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The suprachiasmatic nuclei of the hypothalamus (SCN) are the master circadian clock in mammals. Transcriptional activity in this master clock has a marker in the immediate-early gene c-Fos. Within the SCN, distinct differences in c-Fos in the ventrolateral and the dorsomedial SCN have been reported for rodent species such as rats, mice, and hamsters. We studied C-FOS expression in the common vole (Microtus arvalis) SCN under LD 12:12 h and under constant dim light conditions. In the vole dorsomedial SCN, rhythmic C-FOS expression was seen in LD with a clear peak in the middle of the light period. Under constant dim light, we report constitutive, non-rhythmic expression of C-FOS in the dorsomedial SCN. This pattern is consistent with the circadian organization of behavioral activity, which is weak in voles and may be lost under constant dim-light conditions. In the ventrolateral SCN, we observed a rise in C-FOS expression under LD conditions prior to lights-on, followed by peak expression at lights-on. Another peak was seen at lights-off. In an additional experiment, we subjected animals to LD 16:8 to test the hypothesis that the dawn and dusk peaks in ventrolateral C-FOS expression change phase along with the photoperiod. The peak in C-FOS expression did not shift with the time of lights on, but remained at the same external time 6. The results are consistent with the interpretation that in the vole, c-Fos expression reports transcriptional activity associated more likely with an internal, gating process than with an external effect of light.

Ethylene is a strong controller of root development, and it has been suggested that it is involved in the increase of lateral root development in nutrient-rich soil patches (selective root placement). Here, this contention was tested by comparing selective root placement of an ethylene-insensitive transgenic tobacco (Nicotiana tabacum) genotype (Tetr) with that of wild-type (Wt) plants. Wt and Tetr plants were grown in pots with locally increased nitrate or phosphate concentrations, after which the root growth patterns were compared with those of plants grown in pots with homogeneous nutrient distribution. Tetr plants responded to nutrient patches in a similar way to Wt plants, by placing their roots preferentially at locations with higher nutrient content, and other aspects of plant morphology were also not greatly influenced by ethylene insensitivity. The response of both Wt and Tetr plants to high-nitrate patches was considerably stronger than to locally high phosphate, suggesting that the two responses are not linked in any functional or regulatory way. As the response to nutrient patches was similar in ethylene-sensing and ethylene-insensitive plants, it is concluded that selective root placement in response to nitrate or phosphate is, at least in tobacco, not mediated or modified by ethylene action.

The acquired activation of stem cell leukemia (SCL) during T lymphopoiesis is a common event in T-cell acute lymphoblastic leukemia (T-ALL). Here, we generated tamoxifen (TAM)-inducible transgenic mice (lck-ER(T2)-SCL) to study the consequences of acquired SCL activation during T-cell development. Aberrant activation of SCL in thymocytes resulted in the accumulation of immature CD4(+)CD8(+) (double-positive, DP) cells by preventing normal surface expression of the T-cell receptor alphabeta (TCRalphabeta) complex. SCL-induced immature DP cells were further characterized by up-regulated NOTCH1 and generated noncycling polyclonal CD8(+)TCRbeta(low) cells. The prevalence of these cells was SCL dependent because TAM withdrawal resulted in their disappearance. Furthermore, we observed that SCL activation led to a dramatic up-regulation of NOTCH1 target genes (Hes-1, Deltex1, and CD25) in thymocytes. Strikingly, NOTCH1 target gene up-regulation was already observed after short-term SCL induction, implying that enhanced NOTCH signaling is mediated by SCL and is not dependent on secondary genetic events. These data represent the basis for a novel pathway of SCL-induced leukemogenesis and provide a functional link between SCL and NOTCH1 during this process.

Objective: The aim of this study was to test the consistency (repeatability) of laboratory measures of food intake, including eating microstructure (cumulative food intake curves), in non-obese humans across four eating sessions, each separated by 1 week. A secondary aim was to test the effect of dietary restraint on the food intake of female participants., Methods: Mixed model analyses were used to compare average food intake across sessions, and intraclass correlation coefficients (ICC) were employed to compare within subject variability to total variance. High and low restraint females and low restraint males consumed four lunches in a Universal Eating Monitor (UEM) laboratory. The lunches consisted of one type of sandwich (chicken salad sandwich squares) or three types of sandwiches (chicken salad, ham, and turkey sandwich squares) presented in counterbalanced order., Results: Measures of food intake were stable for men and women, regardless of sandwich variety. In females, level of dietary restraint (high vs. low) did not significantly affect food intake., Discussion: The results indicate that eating behavior in the laboratory is sufficiently stable over time to justify evaluation of interventions designed to alter food intake using within subject designs.

Monoterpenes are an important class of terpenoids that are commonly present in plant essential oils. These can be extracted from plants and are used in the flavouring and perfumery industry. Monoterpene synthases are the key enzymes in monoterpene biosynthesis, as they catalyse the cyclisation of the ubiquitous geranyl diphosphate (GDP) to the specific monoterpene skeletons. Tobacco is one of the most studied model plants, it can easily and efficiently be transformed, and is a suitable model to study the release of plant volatiles. Thus, we have isolated monoterpene synthases from lemon, transformed tobacco with these cDNAs and have used human panelists to study the change in fragrance of the transgenic in comparison to the wild type plants. In a triangle test, we found that subjects were capable of smelling significant differences between leaf samples. However, as a result of variability in panel ratings, no significant difference between two sets of transgenic flowers and the wild type tobacco flowers was found for the generated attributes in a descriptive test.

