Non-arteritic anterior ischemic optic neuropathy (NA-AION) is one of the most common types of ischemic optic neuropathy. Several recent studies suggested that abnormalities of choroidal thickness might be associated with NA-AION. The main objective of this case–control study was to evaluate whether choroidal thickness is an ocular risk factor for the development of NA-AION by evaluating the peripapillary and subfoveal choroidal thicknesses in affected Chinese patients.Forty-four Chinese patients with unilateral NA-AION were recruited and compared with 60 eyes of 60 normal age and refractive-error matched control subjects. Peripapillary and subfoveal choroidal thicknesses were measured by enhanced depth imaging optical coherence tomography. Choroidal thicknesses of eyes with NA-AION and unaffected fellow eyes were compared with normal controls. Choroidal thicknesses of NA-AION eyes with or without optic disc edema were also compared. The correlation between choroidal thickness and retinal nerve fiber layer (RNFL) thickness, logMAR best-corrected visual acuity (BCVA), and the mean deviation (MD) of Humphrey static perimetry in NA-AION eyes were analyzed.P < 0.05). There was no significant difference in the choroidal thicknesses between the unaffected fellow eyes of NA-AION patients and normal eyes of healthy controls; or between the NA-AION eyes with resolved optic disc edema and normal eyes . No significant correlation between choroidal thickness and RNFL thickness, logMAR BCVA and perimetry MD was found in eyes affected by NA-AION .The peripapillary choroidal thicknesses at the nasal, nasal inferior and temporal inferior segments in NA-AION eyes with optic disc edema were significantly thicker compared with that of normal subjects (Increase in peripapillary choroid thickness in some segments was found in NA-ION eyes with optic disc edema. However, our findings do not support the hypothesis that choroidal thickness is abnormal in Chinese patients with NA-AION compared with normal subjects with similar age and refractive error status. Non-arteritic anterior ischemic optic neuropathy (NA-AION) is one of the most common types of ischemic optic neuropathy and can develop bilaterally over an interval of several months to years. The development of NA-AION is caused by acute ischemia of the optic nerve head (ONH), in which the main source of blood supply is derived from the posterior ciliary artery circulation via the peripapillary choroid. Therefore, it has been suggested that the peripapillary choroidal thickness might be altered following ONH ischemic disorders . In addiThis observational case–control study consisted of 44 Chinese patients with unilateral NA-AION and 60 healthy Chinese volunteers. The participants were examined between October 2012 and June 2014. The study adhered to the tenets of the Declaration of Helsinki and was approved by the Institutional Review Board and Ethics Committee of Capital Medical University and informed consent were obtained from all participants. The reporting of the study adhered to the STROBE guidelines for case–control studies. The diagnostic criteria for NA-AION included a history of sudden visual loss; optic disc edema at the disease onset; spontaneous resolution of optic disc edema at subsequent follow-up; optic disc-related visual field defects; and an absence of other ocular, systemic, or neurologic diseases that might influence the patient’s visual symptoms or cause optic disc edema and visual impairment . ExclusiAll participants underwent a thorough ophthalmic examination, which included measurement of logMAR best-corrected visual acuity (BCVA), refraction, tonometry, slit-lamp biomicroscopy, fundus examination, axial length measurements using A-scan ultrasonography, and 24–2 Humphrey automated static perimetry. Peripapillary and subfoveal choroidal thicknesses were measured through natural pupil size using the enhanced depth imaging mode of the Heidelberg Spectralis optical coherence tomography . Subfoveal choroidal thickness was measured by a horizontal scan going directly through the center of the fovea and the peripapillary choroid thickness was measured by a 360°, 3.4 mm diameter peripapillary circular scan using the standard protocol for retinal nerve fiber layer (RNFL) as previously described –12. ChorP values were two-sided and were considered statistically significant when the values were <0.05.Statistical analysis was performed using a commercially available statistical software package . We used independent sample t-test to analyze factors related with choroidal thickness between NA-AION patients and normal subjects including age, gender, refractive error and axial length. Secondly, we performed t-test to compare the choroidal thickness in the affected eyes of NA-AION patients with normal eyes of healthy control subjects, and the unaffected fellow eyes of NA-AION patients with eyes of controls. Further analysis was performed to compare eyes with or without optic disc edema and normal eyes of healthy subjects. Bivariate correlation analysis was performed to evaluate the relationships between choroidal thickness and RNFL thickness, logMAR BCVA, and the mean deviation (MD) of Humphrey static perimetry (24–2) of the affected eyes of NA-AION patients. P > 0.05). Therefore, we compared choroidal thickness between patients in NA-AION and normal control group without adjusting for age, axial length and refractive error.A total of 44 Chinese patients  with unilateral NA-AION were included in the study. None of the patients was excluded due to failure to fulfill the exclusion or inclusion criteria. The mean ± standard deviation (SD) age of the patients was 50.8 ± 10.0 years  and the mean ± SD axial length was 22.69 ± 0.88 mm . The mean ± SD spherical equivalent refractive error of the eyes with NA-AION was 0.78 ± 1.14D (range: +1.00D to −3.00D). The baseline characteristics of the NA-AION patients are shown in Table P<0.05). However, we found no difference between subfoveal choroidal thickness of the NA-AION affected eyes and that of normal subjects; nor between the peripapillary and subfoveal choroidal thickness of the unaffected fellow eyes of NA-AION patients and that of normal subjects .Peripapillary choroidal thicknesses in different segments and subfoveal choroidal thicknesses of the affected and fellow eyes of NA-AION patients and that of normal subjects are shown in Table P<0.05). However, we found no significant difference between the subfoveal choroidal thickness of NA-AION eyes with optic disc edema and that of normal subjects; and between the peripapillary and subfoveal choroidal thicknesses of the affected eyes of NA-AION patients without optic disc edema and that of normal subjects .The subfoveal and peripapillary choroidal thicknesses in various segments in the affected and fellow eyes of NA-AION patients with or without optic disc edema are shown in Table P > 0.05). In addition, no significant correlation was found between the peripapillary choroidal thicknesses in each segment and subfoveal choroidal thicknesses in both the affected and fellow eyes of NA-AION patients .Bivariate correlation analysis was performed to evaluate the association between various parameters and peripapillary and subfoveal choroidal thicknesses. No significant correlation was found between choroidal thickness and peripapillary RNFL thickness, logMAR BCVA, and MD of Humphrey visual field in eyes with NA-AION (all Previous studies on choroidal thickness in various retinal, chorioretinal and optic nerve head disorders have revealed that SFCT was reduced with older age and increasing myopic refractive error, and SFCT was reduced in myopic maculopathy and in eyes with geographic atrophy . MoreoveIn order to evaluate the impact of optic disc edema on the peripapillary choroidal thicknesses, we measured the choroidal thicknesses of NA-AION eyes with optic disc edema and those with resolved optic disc edema and found that the peripapillary choroidal thicknesses in the nasal, nasal inferior and temporal inferior segments in eyes with optic disc edema were thicker than normal subjects. We found no significant difference between the choroidal thickness in eyes with resolved optic disc edema and that in normal subjects, nor between the choroidal thickness of the unaffected fellow eyes of the NA-AION patients and that of normal subjects. The results suggest that abnormal thickening of the peripapillary choroid in some segments may be pathologically attributed to optic disc edema in early stage of NA-AION, rather than contributing to the development of NA-AION. Alterations in the peripapillary choroid observed in eyes with NA-AION are therefore likely to be secondary effects rather than the cause of NA-AION. The reason for the thickening of peripapillary choroid in the nasal, nasal inferior and temporal inferior segments might be related to expansion of peripapillary choroidal blood capillaries. The combination of a relative inferior altitudinal defect with an absolute inferior nasal defect is the most common pattern of visual field defect in NA-AION, suggesting the presence of vascular insufficiency involving the temporal and superior parts of the capillary bed of the ONH, and a vascular compensatory expansion in the nasal and inferior parts of the capillary bed at early stage of NA-AION . As