Search

microRNA-29a (miR-29a) increases with age in humans and mice, and, in the brain, it has a role in neuronal maturation and response to inflammation. We previously found higher miR-29a levels in the human brain to be associated with faster antemortem cognitive decline, suggesting that lowering miR-29a levels could ameliorate memory impairment in the 5×FAD AD mouse model. To test this, we generated an adeno-associated virus (AAV) expressing GFP and a miR-29a "sponge" or empty vector. We found that the AAV expressing miR-29a sponge functionally reduced miR-29a levels and improved measures of memory in the Morris water maze and fear condition paradigms when delivered to the hippocampi of 5×FAD and WT mice. miR-29a sponge significantly reduced hippocampal beta-amyloid deposition in 5×FAD mice and lowered astrocyte and microglia activation in both 5×FAD and WT mice. Using transcriptomic and proteomic sequencing, we identified Plxna1 and Wdfy1 as putative effectors at the transcript and protein level in WT and 5×FAD mice, respectively. These data indicate that lower miR-29a levels mitigate cognitive decline, making miR-29a and its target genes worth further evaluation as targets to mitigate Alzheimer's disease (AD).

* Context.--Regulatory T-cell (Treg) detection in peripheral blood, based on flow cytometry, is invaluable for diagnosis and treatment of immune-mediated diseases. However, there is a lack of reliable methods to verify the performance, which is pivotal toward standardization of the Tregs assay. Objective.--To conduct standardization studies and verify the performance of 3 commercially available reagent sets for the Tregs assay based on flow cytometry and agreement analysis for Treg detection across the different reagent sets. Design.--The analytical performance of Tregs assay using reagent sets supplied by 3 manufacturers was evaluated after establishing the gating strategy and determining the optimal antibody concentration. Postcollection sample stability was evaluated, as well as the repeatability, reproducibility, reportable range, linearity, and assay carryover. Agreement between the different assays was assessed via Bland-Altman plots and linear regression analysis. The relationship between the frequency of [CD4.sup.+][CD25.sup.+][CD127.sup.low/-] Tregs and [CD4.sup.+][CD25.sup.+][Foxp3.sup.+] Tregs was evaluated. Results.--The postcollection sample stability was set at 72 hours after collection at room temperature. The accuracy, repeatability, reproducibility, and accuracy all met the requirements for clinical analysis. Excellent linearity, with [R.sup.2] [greater than or equal to] 0.9 and no assay carryover, was observed. For reportable range, a minimum of 1000 events in the [CD3.sup.+][CD4.sup.+] gate was required for Tregs assay. Moreover, the results for Tregs labeled by antibodies from the 3 manufacturers were in good agreement. The percentage of [CD4.sup.+][CD25.sup.+][CD127.sup.low/-] Tregs was closely correlated with [CD4.sup.+][CD25.sup.+][Foxp3.sup.+] Tregs. Conclusions.--This is the first study to evaluate systematically the measurement performance of Tregs in peripheral blood by flow cytometry, which provides a practical solution to verifying the performance of flow cytometry-based immune monitoring projects in clinical practice. (Arch Pathol Lab Med. 2024;148:1234-1243; doi: 10.5858/arpa.2023-0284-OA), A wide range of immunologic methods is available for monitoring the immune system to aid in the diagnosis and clinical management of immune-mediated diseases, such as autoimmune diseases, infectious diseases, [...]

Nucleotide changes in gene regulatory elements are important determinants of neuronal development and diseases. Using massively parallel reporter assays in primary human cells from mid-gestation cortex and cerebral organoids, we interrogated the cis-regulatory activity of 102,767 open chromatin regions, including thousands of sequences with cell type-specific accessibility and variants associated with brain gene regulation. In primary cells, we identified 46,802 active enhancer sequences and 164 variants that alter enhancer activity. Activity was comparable in organoids and primary cells, suggesting that organoids provide an adequate model for the developing cortex. Using deep learning we decoded the sequence basis and upstream regulators of enhancer activity. This work establishes a comprehensive catalog of functional gene regulatory elements and variants in human neuronal development.

