Mutations in EPHB4 cause human venous valve aplasia.

Venous valve (VV) failure causes chronic venous insufficiency, but the molecular regulation of valve development is poorly understood. A primary lymphatic anomaly, caused by mutations in the receptor tyrosine kinase EPHB4, was recently described, with these patients also presenting with venous insufficiency. Whether the venous anomalies are the result of an effect on VVs is not known. VV formation requires complex "organization" of valve-forming endothelial cells, including their reorientation perpendicular to the direction of blood flow. Using quantitative ultrasound, we identified substantial VV aplasia and deep venous reflux in patients with mutations in EPHB4. We used a GFP reporter in mice to study expression of its ligand, ephrinB2, and analyzed developmental phenotypes after conditional deletion of floxed Ephb4 and Efnb2 alleles. EphB4 and ephrinB2 expression patterns were dynamically regulated around organizing valve-forming cells. Efnb2 deletion disrupted the normal endothelial expression patterns of the gap junction proteins connexin37 and connexin43 (both required for normal valve development) around reorientating valve-forming cells and produced deficient valve-forming cell elongation, reorientation, polarity, and proliferation. Ephb4 was also required for valve-forming cell organization and subsequent growth of the valve leaflets. These results uncover a potentially novel cause of primary human VV aplasia.

Editor's Choice - A Comparison of Computed Tomography Angiography and Colour Duplex Ultrasound Surveillance Post Infrarenal Endovascular Aortic Aneurysm Repair: Financial Implications and Impact of Different International Surveillance Guidelines.

OBJECTIVE: Use of colour duplex ultrasound (CDUS) and computed tomography angiography (CTA) for infrarenal endovascular aortic aneurysm repair (EVAR) surveillance differs in internationally published guidelines. This study aimed firstly to compare CDUS detection of significant sac abnormalities with CTA. Secondly, a sensitivity analysis was conducted to compare financial estimates of the, predominantly CDUS based, local and Society of Vascular Surgery (SVS) protocols, the risk stratified European Society of Vascular Surgery (ESVS) protocol, and the CTA based National Institute of Health and Care Excellence (NICE) protocol. METHODS: Agreement between CDUS and CTA was assessed for detection of significant sac abnormalities. Surveillance protocols were extrapolated from published guidelines and applied to infrarenal EVAR patients active on local surveillance at a large, single centre. Surveillance intensity was dependent on presence of endoleak and subsequent risk of treatment failure in accordance with surveillance recommendations. Estimates for each surveillance protocol were inclusive of a range of published incidences of endoleak, contrast associated acute kidney injury (AKI), and excess hospital bed days, and estimated for a hypothetical five year surveillance period. RESULTS: The kappa coefficient between CDUS and CTA for detecting sac abnormalities was 0.68. Maximum five year surveillance cost estimates for the 289 active EVAR patients were £272 359 for SVS, £230 708 for ESVS, £643 802 for NICE, and £266 777 for local protocols, or £1 270, £1 076, £3 003, and £1 244 per patient. Differences in endoleak incidence accounted for a 1.1 to 1.4 fold increase in costs. AKI incidence accounted for a 3.3 to 6.2 fold increase in costs. CONCLUSION: A combined CTA and CDUS EVAR surveillance protocol, with CTA reserved for early seal assessment and confirmatory purposes, provides an economical approach without compromising detection of sac abnormalities. AKI, as opposed to direct imaging costs, accounted for the largest differences in surveillance cost estimates.

Stroke transcranial Doppler in children with human immunodeficiency virus.

