Development in Danish medical students' empathy: study protocol of a cross-sectional and longitudinal mixed-methods study.

BACKGROUND: Clinical empathy has been associated with positive outcomes for both physicians and patients such as: more accurate diagnosis and treatment, increased patient satisfaction and compliance, and lower levels of burnout and stress among physicians. International studies show mixed results regarding the development of empathy among future physicians associating medical education with decline, stability or increase in empathy levels. These mixed results are due to several study limitations. In Denmark, no investigation of Danish medical students' empathy trajectory has yet been conducted wherefore such a study is needed that optimizes the study design of earlier studies. METHODS: The aim of the study is to examine and analyze empathy levels and empathy changes among Danish medical students from the four medical faculties in Denmark, employing a cross-sectional and longitudinal mixed-methods design including a control group of non-medical students. By supplementing cross-sectional and longitudinal questionnaire studies with a focus group interview study it is the aim to identify and analyze factors (including educational) that are perceived by medical students to influence the development of empathy and its expression in clinical care. DISCUSSION: The results of the study will provide insight into the trajectory of medical students' empathy and in undergraduate and graduate students' experiences with and perceptions of empathy development. In addition, the study will provide evidence to support further research on how targeted educational programmes can best be designed to educate empathic and patient-centered physicians.

[Systematic Review of the Application of Complementary and Alternative Medicine and their Potential Therapeutic Benefits in the Treatment of Ophthalmology Patients]. / Ergebnisse eines systematischen Reviews zu Einsatz und berichtetem therapeutischem Nutzen komplementärmedizinischer Methoden in der Augenheilkunde.

Purpose A systematic review was carried out of the reported therapeutic effects of complementary and alternative medicine methods as supplementary or primary treatments for patients suffering from glaucoma, cataract or age-related macular degeneration (AMD). Material and Methods For the years 1990 to 2013, the following databases were screened for reports of the application of complementary and alternative treatments: PubMed, Cochrane Library, EMBASE, CAMbase and AMED. Both randomised and prospective non-randomised patient trials were included in the review; results were evaluated in the following classes: "phytotherapy", "acupuncture/acupressure", "biofeedback" and "other alternative treatments". The studies were evaluated by measures of clinical effect, statistical significance (p value and/or confidence interval) and the underlying trial design. Results 30 clinical trials were included, including 13 on glaucoma, 5 on cataract and 12 on AMD patients. These trials were based on patient numbers of 6 - 332, 27 - 157 and 6 - 328 patients, respectively. Phytotherapy was applied in 14 trials, including 6 on glaucoma patients (all 6 with a controlled design, and 3 of which reporting statistically significant results); 5 trials were on cataract patients (3 with a controlled design and 2 with a significant result) and 3 on AMD patients (only 1 with a controlled design, with a significant result). Acupuncture/acupressure was investigated in 9 trials, 5 on glaucoma patients (3 with a controlled design, 1 with a significant result); no acupuncture/acupressure trial was found in cataract patients, but 4 trials in AMD patients (none with a controlled design). Biofeedback was studied in 4 trials, all on AMD patients (only one with a controlled design, without statistically significant findings). Conclusion Despite its rigorous inclusion criteria, this review identified several clinical trials on complementary and alternative medicine in ophthalmological patients. Phytotherapeutic methods gave significant results in half of the reported controlled trials, whereas there were few significant benefits with acupuncture or acupressure.

Four-year follow-up on fatigue and sleep quality of a three-armed partly randomized controlled study in breast cancer survivors with cancer-related fatigue.