Abstract DNA methylation modifications are an important mechanism affecting the process of atherosclerosis (AS). Previous studies have shown that Galectin-8 (GAL8) DNA methylation level is associated with sudden death of coronary heart disease or acute events of coronary heart disease. However, the mechanism of GAL8 DNA methylation and gene expression in AS has not been elucidated, prompting us to carry out further research on it. ApoE-/- mice were used to establish an atherosclerosis model, and DNA methylation inhibitor DO05 and MAPK/mTOR inhibitor UO126 were used for intervention. Pyrosequencing was used to detect changes in GAL8 DNA methylation levels of the mouse aorta between groups. ROC curve analysis was performed to assess the relationship between GAL8 DNA methylation and atherosclerosis. Aortic staining with hematoxylin and eosin (H&E) was used to observe the aortic intima, plaque area, and characteristics of secondary lesions within the plaque. Oil Red O staining was used to detect lipid deposition in mouse arterial plaques or macrophages. Movat staining was used to detect the number of foam cells in the plaque. Immunohistochemistry (IHC) and Western blot were used to quantify the localization and expression levels of DNA methyltransferase1 (DNMT1), GAL8, MAPK/mTOR pathway proteins, Light Chain3 (LC3), Beclin1, Sequestosome1 (p62), Tumor Necrosis Factor-α (TNF-α), and other proteins. Immunofluorescence (IF) was used to detect the fluorescence intensity of GAL8, LC3, Monocyte chemoattractant protein-1(MCP-1), and other proteins. Detection of autophagosomes in macrophages by transmission electron microscopy was also performed. The foam cell model was induced with human monocytes (THP-1) and co-cultured with foam cells using siRNAs targeting GAL8, DO05, and UO126. The level of DNMT1 was detected by Western blot; Oil red O staining was used to detect lipid deposition in foam cells in each group, and the localization and expression levels of GAL8, MAPK/mTOR pathway proteins, LC3, Beclin1, p62, and TNF-α were quantitatively determined by Western blot. Immunofluorescence (IF) was used to detect the fluorescence intensity of GAL8, MAPK/mTOR pathway protein, LC3, p62, TNF-α, and other proteins. The GAL-8 promoter region harbors six CpG sites susceptible to DNA methylation. Following DNMT1 inhibition, the DC05 group displayed a significant decrease in methylation across all six CpG sites compared to the C57 and AS groups. Conversely, the UO126 group exhibited increased methylation at the first three CpG loci relative to the AS group. ROC curve analysis revealed GAL8 DNA methylation as an independent risk factor for atherosclerosis: GAL8, along with inflammation-related proteins MCP-1, MMP9, and TNF-α, were upregulated in the mouse lesion group, while expression of autophagy-related proteins LC3 and Beclin1 was downregulated. Additionally, phosphorylated MAPK/mTOR pathway proteins were detected in the mouse model of atherosclerosis. After inhibiting the methylation level of GAL-8 DNA, the expression of GAL-8 was up-regulated, macrophage autophagy was inhibited, inflammation was increased, and atherosclerotic lesions in mice were aggravated. After direct inhibition of the activity of the MAPK/mTOR pathway, macrophage autophagy was further weakened, the inflammatory response was further aggravated, and the atherosclerotic lesions of mice were further aggravated. After the specific knockdown of GAL-8 using siRNA GAL-8 using foam cells, the above phenomenon was reversed, macrophage autophagy was promoted, the inflammatory response was reduced, and the degree of atherosclerosis was alleviated. The degree of GAL8 DNA methylation is related to the progression of atherosclerosis, and its hypomethylation can aggravate atherosclerotic lesions. The mechanism may be through the regulation of MAPK/mTOR pathway to slow down the autophagy of macrophages, and then aggravate the inflammation in plaques. Targeting GAL8 DNA methylation may be a new target for the diagnosis and treatment of atherosclerosis.