AIMS: To describe stroke syndromes and transcranial Doppler (TCD) findings in children with human immunodeficiency virus (HIV) and examine the associations between TCD and clinical and laboratory data. METHOD: We enrolled 42 children (24 males, 18 females) with HIV (median age=7y 6mo; 2y 7mo-15y 6mo), with and without stroke who underwent a TCD examination of the anterior and posterior circulations to derive time-averaged maximum mean velocity (TAMMV) measurements for comparison with previous studies. Clinical and laboratory variables were extracted from the medical records. RESULTS: Of the 42 children with HIV, five had right-sided hemiparesis, three had chronic lung disease, two occurred post-varicella infection, one after herpetic oral ulceration, and one had a poorly functioning left ventricle. Neuroimaging showed middle cerebral artery (MCA) TAMMV greater than 200cm/s, moyamoya-like arteriopathy, left basal ganglia infarction with ipsilateral stenosis, hygroma consistent with venous thrombosis, and a hyperdense left MCA. Eight neurologically asymptomatic children had atypical TCD. The CD4 cell count was non-significantly lower in 6 out of 30 children with atypical TCD (median=21.5; interquartile range=16.1-26.5) compared with the remainder (median=29; interquartile range=21.3-35.0; p=0.09). INTERPRETATION: A variety of stroke syndromes occur in children with HIV. TCD suggests atypical intracranial vessels and/or haemodynamics in some children with HIV infection, consistent with vasculopathy, possibly related directly to immunodeficiency and/or infection. WHAT THIS PAPER ADDS: A range of stroke syndromes are found in children with human immunodeficiency virus (HIV). Transcranial Doppler (TCD) velocities in HIV are commonly outside the range for typically developing children. TCD and neuroimaging data in children with HIV suggest intracranial vasculopathy as one mechanism for stroke. CD4 cell count is non-significantly lower in children with HIV and atypical TCD.

An Educational Study Promoting the Delivery of Transcranial Doppler Ultrasound Screening in Paediatric Sickle Cell Disease: A European Multi-Centre Perspective.

BACKGROUND: Effective stroke prevention in sickle cell disease (SCD) is recommended for children with sickle cell anaemia. Effective implementation relies on the correct stratification of stroke risk using Transcranial Doppler Ultrasound (TCD), prior to committing children to long-term treatment with transfusion. Nevertheless, less than 50% of children with SCD in Europe receive annual TCD-one of the reasons being a lack of trained personnel. The present European multi-centre study was designed to promote the standardisation and delivery of effective screening. METHODS: Fifty-five practitioners from differing professional backgrounds were recruited to the TCD training program. The impact of the training programme was evaluated in three European haematology clinics by comparing stroke risk classification and middle cerebral artery time-averaged maximum velocity (TAMMV) obtained from a cohort of 555 patients, before and after training. RESULTS: 42% (23/55) of trainees successfully completed the program. The TAMMV, used to predict stroke risk at each Centre, demonstrated the highest values in Centre 3 (p < 0.0001) before training. The imaging-TCD TAMMV was also higher in Centre 3 (p < 0.001). Following training, the TAMMV showed closer agreement between centres for both imaging-TCD and non-imaging TCD. The stroke risk distribution of children at each centre varied significantly before training (p < 0.001), but improved after training (Fisher's Exact: no treatment = 5.6, p = 0.41, treatment = 13.8, p < 0.01). The same consistency in stroke risk distribution following training was demonstrated with both non-imaging and imaging-TCD data. CONCLUSION: The attainment of competency in stroke screening using transcranial Doppler scanning (TCD) in sickle cell disease is more feasible for professionals with an ultrasound imaging background. A quality assurance (QA) system is required to ensure that standards are maintained. Further work is in progress to develop an achievable and reproducible QA program.

Human venous valve disease caused by mutations in FOXC2 and GJC2.

Venous valves (VVs) prevent venous hypertension and ulceration. We report that FOXC2 and GJC2 mutations are associated with reduced VV number and length. In mice, early VV formation is marked by elongation and reorientation ("organization") of Prox1hi endothelial cells by postnatal day 0. The expression of the transcription factors Foxc2 and Nfatc1 and the gap junction proteins Gjc2, Gja1, and Gja4 were temporospatially regulated during this process. Foxc2 and Nfatc1 were coexpressed at P0, and combined Foxc2 deletion with calcineurin-Nfat inhibition disrupted early Prox1hi endothelial organization, suggesting cooperative Foxc2-Nfatc1 patterning of these events. Genetic deletion of Gjc2, Gja4, or Gja1 also disrupted early VV Prox1hi endothelial organization at postnatal day 0, and this likely underlies the VV defects seen in patients with GJC2 mutations. Knockout of Gja4 or Gjc2 resulted in reduced proliferation of Prox1hi valve-forming cells. At later stages of blood flow, Foxc2 and calcineurin-Nfat signaling are each required for growth of the valve leaflets, whereas Foxc2 is not required for VV maintenance.

