A rapid method of whole cell sample preparation for scanning electron microscopy using neodymium chloride

Новиков, И. А.; Subbot, A.M.; Turenok, Andrey; Mayanskiy, Nikolay; Chebotar, I. V.

doi:10.1016/j.micron.2019.102687

Cited by 11 publications

(11 citation statements)

References 12 publications

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Section: General Morphology Of the Cell Cell Movement Analysismentioning

confidence: 52%

“…Indeed, both external and internal composition of cells could be clearly visualized, which suggests pervasion of Nd into the interior of the cells with an integration into multiple cellular organelles and membranes. We also noted that cells preserved their 3D shape under vacuum conditions, which suggests a powerful inhibition of membrane channels [9]. In preliminary observations, we noted the lack of organelles movement and decided to study this fact in more detail by observing the dynamics of the lanthanoid treatment process via confocal microscopy time series (video No.…”

Section: General Morphology Of the Cell Cell Movement Analysismentioning

confidence: 98%

“…The lanthanoids are also actively used in microscopy as contrasting agents since they bind to structures responsible for cellular metabolism and architecture. We recently reported a new method of whole-cell sample preparation for scanning electron microscopy (SEM) based on staining/fixation with a rare earth metal alone [9].…”

Section: Introductionmentioning

confidence: 99%

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Life-On-Hold: Lanthanoids Rapidly Induce a Reversible Ametabolic State in Mammalian Cells

Subbot

Kondratieva

et al. 2021

Biology

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Until now, the ability to reversibly halt cellular processes has been limited to cryopreservation and several forms of anabiosis observed in living organisms. In this paper we show that incubation of living cells with a solution containing ~50 mM neodymium induces a rapid shutdown of intracellular organelle movement and all other evidence of active metabolism. We have named this state REEbernation (derived from the terms REE (rare earth elements) and hibernation) and found that the process involves a rapid replacement of calcium with neodymium in membranes and organelles of a cell, allowing it to maintain its shape and membrane integrity under extreme conditions, such as low pressure. Furthermore, phosphate exchange is blocked as a result of non-dissolvable neodymium salts formation, which “discharged” the cell. We further showed that REEbernation is characterized by an immediate cessation of transcriptional activity in observed cells, providing an intriguing opportunity to study a snapshot of gene expression at a given time point. Finally, we found that the REEbernation state is reversible, and we could restore the metabolism and proliferation capacity of the cells. The REEbernation, in addition to being an attractive model to further investigate the basic mechanisms of cell metabolism control, also provides a new method to reversibly place a cell into “on-hold” mode, opening opportunities to develop protocols for biological samples fixation with a minimum effect on the omics profile for biomedical needs.

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Section: General Morphology Of the Cell Cell Movement Analysismentioning

confidence: 52%

Section: General Morphology Of the Cell Cell Movement Analysismentioning

confidence: 98%

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Life-On-Hold: Lanthanoids Rapidly Induce a Reversible Ametabolic State in Mammalian Cells

Subbot

Kondratieva

et al. 2021

Biology

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“…Morphology of the surface and the nearest subsurface layer of samples was examined with SEM using lanthanoid contrast [18]. The treatment protocol included primary flushing, holding for 45 minutes in contrasting solution BioREE (JSC Glaucon, Russia) and final rinsing with distilled water.…”

Section: Scanning Electron Microscopy (Sem) Preparation and Imagingmentioning

confidence: 99%

A Comparison of the Capacity of Mesenchymal Stromal Cells for Cartilage Regeneration Depending on Collagen-Based Injectable Biomimetic Scaffold Type

