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Biomedicine & Pharmacotherapy =... Jun 2024Ketamine, as a non-competitive antagonist of N-methyl-D-aspartate (NMDA) receptors, was originally used in general anesthesia. Epidemiological data show that ketamine...
BACKGROUND
Ketamine, as a non-competitive antagonist of N-methyl-D-aspartate (NMDA) receptors, was originally used in general anesthesia. Epidemiological data show that ketamine has become one of the most commonly abused drugs in China. Ketamine administration might cause cognitive impairment; however, its molecular mechanism remains unclear. The glymphatic system is a lymphoid system that plays a key role in metabolic waste removal and cognitive regulation in the central nervous system.
METHODS
Focusing on the glymphatic system, this study evaluated the behavioral performance and circulatory function of the glymphatic system by building a short-term ketamine administration model in mice, and detected the expression levels of the 5-HT2c receptor, ΔFosb, Pten, Akt, and Aqp4 in the hippocampus. Primary astrocytes were cultured to verify the regulatory relationships among related indexes using a 5-HT2c receptor antagonist, a 5-HT2c receptor short interfering RNA (siRNA), and a ΔFosb siRNA.
RESULTS
Ketamine administration induced ΔFosb accumulation by increasing 5-HT2c receptor expression in mouse hippocampal astrocytes and primary astrocytes. ΔFosb acted as a transcription factor to recognize the AATGATTAAT bases in the 5' regulatory region of the Aqp4 gene (-1096 bp to -1087 bp), which inhibited Aqp4 expression, thus causing the circulatory dysfunction of the glymphatic system, leading to cognitive impairment.
CONCLUSIONS
Although this regulatory mechanism does not involve the Pten/Akt pathway, this study revealed a new mechanism of ketamine-induced cognitive impairment in non-neuronal systems, and provided a theoretical basis for the safety of clinical treatment and the effectiveness of withdrawal.
Topics: Animals; Ketamine; Astrocytes; Cognitive Dysfunction; Mice; Male; Hippocampus; Glymphatic System; Proto-Oncogene Proteins c-akt; Aquaporin 4; Receptor, Serotonin, 5-HT2C; Mice, Inbred C57BL; Cells, Cultured; Proto-Oncogene Proteins c-fos; PTEN Phosphohydrolase
PubMed: 38759288
DOI: 10.1016/j.biopha.2024.116739 -
Science Advances May 2024Understanding how the amino acid sequence dictates protein structure and defines its stability is a fundamental problem in molecular biology. It is especially...
Understanding how the amino acid sequence dictates protein structure and defines its stability is a fundamental problem in molecular biology. It is especially challenging for membrane proteins that reside in the complex environment of a lipid bilayer. Here, we obtain an atomic-level picture of the thermally induced unfolding of a membrane-embedded α-helical protein, human aquaporin 1, using solid-state nuclear magnetic resonance spectroscopy. Our data reveal the hierarchical two-step pathway that begins with unfolding of a structured extracellular loop and proceeds to an intermediate state with a native-like helical packing. In the second step, the transmembrane domain unravels as a single unit, resulting in a heterogeneous misfolded state with high helical content but with nonnative helical packing. Our results show the importance of loops for the kinetic stabilization of the whole membrane protein structure and support the three-stage membrane protein folding model.
Topics: Protein Unfolding; Humans; Membrane Proteins; Aquaporin 1; Nuclear Magnetic Resonance, Biomolecular; Magnetic Resonance Spectroscopy; Models, Molecular; Protein Folding; Kinetics; Thermodynamics
PubMed: 38758787
DOI: 10.1126/sciadv.adm7907 -
Experimental Biology and Medicine... 2024Diabetes mellitus is a prevalent disease that is often accompanied by ocular surface abnormalities including delayed epithelial wound healing and decreased corneal...
