Antenatal depressed mood and child cognitive and physical growth at 18-months in South Africa: a cluster randomised controlled trial of home visiting by community health workers.

AIM: To examine the child outcomes at 18-months post-birth of a population cohort of women with antenatal depressed mood, half of whom were randomly chosen to receive perinatal home visits from community health workers during pregnancy. METHOD: Pregnant women in 24 neighbourhoods (98% participation) were randomised by neighbourhood to: (1) standard clinic care (SC; 12 neighbourhoods; n = 594) or (2) the Philani Intervention Program, a home visiting intervention plus standard care (12 neighbourhoods; n = 644). The physical and cognitive outcomes of children of mothers with antenatally depressed mood (Edinburg Perinatal Depression Scale >13) in the intervention condition were compared at 18-months post-birth to children of mothers without depressed mood in pregnancy in both conditions. RESULTS: More than a third of mothers had heightened levels of antenatal depressed mood (35%), similar across conditions. Antenatal depressed mood was significantly associated with being a mother living with HIV, using alcohol and food insecurity. At 18-months, the overall cognitive and motor scale scores on the Bayley Scales of Development were similar. However, 10.3% fewer children of mothers with antenatal depressed mood in the intervention condition had cognitive scores on the Bayley Scales that were less than 85 (i.e., s.d. = 2 lower than normal) compared with children of mothers with antenatal depressed mood in the SC condition. Intervention children of mothers with antenatal depressed mood were also significantly less likely to be undernourished (Weight-for-Age Z-scores < -2). CONCLUSION: Cognitive development and child growth among children born to mothers with antenatal depressed mood can be improved by mentor mother home visitors, probably resulting from better parenting and care received early in life.

A novel triterpene extract from mistletoe induces rapid apoptosis in murine B16.F10 melanoma cells.

The European mistletoe Viscum album L. is a plant used for remedies in cancer treatment. The benefit of commonly used aqueous extracts is controversial but the plant contains water insoluble triterpene acids providing interesting anticancer properties. Triterpene extracts (TE) from plants and single triterpenoids such as oleanolic acid (OA) or betulinic acid (BA) are known for their cytotoxic effects on cancer cell lines in vitro. We report here cytotoxic effects of a novel OA-rich triterpene extract from mistletoe (V. album L., Santalaceae) solubilized by 2-hydroxypropyl-ß-cyclodextrin (2-HP-ß-CD) on B16.F10 mouse melanoma cells. The 2-HP-ß-CD solubilized triterpene extract (STE) was highly cytotoxic by causing DNA fragmentation, followed by loss of membrane integrity and intracellular adenosine-5'-triphosphate (ATP). Blocking the caspase machinery by inhibitors aborted DNA fragmentation and delayed the cytotoxic effects but did not prevent cell death. The solubilization by 2-HP-ß-CD allows a solvent-free application of triterpene extracts in the in vitro setting. These findings suggest the use of STE from mistletoe as a solvent-free anticancer drug for preclinical animal experiments and clinical trials.

Olea europaea leaf extract exerts L-type Ca(2+) channel antagonistic effects.

ETHNOPHARMACOLOGICAL RELEVANCE: In Southern Europe Olea europaea leafs are known as a folk remedy for hypertension. Cardiovascular diseases are still the leading causes of morbidity and mortality in industrialized countries with hypertension being one of the main risk factors. AIM OF THE STUDY: We investigated effects of a commercial Olea europaea leaf extract (OLE) on isolated hearts and cultured cardiomyocytes. MATERIALS AND METHODS: Isolated rabbit hearts were perfused according to the Langendorff technique and connected to a 256-channel epicardial mapping system. Voltage clamp experiments were performed in cultured neonatal rat cardiomyocytes using a perforated-patch technique. RESULTS: OLE caused a concentration-depended decrease in systolic left ventricular pressure and heart rate as well as an increase in relative coronary flow and a slight, but not significant prolongation of PQ-time. There were no significant changes between the groups in the activation-recovery interval and its dispersion, total activation time, peak-to-peak amplitude, percentage of identical breakthrough-points and similar vectors of local activation. Voltage clamp experiments in cultured neonatal rat cardiomyocytes showed a significant decrease in maximum I(Ca,L) by OLE which was reversible upon wash-out. CONCLUSIONS: OLE suppresses the L-type calcium channel directly and reversibly. Our findings might help to understand the traditional use of OLE in the treatment of cardiovascular disease.

