In Vitro and in Vivo Assays of 3,5-Disubstituted-Tetrahydro-2H-1,3,5-Thiadiazin-2-Thione Derivatives against Trypanosoma cruzi

Cytotoxicity assays of 24 new 3,5-disubstituted-tetrahydro-2H-1,3,5-thiadiazin-2-thione derivatives were performed. The 17 compounds with higher anti-epimastigote activity and lower cytotoxicity were, thereafter, screened against amastigote of Trypanosoma cruzi. Out of these 17 derivatives S-2d was selected to be assayed in vivo, because of its remarkable trypanocidal properties. To determine toxicity against J774 macrophages, a method based on quantification of cell damage, after 24 h, was used. Cell respiration, an indicator of cell viability, was assessed by the reduction of MTT [3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide] to formazan. Anti-amastigote activity was estimated after 48 h by microscopic counts of May Grünwald-Giemsa-stained monolayers. Nifurtimox and benznidazole were used as reference drugs. For the in vivo experiences, mice were infected with 10(4) blood trypomastigotes and then treated during 15 days with S-2d or nifurtimox by oral route. All of the compounds were highly toxic at 100 µg/ml for macrophages and a few of them maintained this cytotoxicity even at 10 µg/ml. Of the derivatives assayed against amastigotes 3k and S-2d showed an interesting activity, that was held even at 1µg/ml. It is demonstrated that the high anti-epimastigote activity previously reported is mainly due to the non-specific toxicity of these compounds. In vivo assays assessed a reduction of parasitemia after administration of S-2d to infected mice.

In Vitro and in Vivo Anti-Trypanosoma cruzi Activity of a Novel Nitro-derivative

Nitroarylidenemalononitriles and their cyanoacetamide derivatives with remarkable anti-epimastigote properties, were synthesized attempting to obtain new 3,5-diamino-4-(5'-nitroarylidene)-4H-thiadiazine 1,1-dioxide derivatives, which in previous reports had shown anti-Trypanosoma cruzi activity. Tests to evaluate the cytotoxicity of compounds were performed on J774 macrophages. 5-nitro-2-thienyl-malononitrile (5NO2TM), was the only product which maintained a high anti-epimastigote activity at concentrations in which it was no longer cytotoxic, thus it was assayed against intracellular amastigotes. Its anti-amastigote activity was similar to that of nifurtimox. Afterwards in vivo toxicity and anti-chagasic activity were determined. A reduction in parasitemia was observed.

Influence of the in vivo deregulation of the expression of ntrk3 (trkc) on cortical development of a murine model of panic/anxiety disorder

Alteracions durant el desenvolupament cerebral produirien canvis en la connectivitat neuronal i la bioquímica cel•lular que podrien resultar en una disfunció cognitiva i/o emocional, desembocant a trastorns psiquiàtrics. Les neurotrofines intervenen en els processos del neurodesenvolupament i en la funcionalitat del cervell adult i, conseqüentment, serien bons candidats com a factors de predisposició en diverses malalties mentals. S’ha suggerit la implicació del receptor de la neurotrofina 3, TrkC, en el trastorn de pànic. Nosaltres proposem que la sobreexpressió del gen NTRK3 (TrkC) és un mediador comú dels desencadenants genètics i ambientals d’aquest trastorn. Concretament, la seva desregulació podria produir canvis estructurals i funcionals a l’escorça cerebral dels pacients pel seu paper durant l’establiment dels circuïts corticals i la neuroplasticitat a l’adult, probablement esdevenint elements de predisposició a patir atacs de pànic. Els objectius principals d’aquest treball han estat: 1/determinar la contribució específica del gen NTRK3 a les alteracions de l’escorça cerebral observades en pacients, utilitzant un model murí modificat genèticament (TgNTRK3), i 2/analitzar l’impacte específic de la sobreexpressió de NTRK3 sobre la corticogènesi durant estadis embrionaris o postnatals estudiant la neurogènesi i la neuritogènesi. Els resultats indiquen que la sobreexpressió de NTRK3 als ratolins produeix una reducció del gruix de l’escorça frontal, recapitulant la hipofrontalitat dels pacients, que comportaria una menor inhibició dels nuclis subcorticals del sistema límbic com l’amígdala, i alteracions citoarquitectòniques a l’escorça prefrontal medial que recolzen la hipòtesi del seu mal funcionament. Tanmateix, els ratolins TgNTRK3 presenten canvis estructurals a l’escorça somatosensorial, suggerint que el processament de la informació sensorial podria estar alterat, el que encara no s’ha explorat en pacients. La sobreexpressió de NTRK3 també afecta la neuritogènesi en cultius primaris corticals i modifica la resposta de les neurones a l’estimulació amb neurotrofines. Per tant, el fenotip cortical adult dels TgNTRK3 podria dependre d’alteracions durant la corticogènesi.

