Expression of the CFH in ARPE-19 cells.

<p>The immunofluorescence results showed CFH expression (green) in the cytoplasm of ARPE-19 cells (A). Immunofluorescence staining was not detected in control wells where only secondary antibody was added (B).</p

CFH RNA interference in ARPE-19 cells.

<p>(A) Western blot assay: CFH expression showed a reduction in RPE cells after being transfected with specific siRNA (100 and 50 nM siRNA) relative to the control (mock transfection with Lipofectamine reagent). The CFH expression level in the 100 nM CFH-siRNA transfected ARPE-19 cells was lower than that in the 50nM CFH-siRNA transfected cells. β-Actin served as an internal control. <b>(B)</b>. Immunofluorescence antibody labeling also showed that CFH immunostaining (green) in ARPE-19 cells transfected with 100 nM specific siRNA was weaker than in controls. The mock transfection served as controls. <b>(C).</b> Real-time PCR results showed CFH expression was knocked down by 88% using specific siRNA. Data are presented as mean ± SD (n = 9; T test; P < 0.01). <b>(D).</b> Complement activation: enzyme immunoassay confirmed that the C3a level in ARPE-19 cells after being transfected with specific siRNA (100 nM siRNA) was increased relative to the controls (mock transfection). Data are presented as mean ± SD (P < 0.01). All the experiments were carried out in triplicate for each condition and repeated three times.</p

Cultured human umbilical vein endothelial cells (HUVECs) confirmed by immunohistochemistry.

<p>The cells have positive speckled immunostaining for CD31 (A) compare to Negative control (B).</p

Complement activation <i>in vitro</i>.

<p>C3a content measured by ELISA in H₂O₂-damaged ARPE-19 cells was significantly higher than that in controls (P < 0.05). Results were expressed as mean ± SD. Experiments were carried out in triplicate for each condition and repeated three times independently.</p

DataSheet_1_Classification of Early and Late Mild Cognitive Impairment Using Functional Brain Network of Resting-State fMRI.pdf

Using the Pearson correlation coefficient to constructing functional brain network has been evidenced to be an effective means to diagnose different stages of mild cognitive impairment (MCI) disease. In this study, we investigated the efficacy of a classification framework to distinguish early mild cognitive impairment (EMCI) from late mild cognitive impairment (LMCI) by using the effective features derived from functional brain network of three frequency bands (full-band: 0.01–0.08 Hz; slow-4: 0.027–0.08 Hz; slow-5: 0.01–0.027 Hz) at Rest. Graphic theory was performed to calculate and analyze the relationship between changes in network connectivity. Subsequently, three different algorithms [minimal redundancy maximal relevance (mRMR), sparse linear regression feature selection algorithm based on stationary selection (SS-LR), and Fisher Score (FS)] were applied to select the features of network attributes, respectively. Finally, we used the support vector machine (SVM) with nested cross validation to classify the samples into two categories to obtain unbiased results. Our results showed that the global efficiency, the local efficiency, and the average clustering coefficient were significantly higher in the slow-5 band for the LMCI–EMCI comparison, while the characteristic path length was significantly longer under most threshold values. The classification results showed that the features selected by the mRMR algorithm have higher classification performance than those selected by the SS-LR and FS algorithms. The classification results obtained by using mRMR algorithm in slow-5 band are the best, with 83.87% accuracy (ACC), 86.21% sensitivity (SEN), 81.21% specificity (SPE), and the area under receiver operating characteristic curve (AUC) of 0.905. The present results suggest that the method we proposed could effectively help diagnose MCI disease in clinic and predict its conversion to Alzheimer’s disease at an early stage.</p

Cell viability of ARPE-19 cells.

<p>The viability of ARPE-19 cells was assessed by WST-1 assay. ARPE-19 cells were subjected to a single dose of 400μM H₂O₂ treatment for 24h and the untreated cells served as controls. The data showed that there was no significant difference in the cell viability between H₂O₂ treated and control cells (defined as 100%) (P > 0.05). The experiment was performed three times independently with triplicate in each group.</p

CFH expression reduced in the H₂O₂ treated ARPE-19 cells.

<p>Real-time PCR showed that the CFH mRNA expression in H₂O_2-damaged ARPE-19 cells was significantly lower than that in control cells (P < 0.05) <b>(A).</b> In Western blots, the CFH band of H₂O_2-damaged ARPE-19 cells was paler than the normal cells <b>(B).</b> The bands’ gray scale was quantified by Quantity One software. Compared to control cells, the CFH expression was reduced significantly in H₂O_2-damaged ARPE-19 cells (P < 0.05) <b>(C).</b> Data are presented as mean ± SD from results done with three independent biological samples, each analyzed with triplicates.</p

The tube formation by HUVECs.

<p>(A) and (B) showed that more tubes were formed by HUVECs in co-cultures with CFH-siRNA transfected ARPE-19 cells compared with control ARPE-19 cells at 24 and 72 h, respectively (P< 0.05). (C) and (D) showed that, in the absence of human serum, there were no significant changes in tube formation at 72h between CFH siRNA transfected RPE- and control non-transfected RPE-HUVEC co-cultures. Data were expressed as mean ± SD from results done with three independent biological samples, each analyzed with triplicates.</p

Effect of injured RPE cells on HUVEC migration.

<p>HUVEC migration was examined with a phase-contrast microscopy (×40) at the indicated time points (0h, 6h, 10h)(A). The wounding width was quantified, and bars represent the migration index of each treatment. The assay showed a significant increase of the distance moved by HUVECs treated with human serum that was cultured with H₂O₂-damaged ARPE-19 cells, when compared to HUVECs treated with the serum that was cultured with control ARPE-19 cells (P < 0.05) (B). The data were expressed as mean ± SD from results done with three independent biological samples, each analyzed with triplicates.</p

Video_2_New aneurysm formation after endovascular embolization of a vertebral epidural AV fistula: a rare sequelae of NF AV fistulae.MP4

BackgroundNeurofibromatosis type 1 (NF-1) is a dominant genetic disorder often accompanied by lesions of the neurovascular system. Patients with NF-1 are predisposed to unique vertebral artery fistula (AVF).Case descriptionWe report on a rare case of multiple neurovascular abnormalities in a 47-year-old man with neurofibromatosis. He was admitted due to a sudden headache and was found to have suffered a subarachnoid hemorrhage from a left vertebral arteriovenous fistula. He underwent two endovascular procedures complicated by a delayed extraspinal mass 7 days after treatment. Angiography revealed a new vascular abnormality, and although we performed another embolization, it failed to respond to further embolization.ConclusionVascular abnormalities in patients with NF-1 can be complex. Endovascular intervention remains feasible for NF-1 related AVF, however, partial occlusion of the fistula should be avoided to limit and iatrogenic damage to the blood vessels.</p