Evaluation of Collagen Fibers, MMP2, MMP9, 8-OHdG and Apoptosis in the Aorta of Ovariectomized LDL Knockout Mice Submitted to Aerobic Exercise / Avaliação de Fibras de Colágeno, MMP2, MMP9, 8-OH-Dg e Apoptose na Aorta de Ratas LDL Nocaute Ovariectomizadas Submetidas a Exercício Aeróbico

Abstract Background: In menopause, there is greater cellular exposure to oxidative stress, related to the decreased antioxidative effects of estrogen. These metabolic changes favor the progression of cardiovascular diseases, such as atherosclerosis. Abnormal function of the aorta - the most important artery - is associated with many cardiovascular diseases. Collagen, especially types I and III, is one of the most important aortic wall components and it can be affected by many factors, including menopause. The 8-OHdG is one of the main markers of DNA oxidative damage induced by reactive oxygen species (ROS). Objective: We aimed to investigate effects of moderate aerobic training on the ascending aorta of LDL-knockout (LDL-KO) and ovariectomized female mice. Methods: A total of 15 C57BL/6 mice and 15 LDL-KO mice were divided into experimental groups. The thickness and volume density of types I and III collagen fibers were performed by morphoquantitative analysis, whereas the MMP-2 and MMP-9 and 8-OHdG were detected by immunohistochemistry and apoptosis was detected by the TUNEL assay. The significance level for all tests was p < 0.05. Results: Exercise causes an increase in the thickness of the aorta in LDL-KO groups, particularly accentuated in the ovariectomized groups. The type I collagen fibers showed an increase in volume density influenced by training in both Control groups and in the LDL-KO group. Type III collagen density decreased in both groups. The MMP-2 showed moderade immunostaining in the tunica media in LDL-KO groups, which did not occur in the control groups and the MMP-9 stained irregularly in all tissues. The marker 8-OhdG was stronger in the exercise training groups. Additionally, the ovariectomy, the exercise training and the LDL-KO treatments increased apoptosis. Conclusion: These results suggest that moderate-intensity aerobic exercise in ovariectomized mice associated to an increase in LDL rate possibly increases oxidative stress and apoptosis induction.

Resumo Fundamento: Na menopausa, há maior exposição celular ao estresse oxidativo, relacionada à diminuição dos efeitos antioxidantes do estrogênio. Essas alterações metabólicas favorecem a progressão das doenças cardiovasculares, como a aterosclerose. A função anormal da aorta - a artéria mais importante - está associada a muitas doenças cardiovasculares. O colágeno, especialmente os tipos I e III, é um dos mais importantes componentes da parede da aorta e pode ser afetado por muitos fatores, incluindo a menopausa. Por sua vez, 8-OHdG é um dos principais marcadores de danos oxidativos do DNA induzidos por espécies reativas de oxigênio (EROS). Objetivo: Investigar os efeitos do treinamento aeróbico moderado na aorta ascendente de camundongos fêmeas, nocaute para LDL (LDL-KO) e ovariectomizadas. Métodos: Um total de 15 animais C57BL/6 e 15 animais LDL-KO foram divididos em grupos experimentais. A espessura e a densidade de volume das fibras de colágeno tipos I e III foram realizadas por análise morfoquantitativa; MMP-2 e MMP-9 e 8-OHdG foram detectadas por imunohistoquímica; e a apoptose foi detectada pelo ensaio TUNEL. O nível de significância adotado para todos os testes realizados foi p < 0,05. Resultados: o exercício causa aumento da espessura da aorta em grupos LDL-KO, particularmente acentuada em grupos ovariectomizados. As fibras de colágeno de tipo I mostraram aumento da densidade de volume influenciado pelo treinamento em animais controle e LDL-KO. A densidade do colágeno tipo III diminuiu em ambos os grupos. A MMP-2 mostrou imunomarcação moderada na túnica média em animais LDL-KO; em grupos controle, a MMP-9 marcou irregularmente em todos os tecidos. O marcador 8-OHdG foi mais forte nos grupos de treinamento de exercícios. Além disso, a ovariectomia, o treinamento físico e os tratamentos de LDL-KO aumentaram a apoptose. Conclusão: Esses resultados sugerem que exercícios aeróbicos de intensidade moderada em camundongos ovariectomizados associados ao aumento da taxa de LDL, possivelmente, aumentam o estresse oxidativo e a indução da apoptose.

