Brain Perivascular Spaces as Biomarkers of Vascular Risk: Results from the Northern Manhattan Study.

BACKGROUND AND PURPOSE: Dilated perivascular spaces in the brain are associated with greater arterial pulsatility. We hypothesized that perivascular spaces identify individuals at higher risk for systemic and cerebral vascular events. MATERIALS AND METHODS: Stroke-free participants in the population-based Northern Manhattan Study had brain MR imaging performed and were followed for myocardial infarction, any stroke, and death. Imaging analyses distinguished perivascular spaces from lesions presumably ischemic. Perivascular spaces were further subdivided into lesions with diameters of ≤3 mm (small perivascular spaces) and >3 mm (large perivascular spaces). We calculated relative rates of events with Poisson models and hazard ratios with Cox proportional models. RESULTS: The Northern Manhattan Study participants who had MR imaging data available for review (n = 1228; 59% women, 65% Hispanic; mean age, 71 ± 9 years) were followed for an average of 9 ± 2 years. Participants in the highest tertile of the small perivascular space score had a higher relative rate of all deaths (relative rate, 1.38; 95% CI, 1.01-1.91), vascular death (relative rate, 1.87; 95% CI, 1.12-3.14), myocardial infarction (relative rate, 2.08; 95% CI, 1.01-4.31), any stroke (relative rate, 1.79; 95% CI, 1.03-3.11), and any vascular event (relative rate, 1.74; 95% CI, 1.18-2.56). After we adjusted for confounders, there was a higher risk of vascular death (hazard ratio, 1.06; 95% CI, 1.01-1.11), myocardial infarction (hazard ratio, 2.22; 95% CI, 1.12-4.42), and any vascular event (hazard ratio, 1.04; 95% CI, 1.01-1.08) with higher small perivascular space scores. CONCLUSIONS: In this multiethnic, population-based study, participants with a high burden of small perivascular spaces had increased risk of vascular events. By gaining pathophysiologic insight into the mechanism of perivascular space dilation, we may be able to propose novel therapies to better prevent vascular disorders in the population.

Predicting different adhesive regimens of circulating particles at blood capillary walls.

A fundamental step in the rational design of vascular targeted particles is the firm adhesion at the blood vessel walls. Here, a combined lattice Boltzmann-immersed boundary model is presented for predicting the near-wall dynamics of circulating particles. A moving least squares algorithm is used to reconstruct the forcing term accounting for the immersed particle, whereas ligand-receptor binding at the particle-wall interface is described via forward and reverse probability distributions. First, it is demonstrated that the model predicts with good accuracy the rolling velocity of tumor cells over an endothelial layer in a microfluidic channel. Then, particle-wall interactions are systematically analyzed in terms of particle geometries (circular, elliptical with aspect ratios 2 and 3), surface ligand densities (0.3, 0.5, 0.7 and 0.9), ligand-receptor bond strengths (1 and 2) and Reynolds numbers (Re = 0.01, 0.1 and 1.0). Depending on these conditions, four different particle-wall interaction regimens are identified, namely not adhering, rolling, sliding and firmly adhering particles. The proposed computational strategy can be efficiently used for predicting the near-wall dynamics of particles with arbitrary geometries and surface properties and represents a fundamental tool in the rational design of particles for the specific delivery of therapeutic and imaging agents.

Effect of mitochondrial ascorbic acid synthesis on photosynthesis.

Ascorbic acid (AA) is synthesized in plant mitochondria through the oxidation of l-galactono-1,4-lactone (l-GalL) and then distributed to different cell compartments. AA-deficient Arabidopsis thaliana mutants (vtc2) and exogenous applications of l-GalL were used to generate plants with different AA content in their leaves. This experimental approach allows determining specific AA-dependent effects on carbon metabolism. No differences in O2 uptake, malic and citric acid and NADH content suggest that AA synthesis or accumulation did not affect mitochondrial activity; however, l-GalL treatment increased CO2 assimilation and photosynthetic electron transport rate in vtc2 (but not wt) leaves demonstrating a stimulation of photosynthesis after l-GalL treatment. Increased CO2 assimilation correlated with increased leaf stomatal conductance observed in l-GalL-treated vtc2 plants.

Point-of-care multiorgan ultrasonography for the evaluation of undifferentiated hypotension in the emergency department.

