Opazovanje zgodnjih napak pri delitvi pljučnih epitelijskih celic po izpostavitvi različnim nanodelcem

Vencelj, Ana (author), Urbančič, Iztok (mentor), Podlipec, Rok (comentor)

Nanodelci so delci s premerom manjšim od 100 nm v vsaj eni dimenziji in so zaradi svoje majhnosti široko uporabljeni. Vse več biološko obstojnih nanomaterialov z visokim razmerjem površine glede na njihov volumen kaže izrazite patogene, predvsem genotoksične učinke, ki se kažejo z vnetjem, fibrozo in rakom. Genotoksični učinki in morebitni nastanek raka so odvisni od oblike, velikosti in vrste nanodelcev, vendar njihov vpliv na nastanek raka še ni povsem razjasnjen. Med rakotvorne snovi so že uvrščena azbestna vlakna, medtem ko pri nanodelcih titanovega dioksida in nanodelcih na osnovi ogljika patološki učinki na biološke sisteme še niso dokončno opredeljeni. Ugotovljeno je bilo, da so prve zgodnje napake, ki jih lahko povzročijo azbestna vlakna, nastanek mikrojeder in večjedrnih celic, kar je povezano z genomsko nestabilnostjo in posledično z razvojem anevploidije – nenormalnega števila kromosomov v celici. V namen ugotavljanja zgodnjih sprememb pri delitvi pljučnih epitelijskih celic smo celice izpostavili različnim nanodelcem, in sicer nanodelcem titanovega dioksida v obliki nanocevk, nanodelcem titanovega dioksida v obliki nanokock in večplastnim ogljikovim nanocevkam, ter spremembe opazovali s konfokalnim fluorescenčnim mikroskopom tekom večurnega in večdnevnega časovnega eksperimenta na živih celicah. Z eksperimenti smo pri celicah, izpostavljenih nanodelcem, pokazali povečano število večjedrnih celic in celic z mikrojedri, katere je hkrati spremljala še deformacija jedra, pri celicah, izpostavljenih nanodelcem. Prav tako smo opazili podaljšan čas mitoze pri celicah, ki so bile izpostavljene nanodelcem titanovega dioksida v obliki nanocevk in večplastnim ogljikovim nanocevkam, kar nakazuje na nepravilnosti v procesu celične delitve. Vse opažene nenormalnosti povezujemo z okvaro citokineze, zadnjo stopnjo celične delitve, v kateri se primarna celica dokončno razdeli na dve hčerinski celici. Kljub temu da rezultati niso nedvoumno potrdili, da so opažene patologije izključno posledica nanodelcev, predvidevamo, da lahko določeni nanodelci vplivajo in spremenijo ključne celične procese in njene morfološke lastnosti. Z in vitro eksperimenti smo pokazali tudi raznolikost vpliva nanodelcev z različnimi fizikalno-kemijskimi lastnostmi na celice, kar zahteva nadaljnje raziskave za boljše razumevanje mehanizmov interakcije na celični ravni.

• Language: Slovenian
• Keywords: nanodelci, mitoza, napake citokineze, mikrojedra, konfokalna fluorescenčna mikroskopija
• Work type: Master's thesis/paper
• Organization: FFA - Faculty of Pharmacy
• Year: 2024
• PID: 20.500.12556/RUL-162369
• Publication date in RUL: 21.09.2024

VENCELJ, Ana, 2024, Opazovanje zgodnjih napak pri delitvi pljučnih epitelijskih celic po izpostavitvi različnim nanodelcem [online]. Master’s thesis. [Accessed 7 April 2025]. Retrieved from: https://repozitorij.uni-lj.si/IzpisGradiva.php?lang=eng&id=162369

Secondary language

• Language: English
• Title: Observation of early defects in lung epithelial cell division after exposure to different nanoparticles

Abstract

Nanoparticles are particles with a diameter of less than 100 nm in at least one dimension and are widely used because of their small size. An increasing number of bio-resistant nanomaterials with a high surface area to volume ratio are exhibiting pronounced pathogenic, mainly genotoxic effects, manifesting in inflammation, fibrosis and cancer. The genotoxic effects and potential cancer formation depend on the shape, size and type of nanoparticles, but their impact on cancer formation is not yet fully understood. Asbestos fibres are already classified as carcinogens, while the pathological effects on biological systems of titanium dioxide nanoparticles and carbon-based nanoparticles have not yet been definitively identified. It has been reported that the first early defects that can be caused by asbestos fibres are the formation of micronuclei and multinucleated cells, which are associated with genomic instability and, consequently, the development of aneuploidy – an abnormal number of chromosomes in a cell. To detect early changes in lung epithelial cell division, we exposed cells to different nanoparticles, namely titanium dioxide nanoparticles in the form of nanotubes, titanium dioxide nanoparticles in the form of nanocubes and multiwalled carbon nanotubes, and observed the changes with a confocal fluorescence microscope over several hours and several days in a time-course experiment on live cells. The experiments showed an increased number of multinucleated and micronucleated cells in cells exposed to nanoparticles, which were accompanied by nuclear deformation in cells exposed to nanoparticles. We also observed prolonged mitosis time in cells exposed to nanoparticles of titanium dioxide nanotubes and multiwalled carbon nanotubes, suggesting abnormalities in the cell division process. All the abnormalities observed are associated with a defect in cytokinesis, the final stage of cell division in which the primary cell finally divides into two daughter cells. Although the results did not unequivocally confirm that the observed pathologies are exclusively due to nanoparticles, we hypothesise that certain nanoparticles may influence and alter key cellular processes and morphological properties. In vitro experiments have also shown the diversity of the impact of nanoparticles with different physicochemical properties on cells, which requires further research to better understand the mechanisms of interaction at the cellular level.

• Keywords: nanoparticles, mitosis, cytokinesis defects, micronuclei, confocal fluorescence microscopy