The effect of high-fructose feeding on hemodynamic behavior and infarct size of isolated rat hearts subjected to low-flow ischemia.

Objective: The aim of this study was to investigate the potentially deleterious effect of in vivo high-fructose feeding of rats on ex vivo hemodynamic recovery and infarct size of isolated rat hearts subjected to low-flow ischemia and reperfusion. Methods: After feeding Sprague-Dawley male rats with a high-fructose (n=9), high-glucose (n=9) or a standard diet (n=9) for four weeks, the hearts were extirpated and perfused ex vivo with a Krebs-Henseleit solution for 15 min; thereafter the hearts were subjected to low flow (0.3 ml/min) ischemia during 30 min followed by 60 min reperfusion. Left ventricular developed pressure (LVDP), first derivative of pressure development during the contraction and relaxation phase and heart rate were recorded prior to ischemia and after reperfusion. Infarct area was measured at the end of the reperfusion period. Results: In isolated hearts, subjected to ex vivo low-flow ischemia followed by 60 min reperfusion, the relative myocardial infarct size did not differ between the three groups. Post-ischemic cardiac contractile recovery appeared to be complete in both the high-fructose and high-glucose group at 60 min reperfusion. In contrast, in the control group LVDP and left ventricular relaxation rate were still depressed at the end of 60 minutes of reperfusion. Conclusion: Feeding with a high-fructose diet of rats appears to have a positive effect on the recovery of left ventricular contractile function after low-flow ischemia, in comparison to a standard diet, without a difference in relative myocardial infarct size. Similar results were obtained in the high-glucose fed rats

Atriyal natriüretik peptid infüzyonunun izole sıçan kalbinde iskemi sonrası oluşturduğu hemodinamik değişikliklerde egzersizin rolü

Amaç: Bu çalışmada düşük akımlı iskemi sonrası reperfüzyon döneminde atriyal natriüretik peptid ANP uygulamasının sol ventrikül hemodinamik yanıtlarına etkisi ve bu etkide egzersizin rolü araştırıldı. Gereç ve yöntem: Çalışmada yer alan tüm sıçanlara 60 dakika düşük akımlı iskemi ve takiben 120 dk reperfüzyon uygulandı. Egzersiz E gruplarına ardışık olarak beş gün süreyle yürüyüş egzersizini takiben iskemi ve reperfüzyon uygulandı. Reperfüzyonun ilk 15 dakikasında 0.1 μM/L ANP infüzyonu yapılan iki grup; ANP n=6 ve Egzersiz-ANP n=6 gruplarını oluşturdu. Tüm gruplarda sol ventrikül gelişim basıncı SVGB , maksimum ve minimum sol ventrikül basınç değişim oranları +dP/dt and -dP/dt kaydedildi. Bulgular: Kontrol K , E, ANP ve E-ANP gruplarının SVGB, +dp/dt, -dp/dt ve kalp hızı değerleri iskemi öncesinde ve iskemi sonrası reperfüzyonun 1, 60 ve 120. dakikalarında karşılaştırıldığında gruplar arasında istatistiksel olarak anlamlı bir farklılık göstermedi. Benzer şekilde infarkt alanlarının yüzde değerleri karşılaştırıldığında dört grup arasında istatistiksel olarak anlamlı farklılık bulunmadı. Sonuç: Kısa süreli yoğun egzersiz, reperfüzyon döneminde ANP uygulamasına bağlı kardiyak kontraktilitede oluşan değişimleri etkilememektedi