Abstract Background We sought to explore the molecular mechanisms underpinning acute lung injury (ALI) caused by poisoning with paraquat (PQ). Methods Selection mice were intraperitoneally injected with PQ at 40 mg/kg, whereas controls were injected with sterile saline. On days 2, 7, and 14 after administration, mice were anesthetized and sacrificed, and lung tissue was removed. Lung pathological changes were observed with conventional staining techniques. Lung tissue components were assessed with tandem mass spectrometry tag technology, and differentially expressed proteins (DEPs) were bioinformatically analyzed and investigated with parallel reaction monitoring. Results The expression of 91, 160, and 78 proteins was significantly altered at days 2, 7, and 14, respectively. Gene Ontology analyses revealed that the DEPs in the PQ-2d and PQ-7d groups were involved primarily in humoral immunity and coagulation-related reactions, whereas those in the PQ-14d group were implicated primarily in chemotactic and regulatory responses. Kyoto Encyclopedia of Genes and Genomes analyses indicated that complement and coagulation cascades were key pathways in the PQ-2d and PQ-7d groups, whereas xenobiotic metabolism by cytochrome P450 was a key pathway in the PQ-14d group. Nine proteins at PQ-2d and eight proteins at PQ-7d were validated through parallel reaction monitoring (PRM). Conclusions PQ-induced ALI depends on over-activation of immune responses by damaged alveolar/endothelial cells, and the complement/coagulation cascade pathway plays a key role during this process. The proteins identified herein might provide new therapeutic targets or biomarkers for PQ poisoning.

Abstract Background Early diagnosis of cleft lip and palate (CLP) requires a multiplane examination, demanding high technical proficiency from radiologists. Therefore, this study aims to develop and validate the first artificial intelligence (AI)-based model (CLP-Net) for fully automated multi-plane localization in three-dimensional(3D) ultrasound during the first trimester. Methods This retrospective study included 418 (394 normal, 24 CLP) 3D ultrasound from 288 pregnant woman between July 2022 to October 2024 from Shenzhen Guangming District People’s Hospital during the 11–13+ 6 weeks of pregnancy. 320 normal volumes were used for training and validation, while 74 normal and 24 CLP volumes were used for testing. Two experienced radiologists reviewed three standard lip and palate planes (mid sagittal, retronasal triangle, and maxillary axial planes) as ground truth (GT) and the CLP-Net was developed to locate these planes. Results In normal test set, mean angle(± SD)° and distance(± SD)mm differences were 6.24 ± 4.83, 9.81 ± 5.48, 15.36 ± 18.14 and 0.86 ± 0.72, 1.36 ± 1.15, 1.96 ± 2.35 for MSP ± SD, RTP ± SD and MAP ± SD, NCC and SSIM were 0.931 ± 0.079, 0.819 ± 0.122, 0.781 ± 0.157 and 0.896 ± 0.058, 0.785 ± 0.076, 0.726 ± 0.088 respectively. In the CLP cases, there were 8.61 ± 5.52, 10.67 ± 5.08, 16.91 ± 17.42 and 1.03 ± 1.20, 1.17 ± 1.08, 1.34 ± 0.95 for mean angle and distance in MSP, RTP, and MAP, respectively. NCC and SSIM were 0.876 ± 0.104, 0.803 ± 0.084, 0.793 ± 0.089 and 0.841 ± 0.105, 0.812 ± 0.085, 0.764 ± 0.100, respectively. CLP-Net predictions had a highly visual acceptance rate among radiologists (MSP: 95%, RTP: 70%, MAP: 70%), with improved localization speed 15s(31.3%) for senior radiologists and 63s(38.9%) for junior radiologists. Conclusions CLP-Net accurately locates three planes for CLP screening, aiding radiologists and enhancing the efficiency of ultrasound examinations.