Transcranial Doppler screening for stroke risk in children with sickle cell disease: a systematic review.

Background Sickle cell disease (SCD) is one of the most common causes of stroke in children worldwide. Based on the results of the Stroke Prevention Trial in Sickle Cell Anemia (STOP), annual transcranial Doppler ultrasound (TCD) screening for affected children is standard practice. However, the need for TCD surveillance programs could override the accuracy of the screening, affecting the correct stratification of stroke risk and subsequent clinical management of the target population. Aims To shed light on this issue, a systematic review of the literature on TCD screening for children and adolescents with SCD was carried out (CRD42016050549), according to a list of clinically relevant questions, with a particular focus on screening practices in European countries. Quality of the evidence was rated using the grading of recommendations assessment, development and evaluation. Summary of review Thirty-three studies published in English or French were included (5 randomized controlled trials, 8 experimental non-randomized, and 20 observational studies). The quality of the retrieved evidence ranged between low and high, but was rated as moderate or high most of the times. TCD is effective as a screening tool for the primary prevention of stroke in SCD children. There is no high-quality evidence on the effectiveness of alternative screening methods, such as imaging-TCD with or without angle correction or magnetic resonance angiography. No evidence was found on effectiveness of the screening on children on hydroxyurea and with genotypes other than HbSS and HbS/ß0. No European data were found on screening rates or adherence of screening practices to the STOP protocol. Conclusions High-quality studies on alternative screening methods that are currently used in real-world practice, and on screening applicability to specific subgroups of patients are urgently needed. Considering the low awareness of the disease in European countries and the lack of data on screening practices and adherence, clinicians need up-to-date guidelines for more uniform and evidence-based surveillance of children with SCD.

A dedicated vascular access clinic for children on haemodialysis: Two years' experience.

BACKGROUND: Arteriovenous fistula (AVF) formation for long-term haemodialysis in children is a niche discipline with little data for guidance. We developed a dedicated Vascular Access Clinic that is run jointly by a transplant surgeon, paediatric nephrologist, dialysis nurse and a clinical vascular scientist specialised in vascular sonography for the assessment and surveillance of AVFs. We report the experience and 2-year outcomes of this clinic. METHODS: Twelve new AVFs were formed and 11 existing AVFs were followed up for 2 years. All children were assessed by clinical and ultrasound examination. RESULTS: During the study period 12 brachiocephalic, nine basilic vein transpositions and two radiocephalic AVFs were followed up. The median age (interquartile range) and weight of those children undergoing new AVF creation were 9.4 (interquartile 3-17) years and 26.9 (14-67) kg, respectively. Pre-operative ultrasound vascular mapping showed maximum median vein and artery diameters of 3.0 (2-5) and 2.7 (2.0-5.3) mm, respectively. Maturation scans 6 weeks after AVF formation showed a median flow of 1277 (432-2880) ml/min. Primary maturation rate was 83 % (10/12). Assisted maturation was 100 %, with two patients requiring a single angioplasty. For the 11 children with an existing AVF the maximum median vein diameter was 14.0 (8.0-26.0) mm, and the median flow rate was 1781 (800-2971) ml/min at a median of 153 weeks after AVF formation. Twenty-two AVFs were used successfully for dialysis, a median kt/V of 1.97 (1.8-2.9), and urea reduction ratio of 80.7 % (79.3-86 %) was observed. One child was transplanted before the AVF was used. CONCLUSIONS: A multidisciplinary vascular clinic incorporating ultrasound assessment is key to maintaining young children on chronic haemodialysis via an AVF.

