Turbulent blood flow plays an essential localizing role in the development of atherosclerotic lesions in experimentally induced hypercholesterolaemia in rats

Taking into account that atherosclerosis is a focal disease and high levels of plasma cholesterol are closely correlated with its pathogenesis, it is a challenge to explain how equal concentrations of cholesterol bathing the endothelium can produce local, rather than global, effects on arteries. The focal distribution of atherosclerotic lesions has been considered to be dependent, at least in part, on hydrodynamic factors. The present study was carried out to further test the hypothesis that these forces are an important localizing factor in rats feeding a hypercholesterolaemic diet and submitted to infra-diaphragmatic aortic constriction. These animals develop a normotensive prestenotic region with laminar blood flow that serves as control for a normotensive poststenotic region with turbulent blood flow. Our findings clearly demonstrated that the combination of turbulent blood flow and low wall shear stress (WSS) in the presence of hypercholesterolaemia and oxidative stress creates conditions to the formation of focally distributed incipient atherosclerotic lesions observed in the poststenotic segment. In contrast, only diffuse fatty streaks could be observed in the normotensive prestenotic segment with laminar blood flow and normal WSS in the presence of hypercholesterolaemia and oxidative stress. Although haemodynamic forces are not by themselves responsible for the pathogenesis of atherosclerosis, they prime the local vascular wall in which the lesion develop. Further studies are required to establish how haemodynamic forces are detected and transduced into chemical signalling by the cells of the artery wall and then converted into pathophysiologically relevant phenotypic changes.

PULSATILITY INDEX CHANGE OF BRACHIAL ARTERY SHOWS BETTER REPRODUCIBILITY THAN FLOW-MEDIATED VASODILATION

We aim to evaluate intra- and interobserver reproducibility of brachial artery flow-mediated dilation (FMD) and of pulsatility index change (PI-C) 1 min after a 5-min forearm compression in pregnant women. Flow-mediated dilation and PI-C were assessed in 31 healthy pregnant women in 3 consecutive days, by two observers. A broader range of response to the stimulus was observed for PI-C (-58.99%-86.71%) than FMD (2.38%-27.01%). The intraclass correlation coefficients were higher for PI-C than for FMD (intra = 0.90 vs. 0.69, p = 0.047; inter = 0.94 vs. 0.61, p = 0.007). Therefore, PI-C measurements were more reliable than FMD. However, studies comprising of subjects with cardiovascular risk factors relating FMD and PI-C become necessary to validate this method for the study of endothelial dysfunction. (E-mail: wpmartins@gmail.com) (C) 2010 World Federation for Ultrasound in Medicine & Biology.

Quantitative three-dimensional power Doppler angiography: a flow-free phantom experiment to evaluate the relationship between color gain, depth and signal artifact

Objectives To evaluate the presence of false flow three-dimensional (3D) power Doppler signals in `flow-free` models. Methods 3D power Doppler datasets were acquired from three different flow-free phantoms (muscle, air and water) with two different transducers and Virtual Organ Computer-aided AnaLysis was used to generate a sphere that was serially applied through the 3D dataset. The vascularization flow index was used to compare artifactual signals at different depths (from 0 to 6 cm) within the different phantoms and at different gain and pulse repetition frequency (PR F) settings. Results Artifactual Doppler signals were seen in all phantoms despite these being flow-free. The pattern was very similar and the degree of artifact appeared to be dependent on the gain and distance from the transducer. False signals were more evident in the far field and increased as the gain was increased, with false signals first appearing with a gain of 1 dB in the air and muscle phantoms. False signals were seen at a lower gain with the water phantom (-15 dB) and these were associated with vertical lines of Doppler artifact that were related to PRF, and disappeared when reflections were attenuated. Conclusions Artifactual Doppler signals are seen in flow-free phantoms and are related to the gain settings and the distance from the transducer. In the in-vivo situation, the lowest gain settings that allow the detection of blood flow and adequate definition of vessel architecture should be used, which invariably means using a setting near or below the middle of the range available. Additionally, observers should be aware of vertical lines when evaluating cystic or liquid-containing structures. Copyright (C) 2010 ISUOC. Published by John Wiley & Sons, Ltd.

