Petru Liuba

Endothelial Dysfunction After Repeated Chlamydia pneumoniae Infection in Apolipoprotein E–Knockout Mice

BackgroundArterial relaxation is largely regulated by endothelial nitric oxide (NO). Its diminished activity has been associated with incipient atherosclerosis. We investigated the endothelium-dependent relaxation of aorta in apolipoprotein E–knockout (apoE-KO) mice exposed to single or repeated Chlamydia pneumoniae inoculation. Methods and ResultsForty-eight apoE-KO mice, 8 weeks old, were inoculated intranasally with C pneumoniae (n=24) or saline (n=24) every 2 weeks over a 6-week period. Twenty mice (10 infected and 10 controls) were killed at 2 weeks and 6 weeks, respectively, after the first inoculation. The smooth muscle tone of aortic rings was measured in vitro at both time points. The norepinephrine-precontracted thoracic aortic rings were successively exposed to methacholine in the absence and presence of NG-nitro-l-arginine methyl ester (L-NAME) and diclofenac. The methacholine-induced relaxation was attenuated in the infected mice at 6 weeks in both the absence and presence of L-NAME (P <0.05 and P <0.01, respectively). When administered together with L-NAME, diclofenac enhanced the relaxation of the L-NAME–pretreated aortas in infected mice at 2 weeks (P <0.05) but not in noninfected mice. The relaxation response from infected mice tended to differ in the same manner at 6 weeks (P <0.1). No intimal thickening was detected at either time point. ConclusionsC pneumoniae impairs arterial endothelial function, and the NO pathway is principally involved. Cyclooxygenase-dependent vasoconstricting products may also account for the infection-induced impaired relaxation. These findings further support the role of C pneumoniae infection in atherosclerosis development.

Acute infections in children are accompanied by oxidative modification of LDL and decrease of HDL cholesterol, and are followed by thickening of carotid intima-media.

BACKGROUND Atherosclerosis begins early in life. Infections might contribute to the pathogenesis of atherosclerosis. In this study, we investigated whether acute infections in children could alter the carotid wall morphology and the lipid profile. METHODS Mean carotid intima-media thickness (IMT) was measured by high-resolution ultrasound in 28 hospitalised children (mean age: 5+/-2 years), who fulfilled the diagnostic criteria of acute infections (body temperature, >38 degrees C; C-reactive protein, >15mg/ml, and clinical), and in 20 age- and gender-matched controls. Antibodies against oxidised low-density lipoprotein (anti-oxLDL antibodies), as well as total and high-density lipoprotein cholesterol (HDL-C) were analysed in all children. The infection group was investigated both during the acute illness and 3 months after clinical recovery (post-infection). RESULTS During the acute illness, the infection group had elevated anti-oxLDL antibodies and decreased HDL-C, as compared to those obtained at 3 months and in controls (p<0.05). These changes in the infection group were followed, at 3 months, by thickening of carotid intima-media. Those who received antibiotics during their acute illness had less carotid thickening than those who were not treated with antibiotics (p<0.05). CONCLUSION Acute infections in children seem to be accompanied by enhanced oxidative modification of LDL and by decrease in HDL-C. These lipid changes may be followed by thickening of carotid artery intima-media. These findings suggest that, in childhood, acute infections could be associated with increased risk of atherosclerosis, and warrant further studies on this topic.

Acute Chlamydia pneumoniae infection causes coronary endothelial dysfunction in pigs.

BACKGROUND Coronary endothelial dysfunction contributes to the pathogenesis of acute coronary syndromes (ACSs). Acute Chlamydia pneumoniae infection has been epidemiologically associated with ACS. In this study, we investigated whether acute C. pneumoniae infection could alter the endothelial vasomotor function of porcine coronary vessels. METHODS AND RESULTS Twenty pigs, 7-9 kg in weight, were inoculated intratracheally with C. pneumoniae (n=12) or saline (n=8), and investigated at 3 days (five infected/four non-infected) and 2 weeks (5+2 infected/four non-infected) after inoculation. The endothelium-dependent reactivity of coronary microcirculation was assessed at both time points by measuring peak coronary flow velocity (CFV) in response to bradykinin, before and after infusions with glutathione, an antioxidant, and L-arginine, a substrate for nitric oxide synthase (NOS). CFV after bradykinin was significantly decreased in infected animals at both time points. At 2 weeks, both glutathione and L-arginine significantly improved CFV after bradykinin. CFV after sodium nitroprusside (SNP) was similar in both groups. At 3 days, the relaxation responses of bradykinin-induced pre-contracted left anterior descending (LAD) coronary rings to bradykinin were significantly less in infected animals. N(G)-nitro-L-arginine-methyl-ester, an NOS inhibitor, had significantly greater inhibitory effect on bradykinin-induced relaxation in infected animals. Plasma nitrate-nitrite and fibrinogen, and NOS activity from LAD coronary samples were significantly increased in infected animals. CONCLUSION Acute C. pneumoniae infection causes endothelial dysfunction of both resistance and epicardial coronary vessels, and favours a pro-coagulant status. These effects could in part account for the epidemiologically suggested association between acute infection and ACS.

