-
Cellular and Molecular Neurobiology Nov 2023Isolated exposure to intermittent hypoxia and permissive hypercapnia activates signaling mechanisms that induce ultrastructural changes in mitochondria and endoplasmic...
Isolated exposure to intermittent hypoxia and permissive hypercapnia activates signaling mechanisms that induce ultrastructural changes in mitochondria and endoplasmic reticulum, accompanied by the development of maximal ischemic tolerance in neurons under the combined influence of these factors. However, there are a lack of data on the combined impact of these factors on the ultrastructure of neuronal organelles. The present study aims to comparatively assess the ultrastructural changes in neurons following isolated and combined exposure to hypoxia and hypercapnia, as well as to correlate these changes with the neuroprotective potential previously observed for these factors. Following a 15-session course of 30-min exposures to permissive hypercapnia (P ≈ 50 mmHg) and/or normobaric hypoxia (P ≈ 150 mmHg), morphometric assessment was conducted to evaluate the extent of ultrastructural changes in hippocampal neurons (mitochondria, perinuclear space, and granular endoplasmic reticulum). It was found that in hippocampal neurons from the CA1 region, permissive hypercapnia resulted in increased mitochondrial size, expansion of membranous compartments of the granular endoplasmic reticulum, and perinuclear space. Normobaric hypoxia affected only mitochondrial size, while hypercapnic hypoxia specifically widened the perinuclear space. These ultrastructural changes objectively reflect varying degrees of the influence of hypoxia and hypercapnia on organelles responsible for energy metabolism, anti-apoptotic, and synthetic functions of neurons. This confirms the effect of potentiation of their neuroprotective effects under combined exposure and highlights the dominant role of the hypercapnic component in this mechanism.
Topics: Humans; Hypercapnia; Hypoxia; Neurons; Cerebral Cortex; Hippocampus
PubMed: 37716927
DOI: 10.1007/s10571-023-01407-8 -
Journal of Lipid Research Aug 2018Neutrophils form neutrophil extracellular traps (NETs), which have been implicated in microcirculatory plugging. NET formation (NETosis) involves the fusion of granule...
Neutrophils form neutrophil extracellular traps (NETs), which have been implicated in microcirculatory plugging. NET formation (NETosis) involves the fusion of granule and nuclear contents, which are then released in the extracellular space. Myeloperoxidase (MPO) plays a major role in NETosis leading to the dissociation of DNA from histones. During neutrophil activation, MPO is released and activated to convert hydrogen peroxide and chloride to hypochlorous acid (HOCl). HOCl targets plasmalogens leading to the production of the chlorinated lipids, 2-chlorofatty aldehyde and 2-chlorofatty acid (2-ClFA). Here, we tested the hypothesis that 2-ClFAs are important lipid mediators of NETosis. Human neutrophils treated with physiological levels of 2-ClFAs formed NETs, characterized by MPO association with DNA and neutrophil elastase (NE) redistribution to the perinuclear area. 2-ClFA-induced NETs reduced colony forming units. 2-ClFA-induced NETosis is calcium- and protein arginine deiminase 4-dependent. Interestingly, unlike PMA, 2-ClFA initiates the NETosis process without neutrophil activation and degranulation. Furthermore, 2-ClFA elicits NETosis in bone-marrow derived neutrophils from MPO-deficient mice. Taken together, these findings suggest 2-ClFA as an MPO product that triggers the NETosis pathway following neutrophil activation.
