In recent years, there has been a growing interest in plant extracellular vesicles (pEVs) due to their immense potential for medical applications, particularly as carriers for drug delivery. To use the benefits of pEVs in the future, it is necessary to identify methods that facilitate their production in sufficient quantities while maintaining high quality. In this study, a comparative analysis of yields of tobacco pEV derived from apoplastic fluid, sterile calli, and suspension cultures, was performed to identify the most suitable plant material for vesicle isolation. Subsequent experiments focused on assessing the efficiency of small interfering RNA (siRNA) loading into callus-derived vesicles, employing various methods such as sonication, incubation, incubation supplemented with saponin, lipofection, and electroporation. Differences in loading efficiency among vesicles derived from apoplastic fluid, calli, and suspension cultures were observed. Moreover, our investigation extended to the presence of tobacco secondary metabolites, specifically anabasine and nicotine, within vesicles originating from three distinct tobacco sources. The outcomes of our study highlight variations not only in vesicle yields based on their source but also in their loadability and the presence of nicotine and anabasine. These findings contribute valuable insights into optimizing the production and application of pEVs for future medicinal purposes.

Centre for Nanomaterials and Biotechnologies Faculty of Science Jan Evangelista Purkyně University in Ústí Nad Labem Ústí Nad Labem Czech Republic

Department of Environmental Chemistry and Technology Faculty of Environment Jan Evangelista Purkyně University in Ústí Nad Labem Ústí Nad Labem Czech Republic

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Di Gioia, S., Hossain, M. N. & Conese, M. Biological properties and therapeutic effects of plant-derived nanovesicles. PubMed DOI PMC

Mu, N. et al. Plant-derived exosome-like nanovesicles: Current progress and prospects. PubMed DOI PMC

Movahed, N. et al. Turnip mosaic virus components are released into the extracellular space by vesicles in infected leaves. PubMed DOI PMC

Cai, Q. PubMed PMC

Regente, M. et al. Plant extracellular vesicles are incorporated by a fungal pathogen and inhibit its growth. PubMed DOI

Liegertová, M. et al. Mucus - derived exosome - like vesicles from the Spanish slug ( PubMed DOI PMC

Woith, E., Fuhrmann, G. & Melzig, M. F. Extracellular vesicles—Connecting kingdoms. PubMed DOI PMC

Nemati, M. et al. Plant - derived extracellular vesicles: A novel nanomedicine approach with advantages and challenges. PubMed DOI PMC

Chen, Y. & Cai, Q. Plant exosome-like nanovesicles and their role in the innovative delivery of RNA therapeutics. PubMed DOI PMC

Karamanidou, T. & Tsouknidas, A. Plant-derived extracellular vesicles as therapeutic nanocarriers. PubMed PMC

Dad, H. et al. Plant exosome-like nanovesicles: emerging therapeutics and drug delivery nanoplatforms. PubMed DOI PMC

Woith, E. et al. Plant extracellular vesicles and nanovesicles: Focus on secondary metabolites, proteins and lipids with perspectives on their potential and sources. PubMed DOI PMC

Rutter, B. D. & Innes, R. W. Extracellular vesicles isolated from the leaf apoplast carry stress-response proteins. PubMed DOI PMC

de la Canal, L. & Pinedo, M. Extracellular vesicles: A missing component in plant cell wall remodeling. PubMed DOI PMC

Weiberg, A. PubMed PMC

Roth, R. et al. Arbuscular cell invasion coincides with extracellular vesicles and membrane tubules. PubMed DOI

Garaeva, L. et al. Delivery of functional exogenous proteins by plant-derived vesicles to human cells in vitro. PubMed DOI PMC

Sundaram, K. PubMed PMC

Teng, Y. et al. Grapefruit-derived nanovectors deliver miR-18a for treatment of liver metastasis of colon cancer by induction of M1 macrophages. PubMed DOI PMC

Wang, Q. et al. Delivery of therapeutic agents by nanoparticles made of grapefruit-derived lipids. PubMed DOI PMC

Tian, Y. et al. A doxorubicin delivery platform using engineered natural membrane vesicle exosomes for targeted tumor therapy. PubMed DOI

Kocholata, M., Maly, J., Martinec, J. & Malinska, H. A. Plant extracellular vesicles and their potential in human health research, the practical approach. PubMed DOI PMC

Alzahrani, F. A., Khan, M. I., Kameli, N., Alsahafi, E. & Riza, Y. M. Plant-derived extracellular vesicles and their exciting potential as the future of next-generation drug delivery. PubMed DOI PMC

Zeng, H. et al. Current strategies for exosome cargo loading and targeting delivery. PubMed DOI PMC

Deng, Z. et al. Broccoli-derived nanoparticle inhibits mouse colitis by activating dendritic cell AMP-activated protein kinase. PubMed DOI PMC