the Studies reported in the literature to date have shown no consensus on the correlation between RNFL and choroidal thicknesses. Zhao et al  believedThere were some limitations in our study. Firstly, our study was a cross-sectional study and we were unable to perform serial EDI-OCT measurements in NA-AION patients. Therefore, the group with optic disc edema and the group without optic disc edema were not self-matching and we lacked data about the longitudinal follow-up of the choroidal thicknesses in the patients with optic disc edema. Secondly, most of the NA-AION patients suffered from one or more systemic diseases such as hypertension, hyperlipidemia and diabetes mellitus. Since SFCT was found not to be significantly associated with blood pressure, serum concentrations of lipids, and serum glucose , we compThe present study showed that increase in peripapillary choroidal thickness occurred in some peripapillary choroid segments in NA-AION eyes with optic disc edema and not in NA-AION eyes without optic disc edema compared with eyes of normal subjects with similar age, refractive error status and axial length. The findings showed there is insufficient evidence to demonstrate that choroidal thickness is abnormal in eyes with NA-AION compared with normal subjects, suggesting that choroidal thickness is unlikely to be an ocular risk factor for the development of NA-AION.APAC, acute primary angle-closure; BCVA, best-corrected visual acuity; EDI-OCT, enhanced depth imaging-optical coherence tomography; MD, mean deviation; NA-AION, non-arteritic anterior ischemic optic neuropathy; NTG, normotension glaucoma; ONH, optic nerve head; RNFL, retinal nerve fiber layer; SFCT, subfoveal choroidal thickness


Inter- and intra-subject variability of the motor evoked potentials (MEPs) to TMS is a well-known phenomenon. Although a possible link between this variability and ongoing brain oscillations was demonstrated, the results of the studies are not consistent with each other. Exploring this topic further is important since the modulation of MEPs provides unique possibility to relate oscillatory cortical phenomena to the state of the motor cortex probed with TMS. Given that alpha oscillations were shown to reflect cortical excitability, we hypothesized that their power and variability might explain the modulation of subject-specific MEPs to single- and paired-pulse TMS . Neuronal activity was recorded with multichannel electroencephalogram. We used spTMS and two ppTMS conditions: intracortical facilitation (ICF) and short-interval intracortical inhibition (SICI). Spearman correlations were calculated within and across subjects between MEPs and the pre-stimulus power of alpha oscillations in low (8–10 Hz) and high (10–12 Hz) frequency bands. Coefficient of quartile variation was used to measure variability. Across-subject analysis revealed no difference in the pre-stimulus alpha power among the TMS conditions. However, the variability of high-alpha power in spTMS condition was larger than in the SICI condition. In ICF condition pre-stimulus high-alpha power variability correlated positively with MEP amplitude variability. No correlation has been observed between the pre-stimulus alpha power and MEP responses in any of the conditions. Our results show that the variability of the alpha oscillations can be more predictive of TMS effects than the commonly used power of oscillations and we provide further support for the dissociation of high and low-alpha bands in predicting responses produced by the stimulation of the motor cortex. While single-pulse TMS (spTMS) allows studying corticospinal excitability , paired-Compared to rather large number of TMS-EEG studies dedicated to spTMS , very feAnother important question is whether the degree of oscillatory neuronal variability by itself can relate with MEP produced by sp and ppTMS. During the last several years a number of studies specifically focused on the role of brain activity fluctuations and their functional relevance to behavioral and clinical outcomes . The varGiven all the considerations presented above, in the present study we hypothesized that: (1) Not only power but also the variability of alpha oscillations would be predictive of TMS responses. (2) Pre-stimulus alpha oscillations would relate more closely to MEP variability in ppTMS than in spTMS protocols.Seventeen right-handed healthy volunteers  between 19 and 34 years of age  participated in the experiment after giving a written informed consent in accordance with the Declaration of Helsinki. Subjects were screened for contraindications to TMS  before tA MagPro X100 (MagVenture) stimulator connected to a water-cooled MCF-B65 induction coil with 75-mm wing radius was used to produce biphasic TMS pulses. A frameless TMS navigation system  was used for MRI-guided navigation, which ensures consistent coil position and orientation in a 3D space through the sequence of stimulations. TMS coil position was optimized accordingly to individual MR scan . Stimulation targeted the left primary motor cortex , a “hotThree blocks of stimuli  were delivered: single-pulse (SP) TMS and two ppTMS protocols: short-interval cortical inhibition (SICI) and ICF. The inter-stimulus intervals (ISIs) for the paired-pulse stimuli were set at 2 ms or 12 ms for SICI and ICF protocols, respectively. The intensity of 110% of RMT was used for spTMS pulses and for the test pulses (S2) in ICF and SICI protocols. Conditioning pulses (S1) for both paired-pulse paradigms had 90% RMT intensity. The interval between the stimuli (or pairs of stimuli) varied randomly from 3 to 10 s and the inter-condition interval varied between 1 to 5 min. We provide the following reasons for determining inter-stimulus intervals. Firstly, we aimed at having a large variability of the intervals in order to avoid the anticipation effect. Secondly, as we used the same distribution for delays in all subjects, the results are unlikely to be due to the chosen inter-stimulus delays but rather would reflect genuine impact of oscillatory activity on cortical excitability. The three blocks were performed in a random order across participants. During all the conditions the coil was held by the operator who constantly monitored its position and orientation with respect to the target using navigation system. Subjects were seated in a comfortable armchair with elbows flexed at 90° and prone hands in a relaxed position, eyes were open and fixed at the mark on the opposite wall.In order to measure EEG, we used BrainAmp DC  amplifier. 91-electrode BrainCap Fast’n Easy Standard Electrode Cap (Brain Products) was used with TMS-compatible electrodes. The reference electrode was at the bridge of the nose, and ground electrode was placed on the left cheekbone. Three electrooculographic (EOG) electrodes were placed above the nasion and below the outer canthi of the eyes as indicated in . The impMEPs to single- and paired-pulse TMS  were recorded from the right APB muscle with surface bipolar EMG, using Ag–AgCl bipolar electrodes in a belly–tendon montage and were also recorded with the BrainAmp DC amplifier.EMG activity was high-pass filtered with a fourth order Butterworth high-pass filter (cut-off frequency at 10 Hz) and with the band-stop filter at 50 Hz to remove power-line noise. MEP peaks were identified within the range of 20–62 ms from the onset of the TMS stimulus for the three conditions . This range was sufficient to include MEPs in all subjects. Peak-to-peak measures of the largest positive–negative deflection were used as the MEP amplitudes. Visual inspection was also performed in parallel to remove possible artifactual trials.Electroencephalographic recordings were segmented using TMS event marker. Pre-stimulus segment was 1200 ms (-1210 ms to -10 ms). Channels with excessive amount of noise were excluded from further analysis (maximum 15 channels per subject). Blink-related artifacts in EEG were removed with fastICA algorithm . After tPre-stimulus alpha power was estimated from the spectrum calculated with Fast Fourier Transform (FFT), Hanning window (duration 1000-ms immediately preceding TMS pulse in spTMS or conditioning pulse in ppTMS). For the single-trial analysis the power was calculated separately for each pre-stimulus interval. For the across-subjects analysis, the power was averaged across all trials in each condition, electrode and subject. As has been performed in previous studies , we consThe variability was estimated from single trials both for the amplitude of MEPs and for the pre-stimulus alpha power obtained in each subject, electrode and condition. It was quantified with the coefficient of quartile variation (CQV)  – a desc1 and Q3 denote the first (lower) and third (upper) quartiles of the data, respectively. Quartiles are the points that divide any ranked data set into four equal groups. Each group contains a quarter of the data. Q3 - Q1 is defined as the interquartile range and it is a measure of the spread. Let vectors jP = 1, p2…pnp contain pre-stimulus power values from n trials in a given subject, condition and jth-electrode . CQV will then be applied iteratively to all vectors jP thus leading to E estimates of CQV. Similarly, CQV was also calculated for the amplitude of MEPs (but only for one bipolar EMG electrode). For each jth EEG electrode, CQV of the pre-stimulus