Cancer-related fatigue (CRF) is a frequent long-term symptom in non-metastasized breast cancer patients (BC). This 4-year follow-up intended to compare the long-term effects of a 10-week multimodal therapy (MT: sleep education, psychoeducation, eurythmy- and painting therapy) and combination therapy [CT: MT plus aerobic training (AT)] to AT-control. BC-patients were randomized or allocated by preference to three arms in a comprehensive cohort study. Primary outcome was a composite score including Pittsburgh Sleep Quality Index (PSQI) and Cancer Fatigue Scale (CFS-D), captured at baseline, after 10 weeks of intervention (T1), 6 months later (T2), and after 4 years (T3). We exploratively tested for superiority of MT and CT versus AT after 4 years (T3) based on the statistical model of the main analysis. Of 126 (65 randomized) BC-patients included, 105 started treatments and 79 were re-assessed for long-term effects (T3). MT and CT were superior over AT after 4 years regarding PSQI/CFS-D and PSQI sum-score, respectively (all p < 0.05), but not for CFS-D. The multimodal and combination treatment for breast cancer patients with CRF indicates sustainable long-term superiority over aerobic training for the outcomes sleep quality and combined sleep quality/fatigue. A confirmative randomized controlled trial is warranted.

[Spiritual needs of patients in an emergency room]. / Spirituelle Bedürfnisse von Patienten eines Notfallzentrums.

BACKGROUND: Spiritual needs (spN) are important for human beings-independently of religious affiliation. They can be a resource for coping with stressful situations, e.g., those triggered by the acute onset of a disease. Emergency rooms are hospital departments with high medical performance which may cause a particular insecurity among emergency patients. The present study is the first to examine spiritual needs in a sample of patients in the emergency room. METHODS: A total of 383 out of 479 patients were approached and asked to complete the German version of the Spiritual Needs Questionnaire (SpNQ-20). All consented to the collection of demographics and clinical data. The analysis encompassed descriptive statistics, correlations analysis, univariate and multiple variance analysis. RESULTS: The needs for inner peace and generative needs (to pass something on to others, to do something for others) were more important than religious (rN) and existential (eN) needs. We did not find a correlation between spN on the one hand and the reason for consultation, the severity, and the number of comorbidities on the other hand. Age did not play a decisive role, rather, patients' needs, especially rN, were significantly more important among women than among men. CONCLUSION: Even in an emergency situation, people are ready to express their spN. Early assessment of these needs exposes important nonmedical aspects of the sick person and helps to consider the assessed needs. Further studies will show whether this has an impact on the further course of treatment and the well-being of the patients.

Spiritual needs of patients with chronic pain diseases and cancer - validation of the spiritual needs questionnaire.

PURPOSE: For many patients confronted with chronic diseases, spirituality/religiosity is a relevant resource to cope. While most studies on patients' spiritual needs refer to the care of patients at the end of life, our intention was to develop an instrument to measure spiritual, existential and psychosocial need of patients with chronic diseases. METHODS: In an anonymous cross-sectional survey, we applied the Spiritual Needs Questionnaire (SpNQ version 1.2.) to 210 patients (75% women, mean age 54 +/- 12 years) with chronic pain conditions (67%), cancer (28%), other chronic conditions (5%). Patients were recruited at the Community Hospital Herdecke, the Institute for Complementary Medicine (University of Bern), and at a conference of a cancer support group in Herten. RESULTS: Factor analysis of the 19-item instrument (Cronbach's alpha +/- .93) pointed to 4 factors which explain 67% of variance: Religious Needs, Need for Inner Peace, Existentialistic Needs (Reflection / Meaning), and Actively Giving. Within the main sample of patients with chronic pain and cancer, Needs for Inner Peace had the highest scores, followed by Self competent Attention; Existentialistic Needs had low scores, while the Religious Needs scores indicate no interest. Patients with cancer had significantly higher SpNQ scores than patients with chronic pain conditions. There were just some weak associations between Actively Giving and life satisfaction (r +/- .17; p +/- .012), and negatively with the symptom score (r +/- -.29; p < .0001); Need for Inner Peace was weakly associated with satisfaction with treatment efficacy (r +/- .24; p < .0001). Regression analyses reveal that the underlying disease (i.e., cancer) was of outstanding relevance for the patients' spiritual needs. CONCLUSION: The preliminary results indicate that spiritual needs are conceptually different from life satisfaction, and can be interpreted as the patients' longing for spiritual well-being. Methods how health care professionals may meet their patients' spiritual needs remain to be explored.

