To determine the factors associated with retinal nerve fiber layer (RNFL) loss in eyes with acute primary angle-closure (APAC), particularly focusing on the influence of the change in the anterior lamina cribrosa surface depth (LCD).After the initial presentation, 30 eyes with unilateral APAC were followed up at the following specific time points over a 12-month period: 1 week, 1~2 months, 2~3 months, 5~6 months, and 11~12 months. These follow-ups involved intraocular pressure measurements, enhanced depth-imaging spectral-domain optical coherence tomography (SD-OCT) scanning of the optic disc, and measurements of the circumpapillary RNFL thickness. The prelaminar tissue thickness (PLT) and LCD were determined in the SD-OCT images obtained at each follow-up visit.p<0.001). The global RNFL thickness decreased continuously throughout the follow-up period, while the PLT decreased until 5~6 months and did not change thereafter. The LCD reduced until 2~3 months and then also remained steady. Multivariable regression analysis revealed that symptoms with a longer duration before receiving laser peripheral iridotomy (LI) (p = 0.049) and a larger LCD reduction (p = 0.034) were significant factors associated with the conversion to an abnormal RNFL thickness defined using OCT normative data.Repeated measures analysis of variance revealed a significant pattern of decrease in the global RNFL thickness, PLT, and LCD (all Early short-term decreases in the PLT and LCD and overall long-term decrease in the peripapillary RNFL were observed during a 12-month follow-up after an APAC episode. A longer duration of symptoms before receiving LI treatment and larger LCD reduction during follow-up were associated with the progressive RNFL loss. The LCD reduction may indicate a prior presence of significant pressure-induced stress that had been imposed on the optic nerve head at the time of APAC episode. Glaucomatous progression should be suspected in eyes showing LCD reduction after the APAC remission. Acute primary angle closure (APAC) is characterized by an acute, symptomatic intraocular pressure (IOP) increase followed by rapid recovery from the symptoms after applying timely treatment. Although APAC generally does not result in significant visual morbidity if it is treated promptly, delayed optic nerve damage still can occur even after the remission of acute episodes.–5 HoweveChanges in the optic nerve head (ONH) in APAC have been described as an initial swelling and hyperemia that resolves after applying timely treatment, or is followed by pallor of the optic disc with diffuse thinning of the axons when the acute episode leads to permanent sequelae in the ONH.–10 HowevThe aim of the study was to elucidate longitudinal changes in the deep ONH and peripapillary tissues after an APAC episode, and to correlate them with the development of RNFL loss after APAC remission.This investigation was based on the data of APAC patients obtained in an ongoing prospective study of patients with glaucoma and healthy subjects being conducted at the Seoul National University Bundang Hospital Glaucoma Clinic, Seoul, South Korea (the Angle Closure Glaucoma Prognosis Study). All of the participants provided written informed consent. This study was approved by the Seoul National University Bundang Hospital Institutional Review Board and followed the tenets of the Declaration of Helsinki.To be included in this study, patients had to have been diagnosed with APAC and had been followed up for at least 12 months after the remission of the APAC episode. The APAC diagnosis was based on the following criteria: (1) presThe following exclusion criteria were applied: (1) secondary angle closure, such as lens-induced glaucoma, neovascular glaucoma, or uveitic glaucoma; (2) pre-existing diagnosis of any forms of glaucoma, (3) presence of coexisting retinal or neurologic disease; (4) history of previous intraocular surgery, or intraocular surgery during the follow-up period; (5) poor-quality spectral-domain optical coherence tomography (SD-OCT) scans of the optic disc  in more than 5 sections that did not allow precise measurement of the ONH parameters ; and (6) best-corrected visual acuity of <20/50 after remission.Measurements of the ONH Parameters). Forty-two APAC patients who met the eligibility criteria were initially recruited. Of these, 12 were excluded due to the poor scan quality preventing precise measurements of the ONH parameters , administration of oral medications, and other systemic diseases. Ocular examinations included visual acuity assessment, Goldmann applanation tonometry, refraction tests, slit-lamp biomicroscopy, gonioscopy, and undilated stereoscopic examination of the optic disc. The patients also underwent color fundus photography , SD-OCT circumpapillary retinal nerve fiber layer (RNFL) scanning, SD-OCT optic disc scanning using the enhanced depth-imaging (EDI) technique , and measurements of corneal curvature , central corneal thickness , and axial length .The participants were followed up at 1 week (FU1), 1~2 months (FU2), 2~3 months (FU3), 5~6 months (FU4), and 11~12 months (FU5) after the initial visit. At each follow-up the participants underwent visual acuity assessment, Goldmann applanation tonometry, slit-lamp biomicroscopy including evaluation of the peripheral anterior chamber depth using the van Herick technique, undilateThe IOP was recorded at the time of the APAC onset, within 30 minutes after LI, and then at every follow-up visit until FU5. The mean and standard deviation values of the follow-up IOP measurements were defined as the mean follow-up IOP and the IOP fluctuation, respectively. IOP re-elevation during the subsequent follow-up was defined when the IOP increased to >21mmHg.The ONH was imaged using the Spectralis SD-OCT system with the EDI technique. The details of the protocol for scanning of the optic nerve using EDI SD-OCT to evaluate the LC are described elsewhere., 28At the time of ONH scanning, the circumpapillary RNFL thickness was also measured using a circular scanning protocol, the details of which have been published elsewhere., 30 The This study measured the prelaminar tissue thickness (PLT), the anterior LC surface depth relative to the Bruch’s membrane (BM) opening (BMO) level (LCD), the juxtapapillary choroidal thickness (JCT), and the BMO diameter on the EDI SD-OCT scans that had been obtained at each visit. Measurements were also performed for contralateral eyes when SD-OCT results were available for both eyes.The PLT, LCD, and JCT were measured in five horizontal B-scans  that divided the optic disc into six equal parts vertically .The LCD was determined by measuring the distance from the BMO plane to the level of the anterior LC surface.–33 A refBecause the LCD measured from the BMO level is influenced by the JCT,, 35 we sMeasurements of the PLT, LCD, and JCT at each time point were performed using the manual caliper tool of the Spectralis viewer , in as close to the same plane as possible. The low reflective shadow within the LC as well as the choroidal shadow was examined to confirm the correspondence of the B-scan image series between images., 33The BMO diameter was defined as the mean of the longest and shortest BMO diameters. The longest and shortest BMO diameters were the longest and shortest distances between the two termination points of the BM, which were measured on the radial B-scan images constructed from the 3 dimensional data set of EDI SD-OCT scan. Measurements were made using the manual caliper tool of the Amira 5.2.2 software .All measurements were performed by two independent observers (E.J.L. and K.M.L.) who were blinded to the clinical information of the participants, including the IOP and the time point of scanning for the follow-up SD-OCT images. Measurements were repeated twice by the two observers, and the mean of the four values was used for the main analysis.The interobserver reproducibility for the measurements of ONH parameters and JCT was evaluated using the intraclass correlation coefficients.t-tests were used to assess changes in ONH measurements during the follow-up. Repeated-measures p values were calculated using the Greenhouse-Geisser correction. Intereye comparisons were performed using paired-samples t-tests. A logistic regression analysis was performed to investigate the factors associated with the conversion to an abnormal OCT RNFL thickness.Repeated-measures multivariable and univariable analysis of variance as well as paired samples p<0.05.Analyses were performed using the Statistical Package for the Social Sciences . Unless stated otherwise, the data are presented as mean±standard deviation values, and the cutoff for statistical significance was set at The duration of symptoms associated with the APAC episode recorded dependent on the patient history was 16.3±19.6 hours  in three patients and at FU2  in the other three. However, IOP normalized in these patients after the cataract surgery. Twenty-five of the 30 eyes received cataract surgery during the follow-up period according to the patients’ preference, once they had been informed about the effects of lens extraction on the anterior chamber angle anatomy and IOP control in eyes with primary angle closure.