Persistent inhibition of FLIP(L) expression by lentiviral small hairpin RNA delivery restores death-receptor-induced apoptosis in neuroblastoma cells.

Neuroblastoma represents the most common and deadly solid tumour of childhood, which disparate biological and clinical behaviour can be explained by differential regulation of apoptosis. To understand mechanisms underlying death resistance in neuroblastoma cells, we developed small hairpin of RNA produced by lentiviral vectors as tools to selectively interfere with FLIP(L), a major negative regulator of death receptor-induced apoptosis. Such tools revealed highly efficient in interfering with FLIP(L) expression and function as they almost completely repressed endogenous and/or exogenously overexpressed FLIP(L) protein and fully reversed FLIP(L)-mediated TRAIL resistance. Moreover, interference with endogenous FLIP(L) and FLIP(S) significantly restored FasL sensitivity in SH-EP neuroblastoma cell line. These results reveal the ability of lentivirus-mediated shRNAs to specifically and persistently interfere with FLIP expression and support involvement of FLIP in the regulation of death receptor-mediated apoptosis in neuroblastoma cells. Combining such tools with other therapeutic modalities may improve treatment of resistant tumours such as neuroblastoma.

Persistent Infections by Leishmania (Viannia) braziliensis

Here we review the phenomenon of persistency in Leishmania (Viannia) braziliensis infections. In other Leishmania species where appropriate animal models exist, considerable advances in the understanding of basic immunologic mechanisms of persistency have been made; for a review see Aebisher (1994). On the contrary, the evidences of persistence in infections with L. braziliensis rest on studies of human clinical cases many of which we summarized and discussed in this work

A new murine model of persistent lung eosinophilic inflammation

We summarize here the main characteristics of a novel model of pulmonary hypersensitivity. Mice were immunized with a subcutaneous implant of a fragment of heat solidified chicken egg white and 14 days later challenged with ovalbumin given either by aerosol or by intratracheal instillation. This procedure induces a persistent eosinophilic lung inflammation, a marked bone marrow eosinophilia, and Th2-type isotypic profile with histopathological findings that resemble human asthma. Further, this model is simple to perform, reproducible in different strains of mice, does not require adjuvants nor multiple boosters. Based on these characteristics we propose it as a suitable murine model of allergic eosinophilic lung inflammation.

Neuro-developmental follow-up of neonates treated with magnesium sulfate for persistent pulmonary hypertension