Decorin is a small proteoglycan that binds to transforming growth factor-beta (TGF-beta) and inhibits its activity. However, its interaction with platelet-derived growth factor (PDGF), involved in arterial repair after injury, is not well characterized. The objectives of this study were to assess decorin-PDGF and decorin-PDGF receptor (PDGFR) interactions, the in vitro effects of decorin on PDGF-stimulated smooth muscle cell (SMC) functions and the in vivo effects of decorin overexpression on arterial repair in a rabbit carotid balloon-injury model. Decorin binding to PDGF was demonstrated by solid-phase binding and affinity cross-linking assays. Decorin potently inhibited PDGF-stimulated PDGFR phosphorylation. Pretreatment of rabbit aortic SMC with decorin significantly inhibited PDGF-stimulated cell migration, proliferation, and collagen synthesis. Decorin overexpression by adenoviral-mediated gene transfection in balloon-injured carotid arteries significantly decreased intimal cross-sectional area and collagen content by approximately 50% at 10 weeks compared to beta-galactosidase-transfected or balloon-injured, non-transfected controls. This study shows that decorin binds to PDGF and inhibits its stimulatory activity on SMCs by preventing PDGFR phosphorylation. Decorin overexpression reduces intimal hyperplasia and collagen content after arterial injury. Decorin may be an effective therapy for the prevention of intimal hyperplasia after balloon angioplasty.

Objectives: To assess sensory irritation levels from isopropanol (IPA) unconfounded by subjective evaluations of odor for comparison against the recommended exposure limits (400 ppm threshold limit value (TLV); American Conference of Governmental Industrial Hygienists)., Method: The lateralization method was used to assess intra-nasal irritation thresholds for IPA, while odor detection thresholds were also measured. Thresholds for 1-butanol and phenyl ethyl alcohol (PEA) were obtained as positive and negative irritant controls. To compare potency and hedonic characteristics, subjects provided subjective ratings of odor, irritation and annoyance intensity for three concentrations of each chemical. Workers occupationally exposed to IPA ( n=26) were compared with previously unexposed controls ( n=26)., Results: The (geometric) mean odor detection threshold for IPA was slightly higher among exposed workers than controls (39 ppm vs. 11 ppm). Lateralization thresholds measuring intra-nasal irritation were elevated when compared with controls (6,083 ppm in exposed workers vs. 3,361 ppm in naïve controls), with a significantly higher proportion of phlebotomists being unable to lateralize the maximum concentration regarded as safe, than controls. Calculations of the 6th percentile for lateralization thresholds revealed that 95% of the sample did not experience sensory irritation below 512 ppm. Thus, while odor detection thresholds were well below the current recommended exposure limits, the irritation thresholds were well above these values. The odor, irritation and annoyance from IPA was perceived, on average, as between weak and almost strong, from lowest to highest concentration., Conclusions: The results indicate that current exposure guidelines would be adequately protective of the acute adverse effect of nasal sensory irritation, as operationally defined by the intra-nasal lateralization threshold. Exposures to higher concentrations should perhaps be evaluated on the basis of existing knowledge about systemic, rather than local (e.g., irritation), toxic effects. IPA appears to be a weak sensory irritant and occupational exposure to IPA appears to elicit small changes in sensitivity that do not generalize to other odorants (e.g., PEA and 1-butanol) and are likely to be reversible.

Objective: Constrictive arterial remodeling following balloon angioplasty has been related to adventitial collagen accumulation and subsequent thickening and can be prevented by matrix metalloproteinase (MMP) inhibition. Following balloon dilation, we examined the effect of MMP inhibition on collagen turnover and the relationship between adventitial area and degree of constrictive remodeling., Methods: In 12 non-atherosclerotic landrace pigs, balloon dilation was performed in 39 peripheral arteries with and without MMP inhibition. Follow up with intravascular ultrasound was performed at 42 days. Collagen content was quantified using polarized light and digital image microscopy. Procollagen expression was determined using immunochemistry and Western blotting., Results: In the MMP inhibitor group, constrictive remodeling was inhibited at 42 days follow up. In control and MMP inhibitor groups, a positive relation was observed between adventitial thickness and degree of constrictive remodeling (P<0.001). Adventitial thickening and adventitial collagen content were reduced in the MMP inhibitor group (P=0.002 and P=0.001, respectively). Procollagen immunostaining, but not protein analysis on Western blotting, was decreased in the MMP inhibitor group., Conclusion: MMP inhibition impaired adventitial thickening by reduction of collagen content 42 days after balloon dilation. This might explain its inhibitory effect on constrictive remodeling.