–38 None EDI SD-OCT optic disc scanning could not be performed at the time of the APAC attack due to corneal edema in all except one eye, and the circumpapillary RNFL thickness could only be evaluated in six eyes. Thus, the SD-OCT measurements made 1 week after the APAC attack were considered as the baseline values for the longitudinal evaluation. The interobserver agreements for the PLT, LCD, JCT, and BMO diameter measurements were good, with intraclass correlation coefficients of 0.978, 0.991, 0.931, and 0.967, respectively.vs. 118.5±34.5 μm, p = 0.031; Wilcoxon signed-rank test). The PLT showed an initial decrease at FU2 and a further decrease at FU4, and then remained steady until FU5 (p<0.001), while neither JCT and nor the BMO diameter showed a significant change .ntil FU5 . The LCDntil FU5 . There wntil FU5 . Both rep = 0.195), global RNFL thickness (p = 0.717), PLT (p = 0.604), LCD (p = 0.581), JCT (p = 0.949), or BMO diameter (p = 0.514)—differed between FU1 and FU5. The intereye comparisons revealed that the global RNFL thickness was larger in the APAC eyes at FU1 (p<0.001) but gradually decreased during the follow-up and became significantly thinner at FU5 , while the difference disappeared at FU5 , but it decreased during the follow-up and became comparable to the LCD of the contralateral eyes , and a larger LCD reduction between the initial visit and FU5  in the univariable analysis (p = 0.049) and the LCD reduction  were significant factors.At FU5, the SD-OCT circumpapillary RNFL thickness measurements revealed a conversion to an abnormal OCT RNFL thickness in 13 out of 30 eyes (43.3%). Logistic regression analysis showed that the conversion to an abnormal OCT RNFL thickness was significantly associated with a longer duration from the symptom onset to receiving LI treatment [odds ratio (OR) = 1.155, analysis . In the The present study has revealed longitudinal changes in the deep ONH tissues and peripapillary RNFL that are present for 1 year after an APAC episode. A short-term reduction of the PLT and LCD during the early follow-up period, and a long-term sustained decrease in the RNFL thickness during the 1-year follow-up were noted after an APAC attack. Eyes with a longer duration from the symptom onset to receiving LI treatment, and a larger LCD reduction during the follow-up were more likely to show progressive RNFL loss. To the authors’ knowledge, this is the first study to show longitudinal changes in the deep ONH tissues after APAC and their influence on the progressive RNFL loss.P = 0.028, Wilcoxon signed-rank test), and then exhibited a continuous decrease from FU1 to FU5, with 40% of the participants showing RNFL loss below the normal range.The initial increase followed by a subsequent decrease in the peripapillary RNFL thickness after APAC has been well described., 39–41 IThe PLT decreased notably during the earlier follow-ups, but did not change at later follow-ups. Given that both the RNFL and prelaminar tissue consist mainly of retinal ganglion cell axons, thinning of the RNFL is thought to accompany a decrease in the PLT. However, this was not true in the later follow-up, when RNFL thinning was not associated with any significant PLT change. This finding is in accordance with a previous finding of the absence of optic disc cupping despite thinning of the RNFL after APAC resolution. We attriIn the present study, the LCD reduced significantly during the earlier follow-ups and then did not change during the later follow-ups. This is comparable to the pattern of longitudinal change in the LCD after trabeculectomy in OAG patients. It is invs. 110.68 μm),[vs. 16.7 mmHg). This may be attributable to the initial disc scan being performed 1 week after LI treatment, when some LCD reduction might have already occurred. The EDI SD-OCT images were not obtainable at the time of the APAC attack in most of the patients due to the presence of corneal edema, which is one of the main limitations of this study (see below). It is also possible that because the IOP elevation is highly symptomatic in APAC, and also decreases rapidly, the duration of IOP elevation is shorter in APAC than in chronic glaucoma, and such a short-term IOP elevation may not induce a large LC displacement as is observed in chronic glaucoma. Alternatively, acute IOP lowering may have induced shrinkage of the scleral canal, leading to the relief of the tensile force that had been stretching the LC within the scleral canal, which in turn moved the LC posteriorly and thus lessened the LCD reduction.[It is noteworthy that the LCD reduction at final FU was smaller in APAC patients than in the OAG eyes included in our previous study (28.84 0.68 μm), despite eduction., 45 AlthIt has been suggested that the LCD measured from the BMO level is influenced by the JCT,, 35 suchWhile the results have been equivocal and variable, a longer duration of symptoms,, 5, 9 grIn contrast to the findings of this study, Jiang et al. did not This study was subject to several limitations. Firstly, the ideal study would perform imaging investigations at the time of the onset of APAC, but this was not possible due to the presence of corneal edema. If SD-OCT images were obtainable at the time of the APAC attack, the change in the ONH parameters might have been more distinct and the effect of the change in glaucoma development might have been clearer. Secondly, our sample may have been too small to detect factors that exerted small effects on glaucoma progression. Thirdly, the participants might have experienced episodes of intermittent angle closure or had a pre-existent chronic angle closure before the APAC episode. Thus, although eyes with pre-existing diagnosis of glaucoma were excluded, some individuals could have had ongoing RNFL damage before the acute attack. This may have influenced the interpretation of the results relating to RNFL changes after APAC. Since the RNFL thickness before APAC attack was not known, the progressive RNFL loss could not be determined based on the amount of RNFL loss relative to its baseline status , but was defined based on the SD-OCT findings at final examination. If the amount of RNFL loss during the follow-up had been considered for defining the progressive RNFL loss, it could not have been differentiated from the resolution of RNFL swelling after APAC attack. Fourthly, 25 of the 30 patients underwent cataract surgery during the follow-up period, which might have influenced the quality of SD-OCT images and caused measurement. However, most of the eyes had mild to moderate degree of cataract, and so any effects on the results might have been minimal. Fifthly, gonioscopy was not performed at all FU points in all patients, thus could not be considered in the analyses. Lastly, the information on the onset of symptom was obtained by self-reporting, which might not have been reliable.In conclusion, early short-term reductions of the PLT and LCD and an overall long-term decrease in the peripapillary RNFL were observed during a 1-year follow-up after an APAC attack. A longer duration from the symptom onset to receiving LI treatment and a larger LCD reduction during the follow-up were associated with the conversion to an abnormal OCT RNFL thickness. The LCD reduction may indicate that a significant level of IOP-induced stress had been imposed on the ONH at the time of APAC episode. Glaucomatous progression should be suspected in eyes showing LCD reduction after the APAC remission.S1 Table(XLSX)Click here for additional data file.


Most people are exposed to at least one traumatic event during the course of their lives, but large numbers of people do not develop posttraumatic stress disorders. Although previous studies have shown that repeated and chronic stress change the brain’s structure and function, few studies have focused on the long-term effects of acute stressful exposure in a nonclinical sample, especially the morphology and functional connectivity changes in brain regions implicated in emotional reactivity and emotion regulation. Forty-one months after the 5/12 Wenchuan earthquake, we investigated the effects of trauma exposure on the structure and functional connectivity of the brains of trauma-exposed healthy individuals compared with healthy controls matched for age, sex, and education. We then used machine-learning algorithms with the brain structural features to distinguish between the two groups at an individual level. In the trauma-exposed healthy individuals, our results showed greater gray matter density in prefrontal-limbic brain systems, including the dorsal anterior cingulate cortex, medial prefrontal cortex, amygdala and hippocampus, than in the controls. Further analysis showed stronger amygdala-hippocampus functional connectivity in the trauma-exposed healthy compared to the controls. Our findings revealed that survival of traumatic experiences, without developing PTSD, was associated with greater gray matter density in the prefrontal-limbic systems related to emotional regulation. Trauma exposure is common and could increase lifetime vulnerability to mental health problems when individuals encounter stress or adversity, especially the most traumatic events, such as a massive earthquake, terrorism or war –3. AlthoExtensive neuroimaging studies of patients with mood disorders have shown significant alternations in brain structure and function –16, whicRather than mass univariate analyses, multivariate-pattern-analysis (MVPA) is sensitive to spatially distributed effects and can be used to separate patients from the healthy controls using structural MRI (magnetic resonance imaging) data or the functional MRI data , 44. PreIn the present study, more than 3 years after the 5/12 Wenchuan earthquakes, we used voxel-based morphometry (VBM) and resting state fMRI (rs-fMRI) to investigate the changes in brain structure and functional connectivity between the stress-related brain areas in trauma survivors compared to healthy controls.p = 0.24), sex (p = 0.58) and length of education (p = 0.62) . Two participants were tested but excluded from the VBM analysis due to problems in the