Mitochondrial morphofunctional alterations in smooth muscle cells of aorta in rats.

In an experimental model of atherogenesis induced by hyperfibrinogenemia (HF), the pharmacological response of vitamin E was studied in order to assess its antioxidant effect on the mitochondrial morphofunctional alterations in aortic smooth muscle cells. Three groups of male rats were used: (Ctr) control, (AI) atherogenesis induced for 120 days, and (AIE) atherogenesis induced for 120 days and treated with vitamin E. HF was induced by adrenalin injection (0.1 mg/day/rat) for 120 days. AIE group was treated with the administration of 3.42 mg/day/rat of vitamin E for 105 days after the first induction. Mitochondria morphology was analyzed by electronic microscopy (EM) and mitochondrial complexes (MC) by spectrophotometry. In group AI the total and mean number of mitochondria reduced significantly, the intermembranous matrix increased, and swelling was observed with respect to Ctr and AIE (P < 0.01). These damages were related to a significant decrease in the activity of citrate synthase and complexes I, II, III, and IV in group AI in comparison to Ctr (P < 0.001). Similar behavior was presented by group AI compared to AIE (P < 0.001). These results show that vitamin E produces a significative regression of inflammatory and oxidative stress process and it resolved the morphofunctional mitochondrial alterations in this experimental model of atherogenic disease.

Effects of Atorvastatin on Oxidative Stress Biomarkers and Mitochondrial Morphofunctionality in Hyperfibrinogenemia-Induced Atherogenesis.

Relationship between hyperfibrinogenemia (HF), oxidative stress, and atherogenesis was established. Effect of atorvastatin (Ator) was assessed. Wistar male (6 months) rats were studied: Ctr, control, without HF induction; Ctr-Ator, without HF treated with atorvastatin; AI, atherogenesis induced, and AI-Ator, atherogenesis induced and treated with atorvastatin. Atherogenesis was induced by daily adrenaline injection (0.1 mL/day/rat) for 90 days; treatment started 15 days after induction. Fibrinogen (mg/dL) and nitric oxide (NO) were measured in plasma (mM) and superoxide dismutase (SOD) (U/mL) in red cell lysate by spectrophotometry. Slices of aorta were analyzed by electron microscopy (EM). ANOVA and chi-square test were used; P < 0.05 was established. There were no significant differences between Ctr and Ctr-Atorv in fibrinogen, NO, and SOD values. Comparing Ctr with AI an increase of fibrinogen is observed (P < 0.001), but it decreased after administration of atorvastatin in AI-Ator (P < 0.001). NO diminished in AI relative to Ctr and increased in AI-Ator (P < 0.001). SOD showed an increase in AI and AI-Ator compared to Ctr (P < 0.001). EM revealed expansion of intermembrane space and disorganization of crests in AI. In AI-Ator mitochondrial areas and diameters were similar to control. Atorvastatin normalizes HF, stabilizes NO, increases SOD, and produces a partial regression of mitochondrial lesions.

Efectos de atioxidantes en un modelo aterogénico por hiperfibrinogenemia inducida / Effects of an atherogenic model atioxidantes induced hyperfibrinogenemia

La hiperfibrinogenemia (HF) sería un marcador precoz de aterogénesis activando la vía fisiopatológica del óxido nítrico (NO) y L- citrulina en la pared vascular, induciendo estrés oxidativo (EO), evento temprano que refleja la disfunción endotelial, repercutiendo sobre la histomorfología vascular y la morfofuncionalidad mitocondrial. Es probable que los sistemas de defensa antioxidantes endógenos, como la enzima superóxido dismutasa (SOD), sean insuficientes para contrarrestar el daño oxidativo vascular, por esta razón se busca incrementar dicha defensa con la administración de fármacos antioxidantes: α-tocoferol y L-ascorbato. Se estudió el efecto de los mismos para evitar la instauración de las primeras lesiones aterogénicas en un modelo experimental de aterogénesis inducida por HF, mediante la determinación de biomarcadores inflamatorios y de estrés oxidativo: fibrinógeno, NO, SOD y L-citrulina por espectrofotometría y evaluando la histopatología de la pared de aorta torácica por microscopia óptica y las alteraciones morfofuncionales mitocondriales en células musculares lisas aórticas por microscopia electrónica. También se valoró la actividad enzimática de Citrato Sintasa y los complejos I, II, III y IV de la cadena respiratoria mitocondrial. Se utilizaron 240 ratas machos, cepa Wistar, distribuidas en 20 grupos constituidos por 12 animales cada uno: control, control + vitaminas E, C ó E + C, inducción de HF por 30, 60, 90 y 120 días, inducción de HF por 30, 60, 90 y 120 días + vitamina E, C ó E + C, respectivamente. La inducción de HF se realizó mediante inyecciones subcutáneas de adrenalina (0.1mg/Kg/rata/día)