Diabetes mellitus is a prevalent disease that is often accompanied by ocular surface abnormalities including delayed epithelial wound healing and decreased corneal sensitivity. The impact of diabetes on the lacrimal functional unit (LFU) and the structures responsible for maintaining tear homeostasis, is not completely known. It has been shown that the Opioid Growth Factor Receptor (OGFr), and its ligand, Opioid Growth Factor (OGF), is dysregulated in the ocular surface of diabetic rats leading to overproduction of the inhibitory growth peptide OGF. The opioid antagonist naltrexone hydrochloride (NTX) blocks the OGF-OGFr pathway, and complete blockade following systemic or topical treatment with NTX restores the rate of re-epithelialization of corneal epithelial wounds, normalizes corneal sensitivity, and reverses dry eye in diabetic animal models. These effects occur rapidly and within days of initiating treatment. The present study was designed to understand mechanisms related to the fast reversal (<5 days) of dry eye by NTX in type 1 diabetes (T1D) by investigating dysregulation of the LFU. The approach involved examination of the morphology of the LFU before and after NTX treatment. Male and female adult Sprague-Dawley rats were rendered hyperglycemic with streptozotocin, and after 6 weeks rats were considered to be a T1D model. Rats received topical NTX twice daily to one eye for 10 days. During the period of treatment, tear production and corneal sensitivity were recorded. On day 11, animals were euthanized and orbital tissues including conjunctiva, eyelids, and lacrimal glands, were removed and processed for histologic examination including immunohistochemistry. Male and female T1D rats had significantly decreased tear production and corneal insensitivity, significantly decreased number and size of lacrimal gland acini, decreased expression of aquaporin-5 (AQP5) protein and decreased goblet cell size. Thus, 10 days of NTX treatment restored tear production and corneal sensitivity to normal values, increased AQP5 expression, and restored the surface area of goblet cells to normal. NTX had no effect on the number of lacrimal gland acini or the number of conjunctival goblet cells. In summary, blockade of the OGF-OGFr pathway with NTX reversed corneal and lacrimal gland complications and restored some components of tear homeostasis confirming the efficacy of topical NTX as a treatment for ocular defects in diabetes.
Topics: Animals; Lacrimal Apparatus; Tears; Naltrexone; Rats, Sprague-Dawley; Male; Diabetes Mellitus, Experimental; Rats; Aquaporin 5; Administration, Topical; Dry Eye Syndromes
PubMed: 38756167
DOI: 10.3389/ebm.2024.10175 -
Journal of Neuroinflammation May 2024Epidemiological studies have unveiled a robust link between exposure to repetitive mild traumatic brain injury (r-mTBI) and elevated susceptibility to develop...
Overexpression of pathogenic tau in astrocytes causes a reduction in AQP4 and GLT1, an immunosuppressed phenotype and unique transcriptional responses to repetitive mild TBI without appreciable changes in tauopathy.
Epidemiological studies have unveiled a robust link between exposure to repetitive mild traumatic brain injury (r-mTBI) and elevated susceptibility to develop neurodegenerative disorders, notably chronic traumatic encephalopathy (CTE). The pathogenic lesion in CTE cases is characterized by the accumulation of hyperphosphorylated tau in neurons around small cerebral blood vessels which can be accompanied by astrocytes that contain phosphorylated tau, the latter termed tau astrogliopathy. However, the contribution of tau astrogliopathy to the pathobiology and functional consequences of r-mTBI/CTE or whether it is merely a consequence of aging remains unclear. We addressed these pivotal questions by utilizing a mouse model harboring tau-bearing astrocytes, GFAP mice, subjected to our r-mTBI paradigm. Despite the fact that r-mTBI did not exacerbate tau astrogliopathy or general tauopathy, it increased phosphorylated tau in the area underneath the impact site. Additionally, gene ontology analysis of tau-bearing astrocytes following r-mTBI revealed profound alterations in key biological processes including immunological and mitochondrial bioenergetics. Moreover, gene array analysis of microdissected astrocytes accrued from stage IV CTE human brains revealed an immunosuppressed astroglial phenotype similar to tau-bearing astrocytes in the GFAP model. Additionally, hippocampal reduction of proteins involved in water transport (AQP4) and glutamate homeostasis (GLT1) was found in the mouse model of tau astrogliopathy. Collectively, these findings reveal the importance of understanding tau astrogliopathy and its role in astroglial pathobiology under normal circumstances and following r-mTBI. The identified mechanisms using this GFAP model may suggest targets for therapeutic interventions in r-mTBI pathogenesis in the context of CTE.
Topics: Astrocytes; Animals; Mice; tau Proteins; Aquaporin 4; Tauopathies; Humans; Excitatory Amino Acid Transporter 2; Mice, Transgenic; Brain Concussion; Male; Phenotype; Mice, Inbred C57BL
PubMed: 38750510
DOI: 10.1186/s12974-024-03117-4 -
MSphere Jun 2024The RNA chaperone Hfq acts as a global regulator of numerous biological processes, such as carbon/nitrogen metabolism and environmental adaptation in plant-associated...
