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Journal of Neurovirology Jun 2024The Rabies virus is a neurotropic virus that manipulates the natural cell death processes of its host to ensure its own survival and replication. Studies have shown that...
The Rabies virus is a neurotropic virus that manipulates the natural cell death processes of its host to ensure its own survival and replication. Studies have shown that the anti-apoptotic effect of the virus is mediated by one of its protein named, rabies glycoprotein (RVG). Alzheimer's disease (AD) is characterized by the loss of neural cells and memory impairment. We aim to examine whether expression of RVG in the hippocampal cells can shield the detrimental effects induced by Aβ. Oligomeric form of Aβ (oAβ) or vehicle was bilaterally microinjected into the dorsal hippocampus of male Wistar rats. One week later, two μl (10 T.U. /ml) of the lentiviral vector carrying RVG gene was injected into their dorsal hippocampus (post-treatment). In another experiment, the lentiviral vector was microinjected one week before Aβ injection (pre-treatment). One week later, the rat's brain was sliced into cross-sections, and the presence of RVG-expressing neuronal cells was confirmed using fluorescent microscopy. Rats were subjected to assessments of spatial learning and memory as well as passive avoidance using the Morris water maze (MWM) and the Shuttle box apparatuses, respectively. Protein expression of AMPA receptor subunit (GluA1) was determined using western blotting technique. In MWM, Aβ treated rats showed decelerated acquisition of the task and impairment of reference memory. RVG expression in the hippocampus prevented and restored the deficits in both pre- and post- treatment conditions, respectively. It also improved inhibitory memory in the oAβ treated rats. RVG increased the expression level of GluA1 level in the hippocampus. Based on our findings, the expression of RVG in the hippocampus has the potential to enhance both inhibitory and spatial learning abilities, ultimately improving memory performance in an AD rat model. This beneficial effect is likely attributed, at least in part, to the increased expression of GluA1-containing AMPA receptors.
PubMed: 38943023
DOI: 10.1007/s13365-024-01221-y -
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 -
Free Radical Biology & Medicine Jun 2024Disturbance in iron homeostasis has been described in Parkinson's disease (PD), in which iron regulatory protein 2 (IRP2) plays a crucial role. IRP2 deletion resulted in...
Disturbance in iron homeostasis has been described in Parkinson's disease (PD), in which iron regulatory protein 2 (IRP2) plays a crucial role. IRP2 deletion resulted in the misregulation of iron metabolism and subsequent neurodegeneration. However, growing evidence showed that the levels of IRP2 were increased in the substantial nigra (SN) in MPTP-induced PD mice. To further clarify the role of increased IRP2 in PD, we developed IRP2-overexpressed mice by microinjecting AAV-Ireb2 in the SN. These mice showed decreased motor ability, abnormal gait and anxiety. Iron deposits induced by increased TFR1 and dopaminergic neuronal loss were observed in the SN. When these mice were treated with MPTP, exacerbated dyskinesia and dopaminergic neuronal loss were observed. In addition, TP53 was post-transcriptionally upregulated by IRP2 binding to the iron regulated element (IRE) in its 3' untranslated region. This resulted in increased lipid peroxidation levels and induced ferroptosis through the SLC7A11-ALOX12 pathway, which was independent of GPX4. This study revealed that IRP2 homeostasis in the SN was critical for PD progression and clarified the molecular mechanism of ferroptosis caused by IRP2.
PubMed: 38936518
DOI: 10.1016/j.freeradbiomed.2024.06.020 -
Reproductive Biology Jun 2024In mammals, early embryogenesis relies heavily on the regulation of maternal transcripts including protein-coding and non-coding RNAs stored in oocytes. In this study,...