Co-Infection with CHLAMYDIA PNEUMONIAE and HELICOBACTER PYLORI Results in Vascular Endothelial Dysfunction and Enhanced VCAM-1 Expression in ApoE-Knockout Mice.

Background: Upregulation of proinflammatory endothelial cell adhesion molecules and decreased bioactivity of endothelial nitric oxide (NO) are important in the pathogenesis of atherosclerosis. We investigated the effects of co-infection with Chlamydia pneumoniae and Helicobacter pylori on these two events in apoE-KO mice. Methods: Thirty-two apoE-KO mice, 8 weeks old, were equally divided into 4 groups. The first 2 groups were infected with either C. pneumoniae or H. pylori, while the 3rd group was infected with both C. pneumoniae and H. pylori. Mice from the 4th group and 4 wild-type mice served as controls. Thoracic and abdominal aortas were harvested after 10 weeks, and staining for vascular cell adhesion molecule-1 (VCAM-1) and intracellular adhesion molecule-1 was analyzed by immunocytochemistry. The endothelial vasomotor responses of thoracic aortas to methacholine were studied in organ chambers in the absence and presence of L-NAME. The plasma levels of nitrate/nitrite were measured. Results: Staining for VCAM-1 was more intense at the branching sites of aortas from mice with co-infection than in mono-infected or noninfected apoE-KO mice. The relaxation responses to methacholine and the plasma levels of nitrate/nitrite were significantly less in the co-infected group than in the other groups (p < 0.05). Conclusion: Co-infection of apoE-KO mice with C. pneumoniae and H. pylori seems to be associated with impaired bioactivity of endothelial NO and increased expression of VCAM-1 at branching sites. The findings may suggest an additive interaction of these pathogens in atherogenesis.

Poor vitamin C status is associated with increased carotid intima-media thickness, decreased microvascular function, and delayed myocardial repolarization in young patients with type 1 diabetes

BACKGROUND Vascular endothelial dysfunction, accelerated thickening of arterial intima, and changes in ventricular repolarization contribute to increased cardiovascular morbidity in type 1 diabetes (T1D). Although vitamin C has important antioxidant functions and increased oxidative stress is a central mechanism of cardiovascular dysfunction in T1D, the relation between vitamin C and the cardiovascular system in young diabetic patients has not been investigated. OBJECTIVE In a cohort of young patients with T1D, we investigated the relation of plasma concentrations of vitamin C with indexes of vascular function and structure and duration of the QT interval corrected for heart rate (QT(c)). DESIGN Carotid artery intima-media thickness, cutaneous microvascular function, and duration of the QT(c) interval were measured in 59 patients (mean age: 17 y; range: 10-22 y) with T1D (diabetes duration: 3-20 y). Plasma vitamin C was analyzed by HPLC with coulometric detection. RESULTS Carotid artery intima-media thickness and duration of the QT(c) interval were higher in patients in the lowest tertile of vitamin C than in those in the highest tertile (P < 0.05 for both). The cutaneous microvascular response to acetylcholine was lower (P = 0.003) in the lowest tertile group than in the highest tertile group, but the response to sodium nitroprusside was not significantly different between these 2 groups. All differences remained significant after adjustment for age, sex, diabetes duration, body mass index, and glycated hemoglobin. CONCLUSIONS In this relatively small-scale cross-sectional study of young patients with T1D, lower plasma concentrations of vitamin C seem to be associated with adverse changes in the microcirculation, peripheral arteries, and ventricular repolarization. Large-scale prospective studies are needed to confirm these results and to clarify the underlying mechanisms.