SUMMARY: Hyperfibrinogenemia (HF) is an early marker of atherogenesis that activates the pathophysiological way of nitric oxide (NO) and L- citrulline in the vascular wall, inducing oxidative stress (OS), an early event that reflects endothelial dysfunction and affects the vascular histomorphology and the mitochondrial morphofunctionality. it is likely that the endogenous antioxidant defense systems, such as the superoxide dismutase enzyme (SOD), are insufficient to counteract the oxidative vascular damage; for this reason, we seek to increase said defense with the administration of antioxidant drugs: α-tocopherol and Lascorbate. The effect of these was studied in order to avoid the first atherogenic injuries in an experimental model of atherogenesis induced by HF and through the establishment of inflammatory biomarkers and oxidative stress: fibrinogen, NO, SOD and L-citrulline by spectrophotometry and considering the histopathology of the thoracic aorta wall by optical microscopy and the mitochondrial morphofunctional alterations in aortic smooth muscle cells by electron microscopy. Also, the enzymatic activity of citrate synthase and the complexes I, II, III and IV of the mitochondrial respiratory chain were considered. We used 240 male rats, Wistar strain, distributed in 20 groups made up of 12 animals each: control, control + vitamins E, C or E + C, HF induction for 30, 60, 90 and 120 days, HF induction for 30, 60, 90 and 120 days + vitamin E, C or E + C, respectively. HF induction was made with subcutaneous injections of adrenaline (0.1mg/Kg/rat/day)

Efectos de atioxidantes en un modelo aterogénico por hiperfibrinogenemia inducida / Effects of an atherogenic model atioxidantes induced hyperfibrinogenemia

La hiperfibrinogenemia (HF) sería un marcador precoz de aterogénesis activando la vía fisiopatológica del óxido nítrico (NO) y L- citrulina en la pared vascular, induciendo estrés oxidativo (EO), evento temprano que refleja la disfunción endotelial, repercutiendo sobre la histomorfología vascular y la morfofuncionalidad mitocondrial. Es probable que los sistemas de defensa antioxidantes endógenos, como la enzima superóxido dismutasa (SOD), sean insuficientes para contrarrestar el daño oxidativo vascular, por esta razón se busca incrementar dicha defensa con la administración de fármacos antioxidantes: α-tocoferol y L-ascorbato. Se estudió el efecto de los mismos para evitar la instauración de las primeras lesiones aterogénicas en un modelo experimental de aterogénesis inducida por HF, mediante la determinación de biomarcadores inflamatorios y de estrés oxidativo: fibrinógeno, NO, SOD y L-citrulina por espectrofotometría y evaluando la histopatología de la pared de aorta torácica por microscopia óptica y las alteraciones morfofuncionales mitocondriales en células musculares lisas aórticas por microscopia electrónica. También se valoró la actividad enzimática de Citrato Sintasa y los complejos I, II, III y IV de la cadena respiratoria mitocondrial. Se utilizaron 240 ratas machos, cepa Wistar, distribuidas en 20 grupos constituidos por 12 animales cada uno: control, control + vitaminas E, C ó E + C, inducción de HF por 30, 60, 90 y 120 días, inducción de HF por 30, 60, 90 y 120 días + vitamina E, C ó E + C, respectivamente. La inducción de HF se realizó mediante inyecciones subcutáneas de adrenalina (0.1mg/Kg/rata/día)(AU)

SUMMARY: Hyperfibrinogenemia (HF) is an early marker of atherogenesis that activates the pathophysiological way of nitric oxide (NO) and L- citrulline in the vascular wall, inducing oxidative stress (OS), an early event that reflects endothelial dysfunction and affects the vascular histomorphology and the mitochondrial morphofunctionality. it is likely that the endogenous antioxidant defense systems, such as the superoxide dismutase enzyme (SOD), are insufficient to counteract the oxidative vascular damage; for this reason, we seek to increase said defense with the administration of antioxidant drugs: α-tocopherol and Lascorbate. The effect of these was studied in order to avoid the first atherogenic injuries in an experimental model of atherogenesis induced by HF and through the establishment of inflammatory biomarkers and oxidative stress: fibrinogen, NO, SOD and L-citrulline by spectrophotometry and considering the histopathology of the thoracic aorta wall by optical microscopy and the mitochondrial morphofunctional alterations in aortic smooth muscle cells by electron microscopy. Also, the enzymatic activity of citrate synthase and the complexes I, II, III and IV of the mitochondrial respiratory chain were considered. We used 240 male rats, Wistar strain, distributed in 20 groups made up of 12 animals each: control, control + vitamins E, C or E + C, HF induction for 30, 60, 90 and 120 days, HF induction for 30, 60, 90 and 120 days + vitamin E, C or E + C, respectively. HF induction was made with subcutaneous injections of adrenaline (0.1mg/Kg/rat/day)(AU)

Papel antioxidante de la vitamina E en la aterogénesis inducida por hiperfibrinogenemia / Antioxidant Role of Vitamin E in Atherogenesis Induced by Hyperfibrinogenemia