Validation of the brief multidimensional life satisfaction scale in patients with chronic diseases.

OBJECTIVE: We intended to determine the reliability of a brief life satisfaction scale in a sample of patients with chronic diseases, and to analyze its external validity. METHODS: Reliability and factor analysis of the 8-item "Brief Multidimensional Life Satisfaction Scale" (BMLSS) were performed according to standard procedures. The test sample contained 979 individuals (mean age 54+/-11 years). Forty-two percent had cancer, 22% chronic pain conditions, 10% depressive disorders, 6% other chronic diseases, and 20% were healthy. RESULTS: Reliability analysis of the 8-item pool revealed a good internal consistency coefficient (Cronbach's alpha=.869), and a single-factor structure which explains 53% of variance. The BMLSS sum scores significantly differed with respect to the underlying disease, family status, duration of disease, and age. The highest scores were found in healthy individuals, and the lowest in patients with chronic pain conditions and depressive disorders. In cancer patients, the BMLSS correlated negatively with Depression/Anxiety (HADS), Fatigue (CFS-D), and positively with SF-12's mental health and to a weaker content also with physical health. Stepwise regression analyses revealed that life satisfaction can be predicted best by (the absence of) depression, but also by Conscious Living (AKU), which is an active cognitive-behavioral style in terms of adaptive coping. CONCLUSIONS: The evaluation of the BMLSS revealed that the instrument has good psychometric properties and can be regarded as a brief, reliable and valid measure of LS in patients with chronic diseases. The instrument can be an important additive to existing health-related quality of life questionnaires, since it captures dimensions that contribute to quality of life but are not health related.

[Assessment of quality of life in health services research - conceptual, methodological and structural prerequisites]. / Die Erfassung von Lebensqualität in der Versorgungsforschung - konzeptuelle, methodische und strukturelle Voraussetzungen.

On July 1, 2009, the German Network for Health Services Research [Deutsches Netzwerk Versorgungsforschung e. V. (DNVF e. V.)] approved the Memorandum III "Methods for Health Services Research", supported by the member societies mentioned as authors and published in this Journal [Gesundheitswesen 2009; 71: 505-510]. This is an in-depth publication on "quality-of-life assessment in health services research". Within the context of the health sciences, quality of life (QL) encompasses the subjective well-being and functioning in the physical, psychological and social domains. QL informs about the aspects of health care that "actually get to the patient". QL is what patients primarily experience, what they talk about and what to a large degree affects the acceptance of health-care services and processes in the society. Therefore, QL can be considered as a highly important endpoint within health services research. The importance of the construct quality of life is also emphasised in German treaties on social law and utility analyses. This paper is the first account on the relations between health services research and the concept and assessment of QL. Our working group has specified key criteria for QL assessment within studies on health services research. (1) Assessment instruments need to comply with standard quality criteria (reliability, validity, sensitivity, interpretability) and the decision for a particular instrument has to be reasonably justified. (2) Study design and study population have to match with the scientific research question and the sample size has to be biometrically sound. (3) QL assessment including time points over the course of the study has to follow a standardized protocol. (4) Criteria for analysis and interpretation have to be prospectively specified. (5) Studies focusing on diagnostic/therapeutic issues need to specify standards for diagnostic criteria and related therapeutic interventions.

Reliability and first validity of the inner correspondence questionnaire for painting therapy (ICPTh) in a sample of breast cancer patients.