Topics: Calcium; DNA; Extracellular Traps; Fatty Acids; Healthy Volunteers; Humans; Lipid Metabolism; Neutrophils; Protein-Arginine Deiminase Type 4; Protein-Arginine Deiminases
PubMed: 29739865
DOI: 10.1194/jlr.M084731 -
Surgical Pathology Clinics Mar 2023CDC73 alterations are associated with three main parathyroid lesions according to the World Health Organization (WHO) classification of tumors of the endocrine system.... (Review)
Review
CDC73 alterations are associated with three main parathyroid lesions according to the World Health Organization (WHO) classification of tumors of the endocrine system. These include hyperparathyroidism-jaw tumor (HPT-JT) syndrome-associated adenomas, atypical parathyroid tumors (APTs), and parathyroid carcinomas (PCs). The loss of nuclear parafibromin expression, which serves as a surrogate marker for the underlying CDC73 alteration, encompasses these tumors under the term parafibromin-deficient parathyroid tumors. They have distinct morphologic features of more abundant eosinophilic cytoplasm with perinuclear clearing surrounding a large nucleus as well as prominent dilated branching "hemangiopericytoma-like" vasculature and a thick capsule as well as variably sized cystic spaces. These tumors include cases that show unequivocal histologic features fulfilling the criteria for PCs with growing data indicating a higher rate of recurrence or metastasis compared with parafibromin intact PCs. More importantly, the loss of parafibromin expression can be used in clinical practice to recognize APTs that fall short of a conclusive diagnosis of PCs, but clinically behave akin to them. Moreover, recognizing these tumors can lead to an underlying germline mutation and a diagnosis of HPT-JT, which impacts long-term treatment and surveillance for patients and close family.
Topics: Humans; Parathyroid Neoplasms; Tumor Suppressor Proteins; Hyperparathyroidism; Jaw Neoplasms; Neoplastic Syndromes, Hereditary; Transcription Factors
PubMed: 36739170
DOI: 10.1016/j.path.2022.09.009 -
Journal of Virology Mar 2020Herpesvirus nucleocapsids leave the nucleus by a vesicle-mediated translocation mediated by the viral nuclear egress complex (NEC). The NEC is composed of two conserved...
Herpesvirus nucleocapsids leave the nucleus by a vesicle-mediated translocation mediated by the viral nuclear egress complex (NEC). The NEC is composed of two conserved viral proteins, designated pUL34 and pUL31 in the alphaherpesvirus pseudorabies virus (PrV). It is required for efficient nuclear egress and is sufficient for vesicle formation and scission from the inner nuclear membrane (INM). Structure-based mutagenesis identified a lysine at position 242 (K242) in pUL31, located in the most membrane distal part of the NEC, to be crucial for efficient nucleocapsid incorporation into budding vesicles. Replacing the lysine by alanine (K242A) resulted in accumulations of empty vesicles in the perinuclear space, despite the presence of excess nucleocapsids in the nucleus. However, it remained unclear whether the defect in capsid incorporation was due to interference with a direct, electrostatic interaction between the capsid and the NEC or structural restrictions. To test this, we replaced K242 with several amino acids, thereby modifying the charge, size, and side chain orientation. In addition, virus recombinants expressing pUL31-K242A were passaged and screened for second-site mutations. Compensatory mutations at different locations in pUL31 or pUL34 were identified, pointing to an inherent flexibility of the NEC. In summary, our data suggest that the amino acid at position 242 does not directly interact with the nucleocapsid but that rearrangements in the NEC coat are required for efficient nucleocapsid envelopment at the INM. Herpesviruses encode an exceptional vesicle formation and scission machinery, which operates at the inner nuclear membrane, translocating the viral nucleocapsid from the nucleus into the perinuclear space. The conserved herpesviral nuclear egress complex (NEC) orchestrates this process. High-resolution imaging approaches as well as the recently solved crystal structures of the NEC provided deep insight into the molecular details of vesicle formation and scission. Nevertheless, the molecular mechanism of nucleocapsid incorporation remained unclear. In accordance with structure-based predictions, a basic amino acid could be pinpointed in the most membrane-distal domain of the NEC (pUL31-K242), indicating that capsid incorporation might depend on a direct electrostatic interaction. Our follow-up study, described here, however, shows that the positive charge is not relevant but that the overall structure matters.
Topics: Active Transport, Cell Nucleus; Animals; Capsid Proteins; Cell Line; Cell Nucleus; Chlorocebus aethiops; DNA Mutational Analysis; Follow-Up Studies; Herpesvirus 1, Suid; Models, Molecular; Mutagenesis, Site-Directed; Mutation; Nuclear Envelope; Nucleocapsid; Protein Conformation; Vero Cells; Viral Proteins; Virion
PubMed: 32051272
DOI: 10.1128/JVI.01910-19 -
Journal of the Royal Society, Interface Nov 2016Angiogenesis, the formation of blood vessels from pre-existing ones, is a key event in pathology, including cancer progression, but also in homeostasis and regeneration....