image registration.A total of forty-two healthy undergraduate students participated in this study about forty-one months after the 5/12 Wenchuan earthquake. Twenty-one individuals  from the Wenchuan earthquake disaster area were identified by the post-traumatic stress disorder self-rating scale (PTSD-SS) (with score < 60) as resilient trauma survivors . The PTSThe Impact of Event Scale Revised (IES-R) is a self-administered, 22-item questionnaire as indicators of PTSD. It should be administered due to trauma and have no other medical basis, and perhaps it is the most widely used self-administered assessment in the field of traumatic stress. The IES-R was administered after scanning for all subjects to record their current subjective distress. They were told: “Please read each item, and then indicate how distressing each difficulty has been for you during the past 7 days with respect to wenchuan earthquake .” The IEThe Spielberger State-Trait Anxiety Inventory (STAI)  was usedAll MRI data were performed on a 3T scanner  at the Brain Imaging Research Central at Southwest University. First, resting-state functional MR images were obtained using an Echo Planar Imaging (EPI) sequence with the following parameters: time repetition [TR] = 2000 ms; time echo [TE] = 40 ms; flip angle [FA] = 90, slices = 28, matrix = 64×64; field of view [FOV] = 192 mm; acquisition voxel size = 3.4 × 3.4 × 4 mm. A total of 242 volumes were collected for each subject. During fMRI scanning, subjects were instructed to close their eyes, not to move, think about anything particular, or fall asleep , 51). Eahttp://resting-fmri.sourceforge.net/). This included the following steps: the first 10 functional images were discarded due to signal instability and the subject’s adaptation to the scanning noise. The slices of the remaining 232 volumes for each subject were corrected for different collection times of signals by slice time correction. Subsequently, the functional image time series were motion-corrected by realigning all images to the middle image volume. The individual structural image of each subject was co-registered to the mean functional image generated after motion correction. Third, the functional images were spatially normalized into the Montreal Neurological Institute (MNI) space using the transformation information generated by segmentation and resampled into 3 mm cubic voxels. The normalized images were smoothed with an isotropic Gaussian kernel (FWHM = 4 mm).The resting-state data preprocessing was performed using a Matlab  toolbox Data Processing Assistant for Resting-State fMRI . For better image registration, the reorientation of the images was manually set to the anterior commissure. The structural image was then segmented into gray matter (GM), white matter (WM), and cerebrospinal fluid (CF). Subsequently, we performed diffeomorphic anatomical registration through exponentiated lie (DARTEL) algebra in SPM8 for registration, normalization and modulation. Then, the registered images were transformed to Montreal Neurological Institute (MNI) space. Finally, the normalized, non-modulated images (gray matter and white matter density images) were smoothed with a 10-mm full-width at half-maximum Gaussian kernel to increase the signal to noise ratio.The structural MRI images were processed using SPM8 software  approach was used to test the gray matter density (GMD) differences between the trauma-exposed group and the healthy controls. To control for the effects of age, gender, and whole brain gray matter density, these variables were added as additional covariates. We also applied absolute threshold masking  to avoid edge effects between gray and white matter. To explore whether there ware structural differences in the amygdala, parahippocampal and dACC, which have been considered to be involved in emotion regulation and stress responsively –24, smalThe PRONTO toolbox based on pattern classification techniques was used for the analysis of neuroimaging data , 57. Thez-transformed r value) between the trauma survivors and the healthy controls. To investigated whether the functional connectivity  would be affected by the IES-R scores between the earthquake exposed group and the control group, we calculated the Pearson correlation coefficients between the z-scores of each ROI-to-ROI and the IES-R scores with controlling the sex and age as regressors of no interest.Functional connectivity was performed by applying a seed region correlation approach  using thThe levels of anxiety were within the normal range across all groups, and the STAI scores were not significantly different between the trauma survivors and healthy controls . Age, seCompared with the healthy controls, the trauma survivors showed that greater GMD in the dorsal medial frontal cortex (dmPFC), dorsal anterior cingulate cortex (dACC) extended to the rostral anterior cingulated cortex (rACC) and bilaterally in the hippocampus/amygdala and lower GMD of the frontoparietal association cortex .We also used MVPA to further investigate the predictive ability of regional gray matter density at the individual level, based on the GMD differences map (GMD map) and regions of interest obtained from the univariate analysis. The applications of SVM to dmPFC clusters classified the trauma-exposed and healthy control groups with a sensitivity of 71.4% and 73.7%, respectively, leading to an overall accuracy of 72.5%; dACC ; hippocampus/amygdala . When all these clusters were considered simultaneously, it classified the trauma-exposed and healthy control groups with a sensitivity of 100% and 68.4%, respectively, leading to an overall accuracy of 85%. Permutation testing indicated that these predictive accuracies were all statistically significant see .t = 2.86; p = 0.007, r = 0.45; p = 0.04, A group comparison analysis revealed that there were no significant differences in the functional connection of the amygdala with the mPFC between the two groups . However, an increased strength of the connection of the amygdala with the hippocampus was found in the trauma survivors compared to the healthy controls  on human brain structure and functional connectivity. We found that resilient trauma-exposed survivors showed greater gray matter density in prefrontal-limbic brain systems and lower gray matter density in the frontoparietal association cortex than controls. Resting-state functional connectivtiy analysis found that resilient trauma-exposed survivors showed strengthened functional connectivity between the amygdala and hippocampus compared to controls.The prefrontal cortex plays an important role in top-down emotion regulation and it can regulate the neural activity of the limbic systems, especially the amygdala and hippocampus , 73–75. It has recently been highlighted that periods of traumatic stress might alter the structural and activity in the human parietal lobule , 80–83. Furthermore, trauma not only induces an anxious state and emotional arousal but can also impair memory through the amygdala’s interactions with other brain regions , 84, 85.However, we observed a significant positive correlation between the connectivity strength and distress. It is not consistent with the notion that greater connectivity serves as a protective role in developing PTSD. Thus, this correlation should generally be interpreted with caution. To begin with, the range of the symptom severity might not large enough to explore the brain-behavioral relationship. Perhaps there would be an inverse association between symptoms and hippocampus-amygdala connectivity in subjects with high-level symptoms. Besides, the trauma-exposed condition is characterized by other three typical symptom clusters: re-experiencing, avoidance and hyperarousal symptoms , 106. ItOur study revealed that resilient trauma survivors showed greater gray matter density in the prefrontal-limbic systems that were implicated in emotional regulation. The emotional regulation ability plays a critical role in preventing the onset of PTSD in those trauma-exposed nonclinical adults. However, there are two possible explanations for the current findings. One possibility is that these structural differences might be a pre-existing factor and those participants did not develop PTSD due to these biological protective factor. Nevertheless, we cannot rule out the possibility that the structural differences are the brain “scar” after the traumatic event. Our study is quite preliminary in nature and only longitudinal studies that examine the changes in behaviors and neuroimaging measures in individuals before and after the traumatic stress could fully rule out one of the possibilities.Of note, there are other limitations in our study. First at all, we did not include the participants who experienced the Wenchuan earthquake and then developed the PTSD in our study. This limitation restricted our ability to differentiate the resilience and vulnerability factor at the neural level. We expect to recruit an additional PTSD patients group and carry out the longitudinal project in the future. Besides, the sample size of our study was modest and the range of symptoms severity is limited. This point largely affects the reliability of our brain-behavior correlational analysis. Finally, the assessments of symptom in this study are not comprehensive enough. The assessments should be multiplex and include different symptoms clusters, such as re-experiencing, avoidance and hyperarousal symptoms along with a high rate of dissociative symptoms , 108.Our results are largely consistent with findings from studies on other stressors  in the literature –24, 77. S1 File(XLSX)Click here for additional data file.