Objectives Art therapy (ArT) such as mindfulness-oriented painting therapy is increasingly used in psychosomatic, oncological integrative and rehabilitative medicine. Though it remains unknown how ArT works, we hypothesize that an engaged participation with painting ('Inner-Correspondence') contributes to improved symptom scores. In the context of a comprehensive cohort study for breast cancer survivors with cancer-related fatigue, we developed a patient-reported outcome measure to assess 'Inner Correspondence' with painting therapy and conducted a first validation study. Design A 24-item questionnaire on 'Inner Correspondence' (ICPTh) was administered after ten weeks of intervention and at six month followup together with concurrent scales (Inner Correspondence and Peaceful Harmony, Cancer Fatigue Scale, Hospital Anxiety and Depression Scale, Internal Coherence Scale). Statistical assessment included reliability- and factor analyses. Results A total of n = 68 BC (mean age, 58.2 years, SD = 8.7) participated in the preliminary validation study. Exploratory factor analysis revealed a robust 22-item scale with an unambiguous four-factor solution explaining 78% of total variance and the following subsales: 1) therapy congruence and relaxation (11 items), 2) inner development and mood (6 items), 3) artistic skill (3 items) and 4) task congruence (2 items). The 22-item ICPTh yielded high reliability (Cronbach's alpha = .966, item-total correlation = .497 - .883, test-retest reliability = .888). Conclusions We present a reliable instrument to measure 'Inner Correspondence' with painting therapy. Due to the small sample size and sample selection further validation studies are indicated.

Adaptation of the Herdecke Quality of Life questionnaire towards quality of life of cancer patients.

Although instruments for the measurement of quality of life (QoL) do exist for cancer patients, factors like sleepiness or digestion are only marginally addressed. We intended to adapt the Herdecke Quality of Life (HLQ) towards these aspects in a multi-centre cross-sectional validation study. A group of 293 subjects [79% female, age: 55.9 +/- 13.4 years; 146 cancer patients, 28 patients with rheumatic diseases and a healthy control group (n = 119)]. Structural relations between the items were detected by factor and reliability analyses. For external validation, correlations with the hospital anxiety and depression scale (HADS), self-regulation score (SRS) and the Marburger short questionnaire on chronotypology (MQC) were performed, and test-retest reliability was calculated. Factor analysis found three sub-scales: physical abilities (PA) (Cronbach's alpha = 0.90), sleep quality (SQ) (Cronbach's alpha = 0.89) and digestive well-being (DWB) (Cronbach's alpha = 0.80). Sleep quality correlated well with HADS-anxiety (r =-0.52), PA with HADS-depression (r =-0.49). We found moderate correlations of PA and SQ with SRS, while the HLQ scales did not correlate with the MQC. Analysis of test-retest reliability resulted in values of r = 0.757 for PA, r = 0.715 for SQ and r = 0.603 for DWB. The HLQ-cancer suits to measure unique features of cancer-related QoL aspects. In future studies it has to be tested in larger samples of cancer patients.

Course of mitogen-stimulated T lymphocytes in cancer patients treated with Viscum album extracts.

BACKGROUND: In a prospective observational study, the impact of two different dose regimes of a commercially available fermented Viscum album L. extract (VA-E, Iscador) on the function of T lymphocytes from cancer patients was investigated. PATIENTS AND METHODS: A total of 71 cancer patients were enrolled. These patients attended two different sections of a tumor outpatient clinic which are used to apply different VA-E escalation schemes. Our hypothesis was that a rapid dose escalation of subcutaneously applied VA-E may induce strong local reactions at the injection side (>3 cm diameter) and may have an effect on the functional competence of T lymphocytes (mitogen-activated interleukin-2 receptor alpha chain), which was recorded over an observation period of six month. RESULTS: Within this observation period, a decline of stimulated T cell function was observed, particularly in patients with colorectal or prostate cancer; this decline was not seen in patients with breast cancer (who received lower mean concentrations per month) nor in patients with dose adaptation in response to too strong local reactions. CONCLUSION: With respect to T-cell function, our results indicate that in patients without local reactions, a long lasting mistletoe extract application should be withheld periodically to allow T-cell reactivity to recover.

Characterisation of granulocyte stimulation by thionins from European mistletoe and from wheat.

