Marcia K. Koike

Omega-6 polyunsaturated fatty acids prevent atherosclerosis development in LDLr-KO mice, in spite of displaying a pro-inflammatory profile similar to trans fatty acids

The development of atherosclerosis and the inflammatory response were investigated in LDLr-KO mice on three high-fat diets (40% energy as fat) for 16 weeks: trans (TRANS), saturated (SAFA) or ω-6 polyunsaturated (PUFA) fats. The following parameters were measured: plasma lipids, aortic root total cholesterol (TC), lesion area (Oil Red-O), ABCA1 content and macrophage infiltration (immunohistochemistry), collagen content (Picrosirius-red) and co-localization of ABCA1 and macrophage (confocal microscopy) besides the plasma inflammatory markers (IL-6, TNF-α) and the macrophage inflammatory response to lipopolysaccharide from Escherichia coli (LPS). As expected, plasma TC and TG concentrations were lower on the PUFA diet than on TRANS or SAFA diets. Aortic intima macrophage infiltration, ABCA1 content, and lesion area on PUFA group were lower compared to TRANS and SAFA groups. Macrophages and ABCA1 markers did not co-localize in the atherosclerotic plaque, suggesting that different cell types were responsible for the ABCA1 expression in plaques. Compared to PUFA, TRANS and SAFA presented higher collagen content and necrotic cores in atherosclerotic plaques. In the artery wall, TC was lower on PUFA compared to TRANS group; free cholesterol was lower on PUFA compared to TRANS and SAFA; cholesteryl ester concentration did not vary amongst the groups. Plasma TNF-α concentration on PUFA and TRANS-fed mice was higher compared to SAFA. No difference was observed in IL-6 concentration amongst groups. Regarding the macrophage inflammatory response to LPS, TRANS and PUFA presented higher culture medium concentrations of IL-6 and TNF-α as compared to SAFA. The PUFA group showed the lowest amount of the anti-inflammatory marker IL-10 compared to TRANS and SAFA groups. In conclusion, PUFA intake prevented atherogenesis, even in a pro-inflammatory condition.

Aortic valve assessment. Anatomical study of 100 healthy human hearts

PURPOSE To assess anatomical characteristics of the aortic valve, so that they may be useful in diagnostic situations and surgical treatment. METHODS The study analyzed 100 healthy fixed human hearts; 84% of them obtained from males, 61% of them from Caucasian individuals. The ages of the individuals ranged from 9 to 86 years (mean 30 +/- 15.5 years). The characteristics assessed related to age, sex, and race were the following: number and height of the cusps, size of the lunulae, internal and external intercommissural distance, position of the coronary ostium in relation to the aortic valve, position of the ventricular septum in relation to the aortic valve, thickness of the cusps. RESULTS All hearts assessed had a tricuspidal aortic valve. In regard to the height of the cusps and size of the lunula, the left coronary cusp was larger, followed by the right coronary cusp and the noncoronary cusp. The internal and external intercommissural distances had mean values of 24.6 +/- 5.7 mm and 19.7 +/- 7 mm, respectively. In regard to the position of the coronary ostia, in one heart two ostia emerged from the left coronary sinus, and in another, the ostium was supracommissural. The mean diameter of the aorta was 21.8 +/- 3.6 mm, and there were no significant sexual or racial differences, but the diameter increased progressively with the increase in age. The thickness of the cusps did not show any significant difference in the 3 points assessed. CONCLUSION The aortic valve annulus did not show a perfect circumference, with some variations in the measurements of the annulus, in the cusps and in the relation with the ventricular septum.

