LipoCardium: endothelium-directed cyclopentenone prostaglandin-based liposome formulation that completely reverses atherosclerotic lesions.

Atherosclerosis is a multifactorial inflammatory disease of blood vessels which decimates one in every three people in industrialized world. Despite the important newest clinical approaches, currently available strategies (e.g. nutritional, pharmacological and surgical) may only restrain the worsening of vascular disease. Since antiproliferative cyclopentenone prostaglandins (CP-PGs) are powerful anti-inflammatory agents, we developed a negatively charged liposome-based pharmaceutical formulation (LipoCardium) that specifically direct CP-PGs towards the injured arterial wall cells of atherosclerotic mice. In the blood stream, LipoCardium delivers its CP-PG contents only into activated arterial wall lining cells due to the presence of antibodies raised against vascular cell adhesion molecule-1 (VCAM-1), which is strongly expressed upon inflammation by endothelial cells and macrophage-foam cells as well. After 4 months in a high-lipid diet, all low-density lipoprotein receptor-deficient adult control mice died from myocardium infarction or stroke in less than 2 weeks, whereas LipoCardium-treated (2 weeks) animals (still under high-lipid diet) completely recovered from vascular injuries. In vitro studies using macrophage-foam cells suggested a tetravalent pattern for LipoCardium action: anti-inflammatory, antiproliferative (and pro-apoptotic only to foam cells), antilipogenic and cytoprotector (via heat-shock protein induction). These astonishing cellular effects were accompanied by a marked reduction in arterial wall thickness, neointimal hyperplasia and lipid accumulation, while guaranteed lifespan to be extended to the elderly age. Our findings suggest that LipoCardium may be safely tested in humans in a near future and may have conceptual implications in atherosclerosis therapy.

Função e apoptose do neutrófilo: modulação pela maturação sexual, exercício e suplementação com glutamina / Neutrophil function and apoptosis: modulation by sexual maturation, exercise and glutamine supplementation

O efeito de uma única sessão de exercício (a 85% da capacidade máxima) na função e apoptose de neutrófilos de ratos de 60 (imaturos sexualmente) e 90 (maduro sexualmente) dias foi estudado. Avaliou-se também o efeito da suplementação com glutamina (1g por kg de peso) na prevenção das alterações induzidas pelo exercício. As funções estudadas foram: capacidade fagocitária, produção de óxido nítrico (NO) e de espécies reativas de oxigênio (EROs). Para avaliar a morte de neutrófilos, os seguintes parâmetros foram determinados: integridade de membrana, condensação de cromatina, fragmentação de DNA, externalização de fosfatidilserina, potencial transmembrânico mitocondrial, expressão de genes anti-apoptóticos (bcl-xL), próapoptóticos (bax e bcl-xS) e caspase 3. A maturação sexual por si só reduziu a capacidade fagocitária, aumentou a expressão dos componentes (gp91, p47 e p22phox) da NADPH-oxidase e aumentou a produção de óxido nítrico (NO). Além disso, a maturação sexual aumentou a proporção de células em apoptose, conforme observado pelo aumento na fragmentação de DNA e despolarização mitocondrial, redução na expressão de Bcl-xL e aumento da expressão de caspase 3. A sessão de exercício, reduziu a produção de NO e aumentou a expressão dos componentes da NADPHoxidase nos neutrófilos de ratos de 90 dias. Em ratos de 60 dias, o exercício não alterou as funções dos neutrófilos (capacidade fagocitária, produção de óxido nítrico e de espécies reativas de oxigênio - EROs). A sessão de exercício aumentou a proporção de neutrófilos em apoptose em ratos de 60 e 90 dias. A suplementação oral com glutamina aumentou a capacidade fagocitária e a produção de EROs nos neutrófilos de ratos de 60 dias e a expressão dos componentes da NADPH-oxidase nos neutrófilos de ratos de 90 dias. Além disso, a suplementação com glutamina reduziu o efeito do exercício na indução de apoptose nos neutrófilos dos ratos de 60 e 90 dias.

The effect of a single session of intense exercise (85% maximal capacity) on apoptosis and function of neutrophils from 60 (sexually immature) and 90 (sexually mature) days-old rats was examined. The possible effect of glutamine supplementation (1 g per kg body weight) to prevent the changes induced by the exercise was also investigated. The functions studied were: phagocytic capacity, nitric oxide (NO) and reactive oxygen species (ROS) production. To evaluate the process of neutrophils death, the following parameters were determined: cell viability, chromatin condensation, DNA fragmentation, phosphatidylserine externalization, mitocondrial transmembrane potential, expression of anti-apoptotic (bcl-xL) and pro-apoptotic (bax, bcl-xS and caspase 3) genes. The sexual maturation per se decreased the phagocytic capacity, raised the expression of the NADPH-oxidase components (gp91, p47 and p22phox) and increased nitric oxide (NO) production. In addition, sexual maturation increased the proportion of cells in apoptosis as indicated by the increase in DNA fragmentation, mitochondrial depolarization and in caspase expression, and a reduction in bcl-xL expression. The exercise session decreased NO production and increased the expression of the NADPH-oxidase components in neutrophils from 90 days old rats. In 60 days old rats, the exercise did not affect neutrophil functions studied: phagocytic capacity, NO and reactive oxygen species (ROS) production. The exercise raised the proportion of neutrophils in apoptosis in both 60 and 90 days-old rats. Oral glutamine supplementation raised the phagocytic capacity and reactive oxygen species (ROS) production in neutrophils from 90 days old rats. In addition, glutamine supplementation decreased the effect of exercise on the induction of apoptosis in neutrophils from both 60 and 90 days old rats.

Beneficial effect of glutamine on exercise-induced apoptosis of rat neutrophils.

INTRODUCTION/PURPOSE: The effect of a single bout of intensive exercise on apoptosis of rat neutrophils and the possible prevention by glutamine administration was examined. The experiments were performed in sexually immature and sexually mature male rats as to examine the possible involvement of sexual maturation in the effect of exercise. METHODS: Exercise was carried out on a treadmill for 1 h before rats were killed by decapitation. Aqueous solution of glutamine was given by gavage (1 g.kg-1 body weight), 1 h before exercise. Neutrophils were obtained by intraperitoneal lavage with phosphate-buffered saline (PBS), 4 h after injection of oyster glycogen solution. The cells were then analyzed for apoptosis by flow cytometry and fluorescence microscopy. Pro- and antiapoptotic gene expression was evaluated by reverse transcriptase chain reaction (RT-PCR). RESULTS: Neutrophils obtained from immature and mature exercised rats showed an increase in DNA fragmentation, chromatin condensation, and phosphatidylserine externalization. This suggests that all neutrophils suffered apoptosis. To study the possible mechanism involved, the production of reactive oxygen metabolites, expression of genes involved in apoptosis and mitochondrial transmembrane potential were examined. Acute exercise raised reactive oxygen metabolites production by neutrophils. Exercise did not change the expression of antiapoptotic (bcl-xL) and apoptotic (bax and bcl-xS) genes in neutrophils from immature rats but caused a significant increase of bax and bcl-xS expression and provoked a significant decrease of bcl-xL expression in cells from mature rats. Exercise also induced a marked loss of mitochondrial depolarization in neutrophils. Oral glutamine supplementation partially prevented the exercise-induced apoptosis in neutrophils from sexually immature and mature rats. CONCLUSION: The protective effect of glutamine on neutrophil apoptosis induced by acute exercise possibly occurs by preservation of mitochondrial function.