Moyamoya disease (MMD) is an important cause of stroke in children and young adults in Asia. To date, diagnosis remains challenging due to varying clinical manifestations and unknown pathogenesis. ...The study aims to identify cerebrospinal fluid (CSF) exosomal microRNAs (exomiRs) that can serve as a novel diagnostic biomarker for diagnosis and assess its clinical applications.
CSF samples were taken from 31 MMD patients and 31 healthy controls. Initial screening of miRNA expression was performed on samples pooled from MMD patients and controls using microarray and validated using quantitative reverse transcription polymerase chain reaction (qRT-PCR). The diagnostic accuracy of the potential exosomal miRNAs was evaluated using receiver operating characteristic curve analyses in an independent patient cohort. The potential pathways regulated by the miRNAs was also determined using bioinformatics analysis.
The microarray results demonstrated that six exomiRs were dysregulated in the MMD patients compared to the controls. Using qRT-PCR, we validated four of the miRNAs (miR-3679-5p, miR-6165, miR-6760-5p, and miR-574-5p) as a biomarker for MMD diagnosis. The four exomiRs showed enhanced sensitivity (75%) and specificity (93.75%) in terms of differentiating MMD patients from healthy subjects area under the curve (AUC) = 0.9453. Pathway enrichment analysis for potential targets of six exomiRs identified proteins involved in cell adhesion and junction formation in the brain.
We identified a novel and highly sensitive exomiRs signature for MMD detection and explored its potential targets using bioinformatics analysis.
Twenty Newcastle disease virus (NDV) isolates from infected chicken flocks during 2001 to 2009 in China were biologically and genetically characterized. All the 20 NDVs were categorized into ...velogenic (
n
=
17) and lentogenic (
n
=
3) viruses, respectively, according to the mean death time (MDT) of chicken embryos. Velogenic viruses carry the motif
112R-R-Q-K-R/F
117 (
n
=
14) or
112G-R-Q-G-R/L
117 (
n
=
3) at the F0 cleavage site, while all the lentogenic virus had a sequence motif of
112G-R-Q-G-R/L
117 (
n
=
3) at the same site. Phylogenetic analysis revealed that at least three distinct genotypes (genotypes I, II and VII) existed in chicken flocks in China and VIId of genotype VII were mainly responsible for the present ND panzootic. Two natural recombinants (XD/Shandong/08 and QG/Hebei/07) were supposed and identified by the SimPlot program, and their two parental-like strains might be from the NDV vaccine lineage and VIId velogenic lineage respectively. The results indicate that recombination play a role in NDV evolution and the live vaccines have capacity to boost NDV evolution by homologous recombination with circulating virus.
The large (L) polymerase proteins of most nonsegmented, negative-stranded (NNS) RNA viruses have conserved methyltransferase motifs, (G)-G-G-D and K-D-K-E, which are important for the stabilization ...and translation of mRNA. However, the function of the (G)-G-G-D and K-D-K-E motifs in the NNS RNA virus Newcastle disease virus (NDV) remains unclear. We observed G-G-D and K-D-K-E motifs in all NDV genotypes. By using the infection cloning system of NDV rSG10 strain, recombinant NDVs with a single amino acid mutated to alanine in one motif (G-G-D or K-D-K-E) were rescued. The intracerebral pathogenicity index and mean death time assay results revealed that the G-G-D motif and K-D-K-E motif attenuate the virulence of NDV to various degrees. The replication, transcription, and translation levels of the K-D-K-E motif-mutant strains were significantly higher than those of wild-type virus owing to their altered regulation of the affinity between nucleocapsid protein and eukaryotic translation initiation factor 4E. When the infection dose was changed from a multiplicity of infection (MOI) of 10 to an MOI of 0.01, the cell-to-cell spread abilities of G-G-D- and K-D-K-E-mutant strains were reduced, according to plaque assay and dynamic indirect immunofluorescence assay results. Finally, we found that NDV strains with G-G-D or K-D-K-E motif mutations had less pathogenicity in 3-week-old specific-pathogen-free chickens than wild-type NDV. Therefore, these methyltransferase motifs can affect virulence by regulating the translation and cell-to-cell spread abilities of NDV. This work provides a feasible approach for generating vaccine candidates for viruses with methyltransferase motifs. IMPORTANCE Newcastle disease virus (NDV) is an important pathogen that is widespread globally. Research on its pathogenic mechanism is an important means of improving prevention and control efforts. Our study found that a deficiency in its methyltransferase motifs (G-G-D and K-D-K-E motifs) can attenuate NDV and revealed the molecular mechanism by which these motifs affect pathogenicity, which provides a new direction for the development of NDV vaccines. In addition to the (G)-G-G-D and K-D-K-E motifs of many nonsegmented, negative-stranded RNA viruses, similar motifs have been found in dengue virus, Zika virus, Japanese encephalitis virus (JEV), and severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2). This suggests that such motifs may be present in more viruses. Our finding also provides a molecular basis for the discovery and functional study of (G)-G-G-D and K-D-K-E motifs of other viruses.
