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A special issue of International Journal of Molecular Sciences (ISSN 1422-0067). This special issue belongs to the section "Biochemistry".

Deadline for manuscript submissions: closed (31 March 2022) | Viewed by 30312

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Special Issue Editor

Prof. Dr. Veronika Kralj-Iglic

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Guest Editor

Biophysics, Faculty of Health Sciences, University of Ljubljana, 1000 Ljubljana, Slovenia
Interests: biological membranes; extracellular vesicles; extracellular particles; small cellular particles; exosomes; clinical biophysics; hip biomechanics
Special Issues, Collections and Topics in MDPI journals

Special Issue Information

Dear Colleagues,

Membranous nanostructures have long dwelt in shadow as regards the majority of scientific society, due to their thinness and fragility. However, with advances in observation techniques that provided proofs of their existence, it has become evident that these tiny structures are key players in physiological and pathophysiological processes. Tunneling nanotubules and nonlamellar lipid-based structures serve as direct transport routes within and between cells while membrane - enclosed bits of cells (cellular vesicles) are more or less free to move with fluids and can mediate also interaction between distant cells, between different tissues and between organisms in global environment. Micro and nano-sized particles, including extracellular vesicles, enveloped viruses and artificial membrane vesicles (liposomes), are all united in one essential feature: they are subjected to natural laws that determine the morphology of their membranes (their shape and composition). Biophysical mechanisms are specific concerning a very general and fundamental issue shaping the world from strings to galaxies - the curvature. This special issue featuring original research, reviews and reflections welcomes experimental and theoretical contributions as well as clinical implications that may elucidate mechanisms involving membranous nanostructures, their effects and relevance in health and disease. Due to emergent COVID-19 issues, virion-based mechanisms are of special interest.

Prof. Veronika Kralj-Iglic
Guest Editor

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Keywords

extracellular vesicles
viruses
liposomes
tunneling nanotubules
enveloped viruses
non-lamellar lipid phases
exosomes
curvature
cell to cell communication
COVID-19

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Published Papers (7 papers)

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Research

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13 pages, 4566 KiB

Open AccessArticle

Different Cutibacterium acnes Phylotypes Release Distinct Extracellular Vesicles

by Anna Chudzik, Paweł Migdał and Mariola Paściak

Int. J. Mol. Sci. 2022, 23(10), 5797; https://doi.org/10.3390/ijms23105797 - 21 May 2022

Cited by 10 | Viewed by 2356

Abstract

Bacterial extracellular vesicles (EVs) perform various biological functions, including those that are critical to microbes. Determination of EVs composition allows for a deep understanding of their role in the bacterial community and communication among them. Cutibacterium acnes, formerly Propionibacteriumacnes, are commensal bacteria responsible for various infections, e.g., prosthesis, sarcoidosis, soft-tissue infections, and the most known but still controversial—acnes lesion. In C. acnes, three major phylotypes represented variable disease associations. Herein, for the first time, we present a comparative analysis of EVs obtained from three C. acnes phylotypes (IA1, IB, and II) to demonstrate the existence of differences in their protein and lipid composition. In the following work, the morphological analysis of EVs was performed, and the SDS-PAGE protein profile and the lipid profile were presented using the TLC and MALDI-TOF MS methods. This study allowed us to show major differences between the protein and lipid composition of C. acnes EVs. This is a clear indication that EVs released by different phylotypes of the one species are not identical to each other in terms of composition and should be separately analyzed each time to obtain reliable results. Full article

(This article belongs to the Special Issue Membranous Nanostructures - Micro World Communication Infrastructure)

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15 pages, 1577 KiB

Open AccessArticle

Investigation of Shape Transformations of Vesicles, Induced by Their Adhesion to Flat Substrates Characterized by Different Adhesion Strength

by Jeel Raval, Aleš Iglič and Wojciech Góźdź

Int. J. Mol. Sci. 2021, 22(24), 13406; https://doi.org/10.3390/ijms222413406 - 14 Dec 2021

Cited by 2 | Viewed by 2042

Abstract

The adhesion of lipid vesicles to a rigid flat surface is investigated. We examine the influence of the membrane spontaneous curvature, adhesion strength, and the reduced volume on the stability and shape transformations of adhered vesicles. The minimal strength of the adhesion necessary to stabilize the shapes of adhered vesicles belonging to different shape classes is determined. It is shown that the budding of an adhered vesicle may be induced by the change of the adhesion strength. The importance of the free vesicle shape for its susceptibility to adhesion is discussed. Full article

(This article belongs to the Special Issue Membranous Nanostructures - Micro World Communication Infrastructure)