Intensity of swimming exercise influences aortic reactivity in rats

Exercise is known to cause a vasodilatory response; however, the correlation between the vasorelaxant response and different training intensities has not been investigated. Therefore, this study evaluated the vascular reactivity and lipid peroxidation after different intensities of swimming exercise in rats. Male Wistar rats (aged 8 weeks; 250-300 g) underwent forced swimming for 1 h whilst tied to loads of 3, 4, 5, 6, and 8% of their body weight, respectively (groups G3, G4, G5, G6 and G8, respectively; n=5 each). Immediately after the test, the aorta was removed and suspended in an organ bath. Cumulative relaxation in response to acetylcholine (10−12-10−4 M) and contraction in response to phenylephrine (10−12-10−5 M) were measured. Oxidative stress was estimated by determining malondialdehyde concentration. The percentages of aorta relaxation were significantly higher in G3 (7.9±0.20), G4 (7.8±0.29), and G5 (7.9±0.21), compared to the control group (7.2±0.04), while relaxation in the G6 (7.4±0.25) and G8 (7.0±0.06) groups was similar to the control group. In contrast, the percentage of contraction was significantly higher in G6 (8.8 ±0.1) and G8 (9.7±0.29) compared to the control (7.1±0.1), G3 (7.3±0.2), G4 (7.2±0.1) and G5 (7.2±0.2%) groups. Lipid peroxidation levels in the aorta were similar to control levels in G3, G4 and G5, but higher in G6 and G8, and significantly higher in G8 (one-way ANOVA). These results indicate a reduction in vasorelaxing activity and an increase in contractile activity in rat aortas after high-intensity exercise, followed by an increase in lipid peroxidation.

The enhanced expression of IL-17-secreting T cells during the early progression of atherosclerosis in ApoE-deficient mice fed on a western-type diet

Atherosclerosis is a chronic progressive inflammatory disorder and the leading cause of cardiovascular mortality. Here we assessed the dynamic changes of T-cell-derived cytokines, such as inteferon (IFN)-gamma, interleukin (IL)-17 and IL-4, during the progression of atherosclerosis in apolipoprotein E-null (ApoE(-/-)) mice, to understand the role of immune responses in different stages of atherosclerosis. Male ApoE(-/-) mice were fed a high-fat, western-type diet (WD: 21% lipid, 1.5% cholesterol) after 5 weeks of age and were compared with C57BL/6 wild-type control mice fed a standard chow diet. Atherosclerotic lesions appeared in the aortic sinus of ApoE(-/-) mice 4 weeks after WD and the lesions progressed and occupied >50% of the total sinus area 16 weeks after WD. Aortic IL-17 mRNA and protein expression started to increase in ApoE(-/-) mice after 4 weeks on the WD and peaked at around 8-12 weeks on the WD. In terms of systemic expression of T-cell-derived cytokines, IL-17 production from splenocytes after anti-CD3/CD28 stimuli increased from 4 weeks on the WD, peaked at 12 weeks and returned to control levels at 16 weeks. The production of IFN-gamma and IL-4 (Th1 and Th2 cytokines, respectively) from splenocytes was delayed compared with IL-17. Taken together, the present data indicate that Th17 cell response may be involved at an early stage in the development of atherosclerosis.

The enhanced expression of IL-17-secreting T cells during the early progression of atherosclerosis in ApoE-deficient mice fed on a western-type diet

Atherosclerosis is a chronic progressive inflammatory disorder and the leading cause of cardiovascular mortality. Here we assessed the dynamic changes of T-cell-derived cytokines, such as inteferon (IFN)-gamma, interleukin (IL)-17 and IL-4, during the progression of atherosclerosis in apolipoprotein E-null (ApoE(-/-)) mice, to understand the role of immune responses in different stages of atherosclerosis. Male ApoE(-/-) mice were fed a high-fat, western-type diet (WD: 21% lipid, 1.5% cholesterol) after 5 weeks of age and were compared with C57BL/6 wild-type control mice fed a standard chow diet. Atherosclerotic lesions appeared in the aortic sinus of ApoE(-/-) mice 4 weeks after WD and the lesions progressed and occupied >50% of the total sinus area 16 weeks after WD. Aortic IL-17 mRNA and protein expression started to increase in ApoE(-/-) mice after 4 weeks on the WD and peaked at around 8-12 weeks on the WD. In terms of systemic expression of T-cell-derived cytokines, IL-17 production from splenocytes after anti-CD3/CD28 stimuli increased from 4 weeks on the WD, peaked at 12 weeks and returned to control levels at 16 weeks. The production of IFN-gamma and IL-4 (Th1 and Th2 cytokines, respectively) from splenocytes was delayed compared with IL-17. Taken together, the present data indicate that Th17 cell response may be involved at an early stage in the development of atherosclerosis.