Background: Chk1 kinase is activated by phosphorylation at serine-345 by Rad3 checkpoint kinase and is required for DNA damage checkpoint in late S and G2 phase of S. pombe cell cycle. We studied the ability of two chk1 mutants, chk1-1 and chk1-2, to undergo phosphorylation and to delay cell cycle progression in response to different types of DNA lesions., Results: Both the Chk1-1 and Chk1-2 mutant proteins are phosphorylated to various extents when DNA is damaged in early G2 phase of cell cycle by either UV irradiation or gamma irradiation. However, chk1-2 mutant does not delay cell cycle progression in a dose dependent manner specifically upon gamma irradiations. This defect is not associated with an important loss of survival. Furthermore, both chk1 mutants survive to Camptothecin treatment despite undetectable Chk1-1 or Chk1-2 phosphorylated forms. We show that both mutant proteins are not phosphorylated in cds1 devoid cells treated with ribonucleotide reductase inhibitor hydroxyurea or when the replisome is affected by a thermosensitive mutation in DNA polymerase delta. This inability is associated with the loss of checkpoint function. We found that an increased level of Crb2/Rhp9 protein specifically complements the defect of the chk1-1 mutant allowing Chk1-1 phosphorylation upon treatment with hydroxyurea of dcds1 cells., Conclusions: Mutants chk1-1 and chk1-2 behave differently according to the type of lesion generated on DNA.

GH promotes not only longitudinal growth in children but is active throughout life in protein, fat, and carbohydrate metabolism. The multiple actions of GH start when GH binds to the cell surface-expressed GH receptor. Effectiveness of the hormone depends both on its presence in the circulation and the availability of receptors at the cell surface of target cells. In this study, we examined the role of the ubiquitin-proteasome pathway in regulating GH receptor availability. We show that receptor turnover is rapid, and almost 3-fold prolonged in the internalization-deficient mutant GH receptor (F327A). Using a monovalent GH antagonist, B2036, we could quantify the internalization of the nonactivated receptor. By comparing internalization of the receptor with shedding of the GH-binding protein, we show that in Chinese hamster lung cell lines, internalization followed by lysosomal degradation is the major pathway for receptor degradation and that the ubiquitin-proteasome pathway controls this process. Inhibition of endocytosis resulted in a 200% increase in receptor availability at the cell surface at steady state.

In this study we have explored the potential of PUVB (8-MOP + UVB) therapy for the reduction of luminal narrowing after arterial injury. In 15 rabbits, balloon dilation of iliac arteries was performed. In 20 arteries, dilation was combined with the delivery of pulsed ultraviolet light B (UVB) irradiation with 10 arteries being previously subjected to sensitizer infusion. Changes in vessel diameter, proliferation and extracellular matrix protein content at 6 weeks were evaluated by means of angiography and histomorphometry-immunohistochemistry. We found that PUVB, applied at the time of dilation, induced reduction in late loss (LL) at 6 weeks (percutaneous transluminal angioplasty vs UVB vs PUVB: 0.64 +/- 0.15 mm vs 0.61 +/- 0.05 mm vs 0.29 +/- 0.05 mm; p = 0.018). The same holds true for constrictive remodeling (0.53 +/- 0.15 mm vs 0.45 +/- 0.06 mm vs 0.15 +/- 0.05 mm; p = 0.016). In the irradiation groups, LL was independent of acute gain (AG), as opposed to the control. Collagen content increased significantly after PUVB in media and adventitia, without increased cellular proliferation in all vessel layers. Thus, PUVB at the time of dilation reduced luminal narrowing at follow-up without effecting proliferation. This effect was independent of AG and was associated with increased collagen content in media and adventitia.

The term "tinea incognito" refers to dermatophyte infections with clinical presentations that have been modified by the administration of corticosteroids. We describe a patient who had pustular inflammatory skin lesions due to Trichophyton rubrum after receiving treatment with potent topical corticosteroid creams.

Objective: This study tested the hypothesis that women with anorexia nervosa (AN) have an inappropriately fatter body image in the left cerebral hemisphere (LH) than in the right cerebral hemisphere (RH)., Method: Women with AN symptomatology were compared with thin controls in a divided visual field experiment. Distorted and undistorted pictures of their own and someone else's body were flashed briefly in the left and right visual fields. Participants judged the pictures as thinner than, equal to, or fatter than the actual body size., Results: The AN participants judged a higher proportion of fatter distortions as equal to their own size. They responded faster when stimuli were presented initially to the LH than when they were presented initially to the RH. In contrast, fewer thinner distortions were judged as equal to their own body size, and were judged more slowly, on LH trials than on RH trials. Controls did not show hemispheric differences when judging their own body and AN participants did not show hemispheric differences when judging pictures of somebody else. Additional analyses revealed that these findings were carried entirely by a subgroup who had AN in the past, not by the subgroup who currently had AN., Discussion: The brain lateralization paradigm may prove useful in understanding body image disturbance in AN patients., (Copyright 2001 by John Wiley & Sons, Inc.)