power was then correlated with CQV of MEP amplitudes (across subjects) to study the relationship between neuronal variability in the cortex and the variability in motor responses. In general, CQV has an advantage compared to frequently used coefficient of variation since it is less sensitive to the deviations from normality (In (1), Qormality .We calculated a repeated measures analysis of variance (ANOVA) in order to compare the CQV of MEP amplitudes among the three TMS conditions. We computed Wilcoxon signed rank test for the comparison of pre-stimulus alpha power among the three TMS conditions. Using the same test, we compared the CQV of pre-stimulus alpha power and CQV of MEP amplitudes between the TMS conditions.We computed a Spearman correlation of MEP amplitudes between conditions. Besides, we computed a Spearman correlation between alpha power and the amplitude of MEPs for each EEG channel and condition within and across subjects. Within-subject correlations are based on single trial analysis where the power of alpha oscillations is correlated with the corresponding MEP amplitude in each channel, subject, and condition. Moreover, in order to address a relationship between cortical and peripheral variabilities, correlations were computed between the CQV of alpha power versus CQV of MEP amplitudes for each EEG channel and condition across subjects (see above the description of CQV).In order to take into account multiple comparisons , for both Wilcoxon signed rank test and Spearman correlations across subjects, a significance was estimated using cluster-based permutation statistics .The analysis was performed with custom scripts implemented in Matlab .Figure 1). This was statistically verified with the t-tests comparing normalized MEPs against the distribution with unitary mean .ppTMS protocols led to robust SICI and ICF phenomena across subjects. ICF protocol resulted in the increase of MEPs amplitudes compared to MEPs amplitudes during SP condition (ICF/SP mean 2.34 ± 0.29). Likewise, SICI protocol resulted in the significant attenuation of MEPs compared to the SP . Pairwise comparisons have been performed with Wilcoxon signed-rank test: significantly higher variability of MEPs amplitudes across subjects (assessed with CQV) was observed for SP compared to ICF sessions (P = 0.005), SICI had higher CQV compared to SP sessions (P = 0.008) and compared to ICF sessions (P < 0.002).Repeated measures ANOVA showed significant differences among conditions ; SP-SICI ; ICF-SICI .CQV of MEP amplitudes between the conditions showed significant positive correlations across subjects between SP and SICI conditions only: SP-ICF . There were no differences of the alpha power between any of the TMS conditions (SP vs. SICI vs. ICF).Figure 3 shows that the CQV of the alpha power in 10–12 Hz frequency range was significantly higher in SP than in SICI condition (P < 0.05). The difference was most pronounced in the right fronto-central area. There were no differences in CQV of alpha power in 8–10 Hz between any of the conditions.Variability (CQV) of the alpha power differed between SP and SICI conditions. Figure 4, a number of subjects with significant correlations is presented for each EEG channel, TMS condition and alpha band. Note that although there are some scattered clusters of electrodes, the number of subjects with significant correlations at each electrode location is not large, i.e., <5 subjects, (<30%). In addition one can see that the correlations with both signs can be present, thus, not demonstrating a consistent tendency between the oscillatory power and the amplitude of MEPs.In single-trial analysis Spearman correlations between EEG alpha power  and the amplitude of MEPs did not reveal consistently similar channels with significant correlations across subjects. In We computed a Spearman correlation between oscillatory pre-stimulus power and the amplitude of MEPs across subjects using cluster-based permutation statistics. No correlation of the alpha power  with the MEP amplitudes across subjects was detected in any TMS condition  or in alpha frequency sub-bands. In addition, no correlation of the alpha power was found with the MEPs variability .P < 0.05). This finding indicates that higher variability of high-alpha (10–12 Hz) power was associated with higher variability of MEP amplitudes . As in the case of the other comparisons, there were no significant correlations for low-alpha (8–10 Hz) band in any of the TMS conditions.Finally we assessed a relationship between the variability at both cortical and peripheral levels using cluster-based permutation statistics. Only for ICF condition we found a significant positive correlation between both CQVs  and theta (4–7 Hz) oscillations and parameters of MEPs. There were no significant correlations across subjects between power in these bands and MEPs amplitudes or between oscillatory-power CQV and MEP CQV.There were four main findings of this study. Firstly, we showed that trial-to-trial variability of the MEP amplitudes differed significantly among three TMS conditions. Secondly, we also found a significant difference of trial-to-trial variability of the high-alpha (10–12 Hz) power in the 1000-ms pre-stimulus EEG for SP-SICI comparison (SP > SICI). At the same time, for alpha power no difference among TMS conditions was observed. Thirdly, neither single trial analysis, nor across-subjects approach revealed any significant correlation between pre-stimulus alpha-power and MEP amplitudes. Finally, in the ICF condition larger variability of the high-alpha power in the pre-stimulus EEG was positively correlated with higher variability of MEP amplitudes.Below we discuss possible explanations of these findings and their significance for the understanding of the within- and across-subject variability of the motor responses in TMS studies.Both SICI and ICF were pronounced across subjects but also demonstrated considerable variability, which agrees with previous studies in healthy volunteers . ImportaWe have not observed significant differences in the pre-stimulus alpha power across conditions, indicating that there were no changes in the overall generation of alpha oscillations often observed for different experimental conditions including attention , sensoriOne of the goals of the present study was to investigate possible relationships between pre-stimulus power of oscillations and MEP amplitudes during prolonged spTMS and ppTMS sessions. Such dependencies are relevant for the development of brain-state triggered stimulation , which cIt is interesting to observe that the variability of MEPs on a single trial level is in contrast to reproducible average MEP responses in ppTMS paradigms  and to TOne of the most remarkable findings of the study is a relationship between two levels of trial-to-trial variability in ICF condition: variability of the high-alpha power over central and right parieto-occipital areas correlated positively with MEP amplitudes variability. Firstly, such a link between central and peripheral levels of the variability corresponds to the previous research connecting brain signal variability with the behavioral variability of both afferent  and effeInterestingly, all our significant results were found for the upper alpha (10–12 Hz) frequency band. In general this agrees with the findings indicating that low (8–10 Hz) and high-alpha (10–12 Hz) sub-bands may be associated with different neuronal processes . Thus, lWhile we did not find a considerable evidence for the previously reported link between the pre-stimulus EEG alpha power and MEP amplitudes either within-or across-subjects, we were able to demonstrate an importance of a pre-stimulus alpha-power variability for ppTMS phenomena, thus, providing a further support for the hypothesis that ongoing neuronal variability may modulate cortical motor output.For both studied ppTMS phenomena only one ISI (2 ms for SICI and 12 for ICF) among several commonly used  was chosBecause of the residual scalp EMG in some of our subjects, we were not able to investigate beta oscillations which are known to be relevant for motor processing. However, our results on alpha rhythm already provide a novel link between the variability in the cortical oscillations and motor responses as tested with ppTMS.At this stage of the study we did not investigate TMS-EEG responses. Firstly, it was due to our original intention to investigate a relationship between ongoing neuronal oscillations and cortical excitability as probed with MEPs in ppTMS phenomena. Secondly, since our amplifiers did not have a technical possibility to be gated during the TMS pulse, we observed considerable artifacts in the post-stimulus interval thus preventing us from analyzing TMS-evoked responses.ZI processed and analyzed the data, and wrote the manuscript. MN designed the study, collected the data and wrote the manuscript. ZI and MN contributed equally to the study. TF wrote codes for the pre-processing and analysis of the data. EB designed the study and collected the data. VN designed and supervised the study, wrote the manuscript.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. The reviewer PL declared a past collaboration with one of the authors (MN) to the handling Editor, who ensured that the process met the standards of a fair and objective review.