A new role for extracellular Ca2+ in gap-junction remodeling: studies in humans and rats.

We wanted to elucidate whether extracellular calcium may regulate the expression of the cardiac gap-junction proteins connexin 40 and connexin43. In the free wall of the left atria of 126 cardiac surgery patients with either sinus rhythm (SR) or chronic atrial fibrillation (AF), we determined the expression of the cardiac gap-junction proteins Cx43 and Cx40 by Western blot and immunohistology. For deeper investigation, we incubated cultured neonatal rat cardiomyocytes at 2 or 4 mM Ca(++) for 24 h and determined intercellular coupling, Cx40, Cx43 protein and mRNA expression, protein trafficking and sensitivity to verapamil (10-100 nM), cyclosporin A (1 microM),and BMS605401 (100 nM), a specific inhibitor of Ca(2+)-sensing receptor (CaSR). We found in patients that both Cx are up-regulated in AF in the left atrium (by 100-200%). Interestingly, Cx40 was mainly up-regulated, if total serum calcium was >or=2.2 mM, while Cx43 was independent from extracellular [Ca(++)]. In cultured cells, 4 mM Ca(++)-exposure lead to up-regulation of Cx40, but not Cx43. We found enhanced Cx40 in the plasma membrane and reduced Cx40 in the Golgi apparatus. The membrane Cx40 up-regulation resulted in enhanced gap-junction intercellular coupling with a shift in the Boltzmann fit of voltage-dependent inactivation indicating a higher contribution of Cx40 as revealed by dual whole cell voltage clamp experiments. BMS605401 could prevent all Ca(2+)-induced changes. Moreover, cyclosporin A completely abolished the Ca(2+)-induced changes, while verapamil was ineffective. We conclude that extracellular calcium (24 h exposure) seems to up-regulate Cx40 but not Cx43.

Induction of antibodies to viscotoxins A1, A2, A3, and B in tumour patients during therapy with an aqueous mistletoe extract.

Mistletoe extracts exert immunomodulatory properties on immunocompetent cells of the innate as well as the specific immune system. These effects have been mainly ascribed to mistletoe lectin 1 (ML-1) present in most of the extracts. However, it became evident that also other components of these extracts may induce immunological reactions, and especially viscotoxins (VT) may be of relevance. Aim of the study was, therefore, to evaluate whether VT like ML-1 could activate B-cells and lead to the production of VT-specific antibodies. Sera from 26 patients with different tumours who were treated with the mistletoe extract ABNOBAviscum Mali (AM) 4 for at least 18 weeks were analysed before therapy and after 3, 6, 9, 12, and 18 weeks. Sera were tested by ELISA against the four viscotoxins A1, A2, A3, B, as well as against ML-1. Within the observation period twenty-four (92%) of the 26 patients developed antibodies to at least one of the four VT and 25 (96%) to ML-1. In most instances, anti-VT antibodies appeared after 6-9 weeks of treatment. The antibodies were predominantly of the IgG type belonging preferentially to the IgG1 and IgG3 subclass. IgE antibodies were found only to VT-B and to ML-1. There was no relation between the development of antibodies to VT and ML-1, and also cross-reactivity could be excluded with high probability. These data indicate that not only ML-1 but also VT induce immunological responses in patients treated with mistletoe extracts. Whether there is any relationship to the postulated anti-tumour effect of mistletoe extracts has, however, still to be evaluated.

Polysaccharides from fresh Viscum album L. berry extract and their interaction with Viscum album agglutinin I.

A special microfiltrated colloidal preparation from fresh Viscum album L. berries was investigated concerning the occurrence and structural features of polymeric carbohydrates. A crude polysaccharide fraction was isolated from lectin-, tannin- and protein-depleted microfiltrates. Further fractionation by exchange chromatography revealed a neutral fraction (average molecular weight 30 kDa) and three acidic fractions (average molecular weights 1300 kDa). Structural analysis of the respective polysaccharide fractions by quantitative and qualitative determination of the sugar composition and linkage analysis indicated that all acidic fractions contained an acidic arabinogalactan with a rhamnose-galactoronic acid backbone and highly branched arabinose-galactose side chains attached by the rhamnose residues to the backbone. The neutral fraction consisted of a neutral arabinogalactan beside minor amounts (about 10%) of a low substituted xyloglucan. Further studies on interaction between the 1340 kDa acidic rhamno-arabinogalactans II and III and mistletoe lectin Viscum album agglutinin I (VAA I) revealed binding capacities between these compounds, while the neutral polymers interacted significantly less with VAA I. Only partial binding of VAA I was observed by incubation of the lectin with polysaccharide II. Similar interactions of polysaccharide fraction III with VAA I was measured in a BIACRORE biosensore system. Using the hemagglutination test, increased agglutination of erythrocytes was observed when mistletoes lectin I and the respective polysaccharide fractions were present together in the assay. All these data indicate clearly strong interaction between VAA I and Viscum polysaccharides.

