Regulation of neurosteroid allopregnanolone biosynthesis in the rat spinal cord by glycine and the alkaloidal analogs strychnine and gelsemine.

The neurosteroid allopregnanolone (3alpha,5alpha-THP) is well characterized as a potentially therapeutic molecule which exerts important neurobiological actions including neuroprotective, antidepressant, anxiolytic, anesthetic and analgesic effects. We have recently observed that neurons and glial cells of the rat spinal cord (SC) contain various key steroidogenic enzymes such as 5alpha-reductase and 3alpha-hydroxysteroid oxido-reductase which are crucial for 3alpha,5alpha-THP biosynthesis. Furthermore, we demonstrated that the rat SC actively produces 3alpha,5alpha-THP. As the key factors regulating neurosteroid production by nerve cells are unknown and because glycine is one of the pivotal inhibitory neurotransmitters in the SC, we investigated glycine effects on 3alpha,5alpha-THP biosynthesis in the rat SC. Glycine markedly stimulated [(3)H]-progesterone conversion into [(3)H]3alpha,5alpha-THP by SC slices. The alkaloid strychnine, well-known as a glycine receptor (Gly-R) antagonist, blocked glycine stimulatory effect on 3alpha,5alpha-THP formation. Gelsemine, another alkaloid containing the same functional groups as strychnine, increased 3alpha,5alpha-THP synthesis. The stimulatory effects of glycine and gelsemine on 3alpha,5alpha-THP production were additive when the two drugs were combined. These results demonstrate that glycine and gelsemine, acting via Gly-R, upregulate 3alpha,5alpha-THP biosynthesis in the SC. The data also revealed a structure-activity relationship of the analogs strychnine and gelsemine on neurosteroidogenesis. Possibilities are opened for glycinergic agents and gelsemine utilization to stimulate selectively 3alpha,5alpha-THP biosynthetic pathways in diseases evoked by a decreased neurosteroidogenic activity of nerve cells.

Effect of homeopathy on analgesic intake following knee ligament reconstruction: a phase III monocentre randomized placebo controlled study.

WHAT IS ALREADY KNOWN ABOUT THIS SUBJECT: The efficacy of homeopathy is still under debate and a recent meta-analysis recommended further randomized double-blind clinical trials to identify any clinical situation in which homeopathy might be effective. WHAT THIS STUDY ADDS: The complex of homeopathy tested in this study (Arnica montana 5 CH, Bryonia alba 5 CH, Hypericum perforatum 5 CH and Ruta graveolens 3 DH) is not superior to placebo in reducing 24 h morphine consumption after knee ligament reconstruction. AIMS: The efficacy of homeopathy is still under debate. The objective of this study was to assess the efficacy of homeopathic treatment (Arnica montana 5 CH, Bryonia alba 5 CH, Hypericum perforatum 5 CH and Ruta graveolens 3 DH) on cumulated morphine intake delivered by PCA over 24 h after knee ligament reconstruction. METHODS: This was an add-on randomized controlled study with three parallel groups: a double-blind homeopathic or placebo arm and an open-label noninterventional control arm. Eligible patients were 18-60 years old candidates for surgery of the anterior cruciate ligament. Treatment was administered the evening before surgery and continued for 3 days. The primary end-point was cumulated morphine intake delivered by PCA during the first 24 h inferior or superior/equal to 10 mg day(-1). RESULTS: One hundred and fifty-eight patients were randomized (66 in the placebo arm, 67 in the homeopathic arm and 25 in the noninterventional group). There was no difference between the treated and the placebo group for primary end-point (mean (95% CI) 48% (35.8, 56.3), and 56% (43.7, 68.3), required less than 10 mg day(-1) of morphine in each group, respectively). The homeopathy treatment had no effect on morphine intake between 24 and 72 h or on the visual analogue pain scale, or on quality of life assessed by the SF-36 questionnaire. In addition, these parameters were not different in patients enrolled in the open-label noninterventional control arm. CONCLUSIONS: The complex of homeopathy tested in this study was not superior to placebo in reducing 24 h morphine consumption after knee ligament reconstruction.

A potentized homeopathic drug, Arsenicum Album 200, can ameliorate genotoxicity induced by repeated injections of arsenic trioxide in mice.