Targetting of the ventro-intermediate nucleus using ultra-high field (7T) MRI for gamma knife surgery purposes: A pilot in vivo study on healthy subjects.

INTRODUCTION: Gamma Knife surgery (GKS) is a non-invasive neurosurgical stereotactic procedure, increasingly used as an alternative to open functional procedures. This includes targeting of the ventro-intermediate nucleus of the thalamus (e.g. Vim) for tremor. We currently perform an indirect targeting, as the Vim is not visible on current 3Tesla MRI acquisitions. Our objective was to enhance anatomic imaging (aiming at refining the precision of anatomic target selection by direct visualisation) in patients treated for tremor with Vim GKS, by using high field 7T MRI. MATERIALS AND METHODSH: Five young healthy subjects were scanned on 3 (T1-w and diffusion tensor imaging) and 7T (high-resolution susceptibility weighted images (SWI)) MRI in Lausanne. All images were further integrated for the first time into the Gamma Plan Software(®) (Elekta Instruments, AB, Sweden) and co-registered (with T1 was a reference). A simulation of targeting of the Vim was done using various methods on the 3T images. Furthermore, a correlation with the position of the found target with the 7T SWI was performed. The atlas of Morel et al. (Zurich, CH) was used to confirm the findings on a detailed analysis inside/outside the Gamma Plan. RESULTS: The use of SWI provided us with a superior resolution and an improved image contrast within the basal ganglia. This allowed visualization and direct delineation of some subgroups of thalamic nuclei in vivo, including the Vim. The position of the target, as assessed on 3T, perfectly matched with the supposed one of the Vim on the SWI. Furthermore, a 3-dimensional model of the Vim-target area was created on the basis of the obtained images. CONCLUSION: This is the first report of the integration of SWI high field MRI into the LGP, aiming at the improvement of targeting validation of the Vim in tremor. The anatomical correlation between the direct visualization on 7T and the current targeting methods on 3T (e.g. quadrilatere of Guyot, histological atlases) seems to show a very good anatomical matching. Further studies are needed to validate this technique, both by improving the accuracy of the targeting of the Vim (potentially also other thalamic nuclei) and to perform clinical assessment.

Differential distribution of thyroid hormone receptor isoform in rat dorsal root ganglia and sciatic nerve in vivo and in vitro.

Using autoradiographic techniques carried out under precise conditions we previously demonstrated that both sensory neurons and peripheral glial cells in dorsal root ganglia (DRG) or sciatic nerve, possess specific [125I]-labeled T3 binding sites. Thyroid hormone receptors (TR) include several isoforms (TR alpha(1), TR alpha(2), TR beta(1), TR beta(2...)) The present study demonstrates that while sensory neurons and peripheral glial cells both possess functional TR, they express a differential expression of TR isoforms. Using a panel of antisera to specific for the TR alpha-common (alpha(1) and alpha(2)), TR alpha-1 or TR beta-1 isoforms, we detected TRs isoform localization at the cellular level during DRG and sciatic nerve development and regeneration. Immunohistochemical analysis revealed that during embryonic life, sensory neurons express TR alpha-common and TR beta-1 rather than TR alpha-1. The number of TR alpha-common and TR beta-1 positive neurons as well as the intensity of labeling increased during the first two postnatal weeks and remained more or less stable in adult life. TR alpha-1 immunoreactivity, which was undetectable in embryonic sensory neurons, became discreetly visible in neurons after birth. In developing DRG and sciatic nerves, Schwann cells exhibited TR alpha-common and TR alpha-1 rather than TR beta-1 immunolabeling. The appearance of TR alpha-common and alpha-1 isoform immunoreactivity in the sciatic nerve was restricted to a short period ranging from E17 up to two postnatal weeks. By comparing TR alpha-common and TR alpha-1 immunostaining we can deduce that Schwann cells primarily express TR alpha-1. Afterwards, in adult rat sciatic nerve TR alpha isoforms was no more detected. However transection of sciatic nerve caused a reexpression of TR alpha isoforms in degenerating nerve. The prevalence of TR alpha in Schwann cells in vivo was correlated with in vitro results. The differential expression of TR alpha and beta by sensory neurons and Schwann cells indicates that the feedback regulation of circulating thyroid hormone could occur by binding to either the alpha or beta TR isoforms. Moreover, the presence of multiple receptor isoforms in developing sensory neurons suggests that thyroid hormone uses multiple signaling pathways to regulate DRG and sciatic nerve development.