We present a locating technique for inorganic materials in polymer-matrix composites through a post-labeling approach based on specific covalent binding. in situ visualization of incorporated inorganic materials in organic–inorganic composites in a post-labeling manner, owing to the lack of specific fluorescent organic dye molecules for targeting inorganic materials. Herein, we observe that the specific covalent B–O binding between the hydroxyl groups of inorganic materials and commercially available aggregation-induced emission (AIE)-active boronic acid could lead to the formation of highly emissive solid-state fluorescent composite materials. The hydroxyl-triggered luminescent probe may serve as a practical method for in situ location of incorporated inorganic materials in polymer-matrix composites by simply dipping the composite film in boronic acid-modified AIE solution. This present work offers a non-invasive avenue to locate inorganic materials which possess hydroxyl-groups in polymer-matrix composites, thereby developing a convenient screening strategy for assessing the advanced properties of composites. This strategy can also be extended to the targeted tracing of other inorganic materials with inherent and functionalized carboxyl, amino, sulfhydryl and other groups via tuning the binding affinity between the inorganic materials and luminescent molecules.There is a long-standing challenge to realize  Furthermore, the visualization of incorporated MMT/LDHs in polymer-matrix composites has been implemented by simply dipping the composite film in AIE-active boronic acid solution without the aid of any pre-modification before the film formation  exhibit different binding affinities towards boronic acid.It has been reported that carbohydrates with different structures  technique. To achieve this goal, we first investigated the binding affinity of TPEDB towards the LDH powder. The results showed that the non-emissive LDHs exhibited a strong cyan luminescence at 470 nm  with theZ-axis were adopted. 2). In comparison with the non-selective physical absorption process, these CFM results reflect the stable and specific location of LDHs in polymer-matrix composites by the proposed hydroxyl-triggered fluorescence method.Next, LDHs were blended in polyethylene (PE) at a ratio of 5% and a stochastic area of the PE–5% LDH film was selected and then dipped into a 100 μM TPEDB solution for 10 min. As shown in Fig. S6,via the B–O bonds. As depicted in Z-axis  through electrostatic interaction to prepare the molded PE–5% (QD@LDH) film. The obtained PE–5% (QD@LDH) film was then dipped into the TPEDB solution to mark the LDHs n Z-axis . These rin situ recognition and visualization of inorganic layered materials in polymer matrices through specific B–O bonds by a rapid and simple approach.We further investigated the universality of this proposed method for mapping the spatial distribution of inorganic layered materials with hydroxyl groups in polymer matrices. The other typical layered material, MMT,in situ targeted tracing of other inorganic materials by changing the binding affinity between the inorganic materials and AIE molecules.In conclusion, we have demonstrated the presence of a strong solid-state emission of layered materials due to the formation of specific B–O bonds between hydroxyl groups on the surfaces of layered materials and AIE-active emissive boronic acid. This hydroxyl fluorescence location technique enabled us to establish a powerful imaging platform to precisely target and trace the incorporated inorganic materials in polymer-matrix composites. This post-labelling approach exhibited superiority in evaluating the inherent structural and functional behaviors of the inorganic–organic composites in comparison with the traditional pre-modification procedures. More importantly, such a unique fluorescent probe can be easily achieved by simply dipping polymer-matrix composite films in AIE-active boronic acid solution combined with high-resolution fluorescent imaging. Our strategy opens up new possibilities for optimization of critical processing conditions, while retaining the initial structural nature of polymer-matrix composites. This facile method can also be applied for the There are no conflicts to declare.Supplementary informationClick here for additional data file.


Members of the ATPases Associated with various cellular Activities (AAA+) superfamily participate in essential and diverse cellular pathways in all kingdoms of life by harnessing the energy of ATP binding and hydrolysis to drive their biological functions. Although most AAA+ proteins share a ring-shaped architecture, AAA+ proteins have evolved distinct structural elements that are fine-tuned to their specific functions. A central question in the field is how ATP binding and hydrolysis are coupled to substrate translocation through the central channel of ring-forming AAA+ proteins. In this mini-review, we will discuss structural elements present in AAA+ proteins involved in protein quality control, drawing similarities to their known role in substrate interaction by AAA+ proteins involved in DNA translocation. Elements to be discussed include the pore loop-1, the Inter-Subunit Signaling (ISS) motif, and the Pre-Sensor I insert (PS-I) motif. Lastly, we will summarize our current understanding on the inter-relationship of those structural elements and propose a model how ATP binding and hydrolysis might be coupled to polypeptide translocation in protein quality control machines. AAA+ proteins harness metabolic energy in form of ATP to facilitate diverse cellular processes, including organelle biogenesis, membrane fusion, transcriptional regulation, and protein quality control (PQC). Members of the AAA+ superfamily can be classified into one of four distinct clades or superclades: (1) the clamp loader clade, (2) the initiator clade, (3) the classic clade, and (4) the Pre-Sensor I insert (PS-I) superclade  impaired protein disaggregation to similar levels to that observed with a ClpB variant featuring a D2 pore loop tyrosine to alanine mutation (ClpBY643A)  of the ISS motif, triggering ATP hydrolysis in the cis-subunit concomitant with substrate translocation. We propose that the PS-I motif communicates with pore loop-1 and controls substrate interaction by either contacting the substrate directly or regulating the ATPase cycle in the D2 ring through communication with the D1 ring. Although the available structural and biochemical evidence provide support for such mechanism, determining the structure of a substrate bound complex will be necessary to provide a more accurate mechanistic understanding how the ATPase cycle is coupled to substrate translocation in PQC machines.The available 3D structures of AAA+ machines involved in PQC have provided snapshots of distinct functional states and have contributed toward our molecular understanding how the ATPase cycle is coupled to conformational changes needed for substrate translocation. Structural evidence suggests that the pore loop-1 conformation optimized for substrate binding is determined by the nucleotide-bound status of the CC, SL, and FT contributed to writing the draft and final version of this mini-review.Research in the FT and SL laboratories is supported by grants from the National Institutes of Health (GM104980 and GM111084) and the Welch Foundation (Q-1530). CC is the recipient of an American Heart Association-Southwest Affiliate Postdoctoral Fellowship.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.


Ice-shelf channels are long curvilinear tracts of thin ice found on Antarctic ice shelves. Many of them originate near the grounding line, but their formation mechanisms remain poorly understood. Here we use ice-penetrating radar data from Roi Baudouin Ice Shelf, East Antarctica, to infer that the morphology of several ice-shelf channels is seeded upstream of the grounding line by large basal obstacles indenting the ice from below. We interpret each obstacle as an esker ridge formed from sediments deposited by subglacial water conduits, and calculate that the eskers' size grows towards the grounding line where deposition rates are maximum. Relict features on the shelf indicate that these linked systems of subglacial conduits and ice-shelf channels have been changing over the past few centuries. Because ice-shelf channels are loci where intense melting occurs to thin an ice shelf, these findings expose a novel link between subglacial drainage, sedimentation and ice-shelf stability. The formation mechanisms of ice-shelf channels remain poorly understood. Here, using ice-penetrating radar data, the authors propose that ice-shelf channel morphology in the Roi Baudouin Ice Shelf, East Antarctica, is seeded by esker ridges, indenting the ice from below. Water beneath the Antarctic Ice Sheet promotes the formation of ice streams that rapidly slide over wet sediments and a lubricated base. Ice streams discharge the majority of Antarctic ice into floating ice shelves, which surround about 74% of the Antarctic perimeter459In this study we use satellite data and ice-penetrating radar to show that ice-shelf channels on the Roi Baudouin Ice Shelf, East Antarctica, are seeded upstream of the grounding line by basal obstacles indenting the ice from below. These obstacles align with predicted hydrological outlets, and thus we interpret them as eskers (sediment ridge composed of gravel and sand) formed by the overlying subglacial water conduits. Our findings confirm a recognized linkage between ice-shelf channel formation and subglacial hydrology6911Here we survey three hydrologically predictedIn full details below, we argue that scenario 3 accords best with our observations, and we interpret each basal obstacle as an actively evolving ramp-shaped esker whose size increases towards the ocean due to subglacial conduit widening and decreasing water flow speed. Eskers, a glacial landform used in the reconstruction of palaeo ice sheets1416We estimate the grounding-line position using satellite-based interferometric synthetic aperture radar by picking the landward limit of the tidal flexure zone in interferograms from 1996, 2007 and 2016 . The lim20212) of reflector A upstream of the grounding line. We see no reflections delineating lateral walls, so either no walls exist or they are too steep to be imaged by our nadir-looking radar. The former interpretation here implies an essentially two-dimensional internal reflector, which is inconsistent with