Plasmodium infection of hepatocytes : role of protein micronemes sporozoltes

L’infection par Plasmodium, parasite responsable du paludisme, débute par l’injection de sporozoïtes par un moustique du genre Anopheles. La première cible des sporozoïtes est le foie, où le parasite se développe avant l’initiation d'une phase d'infection érythrocytaire symptomatique. Dans le foie, les sporozoïtes pénètrent activement les hépatocytes en formant une vacuole parasitophore, dans laquelle le parasite se multiplie. Cette étape, appelée invasion productive, implique des facteurs parasitaires et des protéines de l’hôte, notamment CD81. Toutefois, les mécanismes mis en jeu restent méconnus. À l’aide d’une nouvelle approche génétique développée au laboratoire, nous avons produit de nouvelles souches de parasites transgéniques fluorescents, notamment chez le parasite de rongeurs P. yoelii. L’utilisation des parasites de P. yoelii GFP et d’un système cellulaire de lignées permissives ou non à l’infection, nous a permis de mieux caractériser les mécanismes cellulaires et moléculaires mis en jeu lors de l’invasion. Nous avons confirmé que l’invasion productive est précédée d’une phase de traversée cellulaire. Nous avons découvert et caractérisé la formation de vacuoles transitoires lors de cette phase de traversée cellulaire, distinctes des vacuoles parasitophores productives. Nos résultats montrent que le parasite se sert d’une perforine parasitaire, PLP1 (Perforin-Like Protein 1), pour sortir de cette vacuole transitoire et échapper à la dégradation par les lysosomes cellulaires. Une fois activés, les sporozoïtes passent d’un mode de traversée à un mode d’invasion productive. Nous avons montré que CD81 joue un rôle dans l’invasion productive. CD81 est nécessaire pour induire la sécrétion des rhoptries parasitaires, impliquées dans la formation de la jonction mobile, une structure à travers laquelle le parasite se glisse pour pénétrer dans la cellule. Nous avons pu aussi montrer qu’une autre protéine des hépatocytes, SRBI (scavenger receptor BI), définit une voie d’entrée indépendante de CD81 pour P. berghei et P. vivax. Par une approche génétique originale, nous avons pu montrer que deux protéines des micronèmes des sporozoïtes, P52 et P36, jouent un rôle majeur dans l’entrée via CD81 et SRBI, et mis à jour un lien fonctionnel entre P36 et l’entrée via SRBI. Enfin, nous avons développé plusieurs approches génétiques pour cibler le gène d’ama1 chez P. yoelii, une protéine des micronèmes impliquées dans la formation de la jonction. Nos résultats nous éclairent un peu plus sur les mécanismes d’invasion des sporozoïtes, et ouvrent des perspectives intéressantes vers le développement de nouvelles stratégies vaccinales.Infection with the Plasmodium parasite begins with the injection of sporozoites by an Anopheles mosquito. The first target is the liver where the parasite replicates as a pre-requisite to the development of pathogenic blood stage infection. In the liver, sporozoites penetrate hepatocytes forming a parasitophorous vacuole in which the parasite multiplies. This step, the productive invasion, involves parasitic factors and host proteins, particularly CD81, but the underlying mechanisms remain largely unknown. To facilitate monitoring of sporozoite invasion, we generated novel transgenic fluorescent parasites, using a new selection strategy named GOMO (gene out marker out) in the rodent parasite P. yoelii. The use of this transgenic parasite and of host cell lines permissive or not to infection, has allowed us to better characterize the cellular and molecular mechanisms involved during invasion. We have confirmed that the productive invasion is preceded by a cell traversal phase. We discovered and characterized the formation of transient vacuoles during this step, before formation of the parasitophorous vacuole. Our results uncovered that the perforin-like protein (PLP1) mediates sporozoite egress from transient vacuoles and escape from degradation by the cell lysosomes. Once activated, the sporozoites switch from the mode of cell traversal to productive invasion. We show that CD81 plays a role in the productive invasion. CD81 is necessary to induce the secretion of rhoptries proteins, involved in the formation of the moving junction, a structure through which the parasite glides to enter the cell. We could also show that another hepatocyte protein, SR-B1 (scavenger receptor B1), defines a CD81-independent pathway for P. berghei and P. vivax infection. Using an original genetic approach, we have shown that two sporozoite micronemal proteins, P52 and P36, play a role in the entry via CD81 and SR-B1, and highlighted a functional link between P36 and entry via SR-B1. Finally, we have developed several genetic approaches to target ama1 gene in P. yoelii, which encodes a protein involved in the formation of the moving junction. Altogether, our results contribute to improve our understanding of the mechanisms of sporozoite invasion, and open interesting perspectives for the development of novel vaccine strategies