Groundwater arsenic contamination has become a menacing global problem. No drug is available until now to combat chronic arsenic poisoning. To examine if a potentized homeopathic remedy, Arsenicum Album-200, can effectively combat chronic arsenic toxicity induced by repeated injections of Arsenic trioxide in mice, the following experimental design was adopted. Mice (Mus musculus) were injected subcutaneously with 0.016% arsenic trioxide at the rate of 1 ml/100 g body weight, at an interval of 7 days until they were killed at day 30, 60, 90 or 120 and were divided into three groups: (i) one receiving a daily dose of Arsenicum Album-200 through oral administration, (ii) one receiving the same dose of diluted succussed alcohol (Alcohol-200) and (iii) another receiving neither drug, nor succussed alcohol. The remedy or the placebo, as the case may be, was fed from the next day onwards after injection until the day before the next injection, and the cycle was repeated until the mice were killed. Two other control groups were also maintained: one receiving only normal diet, and the other receiving normal diet and succussed alcohol. Several toxicity assays, such as cytogenetical (chromosome aberrations, micronuclei, mitotic index, sperm head anomaly) and biochemical (acid and alkaline phosphatases, lipid peroxidation), were periodically made. Compared with controls, the drug fed mice showed reduced toxicity at statistically significant levels in respect of all the parameters studied, thereby indicating protective potentials of the homeopathic drug against chronic arsenic poisoning.

A systematic review of the quality of homeopathic pathogenetic trials published from 1945 to 1995.

BACKGROUND: The quality of information gathered from homeopathic pathogenetic trials (HPTs), also known as 'provings', is fundamental to homeopathy. We systematically reviewed HPTs published in six languages (English, German, Spanish, French, Portuguese and Dutch) from 1945 to 1995, to assess their quality in terms of the validity of the information they provide. METHODS: The literature was comprehensively searched, only published reports of HPTs were included. Information was extracted by two reviewers per trial using a form with 87 items. Information on: medicines, volunteers, ethical aspects, blinding, randomization, use of placebo, adverse effects, assessments, presentation of data and number of claimed findings were recorded. Methodological quality was assessed by an index including indicators of internal and external validity, personal judgement and comments of reviewers for each study. RESULTS: 156 HPTs on 143 medicines, involving 2815 volunteers, produced 20,538 pathogenetic effects (median 6.5 per volunteer). There was wide variation in methods and results. Sample size (median 15, range 1-103) and trial duration (mean 34 days) were very variable. Most studies had design flaws, particularly absence of proper randomization, blinding, placebo control and criteria for analysis of outcomes. Mean methodological score was 5.6 (range 4-16). More symptoms were reported from HPTs of poor quality than from better ones. In 56% of trials volunteers took placebo. Pathogenetic effects were claimed in 98% of publications. On average about 84% of volunteers receiving active treatment developed symptoms. The quality of reports was in general poor, and much important information was not available. CONCLUSIONS: The HPTs were generally of low methodological quality. There is a high incidence of pathogenetic effects in publications and volunteers but this could be attributable to design flaws. Homeopathic medicines, tested in HPTs, appear safe. The central question of whether homeopathic medicines in high dilutions can provoke effects in healthy volunteers has not yet been definitively answered, because of methodological weaknesses of the reports. Improvement of the method and reporting of results of HPTs are required. REFERENCES: References to all included RCTs are available on-line at.

A deletion in exon 3 results in a new HLA-A*01 null allele (A*0115N) in a Martinican woman.

We report the identification of a new HLA-A null allele, HLA-A*0115N. This null allele has been identified within the A*01 group by a combination of serological and molecular typing [Polymerase chain reaction (PCR) sequence-specific primers, PCR sequence-specific oligoprobes and sequence-based typing (SBT)] in a potential intrafamilial bone marrow donor from Martinique (French West Indies). To characterize this A*01 null allele, we performed DNA typing by PCR-SBT on genomic DNA from the beginning of exon 2 (position 84) through the end of the exon 4 (position 895) and revealed a nucleotide deletion at the end of the exon 3. This sole difference between the new allele and the HLA-A*0101 generates a premature stop codon (TGA) in the beginning of exon 4. This deletion most likely explains the lack of cell surface expression of the encoded protein despite the presence of A*01 allele. The absence of correct expression of the antigen on the cell surface was confirmed by one-dimensional isoelectric focusing (1D-IEF). To date, this is the fourth null allele described within the A*01 group.