Dietary phytosterol does not accumulate in the arterial wall and prevents atherosclerosis of LDLr-KO mice

SCOPE There have been conflicting reports on the usefulness of phytosterols (PS) in preventing atherosclerosis. We evaluated the effects of dietary PS supplementation in LDLr-KO male mice on the plasma and aorta sterol concentrations and on atherosclerotic lesion development. METHODS AND RESULTS Mice were fed a high fat diet (40% of energy) supplemented with or without PS (2% w/w, n = 10). Plasma and arterial wall cholesterol and PS concentrations, lesion area, macrophage infiltration, and mRNA expression from LOX-1, CD36, ABCA1 and ABCG1 in peritoneal macrophages were measured. After 16 weeks, the plasma cholesterol concentration in PS mice was lower than that in the controls (p = 0.02) and in the arterial wall (p = 0.03). Plasma PS concentrations were higher in PS-fed animals than in controls (p < 0.0001); however, the arterial wall PS concentration did not differ between groups. The atherosclerotic lesion area in the PS group (n = 5) was smaller than that in controls (p = 0.0062) and the macrophage area (p = 0.0007). PS correlates negatively with arterial lipid content and macrophage (r = -0.76; p < 0.05). PS supplementation induced lower ABCG1 mRNA expression (p < 0.05). CONCLUSIONS Despite inducing an increase in PS plasma concentration, PS supplementation is not associated with its accumulation in the arterial wall and prevents atherosclerotic lesion development.

Low coronary driving pressure is associated with subendocardial remodelling and left ventricular dysfunction in aortocaval fistula.

1 The role of haemodynamic changes in left ventricular remodelling has been poorly investigated, especially in the context of volume overload cardiac hypertrophy. Low diastolic blood pressure and high left ventricular filling pressure are expected to affect coronary driving pressure negatively and thereby put in jeopardy subendocardial perfusion in particular. The consequences to global left ventricular remodelling remain undetermined. The aim of the present study was to investigate the role of coronary driving pressure in the development of subendocardial remodelling and the conceivable effects on cardiac function, using a rat model of aortocaval fistula. 2 Wistar rats, weighing 330–350 g, were submitted to aortocaval fistula (ACF group) or sham (control group) operations. Two haemodynamic measurements were determined following surgery, the initial measurement at week 1 and the final measurement at week 8. Cytokine expression, myeloperoxidase (MPO) activity, metalloproteinase expression and activity and fibrosis were assessed in two distinct left ventricular myocardial layers: the subendocardium (SE) and the non‐subendocardium (non‐SE). 3 The ACF group showed lower initial and final coronary driving pressure and lower final +dP/dt and –dP/dt compared with the control group. Multivariate analyses disclosed initial coronary driving pressure as the only haemodynamic parameter independently associated with SE fibrosis (R2 = 0.76; P < 0.0001) and with +dP/dt (R2 = 0.55; P = 0.0004) and –dP/dt (R2 = 0.91; P < 0.0001). Matrix metalloproteinase (MMP)‐2 expression and activity predominated in the SE of ACF animals, particularly in those with low coronary driving pressure. Increased levels of interleukin (IL)‐6 and IL‐1β also predominated in the SE of the ACF group. Otherwise, MPO activity and levels of tumour necrosis factor‐α and IL‐10 were similar in both groups. Final coronary driving pressure correlated with both the expression and activity of MMP‐2. 4 Low coronary driving pressure early in the course of ACF determines SE damage and, by this mechanism, interferes negatively in left ventricular function.

RESETTING OF AORTIC BARORECEPTORS IN RESPONSE TO HYPOTENSION DOES NOT ALTER GAIN SENSITIVITY

1 In chronic hypertension, the baroreceptors reset to hypertensive levels with a decrease in gain sensitivity, but only a few studies have evaluated baroreceptor resetting during chronic hypotension and, under these conditions, no consistent information is available concerning changes in baroreceptor gain sensitivity. Therefore, in the present study, the aortic baroreceptor function curve and the baroreflex control of heart rate (HR) were evaluated in chronic hypotension produced by myocardial infarction (MI) with no heart failure. 2 Aortic baroreceptor function curves were studied in anaesthetized three groups of rats: (i) MI‐7, six rats 7 days after MI; (ii) MI‐30, nine rats 30 days after MI; and (iii) five control animals (SHAM). The pressure–nerve activity relationship was measured during rapid changes in blood pressure by integrating the whole‐nerve activity of the baroreceptors in a computerized beat‐to‐beat analysis. 3 Both long‐term periods (7 or 30 days) of hypotension were accompanied by complete resetting of the baroreceptor in rats (the leftward displacement of the baroreceptor curve matched the decrease in blood pressure). Moreover, the resetting of the baroreceptor function curve was not accompanied by changes in gain sensitivity (1.47, 1.64 and 1.67%/mmHg for SHAM, MI‐7 and MI‐30 groups, respectively) and the baroreflex control of HR was normal comparing SHAM and MI‐30 groups (bradycardic 1.62 ± 0.18 vs 1.99 ± 0.52 b.p.m./mmHg, respectively; tachycardic 3.6 ± 0.5 vs 4.1 ± 0.4 b.p.m./mmHg for, respectively). 4 The data indicate that the resetting of baroreceptors in chronic hypotension is stable and is not accompanied by changes in gain sensitivity, as observed in hypertension. This may account for the normal baroreflex control of HR observed in non‐anaesthetized rats.