Thionins are small basic peptides found in different plant species, which are known to exert cytotoxic properties. In addition, previous data indicated an activation of human granulocytes by thionins from European mistletoe (viscotoxins, VT). To extend these latter findings, we investigated the influence of VT and from thionins from wheat flour (purothionin) on human granulocytes by flow cytometry and tried to characterise the involved molecular structures and mechanisms. Phagocytosis was determined by incorporation of FITC-labelled Escherichia coli and respiratory burst by oxidation of dihydrorhodamine 123 to rhodamine 123. VT and purothionin significantly enhanced E. coli-stimulated phagocytosis and respiratory burst at 25 and 250 microgram/ml. Phagocytosis of damaged lymphocytes by granulocytes was detected by electron microscopy in the VT-stimulated (100 microgram/ml) but not in the control cultures. The poly-cationic structure of the intact molecule seems to be crucial, as evidenced by comparison of the burst and phagocytosis-enhancing effects induced by other poly-cationic (protamine sulphate, histone, poly-l-arginine, poly-l-lysine) and poly-anionic (poly-l-glutamic acid) peptides, while pore forming due to amphipathic properties seems to be less important. Ca2+ and Mg2+ could not inhibit VT-enhanced phagocytosis and, thus, could not inhibit binding of VT to granulocytes. In addition, verapamil at low concentrations inhibited VT activity, suggesting the involvement of Ca2+ channels for granulocyte activation by the VT. Similarly, thionins and histones in contrast to protamine sulphate induced cell death of granulocytes at 250 microgram/ml as demonstrated by an enhanced release of reactive oxygen intermediates in unstimulated granulocytes. From these data one may suggest that activity of VT is induced by strong unspecific ionic binding, probably followed by specific receptor binding, and thionins exhibit stimulatory and cytotoxic effects on immune cells, which have to be further characterised.

Selective killing of CD8+ cells with a 'memory' phenotype (CD62Llo) by the N-acetyl-D-galactosamine-specific lectin from Viscum album L.

As reported previously by our group, among the toxic proteins from Viscum album L. only the mistletoe lectins (MLs) induce the apoptotic killing pathway in human lymphocytes. Although one may expect a homogenous distribution of carbohydrate domains on cell surface receptors for the carbohydrate binding B chains of the toxic protein, the sensitivity of cells to these B chains obviously differ. Here we report a selective killing of CD8+ CD62Llo cells from healthy individuals by the galNAc-specific ML III (and RCA60, which binds to gal and galNAc), while the gal-specific ML I was less effective. This selective killing is not sufficiently explained by protein synthesis inhibition alone, since this subset was not affected by other ribosome inhibiting proteins such as the lectin from Ricinus communis (RCA120), lectin from Abrus precatorus (APA), abrin A, and inhibitors of RNA, DNA and/or protein synthesis such as actinomycin D, mitomycin C, and cycloheximide. We conclude that CD8+ cells with 'memory' phenotype (CD62Llo) are more sensitive to the ML III-mediated killing than their CD8+ CD62Lhi counterparts, CD4+ T cells, and CD19+ B cells. These cells probably express a distinct receptor with galNAc domains that is missing or not active on CD8+ cells with a 'naive' phenotype.

Childhood acute lymphoblastic leukemia (ALL): sister chromatid exchange (SCE) frequency and lymphocyte subpopulations during therapy.

Sister chromatid exchanges (SCE) and lymphocyte subsets of children with acute lymphoblastic leukemia (ALL) were investigated during chemotherapy. The treatment followed protocol ALL-BFM-90. Children with ALL at the time of diagnosis showed statistically significant higher SCE frequencies (4.9 +/- 0.77) than healthy controls (3.6 +/- 0.93; P = 0.002). The in vivo effects of cyclophosphamide (CP) resulted in a dramatic increase of the SCE frequency (20.5 +/- 3.76). This increased SCE level of lymphocytes might reflect an instability of DNA or a deficiency of DNA repair. One could suggest that lymphocytes of children with ALL might have a higher susceptibility to harmful influences; and this could be a co-factor towards the development of the malignant disease. However, immediately 1 week after the administration of CP, the SCE rate decreased. This decline of SCE frequency correlated with a severe reduction of the absolute number of T lymphocytes. The observed reduction of SCE frequency may be due to a loss of T lymphocytes, or SCE became repaired during 1 week.

Induction of apoptosis by mistletoe lectin I and its subunits. No evidence for cytotoxic effects caused by isolated A- and B-chains.