In mammals, early embryogenesis relies heavily on the regulation of maternal transcripts including protein-coding and non-coding RNAs stored in oocytes. In this study, the expression of three bovine oocyte expressed long non-coding RNAs (lncRNAs), OOSNCR1, OOSNCR2, and OOSNCR3, was characterized in somatic tissues, the ovarian follicle, and throughout early embryonic development. Moreover, the functional requirement of each transcript during oocyte maturation and early embryonic development was investigated using a siRNA-mediated knockdown approach. Tissue distribution analysis revealed that OOSNCR1, OOSNCR2 and OOSNCR3 are predominantly expressed in fetal ovaries. Follicular cell expression analysis revealed that these lncRNAs are highly expressed in the oocytes, with minor expression detected in the cumulus cells (CCs) and mural granulosa cells (mGCs). The expression for all three genes was highest during oocyte maturation, decreased at fertilization, and ceased altogether by the 16-cell stage. Knockdown of OOSNCR1, OOSNCR2 and OOSNCR3 in immature oocytes was achieved by microinjection of the cumulus-enclosed germinal vesicle (GV) oocytes with siRNAs targeting these lncRNAs. Knockdown of OOSNCR1, OOSNCR2 and OOSNCR3 did not affect cumulus expansion, but oocyte survival at 12 h post-insemination was significantly reduced. In addition, knockdown of OOSNCR1, OOSNCR2 and OOSNCR3 in immature oocytes resulted in a decreased rate of blastocyst development, and reduced expression of genes associated with oocyte competency such as nucleoplasmin 2 (NPM2), growth differentiation factor 9 (GDF9), bone morphogenetic protein 15 (BMP15), and JY-1 in MII oocytes. The data herein suggest a functional requirement of OOSNCR1, OOSNCR2, and OOSNCR3 during bovine oocyte maturation and early embryogenesis.
PubMed: 38936296
DOI: 10.1016/j.repbio.2024.100915 -
International Journal of Molecular... Jun 2024A large diversity of epigenetic factors, such as microRNAs and histones modifications, are known to be capable of regulating gene expression without altering DNA...
A large diversity of epigenetic factors, such as microRNAs and histones modifications, are known to be capable of regulating gene expression without altering DNA sequence itself. In particular, miR-1 is considered the first essential microRNA in cardiac development. In this study, miR-1 potential role in early cardiac chamber differentiation was analyzed through specific signaling pathways. For this, we performed in chick embryos functional experiments by means of miR-1 microinjections into the posterior cardiac precursors-of both primitive endocardial tubes-committed to sinoatrial region fates. Subsequently, embryos were subjected to whole mount in situ hybridization, immunohistochemistry and RT-qPCR analysis. As a relevant novelty, our results revealed that miR-1 increased , and , while this microRNA diminished and expressions during early differentiation of the cardiac sinoatrial region. Furthermore, we observed in this developmental context that miR-1 upregulated and and downregulated , which are three crucial factors in the retinoic acid signaling pathway. Interestingly, we also noticed that miR-1 directly interacted with and /, as well as with /, which are three key factors actively involved in regulation. Our study shows, for the first time, a key role of miR-1 as an epigenetic regulator in the early differentiation of the cardiac sinoatrial region through orchestrating opposite actions between retinoic acid and , fundamental to properly assign cardiac cells to their respective heart chambers. A better understanding of those molecular mechanisms modulated by miR-1 will definitely help in fields applied to therapy and cardiac regeneration and repair.
Topics: Animals; MicroRNAs; Epigenesis, Genetic; Cell Differentiation; Gene Expression Regulation, Developmental; Chick Embryo; MEF2 Transcription Factors; Sinoatrial Node; Signal Transduction; Heart
PubMed: 38928314
DOI: 10.3390/ijms25126608 -
International Journal of Molecular... Jun 2024Aneurysms pose life-threatening risks due to the dilatation of the arteries and carry a high risk of rupture. Despite continuous research efforts, there are still no...