In vitro evaluation of the activity of aromatic nitrocompounds against Trypanosoma cruzi

Fourteen compounds were evaluated for their activity against Trypanosoma cruzi blood stream forms at the concentration of 500 µg/ml. Six compounds were active and re-tested at lower concentrations.

Diffusion spectrum imaging shows the structural basis of functional cerebellar circuits in the human cerebellum in vivo.

BACKGROUND: The cerebellum is a complex structure that can be affected by several congenital and acquired diseases leading to alteration of its function and neuronal circuits. Identifying the structural bases of cerebellar neuronal networks in humans in vivo may provide biomarkers for diagnosis and management of cerebellar diseases. OBJECTIVES: To define the anatomy of intrinsic and extrinsic cerebellar circuits using high-angular resolution diffusion spectrum imaging (DSI). METHODS: We acquired high-resolution structural MRI and DSI of the cerebellum in four healthy female subjects at 3T. DSI tractography based on a streamline algorithm was performed to identify the circuits connecting the cerebellar cortex with the deep cerebellar nuclei, selected brainstem nuclei, and the thalamus. RESULTS: Using in-vivo DSI in humans we were able to demonstrate the structure of the following cerebellar neuronal circuits: (1) connections of the inferior olivary nucleus with the cerebellar cortex, and with the deep cerebellar nuclei (2) connections between the cerebellar cortex and the deep cerebellar nuclei, (3) connections of the deep cerebellar nuclei conveyed in the superior (SCP), middle (MCP) and inferior (ICP) cerebellar peduncles, (4) complex intersections of fibers in the SCP, MCP and ICP, and (5) connections between the deep cerebellar nuclei and the red nucleus and the thalamus. CONCLUSION: For the first time, we show that DSI tractography in humans in vivo is capable of revealing the structural bases of complex cerebellar networks. DSI thus appears to be a promising imaging method for characterizing anatomical disruptions that occur in cerebellar diseases, and for monitoring response to therapeutic interventions.

Quantification of in vivo short echo-time proton magnetic resonance spectra at 14.1 T using two different approaches of modelling the macromolecule spectrum

Reliable quantification of the macromolecule signals in short echo-time H-1 MRS spectra is particularly important at high magnetic fields for an accurate quantification of metabolite concentrations (the neurochemical profile) due to effectively increased spectral resolution of the macromolecule components. The purpose of the present study was to assess two approaches of quantification, which take the contribution of macromolecules into account in the quantification step. H-1 spectra were acquired on a 14.1 T/26 cm horizontal scanner on five rats using the ultra-short echo-time SPECIAL (spin echo full intensity acquired localization) spectroscopy sequence. Metabolite concentrations were estimated using LCModel, combined with a simulated basis set of metabolites using published spectral parameters and either the spectrum of macromolecules measured in vivo, using an inversion recovery technique, or baseline simulated by the built-in spline function. The fitted spline function resulted in a smooth approximation of the in vivo macromolecules, but in accordance with previous studies using Subtract-QUEST could not reproduce completely all features of the in vivo spectrum of macromolecules at 14.1 T. As a consequence, the measured macromolecular 'baseline' led to a more accurate and reliable quantification at higher field strengths.

Efavirenz in an obese HIV-infected patient - a report and an in vitro-in vivo extrapolation model indicate risk of underdosing.

Here, we report suboptimal efavirenz exposure in an obese patient treated with the standard 600 mg dose. Tripling the dose allowed attainment of therapeutic efavirenz concentrations. We developed an in vitro-in vivo extrapolation model to quantify dose requirements in obese individuals. Obesity represents a risk factor for antiretroviral therapy underdosing.