the seaward extension of these features into ice-shelf channels where ice is lacking compared to the neighbouring areas  bedforms is supported by more evidence on the ice shelf: surface ridges dating from earlier times can be found there, implying past changes in the coupled systems.The seeding of ice-shelf channels A–C at the grounding line means that their morphology can record grounding-line history as well as changes in sub-shelf melt plume dynamics as ice flow advects them towards the shelf front. At sites A and C, the surface ridges decay seawards to grade into the depressed surface of the ice-shelf channels downstream  although−1 as an upper limit, which is an order of magnitude higher than model estimates of sedimentation rates at melt-stream portalsIn this interpretation, the surface ridges at sites A and C are currently below their critical threshold so they do not intrude far into the ice-shelf channels. But the isolated hills suggest that these ridges may soon be advected into the ice shelf. An end-member scenario is that the ramp-shaped esker at site A was completely removed by flooding 175 a ago and has since re-grown to its present size. This requires a sedimentation rate of ∼1.4 m aAt site B, the basal disruption imaged by reflector B is offset from the surface ridge, and we lack ground-based radar data for examining its geometry along flow. In plan view, the corresponding ice-shelf channel is less developed compared to ice-shelf channels A and C and deviates from the ice-flow direction towards ice-shelf channel C. These observations suggest that the esker/conduit B began developing more recently and has been migrating eastward to reach its current position.Numerous subglacial processes in scenario 3 require further investigation. Our mechanism for the formation of ramp-shaped eskers involves a reduction of water flow speed towards the grounding line, which has not been directly measured. Also, without access to the bed, we lack precise information about the current subglacial conduit arrangement and sediment transport regime on/near the eskers, which determine how their form continues to evolve. Since all three sites lack sediment sources on the surface, a negligible sediment delivery to the bed is expected from supraglacial melt-water streamsin situ in their formation environment, and confined by ice, which prevents sediment-flank slumping. If the grounding line retreats, rapid degradation by slumping and erosion would occur, especially as their sediments are probably weakly consolidated in the subaqueous environment; given enough time, a drastic height reduction is hence not inconceivable. These ideas seem to us consistent with the fragmented nature of esker networks from the Wisconsinan glaciation , which typically consist of ridge segments with major gaps in between. Indeed, many of them may be the meagre remnant or core of originally much higher eskers.It is noteworthy that our eskers are an order of magnitude larger than most eskers in deglaciated areas, which usually do not exceed 50 m in heightFinally, the inferred ramp shape of our eskers has counterparts in the deglaciated landform record, notably eskers of type I–III described by BrennandOur findings have several implications. First, the understanding of ice-shelf channel formation is now improved and more complicated than previously assumed. Three mechanisms have been suggested in other studies: first, ice-shelf channels can develop through meltwater plume flow combined with transverse variability in ice thickness1112691188Second, evidence for channelized, subglacial water outlets supports the hypothesis that the meandering of ice-shelf channels seen in many ice shelves is an archive for the history of the subglacial hydrology in the respective drainage area6Third, large ramp-shaped eskers upstream of the grounding line require stability of the subglacial hydrological system for their development. The Roi Baudouin Ice Shelf is a good candidate for this, because many ice-shelf channels extend from the grounding line to the ice-shelf front along flowlines, indicating temporal stability of the respective source at the grounding line over several hundreds of years. Analysis of an ice rise in that ice shelf suggests an even longer period of stability of the large-scale flow regime spanning several thousands of yearsFourth, the surface ridges and the relict ridge in the ice-shelf channel are key to our interpretation of the ramp-shaped esker formation. Therefore, our data provide field evidence for the evolution of a subglacial landform which is extensively investigated in other areas where ice-sheets have retreatedFifth, the ramp-shaped esker/subglacial conduit system is clearly visible in our ground-based and airborne profiles about two ice thicknesses upstream of the grounding line. Their locations can be inferred from remote-sensing data, by aligning calculated water outlets with ice-shelf channelsIn summary, we have discovered an actively evolving system of ramp-shaped eskers and subglacial conduits upstream of the grounding line of an Antarctic ice shelf. Seawards, ice-shelf channels are likely further incised by a buoyant melt-water plume forced by the meltwater exiting the conduit. The ice-shelf channel amplitudes, however, can already largely be determined by the height of the ramp-shaped esker. The latter develop through a generic process of conduit widening at the grounding line where creep closure is small. Diminished water outflow speed increases potential for sedimentation. Some ramp-shaped eskers are large enough to form corresponding ridges at the surface of the overriding ice. Depending on the height-to-width ratio, these ridges are preserved in the adjacent ice shelf and testify to an evolving subglacial hydrological system including rerouting of conduits and potentially flushing/eroding of the ramp-shaped eskers. Our findings improve the understanding of ice-shelf channel formation by exposing a novel link between subglacial hydrology and ramp-shaped esker formation impacting ice-shelf stability. Moreover, our data set is the first evidence for ramp-shaped esker formation beneath a contemporary ice sheet and sheds light on a subglacial landform that has been intensively studied in geomorphology for decades, but for which the formation mechanisms are only poorly constrained. Further research should be directed to more rigorously infer the existence and the type of sediments deposited by the subglacial conduit , which will facilitate quantitative modelling of sediment transport at the basal boundary of ice sheets.−1 and traces were horizontally stacked to common postings (∼12 m) during the post-processing. After bandpass filtering (frequency range between 3 and 9 MHz), the data were migrated to account for off-angle reflections from sloped interfaces  using Kirchoff-Depth migration implemented in the open-source software Seismic Unix. The required radio-wave velocity model varies with depth using a depth–density parameterization8 m s−1) everywhere. We, therefore, chose the parameters (surface density and densification length) of the depth-density function so that the equivalent firn-air content is below 1 m. Picking of internal reflections and 3D-visualization is done using seismic interpretation software (OpenDtect).Ground-based and airborne radars image the thickness and internal stratigraphy of ice by measuring the traveltimes of reflections from actively emitted electromagnetic waves in the MHz range. Reflections originate from changes in ice density, ice acidity or a preferred orientation of the ice crystalsBandpass filtering/migration causes ringing near the strong reflections originating from the bed and the upper surface of the subglacial conduit. These artefacts obstruct analysis of the wavelet's phase structure, which is therefore done using the raw data only. We find no phase reversal for both reflections types, indicating a transition from an optically less dense (with faster radio-wave velocity) to an optically denser (with a slower radio-wave velocity) background medium. This is consistent with a water and or sediment-filled conduit which is overlain by (less-dense) ice.The kinematic GNSS data were collected with a dual-frequency GNSS receiver attached to the radar's receiver. We estimated the position of the moving station at 1 Hz sampling rate using the GAMIT/GLOBK v10.5 softwareThe satellite-derived surface elevations stem from TanDEM-X, a radar satellite pair imaging the ice-sheet surface using the X-band with small signal penetration into the surface. The elevation model is mosaicked out of 40 TanDEM-X single look complex scenes acquired in austral winter 2013. Scenes were processed individually using the SARscape software. The processing includes coregistration with the CryoSat-2 surface elevationAll data are available from the authors on request.How to cite this article: Drews, R. et al. Actively evolving subglacial conduits and eskers initiate ice shelf channels at an Antarctic grounding line. Nat. Commun.8, 15228 doi: 10.1038/ncomms15228 (2017).Publisher's note: Springer Nature remains neutral with regard to jurisdictional claims in published maps and institutional affiliations.Supplementary Notes, Supplementary Table, Supplementary Figures and Supplementary References