When secondary causes are excluded, mechanisms underlying central nervous system angiitis (ACNS) in human immunodeficiency virus (HIV)-infected patients are still not understood and optimal treatment remains undefined. We report here a patient with an untreated HIV infection who presented multiple ischemic strokes probably due to HIV-ACNS. ACNS signs on vessel-wall imaging magnetic resonance monitoring retracted with combined antiretroviral therapy without adjunct immunosuppressive drugs. Between 1 and 5% of patients carrying the human immunodeficiency virus (HIV) will present a stroke and up to 30% will display a cerebral ischemic lesion on autopsy. In these cases, stroke can be a consequence of opportunistic infections, coagulation abnormalities, meningitis, and vasculitis. However, whether HIV-infected patients are predisposed to increased risk of stroke by a direct or indirect virus-associated vascular change or because of the atherogenic effect of antiretroviral agents remains unclear .in situ hybridization and electron microscopy) in the perivascular tissues of patients with HIV-ACNS  is one of the main atiologies of ischemic strokes in HIV-positive patients been accounted for 13–28% of the cases. ACNS in a HIV-positive patient can be secondary (to opportunistic infections) or due to HIV itself either directly or indirectly through modifications in the local immune and inflammatory response. Frequently, the vasculopathy is present without any other discernible disease leading to the diagnosis of HIV-associated ACNS (HIV-ACNS) . HIV-ACNHIV-ACNS . HoweverHIV-ACNS .The optimal treatment for HIV-ACNS remains unknown as only a limited number of case reports are available –13. In tHere, we present the case of a 56-year-old man with an untreated HIV infection presenting multiple ischemic strokes due to a probable case of HIV-ACNS. Vasculitis signs on serial vessel-wall imaging (VWI-MR) retracted with c-ART without adjunct immunosuppressive drugs suggesting that the pathophysiology of HIV-ACNS is more likely being directly related to the HIV itself than to a immune reaction.3 (15.69% lymphocytes) and HIV-1 viral load was 59,375 copies/ml.A 56-year-old man admitted at the emergency room (ER) with a recurrent left leg weakness occurring for more than 24 h. The individual chart indicated the presence of HIV-1 infection since 2012 but without antiretroviral treatment. Anamnesis revealed that the HIV was sexually transmitted and that the patient had never consumed drugs of abuse. On admission, the CD4+ cell count was 313/mmAdditionally, the patient had a history of syphilitic aortic aneurysm treated by Bentall surgery 10 years ago for which he is treated with an anti-vitamin K  and concomitantly presents multiple cardiovascular risk factor including obesity (BMI = 35), glucose intolerance, arterial hypertension, and hypercholesterolemia treated by amlodipine and simvastatin.The weakness in his left leg was first noticed while driving. He went to the ER where he recovered completely within 6 h and was discharged after an unremarkable brain computed tomography angiography. The next day, the patient felt again a left leg weakness and was admitted a second time at the ER.On this second admission, the general physical examination was within normal limits. Neurologic examination disclosed a left leg-predominant hemiparesis sparing the face associated with pyramidal syndrome. Brain MRI indicated restricted diffusion in the right anterior cerebral artery territory, compatible with a recent ischemic lesion, and lacunar lesions in the left temporo-occipital and right superior frontal region Figure . Such atThe combination of HIV infection and multiple ischemic strokes in different arterial territories associated with the enhancement of cerebral wall vessels both at the stroke territory and at other arteries emerging from Willis polygon led to the diagnosis of HIV-ACNS. cART (dolutegravir/abacavir/lamivudine) was immediately introduced but without adjunct immunosuppressive therapy. This was decided based on the patient’s immune status, the immediate and complete neurological recovery of this initial episode, and the absence of recurrence in the 4 days between the patient’s admission and the VWI-MR results.After the first month, most vessels did not show CE anymore Figures J–L. AfteThis study was carried out in accordance with the recommendations of Erasme University Hospital ethics committee with written informed consent from the patient for the publication of this case report.We reported here a case of a probable HIV-ACNS whose evolution was favorable while treated only with c-ART. The case also illustrates the potential contribution of VWI-MR to the diagnosis of ACNS and the monitoring of disease during treatment.The patient was admitted at the hospital with a recurrent left leg weakness occurring over 24 h, without neither headache nor alteration in vigilance, which are both usually seen as classical clinical manifestations of PACNS but only found in 54 and 27%, respectively, of patients . When heThe gold standard for the diagnosis of CNS vasculitis is histopathological evidence of vessel wall inflammation. However, a brain biopsy is an invasive technique and may reveal false negative findings, particularly in patients with predominant or isolated proximal large artery involvement . Thus, fThe CE is visualized with a particular technique based on a 3DT1-weighted TSE sequence (Black Blood), which better differentiate the vessel wall from the lumen blood signal.Recently, many studies tried to evaluate the performances of this technique in PACNS diagnosis and demonstrated that VWI could differentiate vasculitis from other vasculopathy-like atherosclerosis and RCVS offering an excellent specificity , 19. TheSalmonella associated with ACNS in HIV patients argued for primary involvement of the HIV in the vasculopathy of the patient (An analogy could be drawn between the radiological patterns of PACNS and HIV-ACNS, particularly regarding vessel abnormalities. Nevertheless, this patient presented certain differences with the classical MRI abnormalities found in PACNS. Indeed, the patient lacked both the gadolinium meningeal enhancement on MRI that is observed in 40.3% of PACNS  and the  patient , 8.Some authors have proposed, in case of vasculitis associated with HIV infection, to start a systemic immunosuppressive treatment in conjunction with c-ART. These recommendations are based on rare case reports , 11, 12 To the best of our knowledge, this is the first report demonstrating a complete retraction of vessel wall CE after cART initiation without additional immunosuppressive treatment in a so far cART-naive HIV patient. It highlights the potential role of dedicated wall vessel sequences on brain MRI for the diagnosis and follow-up of HIV-ACNS and its mimics. Further studies are required to confirm these preliminary results. The poor outcome, despite HIV-ACNS resolution, highlights the further complexity of the multiple comorbidities of HIV-infected patients.JC: conception, collection of the data, drafting of the article, critical revision, and final approval. CW-T: internistic follow-up of the patient, revision, and final approval. NS: radiological follow-up of the patient, revision of the manuscript and figure, and final approval. GN: neurological follow-up of the patient, critical revision of the manuscript, and final approval.The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest.


Resting state studies of spontaneous fluctuations in the functional magnetic resonance imaging (fMRI) blood oxygen level dependent signal have shown great potential in mapping the intrinsic functional connectivity of the human brain underlying cognitive functions. The aim of the present study was to explore the developmental changes in functional networks of the developing human brain exemplified with the language network in typically developing preschool children. To this end, resting-sate fMRI data were obtained from native Chinese children at ages of 3 and 5 years, 15 in each age group. Resting-state functional connectivity (RSFC) was analyzed for four regions of interest; these are the left and right anterior superior temporal gyrus (aSTG), left posterior superior temporal gyrus (pSTG), and left inferior frontal gyrus (IFG). The comparison of these RSFC maps between 3- and 5-year-olds revealed that RSFC decreases in the right aSTG and increases in the left hemisphere between aSTG seed and IFG, between pSTG seed and IFG, as well as between IFG seed and posterior superior temporal sulcus. In a subsequent analysis, functional asymmetry of the language network seeding in aSTG, pSTG and IFG was further investigated. The results showed an increase of left lateralization in both RSFC of pSTG and of IFG from ages 3 to 5 years. The IFG showed a leftward lateralized trend in 3-year-olds, while pSTG demonstrated rightward asymmetry in 5-year-olds. These findings suggest clear developmental trajectories of the language network between 3- and 5-year-olds revealed as a function of age, characterized by increasing long-range connections and dynamic hemispheric lateralization with age. Our study provides new insights into the developmental changes of a well-established functional network in young children and also offers a basis for future cross-culture and cross-age studies of the resting-state language network. Since the seminal study of Biswal et al.  resting-To date, LFFs analysis has featured several functional networks which are associated with corresponding cognitive functions. One LFFs network that has been identified in rs-fMRI data  and in With the application of the fMRI technique, it has become feasible to investigate the neural basis of language acquisition and processing, even in young infants . For exIt has been widely acknowledged that language processing is based on both ventral and dorsal processing streams –34 and tAs a powerful tool to investigate the spontaneous neuronal activity, rs-fMRI has great advantages in terms of acquiring data from young children who are not able to perform complicated tasks in the scanner, and characterizing functional brain networks independent of specific tasks. Therefore, the rs-fMRI technique is a sound solution to sketch the picture of the development of functional networks of the human brain. It avoids the bias in specific task selection and experimental design on the one hand, and allows exploring the general development at a network level on the other hand. Here, we selected the functional network of language-relevant brain regions because this network is well established and comprises of brain areas across lobes and hemispheres. It has been well-known that language ability develops very fast during the first years of life, and the underlying neural basis might be somewhat unveiled in respect of the language network development with age in early life.Considerable amount of evidence from language fMRI studies has shown the presence of language lateralization in children and its dynamic shift with age associated with developing language skills in children and adolescents . SzaflarIn the present study, RSFC of the language network was investigated in typically developing Chinese preschool children at ages 3 and 5 years given the rapid development during this time. Fifteen rs-fMRI data sets in each age group were acquired while children were in a natural sleep state. Intrinsic connectivity analysis was conducted with four language-related regions of interest (ROIs), including the left aSTG, left pSTG, right aSTG and left IFG, which had been shown to be activated in adult Chinese native speakers and chosen as ROIs in dynamic causal modeling . ExpectiThirty-six typically developing preschool children aged 3 and 5 years participated in this study, including 19 3-year-olds  and 17 5-year-olds , as reported in a separate analysis . All chiAll children were scanned during natural sleep. In order to ensure that children could easily fall asleep, MRI scanning was performed in the afternoons or evenings, and the day before the scanning, they were asked to sleep later in the evening and to get up earlier in the morning than usual. Children were carried into the MRI scanner after falling asleep in the waiting room, and they were accompanied