With the recent availability of novel antibodies against melanoma antigens tyrosinase and MART-1, it is important to validate their usefulness in pathology practice and in screening patients for immunotherapy treatment. In the present study conducted by the Melanoma Cooperative Group of the European Organization for Research and Treatment of Cancer (EORTC-MCG), immunohistochemical staining for gp100 (antibodies NKI-beteb and HMB-45), MART-1 (A103), tyrosinase (T311), and S100 (S100) was compared on formalin-fixed and paraffin-embedded tumour lesions from 80 patients with 130 malignant melanoma lesions, comprising 44 primary tumours, 18 locoregional metastases, 41 lymph node metastases, and 27 visceral metastases from the lung, liver, and brain. A score between 0 and 5 was allocated to each immunohistochemically stained section. These scores were evaluated in a statistical analysis. S100 was by far the most sensitive marker in all four types of lesions tested. Apart from a significantly better performance for T311 in primary melanomas compared with HMB-45, no significant differences were observed between the four remaining antigens tested. Three settings were next investigated to determine whether the expression of melanoma antigens decreases with tumour progression. First, within the primary melanomas, only NKI-beteb and A103 staining showed a nearly significant negative correlation with Clark's level of invasion and a similar tendency was observed for these antibodies with Breslow thickness. Second, when comparing primary melanoma-metastasis pairs from the same patient, lymph node metastases showed less staining with NKI-beteb, HMB-45, A103, and T311, at a level near significance. This difference was not significant when comparing the primary tumour with visceral metastases, probably due to the lower numbers of pairs. Third, regarding tumour progression from primary melanoma to locoregional, to lymph node, to visceral metastasis, a significant decrease with progression was found only for T311. The apparently stable expression of most of the melanoma antigens, and the small contribution of decreased expression in melanoma tumour progression, supports the rationale for immunotherapy based on the melanoma immunogens gp100, MART-1, and tyrosinase.

The feasibility of adding both the multidrug resistance modulator cyclosporin (CsA) and granulocyte colony-stimulating factor (G-CSF) to a standard salvage regimen of idarubicin (IDA) and cytarabine was evaluated in patients with resistant or relapsed acute myeloid leukemia and myelodysplastic syndrome. Three patients received IDA 12 mg/m2/day, the next four patients 9 mg/m2/day. The dose of CsA was 16 mg/kg/day. Six patients showed Pgp expression and none MRP1 expression. Grade III or IV toxicity (CTC-NCIC criteria) was registered in six patients for gastrointestinal, two patients for cardiovascular and one patient for neurological complications. Three patients died in hypoplasia and three patients showed leukemic regrowth. Three control patients were treated with IDA 12 mg/m2/day and cytarabine, but no CsA and G-CSF. The plasma IDA and idarubicinol (ida-ol) area under the curve's of patients treated with IDA 12 mg/m2 plus CsA were higher (P< 0.05) than in controls. Cellular IDA concentrations were almost similar, but cellular ida-ol concentrations were significantly higher (P < 0.05) in the presence of CsA than in controls. We conclude that the toxicity either with IDA 12 or 9 mg/m2/day was too high. The modulating effect of CsA was mainly based on changes in plasma kinetics of IDA and ida-ol, although ida-ol cellular clearance was delayed in the presence of CsA.

Objective: To develop a prototype of the Body Morph Assessment (BMA), and to test the reliability and validity of this new measure of body image. The BMA is a realistic and relatively simple procedure that uses computer morphing for the assessment of body image. For the purposes of this preliminary study, a prototype of the BMA was developed for usage with white women ranging from very thin to obese., Research Methods and Procedures: A total of 72 subjects participated in tests of reliability, content, and convergent validity of the BMA., Results: The reliability and validity of the BMA was supported by the results of this study. In a test of convergent validity, the measures of current, ideal, and reasonable body size were positively correlated with their equivalents from a similar body image assessment procedure. In addition, reliability coefficients were found to be satisfactory for all variables. Participants found the human figural stimuli to be realistic., Discussion: These preliminary findings support the reliability and validity of the BMA with white women. Given these positive findings, we plan to extend the procedure to males and to other racial and ethnic groups.

Expression of the multidrug resistance (MDR) mechanisms P-glycoprotein (Pgp) and MDR-related protein (MRP) decrease cellular retention and consequently cytotoxicity of anthracyclines. MDR is expressed on normal human hematopoietic progenitors and leukemic blasts. Normal CD34(+) progenitors showed rhodamine efflux in 20% to 30% of the cells, which could be blocked by verapamil. These cells appeared noncycling, in contrast to the proliferating rhodamine bright (RhoB) cells. We postulated that MDR expression can be downregulated by proliferation induction. Triggering rhodamine dull (RhoD) CD34(+) cells to proliferate indeed resulted in a higher rhodamine retention and significantly decreased efflux modulation by verapamil (P =.04). Also in acute myeloid leukemia (AML), the proliferation rate (percentage S/G(2)+M and Iododeoxyuridine labelings index) was significantly less in the RhoD blasts (P

Aim of this research was to test the hypothesis that marginal gap measurements by a digital microscopic image processing computer are reproducible. Light microscopic images of the margins of Artglass partial crowns on stone dies were obtained with a 3CCD color video camera, digitized and processed. Ten measuring points were painted on a 0.5 mm section of the finish line of the preparation. The opposite corresponding measuring points on the margin of the partial crown were painted as well. The distance (marginal gap) between two corresponding points was painted and calculated by the computer. Reproducibility (precision) is reflected in the ability to reproduce the same measurement results in repeated measurements (n = 5). The reproducibility error was expressed as the coefficient of variation (CV) in percent (%). The localization of the corresponding measurement points and the calculation of the marginal gap were performed without errors. For a marginal gap between 15-50 microns the average measurement error was 3% or 0.45-1.5 microns. It was concluded that the measurement of the gap between two points by the digital image processing computer was reproducible. The measurement error was neglectable given that a clinically acceptable marginal gap is 100 microns.

