Development of a diagnostic and risk prediction model for Alzheimer's disease through integration of single-cell and bulk transcriptomic analysis of glutamine metabolism.

Background: In this study, we present a novel system for quantifying glutamine metabolism (GM) to enhance the effectiveness of Alzheimer's disease (AD) diagnosis and risk prediction. Methods: Single-cell RNA sequencing (scRNA-seq) analysis was utilized to comprehensively assess the expression patterns of GM. The WGCNA algorithm was applied to investigate the most significant genes related to GM. Subsequently, three machine learning algorithms (Boruta, LASSO, and SVM-RFE) were employed to identify GM-associated characteristic genes and develop a risk model. Patients were divided into high- and low-risk groups based on this model. Moreover, we explored biological properties, distinct signaling pathways, and immunological characteristics of AD patients at different risk levels. Finally, in vitro and in vivo models of AD were constructed to validate the characteristics of the feature genes. Results: Both scRNA-seq and bulk transcriptomic analyses revealed increased GM activity in AD patients, specifically in certain cell subsets (pDC, Tem/Effector helper T cells (LTB), and plasma cells). Cells with higher GM scores demonstrated more significant numbers and strengths of interactions with other cell types. The WGCNA algorithm identified 360 genes related to GM, and a risk score was constructed based on nine characteristic genes (ATP13A4, PIK3C2A, CD164, PHF1, CES2, PDGFB, LCOR, TMEM30A, and PLXNA1) identified through multiple machine learning algorithms displayed reliable diagnostic efficacy for AD onset. Nomograms, calibration curves, and decision curve analysis (DCA) based on these characteristic genes provided significant clinical benefits for AD patients. High-risk AD patients exhibited higher levels of immune-related functions and pathways, increased immune cell infiltration, and elevated expressions of immune modulators. RT-qPCR analysis revealed that the majority of the nine characteristic genes were differentially expressed in AD-induced rat neurons. Knocking down PHF1 could protect against neurite loss and alleviate cell injury in AD neurons. In vivo, down-regulation of PHF1 in AD models decreases GM metabolism levels and modulates the immunoinflammatory response in the brain. Conclusion: This comprehensive identification of gene expression patterns contributes to a deeper understanding of the underlying pathological mechanisms driving AD pathogenesis. Furthermore, the risk model based on the nine-gene signature offers a promising theoretical foundation for developing individualized treatments for AD patients.

Macroscopic Properties and Pore Structure Fractal Characteristics of Alkali-Activated Metakaolin-Slag Composite Cementitious Materials.

To investigate the effects of slag and Na2O content on the macroscopic properties and pore structure characteristics of alkali-activated metakaolin-slag (AAMS) composite cementitious materials, this study used X-ray diffraction (XRD), Fourier transform infrared spectroscopy (FT-IR), scanning electron microscopy (SEM-EDS), and mercury-pressure (MIP) tests for characterization and analyzed the hydration product compositions, microstructures, and pore structure characteristics of AAMS composite cementitious materials. The relationships between the fractal dimension and the pore structure parameters, compressive strengths, and drying shrinkage rates of AAMS composite cementitious materials were investigated with the thermodynamic fractal model. The results showed that at the age of 28 d, the compressive strength and drying shrinkage of the AAMS composite binder increased by 20.57% and 215.11%, respectively, when the slag content increased from 0 to 50%. When the Na2O content increased from 8% to 12%, the compressive strength and drying shrinkage of the AAMS composite increased by 24.37% and 129.40%, respectively. The compressive strengths of AAMS composite cementitious materials increased with increasing slag content and Na2O content, but the drying shrinkage of the system increased, and the volume stability worsened. Microscopic analyses showed that with increases in the slag and Na2O contents, the hydration products of AAMS composite cementitious materials increased, and C-A-S-H and N-A-S-H filled each other so that the internal structures of AAMS composite cementitious materials were denser, and the porosities were significantly reduced. By comparing and analyzing the Menger sponge model and the fractal model based on the thermodynamic relationships, it was found that the fractal model based on the thermodynamic relationship better reflected the pore size distribution over the whole pore size determination range, and the correlation coefficients R2 were above 0.99, indicating that the fractal dimension calculated by the fractal model based on the thermodynamic relationship provided a comprehensive evaluation index for the pore structure characteristics of AAMS composite cementitious materials, and the fractal dimension correlated well with the pore structure parameters, compressive strengths, and drying shrinkage rates of cementitious materials.