Previously, we reported that the mediation of Newcastle disease virus (NDV) pathogenicity by the
524
YLMY
527
motif depends mainly on the regulation of F protein transport to the cell surface. The ...virus and host determinants that govern this intracellular trafficking remain unknown. Here, we confirmed that host adaptor protein (AP) complexes are involved in NDV infection using small interfering RNA. The transport of viral F protein to the cell surface depends on host transport proteins. We observed that the trends for host expression of AP complexes AP1M1 and AP2M1 were similar to those of mutated F proteins, especially in the membrane protein. NDV F protein interacted with AP1M1 and AP2M1, and the YLMY motif influenced this interaction. Knockdown of AP1M1 or AP2M1 suppressed the intracellular and extracellular virus titre of mutated-YLMY-motif NDVs, especially rSG10*-F/Y527A and rSG10*-F/Y524AY527A, to varying degrees. Therefore, the YLMY motif regulates AP-mediated viral F protein transportation from the cytoplasm to the cell surface and subsequently affects viral titer. We further found that the YLMY-motif mutants were differently associated with the process of AAK1 and GAK kinase-mediated AP - viral F protein interaction. These data demonstrate that the essential YLMY motif located in the NDV F protein cytoplasmic tail recruits AP to direct the F protein to the cell surface, which is necessary for its ability to affect virus budding. This study provides support for a deeper understanding of virus and host determinants that facilitate virus trafficking, which can be exploited in the design of novel antiviral therapies.
In order to solve the problem where the amount of screw fertilizer distributor can only be adjusted by rotating speed and poor fertilization uniformity at low rotational speeds, a blocking wheel-type ...screw fertilizer distributor was designed. Single factor and L9(34) orthogonal simulation tests based on EDEM software were carried out to optimize the distributor variables at a speed of 20 r/min. The bench verification test was built under the same conditions as the simulation tests to verify the results of the simulation. Finally, the bench performance tests were carried out to evaluate distributor performance. The results of simulation tests revealed that the minimum coefficient of variation of fertilization uniformity (CVFU) was 19.27%, with the structural parameter combination of the inner diameter (17 mm), pitch (45 mm), outlet distance (40 mm), and number of screw heads (1). The verification test results showed that the changing trend and values of the CVFU were almost the same as the simulation tests. The results of the performance test revealed that when the opening width of the blocking wheel was 10–30 mm and the rotation speed was 20–60 r/min, the amount of fertilizer per lap (FAPL) was in the range of 27.74–38.15 g/r; the maximum CVFU and the coefficient of variation of fertilization stability (CVFS) were 29.43% and 2.18%, respectively, which met the requirements of the industry standard. This research provides a good reference for optimizing the screw fertilizer distribution and for researchers in the field of precision fertilization.