Angiogenesis, the formation of blood vessels from pre-existing ones, is a key event in pathology, including cancer progression, but also in homeostasis and regeneration. As the phenotype of endothelial cells (ECs) is continuously regulated by local biomechanical forces, studying endothelial behaviour in altered gravity might contribute to new insights towards angiogenesis modulation. This study aimed at characterizing EC behaviour after hypergravity exposure (more than 1), with special focus on cytoskeleton architecture and capillary-like structure formation. Herein, human umbilical vein ECs (HUVECs) were cultured under two-dimensional and three-dimensional conditions at 3 and 10 for 4 and 16 h inside the large diameter centrifuge at the European Space Research and Technology Centre (ESTEC) of the European Space Agency. Although no significant tendency regarding cytoskeleton organization was observed for cells exposed to high 's, a slight loss of the perinuclear localization of β-tubulin was observed for cells exposed to 3 with less pronounced peripheral bodies of actin when compared with 1 control cells. Additionally, hypergravity exposure decreased the assembly of HUVECs into capillary-like structures, with a 10 level significantly reducing their organization capacity. In conclusion, short-term hypergravity seems to affect EC phenotype and their angiogenic potential in a time and -level-dependent manner.
Topics: Actins; Human Umbilical Vein Endothelial Cells; Humans; Hypergravity; Neovascularization, Physiologic; Tubulin
PubMed: 28334696
DOI: 10.1098/rsif.2016.0688 -
The Plant Journal : For Cell and... Oct 2020Soybean cyst nematode (SCN; Heterodera glycines) is the largest pathogenic cause of soybean yield loss. The Rhg1 locus is the most used and best characterized SCN...
Soybean cyst nematode (SCN; Heterodera glycines) is the largest pathogenic cause of soybean yield loss. The Rhg1 locus is the most used and best characterized SCN resistance locus, and contains three genes including one encoding an α-SNAP protein. Although the Rhg1 α-SNAP is known to play an important role in vesicle trafficking and SCN resistance, the protein's binding partners and the molecular mechanisms underpinning SCN resistance remain unclear. In this report, we show that the Rhg1 α-SNAP strongly interacts with two syntaxins of the t-SNARE family (Glyma.12G194800 and Glyma.16G154200) in yeast and plants; importantly, the genes encoding these syntaxins co-localize with SCN resistance quantitative trait loci. Fluorescent visualization revealed that the α-SNAP and the two interacting syntaxins localize to the plasma membrane and perinuclear space in both tobacco epidermal and soybean root cells. The two syntaxins and their two homeologs were mutated, individually and in combination, using the CRISPR-Cas9 system in the SCN-resistant Peking and SCN-susceptible Essex soybean lines. Peking roots with deletions introduced into syntaxin genes exhibited significantly reduced resistance to SCN, confirming that t-SNAREs are critical to resisting SCN infection. The results presented here uncover a key step in the molecular mechanism of SCN resistance, and will be invaluable to soybean breeders aiming to develop highly SCN-resistant soybean varieties.
Topics: Animals; Clustered Regularly Interspaced Short Palindromic Repeats; Disease Resistance; Host-Parasite Interactions; Plant Diseases; Plant Proteins; Plant Roots; Plants, Genetically Modified; Qa-SNARE Proteins; Quantitative Trait Loci; SNARE Proteins; Glycine max; Two-Hybrid System Techniques; Tylenchoidea
PubMed: 32645235
DOI: 10.1111/tpj.14923 -
Medical Physics Sep 2023The introduction of Gold NanoParticles (GNPs) in radiotherapy treatments necessitates considerations such as GNP size, location, and quantity, as well as patient...
BACKGROUND
The introduction of Gold NanoParticles (GNPs) in radiotherapy treatments necessitates considerations such as GNP size, location, and quantity, as well as patient geometry and beam quality. Physics considerations span length scales across many orders of magnitude (nanometer-to-centimeter), presenting challenges that often limit the scope of dosimetric studies to either micro- or macroscopic scales.
