Journal of Ultrastructure Research
A low-viscosity epoxy resin embedding medium for electron microscopy
A low-viscosity embedding medium based on ERL-4206 is recommended for use in electron microscopy. The composition is: ERL-4206 (vinyl cyclohexene dioxide) 10 g, D.E.R. 736 (diglycidyl ether of polypropylene glycol) 6 g, NSA (nonenyl succinic anhydride) 26 g, and S-1 (dimethylaminoethanol or DMAE) 0.4 g. The medium is easily and rapidly prepared by dispensing the components, in turn by weight, into a single flask. The relatively low viscosity of the medium (60 cP) permits rapid mixing by shaking and swirling. The medium is infiltrated into specimens after the use of any one of several dehydrating fluids, such as ethanol, acetone, dioxan, hexylene glycol, isopropyl alcohol, propylene oxide, and tert.-butyl alcohol. It is compatible with each of these in all proportions. After infiltration the castings are polymerized at 70°C in 8 hours. Longer curing does not adversely affect the physical properties of the castings. Curing time can be reduced by increasing the temperature or the accelerator, S-1, or both; and the hardness of the castings is controlled by changes in the D.E.R. 736 flexibilizer. The medium has a long pot life of several days and infiltrates readily because of its low viscosity. The castings have good trimming and sectioning qualities. The embedding matrix of the sections is very resistant to oxidation by KMnO4 and Ba(MnO4)2, compared with resins containing NADIC methyl anhydride. Sections are tough under the electron beam and may be used without a supporting membrane on the grids. The background plastic in the sections shows no perceptible substructure at magnifications commonly used for biological materials. The medium has been used successfully with a wide range of specimens, including endosperms with a high lipid content, tissues with hard, lignified cell walls, and highly vacuolated parenchymatous tissues of ripe fruits.
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Loss of mitochondrial adaptation associates with deterioration of mitochondrial turnover and structure in metabolic dysfunction-associated steatotic liver disease
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This cross-sectional study compared individuals with class III obesity (n = 8/group) without (control, OBE CON; NAFLD activity score: 0.4 ± 0.1) or with steatosis (OBE MASL, 2.3 ± 0.4), or MASH (OBE MASH, 5.3 ± 0.3, p < 0.05 vs. other groups). Hepatic mitochondrial ultrastructure was assessed by transmission electron microscopy, mitochondrial respiration by high-resolution respirometry, biomarkers of mitochondrial quality control and endoplasmic reticulum (ER) stress by Western Blot.
Mitochondrial oxidative capacity was 31 % higher in OBE MASL, but 25 % lower in OBE MASH (p < 0.05 vs. OBE CON). OBE MASH showed ~1.5fold lower mitochondrial number, but ~1.2–1.5fold higher diameter and area (p < 0.001 vs. other groups). Biomarkers of autophagy (p62), mitophagy (PINK1, PARKIN), fission (DRP-1, FIS1) and fusion (MFN1/2, OPA1) were reduced in OBE MASH (p < 0.05 vs. OBE CON). OBE MASL showed lower p62, p-PARKIN/PARKIN, and p-DRP-1 (p < 0.05 vs. OBE CON). OBE MASL and MASH showed higher ER stress markers (PERK, ATF4, p-eIF2α-S51/eIF2α; p < 0.05 vs. OBE CON). Mitochondrial diameter associated inversely with fusion/fission biomarkers and with oxidative capacity, but positively with H2O2.
Humans with hepatic steatosis already exhibit impaired mitochondrial turnover, despite upregulated oxidative capacity, and evidence for ER stress. In MASH, oxidative stress likely mediates progressive decline of mitochondrial turnover, ultrastructure and respiration indicating that mitochondrial quality control is key for energy metabolism and may have potential for targeting MASH.
ClinGovTrial: NCT01477957
Gold nanoparticle adsorption alters the cell stiffness and cell wall bio-chemical landscape of Candida albicans fungal cells
2024, Journal of Colloid and Interface ScienceNanomaterials have been extensively investigated for a wide range of biomedical applications, including as antimicrobial agents, drug delivery vehicles, and diagnostic devices. The commonality between these biomedical applications is the necessity for the nanoparticle to interact with or pass through the cellular wall and membrane. Cell-nanomaterial interactions/uptake can occur in various ways, including adhering to the cell wall, forming aggregates on the surface, becoming absorbed within the cell wall itself, or transversing into the cell cytoplasm. These interactions are common to mammalian cells, bacteria, and yeast cells. This variety of interactions can cause changes to the integrity of the cell wall and the cell overall, but the precise mechanisms underpinning such interactions remain poorly understood. Here, we investigate the interaction between commonly investigated gold nanoparticles (AuNPs) and the cell wall/membrane of a model fungal cell to explore the general effects of interaction and uptake.