The coronavirus membrane (M) protein is the key player in virion assembly. One of its functions is to mediate the incorporation of the spikes into the viral envelope. Heterotypic interactions between M and the spike (S) protein can be demonstrated by coimmunoprecipitation and by immunofluorescence colocalization, after coexpression of their genes in eukaryotic cells. Using these assays in a mutagenetic approach, we have mapped the domains in the M protein that are involved in complex formation between M and S. It appeared that the 25-residue luminally exposed amino-terminal domain of the M protein is not important for M-S interaction. A 15-residue deletion, the insertion of a His tag, and replacement of the ectodomain by that of another coronavirus M protein did not affect the ability of the M protein to associate with the S protein. However, complex formation was sensitive to changes in the transmembrane domains of this triple-spanning protein. Deletion of either the first two or the last two transmembrane domains, known not to affect the topology of the protein, led to a considerable decrease in complex formation, but association was not completely abrogated. Various effects of changes in the part of the M protein that is located at the cytoplasmic face of the membrane were observed. Deletions of the extreme carboxy-terminal tail appeared not to interfere with M-S complex formation. However, deletions in the amphipathic domain severely affected M-S interaction. Interestingly, changes in the amino-terminal and extreme carboxy-terminal domains of M, which did not disrupt the interaction with S, are known to be fatal to the ability of the protein to engage in virus particle formation (C. A. M. de Haan, L. Kuo, P. S. Masters, H. Vennema, and P. J. M. Rottier, J. Virol. 72:6838-6850, 1998). Apparently, the structural requirements of the M protein for virus particle assembly differ from the requirements for the formation of M-S complexes.

Currently available data regarding the substrate specificity of the multi-drug resistance (MDR) mechanisms P-glycoprotein (Pgp) and MDR-associated protein (MRP1) for idarubicin are inconclusive. A multiparameter flow cytometry method was developed which allows simultaneous quantitative measurement of total cellular fluorescence and the amount of anthracyclines intercalated into the DNA. Anthracycline DNA intercalation was measured by fluorescence resonance energy transfer (FRET) between Hoechst 33342 and anthracyclines. Daunorubicin and idarubicin accumulation were studied and compared in established cell lines expressing Pgp and MRP1. The data demonstrate that daunorubicin DNA intercalation is affected by both Pgp and MRP1 whereas idarubicin DNA intercalation is affected only by MRP1. MRP1 and Pgp function could be blocked completely by 5 microM PAK 104P, while higher concentrations of verapamil, PSC 833 and cyclosporin A were necessary to attain complete blocking of MRP1 compared to Pgp. Daunorubicin DNA intercalation correlates better with cell survival and is more sensitive at physiological MDR expression as observed in hematopoietic progenitors than daunorubicin levels measured by total cellular fluorescence. In conclusion, idarubicin DNA intercalation is reduced by MRP1 but not by Pgp. PAK-104P is an effective modulator for both Pgp and MRP1 and may further improve idarubicin efficacy.

Objective: To test the feasibility of the technique of "morphing" for body image assessment purposes and to investigate whether females with anorexia nervosa (AN) categorize other female's body sizes similarly to controls., Method: A morphing movie showing the transformation of a thin woman into an obese woman was presented. Females with AN and thin controls selected the sizes corresponding to the transitions between the body size categories thin, normal, fat, and obese. They were instructed to look at the woman in the movie as themselves (Self) versus someone else (Other)., Results: The AN group chose significantly lower transitions associated with the categories thin, normal, fat, and obese under the Self instruction than controls. Similarly, they selected lower settings for all but the obese category when looking at the body as someone else's. It was concluded that females with AN display a harsher judgment not only of their own body size and shape, but also of that of another woman, than women without an eating disorder., Discussion: The morphing instrument was found to be sufficiently realistic, detailed, and sensitive for its intended purposes.

This study investigated whether individuals with anorexia nervosa (AN) visualize themselves as fatter than they are because they perceive themselves as fatter. Females with AN who overestimated their own body size judged size differences between pictures of their own body, and then again of someone else's body. Signal detection analysis of the results showed no differences in perceptual sensitivity between the AN and normal and thin control groups. No significant correlations were found between body size estimates and perceptual sensitivity. The anorexic group did, however, show a bias to report seeing "thin" differences, which was opposite to that of thin controls. Because bias differences between the groups were significant while sensitivity differences were not, it was concluded that abnormalities of body image most probably arise during reconstruction of the visual body image, rather than during perception of the body.

Remission induction therapy fails in 20-30% of the patients with acute myeloid leukemia (AML) despite dose intensification and the use of new and more effective drugs. Primary and acquired drug resistance, metabolic or kinetic are a fundamental problem. Expression of the P-glycoprotein in AML is correlated with therapeutic outcome. Randomized clinical studies with Pgp modulators are currently on-going. Ara-C and anthracyclines, are preferentially cytotoxic to proliferating cells. Proliferation induction of leukemia blasts with growth factors in vitro resulted in an increased toxicity of Ara-C and anthracyclines. Normal hematopoietic blast cells with a high Pgp expression are noncycling and less sensitive to anthracyclines, in contrast to the more proliferating cells with a low Pgp expression. Proliferation induction by growth factors results in a down regulation of Pgp expression. Priming of leukemic cells with growth factors in vivo might be promising and randomized clinical studies are warranted.