Representações sociais de educação em saúde em tempos de AIDS / Social representations of health education in a time of AIDS / Representaciones sociales de educación para la salud en la era del SIDA

Estudo qualitativo e descritivo, cujo objetivo foi identificar e analisar as representações sociais de educação em saúde à pessoa vivendo com HIV entre profissionais de saúde. Os cenários foram três serviços de atenção à DST/HIV/AIDS, em Belém-PA, Brasil, e 37 profissionais de saúde participaram da pesquisa. A coleta de dados deu-se em 2012-2013 por meio de entrevista em profundidade; a análise utilizou o software Alceste 4.10. Com base no conjunto dos resultados foi possível vislumbrar que a educação em saúde pode ser compreendida a partir de categorias: a configuração do agir educativo; as condições sine qua non: educação no trabalho e estrutura da unidade; o processo pedagógico. Conclui-se que as representações sociais configuram-se como orientação-informação para precaução-prevenção e revelam-se no movimento do agir persistente ao emergente, o que suscita uma educação em saúde permanente para se chegar à integralidade nos serviços.

This is a qualitative and descriptive study, which aimed at identifying and analyzing social representations of health education to HIV patients among health professionals. The setting included three healthcare DST/HIV/AIDS services in Belém-PA, Brazil, and 37 health professionals participated in the study. Data collection was conducted in 2012-2013 on the basis of in-depth interviews and analysis was made on Alceste 4.0 software. Final results indicated that health education can be comprehended in light of categories: educational action; sine qua non: education and training at work, and unit structure; teaching-learning process. Conclusions show that social representations are set as guidance-information for precaution-prevention and that they come forth along continuous and emerging action flow, bringing about permanent health education to ensure healthcare services in full.

Estudio cualitativo y descriptivo, que objetivó identificar y analizar las representaciones sociales de educación en salud a la persona viviendo con HIV entre profesionales de salud. Los escenarios fueron tres servicios de atendimiento al DST/HIV/ SIDA, en Belém-PA, Brasil, y 37 profesionales de salud participaran del estudio. La colecta de datos se dio en 2012-2013, por medio de entrevista en profundidad y el análisis utilizo el software Alceste 4.10. Con base en el conjunto de los resultados fue posible vislumbrar que la educación en salud puede ser comprendida a partir de categorías: la configuración del acto educativo; las condiciones sine qua non: educación en el trabajo y estructura de la unidad; el proceso pedagógico. Se concluye que las representaciones sociales se configuran como orientación-información para precaución-prevención y se revelan en el movimiento del acto persistente al emergente, lo que suscita una educación en salud permanente para llegarse a la integralidad en los servicios.

Differentiated miRNA expression and validation of signaling pathways in apoE gene knockout mice by cross-verification microarray platform

The microRNA (miRNA) regulation mechanisms associated with atherosclerosis are largely undocumented. Specific selection and efficient validation of miRNA regulation pathways involved in atherosclerosis development may be better assessed by contemporary microarray platforms applying cross-verification methodology. A screening platform was established using both miRNA and genomic microarrays. Microarray analysis was then simultaneously performed on pooled atherosclerotic aortic tissues from 10 Apolipoprotein E (apoE) knockout mice (apoE-/-) and 10 healthy C57BL/6 (B6) mice. Differentiated miRNAs were screened and cross-verified against an mRNA screen database to explore integrative mRNA-miRNA regulation. Gene set enrichment analysis was conducted to describe the potential pathways regulated by these mRNA-miRNA interactions. High-throughput data analysis of miRNA and genomic microarrays of knockout and healthy control mice revealed 75 differentially expressed miRNAs in apoE-/- mice at a threshold value of 2. The six miRNAs with the greatest differentiation expression were confirmed by real-time quantitative reverse-transcription PCR (qRT-PCR) in atherosclerotic tissues. Significantly enriched pathways, such as the type 2 diabetes mellitus pathway, were observed by a gene-set enrichment analysis. The enriched molecular pathways were confirmed through qRT-PCR evaluation by observing the presence of suppressor of cytokine signaling 3 (SOCS3) and SOCS3-related miRNAs, miR-30a, miR-30e and miR-19b. Cross-verified high-throughput microarrays are optimally accurate and effective screening methods for miRNA regulation profiles associated with atherosclerosis. The identified SOCS3 pathway is a potentially valuable target for future development of targeted miRNA therapies to control atherosclerosis development and progression.

