Separate and variably shaped chromosome arm domains are disclosed by chromosome arm painting in human cell nuclei

Fluorescence in situ hybridization (FISH) with microdissection probes from human chromosomes 3 and 6 was applied to visualize arm and subregional band domains in human amniotic fluid cell nuclei. Confocal laser scanning microscopy and quantitative three-dimensional image analysis showed a pronounced variability of p- and q-arm domain arrangements and shapes. Apparent intermingling of neighbouring arm domains was limited to the domain surface. Three-dimensional distance measurements with pter and qter probes supported a high variability of chromosome territory folding.     

Histological and cytological criteria in the diagnosis of malignant melanomas by cryostat sections

Summary Although cryostat sections in general allow a distinction to be made between malignant melanomas and other pigmented lesions in clinically doubtful cases, the differential diagnosis may be difficult. The histological and cytological criteria taken into account can be classified as major, minor, and insufficient. Knowing the diagnostic value of each makes a conventionally established diagnosis safer. Variance analysis does not contribute to the problem but it can nevertheless be shown that the evaluation of six major criteria makes a quick and reliable cryostat section diagnosis possible. If these results are confirmed in a prospective study it would be a decisive step on the way to a quicker and safer cryostat section diagnosis of malignant melanoma, even for the less experienced histopathologist.The results published here were presented in part at the DDG meeting 1980 at Westerland/SyltWe are grateful to Miss Schubert, Institute of Biomathematics of the University of Munich, for the statistical evaluations     

Psychological reactions in patients with early rheumatoid arthritis

We report the interaction of RA and psychological factors over 2 years in a group of 89 patients with newly established disease. Short-time outcome regarding physical features was fairly good. Disease activity decreased, and disability evaluated by HAQ remained at a low level. Psychological distress as measured by the depression and anxiety subscales of SCL 90 (Symptom Check List) was not very pronounced and not related to disease state factors. A slight decrease of anxiety was recorded after 2 years. A new adjustment test was applied. It contained 13 items focused mainly on negative illness effects such as loss of independence, feelings of guilt, and change of social and leisure time activities. Three factors (regret of lost life values, dysphoric mood, and acceptance) explained 48% of the variance of the 13 items. The validity of the test was acceptable. The patients' degree of adjustment changed slowly or not at all during the 2 years.     

Further studies on the chronotherapy of asthma with inhaled steroids: The effect of dosage timing on drug efficacy

Background: Chronotherapy studies with inhaled corticosteroids have shown optimal therapeutic benefit when steroids are administered four times per day (QID) or once daily at 3 PM.Objective: This study evaluated whether more convenient once-daily dosage times (8 AM and 5:30 PM) produce improvement in asthma equivalent to QID.Methods: Efficacy outcome measures included FEV₁, peak expiratory flow rates, bronchial responsiveness, use of β₂-agonists, nocturnal awakenings, and responses to a quality of life questionnaire. Systemic effects were blood eosinophil count, cortisol level, 24-hour urinary cortisol, and evaluation for oral candidiasis and dysphonia.Results: Baseline measurements for all three treatment groups were similar. For morning peak expiratory flow rate, significant improvement was seen for the QID group (p = 0.001) and the 5:30 PM group (p = 0.003), but not the 8 AM group (p = 0.75). For evening peak expiratory flow rate, significant improvement was seen for the QID group (p = 0.005) and the 5:30 PM group (p = 0.01), but not for the 8 AM group (p = 0.47). There were significant improvements in all other outcome variables for each group except PC₂₀. There was a significant improvement in PC₂₀ only in the QID group. The systemic effects of the three regimens were comparable.Conclusion: Dosing of inhaled steroid at 5:30 PM had no increased systemic effects and produced efficacy similar to QID dosing. Dosing at 8 AM did not produce results consistently comparable to QID dosing. Optimal once-daily dosing of inhaled steroid is between 3 PM and 5:30 PM.     

Drosophila Cyclin B3 is required for female fertility and is dispensable for mitosis like Cyclin B

Cyclin B3 has been conserved during higher eukaryote evolution as evidenced by its identification in chicken, nematodes, and insects. We demonstrate that Cyclin B3 is present in addition to Cyclins A and B in mitotically proliferating cells and not detectable in endoreduplicating tissues of Drosophila embryos. Cyclin B3 is coimmunoprecipitated with Cdk1(Cdc2) but not with Cdk2(Cdc2c). It is degraded abruptly during mitosis like Cyclins A and B. In contrast to these latter cyclins, which accumulate predominantly in the cytoplasm during interphase, Cyclin B3 is a nuclear protein. Genetic analyses indicate functional redundancies. Double and triple mutant analyses demonstrate that Cyclins A, B, and B3 cooperate to regulate mitosis, but surprisingly single mutants reveal that neither Cyclin B3 nor Cyclin B is required for mitosis. However, both are required for female fertility and Cyclin B also for male fertility.     