Mobilized peripheral blood progenitor cells (PBPC) have been shown to differ qualitatively from bone marrow (BM) progenitors. The released progenitor cells are predominantly in G0/G1 and show a relatively high percentage of rhodamine dull cells. Within the BM these last two features are characteristic of the more primitive progenitors. Although the mobilized PB cells can give rise to long-term repopulation and thus contain stem cells, the frequency of stem cells is not much higher if long-term initiating cell (LTC-IC) assays are used. To determine whether quiescent stem cells are selectively released or the low-cycle status of PB progenitors is related to the release from the BM microenvironment, the cell cycle status and rhodamine content in the PB and BM during mobilization were studied and compared with steady-state BM. More differentiated and more primitive progenitors were separated based on differentiation markers and cloned in single cell assay. In mobilized PB 54% of the CD34+ cells (n=5) were rhodamine dull compared to 22% in steady-state BM (P=0.014) [n=6]. The percentage of CD34+ cells in the S/G2M phases of the cell cycle was 2.1% in the mobilized PB (n=11), and 18% in steady-state BM (n=11) [P=0.002]. During mobilization the fraction of cells in the S/G2M phase of the cell cycle was 16% in BM (n=7), similar to steady-state BM (P=0.34). The released progenitors represented a selection of BM progenitors, with significantly more primitive progenitors (CD34+/13+/33dim) and less lymphoid precursors (CD34+/19+). Within the more differentiated CD34+113+/33bright, myelomonocytic precursors, both in PB as well as in BM, the percentage S/G2M was relatively higher than in the CD34+/13+/33dim subfraction: in normal BM: median 18% vs 8% (P=0.006) [n=8]; in mobilized PB 3% vs 2% (P=0.03) [n=10]; and in BM during mobilization 24% vs 7% (P=0.01) [n=6]. The cycle status of mobilized PB progenitors was low both in the primitive and more differentiated subfractions. During the mobilization period the BM progenitors are cycling as in steady-state BM. The low-cycle status of the mobilized PB progenitors may be related to the loss of contact with the micro-environment.

A major problem that has dogged research into body image disturbance in anorexia nervosa concerns the diversity of measurement procedures employed in studies of body size estimation. Many researchers believe that results obtained with different procedures cannot meaningfully be compared with each other, because the methods used measure different aspects of the body image. A meta-analysis of 33 body size estimation studies was conducted to investigate whether methodological differences do indeed influence outcome. This analysis revealed a general overestimation of body size among anorexia patients. Although a difference in mean effect size was found between studies using Body Part and Whole Body methods, this difference was absent when only the most commonly used Body Part method, the visual size estimation procedure, was considered. Among Whole Body methods, silhouette methods could not be distinguished from the rest in terms of effect size. It was concluded that Whole Body methods and the visual size estimation procedure assess correlated aspects of the body image.

Pgp is expressed on normal haemopoietic progenitor cells. The significance of the efflux pump in protecting normal progenitors for anthracycline toxicity is not defined and is the subject of this study. Pgp was measured in CD34+ progenitors with a rhodamine efflux assay. A high efflux, modulated by verapamil, was only found in a distinct subpopulation (20-30%). Pgp measured by the monoclonal antibody antibody (MoAb) MRK-16 was low in the rhodamine dull, but significantly (P < 0.04) higher than in the rhodamine bright cells. Reverse transcriptase polymerase chain reaction (RT-PCR) of MDR1 mRNA showed a very weak signal in both populations. In a single-cell clonogenic assay, rhodamine dull cells appeared less sensitive to anthracyclines (IC50 daunorubicin 0.005 microg/ml; adriamycin 0.03 microg/ml) compared to rhodamine bright cells (IC50 daunorubicin 0.0025 microg/ml; adriamycin 0.01 microg/ml). Furthermore, verapamil significantly (P < 0.05) potentiated anthracycline toxicity only in the rhodamine dull cells, proving its Pgp-specific modulating effect. Rhodamine dull cells gave larger and more mixed colonies compatible with a more primitive origin. Although detection with MoAbs and RT-PCR revealed a low Pgp level, functionally this Pgp appeared to be very important in protecting primitive progenitors against anthracycline toxicity. This protection can be jeopardized by administration of Pgp modulators.

