Rabies control in Europe: an overview of past, current and future strategies.

Europe's step-by-step approach to tackling canine and wildlife-mediated rabies has proven that the disease can be controlled and eliminated at the animal source. A decade of development work and support by the European Union has resulted in a successful oral rabies vaccination campaign for foxes and raccoon dogs, and provided the blueprint for eliminating wildlife-mediated rabies. In this paper, the authors examine the historical stages of animal rabies control and its eventual elimination in parts of Europe, particularly Western Europe. In addition, they consider current control measures and predict future challenges, many of which are faced by the vast and sparsely populated, rabies-endemic areas of Eastern Europe.

L'approche par étapes appliquée en Europe pour combattre la rage canine ainsi que celle transmise par la faune sauvage a démontré que cette maladie peut être contrôlée et éliminée à sa source animale. La stratégie conçue et soutenue par l'Union européenne pendant une décennie s'est concrétisée par une campagne de vaccination antirabique orale des renards et des chiens viverrins qui a été couronnée de succès et constitue un modèle pour l'élimination de la rage transmise par les animaux sauvages. Les auteurs examinent le déroulement des étapes successives de la lutte contre la rage animale et de son élimination dans certaines régions européennes, en particulier en Europe occidentale. Ils examinent également les mesures actuelles de lutte et annoncent les difficultés auxquelles il faudra faire face à l'avenir, en particulier dans les vastes régions d'Europe orientale où la population est clairsemée et la rage endémique.

El método gradual (por etapas) aplicado en Europa para combatir la rabia canina y la transmitida por animales silvestres ha demostrado que es posible controlar y eliminar la enfermedad en su foco animal de origen. Diez años de trabajo de desarrollo y de apoyo de la Unión Europea desembocaron en una fructífera campaña de vacunación antirrábica oral de zorros y mapaches, convertida ahora en un modelo para eliminar la rabia transmitida por animales silvestres. Los autores repasan las etapas históricas de la lucha contra la rabia animal hasta su eliminación final en ciertas partes de Europa, en particular en Europa Occidental. Además, examinan las medidas de control vigentes y auguran futuras dificultades, muchas de ellas en las vastas zonas de Europa Oriental donde la población es escasa y dispersa y la rabia es endémica.

Increased thalamic resting-state connectivity as a core driver of LSD-induced hallucinations.

OBJECTIVE: It has been proposed that the thalamocortical system is an important site of action of hallucinogenic drugs and an essential component of the neural correlates of consciousness. Hallucinogenic drugs such as LSD can be used to induce profoundly altered states of consciousness, and it is thus of interest to test the effects of these drugs on this system. METHOD: 100 µg LSD was administrated orally to 20 healthy participants prior to fMRI assessment. Whole brain thalamic functional connectivity was measured using ROI-to-ROI and ROI-to-voxel approaches. Correlation analyses were used to explore relationships between thalamic connectivity to regions involved in auditory and visual hallucinations and subjective ratings on auditory and visual drug effects. RESULTS: LSD caused significant alterations in all dimensions of the 5D-ASC scale and significantly increased thalamic functional connectivity to various cortical regions. Furthermore, LSD-induced functional connectivity measures between the thalamus and the right fusiform gyrus and insula correlated significantly with subjective auditory and visual drug effects. CONCLUSION: Hallucinogenic drug effects might be provoked by facilitations of cortical excitability via thalamocortical interactions. Our findings have implications for the understanding of the mechanism of action of hallucinogenic drugs and provide further insight into the role of the 5-HT2A -receptor in altered states of consciousness.

Non-allopathic adjuvant management of osteoarthritis by alkalinisation of the diet.

BACKGROUND: Osteoarthritis (OA) is a chronic condition. Nonsteroidal anti-inflammatory drugs recommended for treatment have serious adverse effects. A compelling body of anecdotal evidence alerted the authors to the therapeutic potential of dietary supplementation with Multiforce® (MF) Alkaline Powder for relief of OA symptoms. AIM: The aim of the study was to test the hypothesis that dietary supplementation with MF relieves clinical signs and symptoms of OA of the hands. SETTING: The study was done at the MEDSAC hospital in Somerset West, Western Cape, South Africa. METHODS: The research was conducted in two stages. An open interventional study (n = 40) confirmed the notion that MF 7.5 g twice daily is likely to be an effective alternative or adjunct for relief of symptoms of OA of the hands. The main study was conducted with 100 eligible, consenting volunteers (aged 47-89 years) according to a randomised, placebo-controlled, crossover design. Study duration was 56 days, 28 days per regimen; crossover to alternate regimens took place on day 28. RESULTS: Compared to placebo, MF intake over 28 days was associated with significant reductions ( p < 0.005) in pain, tenderness and stiffness of interphalangeal and metacarpophalangeal joints of the hand. Confirmation of systemic alkalinisation by MF, which is rich in organic anions in the form of citrate salts, was reflected by a significant and sustained increase in urine pH. CONCLUSION: A dietary supplement, Multiforce® Alkaline Powder, containing citrate salts which are converted into bicarbonate in vivo, was efficacious and safe as sole therapeutic intervention, significantly attenuating OA-associated signs and symptoms of the hands.