by their parents and one of the experimenters during the course of scanning. The process would be terminated if they woke up. Finally, MRI data were acquired from 30 children, 15 in each age group .The MRI data acquisition was performed on a 3 Tesla Siemens MRI scanner (Siemens Magnetom Tim Trio) with a 12-channel head coil in Shenzhen Institutes of Advanced Technology, Chinese Academy of Sciences. Functional images were acquired using a T2*-weighted single-shot echo planar imaging (EPI) sequence . After the functional scanning, T1-weighted 3D structural images were obtained with magnetization prepared rapid gradient echo (MPRAGE) sequence . The whole scanning process lasted 15 minutes. For more details on the data acquisition protocol, see Xiao et al. .i) slice timing; ii) head motion correction using a least squares approach and a 6 parameter (rigid body) spatial transformation; iii) producing group-averaged templates via new segment and the diffeomorphic anatomical registration through exponentiated lie algebra (DARTEL) [iv) a component based noise reduction method (CompCor) was performed using the Functional Connectivity Toolbox (CONN) [http://web.mit.edu/swg/software.html) to correct for physiological noise by regressing out principal components (PC) from noise ROIs  [v) band-pass filtering (0.01–0.1 Hz).Before image preprocessing, the first seven EPI volumes were discarded to allow for signal equilibration, and the remaining 185 volumes were included in the analysis. We used the preprocessing procedure employed in a previous study (for details see ), includ(DARTEL) . Each grx (CONN)  (http://ocedure) , includit(28) = -0.55, p = 0.59).In order to further reduce the effect of head motion, framewise displacement (FD) was calculated for each participant following Jenkinson et al.  as suggeIn this analysis, four ROIs were selected based on a previous language study . These rhttp://www.restfmri.net), and voxel-wise functional connectivity was calculated for each ROI. Specifically, the mean time series of each seed were first computed for each participant by averaging the time series of all the voxels in the ROI (a sphere with 6 mm radius), and then the Pearson’s correlation coefficients between the time series of each seed and time series of all other regions within the whole brain were calculated. The individual level RSFC correlation map (r-map) was obtained for each subject, and subsequently, all r-maps were converted into z-maps with application of Fisher’s r-to-z transformation to get approximately normally distributed values for further statistical analysis.RSFC analysis was performed by using REST software ,  theory . Two-sample t-tests were performed for the comparison between 3- and 5-year-olds, controlling for gender and mean FD. Results were corrected at the cluster-level using GRF theory . Finally, all the maps were visualized with the BrainNet Viewer [http://www.nitrc.org/projects/bnv/).One-sample t Viewer ,  regions of these ROIs were defined as right aSTG, right pSTG, as well as right IFG, separately. RSFC maps of these regions were produced using the same procedure as aforementioned. The asymmetry of RSFC in the present study was defined in a same manner as shown in previous studies , 48. Firt-tests were performed to reveal regions which showed significant hemispheric asymmetry. More specifically, the left hemisphere of the t-maps represents the asymmetry of RSFC of left ROIs with the ipsilateral hemisphere , and the right hemisphere of the t-maps demonstrated the asymmetry of connectivity of left ROIs with the contralateral hemisphere .AI maps of all ROIs were calculated voxel by voxel across subjects for both 3- and 5-year-olds. One-sample t-tests. Finally, all the resultant t-maps were corrected for multiple comparison using GRF theory .Moreover, in order to investigate the hemispheric asymmetry changes with age, group comparison between AI maps from 3- and 5-year-olds was carried out for each seed using two-sample Functional connectivity maps seeding in the left aSTG, left pSTG, right aSTG and left IFG revealed similar distributed patterns of connections in both age groups . Left aSt-tests showed developmental changes in children from ages 3 to 5 years   . The conAmong the examined ROIs, functional connectivity of pSTG in both age groups and of IFG in 3-year-olds showed significant hemispheric asymmetry. Asymmetric connectivity with ipsilateral and contralateral hemispheres was obtainable from left and right hemispheres separately . In genet values within each significant cluster  and that of LR-flipped right ROIs . The functional asymmetry patterns are shown in t-value of 7.36 and 6.62 for left and right IFG, respectively), while it showed greater RSFC strength with right pSTG than with left pSTG in 5-year-olds. In addition, three  out of five clusters showed greater connectivity strength with left IFG than right IFG in 3-year-olds.In order to further evaluate the RSFC strength within each region that revealed significant hemispheric asymmetry, mean Furthermore, the group comparison for pSTG revealed increasing ipsilateral connectivity to SPL, angular gyrus, and MFG/SFG; it also demonstrated increasing contralateral connectivity to PCC and decreasing contralateral connectivity to SPL from ages 3 to 5 years . The IFGIn the present study, we employed a seed-based functional connectivity analysis to investigate the developmental changes of functional connectivity with brain regions belonging to the language network. Significant developmental changes were observed comparing preschool children at ages 3 and 5 years. We found stronger right aSTG RSFC in 3-year-olds than in 5-year-olds, and stronger connectivity between left IFG and left temporal cortex in 5-year-olds than in younger children when seeded in the left aSTG, pSTG and IFG. These results show that there is an increasing RSFC between left aSTG and left IFG, between left pSTG and left IFG, as well as between left IFG and left pSTS from ages 3 to 5 years. Moreover, functional hemispheric asymmetry analysis revealed that in 3-year-olds, RSFC of pSTG was almost bilateralized and RSFC of IFG showed leftward lateralization, while the RSFC of pSTG in 5-year-olds was mainly right lateralized. Both RSFC of IFG and of pSTG showed increasing left lateralization with age, but pSTG demonstrated a remaining dominance in rightward asymmetry at the age of 5 years. These findings suggest a clear developmental change of intrinsic brain connectivity in the language network in typically developing children towards a leftward lateralization.The functional lateralization of the language network was examined in order to explore the process of the language lateralization with age. RSFC of pSTG was bilateral in 3-year-olds, but showed unilateral asymmetry in 5-year-olds. The right hemispheric asymmetry in 5-year-olds supports the notion of the engagement of the right hemisphere in language processing , 17, 49 In contrast to the pSTG, RSFC of IFG demonstrated a trend of left lateralization in 3-year-olds and an increase of leftward asymmetry with age. This finding is not only in correspondence with previous rs-fMRI studies in adults that reported the left hemisphere dominance of the language network , 21, butHowever, neither significant hemispheric asymmetry in 3- and 5-year-olds, nor lateralization shift was observed in aSTG. Given the limited data in the present study, further research should shed light on this.As discussed above, conclusion cannot be drawn from the present data that decreasing right aSTG connectivity is attributed to functional lateralization changes during this period, since no significant functional asymmetry in group comparison was observed. Therefore, we would provide an alternative interpretation for this finding.Previous studies in adults have consistently reported activation of the right aSTG being involved in processing pitch information in tonal languages , 55, andThe increasing RSFC between aSTG and IFG in the left hemisphere suggests an enhancement of the ventral processing stream from 3 to 5 years of age. The ventral pathway has been found to be related to sound-to-meaning integration , and itsTherefore, in the present study, we infer that the increasing ventral pathway connectivity with age might be attributed to more demand of sound-to-meaning integration with increasing lexical and semantic input for children at the age of 5 years than children at the age of 3 years when less vocabulary is available. This is supported by a previous behavioral study which showed improvement in vocabulary and word recognition in Chinese preschool children from ages 3 to 6 years . IncreasThe current study also indicates age-related development of long-range functional connectivity between pSTG/STS and IFG in the left hemisphere, reflecting increasing connectivity in the dorsal pathway from ages 3 to 5 years. The finding of fronto-temporal functional connectivity can be related to the observation that the activation of left pSTG/STS has been consistently found to covary with IFG  during processing syntactically complex sentences in language fMRI studies , 64–67. In Chinese, there is a non-canonical construction , where the order of subject and object is reversed. The corresponding canonical construction is termed ‘ba’ construction with the normal order of subject and object. Apparently, the ‘bei’ construction should be more difficult to process than the ‘ba’ construction because it requires additional process for reversed word-order. Research has reported enhanced performance in response to the ‘bei’ construction during childhood , 69. ChaTherefore, we suppose that the increasing fronto-temporal connection in 5-year-old children might be related to the improvement in the processing of syntactic complexity.A few limitations of the current study should be taken into account. Firstly, the interpretation of the results should be confined to -rs-fMRI context. The data shown here were from rs-fMRI rather than task-based fMRI experiment or LFFs analyses based on these. Secondly, a direct relation to language test was not available. Such data would have allowed for further conclusions about behavioral consequences of the observed functional network changes. Thirdly, rs-fMRI data were acquired when children were naturally sleeping. Natural sleep state might differ from regular resting state, but it has been widely used in previous pediatric studies e.g., –11, 70),, 70,11, Considering aforementioned limitations, future studies should include more children in each age group to enhance the robustness, and it is also suggested to investigate more age groups to sketch a complete picture of the developmental trajectory of the language network. Furthermore, it would be interesting to directly identify the relationship between changes in intrinsic brain connectivity and changes in language performance, or to obtain further evidence from task-related fMRI data. In addition, cross-language studies using the -rs-fMRI technique would be essential because it will open new horizons in revealing the similarities and dissimilarities of developmental trajectories of the language network when exposed to different language types .Our data here provide primary evidence for the development and lateralization of the language network in preschool children from task-free fMRI data. We found increasing left language lateralization in IFG from 3- to 5-year-olds and rightward asymmetry for pSTG in 5-year-olds. Moreover, we showed decreasing right aSTG RSFC, and observed greater connections in the ventral pathway between left aSTG and left IFG and increasing connectivity in the dorsal pathway between left IFG and left pSTG/STS in children from ages 3 to 5 years. These results suggest developmental trajectories of intrinsic brain connectivity in the language network. Notably, the present study must leave open whether the decreasing right aSTG RSFC and increasing fronto-temporal connection in the left hemisphere are attributed to the development of language abilities in general, or to specific abilities required by Chinese children and the language they learn. Nevertheless, these findings provide a first basis for cross-language comparisons with children from the different language  environment in future research.