HMG-CoA Reductase Inhibitor Improves Endothelial Dysfunction in Spontaneous Hypertensive Rats Via Down-regulation of Caveolin-1 and Activation of Endothelial Nitric Oxide Synthase

Hypertension is associated with endothelial dysfunction and increased cardiovascular risk. Caveolin-1 regulates nitric oxide (NO) signaling by modulating endothelial nitric oxide synthase (eNOS). The purpose of this study was to examine whether HMG-CoA reductase inhibitor improves impaired endothelial function of the aorta in spontaneous hypertensive rat (SHR) and to determine the underlying mechanisms involved. Eight-week-old male SHR were assigned to either a control group (CON, n=11) or a rosuvastatin group (ROS, n=12), rosuvastatin (10 mg/kg/day) administered for eight weeks. Abdominal aortic rings were prepared and responses to acetylcholine (10-9-10-4 M) were determined in vitro. To evaluate the potential role of NO and caveolin-1, we examined the plasma activity of NOx, eNOS, phosphorylated-eNOS and expression of caveolin-1. The relaxation in response to acetylcholine was significantly enhanced in ROS compared to CON. Expression of eNOS RNA was unchanged, whereas NOx level and phosphorylated-eNOS at serine-1177 was increased accompanied with depressed level of caveolin-1 in ROS. We conclude that 3-Hydroxy-3-methylglutaryl Coenzyme-A (HMG-CoA) reductase inhibitor can improve impaired endothelial dysfunction in SHR, and its underlying mechanisms are associated with increased NO production. Furthermore, HMG-CoA reductase inhibitor can activate the eNOS by phosphorylation related to decreased caveolin-1 abundance. These results imply the therapeutic strategies for the high blood pressure-associated endothelial dysfunction through modifying caveolin status.

Potential role of HMG CoA reductase inhibitor on oxidative stress induced by advanced glycation endproducts in vascular smooth muscle cells of diabetic vasculopathy

Advanced glycation endproducts (AGEs)-induced vascular smooth muscle cell (VSMCs) proliferation and formation of reactive oxygen species (ROS) are emerging as one of the important mechanisms of diabetic vasculopathy but little is known about the antioxidative action of HMG CoA reductase inhibitor (statin) on AGEs. We hypothesized that statin might reduce AGEs-induced intracellular ROS of VSMCs and analyzed the possible mechanism of action of statin in AGEs-induced cellular signaling. Aortic smooth muscle cell of Sprague-Dawley rat (RASMC) culture was done using the different levels of AGEs stimulation in the presence or absence of statin. The proliferation of RASMC, ROS formation and cellular signaling was evaluated and neointimal formation after balloon injury in diabetic rats was analyzed. Increasing concentration of AGEs stimulation was associated with increased RASMC proliferation and increased ROS formation and they were decreased with statin in a dose-dependent manner. Increased NF-kappaB p65, phosphorylated ERK, phosphorylated p38 MAPK, cyclooxygenase-2, and c-jun by AGEs stimulation were noted and their expression was inhibited by statin. Neointimal formation after balloon injury was much thicker in diabetic rats than the sham-treated group but less neointimal growth was observed in those treated with statin after balloon injury. Increased ROS formation, subsequent activation of MAPK system and increased VSMC proliferation may be possible mechanisms of diabetic vasculopathy induced by AGEs and statin may play a key role in the treatment of AGEs-induced diabetic atherosclerosis.

Menor expresión de la enzima convertidora de angiotensinaI omóloga (ECA2) se asocia a mayor remodelamiento de la pared aórtica de ratas con niveles geneticamente elevados de angiotensina II / Decreased expression of homologous angiotensin I converting enzyme (ACE2) is associated to increased remodeling of the aortic arterial wall in rats with genetically elevated levels of angiotensin II

Introducción: El polimorfismo de la enzima convertidora de angiotensina I (ECA) determina mayor actividadde ECA y niveles de angiotensina (Ang) II en ratas Brown Norway (BN) y menor actividad de ECA y niveles de Ang II en ratas Lewis (L). La interregulación entre ECA, ECA2 y su relación con remodelamiento aórtico hipertensivo no ha sido explorada Objetivo: Determinar la expresión de ECA y ECA2 y los parámetros de remodelamiento vascular hipertensivo en la aorta de ratas con niveles genéticamente determinados de ECA. Métodos: A ratas macho homocigotas de 150 grs BN y LL, se les indujo HTA por 6 semanas por el procedimiento Goldblatt (GB, 2 K-1clip). Ratas pseudo-operadas se usaron como controles (sham). Se determinó la presión arterial sistólica (PAS), el grosor de la túnica media (GTM), área de la TM (ATM), expresión génica deECA, ECA2 ,TGF-beta, PAI-1 y MCP-1 por RT-PCR y también proteica de ECA y ECA2 por Western Blot. Resultados: La masa cardiaca relativa y la PAS aumentaron significativamente en los grupos GB respecto a sus controles Sham, sin diferencias por efecto del polimorfismo de la ECA. En condiciones de normotensión las ratas BN mostraron que la pared aórtica expresa mayores niveles génicos y proteicos de la ECA(60% y 134%, respectivamente) y menores de ECA2 (74% y 73%, respectivamente) respecto de las ratas L(p<0.05). Estos resultados se asociaron con mayores GTM y ATM como en los niveles de mRNA de TGF-beta y, PAI-1 en las aortas de ratas BN respecto de las ratas L (p<0,05). En respuesta a un estrés hipertensivo las ratas con mayores niveles de ECA y menores niveles de ECA2 mostraron mayor GTM (p<0,05, respecto de GB-L), sin diferencias en los otros parámetros evaluados...