Vanellus chilensis lampronotus based on adult vocal activity. For this purpose we use passive 14-min and 30-min soundscape recordings (n = 33 201) collected in 24/7 mode between November 2012 and October 2013 in Brazil’s Pantanal wetlands. Time-stamped detections of V. chilensis call events (n = 62 292) were obtained with a species-specific sound recognizer. We demonstrate that the breeding season fell in a three-month period from mid-May to early August 2013, between the end of the flood cycle and the height of the dry season. Several phases of the lapwing’s life history were identified with presumed error margins of a few days: pre-breeding, territory establishment and egg-laying, incubation, hatching, parental defense of chicks, and post-breeding. Diurnal time budgets confirm high acoustic activity levels during midday hours in June and July, indicative of adults defending young. By August, activity patterns had reverted to nonbreeding mode, with peaks around dawn and dusk and low call frequency during midday heat. We assess the current technological limitations of the V. chilensis recognizer through a comprehensive performance assessment and scrutinize the usefulness of automated acoustic recognizers in studies on the distribution pattern, ecology, life history, and conservation status of sound-producing animal species.Computer-assisted species recognition facilitates the analysis of relevant biological information in continuous audio recordings. In the present study, we assess the suitability of this approach for determining distinct life-cycle phases of the Southern Lapwing  Cerrado region are threatening the ecosystem  through adaptive gain control (AGC) amplified weak audio events that otherwise would be disregarded due to their low energy; (ii) down-sampling the signal to sampling rate 24 kHz reduced the computational demands in the subsequent processing steps and removed high-frequency insect sounds ≥ 12 kHz; (iii) filtering with a Butterworth high-pass filter  suppressed low-frequency interference, such as wind noise and some anthropogenic noise of distant cars and airplanes; all remaining sound signals were treated uniformly regardless of their origin; and (iv) segmenting the audio to short 20-ms segments with 75% overlap.2LFCC respectively. Here we kept only the first 20 Δ2LFCCs, as the last 15 were found to be less informative. All feature vectors were post-processed to uniform dynamic range in order to facilitate the pattern recognition process. During the signal parameterization, we treated all target sounds and noises uniformly, so that the abovementioned set of 90 audio descriptors was computed for each segment of audio. Based on the post-processed feature vectors, an acoustic activity detector selects for further processing only those parts of the audio with considerable acoustic activity. Here we set the threshold to –50 dB with respect to the full range input of the SM2+ recorder. The selected segments are used in the pattern recognition process. We only considered detections of acoustic events with amplitudes of at least –50 dB because below that level V. chilensis signals were to faint for being discerned and annotated with confidence, even for a bird sound expert (section Recognizer Performance Assessment).For each segment we computed a set of 90 audio descriptors, including 35 Linear-Frequency Cepstral Coefficients (LFCC)  and theiV. chilensis detector used in this study explicitly modelled the target species and the acoustic environment through purposely developed Gaussian Mixture Models (GMMs). In particular, based on 75 h of soundscape recordings, we built a Universal Background Model (UBM) via an Expectation–Maximization (EM)-based training of a 512-component GMM. The UBM represents the model of everything that is not Southern Lapwing. Next, we created the V. chilensis model via means-only adaptation of the UBM and a purposely-designed training dataset, containing characteristic lapwing vocalization types (section The target species). The training dataset consisted of manually tagged and selectively filtered recordings of the target species .A bird sound expert prepared all training and validation datasets and assessed the detector results. Inspecting the raw detection results of the complete annual cycle of recordings , we deciV. chilensis recognizer performance through a two-pronged approach: (I) For the determination of the false positive rate we drew representative random samples from the V. chilensis detections computed for each month. (II) For all other performance statistics, we selected 26 soundscapes containing at least one correct positive detection. Detailed descriptions of the two validation datasets follow:We evaluated the V. chilensis signals discernible in the background of the same segment. In the case of doubts with respect to the identification of non-target bird species, we consulted xeno-canto (www.xeno-canto.org) and several comprehensive audio publications on Neotropical birds ) were used to correct the number of diurnal and nocturnal detections in The false positive rates (= 100 –(PDF)Click here for additional data file.S3 TableV. chilensis call event detected by the automated recognizer among correct hits and double hits.See (PDF)Click here for additional data file.S4 Table(PDF)Click here for additional data file.


Glioblastoma is the most common and highly malignant primary brain tumor, and patients affected with this disease exhibit a uniformly dismal prognosis. Glioma stem-like cells (GSCs) are a subset of cells within the bulk tumor that possess self-renewal and multi-lineage differentiation properties similar to somatic stem cells. These cells also are at the apex of the cellular hierarchy and cause tumor initiation and expansion after chemo-radiation. These traits make them an attractive target for therapeutic development. Because GSCs are dependent on the brain microenvironment for their growth, and because non-tumorigenic cell types in the microenvironment can influence GSC phenotypes and treatment response, a better understanding of these cell types is needed. In this review, we provide a focused overview of the contributions from the microenvironment to GSC homing, maintenance, phenotypic plasticity, and tumor initiation. The interaction of GSCs with the vascular compartment, mesenchymal stem cells, immune system, and normal brain cell types are discussed. Studies that provide mechanistic insight into each of these GSC–microenvironment interactions are warranted in the future. Our view of cancer has changed ever since the discovery of cancer stem cells (CSCs) . CSCs ar+) populations in extended in vitro cultures. However, CD133 negative populations from freshly dissociated tumors have been also shown to form tumors at similar rates as CD133+ cells and additional markers for defining GSCs have been proposed  are found in the brain and play a role in immune surveillance . The actSome earlier studies describing the role of immune cells in GSCs have been generally shown to silence the immune response, escape immune surveillance, for instance, with an ineffective tumor antigen presentation, or release and recruitment of on-site immune suppressive factors (such as TGF-β) and immune suppressive cells (such as immunosuppressive B-cells and myeloid cells) . Studiesin vivo.In addition to expression of co-inhibitory molecules, alternative mechanisms of T cell inhibition exist. Regulatory T cells (Tregs) are a form of T cells that are immunosuppressive and generally inhibit the expansion of effector T cells. This is attributed to constitutive activation of cytotoxic T-lymphocyte-associated protein 4 (CTLA-4), which contributes to their ability to suppress the immune system . The ligIn addition to secreting immune suppressive cytokines, GSCs are capable of recruiting or modulating immune cells with tumor supportive phenotype. GBMs are characterized by a very high infiltration of macrophages, phagocytic cells that engulf cellular debris and foreign substances. Microglia, are resident macrophages that present the first line of defense in the CNS. Both microglia and infiltrating macrophages (myeloid cells) can represent between 13 and 43% of the tumor mass in different animal models for GBM  and in fin vitro through the secretion of cytokines. These include colony-stimulating factor-1 (CSF-1), C-C motif ligand-2 (CCL-2), and macrophage inhibitory cytokine 1, factors enriched in GSC-conditioned media. Moreover, the secretion of CSF-1 and CCL-2 by the GSCs resulted in a polarization of the macrophages toward the M2 immune suppressive phenotype. Other CSC-secreted factors include IL-10 and TGF-β, which also suppress tumor-associated microglia/macrophage function and generate a more immunosuppressive (M2) phenotype (in vivo shows a reduction in the recruitment of tumor-supportive TAMs (M2 subtype), inhibition of tumor growth, apoptosis of GSCs, and increase of survival in a xenograft mouse model. Another molecule with crucial role in this transition is SPP1 (osteopontin). SPP1 is a secreted protein that shares similarities in features and mechanism of action with Periostin and contributes to the maintenance of an immune suppressive environment. It has been shown, in fact, to be highly expressed in M2 polarized macrophages and GBM-infiltrating CD14+ cells compared to matched blood monocytes and brain microglia  or to chronic inflammatory stimuli, myeloid cells are often grouped into two different immune phenotypes named M2 (which is representative for the chronic inflammatory state) and M1  . In manyhenotype . TAMs inhenotype . Periosticroglia .It is important to note that gene expression studies of TAMs from GBM have shown that such a simplified classification is not feasible in brain tumors, which likely harbor a spectrum of differently polarized myeloid cells that (as a net effect) culminates in a GBM-promoting role of TAMs .Myeloid-derived suppressor cells are immature cells of myeloid origin that possess T cell suppressive properties. Patients with GBM have increased counts of MDSCs compared to healthy subjects . A recenin vitro  . In recein vitro . In an iin vitro . These sin vitro .Dendritic cells are antigen-presenting cells to T lymphocytes and play a major role in initiating and shaping the adaptive response. These cells represent the most potent and versatile way the immune system has evolved to present antigens and develop an immune response . DCs mayStudies involving GSCs have dramatically changed our view of this disease, and we have obtained a wealth of fascinating insight into the cell biology of GBM. The field of CSCs has helped us understand cancer biology from the perspective of developmental biologist. Similarities have been drawn to the cell types and differentiation pathways of normal stem cells. The recent information of GBM heterogeneity retrospectively explains why conflicting data were initially obtained in the GSC research field. It is now well established that GSCs are a central (but not the exclusive) target for therapeutic approaches to treat GBM. This is not only due to the inherent plasticity of GSCs that leads to transdifferentiation but also to the overbearing influence of the microenvironment (as highlighted in this review) that alters their stem cell state, tumorigenic potential, and therapeutic resistance. It is our opinion that future studies must incorporate the microenvironmental aspects while studying GSC biology.AA and SC wrote the basic contents of the manuscript. KB conceived the manuscript, provided input throughout the preparation, and edited for the accuracy of the contents along with RG.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.