Green tea ingestion greatly reduces plasma concentrations of nadolol in healthy subjects.

This study aimed to evaluate the effects of green tea on the pharmacokinetics and pharmacodynamics of the ß-blocker nadolol. Ten healthy volunteers received a single oral dose of 30 mg nadolol with green tea or water after repeated consumption of green tea (700 ml/day) or water for 14 days. Catechin concentrations in green tea and plasma were determined. Green tea markedly decreased the maximum plasma concentration (C(max)) and area under the plasma concentration-time curve (AUC(0-48)) of nadolol by 85.3% and 85.0%, respectively (P < 0.01), without altering renal clearance of nadolol. The effects of nadolol on systolic blood pressure were significantly reduced by green tea. [(3)H]-Nadolol uptake assays in human embryonic kidney 293 cells stably expressing the organic anion-transporting polypeptides OATP1A2 and OATP2B1 revealed that nadolol is a substrate of OATP1A2 (Michaelis constant (K(m)) = 84.3 µmol/l) but not of OATP2B1. Moreover, green tea significantly inhibited OATP1A2-mediated nadolol uptake (half-maximal inhibitory concentration, IC(50) = 1.36%). These results suggest that green tea reduces plasma concentrations of nadolol possibly in part by inhibition of OATP1A2-mediated uptake of nadolol in the intestine.

Potencialidades fitoquímicas do melão (Cucumis melo L. ) na região Noroeste do Rio Grande do Sul - Brasil / Phytochemical potentialities of melon (Cucumis melo L. ) in the northwest region of Rio Grande do Sul - Brazil

O melão (Cucumis melo L.) é uma fruta muito apreciada por suas qualidades e sua produção vem crescendo e ganhando espaço no mercado nacional e internacional. Em regiões como o Noroeste do Rio Grande do Sul, destaca-se como uma nova alternativa de renda para vários agricultores. Neste contexto, o presente trabalho teve como objetivo analisar o potencial fitoquímico de alguns cultivares de melão da região Noroeste do Rio Grande do Sul. A análise fitoquímica utilizando como farmacógeno as folhas, foi realizada para a verificação da presença de metabólitos secundários, tais como: saponinas, cumarinas, cardiotônicos, cianogenéticos, alcalóides, taninos, antraquinonas, flavonoides, e óleos voláteis. Também foi avaliado o teor de suco a partir dos frutos. Dentre os cinco cultivares analisados, Gaúcho, Imperial, Hy Mark, Magelan, e Cantaloupe, o cultivar Gaucho apresentou a maior variedade em metabólitos secundários. Na avaliação do teor de suco a cultivar Magelan se destacou em comparação às demais cultivares testadas.

The melon (Cucumis melo L.) is a fruit highly appreciated for its qualities and its production has been growing and gaining space in the national and in the international market. In regions like the northwest of Rio Grande do Sul - Brazil, it stands out as a new income alternative for farmers. In this context, this study aimed to analyze the phytochemical potential of some melon cultivars in the northwest region of Rio Grande do Sul. The phytochemical analysis, using the leaves as pharmacogen, was performed to verify the presence of secondary metabolites such as saponins, coumarins, cardiac glycosides, cyanogenetic glicosides, alkaloids, tannins, anthraquinones, flavonoids and volatile oils. The juice content from the fruits was also evaluated. Among the five analyzed cultivars, Gaucho, Imperial, Hy Mark, Magelan and Cantaloupe, cultivar Gaucho had the greatest variety of secondary metabolites. In the assessment of the juice content, cultivar Magelan was superior compared to the remaining tested cultivars.

Functional electrical stimulation assisted cycling of patients with subacute stroke: kinetic and kinematic analysis.

