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Cellular Distribution and Ultrastructural Changes in HaCaT Cells, Induced by Podophyllotoxin and Its Novel Fluorescent Derivative, Supported by the Molecular Docking Studies.

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العنوان: Cellular Distribution and Ultrastructural Changes in HaCaT Cells, Induced by Podophyllotoxin and Its Novel Fluorescent Derivative, Supported by the Molecular Docking Studies.
المؤلفون: Strus P; Department of Histology and Embryology, Medical University of Warsaw, Chałubińskiego 5, 02-004 Warsaw, Poland., Sadowski K; Students Scientific Group HESA, Department of Histology and Embryology, Medical University of Warsaw, Chałubinskiego 5, 02-004 Warsaw, Poland., Kostro J; Students Scientific Group HESA, Department of Histology and Embryology, Medical University of Warsaw, Chałubinskiego 5, 02-004 Warsaw, Poland., Szczepankiewicz AA; Laboratory of Electron Microscopy, Nencki Institute of Warsaw, Pasteura 3, 02-093 Warsaw, Poland., Nieznańska H; Laboratory of Electron Microscopy, Nencki Institute of Warsaw, Pasteura 3, 02-093 Warsaw, Poland., Niedzielska M; Laboratory of Natural Products Chemistry, Faculty of Chemistry, University of Warsaw, Pasteura 1, 02-093 Warsaw, Poland., Zlobin A; Laboratory of Natural Products Chemistry, Faculty of Chemistry, University of Warsaw, Pasteura 1, 02-093 Warsaw, Poland., Nawar Ra'idah P; Laboratory of Natural Products Chemistry, Faculty of Chemistry, University of Warsaw, Pasteura 1, 02-093 Warsaw, Poland., Molęda Z; Laboratory of Natural Products Chemistry, Faculty of Chemistry, University of Warsaw, Pasteura 1, 02-093 Warsaw, Poland., Szawkało J; Laboratory of Natural Products Chemistry, Faculty of Chemistry, University of Warsaw, Pasteura 1, 02-093 Warsaw, Poland., Czarnocki Z; Laboratory of Natural Products Chemistry, Faculty of Chemistry, University of Warsaw, Pasteura 1, 02-093 Warsaw, Poland., Wójcik C; Amgen Inc., Thousand Oaks, CA 91320, USA.; Department of Undergraduate Medical Education, OHSU School of Medicine, Portland, OR 97239, USA., Szeleszczuk Ł; Department of Organic and Physical Chemistry, Faculty of Pharmacy, Medical University of Warsaw, Banacha 1, 02-093 Warsaw, Poland., Młynarczuk-Biały I; Department of Histology and Embryology, Medical University of Warsaw, Chałubińskiego 5, 02-004 Warsaw, Poland.
المصدر: International journal of molecular sciences [Int J Mol Sci] 2024 May 29; Vol. 25 (11). Date of Electronic Publication: 2024 May 29.
نوع المنشور: Journal Article
اللغة: English
بيانات الدورية: Publisher: MDPI Country of Publication: Switzerland NLM ID: 101092791 Publication Model: Electronic Cited Medium: Internet ISSN: 1422-0067 (Electronic) Linking ISSN: 14220067 NLM ISO Abbreviation: Int J Mol Sci Subsets: MEDLINE
أسماء مطبوعة: Original Publication: Basel, Switzerland : MDPI, [2000-
مواضيع طبية MeSH: Molecular Docking Simulation* , Podophyllotoxin*/analogs & derivatives , Podophyllotoxin*/pharmacology , Podophyllotoxin*/chemistry , HaCaT Cells* , Tubulin*/metabolism , Keratinocytes*/drug effects , Keratinocytes*/metabolism, Humans ; Cell Survival/drug effects ; Mitochondria/drug effects ; Mitochondria/metabolism ; Mitochondria/ultrastructure ; Fluorescent Dyes/chemistry ; Binding Sites ; Endoplasmic Reticulum Stress/drug effects
مستخلص: Podophyllotoxin (PPT) is an active pharmaceutical ingredient (API) with established antitumor potential. However, due to its systemic toxicity, its use is restricted to topical treatment of anogenital warts. Less toxic PPT derivatives (e.g., etoposide and teniposide) are used intravenously as anticancer agents. PPT has been exploited as a scaffold of new potential therapeutic agents; however, fewer studies have been conducted on the parent molecule than on its derivatives. We have undertaken a study of ultrastructural changes induced by PPT on HaCaT keratinocytes. We have also tracked the intracellular localization of PPT using its fluorescent derivative (PPT-FL). Moreover, we performed molecular docking of both PPT and PPT-FL to compare their affinity to various binding sites of tubulin. Using the Presto blue viability assay, we established working concentrations of PPT in HaCaT cells. Subsequently, we have used selected concentrations to determine PPT effects at the ultrastructural level. Dynamics of PPT distribution by confocal microscopy was performed using PPT-FL. Molecular docking calculations were conducted using Glide. PPT induces a time-dependent cytotoxic effect on HaCaT cells. Within 24 h, we observed the elongation of cytoplasmic processes, formation of cytoplasmic vacuoles, progressive ER stress, and shortening of the mitochondrial long axis. After 48 h, we noticed disintegration of the cell membrane, progressive vacuolization, apoptotic/necrotic vesicles, and a change in the cell nucleus's appearance. PPT-FL was detected within HaCaT cells after ~10 min of incubation and remained within cells in the following measurements. Molecular docking confirmed the formation of a stable complex between tubulin and both PPT and PPT-FL. However, it was formed at different binding sites. PPT is highly toxic to normal human keratinocytes, even at low concentrations. It promptly enters the cells, probably via endocytosis. At lower concentrations, PPT causes disruptions in both ER and mitochondria, while at higher concentrations, it leads to massive vacuolization with subsequent cell death. The novel derivative of PPT, PPT-FL, forms a stable complex with tubulin, and therefore, it is a useful tracker of intracellular PPT binding and trafficking.
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معلومات مُعتمدة: MEiN/2022/DIR/3155 Polish Ministry for Education and Science; 1M15/1/MB/N/22-23 to PS Medical University of Warsaw
فهرسة مساهمة: Keywords: HaCaT ultrastructure; cell death; human keratinocytes; podophyllotoxin; podophyllotoxin docking to tubulin; podophyllotoxin with attached fluorescein
المشرفين على المادة: L36H50F353 (Podophyllotoxin)
0 (Tubulin)
0 (Fluorescent Dyes)
تواريخ الأحداث: Date Created: 20240619 Date Completed: 20240619 Latest Revision: 20240620
رمز التحديث: 20240620
مُعرف محوري في PubMed: PMC11172492
DOI: 10.3390/ijms25115948
PMID: 38892135
قاعدة البيانات: MEDLINE
الوصف
تدمد:1422-0067
DOI:10.3390/ijms25115948