Type II ribosome inactivating proteins (RIP II) are generally known to induce apoptosis in human cells by the inhibition of protein biosynthesis. Recent data from mistletoe RIP II proteins (eg. mistletoe lectin I; ML1) suggest an additional mode of apoptosis induction through the binding of their lectin part to certain cell surface receptors as is known for some human galectins. In order to clarify this possibility, we used highly sensitive flow cytometric apoptosis assays and mistletoe hololectin subunits of proven purity to show that neither human lymphocytes nor Molt-4 cells undergo apoptosis after treatment with isolated lectin-type B-chains. In contrast to earlier investigations, only the hololectin was able to induce apoptosis in these assays. We conclude that direct apoptosis induction by mistletoe lectins occurs only after uptake of the molecules into the cell due to the action of the ribosome inactivating A-chain.

Viscum album L. extracts reduce sister chromatid exchanges in cultured peripheral blood mononuclear cells.

Increasing concentrations of Viscum album L. extracts were shown to significantly reduce sister chromatid exchange (SCE) frequency of phytohaemagglutinin (PHA)-stimulated peripheral blood mononuclear cells (PBMC) of healthy individuals. This decrease of SCE could not be explained either by changes in lymphocyte subpopulations, by cytostatic effects of the drug or by accelerated proliferation of PHA-stimulated PBMC. Currently, no other cells tested have shown this effect. One therapeutic effect of these anti-mutagenic drugs could be a stabilisation of mononuclear blood cell DNA.

Disorders of calculation in aphasic patients--spatial and verbal components.

The aim of the study is to investigate whether the disorders of calculation observed in patients with Broca's aphasia are due to linguistic factors whereas those observed in patients with Wernicke's aphasia may be reduced in part to a disorder in the ability of spatial visualization. Two sets of arithmetic tasks were given to 20 Broca's aphasics, 20 Wernicke's aphasics, 20 patients with right hemispheric retrorolandic cerebral lesions and 40 patients with no brain damage. The solution of the tasks required different degrees of spatial visualization. The statistical evaluation of the data showed, as hypothesized, that Wernicke's aphasics perform significantly poorer than Broca's aphasics in tasks with a more pronounced spatial component than in the tasks having a predominantly verbal component.

Effects of Viscum album L. on cyclophosphamide-treated peripheral blood mononuclear cells in vitro: sister chromatid exchanges and activation/proliferation marker expression.

Based on recently published data, Viscum album L. (VAL) extracts have been shown to provide a DNA stabilizing effect which seems to be restricted to the peripheral blood mononuclear cells (PBMC). We have now investigated whether VAL exerts effects of cellular protection for phytohemagglutinin-activated PBMC treated with cyclophosphamdie (CP) in vitro. The addition of VAL resulted in a slight reduction of CP-induced sister chromatid exchanges of cultured PBMC from healthy individuals. The incubation with CP significantly reduced the expression of the low affinity interleukin-2 receptor (IL-2R alpha chain) and of transferrin receptor (TfR) on PHA-stimulated T lymphocytes. The addition of 10 micrograms/ml VAL was protective against the CP-induced depression of IL-2R alpha chain and TfR expression on these cells. The simultaneous addition of CP and purified VAL components, such as ML I, ML II/III, and viscotoxins did not significantly change expression of IL-2R alpha chain and TfR on T cells. Thus, so far undefined VAL components might be responsible for the observed protection effects of the whole plant extract. The results presented here should encourage investigation of this drug, which might become an interesting adjuvant in cancer therapy.

Induction of apoptosis by the N-acetyl-galactosamine-specific toxic lectin from Viscum album L. is associated with a decrease of nuclear p53 and Bcl-2 proteins and induction of telomeric associations.