Aneurysms pose life-threatening risks due to the dilatation of the arteries and carry a high risk of rupture. Despite continuous research efforts, there are still no satisfactory or clinically effective pharmaceutical treatments for this condition. Accelerated inflammatory processes during aneurysm development lead to increased levels of matrix metalloproteinases (MMPs) and destabilization of the vessel wall through the degradation of the structural components of the extracellular matrix (ECM), mainly collagen and elastin. Tissue inhibitors of metalloproteinases (TIMPs) directly regulate MMP activity and consequently inhibit ECM proteolysis. In this work, the synthesis of TIMP-1 protein was increased by the exogenous delivery of synthetic TIMP-1 encoding mRNA into aortic vessel tissue in an attempt to inhibit MMP-9. In vitro, TIMP-1 mRNA transfection resulted in significantly increased TIMP-1 protein expression in various cells. The functionality of the expressed protein was evaluated in an appropriate ex vivo aortic vessel model. Decreased MMP-9 activity was detected using in situ zymography 24 h and 48 h post microinjection of 5 µg TIMP-1 mRNA into the aortic vessel wall. These results suggest that TIMP-1 mRNA administration is a promising approach for the treatment of aneurysms.
Topics: Tissue Inhibitor of Metalloproteinase-1; Matrix Metalloproteinase 9; RNA, Messenger; Animals; Humans; Rats; Aneurysm; Aorta; Male; Arteries; Matrix Metalloproteinase Inhibitors
PubMed: 38928311
DOI: 10.3390/ijms25126599 -
Environmental Toxicology and Chemistry Jun 2024Benzotriazole ultraviolet stabilizers (BUVSs) are a group of widely used chemicals added to a variety of consumer (e.g., plastics) and industrial (e.g., metal coating)...
Benzotriazole ultraviolet stabilizers (BUVSs) are a group of widely used chemicals added to a variety of consumer (e.g., plastics) and industrial (e.g., metal coating) goods. Although detected globally as an environmentally persistent pollutant, BUVSs have received relatively little toxicological attention and only recently have been acknowledged to affect development and the endocrine system in vivo. In our previous study, altered behavior, indicative of potential neurotoxicity, was observed among rainbow trout alevins (day 14 posthatching) that were microinjected as embryos with a single environmentally relevant dose of 2,4-di-tert-butyl-6-(5-chloro-2H-benzotriazol-2-yl) phenol (UV-327). In the present follow-up study, we performed whole-transcriptome profiling (RNA sequencing) of newly hatched alevins from the same batch. The primary aim was to identify biomarkers related to behavior and neurology. Dose-specifically, 1 to 176 differentially expressed genes (DEGs) were identified. In the group presenting altered behavior (273.4 ng g), 176 DEGs were identified, yet only a fraction was related to neurological functions, including water, calcium, and potassium homeostasis; acetylcholine transmission and signaling; as well insulin and energy metabolism. The second objective was to estimate the transcriptomic point of departure (tPOD) and assess if point estimate(s) are protective of altered behavior. A tPOD was established at 35 to 94 ng UV-327 g egg, making this tPOD protective of behavioral alterations. Holistically, these transcriptomic alterations provide a foundation for future research on how BUVSs can influence rainbow trout alevin development, while providing support to the hypothesis that UV-327 can influence neurogenesis and subsequent behavioral endpoints. The exact structural and functional changes caused by embryonic exposure to UV-327 remain enigmatic and will require extensive investigation before being deciphered and understood toxicologically. Environ Toxicol Chem 2024;00:1-12. © 2024 The Author(s). Environmental Toxicology and Chemistry published by Wiley Periodicals LLC on behalf of SETAC.