Four-copy number alteration (CNA)-related lncRNA prognostic signature for liver cancer.

The objective of this study was to identify CNA-related lncRNAs that can better evaluate the prognosis of patients with liver cancer. Prognostic molecular subtypes were identified, followed by tumor mutation and differential expression analyses. Genomic copy number anomalies and their association with lncRNAs were also evaluated. A risk model was built based on lncRNAs, as well as a nomogram, and the differences in the tumor immune microenvironment and drug sensitivity between the High_ and Low_risk groups were compared. Weighted gene co-expression network analysis was used to identify modules with significant enrichment in prognostic-related lncRNAs. In total, two subtypes were identified, TP53 and CTNNB1 were common high-frequency mutated genes in the two subtypes. A total of 8,372 differentially expressed (DE) mRNAs and 798 DElncRNAs were identified between cluster1 and cluster2. In addition, a four-lncRNA signature was constructed, and statistically significant differences between the Low_ and High_risk groups were found in terms of CD8 T cells, resting memory CD4 T cells, etc. Enrichment analysis showed that prognostic-related lncRNAs were involved in the cell cycle, p53 signaling pathway, non-alcoholic fatty liver disease, etc. A prognostic prediction signature, based on four-CNA-related lncRNAs, could contribute to a more accurate prognosis of patients with liver cancer.

Effect of Slag on the Strength and Shrinkage Properties of Metakaolin-Based Geopolymers.

Metakaolin-based geopolymers possess excellent corrosion and high-temperature resistance, which are advantageous compared to ordinary Portland cement. The addition of slag in metakaolin-based geopolymers is a promising approach to improve their mechanical properties. Thus, this study investigated the effect of slag content on the strength and shrinkage properties of metakaolin-based geopolymers. Increasing the slag content and Na2O content was beneficial to the reaction of alkali-activated metakaolin-based geopolymers, thereby improving their compressive strength and density. After 56 days of aging, a maximum compressive strength of 86.1 MPa was achieved for a metakaolin-based geopolymer with a slag content of 50 mass%. When the Na2O content was 12%, the compressive strength of the metakaolin geopolymers with a slag content of 30% was 42.36% higher than those with a Na2O content of 8%. However, as the slag and alkali contents increased, the reaction rate of the metakaolin-based geopolymers increased, which significantly decreased the porosity, increased the shrinkage, and decreased the volumetric stability of the system. In this paper, in-depth study of the volume stability of alkali-activated metakaolin-based geopolymers plays an important role in further understanding, controlling, and utilizing the deformation behavior of geopolymers.

Heterologous overexpression of Sup35 in Escherichia coli leads to both monomer and complex states.

The heterologous overexpression states of prion proteins play a critical role in understanding the mechanisms of prion-related diseases. We report herein the identification of soluble monomer and complex states for a bakers' yeast prion, Sup35, when expressed in Escherichia coli. Two peaks are apparent with the elution of His-tagged Sup35 by imidazole from a Ni2+ affinity column. Peak I contains Sup35 in both monomer and aggregated states. Sup35 aggregate is abbreviated as C-aggregate and includes a non-fibril complex comprising Sup35 aggregate-HSP90-Dna K, ATP synthase ß unit (chain D), 30S ribosome subunit, and Omp F. The purified monomer and C-aggregate can remain stable for an extended period of time. Peak II contains Sup35 also in both monomer and aggregated (abbreviated as S-aggregate) states, but the aggregated states are caused by the formation of inter-Sup35 disulfide bonds. This study demonstrates that further assembly of Sup35 non-fibril C-aggregate can be interrupted by the chaperone repertoire system in E. coli.

Understanding the Role of Metakaolin towards Mitigating the Shrinkage Behavior of Alkali-Activated Slag.

