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Buzás Edit I

az alábbi absztraktok szerzői között szerepel:

Kovács Dóra Kinga
Nanoinjection of fluorescent nanoparticles to single live cells by robotic fluidic force microscopy

Aug 29 - kedd

12:15 – 12:30

Modern biofizikai módszerek

E13

Nanoinjection of fluorescent nanoparticles to single live cells by robotic fluidic force microscopy

Kinga Dóra Kovács^1,*, Tamás Visnovitz^2,3,*, Tamás Gerecsei¹, Beatrix Peter¹, Sándor Kurunczi¹, Anna Koncz², Krisztina Németh², Dorina Lenzinger², Krisztina V. Vukman², Péter Lőrincz⁴, Inna Székács¹, Edit I. Buzás^2,5,6**, Robert Horvath^1,**

¹ Nanobiosensorics Laboratory, Centre of Energy Research, ELKH, Budapest, Hungary

² Department of Genetics, Cell- and Immunobiology, Semmelweis University, Budapest, Hungary

³ Department of Plant Physiology and Molecular Plant Biology, ELTE Eötvös Loránd University, Budapest, Hungary

⁴ Department of Anatomy, Cell and Developmental Biology, ELTE Eötvös Loránd University, Budapest, Hungary

⁵ HCEMM-SU Extracellular Vesicle Research Group, Budapest, Hungary

⁶ ELKH-SE Translational Extracellular Vesicle Research Group, Budapest, Hungary

*,** equal contributions / **corresponding authors

Direct injection of fluorescent nanoparticles into the cytoplasm of living cells can provide new insights into the intracellular fate of various different fluorescently labelled biologically active particles. Here we used fluorescent nanoparticles to prove the feasibility of nanoinjection into single live HeLa cells by using robotic fluidic force microscopy (FluidFM). This injection platform offers the advantage of high cell selectivity and efficiency. We confirmed the successful injection of both GFP encoding plasmids and GFP tagged fluorescent nanoparticles to the cells by confocal microscopy. We were able track the nanoparticles in the living cells for 20 hours. The injected nanoparticles were initially localized in concentrated spot-like regions within the cytoplasm. Later, they were transported towards the periphery of the cells. Based on our proof-of-principle data, the FluidFM platform is suitable for targeting single living cells by fluorescently labelled biologically active particles and may lead to information about the intracellular cargo delivery at a single-cell level.

Tasvilla Sonallya
Systematic investigation and classification of host defence and cell penetrating peptides based on their affinity for interaction with extracellular vesicles

Aug 30 - szerda

09:55 – 10:10

Membránok és membránfehérjék biofizikája

E24

Systematic investigation and classification of host defence and cell penetrating peptides based on their affinity for interaction with extracellular vesicles.

Tasvilla Sonallya^1,2, Imola Cs. Szigyártó¹ , Tünde Juhász¹, Edit I. Buzas^4,5,6 Delaram Khamari ⁴,Kinga Ilyes^2,3 Zoltán Varga³, and Tamás Beke-Somfai^1*

¹Institute of Materials and Environmental Chemistry, Biomolecular Self-assembly Research Group, Research Centre for Natural Sciences, Budapest H-1117, Magyar tudósok körútja 2, Hungary,

²Hevesy György PhD School of Chemistry, ELTE Eötvös Loránd University, Budapest H-1117, Pázmány Péter sétány 1/A, Hungary.

³Institute of Materials and Environmental Chemistry, Biological Nanochemistry Research Group, Research Centre for Natural Sciences, Budapest, H-1117, Magyar tudósok körútja 2, Hungary.

⁴Department of Genetics, Cell and Immunobiology, Semmelweis University, Budapest, Hungary

⁵HCEMM Extracellular Vesicle Research Group, Semmelweis University, Budapest, Hungary

⁶ELKH-SE Immune-Proteogenomics Extracellular Vesicle Research Group, Budapest, Hungary.