Improvement of flow cytometric analysis of basophil activation inhibition by high histamine dilutions. A novel basophil specific marker: CD 203c.

BACKGROUND: Histamine is known to elicit a negative feedback effect on anti-IgE and allergen-induced basophil activation. A series of experiments performed between 1981 and 1995 using a manual method showed biological activity of highly diluted histamine. Most of the experiments used histamine in the range 10(-30) (15C)-10(-36) M (18C). These results were confirmed by automated flow cytometry, but this method is based on the selection of basophils by anti-IgE and analysis of basophil activation by anti-CD 63, showing significant but relatively low inhibition (approximately 14%), insufficient to convince the scientific community of the reality of the phenomenon. OBJECTIVE: We investigated if the use of CD 203c, a basophil specific, earlier marker than CD 63 of the activation cascade, increased the sensitivity of the method, testing two target histamine dilutions, 10(-4) (2C) and 10(-32) M (16C). METHODS: Basophils, obtained from buffy coats, were pre-incubated with the histamine dilutions and activated by two agonists: anti-IgE and fMLP (formyl-methionyl-leucyl-phenylalanine peptide). Basophil activation was stopped with EDTA. The cells were labelled with anti-IgE, anti-CD 13 and anti-CD 14 for basophil selection, and anti-CD 63 and anti-CD 203c for basophil activation. Results were expressed in up-regulation percentage for CD 63 or mean intensity of fluorescence (MFI) for CD 203c. RESULTS: Histamine 10(-4) M (2C) and histamine 10(-32) M (16C) were capable of inhibiting both IgE-dependent (anti-IgE) and IgE-independent (fMLP) basophil activation. The percentage inhibition depended on the activation marker used. The highest inhibition for histamine dilution 16C was observed with CD 203c (38%, P<0.001), approximately half the inhibition observed with histamine 2C (73%). CONCLUSION: These new flow cytometric protocols confirmed that high dilutions of histamine may inhibit basophil activation and that the inhibitory effect is not restricted to IgE-dependent activation. The use of CD 203c instead of CD 63 increased the magnitude of the response.

Histamine dilutions modulate basophil activation.

BACKGROUND: In order to demonstrate that high dilutions of histamine are able to inhibit basophil activation in a reproducible fashion, several techniques were used in different research laboratories. OBJECTIVE: The aim of the study was to investigate the action of histamine dilutions on basophil activation. METHODS: Basophil activation was assessed by alcian blue staining, measurement of histamine release and CD63 expression. Study 1 used a blinded multi-centre approach in 4 centres. Study 2, related to the confirmation of the multi-centre study by flow cytometry, was performed independently in 3 laboratories. Study 3 examined the histamine release (one laboratory) and the activity of H(2) receptor antagonists and structural analogues (two laboratories). RESULTS: High dilutions of histamine (10(-30)-10(-38) M) influence the activation of human basophils measured by alcian blue staining. The degree of inhibition depends on the initial level of anti-IgE induced stimulation, with the greatest inhibitory effects seen at lower levels of stimulation. This multicentre study was confirmed in the three laboratories by using flow cytometry and in one laboratory by histamine release. Inhibition of CD63 expression by histamine high dilutions was reversed by cimetidine (effect observed in two laboratories) and not by ranitidine (one laboratory). Histidine tested in parallel with histamine showed no activity on this model. CONCLUSIONS: In 3 different types of experiment, it has been shown that high dilutions of histamine may indeed exert an effect on basophil activity. This activity observed by staining basophils with alcian blue was confirmed by flow cytometry. Inhibition by histamine was reversed by anti-H2 and was not observed with histidine these results being in favour of the specificity of this effect We are however unable to explain our findings and are reporting them to encourage others to investigate this phenomenon.

[Homeopathic specialties as substitutes for benzodiazepines: double-blind vs. placebo study]. / Spécialités homéopathiques en substitution de benzodiazépines: étude en double-insu vs. placebo.