Disparities in hepatocellular carcinoma (HCC) incidence and survival have been observed between ethnic groups including African-Americans (AA) and European-Americans (EA). The evaluation of the changes in the levels of metabolites in samples stratified by race could provide a snapshot of ethnically diverse disease related pathways and identify reliable biomarkers. In this study, we considered AA and EA to investigate metabolites that may be associated with HCC in a race-specific manner. The levels of 46 metabolites in plasma samples, collected from patients recruited at MedStar Georgetown University Hospital, were analyzed by Agilent GC-qMS in selected ion monitoring (SIM) mode. A least absolute shrinkage and selection operator (LASSO) regression model was applied to select metabolites with significant changes in HCC vs. cirrhosis in three groups: (1) AA and EA combined; (2) AA separately; and (3) EA separately. In addition, metabolites that distinguish HCC cases from cirrhosis in these three groups were selected by excluding those without HCV infection. The performances of the metabolites selected by LASSO in each group were evaluated through a leave-one-out cross-validation. We identified race-specific metabolites that differentiated HCC cases from cirrhotic controls, yielding better area under the receiver operating characteristics (ROC) curve (AUC) compared to alpha-fetoprotein (AFP), the serological marker widely used for the diagnosis of HCC. This study sheds light on metabolites that could potentially be used as biomarkers for HCC by monitoring their levels in high-risk population of cirrhotic patients in a race-specific manner. Hepatocellular carcinoma (HCC) is the most common type of liver cancer. An estimated 40,710 new cases of liver cancer (including intrahepatic bile duct cancers) will be diagnosed in the US during 2017, approximately three-fourths of which will be HCC .Persistent infections by HBV or HCV are the main recognized risk factors for HCC –8 with tOther than liver imaging, current diagnosis of HCC relies on the measurement of the level of the serum biomarker, α-fetoprotein (AFP). However, when considering AFP levels in patients with HCV infection, AFP appears to be insensitive for the diagnosis of HCC in AA. In a case-control study of 163 HCC patients with HCV infection and 149 control patients with HCV-related cirrhosis, the sensitivity of AFP for the diagnosis of HCC in AA with HCV infection was reported to be lower than that of patients of all other ethnic groups combined . Thus, sMetabolomics has been broadly used for biomarker discovery for many human diseases including cancer . MetabolWe previously conducted several metabolomic studies aimed at identifying HCC biomarkers in cirrhotic patients –26. PartIn this paper, we investigate race-stratified analysis of plasma metabolites measured by gas chromatography coupled with selected ion monitoring mass spectrometry (GC-SIM-MS). The plasma samples were collected from patients recruited at MedStar Georgetown University Hospital (MGUH), Washington, DC. We compared the levels of the metabolites in plasma samples from AA and EA to identify those that distinguish HCC cases from the cirrhotic controls in a race-specific manner. A panel of metabolites was selected using least absolute shrinkage and selection operator (LASSO) logistic regression  by consiThe discovery of exclusive biomarkers in HCC is still a formidable task mainly due to the heterogeneity of the clinical symptoms of cancer and the various etiologic agents that initiate the pathological liver disorders such as abnormal structural nodules with peripheral fibrosis, cirrhosis, chronic inflammation and fatty liver disease that are precursors to end stage liver disease. To our knowledge, this is the first metabolomic study that seeks to identify race-specific biomarkers by comparing the levels of plasma metabolites in HCC cases against cirrhotic patients by stratifying AA and EA. Replication of these findings may contribute to understanding of the racial disparity in HCC and also in improving diagnosis of HCC through race-specific biomarkers.Adult patients were recruited from the Hepatology Clinic at MedStar Georgetown University Hospital (MGUH). All participants provided informed consent to a protocol approved by the Institutional Review Board (IRB) at Georgetown University. The patients were diagnosed to have liver cirrhosis on the basis of established clinical, laboratory and/or imaging criteria. Cases were diagnosed to have HCC based on well-established diagnostic imaging criteria and/or histology. Clinical stages for HCC cases were determined based on the tumor-node-metastasis (TNM) staging system. Controls were required to be HCC free for at least 6 months from the time of study entry. Race information was collected from patients’ self-report. The characteristics of AA and EA, selected from these patients, are summarized in The levels of 46 metabolites in plasma samples from AA and EA in λ was chosen by cross-validation using the R function cv.glmnet. After estimating the coefficient β in , AUC (95%CI).We used multiple support vector machine recursive feature elimination (MSVM-RFE) model to rank all metabolites. MSVM-RFE computed feature-ranking score of multiple linear SVMs and reported the average ranking for each metabolite . Area unpROC [Logistic regression models were built to evaluate the performance of the predictors in HCV+ patients. Leave-one-out cross validation was used to calculate AUC for the ROC curve in logistic regression using a set of metabolites selected in two different ways: (i) selected metabolites from LASSO; and (ii) top-ranked metabolites in MSVM-RFE. The AUC for the ROC curve was then calculated based on testing data in the cross validation within each model. By R package pROC , the conp-value for AA and EA combined (Panel A), AFP was unable to distinguish HCC from cirrhosis in AA (Panel B and B1) or EA (Panel C and C1), emphasizing the need for more potent HCC markers particularly for subjects with HCV+.We evaluated the performance of AFP as a classifier for HCC in AA and EA combined, AA only, and EA only groups, for all subjects (HCV + and HCV-) as well as for HCV+ only subjects. As shown in LASSO regression conducted on AA and EA combined (adjusted or not for race), AA only, and EA only groups, selected a combination of metabolites whose expression levels jointly differentiate HCC cases from cirrhotic controls. The list of metabolites selected in each of the above three groups and considering those with or without HCV infection, is provided in t-test in the comparison between AA and EA within HCC and CIRR groups respectively are indicated. As shown in MSVM-RFE was used to rank all metabolites. Due to the high percentage of HCV+ in AA, this analysis was conducted for subjects that are HCV+ only. The complete list of the ranked metabolites is provided in Leave-one-out cross-validation examined the prediction performance based on these selected metabolites in LASSO and the top-ranked metabolites in MSVM-RFE model. By performing LASSO, 20 metabolites were selected for the AA and EA combined group with adjustment of race factor, 11 for AA and 13 for EA. The top 5 metabolites in all three groups from MSVM-RFE were also selected and evaluated. Among the metabolites selected by LASSO regression, those that were top ranked by MSVM-RFE and showed a consistent fold change between HCC cases from the cirrhotic controls across the three groups  are: alpha tocopherol, selected in all three groups; whereas valine for AA and glycine for EA. Further biological investigation will focus on them. To investigate to what degree these metabolites fluctuate in a healthy population, we analyzed sera from the patients presented in The use of metabolomics to identify potential biomarkers of HCC is greatly advantageous to patients and healthcare providers because the dysregulation of metabolites may be an early indication of dysfunctional metabolic pathways that could offer valuable insight into the mechanism of HCC initiation, development or progression. In this study, we investigated plasma metabolites that may be associated with HCC in a race-specific manner by considering AA and EA from a cohort that we previously examined . The levp-value = 0.0269), its performance declines when evaluated in AA only and EA only groups. Gupta et al. [AFP has been extensively used as a biomarker for HCC. However, its performance in HCC surveillance has been generally low . In adula et al.  examinedWe found statistically significant changes in alpha-tocopherol in HCC cases versus cirrhotic controls similar to our previous report , which wAs candidate biomarkers selected from the analysis based on the larger cohort , we repoAlpha-tocopherol, glycine and valine were previously reported in HCC related metabolomics studies, conducted by our group and others using additional human specimen, complementary metabolomics platforms and multivariate analysis. In addition, validation of the genetic ancestry of the participants should be performed using a panel of ancestry informative markers. We also acknowledge the possibility of confounding variables other than HCV (such as NASH) for the development of HCC in our study patients.In summary, we have demonstrated the inability of AFP in discriminating HCC cases from cirrhotic controls, particularly when its performance is evaluated in HCV+ race specific groups. Among the metabolites selected by LASSO, glycine and valine showed better performance than AFP in EA and AA, respectively. Following further validation in a large cohort of patients and healthy controls matched by their demographic characteristics, the metabolites discovered in this study could contribute to better understanding of the development of HCC and allow early detection of HCC in patients with liver cirrhosis and HCV in a race-specific manner.S1 Table(PDF)Click here for additional data file.S2 Table(PDF)Click here for additional data file.S3 Table(PDF)Click here for additional data file.S1 FigThe individual dot plot, for alpha tocopherol, valine and glycine in AA and EA combined, AA, and EA groups are shown in S1A, S1B, S1C Fig respectively . The changes of the metabolites level are shown from cirrhosis, HCC stage I to HCC stage II.(PDF)Click here for additional data file.