[Value of vasoactive drugs in conservative therapy of peripheral arterial occlusive disease]. / Stellenwert vasoaktiver Pharmaka in der konservativen Therapie der peripheren arteriellen Verschlusskrankheit.

Based on controlled clinical trials, vasoactive drugs (pentoxyfylline, naftidrofuryl, buflomedil) and prostanoids (alprostadil, iloprost) are used in symptomatic peripheral arterial occlusive disease: In stage II, vasoactive drugs are prescribed in order to improve leg hyperemia in response to muscular work; in complicated stage II (e.g., non-healing traumatic skin lesions), and in stages III/IV, prostanoids are given in order to increase forefoot skin blood flow. These drugs should only be applied if neither angioplasty nor vascular surgery are recommendable, possible, or successful after complete angiological diagnostic. It should be considered that these substances have a rather limited clinical efficacy in both stage II and stages III/IV and therefore attempts should be made to select patients with a reasonable chance to benefit. These drugs are also not suitable for primary or secondary prophylaxis against the progression of vascular lesions.

Photohaemolytic activity of chlorophyll degradation products in a mistletoe extract.

An aqueous mistletoe extract containing liposome-like vesicles from chloroplast membranes develops a photohaemolytic activity, which is a function of the temperature. This is correlated with the degradation of chlorophyll. The photolytic activity of the chlorophyllides initially produced in the process was detected by haemolysis. The quantification of the chlorophyllides was performed using HPLC. On being exposed to light, the isolated green vesicle fraction bleaches out extremely rapidly in the presence of traces of oxygen. In this process, the photolytic activity also disappears. In crude extracts, this only occurs to a small extent.

Oligoclonal in vitro response of CD4 T cells to vesicles of mistletoe extracts in mistletoe-treated cancer patients.

Mistletoe (Viscum album) extracts are widely used in adjuvant cancer therapy. We have investigated the in vitro responsiveness of T cells from mistletoe-treated cancer patients and untreated healthy donors to various preparations of mistletoe extracts. Proliferation of peripheral blood mononuclear cells from treated but not from untreated patients was observed in response to therapeutically used mistletoe extracts prepared from apple (mali) or pine (pini) host trees. The strongest proliferation was induced by a vesicle preparation of mali extract. Activation was strongly inhibited by interleukin-10. Using a newly developed flow-cytometry assay, we determined that cell growth was restricted to CD4 T cells. Analysis with a panel of monoclonal antibodies against the variable region of the T cell receptor beta chain (V beta) revealed an oligoclonal pattern of CD4 T cell activation. These results indicate that therapeutic administration of mistletoe extracts sensitizes a restricted set of CD4 T lymphocytes in mistletoe-treated patients.

Stimulation of the specific immune system by mistletoe extracts.

We have investigated the in vitro responsiveness of T cells from mistletoe-treated cancer patients and untreated donors to various preparations of mistletoe extracts. Proliferation of peripheral blood mononuclear cells from treated but not from untreated patients was observed in response to therapeutically used mistletoe extracts prepared from apple (mali) or pine (pini) host trees. The strongest proliferation was induced by vesicle preparation of mali extract. Using a newly developed flow cytometry assay (standard cell dilution assay), we determined that cell growth was restricted to CD4+ T cells. Analysis with a panel of monoclonal antibodies against variable regions of the T cell receptor beta chain (V beta) revealed an oligoclonal pattern of CD4+ T cell activation. These results indicate that therapeutic administration of mistletoe extracts sensitized a restricted set of CD4+ T lymphocytes in mistletoe-treated patients. Lymphocytes from untreated donors are only stimulated with heat-treated mistletoe extracts. The responding T cells are gamma delta T cells and express variable T cell receptor elements V gamma 9 and V delta 2. The gamma delta-stimulating activity of heat-treated mistletoe extracts is sensitive to treatment with alkaline phosphatase but not with proteinase K, indicating that the ligands are non-proteinaceous phosphate-containing compounds.

