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HIV-1 spreads efficiently through direct cell-to-cell transmission at virological synapses (VSs) formed by interactions between HIV-1 envelope proteins (Env) on the surface of infected cells and CD4 receptors on uninfected target cells. Env-CD4 interactions bring the infected and uninfected cellular membranes into close proximity and induce transport of viral and cellular factors to the VS for efficient virion assembly and HIV-1 transmission. Using novel, cell-specific stable isotope labeling and quantitative mass spectrometric proteomics, we identified extensive changes in the levels and phosphorylation states of proteins in HIV-1 infected producer cells upon mixing with CD4+ target cells under conditions inducing VS formation. These coculture-induced alterations involved multiple cellular pathways including transcription, TCR signaling and, unexpectedly, cell cycle regulation, and were dominated by Env-dependent responses. We confirmed the proteomic results using inhibitors targeting regulatory kinases and phosphatases in selected pathways identified by our proteomic analysis. Strikingly, inhibiting the key mitotic regulator Aurora kinase B (AURKB) in HIV-1 infected cells significantly increased HIV activity in cell-to-cell fusion and transmission but had little effect on cell-free infection. Consistent with this, we found that AURKB regulates the fusogenic activity of HIV-1 Env. In the Jurkat T cell line and primary T cells, HIV-1 Env:CD4 interaction also dramatically induced cell cycle-independent AURKB relocalization to the centromere, and this signaling required the long (150 aa) cytoplasmic C-terminal domain (CTD) of Env. These results imply that cytoplasmic/plasma membrane AURKB restricts HIV-1 envelope fusion, and that this restriction is overcome by Env CTD-induced AURKB relocalization. Taken together, our data reveal a new signaling pathway regulating HIV-1 cell-to-cell transmission and potential new avenues for therapeutic intervention through targeting the Env CTD and AURKB activity., Competing Interests: The authors have declared that no competing interests exist., (Copyright: © 2023 Bruce et al. This is an open access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited.)

Background and Purpose: Arterial and neck vessel system characteristics of patients with multiple sclerosis have not been previously investigated. Therefore, the aim of this study was to examine the frequency of neck vessels and their cross-sectional areas (in square millimeters) between patients with MS and healthy controls., Materials and Methods: In this study, 193 patients with MS and 193 age- and sex-matched healthy controls underwent 2D TOF venography at 3T. The main arterial (carotid and vertebral), venous (internal jugular), and secondary neck vessels were examined at 4 separate cervical levels (C2/3, C4, C5/6, and C7/T1). The ANCOVA adjusted for age, body mass index, smoking status, hypertension, and heart disease was used to compare the differences between patients with MS and healthy controls., Results: After controlling for all confounding factors, patients with MS had significantly lower cross-sectional areas of the carotid arteries at the C2/3 ( P = .03), C5/6 ( P = .026), and C7/T1 ( P = .005) levels as well as of the vertebral arteries at the C2/3 ( P = .02), C4 ( P = .012), and C7/T1 ( P = .006) levels, compared with healthy controls. A higher frequency of secondary neck vessels was found at all 4 levels in patients with MS: C2/3 (12.9 versus 10, P < .001), C4 (9.1 versus 7.5, P < .001), C5/6 (7.8 versus 6.8, P = .012), and C7/T1 (8.8 versus 6, P < .001). The total cross-sectional areas of secondary neck vessels were also significantly higher at all 4 levels ( P < .03). No significant differences in the cross-sectional areas of jugular veins were found between patients with MS and healthy controls., Conclusions: Patients with MS showed lower cross-sectional areas of the carotid and vertebral arteries and a higher frequency of secondary neck vessels and their cross-sectional areas compared with healthy controls., (© 2018 by American Journal of Neuroradiology.)

Objectives To assess the relationship between cross-sectional area of internal jugular veins and brain volumes in healthy individuals without neurologic disease. Methods A total of 193 healthy individuals without neurologic disease (63 male and 130 female; age > 20 to < 70 years) received magnetic resonance venography and structural brain magnetic resonance imaging at 3T. The internal jugular vein cross-sectional area was assessed at C2-C3, C4, C5-C6, and C7-T1. Normalized whole brain volume was assessed. Partial correlation analyses were used to determine associations. Results There was an inverse relationship between normalized whole brain volume and total internal jugular vein cross-sectional area (C7-T1: males r = -0.346, p = 0.029; females r = -0.301, p = 0.002). After age adjustment, association of normalized whole brain volume and normalized gray matter volume with internal jugular vein cross-sectional area became positive in males (normalized whole brain volume and right internal jugular vein cross-sectional area (C2-C3) changed from r = -0.163 to r = 0.384, p = 0.002), but not in the females. Conclusion Sex differences exist in the relationship between brain volume and internal jugular vein cross-sectional area in healthy individuals without neurologic disease.

Purpose To assess cerebral microbleed (CMB) prevalence in patients with multiple sclerosis (MS) and clinically isolated syndrome (CIS) and associations with clinical outcomes. Materials and Methods CMBs are associated with aging and neurodegenerative disorders. The prevalence of CMBs has not previously been well established. In this study, 445 patients with MS (266 with relapsing-remitting MS, 138 with secondary progressive MS, and 41 with primary progressive MS), 45 patients with CIS, 51 patients with other neurological diseases, and 177 healthy control subjects (HCs) underwent 3-T magnetic resonance (MR) imaging and clinical examinations. A subset of 168 patients with MS and 50 HCs underwent neuropsychological testing. Number of CMBs was assessed on susceptibility-weighted minimum intensity projections by using the Microbleed Anatomic Rating Scale; volume was calculated by using quantitative susceptibility maps. Differences between groups were analyzed with the χ 2 test, Fisher exact test, Student t test, and analysis of variance; associations of CMBs with clinical and other MR imaging outcomes were explored with correlation and regression analyses. Because CMB frequency increases with age, prevalence was investigated in participants at least 50 years of age and younger than 50 years. Results Significantly more patients with MS than HCs had CMBs (19.8% vs 7.4%, respectively; P = .01) in the group at least 50 years old. A trend toward greater presence of CMBs was found in patients with MS (P = .016) and patients with CIS who were younger than 50 years (P = .039) compared with HCs. In regression analysis adjusted for age, hypertension, and normalized brain volume, increased number of CMBs was significantly associated with increased physical disability in the MS population (R 2 = 0.23, P < .0001). In correlation analysis, increased number of CMBs was significantly associated with deteriorated auditory and verbal learning and memory (P = .006) and visual information processing speed trends (P = .049) in patients with MS. Conclusion Monitoring CMBs may be relevant in patients with MS and CIS at higher risk for developing cognitive and physical disability. © RSNA, 2016 Online supplemental material is available for this article.

