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Journal of Ethnopharmacology Jun 2024The traditional medicinal formulation, Qifu-yin (QFY), has been widely prescribed for Alzheimer's disease (AD) treatment in China, yet the comprehensive mechanisms...
ETHNOPHARMACOLOGICAL RELEVANCE
The traditional medicinal formulation, Qifu-yin (QFY), has been widely prescribed for Alzheimer's disease (AD) treatment in China, yet the comprehensive mechanisms through which QFY mitigates AD pathology remain to be fully delineated.
AIM OF THE STUDY
This study aimed to explore the therapeutic implications of QFY on the synaptic injury and oxidative stress in the hippocampus of APPswe/PS1dE9 (APP/PS1) mice, with a concerted effort to elucidate the molecular mechanisms related to synaptic preservation and memory improvement.
MATERIALS AND METHODS
The components of QFY were identified by ultra-high-performance liquid chromatography-tandem mass spectrometry (UHPLC-MS/MS). The neuroprotective effects of QFY was evaluated using six-month-old male APP/PS1 mice. Subsequent to a 15 days of QFY regimen, spatial memory was assessed utilizing the Morris water maze (MWM) test. Amyloid-beta (Aβ) aggregation was detected via immunostaining, while the quantification of Aβ and Aβ was achieved through enzyme-linked immunosorbent assay (ELISA). Transmission electron microscopy (TEM) was used to investigate the synaptic structure and mitochondrial morphology. Golgi staining was applied to examine dendritic spine density. Reactive oxygen species (ROS), 3-nitrotyrosine (3-NT) and 4-hydroxy-nonenal (4-HNE) assays were employed to assess oxidative stress. The expression profiles of Aβ metabolism-associated enzymes and the Keap1/Nrf2/ARE signaling pathway were determined by Western blot.
RESULTS
A total of 20 principal compounds in QFY were identified. QFY mitigated memory deficits of APP/PS1 mice, including reducing escape latency and search distance and increasing the time and distance spent in the target quadrant. In addition, QFY increased platform crossings of APP/PS1 mice in the probe trial of MWM tests. TEM analysis showed that QFY increased synapse number in the CA1 region of APP/PS1 mice. Further studies indicated that QFY elevated the expression levels of Post synaptic density protein 95 (PSD95) and synaptophysin, and mitigated the loss of dendritic spine density in the hippocampus of APP/PS1 mice. QFY has been shown to ameliorated the structural abnormalities of mitochondria, including mitochondrial dissolution and degradation, up-regulate ATP synthesis and membrane potential in the hippocampus of APP/PS1 mice. Moreover, QFY activated the Keap1/Nrf2/ARE signaling pathway in the hippocampus of APP/PS1 mice, which might contribute to the neuroprotective effects of QFY.
CONCLUSION
QFY activates the Keap1/Nrf2/ARE signaling, and protects against synaptic and mitochondrial dysfunction in APP/PS1 mice, proposing a potential alternative therapeutic strategy for AD management.
PubMed: 38942156
DOI: 10.1016/j.jep.2024.118497 -
Molecular and Cellular Probes Jun 2024Psoriasis is a chronic inflammatory disease characterized by increased keratinocyte proliferation and local inflammation. Long noncoding RNAs (lncRNAs) play important...
Psoriasis is a chronic inflammatory disease characterized by increased keratinocyte proliferation and local inflammation. Long noncoding RNAs (lncRNAs) play important regulatory roles in many immune-mediated diseases, including psoriasis. In this study, we aimed to investigate the role and mechanism of lnc-SPRR2G-2 (SPRR2G) in M5-treated psoriatic keratinocytes. Fluorescence in situ hybridization and quantitative real-time polymerase chain reaction (qRT-PCR) showed that lnc-SPRR2G-2 was significantly upregulated in psoriasis tissues and psoriatic keratinocytes. In psoriatic keratinocytes, functional and molecular experiment analyses demonstrated that SPRR2G regulated proliferation, cell cycle and apoptosis, and induced the expression of S100 calcium binding protein A7 (S100A7), interleukin (IL)-1β, IL-8 and C-X-C motif chemokine ligand 10 (CXCL10). The function of SPRR2G in psoriasis is related to the STAT3 signaling pathway and can be inhibited by a STAT3 inhibitor. Moreover, KH-type splicing regulatory protein (KHSRP) was proved to be regulated by lnc-SPRR2G-2 and to control the mRNA decay of psoriasis-related cytokines (p<0.05). In summary, we reported the functions of lnc-SPRR2G-2 and KHSRP in psoriasis. Our findings provide new insights for the further exploration of the pathogenesis and treatment of psoriasis.