Sardinella aurita fishery off northeastern Venezuela, region of seasonal wind-driven coastal-upwelling, accounts for 90% of the Caribbean Sea small pelagic catch. This law-protected artisanal fishery takes place up to ~10 km offshore. The spatial distribution, number of schools, and biomass of S. aurita were studied using eight hydro-acoustic surveys (1995–1998). The study included the analysis of satellite-derived sea surface temperature and chlorophyll-a. Surveys were grouped by strong, weak, and transitional upwelling seasons. Relationships between these observations were analyzed using Generalized Additive Models. Results show that during the primary upwelling season (January-May) sardines were widely distributed in upwelling plumes that extended up to 70 km offshore. In the other hand, during the weak upwelling season (September-October) higher sardine densities were found within 10 Km off the coastal upwelling foci. The number of small pelagic schools was directly correlated with small pelagic densities; however, regardless of the season, higher numbers of small pelagic schools were always closer to the shoreline, especially during warm conditions. These two behaviors increase the availability and catchability of sardines for the artisanal fishery during the warm season, regardless of the total stock size. Using this evidence, we pose the hypothesis that the collapse of the regional S. aurita fishery in 2005 was due to a combination of stressful habitat conditions sustained since 2004. These included bottom-up factors due to food scarcity caused by weak upwelling, combined with top-down stress due to overfishing, as sardines accumulated in narrow diminished upwelling plumes located close to the coast. The increased catchability within easily accessible upwelling foci led to the demise of this biological resource, which as of 2014 had not yet recovered. Environmental conditions affecting the sardine habitat needs to be taken into account for the management of this stock. For example, during years with weak upwelling, special measures should be taken during the warm season on the second half of the year to avoid further pressure on the stock.The  Sardinella aurita) biomass of 0.6–1.3 million tons [The southern Caribbean Sea  is an arion tons , 6, 7. Tion tons . Despiteion tons , 9, 10.S. aurita responds to changes in their habitat due to the seasonal upwelling in the southeastern Caribbean Sea. Historical acoustic surveys [Changes in habitat conditions have profound impacts on fish spatial distribution . Our res surveys , 12–17 h surveys ). The VE surveys .in situ profiles of water temperature and chlorophyll-a concentration obtained during the VECEP surveys. Those analyses were considered challenging due to the aggregative behavior of sardines [In a previous analysis of the VECEP surveys, Cárdenas and Achury  found nosardines . Aggregasardines , 21. Thesardines , 22.The study area is located in the southeastern Caribbean Sea, off northeastern Venezuela , show sea surface temperatures ~1°C cooler than the central Caribbean Sea due to a weak coastal upwelling process , 19. The2 d-1; ). The di2 d-1; , 26.Spanish sardine density and spatial distribution were obtained from eight acoustic surveys performed during 1995–1998 with the R/V Hermano Ginés , under the Regional Technical Cooperation Fishing Programme VECEP  UE-VECEP ALA 92/43 between the European Union and Venezuela . We refe-1) and the elementary sampling distance unit (ESDU) was one nautical mile (1852 m). The tracks of each cruise were measured twice during day and night conditions. Calibrations of the echosounder were performed before every survey following the sphere protocol described in Foote et al. [Pelagic fish acoustic backscattering strength was measured with a split-beam echo sounder SIMRAD EK500 at 38 and 120 kHz. The study area is characterized by high plankton concentration and relatively shallow depths around the study area (from 10 to 150 m). Most of the processing was based on the data acquired at 120 kHz, since 120 kHz allows a better discrimination of fish echoes from plankton´s and it has a range of about 100 m, in deeper areas the 120 kHz system was mainly used as an aid in the discrimination of fish traces recorded by the 38 kHz system , 28. Thee et al. .A, with units in m2 nmi-2) [Sardinella aurita cannot be distinguished from other small pelagic by visual inspection of the echogram; therefore, its density estimation (sardine-NASC) was calculated by its proportion in the fish community, obtained as the average of the trawling fishing results within three delimited areas  . To minied areas : (1) Theed areas  and it ied areas , with uped areas . Open waed areas . When ex2 pixel) from the Advanced Very High Resolution Radiometer , were collected using a ground-based antenna located at the University of South Florida . SST was derived using the Multi-Channel Sea Surface Temperature (MCSST) split-window techniques [Satellite-derived Sea Surface Temperature (SST) and Chlorophyll-a concentration (Chl) were extracted for the study area . High rechniques , 33, 34.chniques , 36. AVHchniques . The filTo assess whether Spanish sardine has a tendency to aggregate close to upwelling or thermal fronts, we calculated SST gradients (SST-gradient) from the SST imagery covering each acoustic survey. SST-gradient was computed at each grid point  by subtracting the SST from the pixels to the north-south and east-west of the grid point using the following equation:2 pixel), produced with the default NASA OC4 chlorophyll algorithm [We used high resolution satellite Chlorophyll-a concentration , number of small pelagic schools (Schools) and sampling area number 1, 2, or 3 Area, . At eachTo overcome the challenge of the aggregation behavior of sardines and visualize the distribution of sardine-NASC related to the seasonal intensity and dispersion of upwelling (SST), we first computed histograms of SST from satellite data extracted along the cruise track. Then geometric average of sardine-NASC and average of Schools were calculated within each SST bin of 0.3°C.Generalized Additive Models (GAM) were also used to explore the relationship between sardine-NASC and the different variables. A geometric average of sardine-NASC was used because of the log-normal distribution of the sardine abundance. The GAMs analyses were performed using the ‘‘mgcv” library  for the The spatially-averaged SST during each of the VECEP cruises was well within or around one standard deviation of the long-term SST climatology  without an obvious relationship to SST . HoweverRegardless of season, the first four VECEP surveys  had significantly higher sardine-NASC biomass and lower variances than the last four surveys  to obtain the best model that explain the variations of the dependent variable. However, concurvity (non-linear multicollinearity) could produce unstable or inaccurate estimates of the covariates’ functional effects , 50. ConThe influence of some predictors can change when analyzed together or separately. For example, Jablonski and Legey  found anTo contrast the differences in sardine-NASC distribution during different phases of the seasonal upwelling cycle, we analyzed three separate seasons see Figs  and 4:  during the three upwelling seasons was always higher with the spatial location  than with SST or upw-foci  and 8. TSimilarly to the histogram analysis, GAM results showed higher number of small pelagic schools (Schools) in the cooler SST range Figs  and 9. TThe hour of the day was included as a categorical main factor to the GAM analysis, and the GAM summary showed the hour effect on the response variable. For the number of pelagic schools, during “cool” conditions the significant time period with higher number of schools was from 6 am to 8 pm; while during “warm” conditions that period was shorter, from 5 am to 5 pm. GAM analysis of the SST-gradient effects on sardine-NASC and Schools did not show any significance for any of the upwelling conditions .2 of 0.74, 0.64, and 0.71 at p<0.01, <0.05, and <0.01, respectively). Those three correlations had similar rate of change on the number of schools per NASC ; however, ESDUs closer to the upwelling foci had higher number of pelagic schools per biomass . This is expected, as not all the pelagic school detected were from S. aurita. When using the total sum of the pelagic-NASC and of the pelagic schools per cruise—instead of the averages—, correlations were much weaker ; and for the total sum of the sardine-NASC all correlations were no significant.There was no apparent correlation between the number of pelagic schools with pelagic-NASC or with sardine-NASC when we compared the ESDU values directly. This is a result of the highly spatial-temporal variability of the dataset. However, when comparing the cruise averages for these variables, a direct correlation between biomass and the number of schools was discernible . AverageSardinella aurita landings off northeastern Venezuela for 1990–2010 was 110,268 tons  of this stock, with lower CPUE during the principal upwelling (December-April) and higher CPUE during the rest of the year . AdditiSardinella aurita showed strong avoidance of the Orinoco River plume during September-October. No significant numbers of eggs and larvae have been found to the east of this front of fresher and warmer waters [S. aurita to a smaller area: (a) localized cells of upwelling very close to the coast, that induce Spanish sardine concentration closer to shore; and (b) a wider hypohaline front due to the discharge of the Orinoco River to the east, that causes a horizontal migration