicted. Prediction by reception by subjects analysis of variance of CNV amplitude at the 1200 msec point gave a significant F only for prediction, mean amplitude for short being higher than for long. A similar design applied to CNV amplitudes at both the 1200 and 2400 msec points when Ss received the long interval yielded a significant measurement point by interval predicted interaction; at the 1200 msec point, short predictions were followed by higher CNVs than were long predictions; at 2400 msec, the opposite was found. These data combine with those already in the literature to indicate that the relationship between “expectancy” and the CNV is far from simple, and that cognitive and motivational factors play a significant role in determining CNV amplitude.     

White matter hyperintensities and their association with suicidality in depressed young adults

INTRODUCTION: Researchers and clinicians have increasingly recognized that biological markers may help identify patients who are at risk for suicide. The objective of this retrospective, cross-sectional study was to compare the prevalence and location of white matter hyperintensities (WMH) in young inpatients with major depressive disorder (MDD) with and without histories of suicide attempts. METHODS: T2-weighted magnetic resonance images (MRI) of 102 young psychiatric inpatients with MDD were rated for the presence of WMH. Medical charts were reviewed to ascertain history of suicide attempt, demographic and clinical variables. Fisher's Exact Tests and logistic regression modeling were used to test the association between WMH and suicidality. RESULTS: Bivariate analysis showed that the prevalence of periventricular WMH was significantly higher in subjects with past suicide attempts (Fisher's Exact Test, p=0.02). Logistic regression analyses controlling for age, sex, and several clinical risk factors supported this finding (odds ratio=5.7; 95% confidence interval: 1.6, 21.2). LIMITATIONS: Due to the retrospective, cross-sectional design of our study, we are unable to determine if the WMH preceded or followed past suicide attempts. The generalizability of our findings is limited since this group of inpatients is more severely ill than the general psychiatric population. CONCLUSIONS: The increased prevalence of periventricular WMH in young adults with MDD and a history of suicide attempt, compared to similarly depressed adults without such a history, is consistent with our findings in children and youth, and suggests there might be neurobiological in addition to psychosocial risk factors for suicide.     

Contribution of epitope specificity to the binding of monoclonal antibodies to the capsule of Cryptococcus neoformans and the soluble form of its major polysaccharide,glucuronoxylomannan

Incubation of encapsulated cryptococci with monoclonal antibodies (MAbs) specific for glucuronoxylomannan (GXM), the major capsular polysaccharide of Cryptococcus neoformans, produces two distinct capsular quellung-type reactions termed rim and puffy. The type of capsular reaction that occurs is determined by the epitope specificity of the MAb and the serotype of the yeast cell. Several biological activities, including opsonic activity, complement activation, and protective efficacy, are associated with the type of capsular reaction produced by a MAb. The goal of this study was to examine the reactivities of two families of anti-GXM MAbs with serotype A and D capsular polysaccharides in several immunochemical assays, including agglutination, immunofluorescence, quantitative precipitation, and enzyme-linked immunosorbent assay, in an effort to identify serological assays that are predictive of the capsular quellung reaction. The results showed that the type of capsular reaction (rim versus puffy) is a qualitative assessment of antibody-capsule interaction that cannot be predicted on the basis of a serological assay. The results further showed that antibody reactivity demonstrated in one serological assay is not necessarily predictive of results in another assay, particularly in cases where one assay examines antibody-capsule interactions, e.g., agglutination, and another assay examines interaction of antibody with soluble GXM. Taken together, the results suggest caution in interpretation of immunochemical assays for anti-GXM antibodies and recommend the use of multiple assays formats when studying anticryptococcal antibodies.     