Effects of ketogenic diet on vascular function.

BACKGROUND Ketogenic diet is a well-established treatment in children with difficult to treat epilepsy. Very little is known about the long-term effects on vascular atherogenic and biochemical processes of this high-fat and low carbohydrate and protein diet. METHODS We evaluated 26 children after one year and 13 children after two years of ketogenic diet. High resolution ultrasound-based assessment was used for carotid artery intima-media thickness (cIMT), carotid artery distensibility and carotid artery compliance. Blood lipids including high density lipoprotein cholesterol (HDL-C), low density lipoprotein cholesterol, (LDL-C), total cholesterol (TC), apolipoprotein A (apoA), apolipoprotein B (apoB) and high-sensitivity C-reactive protein (hsCRP) were analysed. RESULTS A gradual decrease in carotid distensibility and an increase in LDL-C, apoB and the TC:LDL-C and LDL-C:HDL-C ratios were seen at three and 12 months of KD-treatment. These differences were not significant at 24 months. cIMT, BMI and hsCRP did not show any significant changes. CONCLUSIONS The initial alterations in lipids, apoB and arterial function observed within the first year of KD-treatment appear to be reversible and not significant after 24 months of treatment.

Residual adverse changes in arterial endothelial function and LDL oxidation after a mild systemic inflammation induced by influenza vaccination

Background. Several clinical studies have suggested possible increase in cardiovascular risk during and in the first weeks after an acute inflammatory disease. Using influenza vaccine as inflammatory stimulus, we investigated whether arterial endothelial dysfunction could persist beyond the inflammatory state, and whether amplified oxidative modification of low‐density lipoprotein (LDL) accompanies this vascular disturbance. Methods and subjects. The brachial artery responses to hyperemia (flow‐mediated dilatation (FMD), and to sublingual glyceryl trinitrate (GTN), and the carotid intima‐media thickness were assessed by external ultrasound in eight healthy male volunteers (age 17–30 y) before, and 2 and 14 days after intramuscular administration of influenza vaccine. Plasma levels of high‐sensitivity C‐reactive protein (CRP), fibrinogen, cyclic guanosine monophosphate (cGMP), and antibodies against oxidized LDL (oxLDL) were measured at each time point. Data are means±standard errors of the mean (SEM). Results. Influenza vaccination caused a slight elevation in CRP (from 0.5±0.1 at baseline, to 2±0.6 mg/L, P = 0.01) and fibrinogen (from 2.3±0.1 to 2.7±0.1 g/L, P = 0.01) at 2 days, which completely resolved at 14 days (CRP: 0.6±0.2 mg/L, P = 0.9, and fibrinogen: 2.3±0.1 g/L, P = 0.8 versus baseline). OxLDL antibody levels rose significantly at 2 days (from 1±0.1 at baseline to 2±0.4, P = 0.04), and remained elevated at 14 days (1.7±0.3, P = 0.1 versus baseline). FMD of the brachial artery decreased at 2 days (from 8.3±1.2% at baseline, to 5.4±1%, P = 0.05) with a further decrease at 14 days (4.9±0.8%, P = 0.03 versus baseline). The dilatory responses to GTN and the carotid IMT remained unchanged throughout the study period (P>0.5). Conclusion. Abnormalities in arterial function and LDL oxidation may persist for at least 2 weeks after a slight inflammatory reaction induced by influenza vaccination. These could explain in part the earlier reported increase in cardiovascular risk during the first weeks after an acute inflammatory disorder.

Respiratory infection recurrence and passive smoking in early atherosclerosis in children and adolescents with type 1 diabetes.

Background  Optimal glucose control in juvenile type 1 diabetes mellitus is necessary but not sufficient to reduce the burden of cardiovascular events in later life. This emphasizes the importance of searching for other possible risk factors associated with diabetes. We investigated whether recurrent episodes of acute respiratory infections and exposure to tobacco smoke could influence vascular phenotypes for early atherosclerosis in children and adolescents with type 1 diabetes.