BACKGROUND: Cycling is a safe and functionally effective exercise for patients with early post-stroke and poor balance. Such exercise is considered even more effective when functional electrical stimulation is added. Our principal aim was to determine the biomechanically quantifiable parameters of cycling that can be improved in patients with subacute hemiparesis by incorporating functional electrical stimulation. These parameters were defined as objective goals that can be achieved in clinical applications. A secondary aim was to determine whether they could be used to identify subjects who would benefit from such therapy. METHODS: Using a tricycle testbed, we tested 39 subacute (mean 10.9 weeks post-stroke (SD 5.9)), hemiplegic subjects. During isometric measurements we recorded volitional and electrically evoked crank torques, the latter at maximal tolerable intensity. During ergometric measurements, volitional pedaling was alternated with combined pedaling (volitional supported by stimulation), performed at 30-s intervals. Power, smoothness, and symmetry of cycling were evaluated. FINDINGS: Twenty-six percent of the subjects significantly improved the smoothness of their cycling with functional electrical stimulation. Only 8% and 10% significantly increased their power and symmetry, respectively. The improvement in smoothness significantly correlated with the capability of the individual to generate electrical torque (Spearman's rank correlation coefficient=0.66 at P=0.001). INTERPRETATION: The smoothness of cycling was the most sensitive parameter improved by functional electrical stimulation. This improvement depended on the amount of torque evoked, and the torque achieved, in turn, correlated with the tolerated intensity of stimulation.

[Survival after laser-induced interstitial thermotherapy of colorectal liver metastases--a comparison of first clinical experiences with current therapy results]. / Langzeitüberleben nach laserinduzierter interstitieller Thermotherapie kolorektaler Lebermetastasen--ein Vergleich erster klinischer Erfahrungen mit aktuellen Behandlungsergebnissen.

PURPOSE: Laser-induced interstitial thermotherapy (LITT) is a minimally invasive procedure for local tumor ablation. This report discusses the long-term results of this therapy in non-surgical patients with colorectal liver metastases. MATERIALS AND METHODS: In total, we treated 163 colorectal liver metastases in 85 patients by MR-guided LITT. RESULTS: The overall lethality was 1.1%. The mean survival period of all treated patients was 3.3+/-0.3 years (CI: 2.8; 3.8; Kaplan-Meier method) with a calculation start date of the diagnosis of the metastases. The mean survival period after LITT was 2.6+/-0.2 years (CI 2.2; 3.0). There was a substantial learning curve with respect to the indication for LITT and the technical procedure of the intervention itself. In the first period (50 patients; 1998-2001), the mean survival period was 2.2+/-0.2 years (CI: 1.8; 2.7). Thereafter (35 patients 2001-2006), the mean survival period increased to 2.9+/-0.3 years (CI: 2.4; 3.4; log rank Test; p=0.0097). In the latter group, the 1-year survival rate was 93% and the 3-year survival rate was 56%. CONCLUSION: MR-guided LITT is a treatment suitable for non-surgical patients suffering from colorectal liver metastases. The complication rate of the procedure is comparatively low. The survival periods after the procedure are longer than those reported for similar patient populations after chemotherapy alone.

[Contact dermatitis from polyacrylate in TENS electrode]. / Eczéma de contact aux polyacrylates du gel conducteur des électrodes de neurostimulation.

INTRODUCTION: Transcutaneous electric nerve stimulation (TENS) is useful for many chronic pains. It induces few serious side effects, but skin reactions are not rare. We report on two cases of contact dermatitis due to TENS electrodes by sensitization to the acrylate in TENS conductive gel. CASE REPORTS: A 50 year-old man suffered from post-traumatic lumbar pair. He developed eczematous lesions on the sites where the TENS electrodes were applied. Patch tests were positive with the TENS gel, with ethylene glycol dimethylacrylate (2 p. 100 petrolatum) and ethyl-acrylate (2 p. 100 petrolatum) on day 2 and 4 readings. A 54 Year-old man had a paralysis of the foot elevator following rupture of an aneurysm. After 2 months, he had an eczema on the sites where the TENS electrodes were applied. Patch tests were negative with the TENS electrodes but positive with 2-hydroxyethyl acrylate (0.1 p. 100 petrolatum), triethyleneglycol diacrylate (0.1 p. 100 petrolatum), 2-hydroxyethyl methacrylate (2 p. 100 petrolatum) and 2-hydroxypropyl methacrylate (2 p. 100 petrolatum) on day 2 and 4 readings. DISCUSSION: TENS transmits small electrical currents through the skin that induce the depolarization of the affected sensory nerve endings. They have few serious side effects but skin reactions such as irritation, burns or allergy to propylene glycol in the electrode gel, to the rubber of the electrodes (mercaptobenzothiazole) or to the metallic part of the electrodes, i.e. nickel, are not uncommon. To our knowledge, only one case of an allergy to the polyacrylates of TENS electrode gel has been previously reported in the literature. We emphasize that acrylate could be the main sensitizer in the more recently commercialized TENS electrodes and will propose alternative ways of treating patients sensitized to acrylate and who require treatment with TENS.