PURPOSE: We analyzed the efficacy of a point-of-care ultrasonographic protocol, based on a focused multiorgan examination, for the diagnostic process of symptomatic, non-traumatic hypotensive patients in the emergency department. METHODS: We prospectively enrolled 108 adult patients complaining of non-traumatic symptomatic hypotension of uncertain etiology. Patients received immediate point-of-care ultrasonography to determine cardiac function and right/left ventricle diameter rate, inferior vena cava diameter and collapsibility, pulmonary congestion, consolidations and sliding, abdominal free fluid and aortic aneurysm, and leg vein thrombosis. The organ-oriented diagnoses were combined to formulate an ultrasonographic hypothesis of the cause of hemodynamic instability. The ultrasonographic diagnosis was then compared with a final clinical diagnosis obtained by agreement of three independent expert physicians who performed a retrospective hospital chart review of each case. RESULTS: Considering the whole population, concordance between the point-of-care ultrasonography diagnosis and the final clinical diagnosis was interpreted as good, with Cohen's k = 0.710 (95 % CI, 0.614-0.806), p < 0.0001 and raw agreement (Ra) = 0.768. By eliminating the 13 cases where the final clinical diagnosis was not agreed upon (indefinite), the concordance increased to almost perfect, with k = 0.971 (95 % CI, 0.932-1.000), p < 0.0001 and Ra = 0.978. CONCLUSIONS: Emergency diagnostic judgments guided by point-of-care multiorgan ultrasonography in patients presenting with undifferentiated hypotension significantly agreed with a final clinical diagnosis obtained by retrospective chart review. The integration of an ultrasonographic multiorgan protocol in the diagnostic process of undifferentiated hypotension has great potential in guiding the first-line therapeutic approach.

Endothelial function in mild primary hyperparathyroidism.

BACKGROUND: It is not known if endothelial dysfunction, an important early event in the pathogenesis of atherosclerosis, is present in mild primary hyperparathyroidism (PHPT) and if so, whether it improves following parathyroidectomy. DESIGN: We measured flow-mediated vasodilation (FMD), which estimates endothelial function by ultrasound imaging, in patients prior to and 6 and 12 months after parathyroidectomy. RESULTS: Forty-five patients with mild PHPT [80% female, 61 ± 1 (mean ± SE) years, serum calcium 2·65 ± 0·03 mm (10·6 ± 0·1 mg/dl), PTH 10·5 ± 0·7 pm (99 ± 7 pg/ml), 25-hydroxyvitamin D (25OHD) 70·3 ± 3·7 nm (28·2 ± 1·5 ng/ml)] were studied. Baseline FMD was normal (4·63 ± 0·51%; reference mean: 4·4 ± 0·1%) and was not associated with serum calcium, PTH or 25OHD levels. In the group as a whole, FMD did not change after surgery (6 months: 4·38 ± 0·83%, P = 0·72; 12 months: 5·07 ± 0·74%, P = 0·49). However, in those with abnormal baseline FMD (<2·2%; n = 15), FMD increased by 350%, normalizing by 6 months after surgery (baseline: 0·81± 0·19%; 6 months: 3·18 ± 0·79%, P = 0·02 vs baseline; 12months: 3·68 ± 1·22%, P = 0·04 vs baseline). Baseline calcium, PTH and 25OHD levels did not differ between those with abnormal vs normal FMD, nor did these indices predict postoperative change in FMD. CONCLUSIONS: FMD is generally normal in patients with mild PHPT and is unchanged 1 year after parathyroidectomy. Although FMD may normalize after surgery in patients with baseline abnormalities, data do not support using endothelial dysfunction as an indicator for parathyroidectomy.

Non-invasive measurement of local pulse pressure by pulse wave-based ultrasound manometry (PWUM).