Zhuang, X. PubMed PMC

Berger, E. et al. Use of nanovesicles from orange juice to reverse diet-induced gut modifications in diet-induced obese mice. PubMed DOI PMC

E, Z. PubMed PMC

Dini, L. et al. Microvesicles and exosomes in metabolic diseases and inflammation. PubMed DOI

Rutter, B., Rutter, K. & Innes, R. Isolation and quantification of plant extracellular vesicles. PubMed DOI PMC

Stanly, C. PubMed PMC

Kocholata, M., Prusova, M., Auer Malinska, H., Maly, J. & Janouskova, O. Comparison of two isolation methods of tobacco-derived extracellular vesicles, their characterization and uptake by plant and rat cells. PubMed PMC

Nagata, T. When I encountered tobacco BY-2 cells!. DOI

Srba, M., Černíková, A., Opatrný, Z. & Fischer, L. Practical guidelines for the characterization of tobacco BY-2 cell lines. DOI

Suhas Shinde, S.

Kim, J., Li, S., Zhang, S. & Wang, J. Plant-derived exosome-like nanoparticles and their therapeutic activities. PubMed PMC

Wang, P. et al. Exosomes from M1-polarized macrophages enhance paclitaxel antitumor activity by activating macrophages-mediated inflammation. PubMed DOI PMC

Didiot, M. C. et al. Exosome-mediated delivery of hydrophobically modified siRNA for huntingtin mRNA silencing. PubMed DOI PMC

Thakur, A. et al. Inhibition of glioma cells’ proliferation by doxorubicin-loaded exosomes via microfluidics. PubMed DOI PMC

Fuhrmann, G., Serio, A., Mazo, M., Nair, R. & Stevens, M. M. Active loading into extracellular vesicles significantly improves the cellular uptake and photodynamic effect of porphyrins. PubMed DOI

Cui, Y., Gao, J., He, Y. & Jiang, L. Plant extracellular vesicles. PubMed DOI

Yi, Q. et al. Current understanding of plant-derived exosome-like nanoparticles in regulating the inflammatory response and immune system microenvironment. PubMed DOI

Sun, L. et al. A 3D culture system improves the yield of MSCs-derived extracellular vesicles and enhances their therapeutic efficacy for heart repair. PubMed DOI

Yousafzai, N. A., El Khalki, L., Wang, W., Szpendyk, J. & Sossey-Alaoui, K. Advances in 3D culture models to study exosomes in triple-negative breast cancer. PubMed DOI PMC

Yugay, Y. PubMed PMC

Kumari, A. et al. Metabolomic homeostasis shifts after callus formation and shoot regeneration in tomato. PubMed DOI PMC

Joshi, B. S., Ortiz, D. & Zuhorn, I. S. Converting extracellular vesicles into nanomedicine: Loading and unloading of cargo. DOI

El-Andaloussi, S. et al. Exosome-mediated delivery of siRNA in vitro and in vivo. PubMed DOI

Kooijmans, S. A. A. et al. Electroporation-induced siRNA precipitation obscures the efficiency of siRNA loading into extracellular vesicles. PubMed DOI

Wahlgren, J. PubMed PMC

Lamichhane, T. N. et al. Oncogene knockdown via active loading of small RNAs into extracellular vesicles by sonication. PubMed DOI PMC

Guo, S. et al. Intranasal delivery of mesenchymal stem cell derived exosomes loaded with phosphatase and tensin homolog siRNA repairs complete spinal cord injury. PubMed DOI

Nizamudeen, Z. A. PubMed PMC

Khare, P. et al. Development of lipidoid nanoparticles for siRNA delivery to neural cells. PubMed DOI PMC

Nowak, M. et al. Extracellular vesicles as drug transporters extracellular vesicles as drug transporters. PubMed DOI PMC

Han, Y. et al. Overview and update on methods for cargo loading into extracellular vesicles. PubMed DOI PMC

Wang, B. et al. Targeted drug delivery to intestinal macrophages by bioactive nanovesicles released from grapefruit. PubMed DOI PMC

Dou, W. et al. Protective effect of naringenin against experimental colitis via suppression of Toll-like receptor 4/NF-κB signalling. PubMed DOI PMC

Perut, F. et al. Strawberry-derived exosome-like nanoparticles prevent oxidative stress in human mesenchymal stromal cells. PubMed DOI PMC

Piñol, M. T., Palazon, J. & Serrano, M. Growth and nicotine content of tobacco callus cultures without organogenesis. DOI

Tiburcio, A. F. & Galston, A. W. Analysis of alkaloids in tobacco callus by HPLC. DOI

Shoji, T. & Hashimoto, T. Why does anatabine, but not nicotine, accumulate in jasmonate-elicited cultured tobacco BY-2 cells?. PubMed DOI