Combined surgical and antifungal treatment of a subcutaneous infection due to Paecilomyces lilacinus.

Paecilomyces lilacinus was the causal agent of a case of subcutaneous infection in a patient with liver cirrhosis. Surgical treatment in combination with systemic amphotericin B therapy led to complete recovery. Retrospectively performed microdilution testing revealed dose dependent in vitro susceptibility of the isolate to voriconazole (MIC = 2 g/ml) and terbinafine (MIC = 1 microg/ml).

Hyperpolarization-activated channels HCN1 and HCN4 mediate responses to sour stimuli.

Sour taste is initiated by protons acting at receptor proteins or channels. In vertebrates, transduction of this taste quality involves several parallel pathways. Here we examine the effects of sour stimuli on taste cells in slices of vallate papilla from rat. From a subset of cells, we identified a hyperpolarization-activated current that was enhanced by sour stimulation at the taste pore. This current resembled Ih found in neurons and cardio-myocytes, a current carried by members of the family of hyperpolarization-activated and cyclic-nucleotide-gated (HCN) channels. We show by in situ hybridization and immunohistochemistry that HCN1 and HCN4 are expressed in a subset of taste cells. By contrast, gustducin, the G-protein involved in bitter and sweet taste, is not expressed in these cells. Lowering extracellular pH causes a dose-dependent flattening of the activation curve of HCN channels and a shift in the voltage of half-maximal activation to more positive voltages. Our results indicate that HCN channels are gated by extracellular protons and may act as receptors for sour taste.

Glucosylceramide synthases, a gene family responsible for the biosynthesis of glucosphingolipids in animals, plants, and fungi.

Glucosylceramides are membrane lipids in most eukaryotic organisms and in a few bacteria. The physiological functions of these glycolipids have only been documented in mammalian cells, whereas very little information is available of their roles in plants, fungi, and bacteria. In an attempt to establish appropriate experimental systems to study glucosylceramide functions in these organisms, we performed a systematic functional analysis of a glycosyltransferase gene family with members of animal, plant, fungal, and bacterial origin. Deletion of such putative glycosyltransferase genes in Candida albicans and Pichia pastoris resulted in the complete loss of glucosylceramides. When the corresponding knock-out strains were used as host cells for homologous or heterologous expression of candidate glycosyltransferase genes, five novel glucosylceramide synthase (UDP-glucose:ceramide glucosyltransferase) genes were identified from the plant Gossypium arboreum (cotton), the nematode Caenorhabditis elegans, and the fungi Magnaporthe grisea, Candida albicans, and P. pastoris. The glycosyltransferase gene expressions led to the biosynthesis of different molecular species of glucosylceramides that contained either C18 or very long chain fatty acids. The latter are usually channeled exclusively into inositol-containing sphingolipids known from Saccharomyces cerevisiae and other yeasts. Implications for the biosynthesis, transport, and function of sphingolipids will be discussed.

Laser-induced interstitial thermotherapy of liver metastases in an interventional 0.5 Tesla MRI system: technique and first clinical experiences.

Laser-induced interstitial thermotherapy (LITT) surveyed by magnetic resonance imaging (MRI) has been shown to be effective in various applications. The laser treatment of colorectal liver metastases usually requires a separate device (e.g., ultrasound or CT) to position the laser applicator. In this study, we used an interventional 0.5 T MRI system, allowing both the navigation to the target tissue and on-line thermometry. Laser irradiation was performed using a near-infrared laser source combined with a cooled laser light guide. We treated 20 patients exhibiting a total of 58 colorectal liver metastases. Clinically relevant complications did not occur. No residual tumor was observed after laser irradiation in all metastases with a diameter below 2 cm. Metastases with a mean diameter between 2 and 3 cm demonstrated total necrosis in 71%, while in larger tumors this proportion decreased to 46% (diameter, 3-4 cm) and 30% (diameter, >4 cm), respectively. We conclude that LITT, guided by the employed interventional MRI system, is feasible and safe. The results suggest a more aggressive treatment, especially for larger metastases. J. Magn. Reson. Imaging 2001;13:729-737.