PubMed: 38923588
DOI: 10.1002/etc.5943 -
Insects Jun 2024iGABAR, a member of the Cys-loop ligand-gated ion channel superfamily, is a significant target of the insecticide ivermectin (IVM). GRD is the potential subunit of the...
iGABAR, a member of the Cys-loop ligand-gated ion channel superfamily, is a significant target of the insecticide ivermectin (IVM). GRD is the potential subunit of the insect iGABAR. However, little information about GRD in has been reported. In this study, we involved cloning and characterizing the iGABAR subunit of (). Sequence analysis indicated that , as part of the cysteine-loop ligand-gated ion channel family, is similar to other insect . RNA interference (RNAi) was employed to explore IVM resistance in , resulting in a significant reduction in expression ( < 0.05), and the mortality of adults with knockdown was significantly decreased after exposure to ivermectin. Bioinformatics prediction identified miR-71-5p as a potential regulator of . In vitro, dual-luciferase reporter assays confirmed that expression was regulated by miR-71-5p. Microinjection of miR-71-5p mimics upregulated miR-71-5p expression and downregulated gene expression, reducing mortality by 34.52% following IVM treatment. Conversely, microinjection of a miR-71-5p inhibitor decreased miR-71-5p expression but did not affect the susceptibility to IVM despite increased expression ( < 0.05). In conclusion, , as one of the iGABA receptor subunits, is a potential target of ivermectin. It may influence ivermectin resistance by modulating the GABA signaling pathway. The inhibition of expression by miR-71-5p decreased ivermectin resistance and consequently lowered the mortality rate of mosquitoes. This finding provides empirical evidence of the relationship between and its miRNA in modulating insecticide resistance, offering novel perspectives for mosquito control strategies.
PubMed: 38921167
DOI: 10.3390/insects15060453 -
ACS Nano Jun 2024There is growing concern about the distribution of nanoplastics (NPs) in the central nervous system (CNS), whereas intrusion is poorly understood. In this study,...
There is growing concern about the distribution of nanoplastics (NPs) in the central nervous system (CNS), whereas intrusion is poorly understood. In this study, fluorescent-labeled polystyrene NPs (PS-NPs) were microinjected into different areas of zebrafish embryo to mimic different routes of exposure. PS-NPs were observed in the brain, eyes, and spinal cord through gametal exposure. It indicated that maternally derived PS-NPs were specially distributed in the CNS of zebrafish during early development. Importantly, these NPs were stranded in the CNS but not transferred to other organs during development. Furthermore, using neuron GFP-labeled transgenic zebrafish, colocalization between NPs and the neuron cells revealed that NPs were mostly enriched in the CNS surrounded but not the neurons. Even so, the intrusion of NPs into the CNS induced the significant upregulation of some neurotransmitter receptors, leading to an inhibited effect on the movement of zebrafish larvae. This work provides insights into understanding the intrusion and distribution of NPs in the CNS and the subsequent potential adverse effects.
PubMed: 38918939
DOI: 10.1021/acsnano.4c00625 -
Nature Communications Jun 2024During human embryonic development, early cleavage-stage embryos are more susceptible to errors. Studies have shown that many problems occur during the first mitosis,...
During human embryonic development, early cleavage-stage embryos are more susceptible to errors. Studies have shown that many problems occur during the first mitosis, such as direct cleavage, chromosome segregation errors, and multinucleation. However, the mechanisms whereby these errors occur during the first mitosis in human embryos remain unknown. To clarify this aspect, in the present study, we image discarded living human two-pronuclear stage zygotes using fluorescent labeling and confocal microscopy without microinjection of DNA or mRNA and investigate the association between spindle shape and nuclear abnormality during the first mitosis. We observe that the first mitotic spindles vary, and low-aspect-ratio-shaped spindles tend to lead to the formation of multiple nuclei at the 2-cell stage. Moreover, we observe defocusing poles in many of the first mitotic spindles, which are strongly associated with multinucleation. Additionally, we show that differences in the positions of the centrosomes cause spindle abnormality in the first mitosis. Furthermore, many multinuclei are modified to form mononuclei after the second mitosis because the occurrence of pole defocusing is firmly reduced. Our study will contribute markedly to research on the occurrence of mitotic errors during the early cleavage of human embryos.
Topics: Humans; Spindle Apparatus; Mitosis; Cell Nucleus; Zygote; Embryo, Mammalian; Microscopy, Confocal; Centrosome; Embryonic Development; Female
PubMed: 38918406
DOI: 10.1038/s41467-024-49815-8