In vivo binding of the Cry11Bb toxin of Bacillus thuringiensis subsp. medellin to the midgut of mosquito larvae (Diptera: Culicidae)

Bacillus thuringiensis subsp. medellin produces numerous proteins among which 94 kDa known as Cry11Bb, has mosquitocidal activity. The mode of action of the Cry11 proteins has been described as similar to those of the Cry1 toxins, nevertheless, the mechanism of action is still not clear. In this study we investigated the in vivo binding of the Cry11Bb toxin to the midgut of the insect species Anopheles albimanus, Aedes aegypti, and Culex quinquefasciatus by immunohistochemical analysis. Spodoptera frugiperda was included as negative control. The Cry11Bb protein was detected on the apical microvilli of the midgut epithelial cells, mostly on the posterior midgut and gastric caeca of the three mosquito species. Additionally, the toxin was detected in the Malpighian tubules of An. albimanus, Ae. aegypti, Cx. quinquefasciatus, and in the basal membrane of the epithelial cells of Ae. aegypti midgut. No toxin accumulation was observed in the peritrophic membrane of any of the mosquito species studied. These results confirm that the primary site of action of the Cry11 toxins is the apical membrane of the midgut epithelial cells of mosquito larvae.

Direct in vivo measurement of glycine and the neurochemical profile in the rat medulla oblongata.

The medulla oblongata (MO) contains a high density of glycinergic synapses and a particularly high concentration of glycine. The aims of this study were to measure directly in vivo the neurochemical profile, including glycine, in MO using a spin-echo-based (1)H MRS sequence at TE?=?2.8 ms and to compare it with three other brain regions (cortex, striatum and hippocampus) in the rat. Glycine was quantified in MO at TE?=?2.8 ms with a Cramér-Rao lower bound (CRLB) of approximately 5%. As a result of the relatively low level of glycine in the other three regions, the measurement of glycine was performed at TE?=?20 ms, which provides a favorable J-modulation of overlapping myo-inositol resonance. The other 14 metabolites composing the neurochemical profile were quantified in vivo in MO with CRLBs below 25%. Absolute concentrations of metabolites in MO, such as glutamate, glutamine, ?-aminobutyrate, taurine and glycine, were in the range of previous in vitro quantifications in tissue extracts. Compared with the other regions, MO had a three-fold higher glycine concentration, and was characterised by reduced (p?<?0.001) concentrations of glutamate (-50?±?4%), glutamine (-54?±?3%) and taurine (-78?±?3%). This study suggests that the functional specialisation of distinct brain regions is reflected in the neurochemical profile.

Neurochemical profile of the developing mouse cortex determined by in vivo 1H NMR spectroscopy at 14.1 T and the effect of recurrent anaesthesia.

The neurochemical profile of the cortex develops in a region and time specific manner, which can be distorted by psychiatric and other neurological pathologies. Pre-clinical studies often involve experimental mouse models. In this study, we determined the neurochemical profile of C57BL/6 mice in a longitudinal study design to provide a reference frame for the normal developing mouse cortex. Using in vivo proton NMR spectroscopy at 14 T, we measured the concentrations of 18 metabolites in the anterior and posterior cortex on postnatal days (P) 10, 20, 30, 60 and 90. Cortical development was marked by alterations of highly concentrated metabolites, such as N-acetylaspartate, glutamate, taurine and creatine. Regional specificity was represented by early variations in the concentration of glutamine, aspartate and choline. In adult animals, regional concentration differences were found for N-acetylaspartate, creatine and myo-inositol. In this study, animals were exposed to recurrent isoflurane anaesthesia. Additional experiments showed that the latter was devoid of major effects on behaviour or cortical neurochemical profile. In conclusion, the high sensitivity and reproducibility of the measurements achieved at 14 T allowed us to identify developmental variations of cortical areas within the mouse cortex.

Bacterial flagellin elicits innate immune defenses and cardiac dysfunction in vivo