Background: Angiotensin I converting enzyme (ACE) polymorphism determines increased ACE and Ang IIlevels in Brown Norway rats (BN) and decreased ACE and Ang II levels in Lewis (L) rats. The interactionbetween ACE and ACE2 in relation to aortic remodeling associated to hypertension has not been explored. Aim: to determine the expression of ACE and ACE2 along with parameters of remodeling in hypertensive rats with genetically determined levels of ACE. Methods: BN and L rats weighing 150 g were made hypertensive by the Goldblatt procedure (GB, 2K-1 clip). Sham operated rats were used as controls. Systolic blood pressure (SBP), media thickness (MT), and MT area were measured. RT-PCR was used to determine the genetic expression of ACE, ACE2, TGF-beta, PAI-1 and MCP-1. Western Blot was used to measure the protein fraction of ACE and ACE2 Results: Relative cardiac mass and SBP increases significantly in GB rats compared to controls; ACE polymorphism did not influence this effect. The aortic wall of normotensive BN rats expressed increased genic and protein levels of ACE (60% and 134%, respectively) and decreased levels of ACE2 (74% and 73%, respectively) compared to L rats (p<0.05). These findings were associated to increased MT and MT area as well as increased mRNA for TGF-beta and PAI 1 in BN rats compared to L rats (p<0.05). In response to hypertensive stress, rats with increased ACE and decreased ACE2 levels developed increased MT compared to GB-L rats; other parameters of remodeling were not affected...

Development of a fluorescence stereomicroscope and observation of Bong-Han corpuscles inside blood vessels.

A fluorescence stereomicroscope system was developed in order to observe in situ the distribution of nuclei in intravascular Bong-Han ducts and corpuscle tissues by injecting acridine orange, which stained specifically nuclei. Intravascular Bong-Han corpuscles, connected with Bong-Han ducts could be detected in the aortas of rats, mice, and rabbits.

Expression of NAD(P)H Oxidase Subunits and Their Contribution to Cardiovascular Damage in Aldosterone/Salt-Induced Hypertensive Rat

NAD(P)H oxidase plays an important role in hypertension and its complication in aldosterone-salt rat. We questioned whether NAD(P)H oxidase subunit expression and activity are modulated by aldosterone and whether this is associated with target- organ damage. Rats were infused with aldosterone (0.75 microgram/hr/day) for 6 weeks and were given 0.9% NaCl+/-losartan (30 mg/kg/day), spironolactone (200 mg/kg/ day), and apocynin (1.5 mM/L). Aldosterone-salt hypertension was prevented completely by spironolactone and modestly by losartan and apocynin. Aldosterone increased aortic NAD(P)H oxidase activity by 34% and spironolactone and losartan inhibited the activity. Aortic expression of the subunits p47(phox), gp91(phox), and p22(phox) increased in aldosterone-infused rats by 5.5, 4.7, and 3.2-fold, respectively, which was decreased completely by spironolactone and partially by losartan and apocynin. Therefore, the increased expression of NAD(P)H oxidase may contribute to cardiovascular damage in aldosterone-salt hypertension through the increased expression of each subunit.

La vía transduccional Rho A y Rho kinasa está activada en la pared arterial de ratas con niveles genéticamente elevados de angiotensina II / Transductional Rho A and Rho kynase pathways are activated in the arterial wall of rats with genetically elevated levels of angiotensin II

Antecedentes: El polimorfismo de la enzima convertidora de angiotensina I (ECA) determina mayor actividad de ECA yniveles de angiotensina (Ang) II en ratas Brown Norway (BN) y menor actividad de ECA y niveles de Ang II en ratas Lewis (L). La relación entre niveles de Ang II y la vía transduccional Rho A/Rho kinasa no ha sido explorada.Objetivo: Determinar la participación de la vía Rho A/ Rho kinasa mediante la fosforilación de la proteína blanco 1 de la fosfatasa de la miosina (MYPT1) vascular en ratas con niveles genéticamente bajos (L) y altos de Ang II (BN) y su relación con la expresión de algunos genes que determinan remodelado de la pared arterial: el gen del activador de plasminógeno 1 (PAI-1), el gen de la proteína quimioatractante de monocitos (MCP-1) y el gen del factor de transformación beta 1 (TGF- b1) en la pared arterial. Métodos: Se usaron ratas machos homocigotos de 150 grs BN y L. Para inhibir la vía Rho A / Rho kinasa, se administró fasudil (100 mg/Kg/día por gavage) a ratas BN, durante 7 días. Se determinó la presión arterial sistólica (PAS), la expresión vascular (en la aorta) de MYPT1 total (MYPT1-T) y MYPT1 fosforilada (MYPT1-P) y la relación entre ambas por Western blot además de la expresión génica de PAI-1, MCP-1 y de TGF-b1 por RT-PCR (en unidades de densidad óptica). Resultados como promedio(ES): El antagonista del receptor tipo I de angiotensina II Candesartán por 7 días (10 mg/Kg/d, n = 8) redujo en un 50 por ciento los niveles elevados de MYPT1-P/T en las ratas BN (p <0.05) sin modificar la mayor expresión de los genes evaluados. Conclusión: La vía Rho A/ Rho kinasa se encuentra activada en la pared arterial de ratas con niveles elevados de Ang II (BN) y causa mayor expresión génica de PAI-1 y de MCP-1 ya que la sobre expresión de ambos tiende a normalizarse con fasudil. La mayor expresión génica de TGF-b1 en la aorta en ratas BN parece no estar relacionada con la activación de de Rho A/ Rho kinasa vía Angiotensina II ya que no...

