Chronic Cadmium Exposure Accelerates the Development of Atherosclerosis and Induces Vascular Dysfunction in the Aorta of ApoE-/- Mice.

Cadmium exposure is related to cardiovascular diseases, including hypertension, atherosclerosis, increased oxidative stress, endothelial dysfunction, and specific biochemical changes induced by this metal. Thus, we aimed to investigate whether cadmium exposure induces endothelial dysfunction, accelerates atherosclerotic plaque formation in the aorta, and enhances oxidative stress in apolipoprotein E knockout (ApoE-/-) mice. Experiments were performed in 14-week-old male wild-type and ApoE-/- mice. ApoE-/- mice received cadmium (CdCl2 100 mg/L in drinking water for 28 days) or vehicle (distilled water). After treatment, vascular reactivity to phenylephrine, acetylcholine, and sodium nitroprusside was analyzed using isolated aorta. Bone marrow cells were isolated to assess the production of nitric oxide and reactive oxygen and nitrogen species. ApoE-/- cadmium-treated mice had higher cholesterol levels than non-exposed mice. Cadmium exposure decreased the vasodilatation response to acetylcholine in aortic ring of ApoE-/- mice, though no changes in phenylephrine or sodium nitroprusside responses were observed. L-NAME reduced vasodilator responses to acetylcholine; this effect was lower in ApoE-/- cadmium-treated mice, suggesting reduction in nitric oxide (NO) bioavailability. Moreover, in bone marrow cells, cadmium decreased cytoplasmic levels of NO and increased superoxide anions, hydrogen peroxide, and peroxynitrite in ApoE-/- mice. Morphological analysis showed that cadmium exposure increased plaque deposition in the aorta by approximately 3-fold. Our results suggest that cadmium exposure induces endothelial dysfunction in ApoE-/- mice. Moreover, cadmium increased total cholesterol levels, which may promote the early development of atherosclerosis in the aorta of ApoE-/- mice. Our findings support the hypothesis that cadmium exposure might increase the risk of atherosclerosis.

Computational engineering of cellulase Cel9A-68 functional motions through mutations in its linker region.

Microbial cellulosic degradation by cellulases has become a complementary approach for biofuel production. However, its efficiency is hindered by the recalcitrance of cellulose fibres. In this context, computational protein design methods may offer an efficient way to obtain variants with improved enzymatic activity. Cel9A-68 is a cellulase from Thermobifida fusca that is still active at high temperatures. In a previous work, we described a collective bending motion, which governs the overall cellulase dynamics. This movement promotes the approximation of its CBM and CD structural domains (that are connected by a flexible linker). We have identified two residues (G460 and P461) located at the linker that act as a hinge point. Herein, we applied a new level of protein design, focusing on the modulation of this collective motion to obtain cellulase variants with enhanced functional dynamics. We probed whether specific linker mutations would affect Cel9A-68 dynamics through computational simulations. We assumed that P461G and G460+ (with an extra glycine) constructs would present enhanced interdomain motions, while the G460P mutant would be rigid. From our results, the P461G mutation resulted in a broader exploration of the conformational space, as confirmed by clustering and free energy analyses. The WT enzyme was the most rigid system. However, G460P and P460+ explored distinct conformational states described by opposite directions of low-frequency normal modes; they sampled preferentially closed and open conformations, respectively. Overall, we highlight two significant findings: (i) all mutants explored larger conformational spaces than the WT; (ii) the selection of distinct conformational populations was intimately associated with the mutation considered. Thus, the engineering of Cel9A-68 motions through linker mutations may constitute an efficient way to improve cellulase activity, facilitating the disruption of cellulose fibres.

Deleterious effect of hypothermia in myocardial protection against cold ischemia: a comparative study in isolated rat hearts.

BACKGROUND: There is a growing need to improve heart preservation benefit the performance of cardiac operations, decrease morbidity, and more important, increase the donor pool. Therefore, the objective of this study was to evaluate the cardioprotective effects of Krebs-Henseleit buffer (KHB), Bretschneider-HTK (HTK), St. Thomas No. 1 (STH-1), and Celsior (CEL) solutions infused at 10°C and 20°C. METHODS: Hearts isolated from male albino Wistar rats and prepared according to Langendorff were randomly divided equally into 8 groups according to the temperature of infusion (10°C or 20°C) and cardioprotective solutions (KHB, HTK, STH-1, and CEL). After stabilization with KHB at 37°C, baseline values were collected (control) for heart rate (HR), left ventricle systolic pressure (LVSP), coronary flow (CF), maximum rate of rise of left ventricular pressure during ventricular contraction (+dP/dt) and maximum rate of fall of left ventricular pressure during left ventricular relaxation (-dP/dt). The hearts were then perfused with cardioprotective solutions for 5 minutes and kept for 2 hours in static ischemia at 20°C. Data evaluation used analysis of variance (ANOVA) in all together randomized 2-way ANOVA and Tukey's test for multiple comparisons. The level of significance chosen was P < .05. RESULTS: We observed that all 4 solutions were able to recover HR, independent of temperature. Interestingly, STH-1 solution at 20°C showed HR above baseline throughout the experiment. An evaluation of the corresponding hemodynamic values (LVSP, +dP/dt, and -dP/dt) indicated that treatment with CEL solution was superior at both temperatures compared with the other solutions, and had better performance at 20°C. When analyzing performance on CF maintenance, we observed that it was temperature dependent. However, when applying both HTK and CEL, at 10°C and 20°C respectively, indicated better protection against development of tissue edema. Multiple comparisons between treatments and hemodynamic variable outcomes showed that using CEL solution resulted in significant improvement compared with the other solutions at both temperatures. CONCLUSION: The solutions investigated were not able to fully suppress the deleterious effects of ischemia and reperfusion of the heart. However, these results allow us to conclude that temperature and the cardioprotective solution are interdependent as far as myocardial protection. Although CEL solution is the best for in myocardial protection, more studies are needed to understand the interaction between temperature and perfusion solution used. This will lead to development of better and more efficient cardioprotective methods.