Con el propósito de estudiar el efecto de la vitamina E sobre el estrés oxidativo desencade- nado por hiperfibrinogenemia (HF) en un modelo experimental de aterogénesis y la posible normalización de los indicadores de estrés oxidativo, se evaluaron: óxido nítrico (NO), L-citrulina, superóxido dismutasa (SOD) e involución de lesiones histopatológicas en la aorta torácica. El estudio se realizó en 36 ratas, cepa Wistar, que se dividieron en tres grupos (n = 12 cada uno): A, control; B, HF × 90 días; C, HF × 90 días + vitamina E. La HF se indujo mediante inyecciones de adrenalina (0,1 ml/día/rata) por 90 días. La dosis de vitamina E fue de 2 mg/día/rata durante 75 días. Se dosaron en plasma los niveles de fibrinógeno (mg/dl), NO (uM) y L-citrulina (mM) y en lisado de glóbulos rojos, por espectrofotometría, se determinó la actividad de la SOD (U/ml). Se analizaron cortes de la aorta torácica por microscopia óptica (MO). Para el análisis estadístico se emplearon MANOVA y la prueba de Fisher; se estableció un nivel de significación de p < 0,05. Se observó un aumento significativo de fibrinógeno en el grupo B (407 ± 8,9 mg/dl) en comparación con los grupos A (203 ± 9 mg/dl) y C (191,58 ± 17,79 mg/dl) (p < 0,001). El NO disminuyó significativamente en el grupo B (13,73 ± 1,76 uM) frente a los grupos A (23,58 ± 0,08 uM) y C (26,64 ± 3,65 uM) (p < 0,001). La L-citrulina aumentó en forma significativa en los grupos B (4,99 ± 0,18 mM) y C (6,60 ± 0,16 mM) en comparación con el grupo A (3,03 ± 0,13 mM) (p < 0,001). El SOD incrementó su actividad en los grupos B (251,67 ± 10,34 U/ml) y C (304,75 ± 10,43 U/ml) frente al grupo A (139,44 ± 4,74 U/ml) (p < 0,001). La microscopia óptica mostró denudación endotelial, engrosamiento intimal y protrusión de la pared en el grupo B (90%) y recuperación de la denudación endotelial y disminución del 50% del engrosamiento intimal en el grupo C (p < 0,001). Niveles aumentados de SOD serían insuficientes para impedir alteraciones en la vía del estrés oxidativo inducido por la HF. La vitamina E actuaría deteniendo la reacción en cadena iniciada por los radicales libres y en consecuencia disminuiría el anión superóxido, estimulando de esta manera un incremento en la biodisponibilidad del NO y normalizando las concentraciones de fibrinógeno plasmático.

We used an experimental model of atherogenesis to evaluate the effect of vitamin E on oxidative stress induced by hyperfibrinogenemia (HF) and the possible normalization of oxidative stress markers. The following variables were studied: nitric oxide (NO), L-citrulline, superoxide dismutase (SOD) activity and regression of histopathological lesions in the thoracic aorta. The study was performed in 36 Wistar rats that were divided into three groups of 12 rats each: A, control group; B, HF for 90 days; C, HF for 90 days + vitamin E. Hyperfibrinogenemia was induced by the injection of epinephrine (0.1 ml/day/rat) during 90 days. The dose of vitamin E was 2 mg/day/rat during 75 days. We measured the plasma levels of fibrinogen (mg/dl), NO (uM) and L-citrulline (mM); SOD activity (U/ml) was assayed in red cell lysates using spectrophotometry. The histopathological sections of the thoracic aorta were examined using light microscopy (LM). Statistical analysis was performed using MANOVA and Fisher's test; a p value <0.05 was considered statistically significant. Rats in group B had a significant increase in fibrinogen levels B (407±8.9 mg/dl) compared to groups A (203±9 mg/dl) and C (191.58±17.79 mg/dl) (p<0.001). We observed a significant decrease in NO in group B (13.73±1.76 uM) versus groups A (23.58±0.08 uM) and C (26.64±3.65 uM) (p<0.001). L-citrulline increased significantly in groups B (4.99±0.18 mM) and C (6.60±0.16 mM) compared to group A (3.03±0.13 mM) (p<0.001). SOD activity was greater in groups B (251.67±10.34 U/ml) and C (304.75±10.43 U/ml) versus group A (139.44±4.74 U/ml) (p<0.001). Light microscopic examination revealed the presence of endothelial denudation, intimal thickening and vessel wall protrusion in group B (90%), while recovery of endothelial denudation and a 50% reduction in intimal thickening was observed in group C (p<0.001). High SOD activity might be insufficient to prevent abnormalities in the oxidative stress pathway induced by HF. Vitamin E would stop the chain reaction initiated by free radicals and thus decrease the superoxide anion, stimulating the bioavailability of NO with normalization of fibrinogen plasma levels.