The aim of the present study was to observe the pathological damage in the cerebral cortex of rats under acute morphine exposure (AME) and different durations of morphine dependence (MD), explore ...whether endoplasmic reticulum stress (ERS) is involved in the damage process, and assess the effect of morphine exposure on the proliferation and differentiation of newborn neurons.
Rat models of AME and different durations of MD were established. Pathological changes in cortical neurons were assessed by hematoxylin and eosin (H&E) and thionine staining. The expression of nuclear receptor-related factor 1 (NURR1) and that of the ERS-related proteins glucose-regulated protein 78 (GRP78), p-eIF2α, activating transcription factor 6 (ATF6), and CHOP in cortical neurons was assessed by immunohistochemistry. Double immunofluorescence labeling was used to observe the expression of Ki-67.
H&E and thionine staining revealed that AME resulted in pyknotic changes in cortical neurons. With prolonged morphine exposure, the number of pyknotic neurons was significantly increased, the protein expression of Ki-67 and NURR1 was significantly decreased, and the protein levels of GRP78, p-eIF2α, ATF6, and CHOP showed marked dynamic changes.
AME and different durations of MD caused varying degrees of pathological changes in the cortex. Furthermore, the dynamic changes observed in ERS-related protein expression suggested that ERS may be associated with cortical injury. Different durations of MD inhibited the proliferation, differentiation, and migration of newborn neurons, which may affect the nerve repair process after injury.
Consumer-grade cameras are being increasingly used for remote sensing applications in recent years. However, the performance of this type of cameras has not been systematically tested and well ...documented in the literature. The objective of this research was to evaluate the performance of original and resolution-reduced images taken from two consumer-grade cameras, a RGB camera and a modified near-infrared (NIR) camera, for crop identification and leaf area index (LAI) estimation. Airborne RGB and NIR images taken over a 6.5-square-km cropping area were mosaicked and aligned to create a four-band mosaic with a spatial resolution of 0.4 m. The spatial resolution of the mosaic was then reduced to 1, 2, 4, 10, 15 and 30 m for comparison. Six supervised classifiers were applied to the RGB images and the four-band images for crop identification, and 10 vegetation indices (VIs) derived from the images were related to ground-measured LAI. Accuracy assessment showed that maximum likelihood applied to the 0.4-m images achieved an overall accuracy of 83.3% for the RGB image and 90.4% for the four-band image. Regression analysis showed that the 10 VIs explained 58.7% to 83.1% of the variability in LAI. Moreover, spatial resolutions at 0.4, 1, 2 and 4 m achieved better classification results for both crop identification and LAI prediction than the coarser spatial resolutions at 10, 15 and 30 m. The results from this study indicate that imagery from consumer-grade cameras can be a useful data source for crop identification and canopy cover estimation.
The matrix (M) protein FPIV L-domain is conserved among multiple paramyxoviruses; however, its function and the associated mechanism remain unclear. In this study, the paramyxovirus Newcastle disease ...virus (NDV) was employed to study the FPIV L-domain. Two recombinant NDV strains, each carrying a single amino acid mutation at the Phe (F23) or Pro (P24) site of
FPIV/I
L-domain, were rescued. Growth defects were observed in only the recombinant SG10-F23A (rSG10-F23A) strain. Subsequent studies focused on rSG10-F23A revealed that the virulence, pathogenicity, and replication ability of this strain were all weaker than those of wild-type strain rSG10 and that a budding deficiency contributed to those weaknesses. To uncover the molecular mechanism underlying the rSG10-F23A budding deficiency, the bridging proteins between the FPIV L-domain and endosomal sorting complex required for transported (ESCRT) machinery were explored. Among 17 candidate proteins, only the charged multivesicular body protein 4 (CHMP4) paralogues were found to interact more strongly with the NDV wild-type M protein (M-WT) than with the mutated M protein (M-F23A). Overexpression of M-WT, but not of M-F23A, changed the CHMP4 subcellular location to the NDV budding site. Furthermore, a knockdown of CHMP4B, the most abundant CHMP4 protein, inhibited the release of rSG10 but not that of rSG10-F23A. From these findings, we can reasonably infer that the F23A mutation of the FPIV L-domain blocks the interaction between the NDV M protein and CHMP4B and that this contributes to the budding deficiency and consequent growth defects of rSG10-F23A. This work lays the foundation for further study of the FPIV L-domain in NDV and other paramyxoviruses.