Molecular cloning and characterization of the human protein kinase D2. A novel member of the protein kinase D family of serine threonine kinases.

We have isolated the full-length cDNA of a novel human serine threonine protein kinase gene. The deduced protein sequence contains two cysteine-rich motifs at the N terminus, a pleckstrin homology domain, and a catalytic domain containing all the characteristic sequence motifs of serine protein kinases. It exhibits the strongest homology to the serine threonine protein kinases PKD/PKCmicro and PKCnu, particularly in the duplex zinc finger-like cysteine-rich motif, in the pleckstrin homology domain and in the protein kinase domain. In contrast, it shows only a low degree of sequence similarity to other members of the PKC family. Therefore, the new protein has been termed protein kinase D2 (PKD2). The mRNA of PKD2 is widely expressed in human and murine tissues. It encodes a protein with a molecular mass of 105 kDa in SDS-polyacrylamide gel electrophoresis, which is expressed in various human cell lines, including HL60 cells, which do not express PKCmicro. In vivo phorbol ester binding studies demonstrated a concentration-dependent binding of [(3)H]phorbol 12,13-dibutyrate to PKD2. The addition of phorbol 12,13-dibutyrate in the presence of dioleoylphosphatidylserine stimulated the autophosphorylation of PKD2 in a synergistic fashion. Phorbol esters also stimulated autophosphorylation of PKD2 in intact cells. PKD2 activated by phorbol esters efficiently phosphorylated the exogenous substrate histone H1. In addition, we could identify the C-terminal Ser(876) residue as an in vivo phosphorylation site within PKD2. Phosphorylation of Ser(876) of PKD2 correlated with the activation status of the kinase. Finally, gastrin was found to be a physiological activator of PKD2 in human AGS-B cells stably transfected with the CCK(B)/gastrin receptor. Thus, PKD2 is a novel phorbol ester- and growth factor-stimulated protein kinase.

Selected physical and chemical properties of commercial Hypericum perforatum extracts relevant for formulated product quality and performance.

OBJECTIVE: The complex composition-activity relationship of botanicals such as St John's Wort (SJW) presents a major challenge to product development, manufacture, and establishment of appropriate quality and performance standards for the formulated products. As part of a larger study aimed at addressing that challenge, the goals of the present study are to (1) determine and compare the phytochemical profiles of 3 commercial SJW extracts; (2) assess the possible impact of humidity, temperature, and light on their stability; and (3) evaluate several physical properties important to the development of solid dosage forms for these extracts. METHODS: An adapted analytical method was developed and validated to determine phytochemical profiles and assess their stability. The extract physical properties measured were particle size (Malvern Mastersizer), flow (Carr's compressibility index; minimum orifice diameter), hygroscopicity (method of Callahan et al), and low-pressure compression physics (method of Heda et al). RESULTS: The phytochemical properties differed greatly among the extracts and were extremely sensitive to changes in storage conditions, with marked instability under conditions of elevated humidity. All extracts exhibited moderate to free-flow properties and were very hygroscopic. Compression properties varied among the extracts and differed from a common use excipient, microcrystalline cellulose. CONCLUSIONS: Three commercial sources of SJW extracts exhibited different physical and chemical properties. Standardization to 1 or 2 marker compounds does not ensure chemical equivalence nor necessarily equivalent pharmacological activity. Flow and compression properties appear suitable for automatic capsule-filling machines, but hygroscopicity and the moisture sensitivity of the phytochemical profile are concerns.

Virological and immunological effects of antioxidant treatment in patients with HIV infection.