The interactions between 100 nm citrate-capped AuNPs and the cell wall of Candida albicans fungal cells were studied using a range of advanced microscopy techniques, including atomic force microscopy, confocal laser scanning microscopy, scanning electron microscopy, transmission electron microscopy, and synchrotron-FTIR micro-spectroscopy.
In most cases, particles adhered on the cell surface, although instances of particles being up-taken into the cell cytoplasm and localised within the cell wall and membrane were also observed. There was a measurable increase in the stiffness of the fungal cell after AuNPs were introduced. Analysis of the synchrotron-FTIR data showed significant changes in spectral features associated with phospholipids and proteins after exposure to AuNPs.
Low soil moisture improved shading tolerance by regulating leaf functional traits in Tetracentron sinense Oliv. seedlings
2023, Global Ecology and ConservationTetracentron sinense Oliv. is an endangered relict tree found in south-central China. A low number of seedlings among the natural populations has resulted in the poor regeneration of this species. However, the mechanisms underlying seedling survival and growth remain poorly understood. Leaf functional traits (LFT) likely characterize the growth and development of T. sinense in different ecological environments, playing a crucial role in its adaptive strategies to adversity. In this study, two-factor orthogonal control experiments with two levels of light intensity (L1: 50 % natural light intensity in forest edge; L2: 10 % natural light intensity in understory) and soil moisture content (W1: 30 % moisture content in valley; W2: 15 % moisture content in mountainside) were conducted to analyze the response of the LFT of 7-month-old T. sinense seedlings to different light and soil moisture conditions. With the light intensity decreased, superoxide dismutase activity decreased, and peroxidase activity increased. With a decrease in soil moisture, the contents of chlorophyll (Chl) a, Chl b, Chl t, and soluble sugar increased, and Chl a/b decreased. A decrease in light and soil moisture decreased leaf thickness, leaf dry weight, leaf dry matter content, leaf volume, chloroplast volume, and starch grains, but increased osmophilic grains, the total nitrogen, and phosphorus contents. Moreover, the chloroplast morphology of L2W2 was more complete than that of other treatments. The results showed that the LFT of T. sinense seedlings acted synergistically to adapt to different light and soil moisture conditions, low soil moisture alleviated the effects of shading in T. sinense seedlings, and T. sinense seedlings adopted the “quick investment-return strategy” adaptation strategy under L2W2. Finally, the soil moisture of the mountainside in the natural habitat was favorable to the survival and growth of T. sinense seedlings.
Copper and zinc accumulation, distribution, and tolerance in Pistia stratiotes L.; revealing the role of root caps
2023, Aquatic ToxicologyPollution by potentially toxic trace metals, such as copper or zinc, is global. Both Cu and Zn are essential microelements, which in higher concentrations become toxic. The aquatic plant Pistia stratiotes(L. has great potential for phytoremediation. Also, it has an unusually large and easily detachable root cap, which makes it a suitable model for studying the potential role of the root cap in metal uptake. Plant response to environmentally relevant concentrations of Cu (0.1, 0.3, and 1 μM) and Zn (0.3, 1, and 3 μM) was investigated with the aim of studying their interaction and distribution at the root tissue level as well as revealing their tolerance mechanisms. Changes in the root anatomy and plant ionome were determined using light and fluorescence microscopy, ICP-MS, and μXRF imaging. Alterations in photosynthetic activity caused by Cu or Zn excesses were monitored by direct imaging of fast chlorophyll fluorescence kinetics (OJIP). Fe and Mn were preferentially localized in the root cap, while Ca, Cu, Ni, and Zn were mainly in the root tip regardless of the Cu/Zn treatment. Translocation of Cu and Zn to the leaves increased with higher doses, however the translocation factor was the lowest in the highest treatments. Measurements of photosynthetic parameters showed a higher susceptibility of electron transport flux from QA to QB under increasing Cu than Zn supply. This, along with our findings regarding the root anatomy and the differences in Ca accumulation and distribution, led to the conclusion that P. stratiotes is more effective for Zn remediation than Cu.
N-terminal truncation of PhaC<inf>BP-M-CPF4</inf> and its effect on PHA production
2024, Microbial Cell Factories