Central blood pressure (CBP) has been established as a relevant indicator of cardiovascular disease. Despite its significance, CBP remains particularly challenging to measure in standard clinical practice. The objective of this study is to introduce pulse wave-based ultrasound manometry (PWUM) as a simple-to-use, non-invasive ultrasound-based method for quantitative measurement of the central pulse pressure. Arterial wall displacements are estimated using radiofrequency ultrasound signals acquired at high frame rates and the pulse pressure waveform is estimated using both the distension waveform and the local pulse wave velocity. The method was tested on the abdominal aorta of 11 healthy subjects (age 35.7 ± 16 y.o.). PWUM pulse pressure measurements were compared to those obtained by radial applanation tonometry using a commercial system. The average intra-subject variability of the pulse pressure amplitude was found to be equal to 4.2 mmHg, demonstrating good reproducibility of the method. Excellent correlation was found between the waveforms obtained by PWUM and those obtained by tonometry in all subjects (0.94 < r < 0.98). A significant bias of 4.7 mmHg was found between PWUM and tonometry. PWUM is a highly translational method that can be easily integrated in clinical ultrasound imaging systems. It provides an estimate of the pulse pressure waveform at the imaged location, and may offer therefore the possibility to estimate the pulse pressure at different arterial sites. Future developments include the validation of the method against invasive estimates on patients, as well as its application to other large arteries.

Evaluation of prosthetic-valved devices by means of numerical simulations.

The in vivo evaluation of prosthetic device performance is often difficult, if not impossible. In particular, in order to deal with potential problems such as thrombosis, haemolysis, etc., which could arise when a patient undergoes heart valve replacement, a thorough understanding of the blood flow dynamics inside the devices interacting with natural or composite tissues is required. Numerical simulation, combining both computational fluid and structure dynamics, could provide detailed information on such complex problems. In this work, a numerical investigation of the mechanics of two composite aortic prostheses during a cardiac cycle is presented. The numerical tool presented is able to reproduce accurately the flow and structure dynamics of the prostheses. The analysis shows that the vortical structures forming inside the two different grafts do not influence the kinematics of a bileaflet valve or the main coronary flow, whereas major differences are present for the stress status near the suture line of the coronaries to the prostheses. The results are in agreement with in vitro and in vivo observations found in literature.

Fluid-structure interaction of deformable aortic prostheses with a bileaflet mechanical valve.

Two different aortic prostheses can be used for performing the Bentall procedure: a standard straight graft and the Valsalva graft that better reproduces the aortic root anatomy. The aim of the present work is to study the effect of the graft geometry on the blood flow when a bileaflet mechanical heart valve is used, as well as to evaluate the stress concentration near the suture line where the coronary arteries are connected to graft. An accurate three-dimensional numerical method is proposed, based on the immersed boundary technique. The method accounts for the interactions between the flow and the motion of the rigid leaflets and of the deformable aortic root, under physiological pulsatile conditions. The results show that the graft geometry only slightly influences the leaflets dynamics, while using the Valsalva graft the stress level near the coronary-root anastomoses is about half that obtained using the standard straight graft.

Cardiac structure and diastolic function in mild primary hyperparathyroidism.

CONTEXT: Data on the presence, extent, and reversibility of cardiovascular disease in primary hyperparathyroidism (PHPT) are conflicting. OBJECTIVE: To evaluate the heart in PHPT, we assessed cardiac structure and diastolic function in patients with mild PHPT compared with age- and sex-matched controls. DESIGN: This was a case-control study. SETTINGS: The study was conducted in a university hospital Metabolic Bone Diseases Unit. PARTICIPANTS: Fifty-four men and women with PHPT and 76 controls without PHPT participated in the study. OUTCOME MEASURES: We measured left ventricular mass index (LVMI), the presence of mitral annular calcification, the ratio of early to late diastolic mitral inflow velocities (E/A), and early diastolic velocity of the lateral mitral annulus using Doppler tissue imaging (tissue Doppler e'). RESULTS: Patients had mild disease with mean (+/-sd) serum calcium 10.5 +/- 0.5 mg/dl and PTH 96 +/- 45 pg/ml. LVMI and diastolic function were normal in PHPT. There was no difference in LVMI (98 +/- 23 vs. 96 +/- 24 g/m(2), P = 0.69) or the frequency of mitral annular calcification between PHPT cases and controls. Diastolic function variables (E/A and tissue Doppler e') were higher (better) in cases compared with controls, although both were within the reference range. PHPT patients with low E/A had higher serum PTH (121 +/- 36 vs. 89 +/- 46 pg/ml, P = 0.03) and calcium (10.8 +/- 0.4 vs. 10.5 +/- 0.5 mg/dl, P = 0.05) than those with normal values. Finally, we found LVMI to be inversely associated with serum 25-hydroxyvitamin D in PHPT (r = -0.29, P < 0.05). All findings persisted after adjustment for group differences in cardiovascular risk factors. CONCLUSIONS: Patients with biochemically mild PHPT do not have evidence of increased left ventricular mass, diastolic dysfunction, or increased valvular calcifications. However, the data support an association between low vitamin D levels and the development of left ventricular hypertrophy in this disorder. Finally, the increased serum calcium and PTH levels in those with diastolic dysfunction suggest that disease severity may determine the presence of cardiac manifestations in PHPT.

