Resistant cancer phenotype is a key obstacle in the successful therapy of prostate cancer. The primary aim of our study was to explore resistance mechanisms in the advanced type of prostate cancer cells (PC-3) and to clarify the role of autophagy in these processes. We performed time-lapse experiment (48 hours) with ROS generating plumbagin by using multimodal holographic microscope. Furthermore, we also performed the flow-cytometric analysis and the qRT-PCR gene expression analysis at 12 selected time points. TEM and confocal microscopy were used to verify the results. We found out that autophagy (namely mitophagy) is an important resistance mechanism. The major ROS producing mitochondria were coated by an autophagic membrane derived from endoplasmic reticulum and degraded. According to our results, increasing ROS resistance may be also accompanied by increased average cell size and polyploidization, which seems to be key resistance mechanism when connected with an escape from senescence. Many different types of cell-cell interactions were recorded including entosis, vesicular transfer, eating of dead or dying cells, and engulfment and cannibalism of living cells. Entosis was disclosed as a possible mechanism of polyploidization and enabled the long-term survival of cancer cells. Significantly reduced cell motility was found after the plumbagin treatment. We also found an extensive induction of pluripotency genes expression (NANOG, SOX2, and POU5F1) at the time-point of 20 hours. We suppose, that overexpression of pluripotency genes in the portion of prostate tumour cell population exposed to ROS leads to higher developmental plasticity and capability to faster respond to changes in the extracellular environment that could ultimately lead to an alteration of cell fate.
- MeSH
- analýza hlavních komponent MeSH
- autofagie účinky léků MeSH
- buněčná sebeobnova * účinky léků MeSH
- časosběrné zobrazování MeSH
- endoplazmatické retikulum účinky léků metabolismus MeSH
- entóza účinky léků MeSH
- inhibiční koncentrace 50 MeSH
- lidé MeSH
- metastázy nádorů MeSH
- mezibuněčná komunikace účinky léků MeSH
- mitofagie účinky léků MeSH
- nádorové buněčné linie MeSH
- nádory prostaty genetika patologie MeSH
- naftochinony farmakologie MeSH
- oxidační stres * účinky léků MeSH
- průtoková cytometrie MeSH
- reaktivní formy kyslíku metabolismus MeSH
- regulace genové exprese u nádorů účinky léků MeSH
- stanovení celkové genové exprese MeSH
- velikost buňky účinky léků MeSH
- viabilita buněk účinky léků MeSH
- Check Tag
- lidé MeSH
- mužské pohlaví MeSH
- Publikační typ
- časopisecké články MeSH
- práce podpořená grantem MeSH
The aim of this work was to study the effect of Se(+VI) on viability, cell morphology, and selenomethionine accumulation of the green alga Chlorella sorokiniana grown in batch cultures. Culture exposed to sublethal Se concentrations of 40 mg · L(-1) (212 μM) decreased growth rates for about 25% compared to control. A selenate EC50 value of 45 mg · L(-1) (238.2 μM) was determined. Results showed that chlorophyll and carotenoids contents were not affected by Se exposure, while oxygen evolution decreased by half. Ultrastructural studies revealed granular stroma, fingerprint-like appearance of thylakoids which did not compromise cell activity. Unlike control cultures, SDS PAGE electrophoresis of crude extracts from selenate-exposed cell cultures revealed appearance of a protein band identified as 53 kDa Rubisco large subunit of Chlorella sorokiniana, suggesting that selenate affects expression of the corresponding chloroplast gene as this subunit is encoded in the chloroplast DNA. Results revealed that the microalga was able to accumulate up to 140 mg · kg(-1) of SeMet in 120 h of cultivation. This paper shows that Chlorella sorokiniana biomass can be enriched in the high value aminoacid SeMet in batch cultures, while keeping photochemical viability and carbon dioxide fixation activity intact, if exposed to suitable sublethal concentrations of Se.