The efficacy of salvage interventions on threatened distal bypass grafts.

OBJECTIVE: Infrapopliteal bypass is an established and effective method for limb salvage in patients with critical limb ischemia. Secondary interventions maybe required to maintain graft patency. The aim of this study was to look at the frequency and outcomes of such interventions. METHODS: Consecutive patients undergoing bypasses onto the infrapopliteal vessels for critical limb ischemia (Rutherford 4-6) at a single institution were analyzed between 2009 and 2013. The primary end points were graft patency, amputation-free survival (AFS), and freedom from reintervention at 12 months by Kaplan-Meier analysis. RESULTS: A total of 114 infrapopliteal bypasses were performed in 102 patients. Distal anastomosis was on to the anterior tibial (n = 31), posterior tibial (n = 27), peroneal (n = 24), tibioperoneal trunk (n = 23), or dorsalis pedis artery (n = 9). Primary patency, assisted primary patency, and secondary patency was 57%, 76%, and 82%, respectively, at 12 months and 44%, 70%, and 80%, respectively, at 36 months. AFS was 80% at 12 months and 65% at 36 months. Endovascular salvage interventions were performed on 58 grafts (51%) including angioplasty of inflow/proximal anastomosis (33%), outflow/distal anastomosis (46%), and graft stenosis (20%), with a further 12 grafts (11%) undergoing thrombolysis for occlusion. Surgical salvage interventions included jump grafts (n = 7), revision of anastomotic stenosis (n = 3), and thrombectomy (n = 2). AFS was similar in salvaged threatened and acutely occluded grafts compared with nonthreatened grafts (P = .064) and better in grafts requiring reintervention later (>6 months from bypass) compared with those requiring early reintervention (<6 months; P = .047). CONCLUSIONS: Secondary interventions in threatened distal bypass grafts are successful at maintaining graft patency and AFS when compared with nonthreatened grafts, and are associated with a low morbidity rate.

Use of a New Hybrid Heparin-Bonded Nitinol Ring Stent in the Popliteal Artery: Procedural and Mid-term Clinical and Anatomical Outcomes.

PURPOSE: To report the immediate and mid-term clinical and anatomical outcomes of a novel, hybrid, heparin-bonded, nitinol ring stent (TIGRIS; Gore Medical) when used for the treatment of lesions located in the popliteal artery. MATERIALS AND METHODS: This was a prospective single-centre registry. Patients eligible for inclusion were individuals suffering from symptomatic popliteal arterial occlusive disease (Rutherford-Becker stage 3-6; P1-P3 segments) and treated with placement of the TIGRIS stent(s). Patients were prospectively scheduled for clinical review and duplex ultrasound follow-up after 6 and 12 months. Outcome measures included immediate technical success, primary vessel patency, in-stent binary restenosis (evaluable by Duplex at 50 % threshold; PSVR > 2.0), freedom from target lesion revascularization (TLR) and amputation-free survival (AFS) estimated by Kaplan-Meier (K-M) survival analysis. Cox proportional-hazards regression analysis was also performed to adjust for confounders and search for independent predictors of outcomes. RESULTS: From August 2012 to March 2014, a total of 54 popliteal TIGRIS stents were implanted in 50 limbs of 48 patients (27 men and 21 women; mean age 76.0 ± 1.7 years). Median Rutherford-Becker stage was five at baseline and 37/50 (74.0%) were chronic total occlusions. Technical success was achieved in all cases (100%). Stented lesion length was 114.2 ± 36.9 mm (range 6-20 cm). Median follow-up was 11.8 ± 0.8 months. After 12 months, primary patency of the TIGRIS stent was 69.5 ± 10.2% with an 86.1 ± 5.9% freedom from TLR and 87 ± 5.0% AFS (K-M estimates). CONCLUSION: The TIGRIS hybrid heparin-bonded nitinol ring stent is a safe and effective endovascular option for complex occlusive disease of the popliteal artery.

Nonleg venous thrombosis in critically ill adults: a nested prospective cohort study.