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23 pages, 2031 KiB

Open AccessArticle

Application of Dynamic and Static Light Scattering for Size and Shape Characterization of Small Extracellular Nanoparticles in Plasma and Ascites of Ovarian Cancer Patients

by Ksenija Kogej, Darja Božič, Borut Kobal, Maruša Herzog and Katarina Černe

Int. J. Mol. Sci. 2021, 22(23), 12946; https://doi.org/10.3390/ijms222312946 - 30 Nov 2021

Cited by 10 | Viewed by 2430

Abstract

In parallel to medical treatment of ovarian cancer, methods for the early detection of cancer tumors are being sought. In this contribution, the use of non-invasive static (SLS) and dynamic light scattering (DLS) for the characterization of extracellular nanoparticles (ENPs) in body fluids of advanced serous ovarian cancer (OC) and benign gynecological pathology (BP) patients is demonstrated and critically evaluated. Samples of plasma and ascites (OC patients) or plasma, peritoneal fluid, and peritoneal washing (BP patients) were analyzed. The hydrodynamic radius (R_h) and the radius of gyration (R_g) of ENPs were calculated from the angular dependency of LS intensity for two ENP subpopulations. R_h and R_g of the predominant ENP population of OC patients were in the range 20–30 nm (diameter 40–60 nm). In thawed samples, larger particles (R_h mostly above 100 nm) were detected as well. The shape parameter ρ of both particle populations was around 1, which is typical for spherical particles with mass concentrated on the rim, as in vesicles. The R_h and R_g of ENPs in BP patients were larger than in OC patients, with ρ ≈ 1.1–2, implying a more elongated/distorted shape. These results show that SLS and DLS are promising methods for the analysis of morphological features of ENPs and have the potential to discriminate between OC and BP patients. However, further development of the methodology is required. Full article

(This article belongs to the Special Issue Membranous Nanostructures - Micro World Communication Infrastructure)

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28 pages, 10077 KiB

Open AccessArticle

Stability of Erythrocyte-Derived Nanovesicles Assessed by Light Scattering and Electron Microscopy

by Darja Božič, Matej Hočevar, Matic Kisovec, Manca Pajnič, Ljubiša Pađen, Marko Jeran, Apolonija Bedina Zavec, Marjetka Podobnik, Ksenija Kogej, Aleš Iglič and Veronika Kralj-Iglič

Int. J. Mol. Sci. 2021, 22(23), 12772; https://doi.org/10.3390/ijms222312772 - 25 Nov 2021

Cited by 10 | Viewed by 2879

Abstract

Extracellular vesicles (EVs) are gaining increasing amounts of attention due to their potential use in diagnostics and therapy, but the poor reproducibility of the studies that have been conducted on these structures hinders their breakthrough into routine practice. We believe that a better understanding of EVs stability and methods to control their integrity are the key to resolving this issue. In this work, erythrocyte EVs (hbEVs) were isolated by centrifugation from suspensions of human erythrocytes that had been aged in vitro. The isolate was characterised by scanning (SEM) and cryo-transmission electron microscopy (cryo-TEM), flow cytometry (FCM), dynamic/static light scattering (LS), protein electrophoresis, and UV-V spectrometry. The hbEVs were exposed to various conditions (pH (4–10), osmolarity (50–1000 mOsm/L), temperature (15–60 °C), and surfactant Triton X-100 (10–500 μM)). Their stability was evaluated by LS by considering the hydrodynamic radius (R_h), intensity of scattered light (I), and the shape parameter (ρ). The morphology of the hbEVs that had been stored in phosphate-buffered saline with citrate (PBS–citrate) at 4 °C remained consistent for more than 6 months. A change in the media properties (50–1000 mOsm/L, pH 4–10) had no significant effect on the R_h (=100–130 nm). At pH values below 6 and above 8, at temperatures above 45 °C, and in the presence of Triton X-100, hbEVs degradation was indicated by a decrease in I of more than 20%. Due to the simple preparation, homogeneous morphology, and stability of hbEVs under a wide range of conditions, they are considered to be a suitable option for EV reference material. Full article

(This article belongs to the Special Issue Membranous Nanostructures - Micro World Communication Infrastructure)

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18 pages, 1968 KiB

Open AccessArticle

Single Extracellular Vesicle Analysis Performed by Imaging Flow Cytometry and Nanoparticle Tracking Analysis Evaluate the Accuracy of Urinary Extracellular Vesicle Preparation Techniques Differently

by Marvin Droste, Tobias Tertel, Stefanie Jeruschke, Robin Dittrich, Evangelia Kontopoulou, Bernd Walkenfort, Verena Börger, Peter F. Hoyer, Anja K. Büscher, Basant K. Thakur and Bernd Giebel

Int. J. Mol. Sci. 2021, 22(22), 12436; https://doi.org/10.3390/ijms222212436 - 18 Nov 2021