A study on immunopathogenetic mechanisms of atherosclerotic process caused by chronic infection of Chlamydia pneumoniae in rats (Ratus novergicus).

AIM: this study was aimed to determine the correlations between duration of Chlamydia pneumoniae infection and the development of atherosclerotic process in white-rats' (Ratus novergicus) aorta. METHODS: this is an experimental study which examined the expression of TNFa, IL-1b, IL-8, adhesion molecule of VCAM-1 and the development of foam cells associated with atherosclerotic process in white-rats' aorta. There were 32 male rats, +/- 6 weeks of age, divided into 4 groups: control group (K) without infection, and 3 groups with infection through nasal and oral inoculation of Chlamydia pneumoniae by single dose of 5 x 105 in amount of 35 microl. The first group (P1) was preserved for 5(1/2) months period, the second group (P2) was preserved for 7(1/2) months and the third group (P3) was preserved for 9(1/2) months. At the end of study, histological slides were made from aortic tissues in order to study the development of atherosclerotic process by examining foam cells and cytokines expression of TNFa, IL-1b, IL-8 and VCAM-1. Foam cells examination was performed by Hematoxcillin-eosin staining, while indirect immunohistochemistry staining was used to examine the expression of TNFa, IL-1b, IL-8 and VCAM-1. Afterward, the amount of foam cells and cytokines expression was measured. The study result was analyzed by ANOVA. RESULTS: there was increased expression of TNFa, IL-1b, IL-8, VCAM-1and increased foam cells formation (extended atherosclerosis area) in the aortic tissues infected by Chlamydia pneumoniae (5(1/2) months, 7(1/2) months and 9(1/2) months), which was significantly different compared to the control group. The result of ANOVA revealed that the most important factor in tissue injury is foam cells development induced by VCAM-1 and IL-8 in all of phases (characterized by most abundant neutrophil infiltration). It indicated the infection caused by extracellular pathogenic agent, which established the fatty streak (acute phase) in 5(1/2) months period. In the group with 7(1/2) months infection period, TNFa also had important roles (characterized by increased monocytes and lymphocytes infiltration), indicating that there was negative-gram pathogenic agent with intracellular infection, which caused a progressive atherosclerotic process, and development of fibrosis / atherosclerotic plaque (sub acute phase). In 9(1/2) months infection period, there was large thrombus containing a lot of leukocytes in the aorta (chronic phase). CONCLUSION: based on the result of this study, it may be concluded that Chlamydia pneumoniae may cause atherosclerotic process in aorta. Extracellular infection of Chlamydia pneumoniae occurs in all of phases and intracellular infection begins in sub-acute phase. On 5(1/2) months period, fatty streak is developed (acute phase); on 7(1/2) months period, there is atherosclerotic plaque (sub acute phase); and on 9(1/2) period, there is large thrombus containing a lot of leukocytes (a progressive chronic phase).

Hypolipidaemic efficacy of Capparis decidua fruit and shoot extracts in cholesterol fed rabbits.

High fat diet caused significant (8-fold) increase in serum total cholesterol in rabbits. Administration of C. decidua fruit extract (50% ethanolic) at the dose of 500 mg/kg body weight significantly reduced serum total cholesterol (61%), LDL cholesterol (71%), triglycerides (32%) and phospholipids (25%). Similarly C. decidua shoot extract lowered serum total cholesterol (48%), LDL cholesterol (57%), triglycerides (38%) and phospholipids (36%).The cholesterol content of aorta was decreased by 44 and 28% in fruit and shoot extract treatment respectively. The HDL to total cholesterol ratio and atherogenic index was significantly decreased in plant extract treated groups suggesting antiatherosclerotic nature of these plant extract. These results reveal the hypolipidaemic potential of C. decidua fruit and shoot.