PubMed: 38942130
DOI: 10.1016/j.mcp.2024.101967 -
Nature Protocols Jun 2024Spatial epigenetic mapping of tissues enables the study of gene regulation programs and cellular functions with the dependency on their local tissue environment. Here we... (Review)
Review
Spatial epigenetic mapping of tissues enables the study of gene regulation programs and cellular functions with the dependency on their local tissue environment. Here we outline a complete procedure for two spatial epigenomic profiling methods: spatially resolved genome-wide profiling of histone modifications using in situ cleavage under targets and tagmentation (CUT&Tag) chemistry (spatial-CUT&Tag) and transposase-accessible chromatin sequencing (spatial-ATAC-sequencing) for chromatin accessibility. Both assays utilize in-tissue Tn5 transposition to recognize genomic DNA loci followed by microfluidic deterministic barcoding to incorporate spatial address codes. Furthermore, these two methods do not necessitate prior knowledge of the transcription or epigenetic markers for a given tissue or cell type but permit genome-wide unbiased profiling pixel-by-pixel at the 10 μm pixel size level and single-base resolution. To support the widespread adaptation of these methods, details are provided in five general steps: (1) sample preparation; (2) Tn5 transposition in spatial-ATAC-sequencing or antibody-controlled pA-Tn5 tagmentation in CUT&Tag; (3) library preparation; (4) next-generation sequencing; and (5) data analysis using our customed pipelines available at: https://github.com/dyxmvp/Spatial_ATAC-seq and https://github.com/dyxmvp/spatial-CUT-Tag . The whole procedure can be completed on four samples in 2-3 days. Familiarity with basic molecular biology and bioinformatics skills with access to a high-performance computing environment are required. A rudimentary understanding of pathology and specimen sectioning, as well as deterministic barcoding in tissue-specific skills (e.g., design of a multiparameter barcode panel and creation of microfluidic devices), are also advantageous. In this protocol, we mainly focus on spatial profiling of tissue region-specific epigenetic landscapes in mouse embryos and mouse brains using spatial-ATAC-sequencing and spatial-CUT&Tag, but these methods can be used for other species with no need for species-specific probe design.
PubMed: 38943021
DOI: 10.1038/s41596-024-01013-y -
The Journal of Physical Chemistry. B Jun 2024The ability of small lipophilic molecules to penetrate the blood-brain barrier through transmembrane diffusion has enabled researchers to explore new diagnostics and...
The ability of small lipophilic molecules to penetrate the blood-brain barrier through transmembrane diffusion has enabled researchers to explore new diagnostics and therapies for brain disorders. Until now, therapies targeting the brain have mainly relied on biochemical mechanisms, while electrical treatments such as deep brain stimulation often require invasive procedures. An alternative to implanting deep brain stimulation probes could involve administering small molecule precursors intravenously, capable of crossing the blood-brain barrier, and initiating the formation of conductive polymer networks in the brain through polymerization. This study examines the aggregation behavior of five water-soluble conducting polymer precursors sharing the same conjugate core but differing in side chains, using spectroscopy and various computational chemistry tools. Our findings highlight the significant impact of side chain composition on both aggregation and spectroscopic response.
PubMed: 38942741
DOI: 10.1021/acs.jpcb.4c02891 -
Medicine Jun 2024Hereditary transthyretin-mediated amyloidosis (ATTRv amyloidosis), known as Corino de Andrade disease, is a rare neurodegenerative disorder with a significant global... (Review)
Review
Hereditary transthyretin-mediated amyloidosis (ATTRv amyloidosis), known as Corino de Andrade disease, is a rare neurodegenerative disorder with a significant global impact characterized by the misfolding of transthyretin (TTR) protein leading to amyloid aggregation, ATTRv amyloidosis, especially with polyneuropathy, poses a considerable challenge in managing its rapid progression and debilitating effects. This mini-review focuses on the recent advancements in the treatment landscape for ATTRv amyloidosis with polyneuropathy, specifically the RNA interference therapeutic Vutrisiran and the ligand-conjugated antisense oligonucleotide Eplontersen. We aim to provide a comprehensive overview of the mechanisms, current evidence from clinical trials, and future directions for these novel therapeutic agents. Vutrisiran and Eplontersen have demonstrated significant clinical efficacy in improving neuropathic impairment, quality of life, and serum TTR levels in various trials. The distinct mechanistic approaches of these therapies, coupled with their acceptable safety profiles, offer promising avenues for addressing the complexities of ATTRv amyloidosis with polyneuropathy. The introduction of Vutrisiran and Eplontersen marks a pivotal moment in the quest for effective therapies against ATTRv amyloidosis with polyneuropathy. While clinical evidence is promising, ongoing research is crucial to deepen mechanistic understanding and address research gaps. Future perspectives include the potential expansion of therapeutic options and a more inclusive approach to cater to the diverse needs of individuals globally. This mini-review provides valuable insights into the evolving landscape of ATTRv amyloidosis management and sets the stage for further exploration in this challenging domain.