of the stock away from this front. A completely different scenario might occur during July, when the maximum freshwater extension of the Orinoco River coincides with the mid-year upwelling [r waters  because r waters , 60. Ther waters . Consequpwelling . UnfortuThe largest deviance for every VECEP survey was explained by geographic location . The samSardinella aurita showed a strong avoidance to the coolest temperatures .Higher abundance of small pelagics along thermal fronts has been found in the North Sea  and nortSardinella aurita. School number was directly related to the pelagic-fish density  ringens . The simSardinops sagax is compressed along the coast during summer (non-upwelling season), while it is located offshore during upwelling [Sardinops sagax) distribution is limited to small localized upwelling areas [Sardinops sagax) is found farther offshore during winter/upwelling [Similar preference in distribution tied to upwelling plumes has been reported for other small pelagics. In California, pwelling . During ng areas , 85. In pwelling , but thipwelling . All theSardinella aurita grow fast (K = 1.1 año-1) and reaches sexual maturity around 12–20 months of age [S. aurita biomass and number of schools tend to be higher closer to the coast, and therefore easier to be fished regardless of the total biomass of the stock. This information allows to prepare a management strategy when a weak upwelling is detected during the first trimester, regulating the stock fishery during the warm season on the second half of that year, in order to avoid further pressure on the stock or even overfishing.Off northeastern Venezuela s of age , 89, so Sardinella aurita, was broadly distributed within upwelling plumes that extended from the coast across the continental shelf. In contrast, during the warm season (September-October) Spanish sardine was concentrated close to the coast within the first 10 km from the upwelling foci. During this time, sardines avoided warm waters, areas farther than 30 km from the upwelling foci, and the Orinoco River plume. Regardless of the upwelling season, higher numbers of schools were found in the cooler waters closer to the coast, especially during warm upwelling conditions.Analyses of acoustic fisheries surveys conducted in the southeastern Caribbean Sea under the VECEP cooperative program showed that during upwelling conditions (December-April) the Spanish sardine, The seasonal changes in the distribution of sardine provide insight into the causes of the collapse of the sardine fishery in the region in 2004, a fishery that still had not recovered by 2014. Sardine availability and catchability for the artisanal coastal fishery increases during warm conditions. The southern Caribbean Sea experienced substantial warming and decreased upwelling after 2004 . One of Sardine reproduces fast, so even a limited moratorium of this species can help the fisheries recovery. Fisheries management needs to implement the monitoring and evaluation of environmental conditions affecting the habitat of sardines in order to produce sound regulations of the stock. For example, during years with weak upwelling in the first trimester, particular regulations should be taken during the second half of the year to avoid further pressure and the potential collapse of the sardine stock.


The ceRNAs are also called miRNA “decoys” or miRNA “sponges” and encompass different RNAs competing with each other to attract miRNAs for interactions: mRNA, long non-coding RNAs (lncRNAs), pseudogenes, or circular RNAs. Recently, we developed a computational method for identifying ceRNA-ceRNA interactions in breast invasive carcinoma. We were interested in unveiling which lncRNAs could exert the ceRNA activity. We found a drastic rewiring in the cross-talks between ceRNAs from the physiological to the pathological condition. The main actor of this dysregulated lncRNA-associated ceRNA network was the lncRNA PVT1, which revealed a net biding preference towards the miR-200 family members in normal breast tissues. Despite its up-regulation in breast cancer tissues, mimicked by the miR-200 family members, PVT1 stops working as ceRNA in the cancerous state. The specific conditions required for a ceRNA landscape to occur are still far from being determined. Here, we emphasized the importance of the relative concentration of the ceRNAs, and their related miRNAs. In particular, we focused on the withdrawal in breast cancer tissues of the PVT1 ceRNA activity and performed a gene expression and sequence analysis of its multiple isoforms. We found that the PVT1 isoform harbouring the binding site for a representative miRNA of the miR-200 family shows a drastic decrease in its relative concentration with respect to the miRNA abundance in breast cancer tissues, providing a plausibility argument to the breakdown of the sponge program orchestrated by the oncogene PVT1.Recent findings have identified competing endogenous RNAs (ceRNAs) as the drivers in many disease conditions, including cancers. The ceRNAs indirectly regulate each other by reducing the amount of microRNAs (miRNAs) available to target messenger RNAs (mRNAs). The ceRNA interactions mediated by miRNAs are modulated by a titration mechanism,  This  binding , 45–61.via the base-pairing with miRNA recognition/response elements (MREs) [Competing endogenous RNAs (ceRNAs), also known as miRNA “decoy” or miRNA “sponges”, are RNA transcripts that compete for the binding to the same miRNA s (MREs) –70, subss (MREs) .i.e. copies of real genes that originate from duplications or retro-transpositions) as competitors of their ancestral genes for miRNA binding [The first experimental evidence of lncRNAs acting as miRNA decoys modulating the derepression of miRNA targets has been found in wide variety of human cancers and specifically concerns the functioning of pseudogenes  called linc-MD1 controls muscle differentiation by targeting miR-133 and miR-135 to regulate the expression of MAML1 and MEF2C ; human eMost recently, also the new-appreciate circular RNAs (circRNAs) appear to exert ceRNA activity , 78–80. i.e. tissues that are adjacent to the tumour and taken from the same patient) of breast invasive carcinoma available from The Cancer Genome Atlas (TCGA) [e.g. GATA3, CDH1, TP53, TP63, TP73, RUNX1, and RUNX3).In our previous work , we deves (TCGA) , 88. By i.e. bases 128806779–129113499 within the February 2009 human genome build GRCh37/hg19) on the forward strand of chromosome 8 [i.e. miR-1204, miR-1205, miR-1206, miR-1207-5p, miR-1207-3p, and miR-1208) [PVT1 is a large intergenic non-coding RNA that appears to be strongly conserved between mouse and human –94. The mosome 8 . MoreoveiR-1208) –101. TheiR-1208) , 102–107iR-1208) , 109. IniR-1208) –112.i.e. 6 nucleotides at positions 2-7 from the miRNA 5’-end [CACU) and miR-200b/200c/429 (AAUACU) [et al. [The miR-200 family consists of five members: miR-200a, miR-200b, miR-200c, miR-141 and miR-429. On the basis of the similarities of their seed sequences (A 5’-end ), the mi(AAUACU) , 115. Th(AAUACU) . However [et al.  experimeet al. [i.e. large changes in the ceRNA expression levels that either overcome, or relieve, the miRNA repression on competing ceRNAs; similarly, large changes in the miRNA expression allow miRNAs to escape the recruitment accomplished by ceRNAs.In the dataset we analysed in , all memet al.  suggesteHere, we are interested to analyse the specific conditions required for a ceRNA landscape to occur, betting on the titration mechanism as the main culprit. In particular, inspiring by our amazing results of  and by tThe pipeline of the algorithm for searching putative ceRNAs and for building the MMI-network  in breasData collection and processingi.e. normalized expression data) given in terms of FPKM (i.e. fragments per kilobase of exon per million fragments mapped). The analysis was restricted to 72 individuals for which the complete sets of tumour and matched-normal profiles—for both short and long RNA-seq data—were available. Entries with more than the 10% of missing values were filtered out; coding versus non-coding RNAs based on Entrez gene identifiers and human annotation obtained from NCBI [Collections of tumour and normal expression data from high-throughput RNA- and miRNA-sequencing of breast invasive carcinoma were downloaded from the TCGA data portal , 88. Higrom NCBI  were seprom NCBI . All togrom NCBI .Statistical analysisth percentile of the corresponding overall correlation distribution .The Seed match analysisi.e. without introns) for lncRNAs were obtained querying the Ensembl [biomaRt and by using Entrez gene identifiers [The minimal pairing requirement to predict a miRNA target recognition is a perfect match to positions 2 to 7 (6-mer miRNA seed) at the 5’-end of the mature miRNA sequence . The miR Ensembl  data por Ensembl  interfacntifiers . For eacntifiers , all thentifiers  all the ntifiers .Network buildingρ > 0.7); ii. matching high values of the sensitivity correlation (S > 0.3); iii. sharing binding sites for miRNAs (6-mer miRNA seed match).The MMI-network both in normal and cancer tissues was built by integrating the results of statistical analysis and seed match analysis. Nodes in the networks represent mRNAs and lncRNAs with highly correlated expression profiles while edges represent miRNAs mediating their interactions. Concretely, linked nodes are required to meet three conditions: i. matching high values of the Pearson correlation between their expression profiles (i.e. by using the TCGA dedicated software “gtdownload” to query via controlled access the restricted-access data repository). For