The primary function of terminators is to terminate transcription in gene expression. Although some studies have suggested that terminators also contribute positively to upstream gene expression, the extent and underlying mechanism of this effect remain largely unexplored. Here, the correlation between terminating strength and upstream mRNA stability was investigated by constructing a terminator mutation library through randomizing 5 nucleotides, assisted by FlowSeq technology, terminator variants were categorized based on the downstream fluorescence intensity, followed by high-throughput sequencing. To examine the impact of terminators on mRNA stability, the abundance of downstream gene transcripts for each terminator variant was quantified through cDNA sequencing. The results revealed that the transcript abundance controlled by strong terminators was, on average 2.2 times greater than those controlled by weak terminators on average. Moreover, several distinct features could be ascribed to high relative abundance of upstream gene transcript, including a high GC content at the base region of hairpin, and a high AT content in downstream of the U-tract. Additionally, these terminators showed a free energy between −28 and −22 kcal/mol, and a stem length of 14 nt. Finally, these features ascribed the upstream beneficial terminator were validated across various expression systems. By incorporating the optimal terminator downstream of RSF, GSH and HIS in three different strains, the fermentation productions-NMN SAM and VD13 exhibited a remarkable enhancement of 30 %–70 %. The findings presented here uncovered the terminator characteristics contributed to the upstream mRNA stability, providing guiding principles for gene circuit design.

Cortinarius species are widely distributed in the world and constitute a diverse group of ectomycorrhizal fungi that are associated with a broad range of plant hosts. The species diversity of this genus has been investigated in China, but it remains poorly understood, especially in the subtropical region. In this study, one new section, six novel species, one new combination, one new name, and one new record of Cortinarius are reported from subtropical China based on the morphological and molecular evidences. Cortinarius lilacinicarpus, C. sinocalaisopus, C. sinosalor, C. vividus spp. nov., C. orientisalor nom. nov., and C. illibatus are typical species of the section Delibuti. Cortinarius macroflavicapus sp. nov. belongs to Macroflavicapi sect. nov., a new section in the subgenus Dermocybe. Cortinarius tianbaoyanensis sp. nov. and C. dryadicola comb. nov. belong to the section Spilomei. Detailed descriptions, basidiocarps, and microscopic photographs are provided for these species. In addition, C. leptospermorum comb. nov. in the section Pauperae and subgenus Dermocybe is proposed.

ABSTRACT A scraper conveyor is one of the important equipment to ensure reliable, efficient and stable mining and transportation of coal. As the most important transmission system of the scraper conveyor, the gearbox takes the role of transmitting power and torque. Due to the influence of working conditions in underground coal mines, the gear transmission system is often subject to the impact of nonuniform large loads, which is very prone to failures, and affected by environmental interference, it is difficult to detect the early abnormal signals of the scraper conveyor gearbox in the conventional industrial scenarios of fault monitoring methods. To ensure the stability and reliability of its work, this paper carries out the research on the multi‐parameter fusion of gearbox early fault diagnosis method under strong background noise interference. Aiming at the problem that the change of fluid physical and chemical characteristic parameters can reflect the early health condition of the gear transmission system and the single vibration signal is difficult to be extracted under the strong background noise, a model based on the fluid physical and chemical characteristic parameters and vibration signals is constructed by utilizing the RBF neural network and the Random Forest algorithm, and the body of evidence of the two models is fused at the decision‐making level through the DS evidence theory, which forms the fluid‐vibration multi‐parameter fusion judgment of the early fault diagnosis method of scraper conveyor gearbox. Through comparison, it is found that compared with the fusion methods, such as high‐dimensional variational self‐encoder, and single diagnosis methods, such as the Random Forest Algorithm, the method researched in this paper is more suitable for the early fault warning of the scraper conveyor gearbox of the well coal mine, and the experimental validation finds that the average accuracy rate of the early fault recognition can be up to 96.6%.