Technologies for multimodal digital imaging of vitreoretinal diseases have improved the accuracy of diagnosis and the depth of the knowledge of the mechanisms of disease and their response to treatments. Optic coherence tomography (OCT) has become a mandatory tool for the management and for the follow-up of retinal pathologies. OCT technology evolved in the last two decades from time-domain to spectral domain and recently to the swept-source OCTs (SS-OCT). SS-OCT improved the depth of imaging and the scan speed, thus adding novel algorithms and features such as for vitreous and vitreoretinal interface evaluation, choroid segmentation and mapping, OCT angiography and En-face OCT. The multimodal approach using SS-OCT is expected to advance the understanding of retinal pathologies such as age related macular degeneration, diabetic maculopathy, central serous chorioretinopathy, the pachychoroid spectrum and macular telangiectasia. Surgical vitreoretinal diseases such as vitreo-macular traction syndrome, epiretinal membrane, retinal detachment, proliferative vitreoretinal retinopathy and diabetic traction retinal detachment also will be better understood and documented with SS-OCT. This technology also provides great utility for a broad spectrum of ophthalmic pathologies including glaucoma, uveitis, tumors and anterior segment evaluation. Optic coherence tomography (OCT) is a technology introduced by Huang and associates  that becThe OCT is an imagining modality that enables the documentation of tissue structure or pathologies in real time and in situ with a resolution from 1 to 15 µm . The eyeTime-domain OCT (TD-OCT) uses low-coherence light that split into two beams at a partially reflecting mirror, one is aimed at the tissue and the other at the moving reference arm. The beams then recombine at a photodetector and the interference is assessed to determine the measurement of the tissues . Spectraautomatic real time) mode that improves the resolution with noise reduction and the eye-tracking that follows the saccadic movements and can be used in combination with the ART mode [Other features present in some commercially available SD-OCTs are the ART automaticThe recently introduced SS-OCT uses a short cavity swept laser with a tunable wavelength of operation instead of the diode laser used in spectral-domain OCT . The SS-tm SS-OCT . Its central wavelength is 1050 × nm and the speed scan is 100,000 A-scans per second.The present review highlights some important advances in retinal diagnosis provided by SS-OCT. The SS-OCT commercially available in several countries, including Brazil, is the TritonThere are other SS-OCTs that are still prototypes and to this date are not commercially available such as the Plex Elite Zeiss 9000, the vertical cavity surface emitting laser (VCSEL) OCT angiography from Massachusetts Institute of Technology (MIT) and a swept source megahertz OCT that operates with a 1050 nm wavelength and reaches scan speeds of 1.68 MHz . The VCStm (EVV), the choroid evaluation, En-face SS-OCT and OCT angiography (OCTA) and selected features of the SS-OCT utility in age-related macular degeneration (AMD), in central serous chorioretinopathy (CSCR), in diabetic maculopathy and in macular telangiectasia (Mactel).The scope of this review will include vitreous evaluation using the Enhanced Vitreous VisualizationThe vitreous is a transparent hydrophilic gel, mainly composed of water, occupying the space between the lens and the retina and functions as a pathway for nutrients utilized by the lens, ciliary body and retina . The advtm SS-OCT  uses a feature called EVV  where these different windows for vitreous visualization can be appreciated . EDI is an acquisition software that automatically captures the cross-sectional image with the choroid close to the zero delay line to maximize the sensitivity on the outer limit of the choroid , 44.The SS-OCT has a longer wavelength capable of penetrating tissue more than the SD-OCT, thus both vitreous and choroid can be imaged simultaneously. The first study to measure choroidal thickness (CT) by Margolis and Spaide investigated 54 normal, non-myopic eyes with the EDI of the SD-OCT and found a sub-foveal thickness of 287 µm . RegatieEvaluation of CT is an important diagnostic hallmark of several retinal diseases. Choroidal hyperpermeability is the main feature in CSC , CT is iP = 0.001; 95 % CI 21.88–88.44) [CT may also play a role in diabetic retinopathy. Regatieri and associates evaluated 49 eyes of type 2 diabetic retinopathy patients with SD-OCT, the sub-foveal CT was thinner in eyes with diabetic macular edema, or eyes with proliferative retinopathy and previous pan-photocoagulation . A study8–88.44) .tm  measures the CT with automated segmentation using a viewer software where each sector can be measured separately  and indocyanine green (ICGA) angiographies are the gold standard for imaging the retinal vasculature, they are dynamic and provide the evaluation of the transit of the dye as well as the well-known diagnostic hallmarks of leakage and pooling of dye , 66. HowOCTA is a noninvasive technology that uses the decorrelation motion contrast between rapidly repeated OCT B-scans to visualize blood flow to document the retinal vasculature , 69. OCTNovais and associates showed that En face OCTA images from a 1050-nm-wavelength ultrahigh-speed SS-OCT provided a more accurate representation of CNV lesions compared to 840 nm-wavelength SD-OCT . RoismanThe SS-OCT angiography has great potential both for the diagnosis of the three types of CNV and for the follow-up of the anti-vascular endothelial growth factor (anti-VEGF) treatment outcomes –77. The En face SS-OCT images are useful to diagnose pathological features of CSC Fig. . FerraraThe evaluation of the choroid with SS-OCT En Face and OCTA may help to understand the pachychoroid spectrum disorders. Densingani and associates evaluated 66 eyes with pachychoroid spectrum and found dilated choroidal vessels in all eyes. Other findings associated with chronic disease: focal choriocapillaris atrophy with inward displacement of deep choroidal vessels, were also appreciated .Another pathology involving the choroid that is better diagnosed with SS-OCT is the polypoidal choroidal vasculopathy (PCV). The diagnosis of PCV has been made based on clinical examination  and ICGA , 82. OCTThe depth of the C-scan is an essential element of the usefulness of the OCTA. The possibility to evaluate different levels such as the superficial vascular plexus, the deep vascular plexus, the choroiocapillaris and the choroidal vessels enhances the comprehension of the different retinal pathologies. The SS-OCT angiography improved the multimodal diagnosis of vascular pathologies of the retina such as macular telangiectasia, diabetic maculopathy, proliferative diabetic retinopathy and occlusive vascular diseases .Macular telangiectasia type 2 (MacTel2) is a disease of the central macula that affects all microvascular layers of the retina and includes neovascularization arising from both the retinal and choroidal circulations. The early changes of MacTel2 are in the temporal aspect of the deep vessels, then into microvasculature around the fovea in superficial vessels and into anastomoses between the deep and the superficial plexus. In some patients, the anastomoses progress to form sub-retinal neovascularization. OCTA reveals these changes at every stage of disease progression –91 Fig..Fig. 6SwDiabetic maculopathy and proliferative diabetic retinopathy tomographic features can be appreciated with detail using SS-OCT, En face SS-OCT and OCTA Fig. . StudiesThe abnormalities in perfusion seen both in diabetic retinopathy and in retinal vein or artery occlusion are well documented with OCTA. Density maps determining quantitatively this feature were described and have a potential to be an additional tool for the diagnosis or retinal vascular disease. The SS-OCT angiography prototype using variable inter-scan time analysis (VISTA) using a 1050-nm wavelength, 400-kHz A-scan rate is a promising tool to quantitatively determine flow and to assess its impairments in different retinal pathologies . The usetm) as well as segmentation and mapping of choroidal thickness. The En Face OCT and the SS-OCT angiography have improved the diagnosis and the understanding of the features of retinal pathologies including: CNV of different types and etiologies including AMD, CSC and the pachychoroid spectrum, PVC, MacTel, diabetic retinopathy and retinal vascular occlusions. The quantitative evaluation of flow parameters in different pathologies will add important information, especially the ones with vascular etiology.The SS-OCT has increased scan speed and a deeper penetration than previous modalities of OCT. The deeper penetration allows the simultaneous detailed documentation of the vitreous and the choroid. It visualizes better the choroidal structures and also sub-RPE pathology such as type 1 CNV. The commercially available SS-OCT has different windows for better vitreous visualization (Enhanced Vitreous View