IMPORTANCE: Critically ill patients are at risk of venous thrombosis, and therefore guidelines recommend daily thromboprophylaxis. Deep vein thrombosis (DVT) commonly occurs in the lower extremities but can occur in other sites including the head and neck, trunk, and upper extremities. The risk of nonleg deep venous thromboses (NLDVTs), predisposing factors, and the association between NLDVTs and pulmonary embolism (PE) or death are unclear. OBJECTIVE: To describe the frequency, anatomical location, risk factors, management, and consequences of NLDVTs in a large cohort of medical-surgical critically ill adults. DESIGN, SETTING, AND PARTICIPANTS: A nested prospective cohort study in the setting of secondary and tertiary care intensive care units (ICUs). The study population comprised 3746 patients, who were expected to remain in the ICU for at least 3 days and were enrolled in a randomized clinical trial of dalteparin vs standard heparin for thromboprophylaxis. MAIN OUTCOMES AND MEASURES: The proportion of patients who had NLDVTs, the mean number per patient, and the anatomical location. We characterized NLDVTs as prevalent or incident (identified within 72 hours of ICU admission or thereafter) and whether they were catheter related or not. We used multivariable regression models to evaluate risk factors for NLDVT and to examine subsequent anticoagulant therapy, associated PE, and death. RESULTS Of 3746 trial patients, 84 (2.2%) developed 1 or more non-leg vein thromboses (superficial or deep, proximal or distal). Thromboses were more commonly incident (n = 75 [2.0%]) than prevalent (n = 9 [0.2%]) (P < .001) and more often deep (n = 67 [1.8%]) than superficial (n = 31 [0.8%]) (P < .001). Cancer was the only independent predictor of incident NLDVT (hazard ratio [HR], 2.22; 95% CI, 1.06-4.65). After adjusting for Acute Physiology and Chronic Health Evaluation (APACHE) II scores, personal or family history of venous thromboembolism, body mass index, vasopressor use, type of thromboprophylaxis, and presence of leg DVT, NLDVTs were associated with an increased risk of PE (HR, 11.83; 95% CI, 4.80-29.18). Nonleg DVTs were not associated with ICU mortality (HR, 1.09; 95% CI, 0.62-1.92) in a model adjusting for age, APACHE II, vasopressor use, mechanical ventilation, renal replacement therapy, and platelet count below 50 × 10(9)/L. CONCLUSIONS AND RELEVANCE Despite universal heparin thromboprophylaxis, nonleg thromboses are found in 2.2% of medical-surgical critically ill patients, primarily in deep veins and proximal veins. Patients who have a malignant condition may have a significantly higher risk of developing NLDVT, and patients with NLDVT, compared with those without, appeared to be at higher risk of PE but not higher risk of death. TRIAL REGISTRATION: clinicaltrials.gov Identifier: NCT00182143.

Effectiveness of alternative strategies to define index case phenotypes to aid genetic diagnosis of familial hypercholesterolaemia.

OBJECTIVE: To determine the utility of secondary stratification measures to improve the ascertainment of index cases of familial hypercholesterolaemia (FH). DESIGN: A retrospective study of genotyped index patients with Simon Broome (SB) FH. SETTING: University teaching hospital. PATIENTS: 204 patients aged 55±14 years; 36% had tendon xanthoma (TX), 21% had coronary heart disease (CHD), low-density lipoprotein cholesterol (LDL-C) was 6.20±2.24 mmol/l and 55% had genetic FH. INTERVENTIONS: The effects of different staging systems (SB vs Dutch criteria), presence of TX, use of LDL-C level, personal history of CHD and imaging evidence of atheroma by carotid intima-media thickness or coronary artery calcium score to identify genetic FH was explored. OUTCOME MEASURES: Changes in C-statistic and net reclassification index (NRI). RESULTS: SB criteria gave a C-statistic of 0.64 comprising C=0.65 in TX(+) and C=0.5 in TX(-) patients. Genetic FH was present in 75% of TX(+) compared with 44% in TX(-) patients. The Dutch criteria gave C=0.72. Addition of imaging criteria to prior CHD raised C=0.64 to C=0.65 in all patients with a NRI of 19% (p=0.06). In TX(-) patients imaging raised C=0.50 to C=0.65 with a NRI of 0.38 (p=0.001) and a weighted comparison index of 0.28, implying the detection of 14 more FH cases per thousand. CONCLUSIONS: Patients with tendon xanthoma (definite FH) should be genotyped. In patients with possible FH, the presence of a personal history of CHD or imaging evidence of increased atheroma offers the best method of identifying index patients likely to have monogenic FH.