Multiple viruses utilize a conserved motif, termed the L-domain, to act as a cellular adaptor for recruiting host ESCRT machinery to their budding site. Despite the FPIV type L-domain having been identified in some paramyxoviruses 2 decades ago, its function in virus life cycles and its method of recruiting the ESCRT machinery are poorly understood. In this study, a single amino acid mutation at the F23 site of the
FPIV
L-domain was found to block NDV budding at the late stage. Furthermore, CHMP4B, a core component of the ESCRT-III complex, was identified as a main factor that links the FPIV L-domain and ESCRT machinery together. These results extend previous understanding of the FPIV L-domain and, therefore, not only provide a new approach for attenuating NDV and other paramyxoviruses but also lay the foundation for further study of the FPIV L-domain.
Combined bypass is commonly used in adult Moyamoya disease (MMD) for revascularization purposes. The blood flow from the external carotid artery system supplied by the superficial temporal artery ...(STA), middle meningeal artery (MMA), and deep temporal artery (DTA) can restore the impaired hemodynamics of the ischemic brain. In this study we attempted to evaluate the hemodynamic changes of the STA graft and predict the angiogenesis outcomes in MMD patients after combined bypass surgery by using quantitative ultrasonography.
We retrospectively studied Moyamoya patients who were treated by combined bypass between September 2017 and June 2021 in our hospital. We quantitatively measured the STA with ultrasound and recorded the blood flow, diameter, pulsatility index (PI) and resistance index (RI) to assess graft development preoperatively and at 1 day, 7 days, 3 months, and 6 months after surgery. All patients received both pre- and post- operative angiography evaluation. Patients were divided into either well- or poorly-angiogenesis groups according to the transdural collateral formation status on angiography at 6 months after surgery (W group or P group). Patients with matshushima grade A or B were divided into W group. Patients with matshushima grade C were divided into P group, indicating a poor angiogenesis development.
A total of 52 patients with 54 operated hemispheres were enrolled, including 25 men and 27 women with an average age of 39 ± 14.3 years. Compared to preoperative values, the average blood flow of an STA graft at day 1 postoperation increased from 16.06 ± 12.47 to 117.47± 73.77 (mL/min), diameter increased from 1.14 ± 0.33 to 1.81 ± 0.30 (mm), PI dropped from 1.77 ± 0.42 to 0.76 ± 0.37, and RI dropped from 1.77 ± 0.42 to 0.50 ± 0.12. According to the Matsushima grade at 6 months after surgery, 30 hemispheres qualified as W group and 24 hemispheres as P group. Statistically significant differences were found between the two groups in diameter (
= 0.010) as well as flow (
= 0.017) at 3 months post-surgery. Flow also remained significantly different at 6 months after surgery (
= 0.014). Based on GEE logistic regression evaluation, the patients with higher levels of flow post-operation were more likely to have poorly-compensated collateral. ROC analysis showed that increased flow of ≥69.5 ml/min (
= 0.003; AUC = 0.74) or a 604% (
= 0.012; AUC = 0.70) increase at 3 months post-surgery compared with the pre-operative value is the cut-off point which had the highest Youden's index for predicting P group. Furthermore, a diameter at 3 months post-surgery that is ≥0.75 mm (
= 0.008; AUC = 0.71) or 52% (
=0.021; AUC = 0.68) wider than pre-operation also indicates a high risk of poor indirect collateral formation.
The hemodynamic of the STA graft changed significantly after combined bypass surgery. An increased flow of more than 69.5 ml/min at 3 months was a good predictive factor for poor neoangiogenesis in MMD patients treated with combined bypass surgery.
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