Activation of human gamma delta T-cells by heat-treated mistletoe plant extracts.

Various microorganisms including mycobacteria, other bacteria and parasites such as Plasmodium falciparum are known to activate human gamma delta T-cells in vitro. In this study, we demonstrate that heat-treated (but not untreated) mistletoe extracts similarly stimulate human gamma delta T-cells during in vitro culture. The responding T-cells express the variable T-cell receptor elements V gamma 9 and V delta 2. The gamma delta-stimulating activity of heat-treated mistletoe extracts is sensitive to treatment with alkaline phosphatase but not with proteinase K, indicating that the ligands are non-proteinaceous phosphate-containing compounds. Mistletoe-derived ligands share these features with the previously defined mycobacteria-derived ligands for gamma delta T-cells.

Attractors and quasi-attractors in the cutaneous perfusion in human subjects and patients: "chaotic" or adaptive behaviour?

The causes of the aperiodic fluctuations in the perfusion of the skin (volar hand, measured by the Laser-Doppler (LD) technique) of healthy human subjects were studied were studied by simultaneous recording of the fluctuations of local blood content (reflectiophotoplethysmography (rPPG)) and those in the skin of the glabella. Various thermoregulatory situations were provoked by exposing 12 subjects to 18, 21, 24 and 27 degrees C ambient temperature; in addition, the hands were placed at, below and above heart level. In mentally relaxed subjects (evidenced by a stable approx. 0.015-Hz rhythm in the glabellar rPPG signal), there was perfect temporal correlation between aperiodic LDA and rPPG signal under all thermoregulatory conditions. Clearly identifiable episodes of retardation associated with skin bleaching, asymmetrical shape of LDA and rPPG signals were taken as indicators of episodic sympathetic skin constrictor (SSC) activity. In synergetic terms, the modulated SSC activity operates as transient "quasi-attractor'. A notable exception occurred: when the hand was placed below heart at 27 degrees C ambient temperature, a sinusoidal periodic fluctuation (approx. 0.03 Hz) in the LDA evolved. These were not seen the rPPG signal, i.e., coherence between LDA and rPPG dynamics was lost). Lack of coherency between LDA-rPPG, also observed in patients with autonomic neuropathy and decompensated forms of peripheral arterial disease, suggesting predominance of spontaneously oscillating myogenic vasomotion after removal of temporally variable SSC drive. Stable vasomotion is regarded as a synergetically stereotyped reaction rather than as "well-ordered' stable attractor mode of operation.

Topographical evaluation of skin perfusion patterns in peripheral arterial occlusive disease by means of computer-assisted fluorescein perfusography.

OBJECTIVE: To evaluate the clinical impact of computer-assisted fluorescein perfusography in peripheral arterial occlusive disease (PAOD). DESIGN: Foot and calf skin perfusion was visualised by intravenous fluorescein injection. Fluorescein influx was recorded photographically and converted into functional images of fluorescein appearance times (AT) by means of digital film processing. SETTING: Vascular Laboratory of Clinic for Vascular Disease. MATERIALS: 249 patients with PAOD. Among 481 limbs studied, 83 legs presented with patent arteries, 70 with asymptomatic obstructions (Stage I), 170 with claudication (Stage II) and 158 with rest pain and skin lesions (Stage III/IV). CHIEF OUTCOME MEASURES: Forefoot and calf mean ATs and standard deviations (SD) served as arbitrary measures of regional skin perfusion rates and their homogeneity, respectively. MAIN RESULTS: In the control legs, a homogeneous and fast fluorescence appearance was observed (medians at the foot: AT33.4 s, SD 3.6). In stage II disease, AT (39.9 s, SD 5.6) were slightly impaired as compared to limbs with patent arteries or stage I disease (p < 0.01). Ninety-seven out of the 158 legs in stage III/IV could be managed by conservative therapy. According to fluorescein-perfusography, they did not differ from stage II disease (AT 38.8 s, SD 6.1). Sixty-one limbs were clinically affected by critical ischaemia. They exhibited a markedly delayed and heterogenous fluorescein influx at the foot (AT 77.3 s, SD 26.5, p < 0.01 vs all other groups). Non-fluorescent areas occurred in 53% compared to only 1% of limbs with and without critical ischaemia, respectively. Retrospectively, predictive values of fluorescein perfusography in identifying a critical limb ischaemia (accuracy 93%) were superior to the ankle systolic arterial pressure determination (accuracy 80%). CONCLUSIONS: Fluorescein perfusography seems to be of diagnostic and prognostic use in PAOD in stage III/IV where inflammatory and ischaemic patterns of dye appearance can be distinguished.