Host defence peptides (HDP) are promising biomaterials with antimicrobial and anticancer applications. By disturbing or lysing the cell membrane, they carry out their biological role. These peptides show numerous types of membrane interaction mechanisms i.e., carpet, toroidal pore, and barrel stave. Cell penetrating peptide find application in cargo loading and uptake of small molecules and nanoparticles. The interactive mechanism of these peptides has been studied widely with model membranes however our knowledge with extracellular vesicles (EV) is scarce. There are several aspects where EV – HDP interactions could be relevant, ranging from cooperative presence on infection sites functions to EV cargo loading. Hence, based on their in-depth investigation using biophysical techniques, the binding affinity with extracellular vesicles was studied and categorised as low binding affinity, medium binding affinity and high affinity. This initial categorisation gives further insight into its specific interactive mechanism.

Acknowledgment:

This work was funded by the Ministry of Innovation and Technology of Hungary through the National Research, Development and Innovation Office, financed under the TKP2021-EGA-31, the 2020-1-1-2-PIACI-KFI_2020-00021, 2019-2.1.11-TÉT-2019-00091 and KKP_22 project no. 144180. Support from Eötvös Loránd Research Network, grant no. SA-87/2021 and KEP-5/2021 is also acknowledged. Z.V. was supported by the János Bolyai Research Scholarship of the Hungarian Academy of Sciences.

References

1. Singh, P. et al. Removal and identification of external protein corona members from RBC‐derived extracellular vesicles by surface manipulating antimicrobial peptides. J. Extracell. Biol. 2, (2023).

2. Singh, P. et al. Membrane Active Peptides Remove Surface Adsorbed Protein Corona From Extracellular Vesicles of Red Blood Cells. Front. Chem. 8, (2020).

Varga Zoltán
In vivo Biodistribution of Extracellular Vesicles: Developing Efficient Radiolabeling Techniques

Aug 29 - kedd

14:00 – 14:20

Orvosi biofizika és sugárbiológia

E14

In vivo Biodistribution of Extracellular Vesicles: Developing Efficient Radiolabeling Techniques

Zoltán Varga¹, Kinga Ilyés¹, Dávid Szöllősi², Ildikó Horváth², Domokos Máthé^2,3, Krisztina Németh^4,5, Viola Tamási^4,*, Edit I Buzás^4,5,6, Krisztián Szigeti²

¹ Biological Nanochemistry Research Group, Research Centre for Natural Sciences

² Department of Biophysics and Radiation Biology, Semmelweis University

³ HCMM-SE In Vivo Imaging Advanced Core Facility

⁴ Department of Genetics, Cell- and Immunobiology, Semmelweis University

⁵ ELKH-SE Translational Extracellular Vesicle Research Group

⁶ HCMM-SE Extracellular Vesicle Research Group

^*current affiliation: Department of Molecular Biology, Semmelweis University

The understanding of extracellular vesicle (EV) biodistribution plays an important role in advancing circulating biomarker research. Nuclear imaging techniques like single-photon emission computed tomography (SPECT) hold potential, but the literature on radioisotope labeling of EVs for in vivo studies remains scarce. This presentation explores the evolution of novel radiolabeling methods, focusing on the development and comparative evaluation of various Tc99m radiolabeling strategies.

Our initial method involved the radioisotope labeling of erythrocyte-derived EVs using the Tc99m-tricarbonyl complex [1]. In vivo SPECT/CT biodistribution studies in mice showed that intravenously administered Tc99m-labeled EVs primarily accumulated in the liver and spleen. Our observations suggested good in vivo stability, with a minor fraction of the radioactive label detaching from the EVs. Next, we explored an alternative approach using Tc99m-HYNIC-Duramycin to label cell-derived EVs [2]. Duramycin, a membrane-active peptide, specifically labels EVs, resulting in higher labeling efficiency. Following previous observations, significant uptake of EVs in the liver and the spleen was observed.

The latest experimental focus is on the use of recombinant proteins with His-tag in conjunction with the Tc99m-tricarbonyl complex to label EVs. Preliminary investigations indicate promising superior performance with this method compared to the previous techniques.

This presentation will provide an in-depth comparison of these methods, emphasizing their development process and potential implications for advancing in vivo EV imaging studies. Through a critical evaluation of their advantages and potential limitations, we aim to foster a greater understanding of efficient tracking of EV biodistribution for future research.

References

[1] Varga Z et al. (2016) Cancer Biother. Radiopharm. 31: 168-173.

[2] Németh K et al. (2021) Cell Mol Life Sci 78: 7589–7604