Comparison of high-energy photon and electron dosimetry for various dosimetry protocols

The American Association of Physicists in Medicine Task Group 51 (TG-51) and the International Atomic Energy Agency (IAEA) published a new high-energy photon and electron dosimetry protocol, in 1999 and 2000, respectively. These protocols are based on the use of an ion chamber having an absorbed-dose to water calibration factor with a 60Co beam. These are different from the predecessors, the TG-21 and IAEA TRS-277 protocols, which require a 60Co exposure or air-kerma calibration factor. The purpose of this work is to present the dose comparison between various dosimetry protocols and the AAPM TG-51 protocol for clinical reference dosimetry of high-energy photon and electron beams. The absorbed-dose to water calculated according to the Japanese Association of Radiological Physics (JARP), International Atomic Energy Agency Technical Report Series No. 277 (IAEA TRS-277) and No. 398 (IAEA TRS-398) protocols is compared to that calculated using the TG-51 protocol. For various Farmer-type chambers in photon beams, TG-51 is found to predict 0.6-2.1% higher dose than JARP. Similarly, TG-51 is found to be higher by 0.7-1.7% than TRS-277. For electron beams TG-51 is higher than JARP by 1.5-3.8% and TRS-277 by 0.2-1.9%. The reasons for these differences are presented in terms of the cavity-gas calibration factor, Ngas, and a dose conversion factor, Fw, which converts the absorbed-dose to air in the chamber to the absorbed-dose to water. The ratio of cavity-gas calibration factors based on absorbed-dose to water calibration factors, N60Co(D,w), in TG-51 and cavity-gas calibration factors which are equivalent to absorbed-dose to air chamber factors, N(D,air), based on the IAEA TRS-381 protocol is 1.008 on average. However, the estimated uncertainty of the ratio between the two cavity-gas calibration factors is 0.9% (1 s.d.) and consequently, the observed difference of 0.8% is not significant. The absorbed-dose to water and exposure or air-kerma calibration factors are based on standards traceable to the National Institute of Standards and Technology (NIST). In contrast, the absorbed-dose to water determined with TRS-398 is in good agreement with TG-51 within about 0.5% for photon and electron beams.     

Comparison of colorimetric,fluorometric, and visual methods for determining anti-influenza (H1N1 and H3N2) virus activities and toxicities of compounds

Methods have been developed previously for rapid evaluation of compounds for antiviral activity in 96-well microplates, which include visual quantitation of antiviral activity based upon inhibition of virus-induced cytopathic effect (CPE) or by less subjective colorimetric or fluorometric means. In the present studies we compared a number of colorimetric (crystal violet, MTT [3-[4,5-dimethylthiazol-2-yl]-2,5-diphenyltetrazolium bromide], and neutral red) and fluorometric (Alamar Blue, bisbenzimide [Hoechst 33258], fluorescein diacetate, and rhodamine 6G) methods to visual scoring of antiviral activity in influenza A virus infections in Madin Darby canine kidney (MDCK) cells. Toxicity determinations using these same methods were also made for anti-influenza inhibitors and other compounds known to inhibit cell proliferation. Against influenza A/Texas/36/91 (H1N1) and A/Sydney/05/97 (H3N2) viruses, visual scoring and dye or stain methods produced results that were not significantly different from each other in deriving 50% virus-inhibitory concentrations (EC(50) values) for six anti-influenza compounds (amantadine, rimantadine, ribavirin, RWJ-270201 [BCX-1812], oseltamivir carboxylate, and zanamivir), with the exception of Alamar Blue which quantified lower EC(50) values than expected. In uninfected replicating cells, the visual and Alamar Blue methods underestimated the 50% cytotoxic concentrations (IC(50) values) of ribavirin, 1-beta-D-arabinofuranosylcytosine, and 6-azauridine, but more accurately assessed the toxicities of amantadine, rimantadine, and cycloheximide. Visual scoring, coupled with the use of one of these dyes or stains except Alamar Blue, can be used to accurately and rapidly quantify the anti-influenza virus activities and toxicities of potential new influenza virus inhibitors. These methods should also be applicable to evaluating antiviral effects against other lytic virus infections.