Vesicular acetylcholine transporter (VAChT) in the brain of spontaneously hypertensive rats (SHR): effect of treatment with an acetylcholinesterase inhibitor.

The cholinergic marker vesicular acetylcholine transporter (VAChT) was investigated in different cerebral areas of spontaneously hypertensive rats (SHR) by immunochemistry (Western blot analysis) and by immunohistochemistry. SHR were used as an animal model of hypertensive brain damage. The sensitivity of manipulation of cholinergic system on VAChT was assessed in rats treated for four weeks with the acetylcholinesterase (AChE) inhibitor galantamine (3 mg/Kg/day). VAChT concentrations were increased in the brain of control SHR compared to age-matched normotensive Wistar-Kyoto rats. This increase probably represents an up-regulation of VAChT to oppose cholinergic deficits reported in SHR and is countered by galantamine administration. The possibility that cholinergic neurotransmission enhancement may represent a therapeutic strategy in cerebrovascular disease is discussed.

Impact of recent legislative bills regarding clinical research on Italian ethics committee activity.

AIMS AND BACKGROUND: The present work assessed the impact of two decrees on ethics committees in Italy, aimed at bringing the national laws on the conduct of clinical trials into line with the rest of the EC, and regulating and facilitating not-for-profit research. MATERIAL AND METHODS: Prospectively collected data from an Italian multicentre study were examined with respect to the ethics review process. Administrative and time elements of the review process were audited. Main outcome measures were time between the application submission and the ethics committee definitive opinion, type and number of application submission forms, number of ethics committees that refused fee exemption, and time between the ethics committee approval and the administrative authorisation. RESULTS: A total of 134 local research ethics committees (LRECs) were approached. Application submission procedures and application forms varied greatly; paper submission was mandatory. The median time from submission to approval was 72 days. Only two LRECs refused the fee exemption. The median time from LREC approval to administrative agreement was 50 days and only 9.6% of local authorities came to a verbal agreement with the sponsor. CONCLUSIONS: Italian LRECs are still not sufficiently efficient in complying with the Directive 2001/20/EC requirement (60 days). Better coordination of LRECs work is needed although the optimal level of coordination between them is still not known. In the meantime, national guidelines are needed concerning the application of Directive 2001/20/EC. The behaviour of Italian LRECs towards not-for-profit research was excellent although only the fee exemption was requested.

Impact of valvular thickness on stroke recurrence in medically treated patients with stroke.

BACKGROUND: It remains controversial whether left-sided valvular thickening (VaT) is a risk factor for ischemic stroke. Little is known about the relationship between VaT and the recurrent adverse event rate in medically treated patients with stroke. METHODS: We examined the outcomes of 627 noncardioembolic stroke patients who were double-blindly assigned to either warfarin or aspirin therapy and assessed VaT using transesophageal echocardiography. Endpoints were recurrent ischemic stroke or death from any cause. The Cox proportional hazards model was used to adjust for covariates. RESULTS: VaT was present in 57.3% of the patients (359/627), 34.6% (271/627) involving the aortic valve and 46.4% (291/627) involving the mitral valve. There was no difference in the time to primary endpoints between those with and without VaT of the aortic valve (p = 0.49; hazard ratio, HR: 1.17; 95% CI: 0.74-1.85; 2-year event rates: 18.9 vs. 13.2%) or mitral valve (p = 0.66; HR: 0.91; 95% CI: 0.60-1.38; 2-year event rates: 16.9 vs. 14.7%). Among the patients with VaT, there was no significant difference in the time to primary endpoints between those treated with warfarin and those with aspirin (p = 0.13, HR: 0.65, 95% CI: 0.37-1.14, 2-year event rates: 15.2 vs. 22.7% for the aortic valve; p = 0.22, HR: 0.70, 95% CI: 0.40-1.23, 2-year event rates: 14.2 vs. 19.6% for the mitral valve). CONCLUSIONS: VaT does not appear to increase recurrent adverse event rates in medically treated patients with ischemic stroke, regardless of warfarin or aspirin therapy.