UNLABELLED
The RNA chaperone Hfq acts as a global regulator of numerous biological processes, such as carbon/nitrogen metabolism and environmental adaptation in plant-associated diazotrophs; however, its target RNAs and the mechanisms underlying nitrogen fixation remain largely unknown. Here, we used enhanced UV cross-linking immunoprecipitation coupled with high-throughput sequencing to identify hundreds of Hfq-binding RNAs probably involved in nitrogen fixation, carbon substrate utilization, biofilm formation, and other functions. Collectively, these processes endow strain A1501 with the requisite capabilities to thrive in the highly competitive rhizosphere. Our findings revealed a previously uncharted landscape of Hfq target genes. Notable among these is , encoding an isomerase necessary for nitrogenase reductase solubility; , encoding an ammonium transporter; , encoding a carbohydrate porin; and , encoding a chemotaxis protein. Furthermore, we identified more than 100 genes of unknown function, which expands the potential direct regulatory targets of Hfq in diazotrophs. Our data showed that Hfq directly interacts with the mRNA of regulatory proteins (RsmA, AlgU, and NifA), regulatory ncRNA RsmY, and other potential targets, thus revealing the mechanistic links in nitrogen fixation and other metabolic pathways.
IMPORTANCE
Numerous experimental approaches often face challenges in distinguishing between direct and indirect effects of Hfq-mediated regulation. New technologies based on high-throughput sequencing are increasingly providing insight into the global regulation of Hfq in gene expression. Here, enhanced UV cross-linking immunoprecipitation coupled with high-throughput sequencing was employed to identify the Hfq-binding sites and potential targets in the root-associated A1501 and identify hundreds of novel Hfq-binding RNAs that are predicted to be involved in metabolism, environmental adaptation, and nitrogen fixation. In particular, we have shown Hfq interactions with various regulatory proteins' mRNA and their potential targets at the posttranscriptional level. This study not only enhances our understanding of Hfq regulation but, importantly, also provides a framework for addressing integrated regulatory network underlying root-associated nitrogen fixation.
Topics: Pseudomonas stutzeri; Host Factor 1 Protein; Nitrogen Fixation; Gene Expression Regulation, Bacterial; Plant Roots; RNA, Bacterial; Gene Expression Profiling; Gene Regulatory Networks; Bacterial Proteins; High-Throughput Nucleotide Sequencing; Transcriptome; Rhizosphere
PubMed: 38747590
DOI: 10.1128/msphere.00762-23 -
Nature Communications May 2024Pentamidine and melarsoprol are primary drugs used to treat the lethal human sleeping sickness caused by the parasite Trypanosoma brucei. Cross-resistance to these two...
Pentamidine and melarsoprol are primary drugs used to treat the lethal human sleeping sickness caused by the parasite Trypanosoma brucei. Cross-resistance to these two drugs has recently been linked to aquaglyceroporin 2 of the trypanosome (TbAQP2). TbAQP2 is the first member of the aquaporin family described as capable of drug transport; however, the underlying mechanism remains unclear. Here, we present cryo-electron microscopy structures of TbAQP2 bound to pentamidine or melarsoprol. Our structural studies, together with the molecular dynamic simulations, reveal the mechanisms shaping substrate specificity and drug permeation. Multiple amino acids in TbAQP2, near the extracellular entrance and inside the pore, create an expanded conducting tunnel, sterically and energetically allowing the permeation of pentamidine and melarsoprol. Our study elucidates the mechanism of drug transport by TbAQP2, providing valuable insights to inform the design of drugs against trypanosomiasis.
Topics: Trypanosoma brucei brucei; Aquaglyceroporins; Cryoelectron Microscopy; Molecular Dynamics Simulation; Melarsoprol; Pentamidine; Biological Transport; Trypanocidal Agents; Protozoan Proteins; Humans
PubMed: 38734677
DOI: 10.1038/s41467-024-48445-4 -
Theriogenology Aug 2024Pigs are usually bred through artificial insemination with liquid semen preserved at 15-20 °C. While this method of preservation brings many benefits, including a...