Objectives: Constricted cerebral venous outflow has been linked with increased cerebrospinal fluid (CSF) pulsatility in the aqueduct of Sylvius in multiple sclerosis (MS) patients and healthy individuals. This study investigates the relationship between CSF pulsatility and internal jugular vein (IJV) cross-sectional area (CSA) in these two groups, something previously unknown., Methods: 65 relapsing-remitting MS patients (50.8% female; mean age = 43.8 years) and 74 healthy controls (HCs) (54.1% female; mean age = 43.9 years) were investigated. CSF flow quantification was performed on cine phase-contrast MRI, while IJV-CSA was calculated using magnetic resonance venography. Statistical analysis involved correlation, and partial least squares correlation analysis (PLSCA)., Results: PLSCA revealed a significant difference (p<0.001; effect size = 1.072) between MS patients and HCs in the positive relationship between CSF pulsatility and IJV-CSA at C5-T1, something not detected at C2-C4. Controlling for age and cardiovascular risk factors, statistical trends were identified in HCs between: increased net positive CSF flow (NPF) and increased IJV-CSA at C5-C6 (left: r = 0.374, p = 0.016; right: r = 0.364, p = 0.019) and C4 (left: r = 0.361, p = 0.020); and increased net negative CSF flow and increased left IJV-CSA at C5-C6 (r = -0.348, p = 0.026) and C4 (r = -0.324, p = 0.039), whereas in MS patients a trend was only identified between increased NPF and increased left IJV-CSA at C5-C6 (r = 0.351, p = 0.021). Overall, correlations were weaker in MS patients (p = 0.015)., Conclusions: In healthy adults, increased CSF pulsatility is associated with increased IJV-CSA in the lower cervix (independent of age and cardiovascular risk factors), suggesting a biomechanical link between the two. This relationship is altered in MS patients.

Background: Internal jugular vein (IJV) narrowing has been implicated in central nervous system pathologies, however normal physiological age- and gender-related IJV variance in healthy individuals (HIs) has not been adequately assessed., Objectives: We assessed the relationship between IJV cross-sectional area (CSA) and aging., Materials and Methods: This study involved 193 HIs (63 males and 130 females) who received 2-dimensional magnetic resonance venography at 3T. The minimum CSA of the IJVs at cervical levels C2/C3, C4, C5/C6, and C7/T1 was obtained using a semi-automated contouring-thresholding technique. Subjects were grouped by decade. Pearson and partial correlation (controlled for cardiovascular risk factors, including hypertension, heart disease, smoking and body mass index) and analysis of variance analyses were used, with paired t-tests comparing side differences., Results: Mean right IJV CSA ranges were: in males, 41.6 mm2 (C2/C3) to 82.0 mm2 (C7/T1); in females, 38.0 mm2 (C2/C3) to 62.3 mm2 (C7/T1), while the equivalent left side ranges were: in males, 28.0 mm2 (C2/C3) to 52.2 mm2 (C7/T1); in females, 27.2 mm2 (C2/C3) to 47.8 mm2 (C7/T1). The CSA of the right IJVs was significantly larger (p<0.001) than the left at all cervical levels. Controlling for cardiovascular risk factors, the correlation between age and IJV CSA was more robust in males than in the females for all cervical levels., Conclusions: In HIs age, gender, hand side and cervical location all affect IJV CSA. These findings suggest that any definition of IJV stenosis needs to account for these factors.

Background and Purpose: Aging of the healthy brain is characterized by focal or nonfocal white matter (WM) signal abnormality (SA) changes, which are typically detected as leukoaraiosis (LA). Hypertension is a risk factor for WM lesion formation. This study investigated whether LA might be associated with increased cerebrospinal fluid (CSF) pulsatility linked to arterial hypertension., Methods: A total of 101 individuals without neurologic diseases (53 females and 48 males) aged between 18 and 75 years underwent 3T brain MRI with cine phase contrast imaging for CSF flow estimation, after providing their informed consent. LA was defined as the presence of focal T2 WM SA changes and/or nonfocal uniform areas of signal increase termed dirty appearing white matter (DAWM). Relevant information relating to cardiovascular risk factors was also collected., Results: When controlled for age and hypertension, significant partial correlations were observed between: DAWM volume and: net negative flow (r = -.294, P = .014); net positive flow (NPF) (r = .406, P = .001); and peak positive velocity (r = .342, P = .004). Multiple linear regression analysis revealed DAWM volume to be significantly correlated with CSF NPF (P = .019) and hypertension (P = .007), whereas T2 WM SA volume was only significantly correlated with age (P = .002). Combined DAWM and T2 WM SA volumes were significantly related with age (P = .001) and CSF peak negative velocity (P = .041)., Conclusions: Rarefaction of WM leading to LA is a multifactorial process, in which formation of DAWM induced by hypertension and increased aqueductal CSF pulsatility, may play a contributory role. These two factors appear to act independently of each other in a process that is independent of age., (Copyright © 2015 by the American Society of Neuroimaging.)

Quantifying flow from phase-contrast MRI (PC-MRI) data requires that the vessels of interest be segmented. The estimate of the vessel area will dictate the type and magnitude of the error sources that affect the flow measurement. These sources of errors are well understood, and mathematical expressions have been derived for them in previous work. However, these expressions contain many parameters that render them difficult to use for making practical error estimates. In this work, some realistic assumptions were made that allow for the simplification of such expressions in order to make them more useful. These simplified expressions were then used to numerically simulate the effect of segmentation accuracy and provide some criteria that if met, would keep errors in flow quantification below 10% or 5%. Four different segmentation methods were used on simulated and phantom MRA data to verify the theoretical results. Numerical simulations showed that including partial volumed edge pixels in vessel segmentation provides less error than missing them. This was verified with MRA simulations, as the best performing segmentation method generally included such pixels. Further, it was found that to obtain a flow error of less than 10% (5%), the vessel should be at least 4 (5) pixels in diameter, have an SNR of at least 10:1 and have a peak velocity to saturation cut-off velocity ratio of at least 5:3., (Copyright © 2015 Elsevier Inc. All rights reserved.)

Purpose: To investigate cerebrospinal fluid (CSF) dynamics in the aqueduct of Sylvius (AoS) in chronic cerebrospinal venous insufficiency (CCSVI)-positive and -negative healthy individuals using cine phase contrast imaging., Materials and Methods: Fifty-one healthy individuals (32 CCSVI-negative and 19 age-matched CCSVI-positive subjects) were examined using Doppler sonography (DS). Diagnosis of CCSVI was established if subjects fulfilled ≥2 venous hemodynamic criteria on DS. CSF flow and velocity measures were quantified using a semiautomated method and compared with clinical and routine 3T MRI outcomes., Results: CCSVI was associated with increased CSF pulsatility in the AoS. Net positive CSF flow was 32% greater in the CCSVI-positive group compared with the CCSVI-negative group (P = 0.008). This was accompanied by a 28% increase in the mean aqueductal characteristic signal (ie, the AoS cross-sectional area over the cardiac cycle) in the CCSVI-positive group compared with the CCSVI-negative group (P = 0.021)., Conclusion: CSF dynamics are altered in CCSVI-positive healthy individuals, as demonstrated by increased pulsatility. This is accompanied by enlargement of the AoS, suggesting that structural changes may be occurring in the brain parenchyma of CCSVI-positive healthy individuals., (© 2013 Wiley Periodicals, Inc.)