Brachial artery pulsatility index change 1 minute after 5-minute forearm compression - Comparison with flow-mediated dilatation

Objective. Endothelial impairment evaluation by sonographic measurement of flow-mediated dilatation (FMD) has become broadly used. However, this method has 2 main caveats: the dilatation depends on the baseline arterial diameter, and a high precision level is required. Vasodilatation leads to an amplified fall in impedance. We hypothesized that assessment of the pulsatility index change (PI-C) 1 minute after 5-minute forearm compression might evaluate that fall in impedance. The aim of this study was to compare the PI-C with FMD. Methods. Flow-mediated dilatation and the PI-C were assessed in 51 healthy women aged between 35.1 and 67.1 years. We correlated both FMD and the PI-C with age, body mass index, waist circumference, cholesterol level, high-density lipoprotein level, glucose level, systolic and diastolic blood pressure, pulse pressure, brachial artery diameter, simplified Framingham score, intima-media thickness, and carotid stiffness index. Intraclass correlation coefficients between 2 FMD and PI-C measurements were also examined. Results. Only FMD correlated with baseline brachial diameter (r=-0.53). The PI-C had a high correlation with age, body mass index, waist circumference, cholesterol level, systolic blood pressure, pulse pressure, simplified Framingham score, and intima-media thickness. The correlation between FMD and the PI-C was high (r=-0.66). The PI-C had a higher intraclass correlation coefficient (0.991) than FMD (0.836) but not brachial artery diameter (0.989). Conclusions. The PI-C had a large correlation with various markers of cardiovascular risk. Additionally, PI-C measurement does not require offline analysis, extra software, or electrocardiography We think that the PI-C could be considered a marker of endothelial function. However, more studies are required before further conclusions.

Can eccentric arterial plaques alone cause flow stagnation points and favour thrombus incorporation?

We have used an experimental model of aorta stenosis, with a Plexiglas plug, simulating a stable atheromatous plaque that promotes local turbulence and thrombosis. With animal survival of more than 24 h, we followed the partial fibrinolysis of the thrombus as well as its posterior organization and incorporation to the arterial wall as a neointima for up to 30 days. The mushroom plug form permitted the development of recirculation and stasis areas around it, favouring this evolution. Despite noted limitations, this study demonstrates that thrombus incorporation can contribute to plaque extension, as it can promote recirculation and stasis areas.

Influence of pregnancy and smoking on brachial artery flow-mediated dilation values and time until maximum response

To evaluate the effect of pregnancy and smoking on endothelial function using brachial artery flow-mediated dilation (FMD) and to determine the time necessary until the occurrence of maximum brachial artery dilation after stimulus. This study was an observational study evaluating 133 women, who were grouped as follows: non-smoking pregnant women (N = 47), smoking pregnant women (N = 33), non-smoking women (N = 34), and smoking pregnant women (N = 19). The diameter of the brachial artery was measured at baseline and at 30, 60, 90 and 120 s after stimulus. The relative change of brachial artery was determined for each of these four moments. FMD measured at 60 s after stimulus was compared between the groups. The maximum FMD was observed at 60 s after cuff release in all groups. FMD was greater among non-smoking pregnant women compared to smoking pregnant women (11.50 +/- A 5.77 vs. 8.74 +/- A 4.83; p = 0.03) and also between non-smoking non-pregnant women compared to smoking non-pregnant women (10.52 +/- A 4.76 vs. 7.21 +/- A 5.57; p = 0.03). Maximum FMD was observed approximately 60 s after stimulus in all groups regardless of smoking and pregnancy status. The smoking habit seems to lead to endothelial dysfunction both in pregnant and non-pregnant women, as demonstrated by the lower FMD in smokers.

Analysis of placenta vascularization in patients with uterine altered artery Doppler flow velocity exams

Background: One of the frequent questions in obstetric practice is to determine placental vascular changes that may account for abnormal Doppler flow velocity alterations in maternal uterine vessels from women and fetuses without pregnancy pathology. Methods: A retrospective morphometric study was realized using 27 placentas from patients submitted for Doppler flow velocity exam during pregnancy. The placentas were morphologically examined using hematoxylin-eosin staining. Measurements of villi were made with the use of a video camera coupled to a common light microscope and a computer with automatic image analyzing software. Results: Of the 27 placentas, 13 (48%) were of patients showing unaltered Doppler and 14 (52%) showing altered Doppler. The number of stem villi vessels was significantly larger in the placentas of patients with Doppler exam alterations (P = 0.003). This group also presented greater stem villi vessel thickness, although without significant difference. The number of intermediary and terminal villi vessels was greater in the placentas of patients with altered Doppler exams (P < 0.001), and a greater terminal villi area was observed in these cases (P < 0.001). Conclusion: The morphological proof that uterine artery Doppler flow velocity exam alterations are associated with placental vascular alterations demonstrates the importance of this exam during prenatal care, even in the absence of maternal-fetal alterations.