Infection des hépatocytes par Plasmodium : rôle des protéines de micronèmes des sporozoïtes

Infection with the Plasmodium parasite begins with the injection of sporozoites by an Anopheles mosquito. The first target is the liver where the parasite replicates as a pre-requisite to the development of pathogenic blood stage infection. In the liver, sporozoites penetrate hepatocytes forming a parasitophorous vacuole in which the parasite multiplies. This step, the productive invasion, involves parasitic factors and host proteins, particularly CD81, but the underlying mechanisms remain largely unknown. To facilitate monitoring of sporozoite invasion, we generated novel transgenic fluorescent parasites, using a new selection strategy named GOMO (gene out marker out) in the rodent parasite P. yoelii. The use of this transgenic parasite and of host cell lines permissive or not to infection, has allowed us to better characterize the cellular and molecular mechanisms involved during invasion. We have confirmed that the productive invasion is preceded by a cell traversal phase. We discovered and characterized the formation of transient vacuoles during this step, before formation of the parasitophorous vacuole. Our results uncovered that the perforin-like protein (PLP1) mediates sporozoite egress from transient vacuoles and escape from degradation by the cell lysosomes. Once activated, the sporozoites switch from the mode of cell traversal to productive invasion. We show that CD81 plays a role in the productive invasion. CD81 is necessary to induce the secretion of rhoptries proteins, involved in the formation of the moving junction, a structure through which the parasite glides to enter the cell. We could also show that another hepatocyte protein, SR-B1 (scavenger receptor B1), defines a CD81-independent pathway for P. berghei and P. vivax infection. Using an original genetic approach, we have shown that two sporozoite micronemal proteins, P52 and P36, play a role in the entry via CD81 and SR-B1, and highlighted a functional link between P36 and entry via SR-B1. Finally, we have developed several genetic approaches to target ama1 gene in P. yoelii, which encodes a protein involved in the formation of the moving junction. Altogether, our results contribute to improve our understanding of the mechanisms of sporozoite invasion, and open interesting perspectives for the development of novel vaccine strategies.L’infection par Plasmodium, parasite responsable du paludisme, débute par l’injection de sporozoïtes par un moustique du genre Anopheles. La première cible des sporozoïtes est le foie, où le parasite se développe avant l’initiation d'une phase d'infection érythrocytaire symptomatique. Dans le foie, les sporozoïtes pénètrent activement les hépatocytes en formant une vacuole parasitophore, dans laquelle le parasite se multiplie. Cette étape, appelée invasion productive, implique des facteurs parasitaires et des protéines de l’hôte, notamment CD81. Toutefois, les mécanismes mis en jeu restent méconnus. À l’aide d’une nouvelle approche génétique développée au laboratoire, nous avons produit de nouvelles souches de parasites transgéniques fluorescents, notamment chez le parasite de rongeurs P. yoelii. L’utilisation des parasites de P. yoelii GFP et d’un système cellulaire de lignées permissives ou non à l’infection, nous a permis de mieux caractériser les mécanismes cellulaires et moléculaires mis en jeu lors de l’invasion. Nous avons confirmé que l’invasion productive est précédée d’une phase de traversée cellulaire. Nous avons découvert et caractérisé la formation de vacuoles transitoires lors de cette phase de traversée cellulaire, distinctes des vacuoles parasitophores productives. Nos résultats montrent que le parasite se sert d’une perforine parasitaire, PLP1 (Perforin-Like Protein 1), pour sortir de cette vacuole transitoire et échapper à la dégradation par les lysosomes cellulaires. Une fois activés, les sporozoïtes passent d’un mode de traversée à un mode d’invasion productive. Nous avons montré que CD81 joue un rôle dans l’invasion productive. CD81 est nécessaire pour induire la sécrétion des rhoptries parasitaires, impliquées dans la formation de la jonction mobile, une structure à travers laquelle le parasite se glisse pour pénétrer dans la cellule. Nous avons pu aussi montrer qu’une autre protéine des hépatocytes, SRBI (scavenger receptor BI), définit une voie d’entrée indépendante de CD81 pour P. berghei et P. vivax. Par une approche génétique originale, nous avons pu montrer que deux protéines des micronèmes des sporozoïtes, P52 et P36, jouent un rôle majeur dans l’entrée via CD81 et SRBI, et mis à jour un lien fonctionnel entre P36 et l’entrée via SRBI. Enfin, nous avons développé plusieurs approches génétiques pour cibler le gène d’ama1 chez P. yoelii, une protéine des micronèmes impliquées dans la formation de la jonction. Nos résultats nous éclairent un peu plus sur les mécanismes d’invasion des sporozoïtes, et ouvrent des perspectives intéressantes vers le développement de nouvelles stratégies vaccinales