Pigs are usually bred through artificial insemination with liquid semen preserved at 15-20 °C. While this method of preservation brings many benefits, including a greater reproductive performance compared to frozen-thawed sperm, the period of storage is a limiting factor. As the mitochondrion regulates many facets of sperm physiology, modulating its activity could have an impact on their lifespan. Aligned with this hypothesis, the present study sought to investigate whether inhibition of voltage-dependent anion channels (VDACs), which reside in the outer mitochondrial membrane and regulate the flux of ions between mitochondria and the cytosol in somatic cells, influences the resilience of pig sperm to liquid preservation at 17 °C. For this purpose, semen samples (N = 7) were treated with two different concentrations of TRO19622 (5 μM and 50 μM), an inhibitor of VDACs, and stored at 17 °C for 10 days. At days 0, 4 and 10, sperm quality and functionality parameters were evaluated by flow cytometry and computer-assisted sperm analysis (CASA). The effects of inhibiting VDACs depended on the concentration of the inhibitor. On the one hand, the greatest concentration of TRO19622 (50 μM) led to a decrease in sperm motility, viability and mitochondrial membrane potential, which could be related to the observed intracellular Ca increase. In contrast, total sperm motility was higher in samples treated with 5 μM TRO19622 than in the control, suggesting that when VDACs channels are inhibited by the lowest concentration of the blocking agent the resilience of pig sperm to liquid storage increases. In conclusion, the current research indicates that mitochondrial function, as regulated by ion channels in the outer mitochondrial membrane like VDACs, is related to the sperm resilience to liquid preservation and may influence cell lifespan.
Topics: Anions; Animals; Swine; Semen Preservation; Insemination, Artificial; Voltage-Dependent Anion Channels; Temperature; Cholestenones; Semen; Sperm Motility; Calcium
PubMed: 38723471
DOI: 10.1016/j.theriogenology.2024.05.003 -
Science Advances May 2024Fast collective motions are widely present in biomolecules, but their functional relevance remains unclear. Herein, we reveal that fast collective motions of backbone...
Fast collective motions are widely present in biomolecules, but their functional relevance remains unclear. Herein, we reveal that fast collective motions of backbone are critical to the water transfer of aquaporin Z (AqpZ) by using solid-state nuclear magnetic resonance (ssNMR) spectroscopy and molecular dynamics (MD) simulations. A total of 212 residue site-specific dipolar order parameters and 158 N spin relaxation rates of the backbone are measured by combining the C- and H-detected multidimensional ssNMR spectra. Analysis of these experimental data by theoretic models suggests that the small-amplitude (~10°) collective motions of the transmembrane α helices on the nanosecond-to-microsecond timescales are dominant for the dynamics of AqpZ. The MD simulations demonstrate that these collective motions are critical to the water transfer efficiency of AqpZ by facilitating the opening of the channel and accelerating the water-residue hydrogen bonds renewing in the selectivity filter region.
Topics: Water; Aquaporins; Molecular Dynamics Simulation; Protein Conformation, alpha-Helical; Hydrogen Bonding; Magnetic Resonance Spectroscopy; Nuclear Magnetic Resonance, Biomolecular; Escherichia coli Proteins
PubMed: 38718112
DOI: 10.1126/sciadv.ade9520 -
Porin deficiency or plasmid copy number increase mediated carbapenem-resistant resistance evolution.Emerging Microbes & Infections Dec 2024This study investigated resistance evolution mechanisms of conjugated plasmids and bacterial hosts under different concentrations of antibiotic pressure. Ancestral...
This study investigated resistance evolution mechanisms of conjugated plasmids and bacterial hosts under different concentrations of antibiotic pressure. Ancestral strain ECNX52 was constructed by introducing the -carrying IncX3 plasmid into C600, and was subjected to laboratory evolution under different concentrations of meropenem pressure. Minimal inhibitory concentrations and conjugation frequency were determined. Fitness of these strains was assessed. Whole genome sequencing and transcriptional changes were performed. Ancestral host or plasmids were recombined with evolved hosts or plasmids to verify plasmid or host factors in resistance evolution. Role of the mutation on plasmid copy number was determined. Two out of the four clones (EM2N1 and EM2N3) exhibited four-fold increase in MIC when exposed to a continuous pressure of 2 μg/mL MEM (1/32 MIC), by down regulating expression of outer membrane protein . Besides, all four clones displayed four-fold increase in MIC and higher conjugation frequency when subjected to a continuous pressure of 4 μg/mL MEM (1/16 MIC), attributing to increasing plasmid copy number generated by D140Y (GAT→TAT) mutation. Bacterial hosts and conjugative plasmids can undergo resistance evolution under certain concentrations of antimicrobial pressure by reducing the expression of outer membrane proteins or increasing plasmid copy numbers.