Cited by 26 | Viewed by 4316

Abstract

Small extracellular vesicles isolated from urine (uEVs) are increasingly recognized as potential biomarkers. Meanwhile, different uEV preparation strategies exist. Conventionally, the performance of EV preparation methods is evaluated by single particle quantification, Western blot, and electron microscopy. Recently, we introduced imaging flow cytometry (IFCM) as a next-generation single EV analysis technology. Here, we analyzed uEV samples obtained with different preparation procedures using nanoparticle tracking analysis (NTA), semiquantitative Western blot, and IFCM. IFCM analyses demonstrated that urine contains a predominant CD9⁺ sEV population, which exceeds CD63⁺ and CD81⁺ sEV populations. Furthermore, we demonstrated that the storage temperature of urine samples negatively affects the recovery of CD9⁺ sEVs. Although overall reduced, the highest CD9⁺ sEV recovery was obtained from urine samples stored at −80 °C and the lowest from those stored at −20 °C. Upon comparing the yield of the different uEV preparations, incongruencies between NTA and IFCM data became apparent. Results obtained by both NTA and IFCM were consistent with Western blot analyses for EV marker proteins; however, NTA results correlated with the amount of the impurity marker uromodulin. Despite demonstrating that the combination of ultrafiltration and size exclusion chromatography appears as a reliable uEV preparation technique, our data challenge the soundness of traditional NTA for the evaluation of different EV preparation methods. Full article

(This article belongs to the Special Issue Membranous Nanostructures - Micro World Communication Infrastructure)

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19 pages, 2857 KiB

Open AccessArticle

The Effect of the Osmotically Active Compound Concentration Difference on the Passive Water and Proton Fluxes across a Lipid Bilayer

by Magdalena Przybyło, Dominik Drabik, Joanna Doskocz, Aleš Iglič and Marek Langner

Int. J. Mol. Sci. 2021, 22(20), 11099; https://doi.org/10.3390/ijms222011099 - 14 Oct 2021

Cited by 5 | Viewed by 2131

Abstract

The molecular details of the passive water flux across the hydrophobic membrane interior are still a matter of debate. One of the postulated mechanisms is the spontaneous, water-filled pore opening, which facilitates the hydrophilic connection between aqueous phases separated by the membrane. In the paper, we provide experimental evidence showing that the spontaneous lipid pore formation correlates with the membrane mechanics; hence, it depends on the composition of the lipid bilayer and the concentration of the osmotically active compound. Using liposomes as an experimental membrane model, osmotically induced water efflux was measured with the stopped-flow technique. Shapes of kinetic curves obtained at low osmotic pressure differences are interpreted in terms of two events: the lipid pore opening and water flow across the aqueous channel. The biological significance of the dependence of the lipid pore formation on the concentration difference of an osmotically active compound was illustrated by the demonstration that osmotically driven water flow can be accompanied by the dissipation of the pH gradient. The application of the Helfrich model to describe the probability of lipid pore opening was validated by demonstrating that the probability of pore opening correlates with the membrane bending rigidity. The correlation was determined by experimentally derived bending rigidity coefficients and probabilities of lipid pores opening. Full article

(This article belongs to the Special Issue Membranous Nanostructures - Micro World Communication Infrastructure)

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Review

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20 pages, 2481 KiB

Open AccessReview

Basic Methods for Preparation of Liposomes and Studying Their Interactions with Different Compounds, with the Emphasis on Polyphenols

by Luka Šturm and Nataša Poklar Ulrih

Int. J. Mol. Sci. 2021, 22(12), 6547; https://doi.org/10.3390/ijms22126547 - 18 Jun 2021

Cited by 71 | Viewed by 12860

Abstract

Studying the interactions between lipid membranes and various bioactive molecules (e.g., polyphenols) is important for determining the effects they can have on the functionality of lipid bilayers. This knowledge allows us to use the chosen compounds as potential inhibitors of bacterial and cancer cells, for elimination of viruses, or simply for keeping our healthy cells in good condition. As studying those effect can be exceedingly difficult on living cells, model lipid membranes, such as liposomes, can be used instead. Liposomal bilayer systems represent the most basic platform for studying those interactions, as they are simple, quite easy to prepare and relatively stable. They are especially useful for investigating the effects of bioactive compounds on the structure and kinetics of simple lipid membranes. In this review, we have described the most basic methods available for preparation of liposomes, as well as the essential techniques for studying the effects of bioactive compounds on those liposomes. Additionally, we have provided details for an easy laboratory implementation of some of the described methods, which should prove useful especially to those relatively new on this research field. Full article

(This article belongs to the Special Issue Membranous Nanostructures - Micro World Communication Infrastructure)

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