The ribosome-inhibiting proteins from Viscum album L., i.e. the mistletoe lectins (ML), were recognized to induce apoptosis in various tumour cell lines and human lymphocytes. However, several aspects of ML-induced cell death are unclear. We report that the galNAc-binding ML III incubated with human lymphocytes mediates a very effective death signal resulting in the binding of Annexin-V and expression of mitochondrial membrane proteins Apo2.7, but also in an influx of the DNA intercalating dye propidium iodide. The addition of the ribosome-inhibiting protein Volkensin also induced Apo2.7 molecules, while Momordin, lacking a carbohydrate-binding chain, did not enter the cell membrane and thus did not affect the cells. However, we observed ML III to preferentially affect CD8+ cells with a memory phenotype (CD62L(lo)) as compared to their CD8+ CD62L(hi) counterparts, CD4+ T cells and CD19+ B cells. Furthermore, ML III did not induce sister chromatid exchange-inducing DNA lesions but reduced the intensity of telomeric signals, increased the frequencies of telomeric associations and C-anaphases and reduced nuclear Bcl-2 and p53 proteins. Whatever the exact mechanisms are, our results provide strong evidence that the ML III-mediated cytotoxicity involves distinct killing pathways, i.e. (1) primary cell death via an induction of apoptosis which may not be dependent on protein and/or RNA synthesis and may not involve p53 and Bcl-2 proteins and (2) a loss of telomeres resulting in chromosomal instability in the surviving cells which is incompatible with life. However, we cannot exclude the possibility that this effect is due to a decrease in nuclear p53 proteins.

Induction of mitochondrial Apo2.7 molecules and generation of reactive oxygen-intermediates in cultured lymphocytes by the toxic proteins from Viscum album L.

We analysed mitochondrial alterations in human lymphocytes incubated with toxins exerting RNA and/or protein synthesis/transport inhibitory activity. We found that all toxins known to affect macromolecule synthesis, such as ricin from Ricinus communis, mistletoe lectin I (ML I) from Viscum album, cycloheximide, actinomycin D, and brefeldin A but also the thionins from Viscum album (viscotoxins; VT) generated reactive oxygen intermediates (ROI) and induced expression of newly described mitochondrial membrane proteins Apo2.7, however, with different kinetics. Apart from a rapid permeabilisation of cell membranes by the VT with swelling of mitochondria, loss of their cristae and ROI generation within 2-4 h, the majority of the cells may have received a distinct 'death signal' resulting in an induction of Apo2.7 molecules within 24 h. In contrast, protein synthesis/transport inhibition may signal for apoptosis within 24 h by decreasing distinct 'survival promotors' which remain to be characterised.

Induction of apoptosis in human lymphocytes treated with Viscum album L. is mediated by the mistletoe lectins.

Viscum album L. (VAL) is a phytopreparation used in adjuvant cancer therapy with both immunostimulatory and DNA stabilizing properties at low drug concentrations and cytostatic/cytotoxic properties at higher concentrations. The present work examines the cytotoxic effects of VAL extracts produced from mistletoes grown on different host trees and of purified toxic proteins from VAL, such as the D-galactose-specific lectin I (ML I), the N-acetyl-D-galactosamine-specific ML II and ML III, and crude viscotoxins towards cultured human lymphocytes. The decrease in the number of cultured lymphocytes and blast cells treated with whole plant extracts from VAL was host tree-specific. Nevertheless, there was no close correlation to the content of MLs or viscotoxins. Using the purified proteins, it became obvious that the cell killing was mediated by the induction of apoptosis, as measured by the appearance of a hypodiploid DNA peak using flow cytometry. ML III was the most effective to induce apoptosis, followed by ML II and ML I, while the viscotoxins and oligosaccharides from VAL did not. By measuring the surface expression of IL-2R alpha chains, transferrin receptors and APO-1/Fas molecules on non-apoptotic T cells, no significant changes were observed at low ML concentrations (1 ng/ml), but their decrease at higher ones. Our findings suggest that there might be at least two different ways of cell killing operative in VAL-mediated cytotoxicity: (a) the typical apoptotic cell death with the appearance of hypo-diploid nuclei, and (b) a direct or indirect killing by damaging the cell membrane with subsequent influx of Ca2+ and of the DNA intercalating dye propidium iodide and cell shrinkage. These effects might not be exclusive, as they probably occur simultaneously.