Résumé Les agents pathogènes responsables d'infection entraînent chez l'hôte deux types de réponses immunes, la première, non spécifique, dite immunité innée, la seconde, spécifique à l'agent concerné, dite immunité adaptative. L'immunité innée, qui représente la première ligne de défense contre les pathogènes, est liée à la reconnaissance par les cellules de l'hôte de structures moléculaires propres aux micro-organismes (« Pathogen-Associated Molecular Patterns », PAMPs), grâce à des récepteurs membranaires et cytoplasmiques (« Pattern Recognition Receptors », PRRs) identifiant de manière spécifique ces motifs moléculaires. Les récepteurs membranaires impliqués dans ce processus sont dénommés toll-like récepteurs, ou TLRS. Lorsqu'ils sont activés par leur ligand spécifique, ces récepteurs activent des voies de signalisation intracellulaires initiant la réponse inflammatoire non spécifique et visant à éradiquer l'agent pathogène. Les deux voies de signalisation impliquées dans ce processus sont la voie des « Mitogen-Activated Protein Kinases » (MAPKs) et celle du « Nuclear Factor kappaB » (NF-κB), dont l'activation entraîne in fine l'expression de protéines de l'inflammation dénommées cytokines, ainsi que certaines enzymes produisant divers autres médiateurs inflammatoires. Dans certaines situations, cette réponse immune peut être amplifiée de manière inadéquate, entraînant chez l'hôte une réaction inflammatoire systémique exagérée, appelée sepsis. Le sepsis peut se compliquer de dysfonctions d'organes multiples (sepsis sévère), et dans sa forme la plus grave, d'un collapsus cardiovasculaire, définissant le choc septique. La défaillance circulatoire du choc septique touche les vaisseaux sanguins d'une part, le coeur d'autre part, réalisant un tableau de «dysfonction cardiaque septique », dont on connaît mal les mécanismes pathogéniques. Les bactéries à Gram négatif peuvent déclencher de tels phénomènes, notamment en libérant de l'endotoxine, qui active les voies de l'immunité innée par son interaction avec un toll récepteur, le TLR4. Outre l'endotoxine, la plupart des bactéries à Gram négatif relâchent également dans leur environnement une protéine, la flagelline, qui est le constituant majeur du flagelle bactérien, organelle assurant la mobilité de ces micro-organismes. Des données récentes ont indiqué que la flagelline active, dans certaines cellules, les voies de l'immunité innée en se liant au récepteur TLRS. On ne connaît toutefois pas les conséquences de l'interaction flagelline-TLRS sur le développement de l'inflammation et des dysfonctions d'organes au cours du sepsis. Nous avons par conséquent élaboré le présent travail en formulant l'hypothèse que la flagelline pourrait déclencher une telle inflammation et représenter ainsi un médiateur potentiel de la dysfonction d'organes au cours du sepsis à Gram négatif, en nous intéressant plus particulièrement àl'inflammation et à la dysfonction cardiaque. Dans la première partie de ce travail, nous avons étudié les effets de la flagelline sur l'activation du NF-κB et des MAPKs, et sur l'expression de cytokines inflammatoires au niveau du myocarde in vitro (cardiomyocytes en culture) et in vivo (injection de flagelline recombinante à des souris). Nous avons observé tout d'abord que le récepteur TLRS est fortement exprimé au niveau du myocarde. Nous avons ensuite démontré que la flagelline active la voie du NF-κB et des MAP kinases (p38 et JNK), stimule la production de cytokines et de chemokines inflammatoires in vitro et in vivo, et entraîne l'activation de polynucléaires neutrophiles dans le tissu cardiaque in vivo. Finalement, au plan fonctionnel, nous avons pu montrer que la flagelline entraîne une dilatation et une réduction aiguë de la contractilité du ventricule gauche chez la souris, reproduisant les caractéristiques de la dysfonction cardiaque septique. Dans la deuxième partie, nous avons déterminé la distribution du récepteur TLRS dans les autres organes majeurs de la souris (poumon, foie, intestin et rein}, et avons caractérisé dans ces organes l'effet de la flagelline sur l'activation du NF-κB et des MAPKs, l'expression de cytokines, et l'induction de l'apoptose. Nous avons démontré que le TLRS est exprimé de façon constitutive dans ces organes, et que l'injection de flagelline y déclenche les cascades de l'immunité innée et de processus apoptotiques. Finalement, nous avons également déterminé que la flagelline entraîne une augmentation significative de multiples cytokines dans le plasma une à six heures après son injection. En résumé, nos données démontrent que la flagelline bactérienne (a) entraîne une inflammation et une dysfonction importantes du myocarde et (b) active de manière très significative les mécanismes d'immunité innée dans les principaux organes et entraîne une réponse inflammatoire systémique. Par conséquent, la flagelline peut représenter un médiateur puissant de l'inflammation et de la dysfonction d'organes, notamment du coeur, au cours du choc septique déclenché par les bactéries à Gram négatif. Summary Pathogenic microorganisms trigger two kinds of immune responses in the host. The first one is immediate and non-specific and is termed