Fractionated total body irradiation and high-dose VP-16 with purged autologous bone marrow rescue for children with high risk relapsed acute lymphoblastic leukemia.

Twenty-two children with ALL in high risk second (n = 13), third or subsequent complete remission (n = 9) were treated with high-dose VP-16 60 mg/kg and fractionated total body irradiation (fTBI) 12 Gy, 2 x 2 Gy daily followed by autologous BM rescue. Prior to transplantation all patients had been treated according to intensive German BFM front-line or BFM relapse protocols. In all cases the marrow was purged using monoclonal antibodies attached to magnetic microspheres. All patients engrafted. There was no severe toxicity related to the pre-transplant high-dose chemoradiotherapy. Two patients died in the early course of transplantation from infections (Legionella and Aspergillus). Sixteen patients relapsed within 259 days (median 109 days); 13 died from leukemia. Four patients are alive in CR at a median of 1328 days with a Karnofsky score of 100%. The Kaplan-Meier estimation shows a probability of event-free survival (EFS) of 18% and a probability of relapse of 80%. Considering the otherwise poor prognosis of these children the results are acceptable although the high relapse rate is still disappointing. We conclude that high-dose VP-16 and fTBI combined with ABMT is a curative treatment for some children and should therefore be considered for those who lack an HLA-identical sibling donor. In future better therapy concepts are needed either in pre-transplant conditioning regimens or in post-transplant treatment schedules.

In vitro effects of mistletoe extracts and mistletoe lectins. Cytotoxicity towards tumor cells due to the induction of programmed cell death (apoptosis).

The in vitro effects of therapeutically administered mistletoe extracts (ABNOBAviscum) and pure mistletoe lectins (mainly mistletoe lectin I) on a variety of human and murine tumor cell lines have been investigated. Mistletoe extracts and purified mistletoe lectins inhibited in vitro the growth of all tumor cell lines tested including B cell hybridomas, P815, EL-4, Ke37, MOLT-4 and U937. The mechanism of growth arrest was shown to be due to the induction of programmed cell death (apoptosis). Thus, fragmentation of genomic DNA into oligonucleosomal bands of approximately 200 base pairs in length was observed within 20 h when tumor cells were incubated with mistletoe extracts or lectins. These data point to a rational basis for the direct cytotoxic effects of mistletoe extracts and lectins apart from the postulated immunostimulatory properties of these agents.

[Technical concepts of retrograde venous perfusion with fluorescein]. / Nachweis des technischen Konzepts der retrograden Venenperfusion mittels Fluoreszein.

Regional intravenous drug application has been suggested to increase peripheral tissue concentrations in severe arterial insufficiency. Within this study basic operational characteristics of the retrograde venous perfusion technique (RVP) were evaluated in 10 patients with intermittent claudication. Fluorescein was used to visualize the intracutaneous distribution of the injected fluid. Subjects were divided into 2 groups. The "cocktail-group" received 100 ml of a homogenous fluorescein dilution. To compare the effect of injecting a small sample, 45 ml of unstained solution were given before and after 10 ml of concentrated dye ("fraction-group"). Photographic recording of skin fluorescence distal to the arterial tourniquet revealed a rapid and complete staining of the lower leg including the toes in the cocktail-group. In the fraction-group, however, skin fluorescence appeared markedly delayed and, more over, did not spread down to the toes in 2 cases. In conclusion, drugs can be applied by RVP provided that they are dissolved in a sufficient amount of fluid of 100 ml at least.

[Regional drug-induced sympathicolysis in patients with arterial occlusive disease]. / Die regionale medikamentös induzierte Sympathikolyse bei Patienten mit AVK.

Regional drug-induced sympathectomy is a modification of Bier's block technique. By applying this method skin perfusion in patients with occlusions of lower leg arteries or functional disturbance of peripheral circulation can be improved. Under optimal technical conditions effects of regional and surgical sympathectomy are comparable. Fluorescence angiography can be applied to assess the effectiveness of drug-induced as well as surgical sympathectomy.