Infection-induced coronary dysfunction and systemic inflammation in piglets are dampened in hypercholesterolemic milieu

The synergism of infection with conventional cardiovascular risk factors in atherosclerosis is much debated. We hypothesized that coronary arterial injury correlates with infection recurrence and pathogen burden and is further aggravated by hypercholesterolemia. Forty-two Göttingen minipigs were assigned to repeated intratracheal inoculation of PBS, Chlamydia pneumoniae (Cpn), or both Cpn and influenza virus at 8, 11, and 14 wk of age. Animals were fed either standard or 2% cholesterol diet (chol-diet). At 19 wk of age coronary vasomotor responses to acetylcholine (ACh) and adenosine were assessed in vivo and blood and tissue samples were collected. Nonparametric tests were used to compare the groups. In cholesterol-fed animals, total cholesterol/HDL was significantly increased in infected animals compared with noninfected animals [3.13 (2.17-3.38) vs. 2.03 (1.53-2.41), respectively; P = 0.01]. C-reactive protein (CRP) rose in infected animals [10.60 (4.96-18.00) vs. 2.47 (1.44-3.01) μg/ml in noninfected; P < 0.01] without significant difference between the mono- and coinfected groups. Among coinfected animals, both CRP and haptoglobin were lower in those fed chol-diet than in those fed standard diet (P < 0.05). The vasoconstricting response to ACh was most prominent in coinfected animals {769.3 (594-1,129) cm; P = 0.03 vs. noninfected [342 (309-455) cm] and P = 0.07 vs. monoinfected [415 (252.5-971.8) cm]}. Among monoinfected animals, similar to CRP, a trend for less vasoconstriction was observed in those fed chol-diet (P = 0.08). Coinfection of piglets appears to be associated with more pronounced coronary muscarinic vasomotor dysfunction. In monoinfected animals, use of chol-diet seems to dampen both coronary dysfunction and systemic inflammation induced by infection.

Bradykinin preconditions postischemic arterial endothelial function in humans.

Abstract  Background: Arterial endothelial dysfunction is an important mechanism of tissue injury caused by ischemia‐reperfusion (I/R). Earlier studies of I/R have shown that intracoronary preinfusion with 2.5–5 μg/mL bradykinin (BK) could alleviate the postischemic myocardial damage. Using an experimental human model of I/R, we investigated whether preceding infusion with BK could prevent the I/R‐induced arterial endothelial dysfunction. Methods: The left radial artery (LRA) from 16 healthy male adults, 18 to 30 years old, was submitted to I/R by completely occluding the left brachial artery with a pressure tourniquet for 20 minutes (ischemia), followed by its release (reperfusion). Prior to I/R, half of the subjects were randomly assigned to receive either BK (5 μg/mL) or saline, both being infused into the left brachial artery (0.5 mL/min, 10 min). The infusion was followed by a 10‐minute drug‐free period. The endothelial function of the LRA was studied by measuring the flow‐mediated dilation (FMD) at baseline (prior to drug infusion), and at 15 minutes of reperfusion. In addition, baseline radial artery diameter, plasma nitrate, and von Willebrand factor were measured at these time points, and immediately before I/R (pre‐I/R). Results: BK had no effect on the pre‐I/R plasma nitrate (p > 0.5 vs. saline) and diameter of LRA (p > 0.5 vs. baseline). At 15 minutes of reperfusion, FMD was significantly decreased in the saline group as compared to baseline (absolute dilation: 0.08 ± 0.03 vs. 3.02 ± 0.8 mm, respectively, p < 0.01; percentage dilation: 3 ± 0.6 vs. 8 ± 0.6%, respectively, p < 0.001), but it remained unaffected in the BK group (absolute dilation: 3.06 ± 0.9 vs. 3.27 ± 0.8 mm, respectively, p > 0.5; percentage dilation: 7 ± 0.7 vs. 8 ± 0.8%, respectively, p > 0.5). A similar trend was observed with regard to plasma nitrate, which remained unchanged in the BK group (37.01 ± 4.14 vs. 39.14 ± 4.49 μmol/L, p > 0.5) but decreased in the saline group (35.91 ± 3.03 vs. 28.91 ± 2.81 μmol/L, p < 0.1). Conclusion: Infusion of BK could protect the arterial endothelial function against I/R injury in humans, possibly in part by preserving the endothelial NO availability. The findings support the use of BK in the prevention of tissue injury due to I/R and might reveal an additional mechanism whereby ACE inhibitors exert their preconditioning effects on myocardium.