Topics: Escherichia coli; Plasmids; Microbial Sensitivity Tests; Anti-Bacterial Agents; Porins; Escherichia coli Proteins; Carbapenems; Meropenem; Mutation; Evolution, Molecular; Conjugation, Genetic; Carbapenem-Resistant Enterobacteriaceae; Whole Genome Sequencing; Gene Dosage; beta-Lactamases
PubMed: 38712634
DOI: 10.1080/22221751.2024.2352432 -
Neurology May 2024Knowledge of the evolution of CNS demyelinating lesions within attacks could assist diagnosis. We evaluated intra-attack lesion dynamics in patients with myelin... (Observational Study)
Observational Study
BACKGROUND AND OBJECTIVES
Knowledge of the evolution of CNS demyelinating lesions within attacks could assist diagnosis. We evaluated intra-attack lesion dynamics in patients with myelin oligodendrocyte glycoprotein antibody-associated disease (MOGAD) vs multiple sclerosis (MS) and aquaporin-4 antibody seropositive neuromyelitis optica spectrum disorder (AQP4+NMOSD).
METHODS
This retrospective observational multicenter study included consecutive patients from Mayo Clinic (USA) and Great Ormond Street Hospital for Children (UK). Inclusion criteria were as follows: (1) MOGAD, MS, or AQP4+NMOSD diagnosis; (2) availability of ≥2 brain MRIs (within 30 days of attack onset); and (3) brain involvement (i.e., ≥1 T2 lesion) on ≥1 brain MRI. The initial and subsequent brain MRIs within a single attack were evaluated for the following: new T2 lesions(s); resolved T2 lesion(s); both; or no change. This was compared between MOGAD, MS, and AQP4+NMOSD attacks. We used the Mann-Whitney test and χ/Fisher exact test for statistical analysis.
RESULTS
Our cohort included 55 patients with MOGAD (median age, 14 years; interquartile range [IQR] 5-34; female sex, 29 [53%]) for a total of 58 attacks. The comparison groups included 38 patients with MS, and 19 with AQP4+NMOSD. In MOGAD, the initial brain MRI (median of 5 days from onset [IQR 3-9]) was normal in 6/58 (10%) attacks despite cerebral symptoms (i.e., radiologic lag). The commonest reason for repeat MRI was clinical worsening or no improvement (33/56 [59%] attacks with details available). When compared with the first MRI, the second intra-attack MRI (median of 8 days from initial scan [IQR 5-13]) showed the following: new T2 lesion(s) 27/58 (47%); stability 24/58 (41%); resolution of T2 lesion(s) 4/58 (7%); or both new and resolved T2 lesions 3/58 (5%). Findings were similar between children and adults. Steroid treatment was associated with resolution of ≥1 T2 lesion (6/28 [21%] vs 1/30 [3%], = 0.048) and reduced the likelihood of new T2 lesions (9/28 vs 18/30, = 0.03). Intra-attack MRI changes favored MOGAD (34/58 [59%]) over MS (10/38 [26%], = 0.002) and AQP4+NMOSD (4/19 [21%], = 0.007). Resolution of ≥1 T2 lesions was exclusive to MOGAD (7/58 [12%]).
DISCUSSION
Radiologic lag is common within MOGAD attacks. Dynamic imaging with frequent appearance and occasional disappearance of lesions within a single attack suggest MOGAD diagnosis over MS and AQP4+NMOSD. These findings have implications for clinical practice, clinical trial attack adjudication, and understanding of MOGAD pathogenesis.
Topics: Humans; Female; Male; Myelin-Oligodendrocyte Glycoprotein; Magnetic Resonance Imaging; Adolescent; Child; Retrospective Studies; Brain; Multiple Sclerosis; Aquaporin 4; Neuromyelitis Optica; Young Adult; Autoantibodies; Adult; Disease Progression
PubMed: 38710000
DOI: 10.1212/WNL.0000000000209303