- MeSH
- bioreaktory mikrobiologie MeSH
- Chlorella cytologie účinky léků fyziologie MeSH
- kyselina selenová aplikace a dávkování MeSH
- proliferace buněk účinky léků fyziologie MeSH
- selenomethionin izolace a purifikace metabolismus MeSH
- techniky vsádkové kultivace metody MeSH
- velikost buňky účinky léků MeSH
- viabilita buněk účinky léků fyziologie MeSH
- vztah mezi dávkou a účinkem léčiva MeSH
- Publikační typ
- časopisecké články MeSH
Recently, we have identified two astrocytic subpopulations in the cortex of GFAP-EGFP mice, in which the astrocytes are visualized by the enhanced green-fluorescent protein (EGFP) under the control of the human glial fibrillary acidic protein (GFAP) promotor. These astrocytic subpopulations, termed high response- (HR-) and low response- (LR-) astrocytes, differed in the extent of their swelling during oxygen-glucose deprivation (OGD). In the present study we focused on identifying the ion channels or transporters that might underlie the different capabilities of these two astrocytic subpopulations to regulate their volume during OGD. Using three-dimensional confocal morphometry, which enables quantification of the total astrocytic volume, the effects of selected inhibitors of K⁺ and Cl⁻ channels/transporters or glutamate transporters on astrocyte volume changes were determined during 20 minute-OGD in situ. The inhibition of volume regulated anion channels (VRACs) and two-pore domain potassium channels (K(2P)) highlighted their distinct contributions to volume regulation in HR-/LR-astrocytes. While the inhibition of VRACs or K(2P) channels revealed their contribution to the swelling of HR-astrocytes, in LR-astrocytes they were both involved in anion/K⁺ effluxes. Additionally, the inhibition of Na⁺-K⁺-Cl⁻ co-transporters in HR-astrocytes led to a reduction of cell swelling, but it had no effect on LR-astrocyte volume. Moreover, employing real-time single-cell quantitative polymerase chain reaction (PCR), we characterized the expression profiles of EGFP-positive astrocytes with a focus on those ion channels and transporters participating in astrocyte swelling and volume regulation. The PCR data revealed the existence of two astrocytic subpopulations markedly differing in their gene expression levels for inwardly rectifying K⁺ channels (Kir4.1), K(2P) channels (TREK-1 and TWIK-1) and Cl⁻ channels (ClC2). Thus, we propose that the diverse volume changes displayed by cortical astrocytes during OGD mainly result from their distinct expression patterns of ClC2 and K(2P) channels.
- MeSH
- astrocyty cytologie účinky léků metabolismus MeSH
- biologické modely MeSH
- chloridové kanály metabolismus MeSH
- draslíkové kanály metabolismus MeSH
- gliový fibrilární kyselý protein metabolismus MeSH
- glukosa nedostatek MeSH
- kyslík MeSH
- lidé MeSH
- modulátory membránového transportu farmakologie MeSH
- mozková kůra cytologie MeSH
- myši transgenní MeSH
- myši MeSH
- pohlavní dimorfismus MeSH
- regulace genové exprese účinky léků MeSH
- sodík-draslík-chloridové symportéry metabolismus MeSH
- stanovení celkové genové exprese MeSH
- symportéry metabolismus MeSH
- techniky in vitro MeSH
- velikost buňky účinky léků MeSH
- vezikulární transportní proteiny pro glutamát metabolismus MeSH
- zelené fluorescenční proteiny metabolismus MeSH
- zvířata MeSH
- Check Tag
- lidé MeSH
- mužské pohlaví MeSH
- myši MeSH
- ženské pohlaví MeSH
- zvířata MeSH
- Publikační typ
- časopisecké články MeSH
- práce podpořená grantem MeSH
- MeSH
- astrocyty fyziologie účinky léků MeSH
- extracelulární prostor metabolismus MeSH
- finanční podpora výzkumu jako téma MeSH
- membránové potenciály fyziologie MeSH
- mícha cytologie MeSH
- velikost buňky účinky léků MeSH
- vodní a elektrolytová rovnováha MeSH
- vztah mezi dávkou a účinkem léčiva MeSH
- zvířata MeSH
- Check Tag
- zvířata MeSH