Background and Purpose: It has been demonstrated that increased levels of iron in the brain occur with aging. In this study we investigated the nature of the association between age and SWI-filtered phase values, indicative of iron content, in the subcortical deep gray matter of healthy individuals., Materials and Methods: A total of 210 healthy individuals (men: n = 89, women: n = 121), mean age, 39.8 years (standard deviation = 15.5; range = 6-76 years), were imaged on a 3T scanner. Mean MRI phase, mean phase of low-phase voxels, and normalized volumes were determined for total subcortical deep gray matter, caudate, putamen, globus pallidus, thalamus, pulvinar nucleus, hippocampus, amygdala, nucleus accumbens, red nucleus, and substantia nigra. Linear and nonlinear regression models were used to explore the relationship between phase and volume measures, and aging., Results: Mean phase values of subcortical deep gray matter structures showed a quadratic relationship, with individuals in late middle age (40-59 years) having the lowest mean phase values, followed by a reversal of this trend in the elderly. In contrast, mean phase of low-phase voxel measurements showed strong negative linear relationships with aging. Significantly lower phase values were detected in women compared with men (P < .001), whereas no sex differences were observed for mean phase of low-phase voxels. Normalized volume measurements were also linearly related to aging, and women showed smaller normalized volumes of subcortical deep gray matter structures than men (P < .001). Lower mean phase of low-phase voxels was related to decreased volume measures., Conclusions: A strong association between phase (quadratic effect; phase decreases are followed by increases), mean phase of low-phase voxels (linear effect), volume (linear effect), and age was observed. Low phase was related to brain atrophy.

Background: Activity of disease in patients with multiple sclerosis (MS) is monitored by detecting and delineating hyper-intense lesions on MRI scans. The Minimum Area Contour Change (MACC) algorithm has been created with two main goals: a) to improve inter-operator agreement on outlining regions of interest (ROIs) and b) to automatically propagate longitudinal ROIs from the baseline scan to a follow-up scan., Methods: The MACC algorithm first identifies an outer bound for the solution path, forms a high number of iso-contour curves based on equally spaced contour values, and then selects the best contour value to outline the lesion. The MACC software was tested on a set of 17 FLAIR MRI images evaluated by a pair of human experts and a longitudinal dataset of 12 pairs of T2-weighted Fluid Attenuated Inversion Recovery (FLAIR) images that had lesion analysis ROIs drawn by a single expert operator., Results: In the tests where two human experts evaluated the same MRI images, the MACC program demonstrated that it could markedly reduce inter-operator outline error. In the longitudinal part of the study, the MACC program created ROIs on follow-up scans that were in close agreement to the original expert's ROIs. Finally, in a post-hoc analysis of 424 follow-up scans 91% of propagated MACC were accepted by an expert and only 9% of the final accepted ROIS had to be created or edited by the expert., Conclusion: When used with an expert operator's verification of automatically created ROIs, MACC can be used to improve inter- operator agreement and decrease analysis time, which should improve data collected and analyzed in multicenter clinical trials.

Purpose: To investigate characteristics of cine phase contrast-calculated cerebrospinal fluid (CSF) flow and velocity measures in patients with relapsing-remitting (RR) multiple sclerosis (MS) receiving standard medical treatment who had been diagnosed with chronic cerebrospinal venous insufficiency (CCSVI) and underwent percutaneous transluminal angioplasty (PTA)., Materials and Methods: This case-controlled, magnetic resonance (MR) imaging-blinded study included 15 patients with RR MS who presented with significant stenoses (≥50% lumen reduction on catheter venography) in the azygous or internal jugular veins. Eight patients underwent PTA in addition to medical therapy immediately following baseline assessments (case group) and seven had delayed PTA after 6 months of medical therapy alone (control group). CSF flow and velocity measures were quantified over 32 phases of the cardiac cycle by a semiautomated method. Outcomes were compared between groups at baseline and at 6 and 12 months of the study by mixed-effect model analysis., Results: At baseline, no significant differences in CSF flow or velocity measures were detected between groups. At month 6, significant improvement in flow (P<.001) and velocity (P = .013) outcomes were detected in the immediate versus the delayed group, and persisted to month 12 (P = .001 and P = .021, respectively). Within-group flow comparisons from baseline to follow-up showed a significant increase in the immediate group (P = .033) but a decrease in the delayed group (P = .024). Altered CSF flow and velocity measures were associated with worsening of clinical and MR outcomes in the delayed group., Conclusions: PTA in patients with MS with CCSVI increased CSF flow and decreased CSF velocity, which are indicative of improved venous parenchyma drainage., (Copyright © 2013 SIR. Published by Elsevier Inc. All rights reserved.)

Objectives: To determine the sensitivity and specificity of decreased venous vasculature visibility (VVV) on susceptibility-weighted imaging (SWI) venography in multiple sclerosis (MS) patients versus controls, and to compare this with assessment of whole brain atrophy., Methods: Forty MS patients and 22 controls without known central nervous system (CNS) disease who had non-specific white-matter (WM) lesions were imaged on a 3T GE scanner using SWI venography. Apparent total venous volume (ATVV) and increased average distance from vein (DFV) were calculated for various vein mean diameter categories: <0·3, 0·3-0·6, 0·6-0·9, and >0·9 mm. Principal component analysis (PCA) was used to identify potential discriminatory metrics. Receiver operating characteristics (ROC) of these metrics, along with normalized brain volume (NBV), were calculated to determine sensitivity and specificity values between the groups. The efficacy of the metrics was validated against blinded data from 14 MS patients and 8 controls who had non-specific WM lesions., Results: PCA identified 0·3-0·6 mm venous relative fraction (VRF) and DFV as useful metrics. ROC analysis results in initial sample of 40 MS patients and 22 controls were (sensitivity, specificity): 0·3-0·6 mm VRF (95·0%, 100·0%); DFV (100·0%, 100·0%); and NBV (82·5%, 68·2%). The results in validation sample were: 0·3-0·6 mm VRF (92·9%, 75·0%); DFV (100·0%, 100·0%); and NBV (78·6%, 75·0%)., Discussion: Altered VVV indices on SWI venography showed high sensitivity and specificity for MS. The value of SWI venography for diagnosis of MS has to be further tested at early disease stages and against patients with other neurologic diseases.

Purpose: To investigate cerebrospinal fluid (CSF) dynamics in the aqueduct of Sylvius in multiple sclerosis (MS) patients and healthy controls (HC) using cine phase contrast imaging., Materials and Methods: In all, 67 MS patients (48 relapsing-remitting [RR] and 19 secondary-progressive [SP]), nine patients with clinically isolated syndrome (CIS), and 35 age- and sex-matched HC were examined. CSF flow and velocity measures were quantified using a semiautomated method and compared with clinical and magnetic resonance imaging (MRI) disease outcomes., Results: Significantly decreased CSF net flow was detected in MS patients compared to HC (-3.7 vs. -7.1 μL/beat, P = 0.005). There was a trend for increased net positive flow between SP, RR, and CIS patients. Altered CSF flow and velocity measures were associated with more severe T1 and T2 lesion volumes, lateral and fourth ventricular volumes, and third ventricular width in MS and CIS patients (P < 0.01 for all). In CIS patients, conversion to clinically definite MS in the following year was related to decreased CSF net flow (P = 0.007). There was a trend between increased annual relapse rate and altered CSF flow/velocity measures in RRMS patients (P < 0.05)., Conclusion: CSF flow dynamics are altered in MS patients. More severe clinical and MRI outcomes in RRMS and CIS patients relate to altered CSF flow and velocity measures., (Copyright © 2012 Wiley Periodicals, Inc.)