Role of endothelin-1 in genesis of coronary artery disease.

BACKGROUND: The endothelial cells produce the most potent vasoconstrictor known as endothelin-1. Elevated plasma levels of endothelin have been associated with coronary artery disease, essential hypertension and heart failure. The aims of the present study were, to compare the plasma endothelin-1 levels in coronary artery disease patients and healthy controls, to confirm endothelin-1 as surrogate marker for coronary artery disease and to compare the presence of endothelin-1 like immunoreactivity in aortic and internal mammary artery specimens obtained during coronary artery bypass graft surgery. METHODS AND RESULTS: The circulating levels of endothelin-1 were determined by enzyme-linked immunoassay in patients of coronary artery disease (n=145) and compared with healthy controls (n=70). Tissue endothelin-1 immunoreactivity was examined by immunohistochemical method in aortic and internal mammary artery tissue specimens obtained from 20 patients of coronary artery disease during coronary artery bypass grafting to understand the role of endothelin in atherosclerosis. Significantly higher levels (p < 0.001) of endothelin-1 were observed in all patients of coronary artery disease as compared to healthy controls. The immunoreactivity of endothelin-1 was localized to endothelial cell layer in internal mammary artery whereas in aortic specimens, in addition to endothelial cell layer, immunoreactivity was seen in the cytoplasm of smooth muscle cells of intima and media. CONCLUSIONS: The significant increase in plasma endothelin-1 in coronary artery disease cases as compared to healthy subjects and presence of tissue endothelin-1 immunoreactivity in smooth muscle cells of intimal as well as medial layers of aorta confirms the role of endothelin-1 as a surrogate marker of atherosclerosis.

Proteoglycans production by aortic vascular smooth muscle cells from hypertensive rats

Remodeling of large and small arteries contributes to the development and complications of hypertension. Artery structural changes in chronic sustained hypertension include vascular smooth muscle cells (VSMC) proliferation and extracellular matrix (ECM) modifications. Extracellular constituents such as proteoglycans (PGs), may modulate vascular stiffness and VSMC growth and differentiation. We examined the effect of growth factors on secreted and membrane-bound PGs synthesis by cultured aortic smooth muscle cells (SMC) from 12- to 14- week-old spontaneously hypertensive rats (SHR) and age-matched Wistar rats. After stimulation with platelet-derived growth factor (PDGF-BB), 10% fetal calf serum (FCS) or 0.1% FCS as control, PGs synthesis (dpm/ng DNA) was evaluated in the medium (M-ECM) and in the cell layer (P-ECM) by a double-isotopic label method using both [3H]-glucosamine and [35S]-sodium sulfate which are incorporated into all complex carbohydrates or only into sulfated dysaccharides, respectively. Data are presented as percent of the control (0.1% FCS). SHR VSMC displayed a significantly greater synthesis of M-ECM [3H]-PGs than Wistar rat cells, with both treatments, but no differences in M-ECM [35S] uptake were found in any case. In the P-ECM, both PDGF-BB and 10% FCS produced a greater effect on [3H]-PGs and sulfated PGs synthesis in VSMC from SHR. An important change seen in SHR cells was a significant decreased sulfation, assessed by [35S]/[3H] ratio, in basal and stimulation conditions. Present results indicate the existence of changes in PGS synthesis and modulation in VSMC from a conduit-artery of SHR and support the pathophysiological role proposed for matrix proteoglycans in the vascular wall changes associated to hypertension and related vascular diseases as atherosclerosis.

Enalapril and Verapamil attenuate the aortic wall remodeling in nitric oxide deficiency

Cuatro grupos de 10 ratas cada uno, adultas, machos (Wistar) se mantuvieron vivas por 40 días, divididas en: Control, L-NAME, L-NAME + Enalapril, L-NAME + Verapamil. Tres parámetros de la pared de la aorta fueron analizados: el QA(SMN) (número de núcleos de células musculares lisas de la túnica media por área), la SV(lamellae) (densidad de superficie de las lamelas de la aorta), y AWT ( grosor de la pared de la aorta). Estos tres parámetros mostraron la misma tendencia entre los grupos. Las ratas del grupo L-NAME tuvieron elevación de la presión arterial e hipertrofia cardiaca. El aumento de AWT, o la reducción del QA(SMN) y la SV(lamellae) en las ratas L-NAME no fue completamente prevenido por la administración de enalapril o verapamil. La hipertensión causó aumento del AWT por hipertrofia de las unidades lamelares de la túnica media (SV(lamellae) disminuyó) y la rarefacción de los núcleos de músculo liso (QA(SMN)) disminuyó. El número relativo de núcleos de células musculares lisas (SMC) en ratas L-NAME no fue confirmado cuando este número fue corregido por el área de la pared arterial. La hipótesis de intensa proliferación de SMC en animales con deficiencia de ON, no está confirmada con la dosis de L-NAME usada y la duración del tratamiento de esta investigación.