Topics: Humans; Amyloid Neuropathies, Familial; Polyneuropathies; Oligonucleotides; Oligonucleotides, Antisense; Prealbumin; Quality of Life
PubMed: 38941378
DOI: 10.1097/MD.0000000000038767 -
ACS Sensors Jun 2024Fluorescence-based contrast agents enable real-time detection of solid tumors and their neovasculature, making them ideal for use in image-guided surgery. Several agents...
Fluorescence-based contrast agents enable real-time detection of solid tumors and their neovasculature, making them ideal for use in image-guided surgery. Several agents have entered late-stage clinical trials or secured FDA approval, suggesting they are likely to become the standard of care in cancer surgeries. One of the key parameters to optimize in contrast agents is molecular size, which dictates much of the pharmacokinetic and pharmacodynamic properties of the agent. Here, we describe the development of a class of protease-activated quenched fluorescent probes in which a -(2-hydroxypropyl)methacrylamide copolymer is used as the primary scaffold. This copolymer core provides a high degree of probe modularity to generate structures that cannot be achieved with small molecules and peptide probes. We used a previously validated cathepsin substrate and evaluated the effects of length and type of linker, as well as the positioning of the fluorophore/quencher pair on the polymer core. We found that the polymeric probes could be optimized to achieve increased overall signal and tumor-to-background ratios compared to the reference small molecule probe. Our results also revealed multiple structure-activity relationship trends that can be used to design and optimize future optical imaging probes. Furthermore, they confirm that a hydrophilic polymer is an ideal scaffold for use in optical imaging contrast probes, allowing a highly modular design that enables efficient optimization to maximize probe accumulation and overall biodistribution properties.
PubMed: 38941307
DOI: 10.1021/acssensors.4c00912 -
STAR Protocols Jun 2024A gene-rescue experiment under a mutant background is essential to clarify gene function and the resulting biological potential in vivo. Here, we present a protocol for...
A gene-rescue experiment under a mutant background is essential to clarify gene function and the resulting biological potential in vivo. Here, we present a protocol for determining the change in interferon response by microinjecting plasmids into one-cell-stage zebrafish embryos. We describe steps for comparing the resistance potential to virus infection in wild-type and knockout zebrafish larvae following plasmid microinjection. We then detail how to link the enhanced interferon immunity to the improved resistance in knockout zebrafish larvae by gene-rescue experiments. For complete details on the use and execution of this protocol, please refer to Qu et al..
PubMed: 38941183
DOI: 10.1016/j.xpro.2024.103156 -
MSphere Jun 2024The adaptation of gene deletion methods based on the CRISPR-Cas9 system has facilitated the genetic manipulation of the pathogenic yeast , because homozygous mutants of...
Probing gene function in wild-type strains by Cas9-facilitated one-step integration of two dominant selection markers: a systematic analysis of recombination events at the target locus.
UNLABELLED
The adaptation of gene deletion methods based on the CRISPR-Cas9 system has facilitated the genetic manipulation of the pathogenic yeast , because homozygous mutants of this diploid fungus can now be generated in a single step, allowing the rapid screening of candidate genes for their involvement in a phenotype of interest. However, the Cas9-mediated double-strand breaks at the target site may result in an undesired loss of heterozygosity (LOH) on the affected chromosome and cause phenotypic alterations that are not related to the function of the investigated gene. In our present study, we harnessed Cas9-facilitated gene deletion to probe a set of genes that are constitutively overexpressed in strains containing hyperactive forms of the transcription factor Mrr1 for a possible contribution to the fluconazole resistance of such strains. To this aim, we used gene deletion cassettes containing two different dominant selection markers, and , which confer resistance to nourseothricin and hygromycin, respectively, for simultaneous genomic integration in a single step, hypothesizing that this would minimize undesired LOH events at the target locus. We found that selection for resistance to both nourseothricin and hygromycin strongly increased the proportion of homozygous deletion mutants among the transformants compared with selection on media containing only one of the antibiotics, but it did not avoid undesired LOH events. Our results demonstrate that LOH on the target chromosome is a significant problem when using Cas9 for the generation of gene deletion mutants, which demands a thorough examination of recombination events at the target site.