Background and Purpose: Increasing evidence suggests that iron deposition is present in the later stages of MS. In this study we examined abnormal phase values, indicative of increased iron content on SWI-filtered phase images of the SDGM in CIS patients and HC. We also examined the association of abnormal phase with conventional MR imaging outcomes at first clinical onset., Materials and Methods: Forty-two patients with CIS (31 female, 11 male) and 65 age and sex-matched HC (41 female, 24 male) were scanned on a 3T scanner. Mean age was 40.1 (SD = 10.4) years in patients with CIS, and 42.8 (SD = 14) years in HC, while mean disease duration was 1.2 years (SD = 1.3) in patients with CIS. MP-APT, NPTV, and normalized volume measurements were derived for all SDGM structures. Parametric and nonparametric group-wise comparisons were performed, and associations were determined with other MR imaging metrics., Results: Patients with CIS had significantly increased MP-APT (P = .029) and MP-APT volume (P = .045) in the pulvinar nucleus of the thalamus compared with HC. Furthermore, the putamen (P = .004), caudate (P = .035), and total SDGM (P = .048) displayed significant increases in MP-APT volume, while MP-APT was also significantly increased in the putamen (P = .029). No global or regional volumetric MR imaging differences were found between the study groups. Significant correlations were observed between increased MP-APT volumes of total SDGM, caudate, thalamus, hippocampus, and substantia nigra with white matter atrophy and increased T2 lesion volume (P < .05)., Conclusion: Patients with CIS showed significantly increased content and volume of iron, as determined by abnormal SWI-phase measurement, in the various SDGM structures, suggesting that iron deposition may precede structure-specific atrophy.

Contrast optimization of a three-dimensional (3D) Fluid-attenuated inversion recovery (FLAIR) sequence is examined in the context of multiple sclerosis. In order to develop 3D FLAIR for enhanced detection of lesions, an iterative approach based on theoretical considerations was used. The 3D FLAIR sequence was systematically acquired with incremental parameter changes on a single subject with multiple sclerosis in a 3-T MRI scanner. Region of interest analysis was performed on the images, and the best estimate of the optimal acquisition parameters was established. This process was iterated on subsequent patients with multiple sclerosis until only marginal gains in quality were obtained. Three-dimensional FLAIR for enhanced detection produced increased lesion detection and identification in the supratentorial and infratentorial regions when compared with default GE 3D FLAIR and two-dimensional FLAIR sequences. Lesion border delineation was enhanced as evidenced by better lesion and white matter contrast ratios, demonstrating that the approach is a viable method for improved lesion detection., (Copyright © 2011 Wiley Periodicals, Inc.)

Background and Purpose: It is well known that patients with MS tend to have abnormal iron deposition in and around the MS plaques, in the basal ganglia and the THA. In this study, we used SWI to quantify iron content in patients with MS and healthy volunteers., Materials and Methods: Fifty-two patients with MS were recruited to assess abnormal iron content in their basal ganglia and THA structures. One hundred twenty-two healthy subjects were recruited to establish a baseline of normal iron content in deep GM structures. Each structure was separated into 2 regions: a low-iron-content region and a high-iron-content region. The average phase, the percentage area, and the total phase of the high-iron-content region were evaluated. A weighting was also assigned to each subject depending on the level of iron content and its deviation from the normal range., Results: A clear separation between iron content in healthy subjects versus patients with MS was seen. For healthy subjects 13% and for patients with MS 65% showed an iron-weighting factor >3 SDs from the normal mean (P < .05). The results for those patients younger than 40 years are even more impressive. In these cases, only 1% of healthy subjects and 67% of patients with RRMS showed abnormally high iron content., Conclusions: Iron-weighting factors in the basal ganglia, THA, and the midbrain appeared to be abnormal in roughly two-thirds of patients with MS as measured by SWI.

Background and Purpose: CCSVI was recently described in patients with MS. CCSVI is diagnosed noninvasively by Doppler sonography and invasively by catheter venography. We assessed the role of conventional MRV for the detection of IJV anomalies in patients with MS diagnosed with CCSVI and in healthy controls who underwent MRV and Doppler sonography examinations during 6 months., Materials and Methods: Ten patients with MS underwent TOF, TRICKS, Doppler sonography, and catheter venography at baseline. They were treated at baseline with percutaneous angioplasty and re-evaluated 6 months' posttreatment with MRV and Doppler sonography. In addition, 6 healthy controls underwent a baseline and a 6-month follow-up evaluation by Doppler sonography and MRV., Results: At baseline, the sensitivity, specificity, PPV, and NPV of Doppler sonography for detecting IJV abnormalities relative to catheter venography in patients with MS were calculated, respectively, at 82%, 100%, 99%, and 95%. The figures were 99%, 33%, 33%, 99% for TOF and 99%, 39%, 35%, and 99% for TRICKS. Venous anomalies included the annulus, septum, membrane, and malformed valve. No agreement was found between TOF and catheter venography in 70% of patients with MS and between TRICKS and catheter venography in 60% of patients with MS. At follow-up, 50% of the patients with MS presented with abnormalities on Doppler sonography but only 30% were diagnosed with restenosis., Conclusions: Conventional MRV has limited value for assessing IJV anomalies for both diagnostic and posttreatment purposes.

Background: Several studies have reported hypoperfusion of the brain parenchyma in multiple sclerosis (MS) patients. We hypothesized a possible relationship between abnormal perfusion in MS and hampered venous outflow at the extracranial level, a condition possibly associated with MS and known as chronic cerebrospinal venous insufficiency (CCSVI)., Methods: We investigated the relationship between CCSVI and cerebral perfusion in 16 CCSVI MS patients and 8 age- and sex-matched healthy controls. Subjects were scanned in a 3-T scanner using dynamic susceptibility, contrast-enhanced, perfusion-weighted imaging. Cerebral blood flow (CBF), cerebral blood volume (CBV) and mean transit time (MTT) were measured in the gray matter (GM), white matter (WM) and the subcortical GM (SGM). The severity of CCSVI was assessed according to the venous hemodynamic insufficiency severity score (VHISS) on the basis of the number of venous segments exhibiting flow abnormalities., Results: There was a significant association between increased VHISS and decreased CBF in the majority of examined regions of the brain parenchyma in MS patients. The most robust correlations were observed for GM and WM (r = -0.70 to -0.71, P < 0.002 and P corrected = 0.022), and for the putamen, thalamus, pulvinar nucleus of thalamus, globus pallidus and hippocampus (r = -0.59 to -0.71, P < 0.01 and P corrected < 0.05). No results for correlation between VHISS and CBV or MTT survived multiple comparison correction., Conclusions: This pilot study is the first to report a significant relationship between the severity of CCSVI and hypoperfusion in the brain parenchyma. These preliminary findings should be confirmed in a larger cohort of MS patients to ensure that they generalize to the MS population as a whole. Reduced perfusion could contribute to the known mechanisms of virtual hypoxia in degenerated axons.