Increased ascorbate content delays flowering in long-day grown Arabidopsis thaliana (L.) Heynh.

Flowering requires the integration of different inductive stimuli, including light, temperature and hormones. In an attempt to assess whether ascorbate (ASC) could contribute to the control of flowering time, we analyzed the effects of increased ASC content on the transition to the reproductive stage in the facultative long-day plant Arabidopsis thaliana. ASC content was increased by spraying leaves with the ASC precursor L-galactono-gamma-lactone. Our data show that increased ASC content did not affect vegetative growth, whereas a significant delay (5 days in average) in flower production occurred in ASC-overproducing plants. Higher ASC availability resulted in delayed expression of LEAFY (LFY), the gene encoding for a key transcription factor integrating different flowering-inductive pathways. On the contrary, spraying with gibberellin under the same condition caused both early LFY expression and early flowering. The possible role of ASC in the transition to the reproductive stage is discussed.

P-glycoprotein 170 expression and function as an adverse independent prognostic factor in childhood acute lymphoblastic leukemia.

Little is known about the prognostic role of multidrug resistance (MDR) in newly diagnosed childhood acute lymphoblastic leukemia (ALL). P-glycoprotein 170 (MDR1), a cellular drug efflux pump, is thought to be one of the major causes of MDR. The aim of this retrospective study was to evaluate in 85 children with ALL the impact of the MDR1 product of the mdr-1 gene on the achievement of complete remission (CR) and outcome. MDR1 protein expression was performed by immunocytochemistry (ICC), and flow cytometry (FC). MDR1 functional activity was performed by a rhodamine (Rhd)-123 efflux test with or without verapamil. All patients enrolled in our study were treated with AIEOP ALL 91-95 protocols. At diagnosis, 40 patients (47%) expressed MDR1 protein at significant levels, and 45 (53%) were MDR1 negative. Forty-three of the latter patients were also negative for MDR1 function, while 34/40 (85%) patients MDR1 positive preserved the function. Rhd-123 efflux was inhibited by the MDR modulator verapamil in 12/40 (30%) patients. After induction treatment, CR was achieved in 77/85 children (90.6%). All patients who did not achieve CR were MDR1 positive. Twenty-nine patients relapsed, 17 (58.6%) of whom were MDR1 positive. The 10-year overall survival (OS) rate, and disease-free survival (DFS) for MDR1 negative patients compared to MDR1 positive patients were 75.7% versus 54.8%, and 67.5% versus 46%, respectively. The 10-year event-free survival (EFS) rate was significantly higher (67.5% versus 36.8%) in the MDR1 negative group compared with the MDR1 positive population (p=0.001). Multivariate analysis showed that only EFS was independent of age, WBC count, immunophenotype, FAB subtype and prednisone response (p=0.019). Our results, derived from a monocentric study, demonstrate that MDR1 expression in childhood ALL is an independent adverse prognostic factor on outcome, and could be a useful biological marker of response in these patients. Moreover, MDR1 function was also a predictor of response, but only in univariate analysis.

Hopes, disillusions and more hopes from vitamin C.

In the current view of most biochemists and physiologists, the role of L-ascorbic acid (AA) in cell metabolism would be more or less confined to the scavenging of reactive oxygen species. Nevertheless, many data have been collected in our and other laboratories concerning the involvement of AA in many different aspects of cell metabolism. At the present time the molecular sites of action of AA have not been completely elucidated, but recent findings on the specific requirement of AA for the activity of several 2-oxoacid-dependent dioxygenases involved in cell signalling and the activation of transcription factors open new fascinating perspectives for further research.

Localization of the m5 muscarinic cholinergic receptor in rat circle of Willis and pial arteries.