Rapport de synthèse : DEVENIR NEURO-DEVELOPPEMENTAL DE NOUVEAU-NES TRAITES PAR DU SULFATE DE MAGNESIUM POUR UNE HYPERTENSION PULMONAIRE PERSISTANTE L'hypertension pulmonaire persistante du nouveau-né (HTPP) est un trouble de l'adaptation post-natale de la circulation pulmonaire caractérisé par une défaillance de la diminution normale des résistances vasculaires pulmonaires, accompagné d'un shunt droite-gauche, résultant en une hypoxémie profonde. C'est une pathologie sévère nécessitant des soins intensifs avec un risque augmenté de handicaps neurologiques chez les survivants. Le traitement de l'HTPP du nouveau-né inclut une ventilation mécanique ainsi que différents agents pharmacologiques pour dilater les vaisseaux pulmonaires, dont le sulfate de magnésium (MgSO4) à hautes doses par voie intraveineuse et le monoxyde d'azote par voie inhalée (iN0). Le MgSO4 est une alternative thérapeutique de l'HTPP du nouveau-né avec peu d'effets secondaires et une mortalité basse. Il a aussi été démontré que le MgSO4 est un traitement de l'HTPP du nouveau-né autant efficace que le iN0 et moins coüteux. Des études sur le suivi neuro-développemental de nouveau-nés avec HTPP traités selon différentes méthodes ont été publiées reportant des taux élevés de handicaps majeurs et mineurs. Plus récemment, des études de suivi après traitement par iN0 ont montré des taux plus bas qu'avec des traitements antérieurs. Le devenir neuro-développemental àlong terme d'enfants traités avec du MgSO4 n'a pas été documenté. Le but de cette étude est de décrire le développement des enfants qui ont présenté une HTPP traitée seulement avec du MgS04, de reporter l'incidence de handicaps majeurs et mineurs, et de les comparer à un groupe contrôle d'enfants sains du même âge ainsi qu'aux données de la littérature. La population consiste en 33 nouveau-nés traités pour une HTPP avec seulement du MgSO4 (groupe étude) et 32 nouveau-nés à terme sains (groupe contrôle). Un suivi neurodéveloppemental standardisé et approfondi a été effectué aux âges clés de 18 mois et 5 ans. Les taux de handicaps majeurs à 18 mois et 5 ans dans le groupe étude étaient de 6% et 11,4% respectivement, et de 0% aux deux âges dans le groupe contrôle. Les taux de handicaps mineurs aux mêmes âges étaient de 3% et 26,9% pour le groupe étude, et de 0% et 26,1% pour le groupe contrôle. Les quotients développementaux moyens à 18 mois étaient de 106,6 (DS 1,6) dans le groupe étude et de 118,3 (DS 1,0) dans le groupe contrôle (P < 0,001). L'index général intellectuel en âge préscolaire était de 112.6 (DS 3.7), respectivement de 119.3 (DS 3.1 ), sans différence significative entre les deux groupes. A 18 mois, les taux de handicaps majeurs et mineurs dans les groupes études et contrôle étaient de 6% et 3%. Dans la littérature, des taux entre 0% et 33% ont été décrits. A cet âge, il y avait une différence significative pour tous les scores du test de Griffiths, mëme en tenant compte du status socio-économique de la famille. Ceci suggère un léger retard du développement global et non une altération spécifique. Ces différences n'étaient plus significatives en âge préscolaire, suggérant un rattrapage développemental. Le taux de handicaps majeurs en âge préscolaire pour le groupe étude était de 11.5%, sans aucune infirmité motrice cérébrale. Ces résultats correspondent à ceux d'études de suivi après d'autres traitements jusqu'à l'âge de 24 mois avec des taux variant de 0% à 15%. Le taux de handicaps mineurs était de 26.9% dans le groupe étude et de 26.1% dans le groupe contrôle, sans différence significative entre les deux groupes. L'incidence de handicaps mineurs dans le groupe étude était plutôt élevée en comparaison aux données de la littérature (6 à 22% à 6 ans). Une explication possible est que nous avons considéré des problèmes de langage et de comportement comme handicaps mineurs. Ceci suggère une différence méthodologique et non une plus mauvaise issue dans nos deux groupes. Les évaluations cognitives des enfants des deux groupes se trouvaient dans la norme, ce qui est aussi le cas dans la littérature. En conclusion, cette étude longitudinale non randomisée d'enfants traités avec du MgSO4 seul pour une HTPP sévère ne montre pas de conséquences sur le devenir neuro-développemental à long terme. Cette étude le démontre pour la première fois. Malgré le fait que iN0 soit le traitement actuellement recommandé pour l'HTPP du nopuveau-né, le MgSO4 reste largement utilisé, en particulier dans des pays en voie de développement. L'absence de complications neuro-développementales majeures à long terme permet de considérer l'administration du MgSO4 pour le traitement de l'HTPP du nouveau-né en cas de non réponse ou d'inaccessibilité au iNO.

High bi-atrial organization in patients with long-standing persistent atrial fibrillation terminated within the left atrium