In β-thalassemia major (β-TM) patients, iron chelation therapy is mandatory to reduce iron overload secondary to transfusions. Recommended first line treatment is deferoxamine (DFO) from the age of 2 and second line treatment after the age of 6 is deferiprone (L1). A multicenter randomized open-label trial was designed to assess the effectiveness of long-term alternating sequential L1-DFO vs. L1 alone iron chelation therapy in β-TM patients. Deferiprone 75 mg/kg 4 days/week and DFO 50 mg/kg/day for 3 days/week was compared with L1 alone 75 mg/kg 7 days/week during a 5-year follow-up. A total of 213 thalassemia patients were randomized and underwent intention-to-treat analysis. Statistically, a decrease of serum ferritin level was significantly higher in alternating sequential L1-DFO patients compared with L1 alone patients (p = 0.005). Kaplan-Meier survival analysis for the two chelation treatments did not show statistically significant differences (log-rank test, p = 0.3145). Adverse events and costs were comparable between the groups. Alternating sequential L1-DFO treatment decreased serum ferritin concentration during a 5-year treatment by comparison to L1 alone, without significant differences of survival, adverse events or costs. These findings were confirmed in a further 21-month follow-up. These data suggest that alternating sequential L1-DFO treatment may be useful for some β-TM patients who may not be able to receive other forms of chelation treatment.

Aim: Chronic cerebrospinal venous insufficiency (CCSVI) is a vascular condition characterized by anomalies of primary veins outside the skull that restrict normal outflow of blood from the brain. CCSVI was recently described as highly prevalent in patients with multiple sclerosis (MS), and can be non-invasively diagnosed by Doppler sonography (DS) and invasively by selective venography (SV). The aim of this paper was to investigate the value of neck magnetic resonance venography (MRV) for the diagnosis of CCSVI compared to DS and SV in patients with MS and in healthy controls (HC)., Methods: Ten MS patients and 7 HC underwent DS, 2D-Time-Of-Flight venography (TOF) and 3D-Time Resolved Imaging of Contrast Kinetics angiography (TRICKS). MS patients also underwent SV. The internal jugular veins (IJVs) and the vertebral veins (VVs) were assessed by both MRV sequences, and the findings were validated against SV and DS. SV has been considered the diagnostic gold standard for MS patients., Results: All MS patients and none of the HC presented CCSVI, according to the DS criteria. This was confirmed by SV. For CCSVI diagnosis, DS showed sensitivity, specificity, accuracy, PPV and NPV of 100%, whereas the figures were 40%, 85%, 58%, 80% and 50% for 3D-TRICKS, and 30%, 85%, 52%, 75% and 46% for 2D-TOF in the IJVs. In MS patients, compared to SV, DS showed sensitivity, specificity, accuracy, PPV and NPV of 100%, 75%, 95%, 94% and 100%, whereas the figures were 31%, 100%, 45%, 100% and 26% for 3D-TRICKS and 25%, 100%, 40%, 100% and 25% for 2D-TOF in the IJVs., Conclusion: The use of MRV for diagnosis of CCSVI in MS patients has limited value, and the findings should be interpreted with caution and confirmed by other imaging techniques such as DS and SV.

Aim: Chronic cerebrospinal venous insufficiency (CCSVI) is a vascular phenomenon recently described in multiple sclerosis (MS) that is characterized by stenoses affecting the main extracranial venous outflow pathways and by a high rate of cerebral venous reflux that may lead to increased iron deposition in the brain. Aim of this study was to investigate the relationship between CCSVI and iron deposition in the brain of MS patients by correlating venous hemodynamic (VH) parameters and iron concentration in deep-gray matter structures and lesions, as measured by susceptibility-weighted imaging (SWI), and to preliminarily define the relationship between iron measures and clinical and other magnetic resonance imaging (MRI) outcomes., Methods: Sixteen (16) consecutive relapsing-remitting MS patients and 8 age- and sex-matched healthy controls (HC) were scanned on a GE 3T scanner, using SWI., Results: All 16 MS patients fulfilled the diagnosis of CCSVI (median VH=4), compared to none of the HC. In MS patients, the higher iron concentration in the pulvinar nucleus of the thalamus, thalamus, globus pallidus, and hippocampus was related to a higher number of VH criteria (P<0.05). There was also a significant association between a higher number of VH criteria and higher iron concentration of overlapping T2 (r=-0.64, P=0.007) and T1 (r=-0.56, P=0.023) phase lesions. Iron concentration measures were related to longer disease duration and increased disability as measured by EDSS and MSFC, and to increased MRI lesion burden and decreased brain volume., Conclusion: The findings from this pilot study suggest that CCSVI may be an important mechanism related to iron deposition in the brain parenchyma of MS patients. In turn, iron deposition, as measured by SWI, is a modest-to-strong predictor of disability progression, lesion volume accumulation and atrophy development in patients with MS.

The increase of Hb A(2) (α2δ2) beyond the upper limit [2.0-2.2/3.3-3.4% of the total hemoglobin (Hb)] is an invaluable tool in the hematological screening of β-thalassemia (β-thal) carriers. Factors decreasing Hb A(2) percentages can hinder correct diagnosis. In order to analyze the genotype-phenotype relationship, we characterized δ-, β- and α-globin genotypes in 190 families where the probands had Hb A(2) values of ≤2.0% or were β-thal heterozygotes with normal Hb A(2) levels. Hb A(2) was measured with cation exchange high performance liquid chromatography (HPLC). Mutations were detected with allele-specific methods or DNA sequencing; two multiplex-ARMS (amplification refractory mutation system) assays were set up. The molecular basis underlying the decrease in Hb A(2) was extremely heterogeneous. Nineteen δ-globin alleles (Hb A(2)-S.N. Garganico was new) were detected; their interaction with α- or β-globin alleles (10 and eight, respectively) led us to observe 52 genotypes in 261 carriers. The type of δ-globin mutations, the relative genotypes, the interaction with α(0)-thal traits, are the most important factors in decreasing the Hb A(2) percentage. These results are extremely useful in addressing the molecular diagnosis of hemoglobinopathies and thalassemias.