A differential concentration-dependent effect of IVIg on neutrophil functions: relevance for anti-microbial and anti-inflammatory mechanisms

Background Polymorphonuclear neutrophils (PMN) play a key role in host defences against invading microorganisms but can also potentiate detrimental inflammatory reactions in case of excessive or misdirected responses. Intravenous immunoglobulins (IVIg) are used to treat patients with immune deficiencies and, at higher doses, in autoimmune, allergic and systemic inflammatory disorders. Methodology/Principal Findings We used flow cytometry to examine the effects of IVIg on PMN functions and survival, using whole-blood conditions in order to avoid artifacts due to isolation procedures. IVIg at low concentrations induced PMN activation, as reflected by decreased L-selectin and increased CD11b expression at the PMN surface, oxidative burst enhancement, and prolonged cell survival. In contrast, IVIg at higher concentrations inhibited LPS-induced CD11b degranulation and oxidative burst priming, and counteracted LPS-induced PMN lifespan prolongation. Conclusions/Significance IVIg appears to have differential, concentration-dependent effects on PMN, possibly supporting the use of IVIg as either an anti-microbial or an anti-inflammatory agent

Intravenous Gammaglobulin Inhibits Encephalitogenic Potential of Pathogenic T Cells and Interferes with their Trafficking to the Central Nervous System, Implicating Sphingosine-1 Phosphate Receptor 1–Mammalian Target of Rapamycin Axis

International audienc

Effect of IVIg on human dendritic cell-mediated antigen uptake and presentation: Role of lipid accumulation

International audienc

Malaria Sporozoites Traverse Host Cells within Transient Vacuoles

International audiencePlasmodium sporozoites are deposited in the host skin by Anopheles mosquitoes. The parasites migrate from the dermis to the liver, where they invade hepatocytes through a moving junction (MJ) to form a replicative parasitophorous vacuole (PV). Malaria sporozoites need to traverse cells during progression through host tissues, a process requiring parasite perforin-like protein 1 (PLP1). We find that sporozoites traverse cells inside transient vacuoles that precede PV formation. Sporozoites initially invade cells inside transient vacuoles by an active MJ-independent process that does not require vacuole membrane remodeling or release of parasite secretory organelles typically involved in invasion. Sporozoites use pH sensing and PLP1 to exit these vacuoles and avoid degradation by host lysosomes. Next, parasites enter the MJ-dependent PV, which has a different membrane composition, precluding lysosome fusion. The malaria parasite has thus evolved different strategies to evade host cell defense and establish an intracellular niche for replication

A rapid and robust selection procedure for generating drug-selectable marker-free recombinant malaria parasites

International audienceExperimental genetics have been widely used to explore the biology of the malaria parasites. The rodent parasites Plasmodium berghei and less frequently P. yoelii are commonly utilised, as their complete life cycle can be reproduced in the laboratory and because they are genetically tractable via homologous recombination. However, due to the limited number of drug-selectable markers, multiple modifications of the parasite genome are difficult to achieve and require large numbers of mice. Here we describe a novel strategy that combines positive-negative drug selection and flow cytometry-assisted sorting of fluorescent parasites for the rapid generation of drug-selectable marker-free P. berghei and P. yoelii mutant parasites expressing a GFP or a GFP-luciferase cassette, using minimal numbers of mice. We further illustrate how this new strategy facilitates phenotypic analysis of genetically modified parasites by fluorescence and bioluminescence imaging of P. berghei mutants arrested during liver stage development