PURPOSE
To investigate GNP dose-enhanced radiation Therapy (GNPT) through Monte Carlo (MC) simulations that bridge micro-to-macroscopic scales. The work is presented in two parts, with Part I (this work) investigating accurate and efficient MC modeling at the single cell level to calculate nucleus and cytoplasm Dose Enhancement Factors (n,cDEFs), considering a broad parameter space including GNP concentration, GNP intracellular distribution, cell size, and incident photon energy. Part II then evaluates cell dose enhancement factors across macroscopic (tumor) length scales.
METHODS
Different methods of modeling gold within cells are compared, from a contiguous volume of either pure gold or gold-tissue mixture to discrete GNPs in a hexagonal close-packed lattice. MC simulations with EGSnrc are performed to calculate n,cDEF for a cell with radius µm and nucleus µm considering 10 to 370 keV incident photons, gold concentrations from 4 to 24 mg /g , and three different GNP configurations within the cell: GNPs distributed around the surface of the nucleus (perinuclear) or GNPs packed into one (or four) endosome(s). Select simulations are extended to cells with different cell (and nucleus) sizes: 5 µm (2, 3, and 4 µm), 7.35 µm (4 and 6 µm), and 10 µm (7, 8, and 9 µm).
RESULTS
n,cDEFs are sensitive to the method of modeling gold in the cell, with differences of up to 17% observed; the hexagonal lattice of GNPs is chosen (as the most realistic model) for all subsequent simulations. Across cell/nucleus radii, source energies, and gold concentrations, both nDEF and cDEF are highest for GNPs in the perinuclear configuration, compared with GNPs in one (or four) endosome(s). Across all simulations of the (r , r ) = (7.35, 5) µm cell, nDEFs and cDEFs range from unity to 6.83 and 3.87, respectively. Including different cell sizes, nDEFs and cDEFs as high as 21.5 and 5.5, respectively, are observed. Both nDEF and cDEF are maximized at photon energies above the K- or L-edges of gold by 10 to 20 keV.
CONCLUSIONS
Considering 5000 unique simulation scenarios, this work comprehensively investigates many physics trends on DEFs at the cellular level, including demonstrating that cellular DEFs are sensitive to gold modeling approach, intracellular GNP configuration, cell/nucleus size, gold concentration, and incident source energy. These data should prove especially useful in research as well as treatment planning, allowing one to optimize or estimate DEF using not only GNP uptake, but also account for average tumor cell size, incident photon energy, and intracellular configuration of GNPs. Part II will expand the investigation, taking the Part I cell model and applying it in cm-scale phantoms.
Topics: Humans; Gold; Monte Carlo Method; Metal Nanoparticles; Photons; Radiometry
PubMed: 37211878
DOI: 10.1002/mp.16454 -
EMBO Reports Apr 2019Cyclic dinucleotides (CDNs) are important second messenger molecules in prokaryotes and eukaryotes. Within host cells, cytosolic CDNs are detected by STING and alert the...
Cyclic dinucleotides (CDNs) are important second messenger molecules in prokaryotes and eukaryotes. Within host cells, cytosolic CDNs are detected by STING and alert the host by activating innate immunity characterized by type I interferon (IFN) responses. Extracellular bacteria and dying cells can release CDNs, but sensing of extracellular CDNs (eCDNs) by mammalian cells remains elusive. Here, we report that endocytosis facilitates internalization of eCDNs. The DNA sensor cGAS facilitates sensing of endocytosed CDNs, their perinuclear accumulation, and subsequent STING-dependent release of type I IFN Internalized CDNs bind cGAS directly, leading to its dimerization, and the formation of a cGAS/STING complex, which may activate downstream signaling. Thus, eCDNs comprise microbe- and danger-associated molecular patterns that contribute to host-microbe crosstalk during health and disease.
Topics: Animals; Cell Line; Endocytosis; Extracellular Space; Host-Pathogen Interactions; Humans; Immunity, Innate; Interferon Type I; Macrophages; Membrane Proteins; Mice; Models, Molecular; Nucleotides, Cyclic; Nucleotidyltransferases; Protein Binding; Protein Conformation; Protein Multimerization; Second Messenger Systems; Signal Transduction; Structure-Activity Relationship
PubMed: 30872316
DOI: 10.15252/embr.201846293 -
Journal of Morphology Sep 2023Ovarian follicles of sterlets (Acipenser ruthenus) are composed of a single oocyte surrounded by follicular cells (FCs), basal lamina, and thecal cells. Previtellogenic...