Problems with implementing a standardised transcranial Doppler screening programme: impact of instrumentation variation on STOP classification.

BACKGROUND: The Stroke Prevention Trial in Sickle Cell Anaemia (STOP) demonstrated the value of selective transfusion based on transcranial Doppler (TCD) US screening. This facilitated widespread surveillance, but due to reported differences with non-imaging TCD, imaging velocity thresholds have been reduced in some centres. OBJECTIVE: (1) Retrospectively review velocity measurements obtained by non-imaging and imaging TCD, using a standardised protocol. (2) Determine the impact on STOP classification of different velocity thresholds. MATERIALS AND METHODS: TCD data from 23 children (2-19 years of age) were reviewed. The TCD protocol focused on obtaining the velocity corresponding to the highest audible Doppler frequency. STOP velocity thresholds were the recommended for non-imaging TCD and values reduced by 5-15%. RESULTS: Non-imaging and imaging TCD velocities were correlated closely with little overall bias. Reducing imaging TCD velocity thresholds increased the number of abnormal and conditional classifications. Abnormal TCD imaging classifications ranged from 1.9% to 37% depending on the degree of correction applied to the velocity data. CONCLUSION: The current approach for applying STOP thresholds to imaging TCD data may not be required. Centres need to validate their imaging TCD practice to avoid inappropriate selection of patients for transfusion therapy.

Incidence of aetiological subtypes of stroke in a multi-ethnic population based study: the South London Stroke Register.

BACKGROUND: Stroke risk is higher in black ethnic groups compared with white. Although risk factors for stroke are known to differ between these populations, few population studies have reported on the risk of aetiological stroke subtypes in black ethnic populations. METHODS: Ethnic group differences in incidence of first ever ischaemic stroke by aetiological subtype were investigated with the South London Stroke Register (SLSR). The SLSR is a population based stroke register covering a multi-ethnic inner city population of 271,871 inhabitants comprising 63% white, 28% black and 9% other ethnic group. A modified pathophysiological Trial of ORG 10172 in Acute Stroke Treatment (TOAST) classification of stroke was used to estimate patterns of aetiological subtype and stroke was subtyped into large artery atherosclerosis (LAA), cardioembolism (CE), small vessel occlusion (SVO), other aetiology (OTH), undetermined (UND) and multiple possible or concurrent aetiologies (CONC). RESULTS: Between September 1999 and August 2006, 1181 patients with first ever ischaemic stroke were included in the study. Mean age was 71.4 years, 51% were female and 71% were white patients, 20% were black patients, 6% were other and 3% were of unknown ethnic group. The distribution of the aetiological subtypes was as follows: LAA, 109 (9.3%); CE, 325 (27.8%); SVO, 316 (27.0%); OTH, 40 (3.4%); UND, 283 (24.2%) and CONC, 96 (8.2%). The annual age adjusted incidence rate per 100,000 was for total ischaemic stroke 101.2 (95% CI 82.4 to 122.9) in men and 75.1 (95% CI 59.1 to 94.1) in women; for LAA 10.4 (95% CI 5.1 to 18.9) in men and 6.8 (95% CI 2.7 to 14.2) in women; for CE 23.0 (95% CI 14.6 to 34.5) in men and 21.5 (95% CI 13.4 to 32.8) in women; and for SVO 30.3 (95% CI 20.5 to 43.2) in men and 20.3 (95% CI 12.5 to 31.3) in women. The overall incidence rate ratio (IRR) for black patients was 1.25 (1.07 to 1.46), for black Caribbean (BC) patients 1.31 (1.09 to 1.58), for black African (BA) patients 1.22 (0.93 to 1.61) and for other ethnic groups 1.24 (0.96 to 1.61). IRRs for black ethnic groups as well as for BA and BC were significantly higher for SVO in both sexes, for OTH in black patients for females and in BA for males and females compared with the white ethnic group; IRRs for other ethnic groups compared with white patients were higher for SVO in females and for UND in males. INTERPRETATION: Independent important differences in risk of stroke between different ethnic populations strengthen the evidence base for studying genetic susceptibility and environmental influences in ethnic groups separately.