Chlorhexidine-induced apoptosis or necrosis in L929 fibroblasts: A role for endoplasmic reticulum stress

Chlorhexidine (CHX), widely used as antiseptic and therapeutic agent in medicine and dentistry, has a toxic effect both in vivo and in vitro. The intrinsic mechanism underlying CHX-induced cytotoxicity in eukaryotic cells is, however, still unknown. A recent study from our laboratory has suggested that CHX may induce death in cultured L929 fibroblasts via endoplasmic reticulum (ER) stress. This hypothesis was further tested by means of light and electron microscopy, quantification of apoptosis and necrosis by flow cytometry, fluorescence visualization of the cytoskeleton and endoplasmic reticulum, and evaluation of the expression of 78-kDa glucose-regulated protein 78 (Grp78), a marker of activation of the unfolded protein response (UPR) in cultured L929 fibroblasts. Our finding showing increased Grp 78 expression in CHX-treated cells and the results of flow cytometry, cytoskeleton and endoplasmic reticulum fluorescence visualization, and scanning and transmission electron microscopy allowed us to suggest that CHX elicits accumulation of proteins in the endoplasmic reticulum, which causes ER overload, resulting in ER stress and cell death either by necrosis or apoptosis. It must be pointed out, however, that this does not necessarily mean that ER stress is the only way that CHX kills L929 fibroblasts, but rather that ER stress is an important target or indicator of cell death induced by this drug. (C) 2008 Elsevier Inc. All rights reserved.

Quantification of Nicotine in Commercial Brand Cigarettes HOW MUCH IS INHALED BY THE SMOKER?

The main objective of this experiment is to determine the amount of nicotine in commercial brand cigarettes by means of a nonaqueous acid-base titration. A simple glass device simulating a smoker is proposed, which allows the determination of the volatilized, filter retained, and inhaled portions. Students will readily see that the amount of nicotine/cigarette stated on the label (similar to 0.5-1.0 mg) refers indeed to the inhaled portion only, rather than to the total amount/cigarette (usually more than 10 mg). Even so, values for inhaled nicotine may be significantly higher than those reported for several brands. Students will also be able to make a critical evaluation of the true content of nicotine in the inhaled portion and confront it with the reported value for a given brand. In addition, the theoretical approach, supported by HPLC data, provides an excellent experience on nonaqueous acid-base volumetric analysis.

FURTHER INSIGHTS TOWARD VITAMIN C DETERMINATION AND STABILITY. PROPOSAL OF A NEW QUANTIFICATION METHOD

This work aimed at an evaluation of the classical iodine method for quantification of vitamin C (L-ascorbic acid) in fruit juices, as well as at a search into the stability of this so popular vitamin under different conditions of pH, temperature and light exposition, in addition to a proposal of a new quantification method. Our results point to the persistent reversibility of the blue color of the starch-triiodide complex at the end point when using the classical iodine titration, and the overestimation of the true vitamin concentration in fruit juices. A new quantification method is proposed in order to overcome this problem. Surprising conclusions were obtained regarding the controversial stability of L-ascorbic acid toward atmospheric oxygen, at low pH, even in fruit juice and at room temperature, showing that the major problem concerned with aging of fruit juices is proliferation of microorganisms rather than expontaneous oxidation of L-ascorbic acid.

Effects of type II pyrethroid cyhalothrin on rat innate immunity: A flow cytometric study

Synthetic type II pyrethroids induce anxiety, immunosuppresion or, alternatively, immunostimulatory effects in laboratory animals. Macrophages and neutrophils are known to be key elements in cellular immune responses. The present study was designed to investigate the in vivo effects of cyhalothrin (1.0 and 3.0 mg/kg/once daily for 7 days) on macrophage and neutrophil activities, using a flow cytometry method. Results showed that cyhalothrin treatment decreased the percentage and intensity of phagocytosis performed by macrophages, but did not alter these parameters in neutrophils: and also decreased basal neutrophil oxidative burst and increased S. aureus-induced neutrophil oxidative burst, but did not alter these responses in macrophages. The present results are discussed in the light of a possible indirect action of cyhalothrin on macrophage and neutrophil activities via hypothalamic pituitary adrenal (HPA) axis activation. A possible direct effect of cyhalothrin on macrophage and neutrophil activities is also considered. (C) 2008 Elsevier B.V. All rights reserved.