Efecto de la coartación aórtica en la producción de óxido nítrico por el endotelio vascular / Effect of aortic coarctation on nitric oxide production by vascular endothelium

El óxido nítrico es un potente vasodilatador liberado de endotelio, que se asocia a un efecto regulador del tono vascular y de la presión arterial. En este estudio se evaluó la reactividad vascular a fenilefrina y acetilcolina de anillos de aorta de ratas control e hipertensas por coartación de la aorta, y la liberación de óxido nítrico de los anillos arteriales, en condiciones basales y después de incubarlos con acetilcolina. La reactividad vascular se evalúo en los segmentos superior e inferior a la coartación de la aorta. La vasoconstricción producida por fenilefrina fue mayor en los anillos del segemento superior de las ratas hipertensas en comparación a las ratas control; los anillos de los segmentos inferiores de ambos grupos, no mostraron diferencias significativas. La relajación por acetilcolina mostró una CE50 mayor en los segmentos superiores de las ratas hipertensas; los segmentos inferiores de ambos grupos no mostraron diferencias significativas. Los anillos de aorta de ratas hipertensas tuvieron mayor producción de óxido nítrico en comparación al grupo control, siendo significativamente mayor la producción de los segmentos inferiores, lo cual sugiere que el "shear stress" de estos segmentos estimuló la producción de óxido nítrico

Role of endothelium in angiotensin II formation by the rat aorta and mesenteric arterial bed

We investigated the angiotensin II (Ang II)-generating system by analyzing the vasoconstrictor effect of Ang II, angiotensin I (Ang I), and tetradecapeptide (TDP) renin substrate in the abscence and presence of inhibitors of the renin-angiotensin system in isolated rat aortic rings and mesenteric arterial beds with and without functional endothelium. Ang II, Ang I, and TDP elicited a dose-dependent vasoconstrictor effect in both vascular preparations that was completely blocked by the Ang II receptor antagonist saralasin (50 nM). The angiotensin converting enzyme (ACE) inhibitor captopril (36 muM) completely inhibited the vasoconstrictor effect elicited by Ang I and TDP in aortic rings without affecting that of Ang II. In contrast, captopril (36 muM) significantly reduced (80-90 percent) the response to bolus injection of Ang I, without affecting those to Ang II and TDP in mesenteric arteries. Mechanical removal of the endothelium greatly potentiated (70-95 percent) the vasoconstrictor response to Ang II, Ang I, and TDP in aortic rings while these responses were unaffected by the removal of the endothelium of mesenteric arteries with sodium deoxycholate infusion. In addition, endothelium disruption did not change the pattern of response elicited by these peptides in the presence of captopril. These findings indicate that the endothelium may not be essential for Ang II formation in rat mesenteric arteries and aorta, but it may modulate the response to Ang II. Although Ang II formation from Ang I is essentially dependent on ACE in both vessels, our results suggest the existence of an alternative pathway in the mesenteric arterial bed that may play an important role in Ang II generation from TDP in resistence but not in large vessels during ACE inhibition.

The effect of anti-hypertensive drugs on DNA synthesis and proliferation of ultured rat aortic smooth muscle cells

The aim of this study was to elucidate the effects of anti-hypertensive drugs, nifedipine, furosemide, hydrochlorothiazide, captopril, and atenolol on DNA synthesis and proliferation of cultured rat aortic smooth muscle cells induced by fetal calf serum. Aortic smooth muscle cells from Sprague-Dawley rats were isolated, cultured, and seeded in multi-well plates. When confluent, cells were cultured in a conditioned medium without fetal calf serum. After 72 hours, cells were cultured in the medium retaining 10% fetal calf serum with or without anti-hypertensive drugs by increasing the concentration between 10(-8) and 10(-4) M. DNA synthesis was assessed by [3H]-thymidine uptake and proliferation by cell numbers using a hemocytometer. Nifedipine at a concentration of 10(-5) M and 5 x 10(-5) M inhibited serum-induced DNA synthesis significantly by 50.8% and 86.6%, respectively (p < 0.05). The results of cell numbers paralleled those of 3H-thymidine incorporation. Serum-induced DNA synthesis was also reduced by 32.6% at the highest dose of furosemide (10(-4) M), but there was no statistical significance. Hydrochlorothiazide, captopril, and atenolol did not show anti-proliferative effect throughout any of the doses. In conclusion, among the various anti-hypertensive drugs, nifedipine seems to be most beneficial in view of its direct inhibitory effect on DNA synthesis and proliferation of smooth muscle cells, as well as for its anti-hypertensive effect.