Asymmetry in previtellogenic and early vitellogenic oocytes, ultrastructure of follicular cells and egg envelope in the pigmented sterlet, Acipenser ruthenus L. 1758 (Chondrostei, Acipenseriformes).
Ovarian follicles of sterlets (Acipenser ruthenus) are composed of a single oocyte surrounded by follicular cells (FCs), basal lamina, and thecal cells. Previtellogenic oocytes are polarized. Homogeneous ooplasm (contains ribosomes) and granular ooplasm (comprises nuage aggregations of nuclear origin, rough endoplasmic reticulum (RER), Golgi complexes, ribosomes, and mitochondria) are distinguished. Granular ooplasm is initially located near the nucleus, contacts the plasma membrane of the oocyte (oolemma) and forms a thin layer underneath its entire perimeter. Next, a ring that surrounds the nucleus is formed and sends strands directed toward the oolemma. The lipid body composed of lipid droplets forms adjacent to this ring. Later, the granular ooplasm and strands enlarge toward the oolemma, lipid body disperses, and homogeneous ooplasm is no longer present. A thin cortical ooplasm is formed underneath the oolemma and does not contain any organelles. The oocyte nucleus moves to the center. The nucleoplasm contains lampbrush chromosomes, nuclear bodies, and multiple nucleoli. Early vitellogenic oocytes are polarized, too. Three regions in the ooplasm are distinguished: the perinuclear (contains lipid droplets near the nuclear envelope), the endoplasm (contains yolk platelets and lipid droplets), and the periplasm (contains yolk spheres, pigment granules, and microtubules). In all these regions the RER, Golgi complexes, nuage, and mitochondria are present. Micropinocytotic vesicles, Golgi vesicles and precursors of the internal layer of the egg envelope are in the cortical ooplasm. Some FCs delaminate from the follicular epithelium, degenerate and vesicles are released into the perioocytic space. They may contain precursors of egg envelope and may be involved in "cell-cell" communication. The egg envelope (zona radiata, zona pellucida) is made up of three layers: the vitelline envelope (inner layer), the middle layer, and the outer layer. In its deposition, both the oocyte and FCs are engaged.
Topics: Female; Animals; Oocytes; Ovarian Follicle; Fishes; Cytoplasm; Vitellogenesis
PubMed: 37585228
DOI: 10.1002/jmor.21631 -
Molecular Neurobiology May 2018In conditions of proteasomal impairment, the damaged or misfolded proteins, collectively known as aggresome, can accumulate in the perinuclear space and be subsequently...
In conditions of proteasomal impairment, the damaged or misfolded proteins, collectively known as aggresome, can accumulate in the perinuclear space and be subsequently eliminated by autophagy. Abnormal aggregation of microtubule-associated protein tau in the cytoplasm is a common neuropathological feature of tauopathies. The deficiency in ubiquitin carboxy-terminal hydrolase L1 (UCH-L1), a proteasomal deubiquitinating enzyme, is closely related to tau aggregation; however, the associated mechanisms remain unclear. Here, we showed that UCH-L1 inhibition interrupts proteasomal impairment-induced tau aggresome formation. By reducing the production of lysine (K63)-linked ubiquitin chains, UCH-L1 inhibition decreases HDAC6 deacetylase activity and attenuates the interaction of HDAC6 and tau protein, finally leading to tau aggresome formation impairment. All these results indicated that UCH-L1 plays a key role in the process of tau aggresome formation by regulating HDAC6 deacetylase activity and implied that UCH-L1 may act as a signaling molecule to coordinate the effects of the ubiquitin-proteasome system and the autophagy-lysosome pathway, which mediate protein aggregates degradation in the cytoplasm.
Topics: HEK293 Cells; Histone Deacetylase 6; Humans; Indoles; Leupeptins; Oximes; Proteasome Endopeptidase Complex; Proteasome Inhibitors; Protein Aggregates; Ubiquitin; Ubiquitin Thiolesterase; Ubiquitination; tau Proteins
PubMed: 28540657
DOI: 10.1007/s12035-017-0558-7