The objective of this study was to compare the efficacy of homeopathic drugs Homéogène 46 and Sédatif PC with a placebo as substitute for benzodiazepines in patients treated for at least 3 months with low doses (less than 10 mg/d of diazepam equivalents). A double-blind randomized trial was carried out in general practice. The treatment lasted one month. Several rating scales were used. The main outcome was success/failure defined according to the doctor's clinical judgement and interruption of treatment. A total of 61 patients were randomized, and 19 interrupted their treatment. Comparability between the groups was good. No statistically significant difference between homeopathic drugs and placebo was observed for the main outcome or for the secondary outcomes. The lack of statistical power due to accrual difficulties limits the conclusions of this trial which did not confirm the efficacy of homeopathic drugs in this indication.

[Treatment of pain due to unwanted lactation with a homeopathic preparation given in the immediate post-partum period]. / Traitement de la douleur de la montée laiteuse non souhaitée par homéopathie dans le post-partum immédiat.

Dopaminergic agonists, such as Parlodel((R)), are now widely used to inhibit lactation. However, some countries, such as the United States, no longer use these drugs in this indication because of their sometimes serious adverse effects. In this context, the authors tested a homeopathic treatment designed for parturients unable or not wanting to breastfeed. The APIS MELLIFICA 9 CH and BRYONIA 9 CH combination was chosen for its anti-inflammatory and analgesic effects. 71 patients were included in this double-blind placebo-controlled study. All received basic treatment comprising naproxen and fluid restriction. A significant improvement of lactation pain (main criterion of the study) was observed in parturients treated with homeopathy (p<0.02 on D2 and p<0.01 on D4). A similar effect (p<0.05 on D4) was observed for breast tension and spontaneous milk flow. No significant difference was observed for the other criteria of the study. The homeopathic combination studied was therefore effective on the pain of lactation and should be integrated into the therapeutic armamentarium.

Neurotropic, immunological and gastric effects of low doses of Atropa belladonna L., Gelsemium sempervirens L. and Poumon histamine in stressed mice.

Previous studies realized in the laboratory have indicated that application of experimental stress (such as unavoidable footshock) induced significant behavioral, gastric and immunological alterations in mice. The aim of this study was to evaluate effects of low doses of Atropa belladonna L., Gelsemium sempervirens L. and Poumon histamine on stress-induced behavioral, immunological and gastric alterations. Locomotor, postural and exploratory activities have been evaluated by two behavioral tests: light/dark box and staircase tests. Immunological studies were investigated to count white blood cells subpopulations (lymphocytes, neutrophils, monocytes and basophils) by coulter counter. The severity of gastric erosions was evaluated by microscopic technique in mice after experimental stress. The results have demonstrated that low doses of G. sempervirens L. and A. belladonna L. had a significant neurotropic and protective effects on behavioral and gastric alterations induced by experimental stress. The immunological protective effects observed were probably induced via their neurotropic effects. The P. histamine showed a significant immunoprotective and gastroprotective effect in mice exposed to experimental stress.

Time related neutralization of two doses acetyl salicylic acid.

Aspirin has a well established role in the prevention of arterial thrombosis. Discussion on the efficacy and safety of aspirin in the treatment and prophylaxis of thrombosis has become an important issue. In fact, hemorrhage complications are often associated with its use. On the other hand, previous studies showed unexpected thrombotic potencies associated with the presence of this drug at ultra low doses (ULD) in the circulation. In this study, we aimed to evaluate the effect of aspirin at ULD, injected 1, 2, or 3 hours after the administration of aspirin at 100 mg/kg, on hemostasis and bleeding in rats. We used an experimental model of thrombosis induced by laser beams to evaluate these effects. Platelet aggregation was determined by Cardinal and Flower method. Results from this investigation demonstrate that the neutralizing effect of aspirin at ULD did not operate significantly 1 hour after the injection of aspirin at 100 mg/kg. This effect was observed 2 and 3 hours after. The use of aspirin at ULD to neutralize the side effects of aspirin at high doses will reduce the hemorrhagic risk during extra corporeal circulation. The therapeutic benefit and safety of aspirin therapy in the treatment of cardiovascular diseases can be obtained.