The expression and microanatomical localization of the muscarinic cholinergic m5 receptor subtype was investigated in rat circle of Willis and pial arteries by in situ hybridization, immunoblotting and immunohistochemistry. In situ hybridization histochemistry revealed a strong signal in the endothelium of circle of Willis and pial arteries and a moderate signal in the tunica media of the same arteries, within smooth muscle. Exposure of membranes of arteries to anti-m5 receptor protein antibodies caused the development of a band of approximately 81 kDa. Immunohistochemistry revealed the accumulation of m5 receptor protein immunoreactivity primarily within endothelium of circle of Willis and cerebral arteries and to a lesser extent in the tunica media, within smooth muscle. Medium (external diameter 200-100 microm) and small-sized (external diameter smaller than 100 microm) pial arteries displayed a significantly higher immune staining than large-sized pial arteries or circle of Willis arteries. The above data that are consistent with recent functional studies reporting cholinergic dilation of cerebral blood vessels mediated via a m5 receptor, have shown that both endothelial and muscular components of cerebral arteries synthesize and express a muscarinic m5 receptor. In view of the peculiar localization in cerebral vessels, handling of the muscarinic m5 receptor may be considered as an approach in the treatment of cerebrovascular disease.

Renal cell carcinoma in children: a clinicopathologic study.

PURPOSE: To identify the prognostic factors, treatment, and outcome of children affected by renal cell carcinoma (RCC). PATIENTS AND METHODS: The series included 41 patients (18 males and 23 females) with a median age of 124 months observed at the 11 Italian Association for Pediatric Hematology and Oncology centers from January 1973 to January 2001. Clinical data, surgical notes, pathologic findings, and summaries of therapy were taken from the charts. RESULTS: Seven (17%) of the 41 patients had a papillary histology, and 34 (82.4%) had nonpapillary histology. Eighteen patients (43.9%) had stage I, one patient (2.4%) had stage II, two patients (4.8%) had stage IIIA, 10 patients (24.3%) had stage IIIB, and nine patients (21.9%) had stage IV disease. One patient had a bilateral involvement at diagnosis. Seven patients experienced disease recurrence. Lung and liver were the most common distant lesions and usually were fatal. In this study, the major factor influencing the prognosis was the stage. Event-free survival at 20 years was 53.5% for all patients. Overall survival at 20 years was 54.9% for all patients. CONCLUSION: RCC is a rare disease in children and adolescents. This neoplasm has a different clinical presentation in children compared with adults but the same outcome. In our experience, patients with localized disease could be cured by nephrectomy alone. Prospective studies in a larger number of patients are needed to confirm radiation therapy and biologic response modifiers as effective adjunct therapy in RCC stage III. The alternative therapy seems warranted in patients with advanced disease.

Determination of the in vivo effects of prednisone on Bcl-2 family protein expression in childhood acute lymphoblastic leukemia.

Glucocorticoid resistance is often associated with treatment failure in children with acute lymphoblastic leukaemia (ALL) but the underlying molecular mechanisms are still unclear. In 30 consecutive children with ALL treated with prednisone we determined changes in the expression of Bcl-2, Bax and Bcl-xl proteins in leukemic lymphoblasts and related these to clinical features and rate of prednisone-induced apoptosis. The apoptotic index increased after prednisone therapy in 24 of the 30 patients. At diagnosis, we detected expression of Bcl-2 and Bcl-xl protein in 28 samples, while Bax expression protein was detected in 21 of the 30 patients. Prednisone treatment induced a decrease in Bcl-2 and Bcl-xl levels in 17 and 16 of the 28 patients, respectively, while Bax protein increased in 14 of the 21 patients. Twenty of the 30 patients studied were considered to be good prednisone responders, whereas 10 were poor responders. We observed a statistically significant decrease only for Bcl-xl protein expression in T phenotype ALL, in the poor responder group and in patients with >20000/mm(3) white cell count (WBC) at diagnosis. These data suggest a role of Bcl-xl in the mechanisms of protection of leukemic cells from apoptosis induced by glucocorticoids (GCs).

Acute myocardial infarction caused by paradoxical coronary embolization in a patient with a patent foramen ovale.

We describe the case of a young woman without cardiac risk factors who had an acute inferoapical myocardial infarction. Coronary angiographic appearance was consistent with thrombus in the distal left anterior descending coronary artery. A patent foramen ovale with moderate right-to-left shunting after the Valsalva maneuver was detected by contrast transesophageal echocardiography. No other cardioembolic source was identified. Paradoxical embolization through a patent foramen ovale is a rare phenomenon, which appeared to have resulted in myocardial infarction in this patient.