Sustained atrial fibrillation (AF) is maintained by sites displaying high dominant frequency (DF). In patients (pts) with long-standing persistent AF (LS-pAF), their spatial distribution and the presence of a left-to-right atrial DF gradient remain poorly known. We hypothesized that the pre-ablation bi-atrial frequency characteristics of LS-pAF pts terminated within the left atrium (LT) are different from that of non terminated (NT) ones. Methods: 23 consecutive pts (59±7y, LS-pAF duration 19±12m) underwent stepwise catheter ablation (step-CA) consisting in pulmonary veins isolation, left atrial (LA) defragmentation, and right atrial (RA) ablations for non terminated AF. A quadripolar catheter (CAT) was placed into the RA appendage (RAA), a decapolar CAT into the coronary sinus (CS) and a duodecapolar CAT into the LA divided into 8 segments. For each segment, 20-sec of bipolar recording was acquired. The DF was defined as the largest peak in the power spectrum (3-15 Hz). The inter-atrial DF gradient was defined as the DF difference between LA and RA appendages. Results: LS-pAF was terminated in 83% (19/23) of the pts: 17 LT, 2 during RA ablation and 4 NT. The figure shows that before ablation bi-atrial DF values of LT pts are significantly lower than that of NT pts for each LA segment as well as for the RAA (p < 0.05). No significant LA-to-RA DF gradient was observed both for LT (0.3±0.5 Hz, p=ns) and NT (0.5±0.03 Hz, p=ns) pts. No significant difference in DF values was observed between LA segments. Conclusions: The lower DF of LT pts is suggestive of a higher organization within both atria compared to NT pts. Our findings suggest that low bi-atrial DF values, but not inter-atrial DF gradient, might be of interest for selecting LS-pAF candidates for sinus rhythm restoration by step-CA.

Persistent human parvovirus B19 infection in children under maintenance chemotherapy for acute lymphocytic leukemia.

OBJECTIVE: To report on B19 infection management and chemotherapy schedule consequences in five children treated for acute lymphocytic leukemia (ALL). PATIENTS AND METHODS: Between May 2001 and February 2002, five patients between 4 and 12 years of age, receiving maintenance chemotherapy for ALL, presented with symptoms suggesting B19 infection (pallor, fatigue, petechiae and pancytopenia in four patients; generalized rash in two patients; acute hepatitis in one patient). Qualitative polymerase chain reaction (PCR) on peripheral blood was used for diagnosis and follow-up of infection; quantitative PCR was used for viral load measurement. Intravenous nonspecific high-dose immunoglobulin therapy was administered until PCR was negative. RESULTS: Qualitative B19 DNA was found in the peripheral blood of all patients, confirming the infection. Viral load at diagnosis ranged from 10 to 10 particles/mL blood. B19 DNA was detectable in four patients at 45, 21, 40, and 44 weeks, respectively. Chemotherapy was delayed in all patients. No clear benefit of intravenous immunoglobulin was noted. CONCLUSIONS: Infection with B19 is rarely reported in patients with ALL, but it should be suspected when unexplained pancytopenia occurs during chemotherapy. Persistent B19 infection remains a challenge in the management of patients receiving maintenance chemotherapy for ALL, as no specific therapy such as a specific immunoglobulin or vaccine exists. The role of viral load measurement needs to be established in terms of its use in follow-up and evaluation of the therapeutic response.

Coronary sinus atresia with persistent left superior vena cava: unusual clinical presentation and endovascular management.

Atresia of the coronary sinus (ACS) is a rare congenital anomaly. When associated with persistent left superior vena cava (PLSVC), this defect could have no significant hemodynamic effect, and the patient might remain asymptomatic. However, vascular interventions might induce changes or complications that could show the anomaly. Appropriate management requires a good understanding of this condition. We present the first reported case of ACS and PLSVC occurring after thrombosis of the innominate vein (IV) after central venous catheter placement. The patient presented with atypical subacute chest pain and recurrent extrasystoles. Diagnosis and characterization of vascular anomalies was made by computed tomography phlebography, and the patient was successfully managed by endovascular recanalization of the IV.

Persistent pulmonary arterial hypertension in the newborn (PPHN): a frequent manifestation of TMEM70 defective patients.