each patient the relative two bam files corresponding to the breast tumour and normal sample are used as input for the Cufflinks software [Mapped read data (bam files) for the 72 patients  analysed in  were dowsoftware  in orderThe PVT1 locus assembled by Cufflinks was compared with genome annotations for the same locus provided by Ensembl (release Homo sapiens GRCh37) by running the Cuffcompare utility and by careful inspection of the above assemblies and annotations on the UCSC genome browser.https://genome.ucsc.edu/cgi-bin/hgTracks?hgS_doOtherUser=submit&hgS_otherUserName=pantaleoM&hgS_otherUserSessionName=hg19_pone_S16_46501Targeted reassembly of the PVT1 locus were performed using the Trinity software  with defWhile the overall agreement between the Trinity and Cufflinks assemblies is good, and both methods concur in recovering all of the PVT1 Refseq exons, we notice that some of exons predicted by Cufflinks are not supported by any of Trinity assemblies, and likely constitute false positives. Importantly the most prevalent isoforms reconstructed by Trinity are highly similar if not completely identical to the two most expressed PVT1 isoforms TCONS_147426 and TCONS_147501 predicted by Cufflinks, suggesting that these discrepancies in the assembly are not likely to play a major effect on isoforms abundance estimation. A differential alternative exon usage analysis , performThe PVT1 locus—assembled by the reference-based RNA-Seq transcriptome assembler Cufflinks using TCGA data of breast invasive carcinoma—is composed of 91 different isoforms . The FPKn-by-p data matrix X, whose rows correspond to the n observations and columns to the p variables. The representation of X in the principal component (PC) space is known as matrix of the principal component scores .In order to classify the PVT1 isoforms on the basis of the FPKM data, we used the Principal Component Analysis (PCA) , 130. PCt scores , whose rt scores , whose ri.e. indirect (direct) interaction meaning that the ceRNAs communication is (is not) arbitrated by one or more microRNAs. As already mentioned in [i.e. the Pearson correlation is equal to the partial correlation, leading to the expected conclusion that, in normal breast tissues, the majority of the miRNAs is not arbitrating the cross-talk between long non-coding RNAs and coding RNAs. Thus, the observed high correlations between the expression profiles of the top-correlated lncRNA/mRNA pairs could be presumably ascribable to a common transcriptional regulatory mechanism, rather than to a post-transcriptional regulation program orchestrated by shared miRNAs. Nevertheless, a small pool of miRNAs appears as responsible of the vertical light stripes that unexpectedly stand out from the prevailing red colour of the background of vice-versa, leads to the inference of a marked rewiring in the ceRNA program between normal and pathological breast tissue that confers an interesting character to ceRNAs as potential oncosuppressive, or oncogenic, protagonists in cancer.Inspiring by our previous study presented in , in thisioned in , in physi.e. the node with the largest number of links or the highest degree in the network) in the normal-MMI-network. It is connected to 753 different mRNAs  and the miR-200 family members are arbitrating over the 80% of these interactions . From the other hand, the observation of a simultaneous up-regulation of the PVT1 gene and the miR-200 family members could lead to support the alternative hypothesis of different relative concentrations between each isoform and the miR-200 family members. According to that, a substantial decrease in cancer tissues of the relative variation of the isoform harbouring the binding site for one or more members of the miR-200 family could be due to a huge increase of the miR-200 family associated with a moderate growth in cancer of the expression levels of this PVT1 isoform. This situation, completely different from what occurs in normal tissues where the miRNA/target concentrations are comparable, could give reason of the PVT1 cease-activity as ceRNA in cancer (i.e. a titration mechanism).The analysis of the PVT1 genomic locus showed the existence of multiple isoforms  represenTo shed light on which of the two hypothesised mechanisms lies the origin of the PVT1 stoppage as sponge, we looked at the PVT1 abundance in terms of its isoforms and we found that in both normal  and cancThe observation that both the isoforms, with and without the exons where the MREs of the miR-200 family memebrs reside, resulted expressed in both cancer and normal breast tissues undermine the truthfulness of the hypothesis rested on the exon skipping mechanism and corroborates the proposal based on the relative concentrations of the PVT1 isoforms and the miR-200 family members.i.e. patients)—into a smaller-dimensional subspace—where the dimension of the space is equal to number of PCs that are able to explain the first 100% of the cumulative distribution of the explained variance of the data. The first step of this analysis is to draw a new axis representing the direction of maximum variation through the data (the first PC). Next, another axis is added orthogonal to the first and positioned to represent the next highest variation through the data (the second PC), and so on.Thus, in order to sift through the validity of a titration mechanism, we performed the principal component analysis using the feature abundance levels of all the PVT1 isoforms across samples . The aimWe found that two PCs are able to explain more than the 80% of the variance of the data . In ordeThis suggests the following argument of plausibility of the PCA analysis results: the first PC, which explain by alone about the 60% of the total variance of the analysed data , correspStudying the variation of each PVT1 isoform between normal and cancer breast tissues with respect to the variation of TCONS_147501, the results of PCA seems to be confirmed : the isoi.e. miR-200b) in both normal and cancer tissues. For the TCONS_147501 isoform (missing the binding site) this ratio does not change between normal and breast cancer tissues, while in the case of the TCONS_147426 isoform (harbouring the binding site) this ratio shows a drastic decrease from normal to cancer tissues , we found a drastic drop, in the pathological condition, in the relative concentration of the PVT1 isoform hosting the binding site for the miR-200b. The drastic change observed in the sponge program, which is suggestive of a marked ceRNA rewiring that characterizes the cancer state, could support the testable hypothesis of a titration mechanism regarding the two main isoforms of PVT1 and the miR-200 family members.Starting from the results presented in —where weS1 FigThis figure shows the results of a differential alternative exon usage analysis, performed by comparing the normalized reads counts distributions on the Refseq PVT1 exons. It shows a striking pattern supporting the up-regulation of all the exons downstream of exon 5 in the tumour samples. This observation is highly consistent with our hypothesis that the up-regulation of PVT1 in tumour samples is mostly due to the up-regulation of isoforms of the gene devoid of the key exons exerting the sponge activity on miR-200 family members.(PNG)Click here for additional data file.S2 Figi.e. bases 128,806,789-129,113,603 within the February 2009 human genome build GRCh37/hg19) visualised within the UCSC Genome browser (https://genome.ucsc.edu/) and assembled by the reference-based RNA-Seq transcriptome assembler Cufflinks by using the TCGA breast invasive carcinoma dataset.This figure shows the 91 PVT1 isoforms ((PNG)Click here for additional data file.S1 Filehttps://genome.ucsc.edu/) in order to obtain the This file contains PVT1 gene annotations in GTF (Gene Transfer Format) format provided by the reference-based RNA-Seq transcriptome assembler Cufflinks. This file is a simple tab-delimited text file for describing genomic features and it can be uploaded to a genome browser such as the UCSC Genome browser ((GTF)Click here for additional data file.S2 FileThis file contains the full genome sequences (in FASTA format) of the two PVT1 isoforms that mostly change between normal and cancer tissues: TCONS_147501 (missing the binding site for miR-200 family members) and TCONS_147426 (harbouring the binding site for the miR-200b/200c/429 cluster).(FA)Click here for additional data file.S1 TableThis table reports the FPKM values of PVT1 isoforms across normal and cancer breast tissues in separate and accordingly named sheets.(XLSX)Click here for additional data file.S2 Tablen-by-p data matrix X, whose rows correspond to observations  and columns to variables (i.e. patients), the variance accounted for by each component, and the cumulative function; second sheet) the matrix of the principal component scores, whose rows correspond to observations and columns to components; third sheet) the matrix of factors, whose rows correspond to variables and columns to components.This table reports the results of the principal component analysis, in separate and accordingly named sheets: first sheet) the eigenvalues of the covariance matrix of the (XLSX)Click here for additional data file.S3 TableThis table reports the percentage of PVT1 abundance, showed in (XLSX)Click here for additional data file.S4 TableThis table reports the variations between cancer and normal tissues of the expression levels of all the PVT1 isoforms with respect to the variation of the TCONS_147501 isoform lacking the binding site for the miR-200 family members, showed in (XLSX)Click here for additional data file.