This research investigates the mechanism of metakaolin for mitigating the autogenous and drying shrinkages of alkali-activated slag with regard to the activator parameters, including concentration and modulus. The results indicate that the incorporation of metakaolin can decrease the initial viscosity and setting time. Increasing activator concentration can promote the reaction process and shorten the setting time. An increase in the metakaolin content induces a decrease in compressive strength due to reduced formation of reaction products. However, increasing activator dosage and modulus can improve the compressive strength of alkali-activated slag containing 30% metakaolin. The inclusion of metakaolin can mitigate the autogenous and drying shrinkage of alkali-activated slag by coarsening the pore structure. On the other hand, increases in activator concentration and modulus result in an increase in magnitude of the autogenous and drying shrinkage of alkali-activated slag containing metakaolin. The influence of the activator modulus on the shrinkage behavior of alkali-activated slag-metakaolin binary system should be further investigated.

Functional Foxp3 polymorphisms and the susceptibility to cancer: An update meta-analysis.

BACKGROUND: Forkhead box P3 (Foxp3) plays important roles in the development and pathogensis of cancer. To investigate the association of 3 polymorphisms of Foxp3 (rs3761548, rs 3761549 and rs2280883) and cancer risk, an updated meta-analysis was performed. METHODS: Around 11 studies including 4344 cancer patients and 4665 healthy controls were selected for this meta-analysis. There were nine studies with 3783 cases and 4096 controls for rs3761548, 4 studies with 1669 cases and 1613 controls for rs3761549 and 4 studies with 1821 cases and 1799 controls for rs2280883. Odds radios (ORs) and 95% confidence intervals (CIs) were used to evaluate the cancer risk. RESULTS: Meta-analysis showed that rs3761548 was associated with an increased cancer risk in the overall population under the recessive model (AA vs CA + CC: OR = 1.45, 95%CI = 1.03-2.02, P = .03). No association was found between rs3761549, rs2280883 polymorphisms, and cancer susceptibility in the overall population. Nonetheless, in the genotyping methods subgroup analysis of rs2280883, a lower risk of cancer was found in studies using polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP) under the allelic model (C vs T: OR = 0.70, 95%CI = 0.52-0.95, P = .02), heterozygote model (TC vs TT: OR = 0.60, 95%CI = 0.41-0.87, P = .008) and dominant model (CC + TC vs TT: OR = 0.63, 95%CI = 0.45-0.90, P = .01). In the subgroup analysis by cancer types showed C allele or TC carriers were insusceptible to cancer under 3 genetic models (C vs T: OR = 0.78, 95%CI = 0.64-0.95, P = .01; TC vs TT: OR = 0.50, 95%CI = 0.32-0.79, P = .003; CC + TC vs TT: OR = 0.64, 95%CI = 0.51-0.82, P < .001). CONCLUSION: Our results suggest that rs3761548 polymorphism is associated with cancer risk.

Hypoxia inducible factor-1α regulates autophagy via the p27-E2F1 signaling pathway.

Autophagy is a highly conserved process by which the cell contents are delivered to lysosomes for degradation, or are used to provide macromolecules for energy generation under conditions of nutritional starvation. It has previously been demonstrated that cancer cells in hypoxic regions, with an oxygen concentration below the normal physiological level, express hypoxia inducible factor (HIF)­1α, in order to adapt and survive. HIF­1α is important in the regulation of oxygen homeostasis and the transcription of hundreds of genes in response to conditions of hypoxia, hence maintaining energy and redox homeostasis. To determine if HIF­1α modulates autophagy and the underlying molecular mechanisms regulating this process, the human esophageal cancer EC109 and IMR90 human diploid fibroblast cell lines were exposed to normoxic or hypoxic conditions and the expression levels of various proteins subsequently examined. Small interfering RNA was used to silence p27, in order to investigate its role in the process of HIF­1α regulated autophagy. Hypoxia induced autophagy in IMR90 cells and it was revealed that immature IMR90 cells demonstrated an increased rate of autophagy compared with mature cells. HIF­1α promoted EC109 cell autophagy via positively modulating p27, whereas silencing of p27 abolished the autophagy induced by hypoxia. The present study identified the primary components of the p27­E2F1 signaling pathway by which HIF­1α regulates autophagy. A previously unidentified mechanism is here presented, via which cancer cells may generate energy, or obtain macromolecules for survival.