INTRODUCTION: Mutations in the TMEM70 are the most common cause of nuclear ATP synthase deficiency resulting in a distinctive phenotype characterized by severe neonatal hypotonia, hypertrophic cardiomyopathy (HCMP), facial dysmorphism, severe lactic acidosis, hyperammonemia and 3-methylglutaconic aciduria (3-MGA). METHODS AND RESULTS: We collected 9 patients with genetically confirmed TMEM70 defect from 8 different families. Six were homozygous for the c.317-2A>G mutation, 2 were compound heterozygous for mutations c.317-2A>G and c.628A>C and 1 was homozygous for the novel c.701A>C mutation. Generalized hypotonia, lactic acidosis, hyperammonemia and 3-MGA were present in all since birth. Five patients presented acute respiratory distress at birth requiring intubation and ventilatory support. HCMP was detected in 5 newborns and appeared a few months later in 3 additional children. Five patients showed a severe and persistent neonatal pulmonary hypertension (PPHN) requiring Nitric Oxide (NO) and/or sildenafil administration combined in 2 cases with high-frequency oscillatory (HFO) ventilation. In 3 of these patients, echocardiography detected signs of HCMP at birth. CONCLUSIONS: PPHN is a life-threatening poorly understood condition with bad prognosis if untreated. Pulmonary hypertension has rarely been reported in mitochondrial disorders and, so far, it has been described in association with TMEM70 deficiency only in one patient. This report further expands the clinical and genetic spectrum of the syndrome indicating PPHN as a frequent and life-threatening complication regardless of the type of mutation. Moreover, in these children PPHN appears even in the absence of an overt cardiomyopathy, thus representing an early sign and a clue for diagnosis.

Clonotype selection and composition of human CD8 T cells specific for persistent herpes viruses varies with differentiation but is stable over time.

Protection from reactivation of persistent herpes virus infection is mediated by Ag-specific CD8 T cell responses, which are highly regulated by still poorly understood mechanisms. In this study, we analyzed differentiation and clonotypic dynamics of EBV- and CMV-specific T cells from healthy adults. Although these T lymphocytes included all subsets, from early-differentiated (EM/CD28(pos)) to late-differentiated (EMRA/CD28(neg)) stages, they varied in the sizes/proportions of these subsets. In-depth clonal composition analyses revealed TCR repertoires, which were highly restricted for CMV- and relatively diverse for EBV-specific cells. Virtually all virus-specific clonotypes identified in the EMRA/CD28(neg) subset were also found within the pool of less differentiated "memory" cells. However, striking differences in the patterns of dominance were observed among these subsets, because some clonotypes were selected with differentiation while others were not. Late-differentiated CMV-specific clonotypes were mostly characterized by TCR with lower dependency on CD8 coreceptor interaction. Yet all clonotypes displayed similar functional avidities, suggesting a compensatory role of CD8 in the clonotypes of lower TCR avidity. Importantly, clonotype selection and composition of each virus-specific subset upon differentiation was highly preserved over time, with the presence of the same dominant clonotypes at specific differentiation stages within a period of 4 years. Remarkably, clonotypic distribution was stable not only in late-differentiated but also in less-differentiated T cell subsets. Thus, T cell clonotypes segregate with differentiation, but the clonal composition once established is kept constant for at least several years. These findings reveal novel features of the highly sophisticated control of steady state protective T cell activity in healthy adults.

Disparate activation of the coronary sinus and inferior left atrium during atrial tachycardia after persistent atrial fibrillation ablation: prevalence, pitfalls, and impact on mapping.

INTRODUCTION: Persistent atrial fibrillation (AF) ablation may lead to partial disconnection of the coronary sinus (CS). As a result, disparate activation sequences of the local CS versus contiguous left atrium (LA) may be observed during atrial tachycardia (AT). We aimed to evaluate the prevalence of this phenomenon and its impact on activation mapping. METHODS: AT occurring after persistent AF ablation were investigated in 74 consecutive patients. Partial CS disconnection during AT was suspected when double potentials with disparate activation sequences were observed on the CS catheter. Endocardial mapping facing CS bipoles was performed to differentiate LA far-field from local CS potentials. RESULTS: A total of 149 ATs were observed. Disparate LA-CS activations were apparent in 20 ATs after magnifying the recording scale (13%). The most common pattern (90%) was distal to proximal endocardial LA activation against proximal to distal CS activation, the latter involving the whole CS or its distal part. Perimitral macroreentry was more common when disparate LA-CS activations were observed (67% vs 29%; P = 0.002). Partial CS disconnection also resulted in "pseudo" mitral isthmus (MI) block during LA appendage pacing in 20% of patients as local CS activation was proximal to distal despite distal to proximal activation of the contiguous LA. CONCLUSION: Careful analysis of CS recordings during AT following persistent AF ablation often reveals disparate patterns of activation. Recognizing when endocardial LA activation occurs in the opposite direction to the more obvious local CS signals is critical to avoid misleading interpretations during mapping of AT and evaluation of MI block.