Caloric restriction protects livers from ischemia/reperfusion damage by preventing Ca2+-induced mitochondrial permeability transition

Abstract Caloric restriction (CR) promotes lifespan extension and protects against many pathological conditions, including ischemia/reperfusion injury to the brain, heart and kidney. In the liver, ischemia/reperfusion damage is related to excessive mitochondrial Ca2+ accumulation, leading to the mitochondrial permeability transition. Indeed, liver mitochondria isolated from animals maintained on CR for 4 months were protected against permeability transition and capable of taking up Ca2+ at faster rates and in larger quantities. These changes were not related to modifications in mitochondrial respiratory activity, but rather to a higher proportion of ATP relative to ADP in CR liver mitochondria. Accordingly, both depletion of mitochondrial adenine nucleotides and loading mitochondria with exogenous ATP abolished the differences between CR and ad libitum (AL) fed groups. The prevention against permeability transition promoted by CR strongly protected against in vivo liver damage induced by ischemia/reperfusion. Overall, our results show that CR strongly protects the liver against ischemia/reperfusion and uncover a mechanism for this protection, through a yet undescribed diet‐induced change in liver mitochondrial Ca2+ handling related to elevated intramitochondrial ATP. Graphical abstract Figure. No caption available. HighlightsCaloric restriction protects liver mitochondria against Ca2+ induced permeability transition.Mitochondrial Ca2+ uptake rates are also increased by the intervention.Both of these effects seems to be mediated by elevated ATP/ADP ratios.Decreased susceptibility to permeability transition induced by CR protects mouse livers against ischemia/reperfusion injury.

Effect of low intensity laser therapy in an experimental model of cranio-encephalic trauma in rats

The aim of this study was to analyze the effects of phototherapy with low intensity laser on the inflammatory reaction after rat brain injury. Cryogenic injury was performed at the brain of 16 male Wistar rats (250-300g) using a cooper probe at -80º C. Immediately, 24 h and 48 h later, the rats received laser irradiation using a GaAlAs laser (830 nm, 100 mW). The samples were randomly divided into four groups (n= 4 per group): A: control (non- irradiated); B: energy density of 14.28 J/cm2; C: 28.57 J/cm2; D: 42.85 J/cm2. Three days later, the cerebral vascular permeability and the inflammatory cells at the trauma site were evaluated. For vascular permeability analysis, 2 h prior sacrifice an intra vascular injection of Evans blue stain was done in the rats. For inflammatory cells counting, frozen samples were sectioned and the histological slides were stained with Giemsa. The data were compared by either ANOVA or Kruskall-Wallis complemented by the Dunns test. The irradiated groups presented higher cerebral vascular permeability than controls (A: 2.6 ± 0.8; B:12.0 ± 2.0; C: 13.1 ± 4.1, and D: 12.4 ± 1.8; p=0.016). The inflammatory cell numbers of irradiated samples were similar to controls (A: 65 ± 6; B:85 ± 9; C: 84 ±14, and D: 83 ± 3; p=0.443). The data showed that phototherapy with low intensity laser modulates the inflammatory reaction in the brain by increasing the cerebral vascular permeability after a cryogenic trauma.

Acute aortocaval fistula: role of low perfusion pressure and subendocardial remodeling on left ventricular function.