The prognosis for thalassemia major has dramatically improved in the last two decades. However, many transfusion-dependent patients continue to develop progressive accumulation of iron. This can lead to tissue damage and eventually death, particularly from cardiac disease. Previous studies that investigated iron chelation treatments, including retrospective and prospective non-randomised clinical trials, suggested that mortality, due mainly to cardiac damage, was reduced or completely absent in patients treated with deferiprone (DFP) alone or a combined deferiprone-deferoxamine (DFP-DFO) chelation treatment. However, no survival analysis has been reported for a long-term randomised control trial. Here, we performed a multicenter, long-term, randomised control trial that compared deferoxamine (DFO) versus DFP alone, sequential DFP-DFO, or combined DFP-DFO iron chelation treatments. The trial included 265 patients with thalassemia major, with 128 (48.3%) females and 137 (51.7%) males. No deaths occurred with the DFP-alone or the combined DFP-DFO treatments. One death occurred due to graft versus host disease (GVHD) in a patient that had undergone bone marrow transplantation; this patient was censored at the time of transplant. Only one death occurred with the DFP-DFO sequential treatment in a patient that had experienced an episode of heart failure one year earlier. Ten deaths occurred with the deferoxamine treatment. The main factors that correlated with an increase in the hazard ratio for death were: cirrhosis, arrhythmia, previous episode of heart failure, diabetes, hypogonadism, and hypothyroidism. In a Cox regression model, the interaction effect of sex and age was statistically significant (p-value<0.013). For each increasing year of age, the hazard ratio for males was 1.03 higher than that for females (p-value<0.013). In conclusion, the results of this study show that the risk factors for predicting mortality in patients with thalassemia major are deferoxamine-treatment, complications, and the interaction effect of sex and age.

A multicentre randomized open-label trial was designed to assess the effectiveness of long-term sequential deferiprone-deferoxamine (DFO-DFP) versus DFP alone to treat thalassaemia major (TM). DFP at 75 mg/kg, divided into three oral daily doses, for 4 d/week and DFO by subcutaneous infusion (8-12 h) at 50 mg/kg per day for the remaining 3 d/week was compared with DFP alone at 75 mg/kg, administered 7 d/week during a 5-year follow-up. The main outcome measures were differences between multiple observations of serum ferritin concentrations. Secondary outcomes were survival analysis, adverse events, and costs. Consecutive thalassaemia patients (275) were assessed for eligibility; 213 of these were randomized and underwent intention-to-treat analysis. The decrease of serum ferritin levels during the treatment period was statistically significant higher in sequential DFP-DFO patients compared with DFP-alone patients (P = 0.005). Kaplan-Meier survival analysis for the two chelation treatments did not show any statistically significant differences (long-rank test, P = 0.3145). Adverse events and costs were comparable between the groups. The trial results show that sequential DFP-DFO treatment compared with DFP alone significantly decreased serum ferritin concentration during treatment for 5 years without significant differences regarding survival, adverse events, or costs.

The human delta-globin gene (HBD) is one of the beta-like globin genes expressed in adults. In the Mediterranean countries the carriers of delta-thalassemia defects or Hb A2-variants are >1% and about 40/70 known alleles have been found in families with this ethnic origin. The scope of this study was to investigate the variability of the gene and of the chromosomal background in order to highlight the origin and spreading of the delta-globin gene alleles in the Mediterranean area. We carried out the characterization of the delta-globin gene alleles and of RFLP-haplotypes, SNPs and one microsatellite associated with them in 231 carriers originating principally from East Sicily. Seventeen alleles were identified, of which five were new. The chromosomes associated with mutated alleles from unrelated carriers were 158; the allele Hb A2-Yialousa accounted for about 75% of relative frequency, Hb A2-Mitsero for about 8%. The alleles were associated with RFLP 5'-haplotypes "- - - -" or "+ - + +", prevalent in the Mediterranean area, except Hb A2-Mitsero associated with the 5'-haplotype "Benin" "- - - +" and the Hb A2' associated with "+ - - +", both of African origin. Each allele showed linkage with one haplotype with these exceptions. The Hb A2-Yialousa showed heterogeneity of the 5'-haplotype in 2/58 chromosomes; the Hb A2-Mitsero showed SNPs and (A)gamma-microsatellite typical of a "Benin" haplotype found associated with the Hb C and Hb S chromosomes; the Hb A2-Yialousa (14/58 chromosomes), Hb A2-Mitsero, Hb A2-Pylos, Hb A2-Fitzroy showed heterogeneity in the 3'-haplotypes and beta-globin gene SNPs. The Hb A2-Coburg was found associated with the haplotype "+ - + +/+ +" different from that already reported "- - - -/+ -". With the exception of this last allele, the linkage of each mutation with a core of RFLPs or SNPs around or inside the delta-globin locus suggested the unicentric origin of the mutations followed by recurrent recombination events causing the chromosomal background heterogeneity.

Deferasirox (ICL670) is a once-daily oral iron chelator developed for the treatment of chronic iron overload from blood transfusions. A comparative phase 3 trial was conducted to demonstrate the efficacy of deferasirox in regularly transfused patients with beta-thalassemia aged 2 years or older. Patients were randomized and received treatment with deferasirox (n = 296) or deferoxamine (n = 290), with dosing of each according to baseline liver iron concentration (LIC). The primary endpoint was maintenance or reduction of LIC; secondary endpoints included safety and tolerability, change in serum ferritin level, and net body iron balance. In both arms, patients with LIC values of 7 mg Fe/g dry weight (dw) or higher had significant and similar dose-dependent reductions in LIC and serum ferritin, and effects on net body iron balance. However, the primary endpoint was not met in the overall population, possibly due to the fact that proportionally lower doses of deferasirox relative to deferoxamine were administered to patients with LIC values less than 7 mg Fe/g dw. The most common adverse events included rash, gastrointestinal disturbances, and mild nonprogressive increases in serum creatinine. No agranulocytosis, arthropathy, or growth failure was associated with deferasirox administration. Deferasirox is a promising once-daily oral therapy for the treatment of transfusional iron overload.

Background and Objectives: During the last decade new approaches to the treatment of pulmonary arterial hypertension (PH) have increased symptomatic relief and prolonged survival. PH is a common sequel of the hemoglobinopathies, thalassemia and sickle cell anemia, but the use of standard oral treatment options, such as calcium channel blockers, endothelin receptor antagonists, and long-term anticoagulation therapy, is limited because of toxicity and poor effectiveness. Sildenafil citrate is a selective and potent inhibitor of cGMP-specific phosphodiesterase-5 (PDE5) which promotes selective smooth muscle relaxation in lung vasculature and has been utilized successfully in the treatment of PH. The primary objective of this study was to evaluate the efficacy of sildenafil treatment in the control of PH in patients with hemoglobinopathies., Design and Methods: In this study patients with hemoglobinopathies (thalassemia intermedia n=4; thalassemia major n=2; sickle thalassemia n=1) suffering from severe PH were treated with sildenafil citrate (50 mg b.i.d.) for periods ranging from 4 to 48 months., Results: A significant decrease in pulmonary pressure and improvement in exercise capacity and functional class were observed in all patients. No significant adverse events were reported., Interpretation and Conclusions: These data, in a small group of patients, indicate that sildenafil citrate is effective in the treatment of PH in hemoglobinopathies that cannot be treated with alternative oral drugs and is well tolerated long-term at a daily dose of 100 mg, though studies including more patients may uncover toxicities and limitations of efficacy.