Hypomorphic mutation in the site-1 protease Mbtps1 endows resistance to persistent viral infection in a cell-specific manner.

The prototypic arenavirus lymphocytic choriomeningitis virus (LCMV), which naturally persists in rodents, represents a model for HIV, HBV, and HCV. Cleavage of the viral glycoprotein precursor by membrane-bound transcription factor peptidase, site 1 (Mbtps1 or site-1 protease), is crucial for the life cycle of arenaviruses and therefore represents a potential target for therapy. Recently, we reported a viable hypomorphic allele of Mbtps1 (woodrat) encoding a protease with diminished enzymatic activity. Using the woodrat allele, we examine the role of Mbtps1 during persistent LCMV infection. Surprisingly, Mbtps1 inhibition limits persistent but not acute viral infection and is associated with an organ/cell type-specific decrease in viral titers. Analysis of bone marrow-derived dendritic cells from woodrat mice supports their specific role in resolving persistent viral infection. These results support in vivo targeting of Mbtps1 in the treatment of arenavirus infections and demonstrate a critical role for dendritic cells in persistent viral infections.

A New Method for ECG Tracking of Persistent Atrial Fibrillation Termination during Stepwise Ablation

Stepwise radiofrequency catheter ablation (step-CA) has become the treatment of choice for the restoration of sinus rhythm (SR) in patients with long-standing persistent atrial fibrillation (pers-AF). Its success rate appears limited as the amount of ablation to achieve long term SR is unknown. Multiple organization indexes (OIs) have been previously developed to track the organization of AF during step-CA, however, with limited success. We report an adaptive method for tracking AF termination (AF-term) based on OIs characterizing the relationship between harmonic components of atrial activity from the surface ECG of AF activity. By computing their relative evolution during the last two steps preceding AF-term, we found that the performance of our OIs was superior to classical indices to track the efficiency of step-CA "en route" to AF-term. Our preliminary results suggest that the gradual synchronization between the fundamental and its first harmonic of AF activity appears as a promising parameter for predicting AF-term during step-CA.

Multibacillary leprosy patients with high and persistent serum antibodies to leprosy IDRI diagnostic-1/LID-1: higher susceptibility to develop type 2 reactions

Leprosy inflammatory episodes [type 1 (T1R) and type 2 (T2R) reactions] represent the major cause of irreversible nerve damage. Leprosy serology is known to be influenced by the patient’s bacterial index (BI) with higher positivity in multibacillary patients (MB) and specific multidrug therapy (MDT) reduces antibody production. This study evaluated by ELISA antibody responses to leprosy Infectious Disease Research Institute diagnostic-1 (LID-1) fusion protein and phenolic glycolipid I (PGL-I) in 100 paired serum samples of 50 MB patients collected in the presence/absence of reactions and in nonreactional patients before/after MDT. Patients who presented T2R had a median BI of 3+, while MB patients with T1R and nonreactional patients had median BI of 2.5+ (p > 0.05). Anti-LID-1 and anti-PGL-I antibodies declined in patients diagnosed during T1R (p < 0.05). Anti-LID-1 levels waned in MB with T2R at diagnosis and nonreactional MB patients (p < 0.05). Higher anti-LID-1 levels were seen in patients with T2R at diagnosis (vs. patients with T1R at diagnosis, p = 0.008; vs. nonreactional patients, p = 0.020) and in patients with T2R during MDT (vs. nonreactional MB, p = 0.020). In MB patients, high and persistent anti-LID-1 antibody levels might be a useful tool for clinicians to predict which patients are more susceptible to develop leprosy T2R.