We studied the appearance of retinoblastoma on unenhanced and gadolinium-enhanced images and the accuracy of tumour staging with MR imaging. The MR images were obtained in 18 children with retinoblastoma and compared with histopathological findings after enucleation. The MR imaging included T1-weighted and dual-echo T2-weighted images before, and T1-weighted images after, gadopentetate dimeglumine injection. The contrast between tumour and ipsilateral vitreous strongly increased (57 %) after gadolinium on T1-weighted images (p = 0.004). Tumour was strongly hypointense as compared with ipsilateral vitreous in all patients using heavily T2-weighted (TE = 120 ms) images (p = 0.001). The estimated T2 of tumour (mean 96 + 14 ms) did not correlate with histological grading or degree of calcification. Unenhanced T1-weighted MR images rightfully excluded extrascleral growth in 16 of 16 cases, but its presence was confirmed after enucleation in only one of 2 abnormal MR scans. Invasion of the optic nerve behind the cribriform plate was confirmed in 2 of 3 abnormal gadolinium-enhanced MR scans, but also in 1 of the 15 cases in which MR images were normal. The T2-weighted images were useful in assessing retinal detachment. We conclude that heavily T2-weighted images, unenhanced T1-weighted images and gadolinium-enhanced T1-weighted MR images are complementary in characterizing and staging retinoblastoma.

We performed an open, nonrandomized, multicenter phase-II trial to evaluate the efficacy and toxicity of 1 year of treatment with the oral iron chelator deferiprone in 38 mainly nonthalassemic patients with transfusional iron overload. Initial serum ferritin varied between 996 and 11.644 micrograms/l. Patients were treated with 3-6 g of deferiprone daily. Mean urinary iron excretion (UIE) in 36 evaluable patients was 21.0 mg/24 h and was significantly higher in the patients with thalassemia than in those with myelodysplasia. Negative iron balance was achieved in 20 patients (56%). The median duration of treatment was 10 months; due to side effects and other causes only 20 patients completed 1 year of treatment. Mean serum ferritin levels decreased from 3563 micrograms/l at the start of the trial to 2767 micrograms/l at 6 months (26 patients, p < 0.004) and to 2186 micrograms/l at 12 months (20 patients, p < 0.005). Serum ferritin levels normalized in two patients who were no longer transfusion dependent. Deferiprone was clearly not effective in three patients (two with myelofibrosis, one with myelodysplasia). One patient with myelodysplasia developed agranulocytosis after 12 months of treatment; this was rapidly reversible after stopping deferiprone. Three patients had a mild and transient decrease in white blood cell count. Other side effects leading to withdrawal from the trial consisted mainly of nausea (3 patients), arthralgia (2), and skin rash (1). No clinical signs of zinc deficiency were seen, although zinc excretion was increased in three patients. No changes were seen in liver enzymes, creatinine, antinuclear factor, T-cell subsets, cardiac function, visual acuity, and audiogram. Although our results confirm deferiprone as an effective iron chelator in patients with thalassemia and in some patients with other forms of iron overload, there is still some concern about the safety of this drug, which therefore, at this time, should be used exclusively in well-controlled clinical trials.

The goal of these experiments was to investigate further the relationship between DNA double-strand breaks and cell killing in human tumor cells, first by comparing different cell lines, and second by radiomodification studies. Field-inversion gel electrophoresis was used to quantify double-strand breaks. Two subclones of the radioresistant human squamous cell carcinoma line SQ20B (SQD9 and SQG6) were compared. These subclones differed in DNA index by a factor of 1.7 but showed the same resistance to radiation as cells of the parental cell line. It was found that, although induction of DSBs was not significantly different in the two cell lines, the t1/2 of the fast component of repair was significantly shorter for SQD9 cells, leading to greater overall repair which was not reflected in increased survival. Caffeine and cysteamine were tested as modifiers of radiosensitivity, using the radioresistant SQ20B line and the radiosensitive SCC61 cell line. No effect of caffeine was seen when the drug was present only during irradiation. Postirradiation incubations with caffeine, however, resulted in a dose reduction factor greater than 2.0 in cell survival for both cell lines. In contrast, induction of DSBs was reduced by caffeine, and no effect on DSB repair was observed. Cysteamine led to a dose protection factor greater than 1.8 in cell survival in both cell lines. A reduction in induced DSBs was found at high doses corresponding approximately with the increase in cell survival. Over the same (low) dose range, however, the correlation between DSB induction and cell killing was poor. These data indicate that DSB induction does not correlate well with cell killing either for different cell lines, for radiochemical modification (cysteamine) or for some other types of modification (caffeine).

Familial hemiplegic migraine (FHM) is an autosomal dominant type of migraine and probably represents the most extreme end of migraine with aura. Reduced magnesium facilitates the development of spreading depression and possibly aura. Cellular magnesium levels are under genetic control. We hypothesized that FHM patients would have significantly reduced intracellular magnesium levels. We determined intracellular and plasma magnesium levels in blood of 38 afflicted and 11 non-afflicted members of three families with FHM and in 32 migraine patients (9 with and 23 without aura) and 32 age and sex matched healthy controls. We found no significant differences between the magnesium levels in the five study groups. We conclude that reduced blood magnesium is unlikely to be related to migraine pathophysiology.