External quality control assessment in PCR diagnostics of dengue virus infections

BACKGROUND: Increased travelling to countries endemic for dengue fever (DF) demands efficient laboratory diagnostics. Nucleic acid amplification techniques (NAT) are now frequently used for rapid diagnosis of imported viral diseases. Different PCR systems are available. OBJECTIVES: In order to assess the quality of molecular diagnostics of dengue virus infections, an external quality assurance (EQA) in PCR diagnostics was conducted. Study design: A panel of 10 human plasma samples was prepared and spiked with dengue virus types DEN-1 to DEN-4. In addition, a 10-fold dilution series (1:10-1:10(4) ) of DEN-3 virus was included. The panel was pre-tested by nested RT-PCR, in-house real-time PCR, and a commercial real-time PCR kit. The samples were inactivated by gamma irradiation and shipped in freeze dried state. Thirteen laboratories, within the European network for the diagnostics of imported viral diseases (ENIVD) took part using either single-round, nested, or real-time RT-PCR methods. Two laboratories used two methods in parallel, summarising up to 15 comparable results. RESULTS: 33-100% correct results were achieved. All laboratories detected DEN-2 correctly, followed by DEN-1 (14 positive results of 15), DEN-3 (12/15) and DEN-4 (11/15). Testing of the serial dilution revealed low sensitivity in many labs, with results ranging from 33 to 80% of correctly tested samples. CONCLUSION: The EQA gives a feedback of the quality of the RT-PCR system used by each respective laboratory. The different test systems and amplification conditions demonstrate the importance of external quality control measures.     

Visuomotor control within a distributed parieto-frontal network

The aim of this functional magnetic resonance imaging study was to investigate differences in visuomotor control with increasing task complexity. Twelve right-handed volunteers were asked to perform their signature under different degrees of visual control: internally generated movement with closed eyes, signing with open eyes, tracking the line of the projected signature forwards, and tracking the line of the projected signature backwards. There was a gradual onset and disappearance of activation within a distributed network. Parietal, lateral and medial frontal brain areas were activated during all conditions, confirming the involvement of a parieto-frontal system. The weight of activation shifted with increasing task complexity. Internally generated movements activated predominantly the inferior parietal lobule and the ventral premotor cortex, as well as the rostral cingulate area, pre-supplementary motor area (pre-SMA) and SMA proper. Opening the eyes reduced SMA and cingulate activation and activated increasingly the occipito-parietal areas with higher task complexity. Visually guided movements produced an activation predominantly in the superior parietal lobule and dorsal premotor cortex. This study bridges human activation studies with the results of neurophysiological studies with monkeys. It confirms a gradual transition of visuomotor control with increasing task complexity within a distributed parieto-frontal network.     

Reduction of FK-506 requirements by combination with polyethylene glycol superoxide dismutase in orthotopic rat liver transplantation

Reperfusion after ischemia results in endothelial cell injury and Kupffer cell activation. Inflammatory cytokines thus released can induce major histocompatibility complex antigens and increase the immunogenecity of the graft. An orthotopic rat liver allotransplant model was used to test the hypothesis that prevention of reperfusion injury by infusion of polyethylene glycol superoxide dismutase (PEG-SOD) would result in long-term allograft survival in the presence of subthreshold immunosuppressive dosages. ACI rats were used as donors, and Lewis strain rats as recipients. Orthotopic liver transplantation was initially performed to identify a subthreshold dose of the immunosuppressant FK-506, which would be unable to extend survival longer than control untreated rats with this strain combination. After testing three intramuscular FK-506 doses of 0.04, 0.08, and 0.16 mg/kg, it was observed that an FK-506 dose of 0.04 mg/kg/day for 14 days was unable to extend survival longer than in untreated recipients. This dose of FK-506 was used in combination with PEG-SOD at doses of 1000, 3000, 10,000, or 30,000 units. Recipient animals were treated intravenously with PEG-SOD as a loading dose to facilitate tissue penetration on day 1, and beginning on the day of transplantation, every 2 days for the duration of the study. Results of histologic studies and mean survival time were compared in untreated recipients and in rats treated with PEG-SOD plus 0.04 mg/kg/day FK-506. Mean survival time was increased significantly in these animals (p < 0.007) to 40.6 ± 25.6 days as compared with either untreated rats (10.0 ± 2.7 days) or rats treated with 0.04 mg/kg FK-506 alone (13.7 ± 4.2 days). Histologic examination demonstrated a significant reduction in the cellular infiltrate in rats treated with PEG-SOD plus FK-506, as compared with recipients treated with either agent alone or left untreated. Our results therefore suggest a potential approach to reducing immunosuppression in transplantation. (J ALLERGY CLIN IMMUNOL 1995;95:1276-81.)     

OH treatment of tetanus toxin reduces its susceptibility to limited proteolysis with more efficient presentation to specific T cells

At inflammatory sites, before their processing, antigens are exposed to oxygen free radicals released by activated cells. The effect of hydroxyl radicals (OH·) on the structure of a protein antigen, tetanus toxin (TT) was investigated, as well as the consequences on processing and presentation. A chemical system composed of Fe-EDTA, ascorbate and H₂O₂ was used to produce physiological amounts of OH^• radicals. TT exposed to OH· radicals presented a marked decrease of its intrinsic fluorescence with a concomitant increase of the content of bityrosine, but no fragmentation of the protein was detected by SDS-PAGE. Processing of the modified TT was analysed, by incubating TT at acidic pH with fractions enriched in plasma membranes and lysosomes obtained from a lymphoblastoid cell line (LCL). Proteolysis of OH·-treated TT was less important than proteolysis of native TT, especially upon prolonged incubations. Oxidized TT presented by LCL cells induced a greater proliferation of three different TT specific T cell clones, compared to native TT. When proteolytic digests of TT were presented by fixed LCL cells to a homologous T cell line, the proliferative response obtained in the presence of digests of OH·-treated TT was sustained, even in the case of prolonged proteolysis, whereas the response to digests of native TT fell rapidly. The relative resistance of OH·-treated TT to proteolysis appears thus responsible for its greater presentation to specific T cells, probably by protecting epitopes.