Serum non-transferrin-bound iron (NTBI) levels assessed at arbitrary time points during transfusion cycles may not be representative if NTBI is undergoing significant changes during transfusion cycles. In 15 patients with beta-thalassemia major (age: 21 +/- 6 years, liver iron concentration: 2200 +/- 1200 microg/g-liver), NTBI and other hematologic parameters (transferrin saturation, transferrin receptor) were measured weekly. The largest variation of NTBI levels between individual patients was observed at midcycle. For long-term monitoring of NTBI levels, a particular time point relative to the last blood transfusion should be selected for blood drawing.

Background and Objectives: Updated information on liver disease in transfusion-dependent beta-thalassemia is lacking. We conducted a multicenter study within the Cooleycare Group to describe the clinical and histopathological features of liver disease in currently treated thalassemics., Design and Methods: Two-hundred and three thalassemics with laboratory signs of liver disease were eligible. Liver biopsy was performed in the 129 (63.5%) who consented (age 26+/-7 years). Biological samples were sent to the central laboratory., Results: Anti-hepatitis C virus (HCV) antibodies were found in 118 patients (91%), 85 (72%) of whom were viremic. Ninety-one patients (70%) had abnormal aminotransferase concentrations. In the 117 liver biopsies that met the criteria for evaluation (88%), the median Ishak's necroinflammatory and fibrosis scores were 4 (range, 0-9) and 2 (range, 0-6), respectively. Significant fibrosis (score >or=3) was found in 53 (45%); 9 (8%) had cirrhosis. At multivariate analysis, necroinflammation was related to HCV viremia, and fibrosis to increased serum aminotransferases, higher iron stores (including serum ferritin, Deugnier's total iron score, and liver iron content) and male gender (p<0.05). In HCV-RNA negative subjects, the median total iron score was 27 (range, 0-52). Iron accumulated in both mesenchymal cells and hepatocytes, and the presence of a lobular gradient was interpreted to indicate intestinal hyperabsorption., Interpretation and Conclusions: Transfusion-dependent thalassemics have mild liver necroinflammation, mainly attributable to HCV infection. Significant fibrosis is frequent, and its progression is mostly influenced by iron overload which, with current therapy regimens, may be attributable to both erythrocyte catabolism and iron hyperabsorption.

Background: The cardiac function in thalassemia major has never been studied at ultrasonic backscatter techniques. We assessed the utility of acoustic densitometry in thalassemic patients without clinical or echocardiographic signs of heart failure., Methods: Three groups of subjects with comparable age, sex and body surface area were analyzed: 25 with beta-thalassemia major (group A), 14 with thalassemia intermedia (group B) and 10 healthy subjects (group C). All patients were asymptomatic and without conventional echocardiographic signs of systo-diastolic dysfunction. The left ventricular mass and volumes were echocardiographically evaluated. The ultrasonic myocardial integrated backscatter signal (IBS) was recorded and analyzed by means of acoustic densitometry in the parasternal long-axis view at the septum and posterior wall, both at the basal and intermediate levels. Both the average image intensity and the systo-diastolic variations of the IBS (cyclic variation index-CVIibs and peak-to-peak intensity-PPI), respectively related to the structure and contractility of the myocardium, were calculated. The serum ferritin and liver iron concentrations were also measured, as markers of tissue iron storage., Results: The CVIibs was significantly lower in groups A and B than in group C at basal (22.7 +/- 8.4 vs 22.1 +/- 7.8 vs 31.8 +/- 10.2%; p = 0.001) and intermediate septum (24.4 +/- 7.6 vs 25.3 +/- 8.1 vs 30 +/- 9.8%; p = 0.03) and at basal (25.9 +/- 7.6 vs 24.5 +/- 6.1 vs 31.1 +/- 10.6%; p = 0.02) and intermediate posterior wall (25.1 +/- 5.1 vs 24.3 +/- 6.2 vs 30.2 +/- 6.6%; p = 0.02). The PPI was also significantly lower in groups A and B than in group C. Both CVIibs and PPI were comparable in groups A and B. The average image intensity and left ventricular mass and volumes were not significantly different in the three groups. No correlation was found between the densitometric findings and markers of tissue iron storage., Conclusions: In asymptomatic patients with thalassemia major with normal conventional indexes of systo-diastolic cardiac function, acoustic densitometry may show a reduced cyclic variation of the IBS as a possible marker of initial myocardial contractile deficiency. On the contrary, neither structural alterations nor the extent of myocardial iron stores are detectable by this technique in this type of patients.

The purpose of this study was to evaluate if the variations of heart magnetic resonance imaging in beta-thalassemia major patients treated with Deferoxamine B mesylate (DF) or Deferiprone (L1) chelation therapy is a useful tool of the indirect myocardial iron content determination. For this reason, a prospective study was carried out. Seventy-two consecutive patients with beta-thalassemia major (35 treated with DF and 37 with L1) were studied. The main outcome results were laboratory parameters including determination of the liver iron concentration (LIC) and magnetic resonance imaging (MRI) of the heart and liver. The heart to muscle signal intensity ratios (HSIRs) were significantly increased in both the DF (t = -2.8; p < 0.01) and L1 (t = -3.1; p < 0.01) groups after one year of treatment No statistically significant difference in the values of HSIRs was present between the two groups at the beginning of treatment (p = 0.25; t = 1.13), and after one year of treatment (p = 0.20; t = 1.28). The HSIR were inversely correlated to the LIC (r = -0.52; p < 0.001) but not with ferritin levels (r = 0.10; p = 0.18). A positive correlation was found between the variation of HSIRs and that of the liver signal intensity ratios (r=0.52; p < 0.001), and a mild correlation (r = 0.40; p < 0.001) was found between the gamma glutamyltransferase (gammaGt) levels and the HSIRs values. Our data confirm that heart MRI is sensitive enough to detect significant variations of the mean HSIR during iron chelation with DF or L1.

Deferiprone has been suggested as an effective oral chelation therapy for thalassemia major. To assess its clinical efficacy, we compared deferiprone with deferoxamine in a large multicenter randomized clinical trial. One-hundred forty-four consecutive patients with thalassemia major and serum ferritin between 1500 and 3000 ng/ml were randomly assigned to deferiprone (75 mg/kg/day) (n = 71) or deferoxamine (50 mg/kg/day) (n = 73) for 1 year. The main measure of efficacy was the reduction of serum ferritin. Liver and heart iron contents were assessed by magnetic resonance. Liver iron content and fibrosis stage variations were assessed on liver biopsy by the Ishak score in all patients willing to undergo liver biopsy before and after treatment. The mean serum ferritin reduction was 222 +/- 783 ng/ml in the deferiprone and 232 +/- 619 ng/ml in the deferoxamine group (P = 0.81). No difference in the reduction of liver and heart iron content was found by magnetic resonance between the two groups. Thirty-six patients accepted to undergo repeat liver biopsy: 21 in the deferiprone and 15 in the deferoxamine group. Their mean reduction of liver iron content was 1022 +/- 3511 microg/g of dry liver and 350 +/- 524, respectively (P = 0.4). No difference in variation of the Ishak fibrosis stage was observed between the two groups. Treatment was discontinued because of reversible side effects in 5 patients in the deferiprone group (3 hypertransamin/asemia and 2 leukocytopenia) and in none in the deferoxamine group. These findings suggest that deferiprone may be as effective as deferoxamine in the treatment of thalassemia major with few mild and reversible side effects.