The experimental model of aortocaval fistula is a useful model of cardiac hypertrophy in response to volume overload. In the present study it has been used to investigate the pathologic subendocardial remodeling associated with the development of heart failure during the early phases (day 1, 3, and 7) following volume overload. Compared with sham treated rats, aortocaval fistula rats showed lower systemic blood pressure and higher left ventricular end‐diastolic pressure This resulted in lower coronary driving pressure and left ventricular systolic and diastolic dysfunction. Signs of myocyte necrosis, leukocyte cell infiltration, fibroplasia and collagen deposition appeared sequentially in the subendocardium where remodeling was more prominent than in the non‐subendocardium. Accordingly, increased levels of TNF‐alpha, IL‐1 beta, and IL‐6, and enhanced MMP‐2 activity were all found in the subendocardium of rats with coronary driving pressure ≤60 mmHg. The coronary driving pressure was inversely correlated with MMP‐2 activity in subendocardium in all time‐points studied, and blood flow in this region showed positive correlation with systolic and diastolic function at day 7. Thus the predominant subendocardial remodeling that occurs in response to low myocardial perfusion pressure during the acute phases of aortocaval fistula contributes to early left ventricular dysfunction.

Effects of terlipressin and naloxone compared with epinephrine in a rat model of asphyxia-induced cardiac arrest

OBJECTIVE: To evaluate the hemodynamic and metabolic effects of terlipressin and naloxone in cardiac arrest. METHODS: Cardiac arrest in rats was induced by asphyxia and maintained for 3.5 minutes. Animals were then resuscitated and randomized into one of six groups: placebo (n = 7), epinephrine (0.02 mg/kg; n = 7), naloxone (1 mg/kg; n = 7) or terlipressin, of which three different doses were tested: 50 µg/kg (TP50; n = 7), 100 µg/kg (TP100; n = 7) and 150 µg/kg (TP150; n = 7). Hemodynamic variables were measured at baseline and at 10 (T10), 20 (T20), 30 (T30), 45 (T45) and 60 (T60) minutes after cardiac arrest. Arterial blood samples were collected at T10, T30 and T60. RESULTS: The mean arterial pressure values in the TP50 group were higher than those in the epinephrine group at T10 (165 vs. 112 mmHg), T20 (160 vs. 82 mmHg), T30 (143 vs. 66 mmHg), T45 (119 vs. 67 mmHg) and T60 (96 vs. 66.8 mmHg). The blood lactate level was lower in the naloxone group than in the epinephrine group at T10 (5.15 vs. 10.5 mmol/L), T30 (2.57 vs. 5.24 mmol/L) and T60 (2.1 vs. 4.1 mmol/L). CONCLUSIONS: In this rat model of asphyxia-induced cardiac arrest, terlipressin and naloxone were effective vasopressors in cardiopulmonary resuscitation and presented better metabolic profiles than epinephrine. Terlipressin provided better hemodynamic stability than epinephrine.

Induction and maintenance of in vivo ventricular fibrillation in rabbits

AIM To provide new sustainable in vivo models of ventricular fibrillation in rabbits. METHODS New Zealand rabbits were submitted to anaesthesia and mechanical ventilation, after which ventricular fibrillation was induced through electrical stimulation (for 2min at 100Hz, with 2-ms pulses, 10mA, and 10V) directly to the heart. To that end, the animals were divided into two groups: right ventricle (n=11) and left ventricle (n=11). In group right ventricle, the thoracic cavity was exposed, and a catheter was introduced into the right ventricle via the right jugular vein. In group left ventricle, the thorax remained closed, and the catheter was introduced into the left ventricle via the left common carotid artery (cervical access). RESULTS Sustained ventricular fibrillation was achieved in 100% of group right ventricle rabbits (n=11) and in 82% of group left ventricle rabbits (n=9). CONCLUSION Both models proved appropriate for achieving sustained ventricular fibrillation. However, in view of the invasiveness of the procedure adopted in group right ventricle, the experimental conditions used in group left ventricle seemed more physiological and more effective in inducing sustained ventricular fibrillation.