Generalized arterial calcification of infancy: phenotypic spectrum among three siblings including one case without obvious arterial calcifications.

Generalized arterial calcification of infancy (GACI) (OMIM no. 208000) is characterized by calcification of the major arteries and soft tissues and associated with mutations in the ENPP1 gene. Most affected patients die within the first 6 months of life although prolonged survival is increasingly recognized. We report on three siblings with GACI and striking phenotypic variability. Two siblings (including the sibling survivor) were compound heterozygotes for mutations in exon 7 (c.783C>G (p.Y261X)) and exon 8 (c. 878_879delAA (p.K293fsX5)) of the ENPP1 gene confirming the diagnosis of GACI. The sibling survivor did not have calcification on X-ray studies or evidence of hypophosphatemic rickets. GACI may be under recognized and we emphasize consideration of this condition in patients with multiple arterial stenosis even in the absence of radiographic calcification. This adds to the expanding phenotype of GACI and supports a potential role for modifying genes.

Novel candidate genes in unstable areas of human atherosclerotic plaques.

OBJECTIVE: Comparison of gene expression in stable versus unstable atherosclerotic plaque may be confounded by interpatient variability. The aim of this study was to identify differences in gene expression between stable and unstable segments of plaque obtained from the same patient. METHODS AND RESULTS: Human carotid endarterectomy specimens were segmented and macroscopically classified using a morphological classification system. Two analytical methods, an intraplaque and an interplaque analysis, revealed 170 and 1916 differentially expressed genes, respectively using Affymetrix gene chip analysis. A total of 115 genes were identified from both analyses. The differential expression of 27 genes was also confirmed using quantitative-polymerase chain reaction on a larger panel of samples. Eighteen of these genes have not been associated previously with plaque instability, including the metalloproteinase, ADAMDEC1 (approximately 37-fold), retinoic acid receptor responder-1 (approximately 5-fold), and cysteine protease legumain (approximately 3-fold). Matrix metalloproteinase-9 (MMP-9), cathepsin B, and a novel gene, legumain, a potential activator of MMPs and cathepsins, were also confirmed at the protein level. CONCLUSIONS: The differential expression of 18 genes not previously associated with plaque rupture has been confirmed in stable and unstable regions of the same atherosclerotic plaque. These genes may represent novel targets for the treatment of unstable plaque or useful diagnostic markers of plaque instability.

The impact of theoretical errors on velocity estimation and accuracy of duplex grading of carotid stenosis.

Two potential errors in velocity estimation, Doppler angle misalignment and intrinsic spectral broadening (ISB), were determined and used to correct recorded blood velocities obtained from 20 patients (38 bifurcations). The recorded and corrected velocities were used to grade stenoses of greater than 70% using two duplex classification schemes. The first scheme used a peak systolic velocity (PSV) of > 250 cm/s in the internal carotid artery (ICA), and the second a PSV ratio of > 3.4 (ICA PSV/common carotid artery PSV). The "gold standard" was digital subtraction angiography (DSA). The maximum error in velocity estimation due to Doppler angle misalignment was 33 cm/s, but this did not alter sensitivity of stenosis detection. ISB correction caused a reduction in PSV that decreased the sensitivity of the PSV scheme from 65% to 45%. The PSV ratio classification was not affected by ISB errors. Centres using a PSV criterion for grading stenosis should use a fixed Doppler angle and should establish velocity thresholds in-house.