Effect of defocused infrared diode laser on salivary flow rate and some salivary parameters of rats

This study aims to investigate whether infrared diode low-level laser therapy (LLLT) increased salivary flow rate and altered pH value, protein concentration, and peroxidase and amylase activities in saliva of rats. Wistar rats were used and divided into three groups. Experimental groups (A and 13) had their parotid, submandibular and sublingual glands submitted to diode laser, 808-nm wavelength, on two consecutive days. The dose results were 4 and 8 J/cm(2), respectively. A red guide light was used to visualize the irradiated area. Group C was irradiated only with red pilot beam and served as control. The saliva samples were collected after each irradiation step (first and second collection days) and 1 week after the first irradiation (seventh day). Statistical analysis was performed, and differences were observed according to different days of salivary collection. The results showed that salivary flow rate for groups A and B was higher on the seventh day if it is compared to data obtained for the first day (p<0.05). LLLT applications on salivary glands are a therapy procedure that requires further studies.

Comparison of the effects of pilocarpine and cevimeline on salivary flow

Objective: The aim of the present study was to compare the effect of low-dose pilocarpine and cevimeline as stimulants for salivary flow in healthy subjects. Methods: In this cross-over clinical trial with a 1-week washout period, 40 male volunteers were submitted to an oral dose of pilocarpine 1% (Salagen (TM)) -60 mu g kg(-1) body-weight (Group 1) or Cevimeline (Evoxac (TM)) -30 mg (Group 2). Saliva samples were collected and the salivary flow rate was measured (ml min(-1)) at baseline and 20, 40, 60, 80, 140 and 200 min after administration of drugs. In addition, salivary secretion was also measured under mechanical stimulation to observe salivary gland function. Results: The data were analyzed by Friedman and Wilcoxon signed-rank tests (significance level = 5%). Pilocarpine and cevimeline significantly increased salivary flow 140 min after intake. There was a significant higher secretion with cevimeline 140 and 200 min after administration. There were no differences seen among subjects in the salivary glands function by mechanical stimulation. Conclusion: Both drugs showed efficacy in increasing the salivary flow in healthy volunteers, but cevimeline was more effective than pilocarpine.

Evaluation of the flow rate of 3 endodontic sealers: Sealer 26, AH Plus, and MTA Obtura

Objective. The aim of this study was to evaluate the flow rate of 3 endodontic sealers: Sealer 26, AH Plus, and MTA Obtura. Study design. According to the method proposed by the American Dental Association (ADA specification no. 57), the sealers were placed between 2 glass slabs under a weight of 120 g. The diameters of the formed discs were measured with a digital paquimeter. The test was repeated 5 times for each sealer. Results. The results were expressed as arithmetic means, and the statistical analysis was performed through Tukey test. AH Plus showed significantly superior flow rate compared with Sealer 26 and MTA Obtura. There was no statistically significant difference between flow rates presented by Sealer 26 and MTA Obtura. Conclusion. Within the limitations of this in vitro study, it was concluded that all of the the endodontic sealers tested presented greater flow than the minimum recommended in the ADA 57 specification. (Oral Surg Oral Med Oral Pathol Oral Radiol Endod 2010;109:e47-e49)

GP5+/6+ SYBR Green methodology for simultaneous screening and quantification of human papillomavirus

Background: Detection and quantification of human papillomavirus (HPV) may help in predicting the evolution of HPV infection and progression of associated lesions. Objectives: We propose a novel protocol using consensus primers GP5+/6+ in a SYBR Green quantitative real-time (Q-RT) polymerase chain reaction (PCR). The strategy permits screening for HPV infection and viral load quantification simultaneously. Study design: DNA from 153 archived cervical samples, previously tested for HPV detection by GP5+/6+ PCR and typed by EIA-RLB (enzyme immunoassay-reverse line blot) or sequence analysis, was analysed using SYBR Green Q-RT PCR. Melting temperature assay (T(m)) and cycle threshold (C(t)) were used to evaluate HPV positivity and viral load. The T(m) in the range of 77-82 degrees C was considered to be positive for HPV-DNA. HPV results generated through GP5+/6+ conventional PCR were considered the gold standard against which sensitivity and specificity of our assay were measured. Results: Out of 104 HPV positive samples, 100 (96.2%) were also determined as positive by SYBR Green Q-RT PCR; of the 49 HPV-negative samples, all were determined as negative. There was an excellent positivity agreement (K = 0.94) between the SYBR Green Q-RT and the previous methods employed. The specificity and sensitivity were 100% and 96.2%, respectively. Comparison of SYBR Green Q-RT and TaqMan oligo-probe technologies gave an excellent concordance (pc = 0.95) which validated the proposed strategy. Conclusions: We propose a sensitive and easy-to-perform technique for HPV screening and viral load quantification simultaneously. (C) 2009 Elsevier B.V. All rights reserved.