BACKGROUND: Intracellular oxidative stress in CD4+ lymphocytes due to disturbed glutathione homeostasis may lead to impaired lymphocyte functions and enhanced HIV replication in patients with HIV infection, especially in those with advanced immunodeficiency. The aim of the present study was to assess whether short-term, high-dose antioxidant treatment might have effects on immunological and virological parameters in patients with HIV infection. MATERIALS AND METHODS: In this pilot study, we examined virological and immunological effects of antioxidant combination treatment for 6 days with high doses of N-acetylcysteine (NAC) and vitamin C in 8 patients with HIV infection. The following were assayed before, during and after antioxidant treatment: HIV RNA plasma levels; numbers of CD4+, CD8+, and CD14+ leukocytes in blood; plasma thiols; intracellular glutathione redox status in CD4+ lymphocytes and CD14+ monocytes; lymphocyte proliferation; lymphocyte apoptosis and plasma levels of tumour necrosis factor (TNF)alpha; soluble TNF receptors and neopterin in plasma. RESULTS: No significant changes in HIV RNA plasma levels or CD4+ lymphocyte counts in blood were noted during antioxidant treatment in the patient group. However, in the 5 patients with the most advanced immunodeficiency (CD4+ lymphocyte counts < 200 x 106 L(-1)), a significant rise in CD4+ lymphocyte count, a reduction in HIV RNA plasma level of 0.8 log, an enhanced lymphocyte proliferation and an increased level of intracellular glutathione in CD4+ lymphocytes were found. No change in lymphocyte apoptosis was noted. CONCLUSIONS: Short-term, high-dose combination treatment with NAC and vitamin C in patients with HIV infection and advanced immunodeficiency lead to immunological and virological effects that might be of therapeutic value.

Decreased vitamin A levels in common variable immunodeficiency: vitamin A supplementation in vivo enhances immunoglobulin production and downregulates inflammatory responses.

BACKGROUND: Vitamin A has a broad range of immunological effects, and vitamin A deficiency is associated with recurrent infections. Common variable immunodeficiency (CVI) is a group of B-cell deficiency syndromes with impaired antibody production and recurrent bacterial infections as the major manifestations, but the immunological dysfunctions may also include T cells and macrophages. In the present study we examined the possible role of vitamin A deficiency in CVI. PATIENTS AND METHODS: We analysed plasma vitamin A levels in 20 CVI patients and 16 controls, and examined the relationships between vitamin A and clinical, immunological and metabolic parameters in CVI. In the six CVI patients with the lowest vitamin A levels we also studied the effect of vitamin A supplementation in vivo on several immunological functions in these patients. RESULTS: (i) The majority of CVI patients had decreased vitamin A levels compared with healthy controls, as found in both cross-sectional and longitudinal testing. (ii) Low vitamin A levels were associated with the occurrence of chronic bacterial infections and splenomegaly as well as high neopterin levels. Decreased levels of carrier protein and malabsorption were not observed. (iii) Vitamin A supplementation in patients with low vitamin A levels resulted in increased interleukin-10 (IL-10) and decreased tumour necrosis factor-alpha (TNFalpha) levels, as found in both plasma and monocyte supernatants, possibly favouring anti-inflammatory net effects. (iv) Vitamin A supplementation in vivo also enhanced anti-CD40-stimulated IgG production, serum IgA levels and phytohaemagglutinin (PHA)-stimulated peripheral blood mononuclear cell (PBMC) proliferation. CONCLUSION: A considerable subgroup of CVI patients appears to be characterized by low vitamin A levels. Our findings support a possible role for vitamin A supplementation in CVI, perhaps resulting in enhanced immunoglobulin synthesis and downregulated inflammatory responses.

Correlations of genotype and phenotype in hypophosphatasia.

Hypophosphatasia, a rare inherited disorder characterized by defective bone mineralization, is highly variable in its clinical expression. The disease is due to various mutations in the tissue-non-specific alkaline phosphatase ( TNSALP ) gene. We report here the use of clinical data, site-directed mutagenesis and computer-assisted modelling to propose a classification of 32 TNSALP gene mutations found in 23 European patients, 17 affected with lethal hypophosphatasia and six with non-lethal hypophosphatasia. Transfection studies of the missense mutations found in non-lethal hypophosphatasia showed that six of them allowed significant residual in vitro enzymatic activity, suggesting that these mutations corresponded to moderate alleles. Each of the six patients with non-lethal hypophosphatasia carried at least one of these alleles. The three-dimensional model study showed that moderate mutations were not found in the active site, and that most of the severe missense mutations were localized in crucial domains such as the active site, the vicinity of the active site and homodimer interface. Some mutations appeared to be organized in clusters on the surface of the molecule that may represent possible candidates for regions interacting with the C-terminal end involved in glycosylphosphatidylinositol (GPI) attachment or with other dimers to form tetramers. Finally, our results show a good correlation between clinical forms of the disease, mutagenesis experiments and the three-dimensional structure study, and allowed us to clearly distinguish moderate alleles from severe alleles. They also confirm that the extremely high phenotypic heterogeneity observed in patients with hypophosphatasia was due mainly to variable residual enzymatic activities allowed by missense mutations found in the human TNSALP gene.