Effects of acetyl salicylic acid therapy on an experimental thrombosis induced by laser beam.

Aspirin inhibits the synthesis of both platelet and vascular arachidonic acid metabolism which have opposite effects on platelet functions. The rationale for its clinical use as an antithrombotic drug has therefore been questioned. Therefore, we investigated the effects of acetylsalicylic acid (ASA) at 100 mg/kg on an experimental thrombosis induced by laser beams using different groups of rats that were previously treated with the same dose (100 mg/kg), according to the delay between the first and second injections. A partial occlusion was induced by laser beams in the rat mesenteric microvessels (15-25 m). The thrombus formed within seconds after the laser lesion; both it and the embolization which began within minutes after, were continuously accounted. Experiments were done on 11 groups of 5 animals each: 45 rats received a first injection of ASA at j(0) and a second injection 30 minutes before thrombosis induction at j(0)+x (x=2, 4, 6, 8, 9, 10, 12, 14 and 16 days). Different groups are defined according to the x value. The rats receiving NaCl 0.9% or a single injection of ASA at 100 mg/kg 30 minutes before thrombosis induction were used as control (Group I) and reference group (Group II) respectively. In this study, ASA treatment showed two types of results. The administration of ASA (100 mg/kg) 30 minutes before laser-induced thrombosis prevented thrombus formation. In the same way, ASA injected to rats already treated with the same dose 2 or 4 day later also demonstrated a potent antithrombotic effect. The same trends were observed with animals receiving the second injection (100 mg ASA) at j(0+8), j(0+12), j(0+14), and j(0+16). However, when injected to rats at j(0+6) and at j(0+10), ASA did not shown any effects on thrombus formation compared to the control (p>/=0.05). The same phases of ASA action were observed on the induced hemorrhagic time. The antithrombotic effects of the later second injection of ASA (100 mg/kg) were neutralized in rats previously receiving the same dose of this drug. This phenomenon seems to be periodic and is of great importance for the observance of ASA treatment.

Combination of two doses of acetyl salicylic acid: experimental study of arterial thrombosis.

The antithrombotic effect of high dose acetylsalicylic acid is well known, and recently, in vitro studies hinted the potent thrombotic effect of ultra-low dose of acetylsalicylic acid (<1mg/day) showing a significant decrease in bleeding time. In this study, we investigated the effect of a combination between a high and an ultra-low dosage (100 mg/kg+ 10(-30) mg/kg) on an arterial thrombosis induced by a laser beam. We used an intravital microscopic technique, allowing to evaluate (anti)-thromboembolic events at previously determined locations of microvasculature. Thrombus formation was induced by argon-laser shot. The instrumental test setup was completed with a video system, to select mesenteric arterioles with the same diameter (between 15 and 25 microm). The changes in platelet aggregability were determined by Cardinal and Flower method, and the concentration of acetylsalicylic acid in the plasma was measured by high pressure liquid chromatography. Antithrombotic effect of high dose (100 mg/kg) acetylsalicylic acid was confirmed in all results obtained. Asa injected at ultra-low dose (10(-30) mg/kg) had a potent thrombotic properties and decreased significantly the bleeding time. The subcutaneous administration of the combination of the two doses permitted to come back to the control values, and the bleeding time was shortened compared to control group.

Thromboembolic complications several days after a single-dose administration of aspirin.

The antithrombotic properties of acetyl salicylic acid (ASA) used at current doses are largely demonstrated. However, our previous study showed unexpected thrombotic potencies associated with the use of this drug. In this study we investigate the effect of aspirin on an experimental thrombosis induced by laser beams, according to its in vivo plasma concentration. Experiments were done on nine groups of seven Wistar male rats. The groups are defined by the delay between aspirin administration time and the laser-induced thrombosis time. Results from this study showed an enhancement of thromboembolic complications when thrombosis was induced 8 or 10 days after aspirin administration; the number of emboli and the duration of embolization are increased, compared to the control group. The prothrombotic properties of ASA demonstrated in this study, might limit its therapeutic benefit and might explain thromboembolic complications observed in some ASA-treated patients. These results also suggest a biological monitoring several days after aspirin administration to patients.