Echocardiographic and Doppler assessment of maternal cardiovascular function in normal and abnormal canine pregnancies.

The objective was to verify maternal hemodynamic differences between normal and abnormal pregnancies in dogs. Brucella-negative pregnant bitches (n = 31) were retrospectively classified into abnormal (which had either their pregnancy interrupted between Days 52 and 60 or perinatal death of more than 50% of the litter; n = 14) and normal (which had delivered healthy puppies at term; n = 17). These dogs were evaluated with echocardiography every 10 days from Days 0 to 60 of gestation (Day 0 = estimated day of LH peak). Systolic blood pressure was also assessed. At Day 50 of gestation, left ventricular free wall in systole increased in the normal but not in the abnormal group (P < 0.01). In contrast, end systolic stress (P < 0.01) and systolic blood pressure (P < 0.01) diminished only in normal animals. We concluded that signs of altered maternal cardiovascular adaptation to pregnancy may be predictors of obstetrical complications in dogs.

Electrocardiographic changes in normal and abnormal canine pregnancy.

The aim of this study was to describe the canine electrocardiographic changes in the course of normal and abnormal pregnancy. Twenty-three Brucellosis-negative pregnant bitches were retrospectively classified as normal (n = 12) or abnormal (n = 11). A control group of non-pregnant dioestrous bitches (n = 10) was also included. Normal pregnant females delivered healthy puppies at term while abnormal animals interrupted their pregnancy between days 52-60 (from estimated luteinizing hormone peak) or presented perinatal litter death higher than 60%. All the bitches were electrocardiographically evaluated every 10 days from day 0 to day 65 of the oestrous cycle, to parturition or abortion. Percentage heart rate change increased 31.3% from day 40 to 60 in normal gestation while it decreased -1.8% in dioestrous bitches, although it did not change in the abnormal group (p < 0.01). In the abnormal pregnant group but not in the others, percentage QRSa change fell to -34% on day 60 (p < 0.01). At the same time point, percentage QRSd change was 6.2% vs -4.9% in normal gestations and dioestrous animals, respectively (p < 0.05). Corrected QT interval augmented from day 40 onwards up to 9.9% and 4.3% in the normal pregnant and dioestrous groups, respectively, while it remained unchanged in abnormal gestations (p < 0.05). It is concluded that during normal canine pregnancy, some electrocardiographic parameters begin changing from day 40 onwards, and that pathological gestations differ from normality from day 30. The use of electrocardiography in canine obstetrics might contribute to identify abnormal outcomes before they become clinically evident.

Hypertensive effects of the iv administration of picomoles of ouabain.

Ouabain, an endogenous digitalis compound, has been detected in nanomolar concentrations in the plasma of several mammals and is associated with the development of hypertension. In addition, plasma ouabain is increased in several hypertension models, and the acute or chronic administration of ouabain increases blood pressure in rodents. These results suggest a possible association between ouabain and the genesis or development and maintenance of arterial hypertension. One explanation for this association is that ouabain binds to the α-subunit of the Na(+) pump, inhibiting its activity. Inhibition of this pump increases intracellular Na(+), which reduces the activity of the sarcolemmal Na(+)/Ca(2+) exchanger and thereby reduces Ca(2+) extrusion. Consequently, intracellular Ca(2+) increases and is taken up by the sarcoplasmic reticulum, which, upon activation, releases more calcium and increases the vascular smooth muscle tone. In fact, acute treatment with ouabain enhances the vascular reactivity to vasopressor agents, increases the release of norepinephrine from the perivascular adrenergic nerve endings and promotes increases in the activity of endothelial angiotensin-converting enzyme and the local synthesis of angiotensin II in the tail vascular bed. Additionally, the hypertension induced by ouabain has been associated with central mechanisms that increase sympathetic tone, subsequent to the activation of the cerebral renin-angiotensin system. Thus, the association with peripheral mechanisms and central mechanisms, mainly involving the renin-angiotensin system, may contribute to the acute effects of ouabain-induced elevation of arterial blood pressure.

Hypertensive effects of the iv administration of picomoles of ouabain

Ouabain, an endogenous digitalis compound, has been detected in nanomolar concentrations in the plasma of several mammals and is associated with the development of hypertension. In addition, plasma ouabain is increased in several hypertension models, and the acute or chronic administration of ouabain increases blood pressure in rodents. These results suggest a possible association between ouabain and the genesis or development and maintenance of arterial hypertension. One explanation for this association is that ouabain binds to the α-subunit of the Na+ pump, inhibiting its activity. Inhibition of this pump increases intracellular Na+, which reduces the activity of the sarcolemmal Na+/Ca2+ exchanger and thereby reduces Ca2+ extrusion. Consequently, intracellular Ca2+ increases and is taken up by the sarcoplasmic reticulum, which, upon activation, releases more calcium and increases the vascular smooth muscle tone. In fact, acute treatment with ouabain enhances the vascular reactivity to vasopressor agents, increases the release of norepinephrine from the perivascular adrenergic nerve endings and promotes increases in the activity of endothelial angiotensin-converting enzyme and the local synthesis of angiotensin II in the tail vascular bed. Additionally, the hypertension induced by ouabain has been associated with central mechanisms that increase sympathetic tone, subsequent to the activation of the cerebral renin-angiotensin system. Thus, the association with peripheral mechanisms and central mechanisms, mainly involving the renin-angiotensin system, may contribute to the acute effects of ouabain-induced elevation of arterial blood pressure.