innate immunity, whereas the second one, specifically targeted at the invading agent, is termed adaptative immunity. Innate immunity, which represents the first line of defense against invading pathogens, confers the host the ability to recognize molecular structures common to many microbial pathogens, ("Pathogen-Associated Molecular Patterns", PAMPs), through cytosolic or membrane-associated receptors ("Pattern Recognition Receptors", PRRs), the latter being represented by a family of receptors termed "toll-like receptors or TLRs". Once activated by the binding of their specific ligand, these receptors activate intracellular signaling pathways, which initiate the non-specific inflammatory response aimed at eradicating the pathogens. The two pathways implicated in this process are the mitogen-activated protein kinases (MAPK) and the nuclear factor kappa B (NF-κB) signaling pathways, whose activation elicit in fine the expression of inflammatory proteins termed cytokines, as well as various enzymes producing a wealth of additional inflammatory mediators. In some circumstances, the innate immune response can become amplified and dysregulated, triggering an overwhelming systemic inflammatory response in the host, identified as sepsis. Sepsis can be associated with multiple organ dysfunction (severe sepsis), and in its most severe form, with cardiovascular collapse, defming septic shock. The cardiovascular failure associated with septic shock affects blood vessels as well as the heart, resulting in a particular form of acute heart failure termed "septic cardiac dysfunction ", whose pathogenic mechanisms remain partly undefined. Gram-negative bacteria can initiate such phenomena, notably by releasing lipopolysaccharide (LPS), which activates innate immune signaling by interacting with its specific toll receptor, the TLR4. Besides LPS, most Gram-negative bacteria also release flagellin into their environment, which is the main structural protein of the bacterial flagellum, an appendage extending from the outer bacterial membrane, responsible for the motility of the microorganism. Recent data indicated that flagellin activate immune responses upon binding to its receptor, TLRS, in various cell types. However, the role of flagellin/TLRS interaction in the development of inflammation and organ dysfunction during sepsis is not known. Therefore, we designed the present work to address the hypothesis that flagellin might trigger such inflammatory responses and thus represent a potential mediator of organ dysfunction during Gram-negative sepsis, with a particular emphasis on cardiac inflammation and contractile dysfunction. In the first part of this work, we investigated the effects of flagellin on NF-κB and MAPK activation and the generation of pro-inflammatory mediators within the heart in vitro (cultured cardiomyocytes) and in vivo (injection of recombinant flagellin into mice). We first observed that TLRS protein is strongly expressed by the myocardium. We then demonstrated that flagellin activates NF-κB and MAP kinases (p38 and JNK), upregulates the transcription of pro-inflammatory cytokines and chemokines in vitro and in vivo, and stimulates the activation of polymorphonuclear neutrophils within the heart in vivo. Finally, we demonstrated that flagellin triggers acute cardiac dilation, and a significant reduction of left ventricular contractility, mimicking characteristics of clinical septic cardiac dysfunction. In the second part, we determined the TLRS distribution in other mice major organs (lung, liver, gut and kidney) and we characterized in these organs the effects of flagellin on NF-κB and MAPK activation, on the expression of pro-inflammatory çytokines, and on the induction of apoptosis. We demonstrated that TLRS protein is constitutively expressed and that flagellin activates prototypical innate immune responses and pro-apoptotic pathways in all these organs. Finally, we also observed that flagellin induces a significant increase of multiple cytokines in the plasma from 1 to 6 hours after its intravenous administration. Altogether, these data provide evidence that bacterial flagellin (a) triggers an important inflammatory response and an acute dysfunction of the myocardium, and (b) significantly activates the mechanisms of innate immunity in most major organs and elicits a systemic inflammatory response. In consequence, flagellin may represent a potent mediator of inflammation and multiple organ failure, notably cardiac dysfunction, during Gram-negative septic shock.

Leishmania (Viannia) lainsoni: occurrence of intracellular promastigote forms in vivo and in vitro

Experimental chronic (45-day-old) skin lesion in hamster hind foot induced by Leishmania (Viannia) lainsoni infection showed the presence of promastigote forms in the tissue, inside parasitophorous vacuoles, as assessed by transmission electron microscopy. Experimental in vitro interaction (24 and 48 h) between Leishmania (V.)lainsoni and J774-G8 macrophage cells also demonstrated the same profile. This morphological aspect is unusual, since in this parasite genus only amastigote forms have been described as the resistant and obligate intracellular forms.