Hb G-San Josè or beta7(A4)Glu-->Gly has been reported in Southern Italian or Mexican families. We have studied four families from Sicily and Campania, Southern Italy. In six carriers, the hemoglobin variant level ranged from 32 to 38%. In four double heterozygotes for Hb G-San Josè and alpha-thalassemia the variant level showed a strong correlation with the alpha-thalassemia genotype. In fact, the variant level was 15% when interacting with the - (alpha)20.5/alphaalpha, 19.6% with the alphaalpha/alphaPoly Aalpha, and 24.8% with alphaalpha/alpha(-5) ntalpha genotypes. In two double heterozygotes for Hb G-San Josè and beta+ -IVS-I-6 (T-->C) the hemoglobin variant level was 67%. These data show that the reduced synthesis of alpha chains causes drastic reduction of probability to form Hb G-San Josè in favor of the formation of Hb A. Moreover, this reduction, (i) correlates with the type of alpha-thalassemia genotype and with the degree of the alpha chain deficiency, and (ii) is, most probably, more marked than the degree of alpha chain reduction. The minor affinity of the beta chain variant for the alpha chains associated with the reduced synthesis of the alpha chains is probably the principal cause of the variant hemoglobin reduction. Moreover, the rapid removal of the abnormal chains by proteolytic enzymes must have an essential role in order to reduce the chain variant pool. These conclusions are in agreement with the results obtained in reticulocyte and in vitro recombination experiments.

To evaluate the risk of transmitting blood-borne GB virus C/hepatitis G virus (GBV-C/HGV) and to define the natural course of infection, we performed a prospective study in a cohort of multitransfused beta-thalassemics during a 6-year follow-up period. We analyzed serum samples of 150 patients collected at 3-year intervals from 1990 to 1996. GBV-C/HGV RNA was determined by reverse transcriptase-polymerase chain reaction and antibodies to E2-protein by an enzyme immunoassay. At baseline, 14.5% of patients had viremia and 18.5% anti-E2. None of the patients with anti-E2 in 1990 subsequently became viremic. Of the 100 GBV-C/HGV RNA-, anti-E2- patients, 10 acquired infection during follow-up, as indicated by positivity of GBV-C/HGV RNA (n = 2), anti-E2 (n = 7), or both markers (n = 1) in 1996. The incidence was 1.7 per 100 person-years (95% confidence interval [CI], 0.8 to 3). Since approximately 19,000 blood units were transfused to these patients during follow-up, the risk of infection was 5.3 in 10,000 units (95% CI, 2 to 8.5). Six of 22 viremic patients cleared the virus during follow-up; 4 of them became anti-E2+. Twelve of 28 patients lost anti-E2 reactivity during follow-up. In conclusion, more than 25% of infections resolve within 6 years; the presence of anti-E2 seems to be protective against infection. Anti-E2 reactivity may decrease with time.

To assess whether cocaine exposure in utero affected adrenocortical responsiveness in the neonatal period, salivary cortisol levels were compared between noncocaine-exposed (n = 35) and cocaine-exposed (n = 11) healthy preterm infants just prior to hospital discharge. Cortisol levels were measured under three conditions: Basal--120 min after no disrupting event or behavioral distress; Noninvasive Stressor--30 min after a neurobehavioral examination; Invasive Stressor--30 min after a heel-stick procedure. There were no differences in Basal cortisol levels between the noncocaine-exposed and cocaine-exposed infants, but the cocaine-exposed infants had significantly lower levels in both the Noninvasive and Invasive Stressor conditions. The suppressed cortisol responding to stressful events in cocaine-exposed infants suggests that these infants may have decreased modulation capability to normally stressful events, which could underlie some of the subtle state regulation problems reported.

The authors have evaluated the skull, thorax, wrist and hand standard radiographs of 29 patients affected by Cooley's disease who were treated up to 1983 with a low transfusion regimen, rather with a high transfusion regimen (Hb level greater than 9.5 g/dl). The patients have been divided into two groups, according to the prevalence of the first or the second transfusion regimen. Almost all the changes described in the literature have been found in the radiographs, but completely modified; some of these, like rib notchings, in higher percentage than known. In the youngest patients the most significant appearances were ribs and hand changes, that can be considered the most important feature in their radiological monitoring. However this new transitional radiological syndrome will rapidly modify with the establishment "ab initio" of the high transfusional regimen.

Based on a review of the literature and personal observation, the authors discuss the most frequent endocrine complication that occur in patients with thalassemia major given blood transfusion and chelating therapy. All the complication are attributed to iron overload in the endocrine glands. The authors conclude that an adequate chelating therapy protects against hemosiderosis and thus against endocrine pathologies.

In order to investigate possible contamination of saliva samples with formula or breast milk obtained from young infants for cortisol assays, we measured cortisol concentrations in 3 common infant milk formulas and in breast milk before and after defatting and extraction using a commercial radioimmunoassay kit. Cortisol concentrations obtained prior to defatting and extraction were equal to or higher than (1.64-82.80 micrograms/dL) levels reported for salivary cortisol levels reported in newborn infants (0.09-2.08 micrograms/dL). Cortisol concentrations obtained after the defatting and extraction were lower (0.02-0.15 micrograms/dL), indicating that values obtained prior to defatting and extraction were due to cross-reacting substances as well as cortisol. As saliva samples are not routinely defatted or extracted prior to being assayed, high cortisol levels and interfering substances in formula and breast milk could contaminate salivary cortisol measurements in young infants. The present study suggests that appropriate controls should be taken when making salivary cortisol measurements in young infants to help ensure accurate results.

The relationship between cranial ultrasonograms (SONOs) and brain-stem auditory evoked responses (BAERs) was evaluated in 2 independent samples of newborn infants at risk for brain injury (n = 113 and 203). Features of the BAER wave forms subjected to stepwise linear discriminant analysis formed the basis of an algorithm used to detect and follow early brain injury. Using this algorithm, information derived from BAERs reliably predicted SONO abnormalities at least 82.3% of the time in the initial study which was replicated with the second sample (77.3%). The wave I component latency (CL) and the wave III-V inter-peak latency interval (IPL) were independent of each other, and both contributed to a prediction of SONO abnormality. Possible mechanisms for these BAER results include compromise to the cochlear membrane or to the auditory nerve itself as well as prolongation of transmission in the brain-stem due to brain-stem hemorrhage, edema, or compression. Normative BAER values and non-linear regression functions for the wave I, III and V CLs, and the I-III, III-V, and I-V IPLs were calculated across age using data from 109 infants who demonstrated normal BAER patterns and had no history of SONO abnormalities. Our analyses indicate BAER techniques, where a single higher intensity is used to produce the BAER wave form, are both valid and efficient for use in the evaluation of early brain injury.