IMPORTANCE
is one of the medically most important fungi and a model organism to study fungal pathogenicity. Investigating gene function in this diploid yeast has been facilitated by the adaptation of gene deletion methods based on the bacterial CRISPR-Cas9 system, because they enable the generation of homozygous mutants in a single step. We found that, in addition to increasing the efficiency of gene replacement by selection markers, the Cas9-mediated double-strand breaks also result in frequent loss of heterozygosity on the same chromosome, even when two different selection markers were independently integrated into the two alleles of the target gene. Since loss of heterozygosity for other genes can result in phenotypic alterations that are not caused by the absence of the target gene, these findings show that it is important to thoroughly analyze recombination events at the target locus when using Cas9 to generate gene deletion mutants in .
PubMed: 38940507
DOI: 10.1128/msphere.00388-24 -
International Journal of Molecular... Aug 2024Naringenin (NAR) is a prominent flavanone that has been recognized for its capacity to promote the osteogenic differentiation of human periodontal ligament stem cells...
Naringenin (NAR) is a prominent flavanone that has been recognized for its capacity to promote the osteogenic differentiation of human periodontal ligament stem cells (hPDLSCs). The present study aimed to explore how NAR promotes the osteogenic differentiation of hPDLSCs and to assess its efficacy in repairing alveolar bone defects. For this purpose, a protein‑protein interaction network of NAR action was established by mRNA sequencing and network pharmacological analysis. Gene and protein expression levels were evaluated by reverse transcription‑quantitative and western blotting. Alizarin red and alkaline phosphatase staining were also employed to observe the osteogenic capacity of hPDLSCs, and immunofluorescence was used to examine the co‑localization of NAR molecular probes and AKT in cells. The repair of mandibular defects was assessed by micro‑computed tomography (micro‑CT), Masson staining and immunofluorescence. Additionally, computer simulation docking software was utilized to determine the binding affinity of NAR to the target protein, AKT. The results demonstrated that activation of the nitric oxide (NO)‑cyclic guanosine monophosphate (cGMP)‑protein kinase G (PKG) signaling pathway could promote the osteogenic differentiation of hPDLSCs. Inhibition of AKT, endothelial nitric oxide synthase and soluble guanylate cyclase individually attenuated the ability of NAR to promote the osteogenic differentiation of hPDLSCs. Micro‑CT and Masson staining revealed that the NAR gavage group exhibited more new bone formation at the defect site. Immunofluorescence assays confirmed the upregulated expression of Runt‑related transcription factor 2 and osteopontin in the NAR gavage group. In conclusion, the results of the present study suggested that NAR promotes the osteogenic differentiation of hPDLSCs by activating the NO‑cGMP‑PKG signaling pathway through its binding to AKT.
Topics: Humans; Osteogenesis; Flavanones; Proto-Oncogene Proteins c-akt; Signal Transduction; Cell Differentiation; Nitric Oxide; Cyclic GMP-Dependent Protein Kinases; Stem Cells; Cyclic GMP; Animals; Male; Cells, Cultured
PubMed: 38940332
DOI: 10.3892/ijmm.2024.5391 -
Chemical & Biomedical Imaging Jun 2024Epicuticular wax is the outmost layer of plant leaves that protects them from desiccation and penetration of harmful reagents. There is an intense industrial effort in...
Epicuticular wax is the outmost layer of plant leaves that protects them from desiccation and penetration of harmful reagents. There is an intense industrial effort in the development of softening agents, adjuvants, that can adjust the permeability of the wax toward pesticides and, thus, play an important role in sustainable agriculture. However, mechanistic understanding of the structure and dynamic properties within the plant wax, particularly upon the application of adjuvants, is currently lacking. In this work, we demonstrate that fluorescence lifetime imaging microscopy (FLIM) combined with molecular rotors, fluorescent probes sensitive to viscosity, can directly probe the microviscosity of amorphous and crystalline phases of model plant wax layers. Moreover, this approach is able to quantify the changes in viscosity in both phases upon the addition of water and adjuvant solutions on top of the wax. We show that water permeation mostly perturbs the crystalline phase of the wax, while our chosen adjuvant, Plurafac LF431, mainly softens the amorphous phase of the wax. Our technique provides a facile and quantitative way to monitor dynamic properties within plant waxes with diffraction-limited resolution and reveals the effect of organic substances on wax structure and rigidity, crucial for designing next-generation agents to improve agricultural efficiency.
PubMed: 38939873
DOI: 10.1021/cbmi.4c00005