Purpose: Christmas tree cataract consists of highly refractile multicolored "needles" crisscrossing the lens fibers of the deep cortex. The fact that the colors vary according to the angle of the incident light, and that in retroillumination only a dim outline of the cataract is seen, would suggest that Christmas tree cataract is a diffractive phenomenon. This study was performed to unravel the ultrastructure and chemical composition of the Christmas tree needles., Methods: Eight lenses from donor eyes and four extracapsularly extracted lenses with Christmas tree cataract were investigated by scanning and transmission electron microscopy. The chemical composition was studied with energy-dispersive x-ray microanalysis and Raman microspectroscopy., Results: Scanning electron microscope examination showed that the needles are smooth, rectangular, plate-like elements bordered by membranes and amorphous material and running crisscross through the lens. In the specimens for transmission electron microscopic examination, the needles proved to be largely dissolved, but the remains showed regular spacings of approximately 5 nm. Material identical in spacing and electron density was found in neighboring cells bound to a reticular membranous network originating from the fiber-limiting membranes. Energy-dispersive x-ray and Raman microanalysis showed that the needles have a high sulfur content and pronounced S-S, CS-SC, and C-S vibrations. The cytoplasm adjacent to the needles and reticular meshwork had an elevated Ca++ content., Conclusions: It is concluded that cystine is the most likely candidate for the Christmas tree needles and that the needles probably are formed as the result of an age-related aberrant breakdown of crystallins induced by elevated Ca++ levels.

Confocal Raman microspectroscopy was used to monitor local and age-related changes in protein conformation in human eye lenses. In clear human lenses of varying age (range 17-80 years) spectra were recorded along the visual axis, using laser light of 660 nm wavelength. The Raman vibrations in the 650-1750 cm-1 spectral region were analyzed. Difference spectra between central core and different positions along the visual axis were calculated after calibration for protein content using the I(1450) cm-1 CH2/CH3 vibration peak. Tryptophan content was quantified using the peak at 760 cm-1 calibrated for protein. Changes in the 'exposed' vs. 'buried' position of tryptophan were analyzed using the peak heights at I(880) and I(760) cm-1. The difference spectra revealed an excess of tryptophan, tyrosine, phenylalanine, beta-sheet conformation and molecules or molecular groups responsible for a 1425 cm-1 peak in the core region in all lenses investigated. The excess peaks disappeared at about 0.6-0.9 mm below the surface. The tryptophan content increased from superficial to deep layers, levelling off between 0.4-0.8 mm below the surface. Upon aging, the tryptophan content increases in the core not in the cortex. No changes in the 'exposed' vs. 'buried' position of tryptophan were observed. Changes in tryptophan and tyrosine probably reflect the maturational shift from cortex to core in the relative content of alpha, beta and gamma crystallines. The age-related increase in tryptophan in the core may reflect the preferential breakdown by endo- and exopeptidases of alpha-crystallins damaged upon aging. The increase in beta-sheet conformation may indicate a post-translational shift in secondary conformation upon aging. These changes in protein conformation are largely completed in a small superficial zone, i.e., in the early life span of the crystallins.

Based on the mode of action of iron chelators, one might expect a decrease in bone marrow iron availability, resulting in worsening of the anaemia in certain types of iron-loading anaemia. However, improvement of anaemia or reduction in transfusion requirements during chelation treatment has been reported in various types of iron-loading anaemia. It is suggested that iron chelators act as mediators facilitating iron release from storage sites and its delivery to haematopoietic tissues. In addition, a reduction of iron stores may upregulate erythropoietin response and bring about a decrease of disease activity in inflammatory disorders, resulting in a haemoglobin rise. Large trials with (oral) iron chelators are required to verify these possible effects.

The taxonomic relations of a number of tospovirus isolates, collected in different geographical areas and from different host plants, were studied. To delineate these isolates, properties such as susceptibility of a limited range of host plants, symptomatology, cytopathology, nucleocapsid composition, serology of their nucleocapsid proteins, and nucleotide sequence homology were compared. The results show that isolates which have previously been discriminated as members of three different serogroups, should in fact be regarded as representatives of at least three distinct virus species in the tospovirus genus.

In an 86-year-old woman a neglected pessary and extreme coprostasis gave rise to anuria, urosepsis and bilateral ureteropelvic dilatation with an empty bladder. This seems to be the first reported case of its kind.

Effects of various doses (0-250 micrograms/kg, SC) of estradiol-17 beta (E2) in a two-bottle choice conditioned taste aversion and a two-compartment conditioned place preference procedure were studied in male and female rats. Dose-dependent taste aversion and place aversion effects of E2 were established, and the conditioned taste aversion procedure was found to be more sensitive in detecting aversive properties of E2 than the conditioned place preference procedure. Although aversive properties of E2 were found in both sexes, the effects were clearly more prominent in males as compared to females. From this study, it was concluded that E2 acts as an unconditioned aversive stimulus in both male and female rats capable of gaining control over different types of behavior by associative learning.

An influenza-like illness appeared recently among workers in a plant processing synthetic yarn. A humidifier, a cold-water spraying system, was the suspected cause. Lung function changes over the day and week and changes in blood leucocytes were studied among the workers from the suspected department and two reference populations. Exposure to colony-forming units of bacteria and fungi and to endotoxins was also monitored. The workers from the suspected department had statistically significantly lower lung functions on the first workday of the week than the referents. Their blood leucocytes were also raised statistically significantly. The exposure to fungi, bacteria, and endotoxins differed significantly between the various departments, but the measured levels were low. It was concluded that the observed effects were suggestive of a "Monday morning fever" type of reaction and that adverse effects occurred at exposure levels lower than those found to date in the literature.