Sevastianov

Basok

Кирсанова

et al. 2021

Preprint

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Mesenchymal stromal cells (MSCs) have shown a high potential for cartilage repair. Collagen-based scaffolds are used to deliver and retain cells at the site of cartilage damage. The aim of the work was a comparative analysis of the capacity of the MSCs from human adipose tissue to differentiate into chondrocytes in vitro and to stimulate the regeneration of articular cartilage in an experimental model of rabbit knee osteoarthrosis when cultured on microheterogenic collagen-based hydrogel (МCH) and the microparticles of decellularized porcine articular cartilage (DPC). The morphology of samples was evaluated using scanning electron microscopy and histological staining methods. On the surface of the DPC, the cells were distributed more uniformly than on the MCH surface. On day 28, the cells cultured on the DPC produced glycosaminoglycans more intensely compared to the MCH with the synthesis of collagen type II. However, in the experimental model of osteoarthrosis, the stimulation of the cartilage regeneration was more effective when the MSCs were administered to the MCH carrier. The present study demonstrates the way to regulate the action of the MSCs in the area of cartilage regeneration: the MCH is more conducive to stimulating cartilage repair by the MSCs, while the DPC is an inducer for a formation of a cartilage-like tissue by the MSCs in vitro.

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“…Метод лантаноидного контрастирования позволяет наблюдать нефиксированные биологические образцы в режиме низкого вакуума после выдержки их в насыщенном растворе редкоземельного металла. При этом сохраняется максимально нативное состояние исследуемого объекта, а изображение, полученное в режиме детекции обратно рассеянных электронов, несет расширенную информацию о клеточных структурах [15].…”

Section: материалы и методыunclassified

Experimental approaches to creating a tissue-specific matrix for a bioartificial liver

Григорьев

Басок

Kirillova

et al. 2020

RJTAO

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Shortage of donor organs for liver transplantation in the treatment of end-stage liver disease dictates the need to develop alternative methods that include technologies on tissue engineering and regenerative medicine. Objective: to study the ability of a tissue-specific matrix from decellularized human liver fragments (DHLF) to maintain adhesion and proliferation of human adipose tissue-derived mesenchymal stem cells (hAT-MSCs) and HepG2 under static conditions and in a flow-through bioreactor. Materials and methods. Treatment with surfactants (SAS) – sodium dodecyl sulfate, Triton X-100 – followed by exposure to DNase was used for decellularization of human liver fragments (no more than 8 mm3). Biochemical screening included the determination of DNA quantity in the test samples. Efficiency of surfactant washing was assessed by the cytotoxicity of the matrix in the NIH 3T3 fibroblast culture. Viability and metabolic activity of cells were assessed via vital staining with a complex of fluorescent dyes LIVE/DEAD ® and PrestoBlue™ (Invitrogen, USA). Morphological examination of the liver cell-engineered constructs was carried out through histological staining and scanning electron microscopy with lanthanide contrast. Results. It was shown that the liver decellularization method used allows to obtain a biocompatible matrix with a residual DNA quantity <1%, which is capable of maintaining adhesion and proliferation of hAT-MSCs and HepG2. On day 7 of cultivation in the bioreactor, there was formation of a single conglomerate of the DHLF matrix with numerous groups of viable cells with a high nuclear-cytoplasmic ratio. The urea content in the culture medium is 1.5 ± 0.1 mmol/L, exceeding that of samples obtained under static conditions. This indicates the metabolic activity of HepG2 in the composition of the obtained culture systems. It was shown that constant flow of the culture medium in the perfusion bioreactor increased the proliferative activity of HepG2 and allowed to provide a more uniform colonization by matrix cells in comparison with static cultivation conditions. Conclusion. The conditions for uniform colonization of DHLFs in a flow-through bioreactor with cell cultures were established. The ability of the matrix to maintain adhesion and proliferation of hADSCs and HepG2 for 11 days indicates that it could be used in liver tissue engineering.

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A rapid method of whole cell sample preparation for scanning electron microscopy using neodymium chloride

Cited by 11 publications

References 12 publications

Life-On-Hold: Lanthanoids Rapidly Induce a Reversible Ametabolic State in Mammalian Cells

Life-On-Hold: Lanthanoids Rapidly Induce a Reversible Ametabolic State in Mammalian Cells

A Comparison of the Capacity of Mesenchymal Stromal Cells for Cartilage Regeneration Depending on Collagen-Based Injectable Biomimetic Scaffold Type

Experimental approaches to creating a tissue-specific matrix for a bioartificial liver

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