A cypovirus VP5 displays the RNA chaperone-like activity that destabilizes RNA helices and accelerates strand annealing.

For double-stranded RNA (dsRNA) viruses in the family Reoviridae, their inner capsids function as the machinery for viral RNA (vRNA) replication. Unlike other multishelled reoviruses, cypovirus has a single-layered capsid, thereby representing a simplified model for studying vRNA replication of reoviruses. VP5 is one of the three major cypovirus capsid proteins and functions as a clamp protein to stabilize cypovirus capsid. Here, we expressed VP5 from type 5 Helicoverpa armigera cypovirus (HaCPV-5) in a eukaryotic system and determined that this VP5 possesses RNA chaperone-like activity, which destabilizes RNA helices and accelerates strand annealing independent of ATP. Our further characterization of VP5 revealed that its helix-destabilizing activity is RNA specific, lacks directionality and could be inhibited by divalent ions, such as Mg(2+), Mn(2+), Ca(2+) or Zn(2+), to varying degrees. Furthermore, we found that HaCPV-5 VP5 facilitates the replication initiation of an alternative polymerase (i.e. reverse transcriptase) through a panhandle-structured RNA template, which mimics the 5'-3' cyclization of cypoviral positive-stranded RNA. Given that the replication of negative-stranded vRNA on the positive-stranded vRNA template necessitates the dissociation of the 5'-3' panhandle, the RNA chaperone activity of VP5 may play a direct role in the initiation of reoviral dsRNA synthesis.

The nonstructural protein 2C of a Picorna-like virus displays nucleic acid helix destabilizing activity that can be functionally separated from its ATPase activity.

Picorna-like viruses in the Picornavirales order are a large group of positive-strand RNA viruses that include numerous important pathogens for plants, insects, and humans. In these viruses, nonstructural protein 2C is one of the most conserved proteins and contains ATPase activity and putative RNA helicase activity. Here we expressed 2C protein of Ectropis obliqua picorna-like virus (EoV; genus Iflavirus, family Iflaviridae, order Picornavirales) in a eukaryotic expression system and determined that EoV 2C displays ATP-independent nucleic acid helix destabilizing and strand annealing acceleration activity in a concentration-dependent manner, indicating that this picornaviral 2C is more like an RNA chaperone than like the previously predicted RNA helicase. Our further characterization of EoV 2C revealed that divalent metal ions, such as Mg(2+) and Zn(2+), inhibit 2C-mediated helix destabilization to different extents. Moreover, we determined that EoV 2C also contains ATPase activity like that of other picornaviral 2C proteins and further assessed the functional relevance between its RNA chaperone-like and ATPase activities using mutational analysis as well as their responses to Mg(2+). Our data show that, when one of the two 2C activities was dramatically inhibited or almost abolished, the other activity could remain intact, showing that the RNA chaperone-like and ATPase activities of EoV 2C can be functionally separated. This report reveals that a picorna-like virus 2C protein displays RNA helix destabilizing and strand annealing acceleration activity, which may be critical for picornaviral replication and pathogenesis, and should foster our understanding of picorna-like viruses and viral RNA chaperones.

Identification and characterization of Iflavirus 3C-like protease processing activities.

Viral replication and capsid assembly in the viruses in the order Picornavirales requires polyprotein proteolytic processing by 3C or 3C-like (3CL) proteases. We identified and characterized the 3CL protease of Ectropis obliqua virus (EoV) of the newly established family Iflaviridae (order Picornavirales). The bacterially expressed EoV 3CL protease domain autocatalytically released itself from larger precursors by proteolytic cleavage, and cleavage sites were determined via N-terminal sequencing of the cleavage products. This protease also mediated trans-proteolytic activity and cleaved the polyprotein at the same specific positions. Moreover, we determined the critical catalytic residues (H2261, D2299, C2383) for the protease activity, and characterized the biochemical properties of EoV 3CL and its responses to various protease inhibitors. Our work is the first study to identify an iflaviral 3CL protease and further characterize it in detail and should foster our understanding of EoV and other iflaviruses.